WO2023077447A1 - 环肽玻璃及含有环肽的药物组合物玻璃 - Google Patents

环肽玻璃及含有环肽的药物组合物玻璃 Download PDF

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WO2023077447A1
WO2023077447A1 PCT/CN2021/129065 CN2021129065W WO2023077447A1 WO 2023077447 A1 WO2023077447 A1 WO 2023077447A1 CN 2021129065 W CN2021129065 W CN 2021129065W WO 2023077447 A1 WO2023077447 A1 WO 2023077447A1
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glass
cyclic peptide
temperature
pharmaceutical composition
cyclopeptide
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PCT/CN2021/129065
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English (en)
French (fr)
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闫学海
邢蕊蕊
袁成前
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中国科学院过程工程研究所
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/439—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom the ring forming part of a bridged ring system, e.g. quinuclidine
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/04—Peptides having up to 20 amino acids in a fully defined sequence; Derivatives thereof
    • A61K38/05—Dipeptides
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/04—Peptides having up to 20 amino acids in a fully defined sequence; Derivatives thereof
    • A61K38/12—Cyclic peptides, e.g. bacitracins; Polymyxins; Gramicidins S, C; Tyrocidins A, B or C
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/04—Peptides having up to 20 amino acids in a fully defined sequence; Derivatives thereof
    • A61K38/14—Peptides containing saccharide radicals; Derivatives thereof, e.g. bleomycin, phleomycin, muramylpeptides or vancomycin
    • 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
    • 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
    • A61K47/30—Macromolecular organic or inorganic compounds, e.g. inorganic polyphosphates
    • A61K47/42—Proteins; Polypeptides; Degradation products thereof; Derivatives thereof, e.g. albumin, gelatin or zein
    • 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
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P3/00—Drugs for disorders of the metabolism
    • A61P3/08—Drugs for disorders of the metabolism for glucose homeostasis
    • A61P3/10—Drugs for disorders of the metabolism for glucose homeostasis for hyperglycaemia, e.g. antidiabetics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/04—Antibacterial agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/12—Antivirals
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • 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
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D498/00—Heterocyclic compounds containing in the condensed system at least one hetero ring having nitrogen and oxygen atoms as the only ring hetero atoms
    • C07D498/12—Heterocyclic compounds containing in the condensed system at least one hetero ring having nitrogen and oxygen atoms as the only ring hetero atoms in which the condensed system contains three hetero rings
    • C07D498/18—Bridged systems
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D498/00—Heterocyclic compounds containing in the condensed system at least one hetero ring having nitrogen and oxygen atoms as the only ring hetero atoms
    • C07D498/22—Heterocyclic compounds containing in the condensed system at least one hetero ring having nitrogen and oxygen atoms as the only ring hetero atoms in which the condensed system contains four or more hetero rings
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K11/00—Depsipeptides having up to 20 amino acids in a fully defined sequence; Derivatives thereof
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C07K14/575—Hormones
    • C07K14/605—Glucagons
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K5/00—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof
    • C07K5/04—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing only normal peptide links
    • C07K5/10—Tetrapeptides
    • C07K5/1002—Tetrapeptides with the first amino acid being neutral
    • C07K5/1005—Tetrapeptides with the first amino acid being neutral and aliphatic
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K5/00—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof
    • C07K5/04—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing only normal peptide links
    • C07K5/12—Cyclic peptides with only normal peptide bonds in the ring
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K7/00—Peptides having 5 to 20 amino acids in a fully defined sequence; Derivatives thereof
    • C07K7/04—Linear peptides containing only normal peptide links
    • C07K7/06—Linear peptides containing only normal peptide links having 5 to 11 amino acids
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K7/00—Peptides having 5 to 20 amino acids in a fully defined sequence; Derivatives thereof
    • C07K7/50—Cyclic peptides containing at least one abnormal peptide link
    • C07K7/54—Cyclic peptides containing at least one abnormal peptide link with at least one abnormal peptide link in the ring
    • C07K7/56—Cyclic peptides containing at least one abnormal peptide link with at least one abnormal peptide link in the ring the cyclisation not occurring through 2,4-diamino-butanoic acid
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K7/00—Peptides having 5 to 20 amino acids in a fully defined sequence; Derivatives thereof
    • C07K7/50—Cyclic peptides containing at least one abnormal peptide link
    • C07K7/54—Cyclic peptides containing at least one abnormal peptide link with at least one abnormal peptide link in the ring
    • C07K7/56—Cyclic peptides containing at least one abnormal peptide link with at least one abnormal peptide link in the ring the cyclisation not occurring through 2,4-diamino-butanoic acid
    • C07K7/58—Bacitracins; Related peptides
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K7/00—Peptides having 5 to 20 amino acids in a fully defined sequence; Derivatives thereof
    • C07K7/50—Cyclic peptides containing at least one abnormal peptide link
    • C07K7/54—Cyclic peptides containing at least one abnormal peptide link with at least one abnormal peptide link in the ring
    • C07K7/60—Cyclic peptides containing at least one abnormal peptide link with at least one abnormal peptide link in the ring the cyclisation occurring through the 4-amino group of 2,4-diamino-butanoic acid
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K7/00—Peptides having 5 to 20 amino acids in a fully defined sequence; Derivatives thereof
    • C07K7/50—Cyclic peptides containing at least one abnormal peptide link
    • C07K7/54—Cyclic peptides containing at least one abnormal peptide link with at least one abnormal peptide link in the ring
    • C07K7/60—Cyclic peptides containing at least one abnormal peptide link with at least one abnormal peptide link in the ring the cyclisation occurring through the 4-amino group of 2,4-diamino-butanoic acid
    • C07K7/62—Polymyxins; Related peptides
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K7/00—Peptides having 5 to 20 amino acids in a fully defined sequence; Derivatives thereof
    • C07K7/64—Cyclic peptides containing only normal peptide links
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K7/00—Peptides having 5 to 20 amino acids in a fully defined sequence; Derivatives thereof
    • C07K7/64—Cyclic peptides containing only normal peptide links
    • C07K7/66—Gramicidins S, C; Tyrocidins A, B, C; Related peptides
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K9/00—Peptides having up to 20 amino acids, containing saccharide radicals and having a fully defined sequence; Derivatives thereof

Definitions

  • the invention belongs to the technical field of medicine, and relates to a new dosage form and a new auxiliary material of cyclopeptide medicine, in particular to a cyclopeptide glass and a cyclopeptide medicine composition glass.
  • it involves related technologies for effectively utilizing insoluble drugs, and in particular relates to a preparation method for converting insoluble cyclic peptide drugs into glass states to form glass drugs.
  • This cyclic peptide glass can be used in the field of drug delivery such as anti-tumor, anti-virus/anti-bacterial, blood sugar control, immune regulation, neuromodulation, etc., especially to achieve drug sustained release therapy.
  • Cyclic peptides are formed by condensation and cyclization of multiple amino acids through peptide bonds, and their structures have certain conformational constraints. The unique topology of cyclic peptides makes them exceptionally stable against chemical, thermal and biological degradation. There are a large number of hydrogen bond acceptors and donors in cyclic peptides, and hydrogen bonds are one of the main ways for drugs to interact with receptors. Therefore, cyclic peptides often have certain biological and pharmacological activities and become an important component in medicinal chemistry. Pharmacophore.
  • cyclic peptides exhibit a wide range of biological and pharmacological activities, such as antitumor, antiviral, antibacterial, antiaging, Immunomodulation, memory enhancement, blood sugar regulation, etc.
  • the glassy state is a metastable amorphous structure that retains the biological and pharmacological activities of cyclic peptide molecules.
  • glassy cyclic peptide drugs exhibit high surface free energy and high dispersion, which can effectively improve the dissolution rate and bioavailability of cyclic peptides.
  • the cyclopeptide glass of the present invention can completely replace traditional pharmaceutical excipients and realize functions such as dispersion, solubilization, adhesion, and controlled release.
  • it can be used as a substitute for castor oil pharmaceutical excipients to effectively avoid severe allergic reactions caused by histamine.
  • cyclic peptide glass especially the glass based on pharmaceutical composition with biological/pharmacological activity and sustained release function, and the method of converting insoluble cyclic peptide into glass state.
  • the present invention finds that cyclic peptide and its derivatives can be processed into cyclic peptide glass and cyclic peptide pharmaceutical composition glass through a specific preparation process.
  • the present invention also discovers a method for converting insoluble cyclic peptide drugs into a glass state, which effectively increases the dissolution rate and bioavailability of cyclic peptide drugs.
  • the present invention has been accomplished based on this discovery. Based on the cyclic peptide glass discovered in the present invention, it is expected to be used as an active drug or a pharmaceutical excipient, and is widely used in the fields of drug delivery and sustained release in antitumor, antiviral/antibacterial, blood sugar control, immune regulation, neuromodulation, etc.
  • the primary purpose of the present invention is to provide a cyclopeptide glass and a cyclopeptide pharmaceutical composition glass. And the method of converting insoluble cyclic peptide into glassy state.
  • the glass refers to an amorphous solid that exhibits a glass transition phenomenon
  • the glass state refers to a disordered organizational structure that maintains glass-like properties, and has no crystal structure in X-ray diffraction detection, and There is a defined glass transition temperature.
  • the present invention provides a method for transforming an insoluble cyclic peptide from a crystalline state to a glass state, effectively improving the in vitro dissolution rate and solubility of the insoluble cyclic peptide, and increasing the bioavailability of the insoluble cyclic peptide drug.
  • the first aspect cyclic peptide-based glass, characterized in that, the cyclic peptide is a cyclic peptide and/or its salt in the form of structural formula 1, and the cyclic peptide is one or a combination of two or more,
  • Said A 1 -A n are independently selected from:
  • Glycine Alanine, Valine, Leucine, Isoleucine, Methionine (Methionine), Proline, Tryptophan, Serine, Tyrosine, Cysteine, Phenylalanine
  • R 1 -R n are independently selected from H or other modifiable groups, and the modifiable groups are preferably methyl, alkyl, phosphoric acid, acetyl, formyl, fatty acid, benzoyl, amide, ester, 9-fluorenyl Methoxycarbonyl, tert-butoxycarbonyl;
  • n ⁇ 2 preferably, 2 ⁇ n ⁇ 15, A 1 -A n are connected through amino acid condensation.
