WO2022002186A1 - 用于修复黏膜损伤或皮肤创伤的多肽及其应用 - Google Patents

用于修复黏膜损伤或皮肤创伤的多肽及其应用 Download PDF

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WO2022002186A1
WO2022002186A1 PCT/CN2021/103943 CN2021103943W WO2022002186A1 WO 2022002186 A1 WO2022002186 A1 WO 2022002186A1 CN 2021103943 W CN2021103943 W CN 2021103943W WO 2022002186 A1 WO2022002186 A1 WO 2022002186A1
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pro
val
ala
compound
glu
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PCT/CN2021/103943
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English (en)
French (fr)
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耿福能
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四川好医生攀西药业有限责任公司
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Priority to IL299498A priority Critical patent/IL299498A/en
Priority to KR1020237002409A priority patent/KR20230031898A/ko
Priority to JP2022580465A priority patent/JP7531082B2/ja
Priority to CN202180044414.2A priority patent/CN116133673A/zh
Priority to AU2021302188A priority patent/AU2021302188C1/en
Priority to US18/013,344 priority patent/US20230295233A1/en
Priority to CA3184710A priority patent/CA3184710A1/en
Priority to EP21834686.4A priority patent/EP4177259A4/en
Publication of WO2022002186A1 publication Critical patent/WO2022002186A1/zh

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    • 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
    • A61K38/07—Tetrapeptides
    • 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
    • A61K38/08—Peptides having 5 to 11 amino acids
    • 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
    • A61K38/10—Peptides having 12 to 20 amino acids
    • 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
    • A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
    • A61P1/02—Stomatological preparations, e.g. drugs for caries, aphtae, periodontitis
    • 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
    • A61P1/04—Drugs for disorders of the alimentary tract or the digestive system for ulcers, gastritis or reflux esophagitis, e.g. antacids, inhibitors of acid secretion, mucosal protectants
    • 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
    • A61P1/14—Prodigestives, e.g. acids, enzymes, appetite stimulants, antidyspeptics, tonics, antiflatulents
    • 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
    • A61P17/02—Drugs for dermatological disorders for treating wounds, ulcers, burns, scars, keloids, or the like
    • 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
    • C07K5/10—Tetrapeptides
    • C07K5/1002—Tetrapeptides with the first amino acid being neutral
    • C07K5/1005—Tetrapeptides with the first amino acid being neutral and aliphatic
    • C07K5/1008—Tetrapeptides with the first amino acid being neutral and aliphatic the side chain containing 0 or 1 carbon atoms, i.e. Gly, Ala
    • 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
    • C07K5/101—Tetrapeptides with the first amino acid being neutral and aliphatic the side chain containing 2 to 4 carbon atoms, e.g. Val, Ile, Leu
    • 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
    • C07K5/1024—Tetrapeptides with the first amino acid being heterocyclic
    • 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
    • C07K7/04—Linear peptides containing only normal peptide links
    • C07K7/08—Linear peptides containing only normal peptide links having 12 to 20 amino acids
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides

Definitions

  • the present invention relates to a novel polypeptide for repairing mucosal injury or skin wound and its application.
  • the polypeptide of the present invention has no homology with known polypeptides, and can exist stably in vivo and in vitro. It has the functions of regulating stem cell proliferation and differentiation, and repairing mucosa The effect of injury or skin injury; and the use of the novel polypeptide to repair mucosal injury or skin wound by regulating the proliferation and differentiation of stem cells, and the use of preventing, alleviating or treating gastrointestinal diseases.
  • Skin trauma and/or mucosal damage are common pathological features of many diseases.
  • Skin trauma or skin injury refers to the damage of normal skin (tissue) caused by external injury factors such as surgery, external force, heat, current, chemical substances, low temperature and internal factors in the body such as local blood supply disorders.
  • Skin damage is often accompanied by disruption of skin integrity and loss of a certain amount of normal tissue, while at the same time, the normal function of the skin is impaired. Also called a wound or trauma.
  • protein/polypeptide drugs including basic fibroblast growth factor, epidermal growth factor, platelet growth factor, granulocyte-macrophage colony-stimulating factor and growth hormone that have obvious effects on wound repair, skin care, anti-wrinkle and anti-aging, but these
  • the long amino acid sequence of protein/polypeptide drugs leads to the disadvantages of high preparation cost and poor stability, so their application is limited to a certain extent.
  • Human mucosa refers to the inner layer of cavities or cystic muscular organs such as respiratory system, digestive system, genitourinary system, etc. It is the second largest barrier of human body after skin, including oral cavity, pharynx, trachea, esophagus, stomach and intestines. , vagina, bladder, etc.
  • the wall or cyst wall of these organs has a common stratification law, and has the characteristics of adapting to its function, and its embryonic origin, tissue structure, pathological process, clinical manifestations, prognosis, etc. have the same characteristics.
  • Chronic gastritis is a chronic inflammation of the gastric mucosa, which is a common and frequently-occurring disease in the department of gastroenterology. Atrophic lesions of the glands are called chronic gastritis.
  • Chronic atrophic gastrifis (CAG), characterized by gastric mucosal epithelial and gland atrophy, has an insidious onset, a long course of disease, nonspecific symptoms, and is difficult to cure, accounting for 11% to 31% of chronic gastritis. It usually occurs in middle-aged and elderly people. The onset is related to age but not gender. The onset is slow and the disease is lingering, protracted and difficult to cure, and the treatment is difficult.
  • Chronic atrophic gastritis is an important stage in the occurrence and development of gastric cancer and is regarded as a precancerous lesion of gastric cancer.
  • WHO World Health Organization
  • Precancerous lesions such as typical hyperplasia stage. Especially those with diffuse intestinal metaplasia or atypical hyperplasia are more likely to become cancerous.
  • Most professionals in the industry agree that chronic superficial gastritis, atrophic gastritis, intestinal metaplasia, dysplasia, intestinal type Such a developmental model of gastric cancer.
  • CAG is an important stage in the transformation of chronic gastritis into gastric cancer. Active treatment of CAG has far-reaching significance in preventing its carcinogenesis and reducing the incidence of gastric cancer. Finding an effective treatment for atrophic gastritis is one of the important measures to better prevent gastric cancer.
  • the damage of mucosal tissue can clinically lead to gastrointestinal diseases such as chronic gastritis and peptic ulcer.
  • the repair of mucosal epithelium has two different mechanisms: restitution and regeneration (Cur. Med. Chem., 2008, 15, 3133-3144): repair or recovery generally begins within minutes after injury , rapid repair of superficial lesions through cell migration; regeneration is continuous regeneration through the differentiation and proliferation of stem and progenitor cells for days to months.