  • the cyclic peptide has biological and/or pharmacological activity; more preferably, it has antibacterial/antiviral, antitumor, blood sugar regulation, and immune regulation activities, wherein,
  • Antibacterial/antiviral cyclic peptides include but are not limited to the following structures and their analogues:
  • Anti-tumor cyclic peptides include but are not limited to the following structures and similar structures:
  • Immunomodulatory cyclic peptides include but are not limited to the following structures and similar structures:
  • Blood glucose regulating cyclic peptides include but are not limited to the following structures and similar structures:
  • Cardiovascular and blood-related cyclic peptides include, but are not limited to, the following structures and their analogues:
  • active cyclic peptides include, but are not limited to, the following structures and their analogs:
  • the cyclic peptide derivatives are molecules, isomers and salts thereof similar to the above-mentioned cyclic peptide molecular structure skeleton.
  • the cyclic peptide is a cyclic peptide with poor water solubility, and the poor water solubility means that the maximum dissolved concentration of the cyclic peptide in pure water at normal temperature and pressure is ⁇ 5wt%.
  • the cyclic peptide-based glass is characterized in that it is completely prepared from cyclic peptides and salts; or, other pharmaceutical excipients and/or pharmaceutical active ingredients are added to the glass.
  • the pharmaceutical composition glass is composed of one or more of the following components: cyclic peptide or cyclic peptide derivatives, other pharmaceutical active ingredients or their pharmaceutically acceptable salts, pharmaceutically acceptable excipients; preferably, at least one active ring peptide;
  • the active ingredients of the drug include one or more mixtures of acyclic peptide drug molecules with antibacterial/antiviral, antitumor, blood sugar regulation, immune regulation, antipsychotic and other effects.
  • the antibacterial/antiviral drug is penicillin , cephalexin, amikacin, norfloxacin, nitrofurantoin, metronidazole, amantadine, acyclovir, zidovudine, ribavirin; antineoplastic drugs are cisplatin, doxorubicin, Vincristine, paclitaxel, docetaxel, gemcitabine, camptothecin, hydroxycamptothecin, irinotecan, etoposide, dexamethasone, fluorouracil, cyclophosphamide; blood glucose regulating drugs are metformin, repaglinide, Nateglinide, insulin; immunomodulatory drugs include thymopentin, tripterygium glycosides, tript
  • compositions include fillers, wetting agents, binders, disintegrants, lubricants and others, preferably, fillers are starch, lactose, mannitol; wetting agents/binders are sodium carboxymethylcellulose, Hydroxypropyl cellulose; disintegrants are sodium carboxymethyl starch, low-substituted hydroxypropyl cellulose, cross-linked polyvinylpyrrolidone, cross-linked sodium carboxymethyl cellulose; lubricants are polyethylene glycol, stearic acid Magnesium, hydrogenated vegetable oil; others are sodium lauryl sulfate;
  • the mass ratio of cyclic peptide is between 1-100%, preferably 1%-50%;
  • the mass ratio of other pharmaceutical active ingredients is between 1-20%, preferably 1-10%;
  • the mass ratio of pharmaceutically acceptable excipients is between 0-50%, preferably 0-5%, more preferably 0%, that is, cyclopeptide glass completely replaces traditional pharmaceutical excipients.
  • the cyclic peptide-based glass is characterized in that it can be used as a pharmaceutical excipient, and can completely or partially replace the pharmaceutical excipient.
  • a method for converting an insoluble cyclic peptide into a glass state to prepare a cyclic peptide glass drug or a cyclic peptide glass excipient is characterized in that it mainly includes the following steps:
  • the fully ground cyclic peptide raw material is heated to a temperature near the melting point in an inert gas atmosphere, and after a period of heat preservation treatment, it is transferred to an annealing furnace for annealing treatment, that is, the prepared Cyclopeptide glass drugs or cyclopeptide glass excipients;
  • Heating temperature refers to the melting point temperature (T m ) ⁇ 50 ⁇ 250K temperature, preferably, higher than T m 50 ⁇ 100K;
  • the holding time is 0min-30h, preferably 15-30min;
  • Annealing temperature refers to the glass transition temperature (T g ) ⁇ 50 ⁇ 150K temperature, preferably, is lower than T g 50 ⁇ 100K;
  • the annealing treatment time is 30min-2h, preferably, 30min-1h;
  • T m and T g are measured by thermogravimetric analysis and differential scanning calorimetry, and the heating and cooling rates are 2-50Kmin -1 .
  • a method for preparing glass of an insoluble cyclic peptide pharmaceutical composition is characterized in that it mainly includes the following steps:
  • step (3) The glass obtained in step (2) and pharmaceutically acceptable excipients are co-made into a pharmaceutical composition by tableting, wet granulation, fluidized bed granulation, coating, spray granulation, programmed casting, 3D One or more combination of printing.
  • the heating temperature is a melting point temperature (T m ) ⁇ 50 ⁇ 250K temperature, preferably, 50 ⁇ 100K higher than T m ;
  • the holding time is 0min-30h, preferably 15-30min;
  • the good solvent is preferably ethanol, hexafluoroisopropanol, dichloromethane, carbon tetrachloride, acetone, ethyl acetate
  • Cosolvent is preferably one or more mixtures of sodium benzoate, dimethylacetamide, urea, Tween;
  • step (3) (4) mixing the active pharmaceutical ingredient solution obtained in step (3) with the molten cyclic peptide in step (1);
  • step (4) The mixture obtained in step (4) is placed at the temperature set in step (2), and the solvent is rotary evaporated under reduced pressure;
  • the annealing temperature is a glass transition temperature (T g ) ⁇ 50-150K, preferably 50-100K lower than T g ;
  • the annealing treatment time is 30min-2h, preferably, 30min-1h;
  • step (6) The glass obtained in step (6) and pharmaceutically acceptable excipients are co-made into a pharmaceutical composition by tableting, wet granulation, fluidized bed granulation, coating, spray granulation, programmed casting, 3D One or more combination of printing.
  • the insoluble cyclic peptide and other pharmaceutical active ingredients together in a good solvent and a co-solvent to dissolve completely
  • the good solvent is preferably ethanol, hexafluoroisopropanol, methylene chloride, carbon tetrachloride, acetone , ethyl acetate
  • the cosolvent is preferably one or more mixtures of sodium benzoate, dimethylacetamide, urea, Tween;
  • step (2) The mixed solution obtained in step (1) is heated up under an inert gas atmosphere, and the solvent is evaporated by rotary evaporation under reduced pressure, followed by heat preservation treatment.
  • T m 50-100K holding time is 0min-30h, preferably 15-30min;
  • the annealing temperature is a glass transition temperature (T g ) ⁇ 50-150K, preferably 50-100K lower than T g ;
  • the annealing treatment time is 30min-2h, preferably, 30min-1h;
  • step (3) The glass obtained in step (3) and pharmaceutically acceptable excipients are co-made into a pharmaceutical composition by tableting, wet granulation, fluidized bed granulation, coating, spray granulation, programmed casting, 3D One or more combination of printing.
  • the glass pharmaceutical dosage form can be oral preparation, patch, subcutaneous pack Implants, scaffold materials and microneedle devices.
  • the processes involved are tablet compression, dry granulation, high-shear wet granulation, fluidized bed granulation, capsule filling, microcapsule embedding, coating , spray drying, spray condensation, photolithography, programmed pouring, microneedle array, 3D printing, or one or more of them.
  • the cyclic peptide glass, the cyclic peptide drug composition glass, and the cyclic peptide drug glass dosage form of the present invention have the following advantages and beneficial effects:
  • the cyclic peptide glass, the cyclic peptide drug composition glass, and the cyclic peptide drug glass dosage form of the present invention have the following applications: drug delivery for anti-tumor, anti-viral/antibacterial, blood sugar control, immune regulation, neuromodulation, etc. field, especially to achieve drug sustained release therapy.
  • the raw materials of the cyclic peptide glass of the present invention include cyclic peptides, especially any one or more combinations of cyclic peptides with biological/pharmacological activity or their pharmaceutically acceptable derivatives and their salts, which are generally water-soluble Poor stability, easy crystallization, low bioavailability.
  • the present invention creatively proposes a method for converting an insoluble cyclic peptide drug into a glass state, including preparing the insoluble cyclic peptide into a glass, and co-making the insoluble cyclic peptide into a glass with other active pharmaceutical ingredients.
  • the invention provides a method for converting poorly soluble drugs into glass states to form drug glass dosage forms to realize the slow release of poorly soluble active cyclic peptides and drugs.
  • the obtained cyclic peptide glass has high biocompatibility, degradability and high thermal/chemical stability, and can be further processed into oral agents, patches, subcutaneous embedding agents, and stents by spray drying, solid dispersion, and additive manufacturing technologies materials and microneedle devices.
  • the obtained cyclopeptide glass can play the role of drug efficacy and drug excipients at the same time. Compared with crystals and traditional drug dosage forms or excipients, cyclopeptide glass can effectively increase the dissolution rate of drugs and improve the bioavailability of drugs. Drug delivery and sustained release fields such as blood sugar control, immune regulation, and neuromodulation are widely used.
  • Figure 1 is the physical picture (a) of the CsA glass prepared in Example 1 at room temperature and the physical picture (b) of the CsA glass obtained by the pouring process, which proves the processability of the cyclopeptide glass.
  • Figure 2 is the H NMR spectrum of the CsA glass prepared in Example 1. Compared with the CsA raw material, the peaks have not changed significantly, indicating that the chemical composition of the cyclic peptide raw material molecules has not changed after heating, melting and annealing.
  • DSC differential scanning calorimetry
  • FIG. 4 is an X-ray diffraction (XRD) spectrum of the CsA glass prepared in Example 1, which shows that the CsA glass has an amorphous structure.
  • XRD X-ray diffraction
  • Fig. 5 is the stability test result of the CsA glass prepared in Example 1, confirming its high stability performance.
  • Fig. 6 is the drug sustained release curve of the CsA glass prepared in Example 1, which proves that the drug can be slowly released.
  • FIG. 7 shows the immune-related indicators of mice after oral administration of the CsA glass prepared in Example 1.
  • Fig. 8 is the XRD spectrum of the glass of the pharmaceutical composition based on CFP and the tumor chemotherapeutic drug PTX in Example 2, which is an amorphous structure.
  • Figure 9 shows the degradation of the glass capsule of the pharmaceutical composition prepared in Example 2 in artificial gastric juice (configured according to the preparation method of the Chinese Pharmacopoeia), confirming its degradable property.
  • Fig. 10 is the drug-time curve of the glass capsule of the pharmaceutical composition prepared in Example 2 in the mouse body over time after intragastric administration.
  • Fig. 11 is the tumor inhibition curve of experimental mice after oral administration of the glass capsule of the pharmaceutical composition prepared in Example 2, which proves to have an anti-tumor effect.
  • Fig. 12 is the body weight change curve of the experimental mice after oral administration of the glass capsule of the pharmaceutical composition prepared in Example 2, which proves the high biological safety.
  • Fig. 13 shows the changes of organ coefficients of experimental mice after oral administration of the glass pharmaceutical composition capsule prepared in Example 2, demonstrating high biological safety.
  • Figure 14 is the melting image and XRD data of the glass of the pharmaceutical composition of Example 3, which confirms the transformation from crystal to glass.
  • Fig. 15 is the biocompatibility test result of the pharmaceutical composition glass prepared in Example 3, which proves to have high biocompatibility.
  • Fig. 16 is the curve of inhibiting bacterial growth of the glass coating of the pharmaceutical composition prepared in Example 3.
  • Fig. 17 is the antibacterial effect of the pharmaceutical composition glass tablet prepared in Example 3 on Escherichia coli and Staphylococcus aureus.
  • Fig. 18 is the degradation curve over time after subcutaneous embedding of the embedding agent of the glass pharmaceutical composition prepared in Example 3.
  • Fig. 19 is the control curve of the glass tablet of the pharmaceutical composition prepared in Example 4 on the blood sugar of spontaneous type II diabetic mice, which proves that the blood sugar of the mice can be effectively controlled.
  • Fig. 20 is the glass tablet of the pharmaceutical composition prepared in Example 4, administered to spontaneous type II diabetic mice by intragastric administration, and the change curve of the mouse body weight over time, which proves that the mouse body weight can be effectively controlled.
  • Figure 21 is the tablet of the pharmaceutical composition glass prepared in Example 4, after intragastric administration to spontaneous type II diabetic mice for 30 days, the tolerance of oral high-concentration glucose, the glucose concentration is 2.5g kg - 1.
  • Figure 22 is the DSC curve of the CPY-CFP-CLP cyclic peptide mixed glass prepared in Example 5, and the glass transition temperatures of the single and mixed glasses are obtained, indicating that mixing multiple cyclic peptides can effectively inhibit the crystallization tendency of a single cyclic peptide.
  • Figure 23 is the release curve of CLP molecules from the mixed cyclic peptide glass and CLP crystals prepared in Example 5, indicating that the mixed cyclic peptide glass can increase the dissolution rate of the CLP active cyclic peptide.
  • Fig. 24 is the XRD pattern of the glass of the pharmaceutical composition prepared in Example 5, which is amorphous.
  • Fig. 25 is the statistical data of autonomous activity ability and anxiety-like behavior of mice in the open field experiment. The results showed that compared with normal mice, the distance of movement in the central area of the early social isolation mice was significantly reduced, and it was significantly improved after treatment with the glass microneedle patch.
  • Figure 26 shows the number of times mice entered the open arm and closed arm within 5 minutes and the residence time in the two arms in the maze experiment. The results showed that compared with normal mice, the number of times of entering the open arm and the residence time in the open arm were significantly reduced in the early social isolation mice, which were significantly improved after being treated with the glass microneedle patch.
  • a kind of preparation method of the cyclopeptide glass based on cyclosporine A comprises the steps:
  • step (1) (2) the crucible that CsA powder is housed in step (1) is placed in heating equipment under N atmosphere;
  • step (2) Heat the equipment in step (2), heat the crucible from room temperature to a temperature of 573.15K at a heating rate of 10K min -1 , and heat-insulate at this temperature for 20 minutes;
  • step (3) Cooling the equipment in step (3), lowering the crucible to a constant temperature of 273.15K in an annealing furnace for 30 minutes at a cooling rate of 10K min -1 , and annealing the glass to obtain CsA glass.
  • Figure 1 is the physical picture (a) of the CsA glass prepared in Example 1 at room temperature and the physical picture (b) of the CsA glass obtained by the pouring process, which proves the processability of the cyclopeptide glass.
  • Figure 2 is the H NMR spectrum of the CsA glass prepared in Example 1. Compared with the CsA raw material, the peaks have not changed significantly, indicating that the chemical composition of the cyclic peptide raw material molecules has not changed after heating, melting and annealing.
  • FIG. 4 is an X-ray diffraction (XRD) spectrum of the CsA glass prepared in Example 1, which shows that the CsA glass has an amorphous structure.
  • XRD X-ray diffraction
  • Fig. 5 is the stability test result of the CsA glass prepared in Example 1, confirming its high stability performance.
  • Fig. 6 is the drug sustained release curve of the CsA glass prepared in Example 1, which proves that the drug can be slowly released.
  • the initial mass of CsA glass is 20 ⁇ 1mg
  • the stirring rate is 100r min -1 , through the dissolution tester and high performance liquid chromatography The dissolved CsA was detected, and the sustained-release curve of the drug was drawn.
  • FIG. 7 shows the immune-related indicators of mice after oral administration of the CsA glass prepared in Example 1.
  • healthy male Kunming white mice (body weight 20 ⁇ 1 g, 4-5 weeks old) were randomly divided into two groups, 8 rats in each group, and fed a normal diet.
  • the mice in the experimental group were intragastrically administered every other day, the dosage was 40 mg kg -1 , and the administration times were 5, and the mice in the control group were intragastrically administered normal saline of equal quality every other day.
  • the day after the last oral administration the mice were euthanized to detect the corresponding indicators.
  • mice The thymus and spleen of the mice were dissected, the surface blood was washed away with phosphate buffered saline (PBS), the water was blotted with filter paper, and then weighed on an electronic balance, and the organ weight (mg) was divided by the total weight of the mouse (g ) to calculate the organ weight index.
  • PBS phosphate buffered saline
  • the organ weight index was divided by the total weight of the mouse (g ) to calculate the organ weight index.
  • a kind of preparation method of the pharmaceutical composition glass based on phenylalanine-proline cyclic dipeptide (CFP) and paclitaxel (PTX) comprises the steps:
  • step (1) heat-treating the container of step (1), heating the container from room temperature to a temperature of 473.15K at a heating rate of 20K min -1 , and heat-insulating at this temperature for 30 minutes;
  • step (3) The container of step (2) is cooled, and the container is reduced to a temperature of 333.15K at a cooling rate of 10K min -1 ;
  • step (4) Add the solution of step (4) dropwise into the container of step (3), place the container in a vacuum rotary evaporator, maintain the temperature at 333.15K, and remove the solvent;
  • step (5) The equipment in step (5) is cooled, and the container is lowered to a constant temperature of 315.15K in an annealing furnace for 30 minutes at a cooling rate of 10K min ⁇ 1 to obtain a pharmaceutical composition glass based on CFP and the tumor chemotherapy drug PTX , the above glass is completely dissolved in the HFIP organic solution, the concentration of PTX is quantified by high performance liquid chromatography, and the drug loading amount of PTX is calculated to be 3.5 ⁇ 0.2%;
  • Fig. 8 is the XRD spectrum of the glass of the pharmaceutical composition based on CFP and tumor chemotherapy drug PTX in Example 2, which proves that the pharmaceutical composition of the present invention has an amorphous structure.
  • Figure 9 shows the degradation of the glass capsule of the pharmaceutical composition prepared in Example 2 in artificial gastric juice (following the preparation method of the Chinese Pharmacopoeia), confirming its degradable property.
  • the preparation steps of the glass capsule of the pharmaceutical composition include: grinding the glass of the pharmaceutical composition into powder through a homogenizer, and filling the powder into a gelatin capsule shell.
  • Fig. 10 is the drug-time curve of the glass capsule of the pharmaceutical composition prepared in Example 2 in the mouse body over time after intragastric administration. Specifically, mice were given a dosage of 8 mg kg -1 , and the peripheral blood of the mice was collected by taking blood from the orbit over time after intragastric administration, cell lysis and solvent dissolution were performed, and the filtered solution was detected by high performance liquid chromatography , calculate the content of PTX in the blood, and draw the drug-time curve.
  • Fig. 11 is the tumor inhibition curve of experimental mice after oral administration of the glass capsule of the pharmaceutical composition prepared in Example 2, which proves to have an anti-tumor effect.
  • healthy BALB/c nude mice weight 15 ⁇ 1 g, 4-6 weeks old
  • 4T1 cells were subcutaneously inoculated in the axilla to establish a breast cancer 4T1 mouse xenograft model.
  • mice were treated with drugs.
  • Mice in the experimental group were administered intragastrically every other day, with a dose of 8 mg kg -1 , for a total of 5 administration times.
  • mice in the control group were intragastrically administered PBS of equal quality every other day.
  • Fig. 12 is the body weight change curve of the experimental mice after oral administration of the glass capsule of the pharmaceutical composition prepared in Example 2, which proves the high biological safety.
  • Fig. 13 shows the changes of organ coefficients of experimental mice after oral administration of the glass capsule of the pharmaceutical composition prepared in Example 2, demonstrating high biological safety.
  • the important organs including heart, liver, spleen, lung, and kidney
  • the superficial blood was washed away with PBS, and the water was blotted with filter paper, and then weighed on an electronic balance.
  • the organ coefficient was calculated by dividing the weight (mg) by the total weight of the mouse (g).