  • Epidermal growth factor is a polypeptide consisting of 53 amino acid residues, which is widely present in various tissues, organs and body fluids. It can promote the proliferation of epithelial cells and protect the skin. Epidermal growth factor mainly promotes the proliferation and growth of skin tissue cells, so that new cells can replace aging cells, so as to play anti-aging and skin care and health care functions. Epidermal growth factor has been reported to have the effect of repairing wounds. When skin wounds need to be disinfected and debridement, disinfectants containing iodine or hydrogen peroxide will be applied. EGF is unstable under these conditions. Growth factors are related to the healing of gastrointestinal anastomosis (J.Surgical Res.2014; 17:202-210), but when EGF is orally administered orally, it will be degraded after entering the body, and the therapeutic effect cannot be achieved.
  • the purpose of the present invention is to provide a new type of polypeptide.
  • the present invention provides a compound of formula (I), or a physiologically compatible salt thereof, wherein said compound of formula (I) is as follows:
  • Xaa 1 is Pro, Gly, Ala or missing;
  • Xaa 2 is Ala, Leu, Ile, Gly, Cys, Ser or missing;
  • Xaa 3 is Ala, Pro, Gly, Leu, Ile, Cys, Ser or missing;
  • Xaa 4 is Glu, Gln, Asp, Asn, Leu, Ile, Val or missing;
  • Xaa 5 is Pro, Gly, Ala, Val or missing;
  • Xaa 6 is Pro, Gly or Ala
  • Xaa 7 is Leu, Phe, Val, Ala, Tyr, Glu, Lys, Asp, Ile, Met or missing;
  • Xaa 8 is Val, Leu, Gln, Ile, Met or missing;
  • Xaa 9 is Lys, Arg, His, Asp, Val or missing;
  • Xaa 10 is Glu, Gln, Asp, Asn or missing;
  • Xaa 11 is Glu, Asp, Asn, Gln or missing
  • X aa5 is Pro.
  • X aa6 is Pro.
  • X aa7 is Leu.
  • the compound of formula (I) has the following structure of formula (II),
  • Xaa 1 is Pro, Gly, Ala or missing;
  • Xaa 2 is Ala, Leu, Ile, Gly, Cys, Ser or missing;
  • Xaa 3 is Ala, Pro, Gly, Leu, Ile, Cys, Ser or missing;
  • Xaa 4 is Glu, Gln, Asp, Asn, Leu, Ile, Val or missing;
  • Xaa 8 is Val, Leu, Gln, Ile, Met or missing;
  • Xaa 9 is Lys, Arg, His, Asp, Val or missing;
  • Xaa 10 is Glu, Gln, Asp, Asn or missing;
  • Xaa 11 is Glu, Asp, Asn, GIn or deletion.
  • Xaa 1 in formula (I) or (II) is Pro or absent; preferably Pro.
  • Xaa 2 in formula (I) or (II) is Ala or absent; preferably Ala.
  • Xaa 3 in formula (I) or (II) is Ala, Gly or deletion; preferably Ala.
  • Xaa 4 in formula (I) or (II) is Glu, GIn, Asp or deletion; preferably Glu, Asp or GIn.
  • Xaa 8 in formula (I) or (II) is Val or absent; preferably Val.
  • Xaa 9 in formula (I) or (II) is Lys, Arg or deletion; preferably Lys or deletion.
  • Xaa 10 in formula (I) or (II) is Glu, GIn or deletion; preferably GIn.
  • Xaa 11 in formula (I) or (II) is Asp or absent; preferably Asp.
  • one of Xaa 1 , Xaa 2 , Xaa 3 and Xaa 4 in formula (I) or (II) is deleted; or two are deleted; or three are deleted; or are missing.
  • one of Xaa 8 , Xaa 9 , Xaa 10 and Xaa 11 in formula (I) or (II) is deleted; or two are deleted; or three are deleted; or are missing.
  • the compound is selected from any one of Compounds 1-73 of the present invention.
  • the compound is selected from:
  • the compound is Pro-Ala-Ala-Glu-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp (Compound 1).
  • the present invention provides a method of repairing mucosal damage, the method comprising administering to a subject a compound of the present invention or a physiologically compatible salt thereof or making the mucosal damage a compound of the present invention or a physiologically compatible salt thereof salt exposure.
  • the repair is by modulating stem cell proliferation and differentiation.
  • the mucosal damage is mucosal damage in the lumen of the digestive system, respiratory system and the like.
  • Digestive system mucosal damage is related to oral cavity, esophagus, and gastrointestinal diseases, and the oral diseases include oral ulcers, stomatitis, gingivitis, periodontitis, etc.; the esophageal diseases include esophagitis, esophageal ulcers, etc.; the gastrointestinal diseases Including chronic gastritis, chronic atrophic gastritis, acute gastritis, gastroduodenal ulcer, functional gastrointestinal diseases, dyspepsia, precancerous lesions, digestive system tumors, gastrointestinal bleeding, gastroesophageal reflux disease, acute and chronic Enteritis, ulcerative colitis, Crohn's disease, and mucosal damage from chemoradiation, but not limited thereto.
  • the digestive tract mucosa includes gastric mucosa and intestinal mucosa.
  • chronic gastritis includes chronic atrophic gastritis.
  • the mucosal damage is gastric mucosal damage caused by irritating substances or drugs, stress states.
  • the irritant substances are such as hydrochloric acid, ethanol or alcohol, and the drugs are such as non-steroidal anti-inflammatory drugs aspirin or indomethacin.
  • the present invention provides a method of preventing, alleviating or treating a disease of the digestive tract or eliminating inflammatory edema, the method comprising administering to a subject a compound of the present invention or a physiologically compatible salt thereof.
  • the gastrointestinal diseases include oral cavity, esophagus, and gastrointestinal diseases, and the oral diseases include oral ulcers, stomatitis, gingivitis, periodontitis, etc.; the esophageal diseases include esophagitis, esophageal ulcers, etc.; the gastrointestinal diseases include Diseases include chronic gastritis, chronic atrophic gastritis, acute gastritis, gastroduodenal ulcer, functional gastrointestinal disease, dyspepsia, precancerous lesions, digestive system tumors, gastrointestinal bleeding, gastroesophageal reflux disease, acute Chronic enteritis, ulcerative colitis, Crohn's disease and mucosal damage from chemoradiation; but not limited thereto.
  • the prevention, alleviation or treatment of the digestive tract disease is performed by modulating stem cell proliferation and differentiation.