  • a kind of preparation method of the pharmaceutical composition glass based on vancomycin, gramicidin S and penicillin comprises the steps:
  • step (1) Add a mixed organic solvent of hexafluoroisopropanol and ethanol to the mixed powder in step (1), and stir until completely dissolved;
  • step (3) Place the mixed solution obtained in step (2) in a reduced-pressure rotary evaporator, raise the temperature from room temperature to 443.15K at a heating rate of 50K min -1 , and heat-preserve at this temperature for 30 minutes to remove the organic solvent;
  • step (3) The equipment in step (3) is cooled down to a temperature of 243.15K at a cooling rate of 10K min ⁇ 1 , and kept at this temperature for 20 minutes;
  • step (4) Add a certain quality of pharmaceutically acceptable excipients to the product obtained in step (4), including 1.5 parts of starch, 1 part of sodium carboxymethylcellulose, 2.5 parts of polyethylene glycol and 6.5 parts of hydrogenated vegetable oil, the total mass of the excipients The ratio is 92.5%;
  • step (5) The pharmaceutical composition obtained in step (5) is homogenized, combined with tabletting and vacuum spray-drying techniques to prepare an embedding agent for a glass pharmaceutical composition.
  • Figure 14 is the melting image and XRD data of the glass of the pharmaceutical composition of Example 3, which confirms the transformation from crystal to glass.
  • Fig. 15 is the biocompatibility test result of the pharmaceutical composition glass prepared in Example 3, which proves to have high biocompatibility. Specifically, the above-mentioned glass was machined into a cell culture dish coating with a diameter of 60 mm and a thickness of 0.2 mm for cell culture (fibroblast 3T3). After co-incubating for 48 hours, the survival rate of the cells was tested by the MTT method.
  • Fig. 16 is the curve of inhibiting bacterial growth of the glass coating of the pharmaceutical composition prepared in Example 3.
  • the above-mentioned medicinal composition glass is processed into a tablet of 1cm*1cm*0.1cm, and pasted on the center of the bacterial culture dish, the total concentration of the drug is calculated to be 10ug/dish, and Escherichia coli and Staphylococcus aureus are mixed in a liquid medium After mixing evenly in the medium, drop it into the bacterial culture dish, incubate with shaking at 37°C, and measure the concentration of bacteria in the bacterial culture dish over time by using the OD value method.
  • Fig. 17 is the antibacterial effect of the pharmaceutical composition glass tablet prepared in Example 3 on Escherichia coli and Staphylococcus aureus.
  • the above-mentioned pharmaceutical composition glass is processed into a tablet with a diameter of 5 mm and a thickness of 0.2 mm, and an Oxford cup (6 mm in diameter) is placed on an agar petri dish, and the tablet is placed in the Oxford cup to fit tightly with the agar.
  • Escherichia coli and Staphylococcus aureus evenly with PBS, spread them on a petri dish, place them in a 37°C incubator and cultivate them.
  • After 24 hours observe whether the antibacterial ring is produced or not, and judge the inhibitory effect of the glass of the pharmaceutical composition. bacteria effect.
  • Fig. 18 is the degradation curve over time after subcutaneous embedding of the embedding agent of the pharmaceutical composition glass prepared in Example 3.
  • healthy male Kunming white mice (body weight 20 ⁇ 1 g, 4-5 weeks old) were randomly divided into two groups, 15 rats in each group.
  • a 1 cm window was surgically cut on the back of the mice in the experimental group, and 30 ⁇ 2.5 g of glass of the pharmaceutical composition was buried subcutaneously, and then sutured.
  • Take any mouse over time take out the glass of the medicinal composition, weigh the remaining mass, and draw the degradation curve.
  • Fig. 19 is the control curve of the glass tablet of the pharmaceutical composition prepared in Example 4 on the blood sugar of spontaneous type II diabetic mice, which proves that the blood sugar of the mice can be effectively controlled.
  • ordered spontaneous type II diabetes model mice male, 8 weeks old
  • mice in the experimental group were fed with glass of the medicinal composition every day at a dose of 4 mg kg -1
  • the mice in the control group were fed with normal saline of equal quality, and the test period was 30 days.
  • the tail blood of the mice was taken every day, and the fasting blood glucose of the mice was measured with a blood glucose tester.
  • the abscissa is time, and the ordinate is the fasting blood glucose level of the experimental mice to draw a curve.
  • Fig. 20 is the glass tablet of the pharmaceutical composition prepared in Example 4, administered to spontaneous type II diabetic mice by intragastric administration, and the change curve of the mouse body weight over time, which proves that the mouse body weight can be effectively controlled.
  • Figure 21 is the tablet of the pharmaceutical composition glass prepared in Example 4, after intragastric administration to spontaneous type II diabetic mice for 30 days, the tolerance of oral high-concentration glucose, the glucose concentration is 2.5g kg - 1.
  • Figure 22 is the DSC curve of the CPY-CFP-CLP cyclic peptide mixed glass prepared in Example 5, and the glass transition temperatures of the single and mixed glasses are obtained, indicating that mixing multiple cyclic peptides can effectively inhibit the crystallization tendency of a single cyclic peptide.
  • Figure 23 is the release curve of CLP molecules from the mixed cyclic peptide glass and CLP crystals prepared in Example 5, indicating that the mixed cyclic peptide glass can increase the dissolution rate of the CLP active cyclic peptide.
  • the initial mass of the cyclic peptide mixed glass is 15 ⁇ 1mg
  • the stirring rate is 100r min -1
  • the dissolution tester and high performance liquid chromatography The dissolved CLP is detected, and the sustained release curve of the cyclic peptide mixed glass is drawn.
  • CLP crystals of the same mass were weighed and placed in the above slow-release solution for incubation with stirring, and the concentration of CLP in the slow-release solution was detected.
  • Fig. 24 is the XRD spectrum of the glass of the pharmaceutical composition prepared in Example 6, which is amorphous.
  • Fig. 25 is the statistical data of autonomous activity and anxiety-like behaviors of mice in the open field experiment of small animals in which the pharmaceutical composition glass prepared in Example 6 is applied.
  • the results showed that compared with normal mice, the distance of activity in the central area of early socially isolated mice was significantly reduced, and it was significantly improved after treatment with the glass microneedle patch.
  • C57BL/6 early social isolation mice (8 weeks old, male) were evenly divided into 2 groups, 6 mice in each group. One group of mice did not receive special treatment, and the other group of mice had glass microneedle patches pasted on their backs after depilation.
  • C57BL/6 group housed 6 normal mice as blank control group.
  • a black mine device (45cm*45cm*45cm) was prepared, the mice were placed in the mine, and the autonomous activity behavior of the mice was tracked and recorded.
  • the ordinate is the statistical value of the activity distance in the central area of the mice in the mine field for 1 hour.
  • Figure 26 shows the number of times the mouse enters the open arm and the closed arm within 5 minutes and the residence time in the two arms when the glass of the pharmaceutical composition prepared in Example 6 is applied to the mouse maze experiment.
  • the results showed that compared with normal mice, the number of times of entering the open arm and the residence time in the open arm were significantly reduced in the early social isolation mice, which were significantly improved after treatment with the microneedle patch.
  • a maze device with an arm width of 5 cm, an arm length of 35 cm, a closed arm height of 15 cm, a central platform of 5 cm*5 cm, and a maze height of about 40-55 cm from the ground. Place the mouse in the center of the platform, record the number of times the mouse enters the open arm and the closed arm and the time it stays in the two arms, and evaluate the indicators of anxiety.
  • a kind of preparation method of the pharmaceutical composition glass based on histidine-proline cyclic dipeptide and Siglipide comprises the steps:
  • step (3) Place the mixed solution obtained in step (2) in a reduced-pressure rotary evaporator, raise the temperature from room temperature to 535.15K at a heating rate of 10K min -1 , and heat-preserve at this temperature for 30 minutes to remove the organic solvent;
  • step (3) The equipment in step (3) is cooled down to a temperature of 283.15K at a cooling rate of 10K min -1 , and kept at this temperature for 1 hour to obtain a histidine-proline cyclic dipeptide and a Gliptide pharmaceutical composition glass;
  • step (5) After the glass of the pharmaceutical composition obtained in step (5) is subjected to homogenization treatment, it is prepared into tablets through tablet compression and programmed pouring, which are used for oral administration to regulate blood sugar.
  • step (2) Place the mixed solution obtained in step (1) in a reduced-pressure rotary evaporator, raise the temperature from room temperature to 473.15K at a heating rate of 10K min -1 , and heat-preserve at this temperature for 30 minutes to remove the organic solvent;
  • step (2) The equipment in step (2) is cooled down to 243.15K at a cooling rate of 10K min -1 , and kept at this temperature for 30 minutes to obtain a CPY-CFP-CLP cyclopeptide mixed glass.
  • a method for preparing a pharmaceutical composition glass based on tryptophan-tryptophan cyclic dipeptide (CWW) and risperidone comprises the steps:
  • step (2) Place the mixed solution obtained in step (1) in a reduced-pressure rotary evaporator, raise the temperature from room temperature to 573.15K at a heating rate of 10K min -1 , and heat-preserve at this temperature for 30 minutes to remove the organic solvent;
  • step (2) The equipment in step (2) is cooled down to a temperature of 283.15K at a cooling rate of 50K min -1 , and kept at this temperature for 30 minutes to obtain a glass pharmaceutical combination based on CWW and paliperidone thing;
  • the glass pharmaceutical composition obtained in step (3) is processed into a microneedle patch with a size of 0.8*0.8cm for transdermal drug delivery.
  • the early social isolation mouse model of C57BL/6 was established, treated with subcutaneous microneedle patch, and the dose of risperidone was 5 mg kg -1 . After 14 days, the schizophrenia-like behavior of the mice was evaluated.

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Abstract

本发明公开了一种环肽玻璃、及含有环肽的药物组合物玻璃。本发明的环肽玻璃可同时发挥药效及药物辅料功能，与晶体及传统药物剂型或辅料相比，环肽玻璃可有效增加药物溶出速率、提高药物生物利用度，在抗肿瘤、抗病毒/抗菌、血糖控制、免疫调节、神经调节等的药物递送及缓释领域应用广泛。

Description

环肽玻璃及含有环肽的药物组合物玻璃 技术领域：
本发明属于医药技术领域，涉及环肽药物新剂型及新辅料，特别涉及一种环肽玻璃、及环肽药物组合物玻璃。同时涉及有效地利用难溶性药物的相关技术，特别涉及一种将难溶性环肽药物转化为玻璃态，形成玻璃药物的制备方法。此环肽玻璃可应用于抗肿瘤、抗病毒/抗菌、血糖控制、免疫调节、神经调节等的药物递送领域，特别是实现药物缓释治疗。
背景技术：
环肽是多个氨基酸通过肽键缩合环化形成的，其结构具有一定的构象约束作用。环肽独特的拓扑学结构使得它们对化学、热和生物降解异常稳定。环肽中存在大量的氢键受体和给体，而氢键是药物与受体相互作用的主要方式之一，因此，环肽往往具有一定的生物和药理活性，成为药物化学中一个重要的药效团。此外，环肽的受限构象使其表现出较线性肽更好的特异性和靶向亲和力，因此，环肽表现出广泛的生物和药理活性，如抗肿瘤、抗病毒、抗菌、抗衰老、免疫调节、增强记忆力、血糖调节等。
然而，环肽分子显著的刚性结构和极强的氢键形成能力，导致其存在水溶性差、易结晶和生物利用度低的缺陷，严重影响其药用活性。目前，国内外研究者采取了多种制剂学手段针对上述缺点进行改善，包括对环肽进行直接化学修饰改性(如N-甲基化修饰)或纳米载体包封(如脂质或胶束)在内的几种策略。然而，复杂的合成工艺、潜在的毒性以及环肽装载量低等问题仍然是不可避免的。
相比于热力学上更加稳定的晶体，玻璃态是一种亚稳态的无定形结构，其能够保留环肽分子的生物和药理活性。玻璃态环肽药物与晶态环肽药物相比，表现出高表面自由能及高分散度，能有效提高环肽的溶出速率和生物利用度。
特别需要说明的是，本发明的环肽玻璃可完全替代传统药用辅料，实现分散、增溶、粘合、控释等功能。特别是，可作为蓖麻油类药用辅料的替代品，有效避免组胺引发的致严重过敏反应。
目前尚未公开报道环肽玻璃，特别是尚未公开基于具有生物/药理活性和缓释功能的药物组合物玻璃、及将难溶性环肽转化为玻璃态的方法。
出人意料的，本发明发现环肽及其衍生物通过特定的制备工艺，可被加工成环肽玻璃、及环肽药物组合物玻璃。本发明同时发现了一种将难溶性环肽药物转化为玻璃态的方法，有效的增加了环肽药物的溶出速率和生物利用度。
本发明正是基于这一发现得以完成。基于本发明所发现的环肽玻璃，其有望作为活性药物或药用辅料，广泛应用于抗肿瘤、抗病毒/抗菌、血糖控制、免疫调节、神经调节等的药物递送及缓释领域。
发明内容：
本发明的首要目的在于提供一种环肽玻璃、及环肽药物组合物玻璃。及将难溶性环肽转化为玻璃态的方法。在本发明中，所述玻璃是指呈现玻璃化转变现象的非晶态固体，所述玻璃态是指保持类玻璃特性的无序组织结构，在X-射线衍射检测中无晶型结构，并且存在确定的玻璃化转变温度。即，本发明提供一种将难溶性环肽从晶态到玻璃态转变方法，有效改善难溶性环肽的体外溶出速率和溶解度，提高难溶性环肽药物的生物利用度。
第一方面：基于环肽的玻璃，其特征在于，所述环肽为形如结构式1的环肽及/或其盐，所述环肽是一种或两种以上的组合，
Figure PCTCN2021129065-appb-000001
所述A 1-A n独立地选自：
甘氨酸、丙氨酸、缬氨酸、亮氨酸、异亮氨酸、甲硫氨酸(蛋氨酸)、脯氨酸、色氨酸、丝氨酸、酪氨酸、半胱氨酸、苯丙氨酸、天冬酰胺、谷氨酰胺、苏 氨酸、天冬氨酸、谷氨酸、赖氨酸、精氨酸、组氨酸、硒半胱氨酸和吡咯赖氨酸中的一种或者多种的组合；
R 1-R n独立地选自H或其它可修饰基团，可修饰基团优选为甲基、烷基、磷酸、乙酰、甲酰、脂肪酸、苯甲酰、酰胺、酯、9-芴基甲氧基羰基、叔丁氧羰基；
n≥2，优选地，2≤n≤15，A 1-A n通过氨基酸缩合连接。
优选地，所述的环肽具有生物和/或药理活性；再优选地，具有抗菌/抗病毒、抗肿瘤、血糖调节、免疫调节活性，其中，
抗菌/抗病毒环肽包括但不限于下述结构及其类似结构：
Figure PCTCN2021129065-appb-000002
Figure PCTCN2021129065-appb-000003
Figure PCTCN2021129065-appb-000004
抗肿瘤环肽包括但不限于下述结构及其类似结构：
Figure PCTCN2021129065-appb-000005
Figure PCTCN2021129065-appb-000006
免疫调节环肽包括但不限于下述结构及其类似结构：
Figure PCTCN2021129065-appb-000007
血糖调节环肽包括但不限于下述结构及其类似结构：
Figure PCTCN2021129065-appb-000008
心血管及血液相关环肽包括但不限于下述结构及其类似结构：
Figure PCTCN2021129065-appb-000009
其他活性环肽包括但不限于下述结构及其类似结构：
Figure PCTCN2021129065-appb-000010
Figure PCTCN2021129065-appb-000011
优选地，所述的环肽衍生物为与上述环肽分子结构骨架类似的分子、异构体及其盐。
优选地，所述环肽为水溶性差的环肽，所述水溶性差是指在常温常压下环肽在纯水中的最大溶解浓度≤5wt％。
第二方面，基于环肽的玻璃，其特征在于，其完全由环肽及盐制备而得；或者，在玻璃中还加入其它药用辅料和/或药物活性成分。
该药用组合物玻璃由以下一种或多种成分组成：环肽或环肽衍生物、其它药物活性成分或其可药用盐、可药用的辅料；优选地，至少含有一种活性环肽；
药物活性成分包括具有抗菌/抗病毒、抗肿瘤、血糖调节、免疫调节、抗精神病等功效的非环肽类药物分子中的一种或两种以上混合物，优选地，抗菌/抗病毒药物为青霉素、头孢氨苄、阿米卡星、诺氟沙星、呋喃妥因、甲硝唑、金刚烷胺、阿昔洛韦、齐多夫定、利巴韦林；抗肿瘤药物为顺铂、阿霉素、长春新碱、紫杉醇、多烯紫杉醇、吉西他滨、喜树碱、羟基喜树碱、伊立替康、依托泊苷、地塞米松、氟尿嘧啶、环磷酰胺；血糖调节药物为二甲双胍、瑞格列奈、那格列奈、胰岛素；免疫调节药物为胸腺五肽、雷公藤多苷、雷公藤甲素、他克莫司、干扰素、云芝多糖；抗精神病药物为氯丙嗪、氟哌啶醇、利培酮、帕利哌酮、齐拉西酮；
药用辅料包括填充剂、湿润剂、粘合剂、崩解剂、润滑剂及其它，优选地，填充剂为淀粉、乳糖、甘露醇；湿润剂/粘合剂为羧甲基纤维素钠、羟丙基纤维素；崩解剂为羧甲基淀粉钠、低取代羟丙基纤维素、交联聚乙烯吡咯烷酮、交联羧甲基纤维素钠；润滑剂为聚乙二醇、硬脂酸镁、氢化植物油；其它为十二烷基硫酸钠；
环肽的质量比在1-100％之间，优选地，为1％-50％；
其它药物活性成分的质量比在1-20％之间，优选地，为1-10％；
可药用辅料的质量比在0-50％之间，优选地，为0-5％，再优选地，为0％，即环肽玻璃完全替代传统药用辅料。
第三方面，基于环肽的玻璃，其特征在于，可作为药用辅料、可完全或部分替代药用辅料。
第四方面，一种将难溶性环肽转化为玻璃态，制备环肽玻璃药物或环肽玻璃辅料的方法，其特征在于，主要包括如下步骤：
(1)将一种或多种难溶性环肽充分球磨处理，控制球磨过程中原料温度为0-50℃，优选地，为10-30℃；
(2)通过“升温-淬火”方法，具体地，将充分研磨的环肽原料在惰性气体氛围下升温到熔点温度附近，保温处理一段时间后，转移至退火炉中进行退火处理，即制备得到环肽玻璃药物或环肽玻璃辅料；
其特征在于，
升温温度是指熔点温度(T m)±50～250K的温度，优选地，为高于T m 50～100K；
保温时间为0min～30h，优选地，为15～30min；
退火温度是指玻璃化转变温度(T g)±50～150K的温度，优选地，为低于T g 50～100K；
退火处理时间为30min～2h，优选地，为30min～1h；
T m及T g通过热重分析及差式扫描量热方法测得，升温及降温速率为2-50Kmin -1。
第五方面，一种难溶性环肽药用组合物玻璃的制备方法，其特征在于，主要包括如下步骤：
(1)将难溶性环肽、其它药物活性成分混合后，充分球磨处理，控制球磨过程中原料温度为0-50℃，优选地，为10-30℃；
(2)遵循第四方面的步骤(2)；
(3)将步骤(2)得到的玻璃与可药用辅料共制成药用组合物，方法为压片、湿法制粒、流化床制粒、包衣、喷雾制粒、程序浇注、3D打印中的一种或两种以上联用。
或通过如下制备方法，其特征在于，主要包括如下步骤：
(1)将难溶性环肽、其它药物活性成分混合后，充分球磨处理，控制球磨过程中原料温度为0-50℃，优选地，为10-30℃；
(2)将步骤(1)的粉末在惰性气体氛围下升温，保温处理一段时间，升温温度为熔点温度(T m)±50～250K的温度，优选地，为高于T m 50～100K；保温时间为0min～30h，优选地，为15～30min；
(3)将其它药物活性成分，置于良溶剂及助溶剂中完全溶解，所述的良溶剂优选为乙醇、六氟异丙醇、二氯甲烷、四氯化碳、丙酮、乙酸乙酯，助溶剂优选为苯甲酸钠、二甲基乙酰胺、尿素、吐温中的一种或两种以上混合物；