  • the compounds of the present invention or their physiologically compatible salts play a protective effect on gastrointestinal mucosa such as gastric mucosa or intestinal mucosa, or repair the damage of digestive tract mucosa such as gastric mucosa or intestinal mucosa, thereby preventing and reducing the damage to the digestive tract mucosa. Or the role of the treatment of gastrointestinal diseases.
  • the present invention provides a method of repairing a mucosal or skin wound comprising administering to a subject a compound of the present invention or a physiologically compatible salt thereof.
  • the repair of the mucosal or skin wound comprises regulating stem cell proliferation and differentiation.
  • the compound of the present invention or a physiologically compatible salt thereof is administered orally, by injection, subcutaneously, or the like.
  • the present invention provides a method of repairing a skin wound, the method comprising contacting the skin wound with a compound of the invention or a physiologically compatible salt thereof.
  • the skin wound is associated with, but not limited to, epidermal inflammation, mechanical and surgical wounds, burns and scalds, ulcers, fistulas, bedsores, and skin lesions caused by chemoradiotherapy.
  • the skin wound refers to the damage to normal skin caused by external injury factors such as surgery, external force, heat, current, chemical substances, low temperature, and internal factors in the body such as local blood supply disorders. .
  • the skin wound is often accompanied by disruption of skin integrity and loss of a certain amount of normal tissue.
  • the skin wound comprises impairment of the normal function of the skin.
  • the recovery of the skin wound is by modulating stem cell proliferation and differentiation.
  • the present invention provides a method of promoting the proliferation of HaCAT cells, the method comprising contacting the cells with a compound of the present invention or a physiologically compatible salt thereof.
  • the present invention provides a method of regenerating damaged blood vessels, the method comprising contacting the damaged blood vessel with a compound of the present invention or a physiologically compatible salt thereof.
  • the damaged blood vessels include vascular damage caused by mucosal damage of the digestive tract and skin trauma.
  • the present invention provides a pharmaceutical, food, health care product or cosmetic, daily necessities composition
  • a pharmaceutical, food, health care product or cosmetic, daily necessities composition comprising the compound of the present invention or a physiologically compatible salt thereof and a physiologically acceptable carrier.
  • the physiologically acceptable carrier includes a pharmaceutically acceptable carrier or a cosmetically acceptable carrier.
  • the pharmaceutical food, health care product or cosmetic, daily necessities composition can be prepared according to the conventional techniques of formulation or cosmetics, including mixing the compound of the present invention as an active ingredient with a carrier, and preparing the desired dosage form according to conventional techniques.
  • the composition of the present invention can be formulated into oral administration preparations, mucosal administration preparations, injection preparations, inhalation preparations and external preparations, as required.
  • the polypeptide of the present invention has no homology with known polypeptides, which is convenient for artificial polypeptide synthesis to obtain a high-purity polypeptide. Compared with the epidermal growth factor polypeptide, the polypeptide of the present invention is stable in vivo because it only consists of at most 12 amino acid residues.
  • the polypeptide of the invention can promote the proliferation and differentiation of stem cells, especially can promote the proliferation and differentiation of gastric organoids, participate in and regulate the proliferation and differentiation of gastric epithelial stem cells, thereby repairing the damage of gastrointestinal stem cells and epidermal stem cells.
  • the polypeptide of the present invention when applied to the skin wound on the body surface, it can work even after being sterilized by iodine preparation or hydrogen peroxide, or can exist stably in artificial gastric juice, artificial intestinal juice, etc., and when the epidermal growth factor is used on the body surface skin, After disinfection by iodine preparation or hydrogen peroxide, the structure will be destroyed and cannot function.
  • Figure 1 shows a schematic diagram of the steps of solid-phase synthesis of peptides.
  • Figure 2 shows the antiulcer effect of compound 1 on ethanol-induced gastric ulcer in mice.
  • Figure 3 shows the results of Compound 1 on promoting proliferation and differentiation of gastric organoids.
  • Figure 4 shows the results of Compound 1 on promoting the differentiation of gastric organoids.
  • Figure 5 shows the therapeutic effect of compound 1 on chronic atrophic gastritis mice.
  • Figure 6 shows the effect of compound 1 on gastric epithelial stem cells in chronic atrophic gastritis rats.
  • Figure 7 shows the therapeutic effect of compound 1 on chronic atrophic gastritis rats.
  • Figure 8 shows the pro-proliferative effect of compound 1 on HaCAT cells.
  • Figure 9 shows a schematic diagram of the analysis site of the zebrafish subintestinal vessel area.
  • Figure 10 shows a typical graph of the area of the subintestinal blood vessels in zebrafish treated with compounds of the present invention.
  • Figure 11 shows the number of branches of the subintestinal blood vessels of zebrafish treated with compounds of the present invention.
  • physiologically compatible salts refers to salt forms that are physiologically compatible (ie, pharmacologically acceptable) and that are substantially non-toxic to the individual to which the compounds of the present invention will be administered.
  • Physiologically compatible salts of the compounds of the present invention include conventional and stoichiometric acid or base addition salts formed from suitable, non-toxic, organic or inorganic acids or inorganic bases.
  • subject refers to an animal, preferably a mammal, most preferably a human.
  • subject refers to a mammal or a human with skin trauma and/or mucosal damage. It will be understood by those skilled in the art that the repair of skin wounds and/or mucosal lesions of the present invention may be administered for cosmetic (ie, non-therapeutic) as well as therapeutic purposes.
  • skin damage in the present application also includes skin damage repaired for cosmetic purposes, for example, wrinkles (such as those caused by UV exposure), skin lines, cracks, bumps, large pores (such as those associated with accessory structures such as sweat gland ducts) , sebaceous glands or hair follicles), or uneven or rough, loss of skin elasticity (loss and/or inactivation of functional skin elastin), sagging (including eye and jaw puffiness), loss of skin firmness, loss of skin firmness, Loss of recovery from skin deformation, discoloration (including dark circles), blemishes, sallow complexion, hyperpigmented skin areas such as age spots and freckles, keratin, abnormal differentiation, hyperkeratinization, elastin degeneration, destruction of collagen , and skin keratin, dermis, epidermis, skin vasculature (eg, telangiectasias or polyfurcations), and other tissue changes in the subcutaneous tissue, especially near the skin.
  • wrinkles such as those caused
  • Polypeptide compounds are synthesized by conventional solid-phase synthesis methods, and go through multiple cycles of resin swelling, substitution, deprotection, washing, amino acid dissolution, amino acid activation and condensation, washing, re-deprotection, and finally cleavage and side chain deprotection.
  • Cl-2-Cl-Resin represents 2-chlorotrityl chloride resin (2-Chlorotrityl Chloride Resin); Fmoc-Aa(n) etc.