(4)将步骤(3)得到的药物活性成分溶液，与步骤(1)熔融态环肽混合均匀；
(5)将步骤(4)得到的混合物置于步骤(2)所设置的温度下，减压旋转蒸发溶剂；
(6)将步骤(5)得到的混合物转移至退火炉中进行退火处理，退火温度为玻璃化转变温度(T g)±50～150K的温度，优选地，为低于T g 50～100K；退火处理时间为30min～2h，优选地，为30min～1h；
(7)将步骤(6)得到的玻璃与可药用辅料共制成药用组合物，方法为压片、湿法制粒、流化床制粒、包衣、喷雾制粒、程序浇注、3D打印中的一种或两种以上联用。
或通过如下制备方法，其特征在于，主要包括如下步骤：
(1)将难溶性环肽及其它药物活性成分共同置于良溶剂及助溶剂中完全溶解，所述的良溶剂优选为乙醇、六氟异丙醇、二氯甲烷、四氯化碳、丙酮、乙酸乙酯，助溶剂优选为苯甲酸钠、二甲基乙酰胺、尿素、吐温中的一种或两种以上混合物；
(2)将步骤(1)得到的混合溶液在惰性气体氛围下升温，减压旋转蒸发溶剂，后保温处理，升温温度为熔点温度(T m)±50～250K的温度，优选地，为高于T m 50～100K；保温时间为0min～30h，优选地，为15～30min；
(3)将步骤(2)得到的混合物转移至退火炉中进行退火处理，退火温度为玻璃化转变温度(T g)±50～150K的温度，优选地，为低于T g 50～100K；退火处理时间为30min～2h，优选地，为30min～1h；
(4)将步骤(3)得到的玻璃与可药用辅料共制成药用组合物，方法为压片、湿法制粒、流化床制粒、包衣、喷雾制粒、程序浇注、3D打印中的一种或两种以上联用。
第六方面，根据第一至五方面所述的环肽玻璃药物、环肽玻璃辅料、环肽药用组合物玻璃及制备方法，所述的玻璃药物剂型可为口服剂、贴剂、皮下包埋剂、支架材料及微针器件。
第七方面，根据第六方面所述的药物剂型，涉及到的工艺为压片、干法制粒、高剪切湿法制粒、流化床制粒、胶囊罐装、微胶囊包埋、包衣、喷雾干燥、喷雾凝结、光刻、程序浇注、微针阵列、3D打印中的一种或两种以上。
第八方面，本发明的环肽玻璃、及环肽药物组合物玻璃、及环肽药物玻璃态剂型具有如下优点及有益效果：
(1)可同时发挥药效及药物辅料功能；
(2)可有效增加药物溶出速率、提高药物生物利用度，降低药物副作用；
(3)具有良好的生物相容性和可生物降解性；
(4)具有较高的热/化学稳定性。
第九方面，本发明的环肽玻璃、及环肽药物组合物玻璃、及环肽药物玻璃态剂型有以下应用：抗肿瘤、抗病毒/抗菌、血糖控制、免疫调节、神经调节等的药物递送领域，特别是实现药物缓释治疗。
本发明的环肽玻璃的原料包括环肽，特别是具有生物/药理活性的环肽或其药学上可接受的衍生物及其盐类中的任意一种或者多种的组合，一般表现为水溶性差、易结晶、生物利用度低。本发明创造性的提出将难溶性环肽药物转化为玻璃态的方法，包括将难溶性环肽制备成玻璃，及将难溶性环肽与其它活性药物成 分共制成玻璃的方法。本发明提供了一种将难溶性药物转化为玻璃态，形成药物玻璃态剂型，实现难溶性活性环肽及药物缓释的方法。所得的环肽玻璃具有高生物相容性、可降解性及高热/化学稳定性，可进一步经喷雾干燥、固体分散、增材制造等技术加工成口服剂、贴剂、皮下包埋剂、支架材料及微针器件。所得环肽玻璃可同时发挥药效及药物辅料功能，与晶体及传统药物剂型或辅料相比，环肽玻璃可有效增加药物溶出速率、提高药物生物利用度，在抗肿瘤、抗病毒/抗菌、血糖控制、免疫调节、神经调节等的药物递送及缓释领域应用广泛。
附图说明
图1为实施例1所制备的CsA玻璃在室温下的实物图(a)及经浇筑工艺得到的CsA玻璃实物图(b)，证实环肽玻璃的可加工性。
图2为实施例1所制备的CsA玻璃的核磁氢谱，对比于CsA原料，峰未发生明显变化，说明环肽原料分子经加热熔融及退火处理，化学成分并未发生改变。
图3为实施例1所制备的CsA玻璃的差式扫描量热(DSC)图，证实其玻璃态结构，其玻璃化转变温度为T g＝100.4K。
图4为实施例1所制备的CsA玻璃的X-射线衍射(XRD)图谱，表明CsA玻璃为非晶态结构。
图5为实施例1所制备的CsA玻璃的稳定性测试结果，证实其高的稳定性能。
图6为实施例1所制备的CsA玻璃的药物缓释曲线，证实可实现药物的缓慢释放。
图7为实施例1所制备的CsA玻璃经口服给药后，小鼠的免疫相关指标。
图8为实施例2基于CFP和肿瘤化疗药物PTX的药用组合物玻璃的XRD谱图，为非晶态结构。
图9为实施例2所制备的药用组合物玻璃胶囊在人工胃液(遵循中国药典制备方法配置)中的降解情况，证实其可降解性质。
图10为实施例2所制备的药用组合物玻璃胶囊经灌胃后，在小鼠体内随时间的药物-时间曲线。
图11为实施例2所制备的药用组合物玻璃胶囊经口服给药后，实验小鼠的肿瘤抑制曲线，证实具有抗肿瘤效果。
图12为实施例2所制备的药用组合物玻璃胶囊经口服给药后，实验小鼠的体重变化曲线，证实高的生物安全性。
图13为实施例2所制备的玻璃药用组合物胶囊经口服给药后，实验小鼠的脏器系数变化，证实高的生物安全性。
图14为实施例3药用组合物玻璃的熔融图片及XRD数据，证实由晶体转化为玻璃态。
图15为实施例3所制备的药用组合物玻璃的生物相容性测试结果，证实具有高的生物相容性。
图16为实施例3所制备的药用组合物玻璃涂层，抑制细菌生长的曲线。
图17为实施例3所制备的药用组合物玻璃药片，对大肠杆菌和金黄色葡萄球菌的抑菌效果。
图18为实施例3所制备的玻璃药用组合物的包埋剂，皮下包埋后随时间的降解曲线。
图19为实施例4所制备的药用组合物玻璃的片剂，对自发性II型糖尿病小鼠血糖的调控曲线，证实可有效控制小鼠血糖。
图20为实施例4所制备的药用组合物玻璃的片剂，对自发性II型糖尿病小鼠进行灌胃给药，小鼠体重随时间的变化曲线，证实可有效控制小鼠体重。
图21为实施例4所制备的药用组合物玻璃的片剂，对自发性II型糖尿病小鼠进行灌胃给药30天后，口服高浓度葡萄糖的耐受情况，葡萄糖浓度为2.5g kg -1，通过测量小鼠足底血的血糖浓度绘制曲线，证实实验组小鼠具有更好的葡萄糖耐受能力。
图22为实施例5所制备的CPY-CFP-CLP环肽混合玻璃的DSC曲线，得到单一及混合玻璃的玻璃化转变温度，表明多种环肽混合后可有效抑制单一环肽的结晶倾向。
图23为实施例5所制备的环肽混合玻璃和CLP晶体释放CLP分子的曲线，表明环肽混合玻璃可提升CLP活性环肽的溶出速率。
图24为实施例5所制备的药用组合物玻璃的XRD图谱，为非晶态。
图25为旷场实验中，小鼠的自主活动能力和焦虑样行为统计数据。结果表明，与正常小鼠相比，早期社会隔离小鼠的中心区域活动距离明显减小，玻璃微针贴片进行治疗后有明显的改善。
图26为迷宫实验中，小鼠5min内进入开臂和闭臂的次数和在两臂滞留时间。结果表明，与正常小鼠相比，早期社会隔离小鼠进入开臂的次数明显减少且在开臂中的滞留时间减少，经玻璃微针贴片进行治疗后有明显的改善。
具体实施方式
下面通过实施例对本发明的技术方案进行详细说明，但本发明所保护内容不仅限于此。
实施例1
一种基于环孢菌素A(CsA)的环肽玻璃的制备方法包括如下步骤：
(1)将CsA粉末置于球磨机中研磨均匀，控制球磨机恒温为25℃，后转移至坩埚中；
(2)将步骤(1)中装有CsA粉末的坩埚在N 2氛围下放置在加热设备内；
(3)对步骤(2)的设备进行加热处理，以10K min -1的升温速率将坩埚由室温升温至573.15K温度，在此温度下保温处理20min时间；
(4)对步骤(3)的设备进行降温处理，以10K min -1的降温速率将坩埚降至273.15K的退火炉中恒温30min，进行玻璃的退火处理，即得到CsA玻璃。
图1为实施例1所制备的CsA玻璃在室温下的实物图(a)及经浇筑工艺得到的CsA玻璃实物图(b)，证实环肽玻璃的可加工性。
图2为实施例1所制备的CsA玻璃的核磁氢谱，对比于CsA原料，峰未发生明显变化，说明环肽原料分子经加热熔融及退火处理，化学成分并未发生改变。
图3为实施例1所制备的CsA玻璃的DSC图，证实其玻璃态结构，其玻璃化转变温度为T g＝100.4K。
图4为实施例1所制备的CsA玻璃的X-射线衍射(XRD)图谱，表明CsA玻璃为非晶态结构。
图5为实施例1所制备的CsA玻璃的稳定性测试结果，证实其高的稳定性能。
图6为实施例1所制备的CsA玻璃的药物缓释曲线，证实可实现药物的缓慢释放。具体地，CsA玻璃的初始质量为20±1mg，缓释溶剂为10mL磷酸盐缓冲液(0.01M，pH＝6.8)，搅拌速率为100r min -1，通过溶出度试验仪及高效液相色谱仪对溶出的CsA进行检测，绘制药物缓释曲线。
图7为实施例1所制备的CsA玻璃经口服给药后，小鼠的免疫相关指标。具体地，将健康雄性昆明白鼠(体重20±1g，4～5周龄)随机分为两组，每组 8只，正常饮食。实验组小鼠隔天灌胃给药，给药剂量为40mg kg -1，共给药次数5，对照组小鼠隔天灌胃等质量的生理盐水。末次口服给药后的次日对小鼠进行安乐死检测相应指标。解剖取得小鼠的胸腺及脾脏，用磷酸盐缓冲液(PBS)冲去表面血液，滤纸吸干水分，后置于电子天平上称重，将器官重量(mg)除以小鼠总重(g)计算得到器官重量指数。取小鼠外周血，与CD3 +(PE-Cy5)单抗、CD4 +(PE)单抗和CD8 +(APC)单抗混合摇匀后避光孵育，经细胞裂解、离心后经流式细胞仪进行分析。
实施例2
一种基于苯丙氨酸-脯氨酸环二肽(CFP)和紫杉醇(PTX)的药用组合物玻璃的制备方法包括如下步骤：
(1)将CFP粉末置于球磨机中研磨混合均匀，控制球磨机恒温为37℃，后转移至容器中；
(2)对步骤(1)的容器进行加热处理，以20K min -1的升温速率将容器由室温升温至473.15K温度，在此温度下保温处理30min时间；
(3)对步骤(2)的容器进行降温处理，以10K min -1的降温速率将容器降至333.15K温度；
(4)将PTX粉末置于球磨机中研磨均匀，控制球磨机恒温为37℃，后加入乙醇完全溶解，CFP与PTX的摩尔比为25：1；
(5)将步骤(4)的溶液滴加入步骤(3)的容器内，将容器置于减压旋转蒸发仪内，维持温度在333.15K，除去溶剂；
(6)对步骤(5)的设备进行降温处理，以10K min -1的降温速率将容器降至315.15K的退火炉中恒温30min，即得到基于CFP和肿瘤化疗药物PTX的药用组合物玻璃，将上述玻璃完全溶解于HFIP有机溶液中，通过高效液相色谱定量PTX的浓度，计算PTX的载药量为3.5±0.2％；
(7)经喷雾干燥及胶囊灌注的制备工艺，制备得到环肽玻璃口服剂型。
图8为实施例2基于CFP和肿瘤化疗药物PTX的药用组合物玻璃的XRD谱图，证明本发明的药物组合物为非晶态结构。
图9为实施例2所制备的药用组合物玻璃胶囊在人工胃液(遵循中国药典制备方法)中的降解情况，证实其可降解性质。药用组合物玻璃胶囊的制备步骤包括，将药物组合物玻璃经均质机研磨成粉末，将上述粉末灌装于明胶胶囊壳。
图10为实施例2所制备的药用组合物玻璃胶囊经灌胃后，在小鼠体内随时间的药物-时间曲线。具体地，小鼠给药剂量为8mg kg -1，灌胃后随时间采用眼眶取血方式采集小鼠外周血，进行细胞裂解及溶剂溶出，用高效液相色谱仪对过滤后的溶液进行检测，计算血液中PTX的含量，绘制药物-时间曲线。
图11为实施例2所制备的药用组合物玻璃胶囊经口服给药后，实验小鼠的肿瘤抑制曲线，证实具有抗肿瘤效果。具体地，将健康BALB/c nude裸鼠(体重15±1g，4～6周龄)随机分为两组，每组8只，采用腋窝皮下接种4T1细胞建立乳腺癌4T1小鼠移植瘤模型。待小鼠肿瘤增长至80±10mm 3体积后，对小鼠进行药物治疗。实验组小鼠隔天灌胃给药，给药剂量为8mg kg -1，共给药次数5，对照组小鼠隔天灌胃等质量的PBS。每日用游标卡尺测量并记录小鼠肿瘤体积变化，小鼠肿瘤体积按如下公示计算，V＝ab 2/2，其中a为肿瘤的长边距离，b为肿瘤短边距离。
图12为实施例2所制备的药用组合物玻璃胶囊经口服给药后，实验小鼠的体重变化曲线，证实高的生物安全性。
图13为实施例2所制备的药用组合物玻璃胶囊经口服给药后，实验小鼠的脏器系数变化，证实高的生物安全性。具体地，在第35天解剖取得小鼠的重要脏器(包括心脏、肝脏、脾脏、肺、肾脏)，用PBS冲去表面血液，滤纸吸干水分，后于电子天平上称重，将器官重量(mg)除以小鼠总重(g)计算得到脏器系数。
实施例3
一种基于万古霉素、短杆菌肽S和青霉素的药用组合物玻璃的制备方法包括如下步骤：
(1)将万古霉素、短杆菌肽S和青霉素置于球磨机中研磨混合均匀，控制球磨机恒温为37℃，后转移至容器中，三者的质量比为2：2：1；
(2)向步骤(1)的混合粉末中加入六氟异丙醇、乙醇的混合有机溶剂，搅拌至完全溶解；
(3)将步骤(2)得到的混合溶液置于减压旋转蒸发仪内，以50K min -1的升温速率由室温升温至443.15K温度，在此温度下保温处理30min时间，去除有机溶剂；
(4)对步骤(3)的设备进行降温处理，以10K min -1的降温速率降至243.15K温度，并在该温度下恒温20min；
(5)向步骤(4)得到的产物中加入一定质量的可药用辅料，包括1.5份淀粉、1份羧甲基纤维素钠、2.5份聚乙二醇和6.5份氢化植物油，辅料的总质量比为92.5％；
(6)将步骤(5)得到的药物组合物经均质处理，联合压片、真空喷雾干燥技术制备成玻璃药用组合物的包埋剂。
图14为实施例3药用组合物玻璃的熔融图片及XRD数据，证实由晶体转化为玻璃态。
图15为实施例3所制备的药用组合物玻璃的生物相容性测试结果，证实具有高的生物相容性。具体地，将上述玻璃经涂膜机加工成直径60mm、厚度0.2mm的细胞培养皿涂层，用于细胞(成纤维细胞3T3)培养，共孵育48h后，采用MTT法测试细胞的存活率。
图16为实施例3所制备的药用组合物玻璃涂层，抑制细菌生长的曲线。具体地，将上述药用组合物玻璃加工成1cm*1cm*0.1cm的药片，贴于细菌培养皿中央，药物总浓度经计算为10ug/皿，将大肠杆菌和金黄色葡萄球菌在液体培养基中混合均匀后，滴加入细菌培养皿内，37℃震荡孵育，随时间采用OD值法对细菌培养皿中的细菌浓度进行测量。
图17为实施例3所制备的药用组合物玻璃药片，对大肠杆菌和金黄色葡萄球菌的抑菌效果。具体地，将上述药用组合物玻璃加工成直径为5mm，厚度为0.2mm药片，牛津杯(直径为6mm)放置于琼脂培养皿，将药片置于牛津杯内，与琼脂贴合紧密。将大肠杆菌及金黄色葡萄球菌经PBS混合均匀后，涂布于培养皿内，置于37℃培养箱中培养，24h后通过观察抑菌环的产生与否，判断药用组合物玻璃的抑菌效果。
图18为实施例3所制备的药用组合物玻璃的包埋剂，皮下包埋后随时间的降解曲线。具体的，将健康雄性昆明白鼠(体重20±1g，4～5周龄)随机分为两组，每组15只。在实验组小鼠的背部手术划开1cm窗口，将药用组合物玻璃30±2.5g埋于皮下，后缝合。随时间取任意一只小鼠，取出药用组合物玻璃并称量剩余质量，绘制降解曲线。
图19为实施例4所制备的药用组合物玻璃的片剂，对自发性II型糖尿病小鼠血糖的调控曲线，证实可有效控制小鼠血糖。具体地，将订购的自发性II型糖尿病模型小鼠(雄性、8周龄)分为两组，每组6只。实验组小鼠每日灌胃药用组合物玻璃，给药剂量为4mg kg -1，对照组小鼠灌胃等质量的生理盐水，测试周期为30天。每日取小鼠尾部血，用血糖测试仪测定小鼠的空腹血糖值。横坐标为时间，纵坐标为实验小鼠的空腹血糖值绘制曲线。
图20为实施例4所制备的药用组合物玻璃的片剂，对自发性II型糖尿病小鼠进行灌胃给药，小鼠体重随时间的变化曲线，证实可有效控制小鼠体重。
图21为实施例4所制备的药用组合物玻璃的片剂，对自发性II型糖尿病小鼠进行灌胃给药30天后，口服高浓度葡萄糖的耐受情况，葡萄糖浓度为2.5g kg -1，通过测量小鼠足底血的血糖浓度绘制曲线，证实实验组小鼠具有更好的葡萄糖耐受能力。
图22为实施例5所制备的CPY-CFP-CLP环肽混合玻璃的DSC曲线，得到单一及混合玻璃的玻璃化转变温度，表明多种环肽混合后可有效抑制单一环肽的结晶倾向。
图23为实施例5所制备的环肽混合玻璃和CLP晶体释放CLP分子的曲线，表明环肽混合玻璃可提升CLP活性环肽的溶出速率。具体地，环肽混合玻璃的初始质量为15±1mg，缓释溶剂为10mL PBS(0.01M，pH＝6.8)，搅拌速率为100r min -1，通过溶出度试验仪及高效液相色谱仪对溶出的CLP进行检测，绘制环肽混合玻璃的缓释曲线。作为对照，称取同等质量的CLP晶体置于上述缓释溶液中搅拌孵育，检测CLP在缓释液中的浓度。
图24为实施例6所制备的药用组合物玻璃的XRD图谱，为非晶态。
图25为实施例6所制备的药物组合物玻璃应用于小动物旷场实验中，小鼠的自主活动能力和焦虑样行为统计数据。结果表明，与正常小鼠相比，早期社会 隔离小鼠的中心区域活动距离明显减小，玻璃微针贴片进行治疗后有明显的改善。具体地，将C57BL/6早期社会隔离小鼠(8周龄，雄性)平均分为2组，每组6只。其中一组不作特殊处理，另一组小鼠在背部脱毛后粘贴玻璃微针贴片。C57BL/6群养正常小鼠6只作为空白对照组。按照标准，准备黑色矿场装置(45cm*45cm*45cm)，将小鼠置于矿场中，追踪并记录小鼠的自主活动行为。纵坐标为小鼠在矿场中运动1h的中心区域活动距离统计值。
图26为实施例6所制备的药物组合物玻璃应用于小鼠迷宫实验中，小鼠5min内进入开臂和闭臂的次数和在两臂滞留时间。结果表明，与正常小鼠相比，早期社会隔离小鼠进入开臂的次数明显减少且在开臂中的滞留时间减少，经微针贴片进行治疗后有明显的改善。具体地，按照标准，准备迷宫装置，臂宽5cm，臂长35cm，闭臂高15cm，中央平台5cm*5cm，迷宫离地高度约40-55cm左右。将小鼠置于平台中央，记录小鼠进入开臂和闭臂的次数和在两臂滞留时间，评价焦虑的指标。
实施例4
一种基于组氨酸-脯氨酸环二肽和司格列肽的药用组合物玻璃的制备方法包括如下步骤：
(1)将组氨酸-脯氨酸环二肽和司格列肽置于球磨机中研磨混合均匀，控制球磨机恒温为25℃，后转移至容器中，二者的质量比为10：1；
(2)向步骤(1)的混合粉末中加入二氯甲烷溶剂，搅拌至完全溶解；
(3)将步骤(2)得到的混合溶液置于减压旋转蒸发仪内，以10K min -1的升温速率由室温升温至535.15K温度，在此温度下保温处理30min时间，去除有机溶剂；
(4)对步骤(3)的设备进行降温处理，以10K min -1的降温速率降至283.15K温度，并在该温度下恒温1h，得到基于组氨酸-脯氨酸环二肽和司格列肽的药用组合物玻璃；
(5)将步骤(5)得到的药物组合物玻璃经均质处理后，经压片及程序浇注制备成片剂，用于口服给药血糖调控。
实施例5
一种基于脯氨酸-酪氨酸环二肽(CPY)、苯丙氨酸-脯氨酸环二肽(CFP)和亮氨酸-脯氨酸环二肽(CLP)混合玻璃的制备方法包括如下步骤：
(1)称取等摩尔比的CPY、CFP、CLP粉末，分别溶于六氟异丙醇中形成单一环肽溶液；