  • DIPEA is N,N-diisopropylethylamine
  • DCM is dichloromethane
  • PIP is piperidine
  • DMF is N,N-dimethylformamide
  • HOBt is 1-hydroxybenzotriazole
  • DIC is N,N' - diisopropylcarbodiimide
  • TFA is trifluoroacetic acid
  • TIPS is triisopropylsilane.
  • the following takes compound 1 (Pro-Ala-Ala-Glu-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp) as an example to illustrate the synthesis and purification method of compound 1.
  • the method includes steps 1, Preparation of fully protected peptide resin; 2. Cleavage and deprotection; 3. Purification (salt exchange) lyophilization.
  • steps (3) to (5) are repeated to extend the peptide chain according to the amino acid sequence until the coupling of the last amino acid is completed.
  • the crude peptide obtained in step 2 was subjected to HPLC purification using the following chromatographic parameters Condition A. Specifically, dissolve the crude peptide obtained in step 2 with water and/or acetonitrile, and filter through a 0.45 ⁇ m filter; inject the sample; elute with an acetonitrile-water mobile phase gradient; collect the target peptide eluent; and finally perform rotary evaporation concentrate.
  • Chromatographic column YMC-Actus Triart C18 30*250mm;
  • UV detection wavelength 220nm UV detection wavelength 220nm
  • the product obtained in the above step is salt-exchanged by HPLC to obtain the final peptide compound 1.
  • the structure of Compound 1 was confirmed by MS and 1 H-NMR.
  • Double charge means 1/2 of the target peak in mass spectrometry
  • triple charge means 1/3 of the target peak in mass spectrometry
  • N/A means it is difficult to weigh, excluding the actual weight.
  • Example 2 Anti-ulcer effect of polypeptide (compound 1) on ethanol-induced gastric ulcer model in mice
  • mice were randomly divided into 4 groups with 10 mice in each group, namely: the control group (blank group), the model group (modeling with absolute ethanol), and the positive drug group (replacement group). Prednone 160 mg/kg), compound 1 group (1 mg/kg). After the animals were grouped, the control group and the model group were given pure water treatment, and the other treatment groups were given corresponding drug treatment, once a day for 8 consecutive days. After the administration on the 7th day, all animals were fasted for 24 hours. After 30 minutes of administration on the 8th day, mice in each group (except the control group) were given 0.15 mL of absolute ethanol by oral gavage to establish a model. After 1 hour, CO 2.
  • the animals were killed by overdose, the gastric cardia was ligated and the pylorus was clipped, and the whole stomach was extracted. 1 mL of 1% formaldehyde solution was injected into the stomach, the cardia was ligated, and the stomach was taken out and then placed in 1% formaldehyde solution for 25 minutes. Cut along the greater curvature of the stomach, wash the gastric contents with normal saline, and calculate the ulcer index after flattening.
  • Ulcer index calculation method If the length of the cord-like injury is greater than 1mm, the length is measured, and each millimeter is counted as 1 point; if the width is greater than 1mm, the score will be doubled according to the number of millimeters of the width; if the length is less than 1mm, 0.5 points will be scored. Addition gives the animal's ulcer index.
  • Example 3 Anti-ulcer effect of some polypeptide samples obtained in Example 1 on ethanol-induced gastric ulcer model in mice
  • mice were randomly divided into groups: 5 in the blank group, 10 in the model group, and 10 in each administration group. Except for the blank group and the model group, which were given purified water by gavage, the administration group was administered 0.2 mg/kg. The different test samples were given by gavage at the same dosage. After 1 hour of administration, the mice in each group were given 0.9 ml/kg of absolute ethanol by oral gavage to make a model. After 1 h, the animals were killed by de-neck method, and their stomachs were ligated. Cardia and pylorus were clipped, and the whole stomach was extracted.
  • Calculation method of ulcer index If the length of the cord-shaped injury is greater than 1mm, the length of the cord-like injury is measured, and each millimeter is counted as 1 point; if the width is greater than 1mm, the score will be doubled according to the number of millimeters of the width; if the length is less than 1mm, 0.5 points will be scored. Addition gives the animal's ulcer index.
  • Ulcer inhibition rate (Ulcer index in model group-Ulcer index in drug administration group)/Ulcer index in model group ⁇ 100%;
  • Relative ulcer inhibition rate (Ulcer inhibition rate of test compound)/(Ulcer inhibition rate of compound 1).
  • the antiulcer effect of each compound was tested in multiple batches.
  • the anti-ulcer activity was expressed as the mean value of the relative ulcer inhibition rate (compound 1 was used as the control group for each batch of experiments).
  • Relative ulcer inhibition rate (test compound ulcer inhibition rate) / (compound 1 ulcer inhibition rate)
  • the relative ulcer inhibition rate was 0.9-1.20, expressed as "+++";
  • the relative ulcer inhibition rate was 0.6–0.9, denoted as "++";
  • the relative ulcer inhibition rate was 0.3–0.6, denoted as "+";
  • ND means that no comparison with compound 1 was performed.
  • the gastric antrum epithelial tissue of healthy wild-type mice was used for organoid culture.
  • the gastric antrum epithelial layer and the muscle layer of the mice were separated under a stereoscope, cut into small pieces about 0.5 mm in size, and placed in the Digestion was performed in 2.5mM EDTA/DPBS at 4°C for about 1 h.
  • the digested gastric gland epithelial cells were filtered and centrifuged, and the supernatant was discarded.
  • the obtained cell pellet was resuspended in Matrigel, and cultured in a petri dish by adding growth factor-containing organoid medium.
  • test drugs were added, and compound 1 was added to the experimental group under the same culture conditions as the control group, and the spheres and growth conditions were observed and photographed with an inverted microscope on Day1, Day3, Day5, and Day7, and the effect of compound 1 on the growth of organoids was observed.
  • RESULTS Compared with the control group, the growth rate of organoids in the medium containing compound 1 was significantly accelerated over time. Through quantitative analysis of the diameter of the organoids, the results showed that the volume of the organoids treated with compound 1 was significantly larger than that of the control group.
  • the control group (see Figure 3, compared with the control group in Figure 3, * represents p ⁇ 0.05), can significantly promote the proliferation of organoids (there are also significant differences in statistical data).
  • the RNA of organoids treated with compound 1 was extracted, and the expression of differentiation-related genes was detected by real-time fluorescence quantitative PCR. It was found that the mRNA levels of endocrine cell markers SST and Gastrin and neck mucus cell marker TFF2 were significantly increased (see Figure 4).
  • Example 5 Therapeutic effect of compound 1 on a mouse model of chronic atrophic gastritis
  • mice Lgr5-GFP-CreERT mice
  • MNNG N-methyl-N-nitro-N-nitrosoguanidine
  • ranitidine the mice were free to drink
  • mice were given 150 mg/kg of ranitidine (8 mg/ml) in water by gavage at fixed times daily for 20 weeks, containing MNNG (100 mg/ml) in water.
  • the mice were given compound 1 (5 mg/kg) by gavage every day on the basis of drinking ordinary distilled water, and the therapeutic effect of compound 1 on chronic atrophic gastritis was observed after 2 weeks of administration.
  • results of tissue staining showed that the structure of the glands in the gastric corpus and gastric antrum of the model group was disordered, accompanied by a decrease in the number of gastric acid cells (H + -K + -ATPase positive) cells, and the mucosal epithelium in the gastric antrum was altitude drop.
  • the structure of the gastric glands of the mice in the compound 1 group returned to normal, the number of gastric acid cells increased significantly, and the height of the gastric antrum mucosa basically returned to normal.
  • the results are shown in Figure 5. These results indicate that compound 1 can promote the repair of chronic atrophic gastritis in mice.
  • Example 6 Therapeutic effect of compound 1 on chronic atrophic gastritis rats
  • the blank control group was given daily gavage of 5ml/kg of deionized water, normal diet, and free drinking water.
  • the CAG group was given a daily gavage of 120 ⁇ g/mL MNNG (N-methyl-N-nitro-N-nitrosoguanidine) aqueous solution, 5 ml/kg, 0.03% ranitidine diet and 2% ranitidine freely.
  • MNNG N-methyl-N-nitro-N-nitrosoguanidine
  • the body weight changes of the rats were regularly detected every week, the food intake and water intake were determined, and the activity status of the rats was observed.
  • the rats were fasted for 24 hours and anesthetized by intraperitoneal injection of 10% chloral hydrate 3.5ml/kg.
  • the whole stomach was removed, quickly cut along the greater curvature, rinsed with normal saline, and the gastric mucosa tissue from the lesser curvature and the near greater curvature up to the esophagus and down to the duodenum was collected in 10% neutral fur.
  • the body hair of the rats in the normal group was smooth and thick, and the hair color was white and shiny. It has a high degree of activity, and has a high degree of response to activities such as moving the cage, feeding water, and sound.
  • the mental state is good, and the mood is stable during operations such as gavage and weighing.
  • the hair of the rats in the model group was sparse and sparse, easy to fall off, and the hair color was dark and beige.
  • Example 7 Promoting effect of compound 1 on the proliferation of HaCAT cells
  • the concentration of HaCAT cells was adjusted to 1.0 ⁇ 10 5 ⁇ 5.0 ⁇ 10 5 /mL for subculture, and cultured at 37°C under 5% CO 2 for 24-36 hours for the detection of biological activity.
  • Cells were digested with trypsin and collected, and inoculated into a 96-well cell culture plate with serum-free medium at a concentration of 2.5 ⁇ 10 4 /mL, 100 ⁇ L per well, that is, 2500 cells/well, at 37°C, 5% CO 2 . Incubate overnight under conditions.
  • Example 8 Repairing effects of compound 1 and compound 26 on zebrafish vascular injury
  • the normal control group was treated with standard diluted water, and the other experimental groups were treated with simvastatin in water for 3 hours to establish the zebrafish microvessel deletion model. After 3 hours, the aqueous solutions of all groups were replaced with standard dilution water to terminate simvastatin induction.
  • the test drug groups were given compound 1 (500ng/tail) or compound 26 (500ng/tail) by intravenous injection respectively, and were treated at 28°C for 2 days. 10 zebrafish were randomly selected from each group to be photographed under a fluorescence microscope, and the data were analyzed and collected by NIS-Elements D 3.20 advanced image processing software to analyze the number of branches of lower intestinal blood vessels.
  • Example 9 Gastric and intestinal stability test of some polypeptide samples
  • Method Take 1 mg of each sample to be tested (Compound 1, Compound 26, Compound 27, Compound 28 and EGF), add 1 ml of water to dissolve. Take 100ul of the sample solution, add 900ul of water, mix well, and use it as the reference solution. Take 100ul of each sample solution, add 900ul of artificial gastric juice (W), artificial intestinal juice (X), povidone-iodine solution (I), and hydrogen peroxide solution (O) respectively, 37 ° C constant temperature water bath for 1 hour, let cool, filter , as the test solution, the peak areas of the samples before and after the treatment were detected by high performance liquid chromatography, and the test results were calculated by comparing the peak areas of the samples. Take the stock solution without any treatment after dilution with water as the control, and compare and count the changes of the peak area (content) at the corresponding position of the other test solutions.
  • W gastric juice
  • X artificial intestinal juice
  • I povidone-iodine solution
  • OF hydrogen peroxide solution
  • W stands for artificial gastric juice
  • X stands for artificial intestinal fluid
  • I stands for povidone-iodine solution
  • O stands for hydrogen peroxide solution
  • test articles As shown in Table 9, 4 test articles (compounds 1, 26, 27 and 28) were 100% retained for artificial gastric juice (W), povidone-iodine solution (I) and hydrogen peroxide solution (O) , indicating that it is very stable; compound 26 is also extremely stable to artificial intestinal fluid (X); EGF is not retained in gastric and intestinal fluids, indicating that it is unstable in gastric and intestinal fluids. Povidone-iodine solution and hydrogen peroxide solution are applied after disinfection. EGF topical can also be destroyed.
  • the animals were sacrificed, the cardia was ligated and the pylorus was clipped, and the whole stomach was removed. Inject 8 mL of 1% formaldehyde solution into the stomach, ligate the pylorus, take out the stomach and put it in 1% formaldehyde solution to fix it, cut it along the greater curvature of the stomach after 30 minutes, wash the stomach contents with normal saline, flatten the stomach, and observe it. And take panoramic photos of the stomach to measure the ulcer area.