(2)将步骤(1)得到的混合溶液置于减压旋转蒸发仪内，以10K min -1的升温速率由室温升温至473.15K，在此温度下保温处理30min时间，去除有机溶剂；
(3)对步骤(2)的设备进行降温处理，以10K min -1的降温速率降至243.15K温度，并在该温度下恒温30min，即得到CPY-CFP-CLP环肽混合玻璃。
实施例6
一种基于色氨酸-色氨酸环二肽(CWW)和利培酮的药用组合物玻璃制备方法包括如下步骤：
(1)称取等摩尔比的CWW、利培酮粉末，分别溶于六氟异丙醇和甲醇中形成单分散溶液；
(2)将步骤(1)得到的混合溶液置于减压旋转蒸发仪内，以10K min -1的升温速率由室温升温至573.15K，在此温度下保温处理30min时间，去除有机溶剂；
(3)对步骤(2)的设备进行降温处理，以50K min -1的降温速率降至283.15K温度，并在该温度下恒温30min，即得到基于CWW和帕利哌酮的玻璃药用组合物；
(4)基于3D打印工艺及浇筑工艺，将步骤(3)得到的玻璃药物组合物加工为微针贴片，贴片尺寸为0.8*0.8cm，用于经皮药物递送。建立C57BL/6的早期社会隔离小鼠模型，采用皮下微针贴片进行治疗，利培酮剂量为5mg kg -1，14天后，评估小鼠的精神分裂样行为。

Claims (10)

  1. 一种基于环肽的玻璃，其特征在于，所述环肽为形如结构式1的环肽及其盐，所述环肽为一种或两种以上的组合，
    Figure PCTCN2021129065-appb-100001
    其中，A 1-A n独立地选自：
    甘氨酸、丙氨酸、缬氨酸、亮氨酸、异亮氨酸、甲硫氨酸(蛋氨酸)、脯氨酸、色氨酸、丝氨酸、酪氨酸、半胱氨酸、苯丙氨酸、天冬酰胺、谷氨酰胺、苏氨酸、天冬氨酸、谷氨酸、赖氨酸、精氨酸、组氨酸、硒半胱氨酸和吡咯赖氨酸；
    R 1-R n独立地选自H或其它可修饰基团，可修饰基团独立地选自甲基、烷基、磷酸、乙酰、甲酰、脂肪酸、苯甲酰、酰胺、酯、9-芴基甲氧基羰基、叔丁氧羰基；
    n≥2，优选地，2≤n≤15，A 1-A n通过氨基酸缩合连接；
    优选地，所述环肽具有生物和/或药理活性；进一步优选地，所述环肽具有抗菌/抗病毒、抗肿瘤、血糖调节、免疫调节活性。
  2. 根据权利要求1所述的基于环肽的玻璃，其中，
    所述抗菌/抗病毒环肽选自如下环肽中的一种或者多种的组合：
    Figure PCTCN2021129065-appb-100002
    Figure PCTCN2021129065-appb-100003
    Figure PCTCN2021129065-appb-100004
    所述抗肿瘤环肽选自如下环肽中的一种或者多种的组合：
    Figure PCTCN2021129065-appb-100005
    Figure PCTCN2021129065-appb-100006
    Figure PCTCN2021129065-appb-100007
    所述免疫调节环肽选自如下环肽中的一种或者多种的组合：
    Figure PCTCN2021129065-appb-100008
    所述血糖调节环肽选自如下环肽中的一种或者多种的组合：
    Figure PCTCN2021129065-appb-100009
    所述心血管及血液相关环肽选自如下环肽中的一种或者多种的组合：
    Figure PCTCN2021129065-appb-100010
    所述其他活性环肽选自如下环肽中的一种或者多种的组合：
    Figure PCTCN2021129065-appb-100011
  3. 根据权利要求1所述的基于环肽的玻璃，其特征在于所述环肽为水难溶性环肽，也包含水难溶性环肽的肽链骨架经过修饰后的水溶性环肽衍生物。
  4. 一种含有权利要求1-3任意一项所述玻璃的药用组合物，其特征在于，所述药用组合物为玻璃形态，所述组合物完全由环肽制备而得或者还含有药用辅料和/或药物活性成分。
  5. 制备权利要求1-3任意一项所述环肽玻璃的方法，其特征在于，包括如下步骤：
    (1)将一种或多种环肽进行球磨处理，控制球磨过程中原料温度为0-50℃，优选地，为10-30℃；
    (2)通过“升温-淬火”方法制备环肽玻璃，将研磨的环肽原料在惰性气体氛围下升温到熔点温度附近，保温处理一段时间后，转移至退火炉中进行退火处理。
  6. 根据权利要求5所述的方法，其特征在于：
    升温温度是指熔点温度(T m)±50～250K的温度，优选地，为高于T m50～100K；
    保温时间为0min～30h，优选地，为15～30min；
    退火温度是指玻璃化转变温度(T g)±50～150K的温度，优选地，为低于T g50～100K；
    退火处理时间为30min～2h，优选地，为30min～1h；
    T m及T g通过热重分析及差式扫描量热方法测得，升温及降温速率为2-50K min -1。
  7. 权利要求4所述的药物组合物的制备方法，其特征在于，包括如下步骤：
    (1)将环肽与其它药物活性成分混合后，进行球磨处理，控制球磨过程中原料温度为0-50℃，优选地，为10-30℃；
    (2)通过“升温-淬火”方法制备环肽玻璃，将研磨的环肽原料在惰性气体氛围下升温到熔点温度附近，保温处理一段时间后，转移至退火炉中进行退火处理得到玻璃；
    (3)任选地，将步骤(2)得到的玻璃与可药用辅料共制成药用组合物。
  8. 权利要求4所述的药物组合物的制备方法，其特征在于，包括如下步骤：
    (1)将一种或多种环肽进行球磨处理，控制球磨过程中原料温度为0-50℃，优选地，为10-30℃；
    (2)将步骤(1)的粉末在惰性气体氛围下升温，保温处理一段时间，升温温度为熔点温度(T m)±50～250K的温度，优选地，为高于T m50～100K；保温时间为0min～30h，优选地，为15～30min；
    (3)将其它药物活性成分，置于良溶剂及助溶剂中完全溶解；
    (4)将步骤(3)得到的药物活性成分溶液，与步骤(1)熔融态环肽混合均匀；
    (5)将步骤(4)得到的混合物置于步骤(2)所设置的温度下，减压旋转蒸发溶剂；
    (6)将步骤(5)得到的混合物转移至退火炉中进行退火处理，退火温度为玻璃化转变温度(T g)±50～150K的温度，优选地，为低于T g50～100K；退火处理时间为30min～2h，优选地，为30min～1h；
    (7)任选地，将步骤(2)得到的玻璃与可药用辅料共制成药用组合物。
  9. 权利要求4所述的药物组合物的制备方法，其特征在于，包括如下步骤：
    (1)将环肽及其它药物活性成分共同置于良溶剂及助溶剂中完全溶解；
    (2)将步骤(1)得到的混合溶液在惰性气体氛围下升温，减压旋转蒸发溶剂，后保温处理，升温温度为熔点温度(T m)±50～250K的温度，优选地，为高于T m50～100K；保温时间为0min～30h，优选地，为15～30min；
    (6)将步骤(5)得到的混合物转移至退火炉中进行退火处理，退火温度为玻璃化转变温度(T g)±50～150K的温度，优选地，为低于T g50～100K；退火处理时间为30min～2h，优选地，为30min～1h；
    (7)任选地，将步骤(2)得到的玻璃与可药用辅料共制成药用组合物。
  10. 根据权利要求4所述的药物组合物，所述组合物玻璃形态的药物组合物进一步制备为口服剂、贴剂、皮下包埋剂、支架材料及微针器件。
PCT/CN2021/129065 2021-11-05 2021-11-05 环肽玻璃及含有环肽的药物组合物玻璃 WO2023077447A1 (zh)

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WO1996003978A1 (en) * 1994-08-04 1996-02-15 Quadrant Holdings Cambridge Limited Solid delivery systems for controlled release of molecules incorporated therein and methods of making same
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WO1996003978A1 (en) * 1994-08-04 1996-02-15 Quadrant Holdings Cambridge Limited Solid delivery systems for controlled release of molecules incorporated therein and methods of making same
WO1999001463A2 (en) * 1997-07-03 1999-01-14 Quadrant Holdings Cambridge Limited Modified glycosides, compositions comprised thereof and methods of use thereof
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