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Abstract

一种用于修复黏膜损伤或皮肤创伤的新型多肽及其应用，所述多肽与已知多肽无同源性，而且能够在体内和体外均稳定存在，具有调控干细胞增殖和分化修复黏膜损伤或皮肤创伤的效果；以及涉及所述新型多肽通过调控干细胞增殖和分化修复黏膜损伤或皮肤创伤的用途，以及用于预防、减轻或治疗胃肠疾病的用途。

Description

用于修复黏膜损伤或皮肤创伤的多肽及其应用 技术领域
本发明涉及用于修复黏膜损伤或皮肤创伤的新型多肽及其应用，本发明的多肽与已知多肽无同源性，而且能够在体内和体外均稳定存在，具有调控干细胞增殖和分化，修复黏膜损伤或皮肤损伤的效果；以及涉及所述新型多肽通过调控干细胞增殖和分化修复黏膜损伤或皮肤创伤的用途，以及用于预防、减轻或治疗胃肠疾病的用途。
背景技术
皮肤创伤和/或黏膜损伤是很多疾病的共同病理特征。皮肤创伤或皮肤损伤是指正常皮肤(组织)在外界致伤因子如外科手术、外力、热、电流、化学物质、低温以及机体内在因素如局部血液供应障碍等作用下所导致的损害。皮肤损伤常伴有皮肤完整性的破坏以及一定量正常组织的丢失，同时，皮肤的正常功能受损。也称为伤口或者创伤。目前有包括碱性成纤维生长因子、表皮生长因子、血小板生长因子、粒-巨噬细胞集落刺激因子及生长激素等蛋白质/多肽药物具有明显的修复创面、护肤、抗皱和防衰老作用，但这些蛋白质/多肽药物的氨基酸序列较长导致制备成本高、稳定性较差等缺点，因而其应用受到了一定的限制。
人体黏膜是指呼吸系统、消化系统、生殖泌尿系统等腔道或囊状肌性器官的内层，是仅次于皮肤的人体第二大屏障，包括口腔、咽、气管、食道、胃、肠道、阴道、膀胱等，这些器官的管壁或囊壁均有共同的分层规律，又具有与其功能相适应的特点，其胚胎起源、组织结构、病理过程、临床表现、愈后等均有共同特点。
慢性胃炎是一种胃黏膜慢性炎症，是消化内科的常见病和多发病，临床上将不同原因引起的胃黏膜慢性炎症(即在病理上表现为单核细胞和淋巴细胞浸润)和(或)腺体萎缩性病变称为慢性胃炎。以胃黏膜上皮和腺体萎缩为特征的慢性萎缩性胃炎(chronic atrophic gastrifis,CAG)，其起病隐匿、病程长、症状无特异性、难以治愈，在慢性胃炎中占11％～31％，好发于中老年人群，发病和年龄有一定的关联但与性别没有关系，发病缓慢且病势缠绵，迁延难愈，治疗棘手。慢性萎缩性胃炎是胃癌发生发展重要阶段被视为胃癌前病变，1978年世界卫生组织％WHO)已将CAG列为癌前状态，其常伴假幽门腺化生和肠上皮化生，或不典型增生阶段等癌前病变。尤其是伴有弥漫的肠上皮化生或不典型增生者，癌变可能性更大，业界大多数专业人士都赞同通过慢性浅表性胃炎、萎缩性胃炎、肠上皮化生、异型增生、肠型胃癌这样的发展模式。故早期准确的诊断和治疗对慢性萎缩性胃炎患者有着重要的意义。目前现代医学治疗本病主要以改善症状、手术介入为主，在腺体萎缩、肠化生方面尚无良策。CAG为慢性胃炎转变为胃癌的重要阶段，积极治疗CAG对预防其癌变、降低胃癌发病率有 着深远意义。寻求萎缩性胃炎的有效治疗方法，是更好地开展胃癌预防的重要措施之一。
黏膜组织的损伤临床上会导致慢性胃炎和消化道溃疡等胃肠道疾病。黏膜上皮的修复有修复(restitution)和再生(regeneration或renewal)两种不同的机制(Cur.Med.Chem.,2008,15,3133-3144)：修复或恢复一般在损伤后几分钟内就开始，通过细胞迁移快速修复浅表病变；再生是通过干细胞和祖细胞的分化和增殖持续再生，持续数天至数月。
表皮生长因子(EGF)是一种由53个氨基酸残基组成的多肽，广泛存在于多种组织器官和体液中，其可促进上皮细胞增殖从而对皮肤起到保护作用。表皮生长因子主要是促进皮肤组织细胞的增殖与生长，使新生的细胞替代衰老的细胞，从而起到抗衰老和护肤保健等功能。表皮生长因子已经被报道具有修复创面作用，因皮肤创面需要消毒、清创时，会应用含碘或双氧水的消毒剂，EGF在此条件下不稳定。生长因子与胃肠吻合口愈合有关(J.Surgical Res.2014；17:202-210)，但是EGF经胃肠口服给药时，进入体内后会发生降解，不能达到治疗效果。
因此，需要一种体内外稳定、能够治疗皮肤和/或黏膜损伤，特别是胃肠道黏膜损伤的肽类物质。
发明内容
为了克服现有技术的不足和缺陷，本发明的目的在于提供一类新型多肽。
第一方面，本发明提供式(I)的化合物或其生理学上相容的盐，其中所述式(I)的化合物如下：
H-Xaa 1-Xaa 2-Xaa 3-Xaa 4-Xaa 5-Val-Xaa 6-Xaa 7-Xaa 8-Xaa 9-Xaa 10-Xaa 11-OH(I)
其中
Xaa 1为Pro、Gly、Ala或缺失；
Xaa 2为Ala、Leu、Ile、Gly、Cys、Ser或缺失；
Xaa 3为Ala、Pro、Gly、Leu、Ile、Cys、Ser或缺失；
Xaa 4为Glu、Gln、Asp、Asn、Leu、Ile、Val或缺失；
Xaa 5为Pro、Gly、Ala、Val或缺失；
Xaa 6为Pro、Gly或Ala；
Xaa 7为Leu、Phe、Val、Ala、Tyr、Glu、Lys、Asp、Ile、Met或缺失；
Xaa 8为Val、Leu、Gln、Ile、Met或缺失；
Xaa 9为Lys、Arg、His、Asp、Val或缺失；
Xaa 10为Glu、Gln、Asp、Asn或缺失；以及
Xaa 11为Glu、Asp、Asn、Gln或缺失，
条件是：Xaa 1、Xaa 2、Xaa 3、Xaa 4、Xaa 5、Xaa 7、Xaa 8、Xaa 9、Xaa 10和Xaa 11中至多有9个是缺失的。
在一实施方案中，X aa5为Pro。
在一实施方案中，X aa6为Pro。
在一实施方案中，X aa7为Leu。
在一实施方案中，式(I)化合物具有以下式(II)结构，
H-Xaa 1-Xaa 2-Xaa 3-Xaa 4-Pro-Val-Pro-Leu-Xaa 8-Xaa 9-Xaa 10-Xaa 11-OH(II)，
其中
Xaa 1为Pro、Gly、Ala或缺失；
Xaa 2为Ala、Leu、Ile、Gly、Cys、Ser或缺失；
Xaa 3为Ala、Pro、Gly、Leu、Ile、Cys、Ser或缺失；
Xaa 4为Glu、Gln、Asp、Asn、Leu、Ile、Val或缺失；
Xaa 8为Val、Leu、Gln、Ile、Met或缺失；
Xaa 9为Lys、Arg、His、Asp、Val或缺失；
Xaa 10为Glu、Gln、Asp、Asn或缺失；以及
Xaa 11为Glu、Asp、Asn、Gln或缺失。
在一实施方案中，式(I)或(II)中的Xaa 1为Pro或缺失；优选为Pro。
在一实施方案中，式(I)或(II)中的Xaa 2为Ala或缺失；优选为Ala。
在一实施方案中，式(I)或(II)中的Xaa 3为Ala、Gly或缺失；优选为Ala。
在一实施方案中，式(I)或(II)中的Xaa 4为Glu、Gln、Asp或缺失；优选为Glu、Asp或Gln。
在一实施方案中，式(I)或(II)中的Xaa 8为Val或缺失；优选为Val。
在一实施方案中，式(I)或(II)中的Xaa 9为Lys、Arg或缺失；优选为Lys或缺失。
在一实施方案中，式(I)或(II)中的Xaa 10为Glu、Gln或缺失；优选为Gln。
在一实施方案中，式(I)或(II)中的Xaa 11为Asp或缺失；优选为Asp。
在一实施方案中，式(I)或(II)中的Xaa 1、Xaa 2、Xaa 3和Xaa 4中有一个是缺失的；或者有两个是缺失的；或者有三个是缺失的；或者都是缺失的。
在一实施方案中，式(I)或(II)中的Xaa 8、Xaa 9、Xaa 10和Xaa 11中有一个是缺失的；或者有两个是缺失的；或者有三个是缺失的；或者都是缺失的。
在一实施方案中，所述化合物选自本发明化合物1-73中任一个。
在一实施方案中，所述化合物选自：
Pro-Ala-Ala-Glu-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物1)；
Pro-Ala-Ala-Glu-Pro-Val-Pro-Leu-Val-Lys(化合物3)；
Pro-Ala-Ala-Gln-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物10)；
Pro-Ala-Ala-Glu-Pro-Val-Pro-Leu(化合物26)；
Ala-Glu-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物27)；
Ala-Glu-Pro-Val-Pro-Leu(化合物30)；
Glu-Pro-Val-Pro-Leu(化合物31)；
Pro-Val-Pro-Leu(化合物32)；
Pro-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物28)；
Pro-Ala-Ala-Asp-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物11)；
Pro-Ala-Gly-Glu-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物48)；
Pro-Ala-Ala-Glu-Pro-Val-Pro-Leu-Val-Lys-Glu-Asp(化合物57)；或
Glu-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物38)。
为方便起见，在本申请中描述本发明的化合物时，省略左侧的H和右侧的OH。
在一实施方案中，所述化合物为Pro-Ala-Ala-Glu-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物1)。
第二方面，本发明提供一种修复黏膜损伤的方法，所述方法包括对受试者给予本发明的化合物或其生理学上相容的盐或使黏膜损伤与本发明的化合物或其生理学上相容的盐接触。在一实施方案中，所述修复是通过调控干细胞增殖和分化进行的。
在一实施方案中，所述黏膜损伤是消化系统、呼吸系统等腔道内黏膜损伤。
消化系统黏膜损伤与口腔、食道、胃肠疾病相关，所述口腔疾病包括口腔溃疡、口腔炎、牙龈炎、牙周炎等；所述食道疾病包括食管炎、食管溃疡等；所述胃肠疾病包括慢性胃炎、慢性萎缩性胃炎、急性胃炎、胃十二指肠溃疡、功能性胃肠道疾病、消化不良、癌前病变、消化系统肿瘤、胃肠道出血、胃食管返流疾病、急慢性肠炎、溃疡性结肠炎、克罗恩病和放化疗引起的黏膜损伤，但不限于此。
在优选的实施方案中，所述消化道黏膜包括胃黏膜和肠黏膜。在优选的实施方案中，慢性胃炎包括慢性萎缩性胃炎。在优选的实施方案中，所述黏膜损伤是由刺激性物质或药物、应激状态引起的胃黏膜损伤。所述刺激性物质如盐酸、乙醇或酒精等，所述药物例如非类固醇类抗炎药阿司匹林或吲哚美辛等。
本发明提供一种预防、减轻或治疗消化道疾病或消除炎症水肿的方法，所述方法包括对受试者给予本发明的化合物或其生理学上相容的盐。所述消化道疾病包括口腔、食道、胃肠 疾病相关，所述口腔疾病包括口腔溃疡、口腔炎、牙龈炎、牙周炎等；所述食道疾病包括食管炎、食管溃疡等；所述胃肠疾病包括慢性胃炎、慢性萎缩性胃炎、急性胃炎、胃十二指肠溃疡、功能性胃肠道疾病、消化不良、癌前病变、消化系统肿瘤、胃肠道出血、胃食管返流疾病、急慢性肠炎、溃疡性结肠炎、克罗恩病和放化疗引起的黏膜损伤；但不限于此。在一实施方案中，所述消化道疾病的预防、减轻或治疗是通过调控干细胞增殖和分化进行的。所述方法通过本发明的化合物或其生理学上相容的盐对胃黏膜或肠黏膜等消化道黏膜起到保护作用或修复胃黏膜或肠黏膜等消化道黏膜的损伤，从而起到预防、减轻或治疗胃肠疾病的作用。
本发明提供一种修复黏膜或皮肤创面的方法，所述方法包括对受试者给予本发明的化合物或其生理学上相容的盐。在一实施方案中，所述黏膜或皮肤创面的修复包括调控干细胞增殖和分化。
在上述本发明的方法中，本发明的化合物或其生理学上相容的盐是通过口服、注射、皮下等方式给药的。
第三方面，本发明提供一种修复皮肤创伤的方法，所述方法包括使皮肤创伤与本发明的化合物或其生理学上相容的盐接触。在优选的实施方案中，所述皮肤创伤与表皮炎症、机械及手术创面、烧伤及烫伤、溃疡、瘘管、褥疮、放化疗引起的皮肤损伤相关，但不限于此。在一实施方案中，所述皮肤创伤是指正常皮肤在外界致伤因子如外科手术、外力、热、电流、化学物质、低温以及机体内在因素如局部血液供应障碍等作用下所导致的损害。在一实施方案中，所述皮肤创伤常伴有皮肤完整性的破坏以及一定量正常组织的丢失。在另一实施方案中，所述皮肤创伤包括皮肤的正常功能受损。在一实施方案中，所述皮肤创伤的恢复是通过调控干细胞增殖和分化进行的。
本发明提供一种促进HaCAT细胞增殖的方法，所述方法包括使所述细胞与本发明的化合物或其生理学上相容的盐接触。
第四方面，本发明提供一种使损伤的血管再生的方法，所述方法包括使损伤的血管与本发明的化合物或其生理学上相容的盐接触。在优选的实施方案中，所述损伤的血管包括消化道黏膜损伤和皮肤创伤导致的血管损伤。
第五方面，本发明提供一种药物、食品、保健品或化妆品、日用品组合物，所述组合物包括本发明的化合物或其生理学上相容的盐以及生理学上可接受的载体。在一实施方案中，所述生理学上可接受的载体包括药学上可接受的载体或化妆品可接受的载体。所述药物食品、保健品或化妆品、日用品组合物可以按照制剂学或化妆品常规技术制备，包括将作为活性成分的本发明的化合物与载体混合，按照常规技术制成所需要的剂型。可以根据需要，将本发 明的组合物配制成口服施用制剂、黏膜施用制剂、注射制剂、吸入制剂和外用制剂。
本发明的多肽与已知多肽无同源性，便于人工多肽合成以获得高纯度多肽，与表皮生长因子多肽相比本发明的多肽由于仅由至多12个氨基酸残基组成在体内稳定，而且本发明的多肽能促进干细胞增殖分化，特别是能促进胃类器官的增殖和分化，参与并调控胃上皮干细胞的增殖和分化，从而可对胃肠干细胞，表皮干细胞损伤进行修复，具有显著的减轻诸如慢性胃炎及消化道溃疡的胃肠疾病病理发展、消除炎症水肿、促进消化道黏膜损伤修复和皮肤创伤修复、缩短创面愈合时间、调解免疫功能等作用。此外，本发明的多肽用于体表皮肤创面时即使经碘制剂或双氧水消毒之后也能够起作用或者在人工胃液、人工肠液等中也能稳定存在，而表皮生长因子用于体表皮肤时，经碘制剂或双氧水消毒之后结构会破坏，不能发挥作用。
附图说明
图1示出了多肽固相合成步骤的示意图。
图2示出了化合物1对乙醇诱导的小鼠胃溃疡的抗溃疡作用。
图3示出了化合物1对促进胃类器官的增殖和分化的结果。
图4示出了化合物1对促进胃类器官的分化的结果。
图5示出了化合物1对慢性萎缩性胃炎小鼠的治疗作用。
图6示出了化合物1对慢性萎缩性胃炎大鼠胃上皮干细胞的影响。
图7示出了化合物1对慢性萎缩性胃炎大鼠的治疗作用。
图8示出了化合物1对HaCAT细胞的促增殖作用。
图9示出了斑马鱼肠下血管面积分析部位示意图。
图10示出了经本发明化合物处理后的斑马鱼肠下血管面积典型图。
图11示出了经本发明化合物处理后的斑马鱼肠下血管分支数。
具体实施方式
术语“生理学上相容的盐”是指生理学上相容的(即药理学上可接受的)并且对将被施用本发明化合物的个体基本上无毒的盐形式。本发明化合物的生理学上相容的盐包括由适宜的、无毒的有机或无机酸或无机碱形成的常规的和化学计量的酸加成盐或碱加成盐。
术语“受试者”是指动物，优选为哺乳动物，最优选为人。具体地讲，术语“受试者”涉及具有皮肤创伤和/或黏膜损伤的哺乳动物或人。本领域技术人员应理解，本发明的皮肤创伤和/或黏膜损伤的修复可出于美容目的(即非治疗性目的)和治疗性目的施用。为此，本申请 的术语“皮肤损伤”还包括出于美容目的修复的皮肤损伤例如，皱纹(例如紫外线照射导致的皱纹)、皮肤纹、裂缝、肿块、大毛孔(例如与附件结构如汗腺管、皮脂腺或毛囊相关的)，或不平或粗糙，皮肤失去弹性(功能性皮肤弹性蛋白丧失和/或失活)、下垂(包括眼部和下颌浮肿)、皮肤硬度丧失、皮肤紧实度丧失、皮肤变形后的回复能力丧失、变色(包括黑眼圈)、斑疱、肤色灰黄、色素过量皮肤区域例如老年斑和雀斑、角质物、异常的分化、过度角质化、弹性组织变性、胶原蛋白的破坏，以及皮肤角质，真皮，表皮层，皮肤血管系统(例如毛细血管扩张或多叉血管)，以及皮下组织，特别是靠近皮肤的皮下组织中的其它组织变化。
以下是结合具体试验对本发明的说明，并不是对本发明保护范围的限制。
表1书写中所使用到试剂、溶剂的英文名或缩写
英文名或缩写 中文名
HBTU 苯并三氮唑-N,N,N',N'-四甲基脲六氟磷酸盐
Methanol 甲醇
Tert-Butyl methyl ether 甲基叔丁基醚
Ethanol 乙醇
AA 氨基酸
实施例1：多肽的化学合成
多肽化合物的合成采用常规固相合成方法，经过树脂溶胀、取代、脱保护、洗涤、氨基酸溶解、氨基酸活化和缩合过程、洗涤、再脱保护多个循环过程，以及最后裂解和侧链脱保护。
多肽固相合成步骤示意图参见图1。图1中，Cl-2-Cl-Resin表示2-氯三苯甲基氯树脂(2-Chlorotrityl Chloride Resin)；Fmoc-Aa(n)等表示9-芴基甲氧基羰基的氨基酸；DIPEA为N,N-二异丙基乙胺；DCM为二氯甲烷；PIP为哌啶；DMF为N,N-二甲基甲酰胺；HOBt为1-羟基苯并三唑；DIC为N,N'-二异丙基碳二亚胺；TFA为三氟乙酸；TIPS为三异丙基硅烷。
以下以化合物1(Pro-Ala-Ala-Glu-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp)为例，示出化合物1的合成和纯化的方法，所述方法包括步骤1、全保护肽树脂的制备；2、切割、脱保护；3、纯化(换盐)冻干。
步骤1、全保护肽树脂的制备
(1)树脂溶胀：称取2-Chlorotrityl Chloride Resin 2.0192g(S＝0.73mmol/g)，加入到有筛板的合成管中，用40ml二氯甲烷溶胀30min，抽滤除去二氯甲烷。
(2)制备Fmoc-Asp(OtBu)-树脂：按树脂、Fmoc-Asp(OtBu)-OH、DIPEA，以1：1.5： 1.65的摩尔比分别称取Fmoc-Asp(OtBu)-OH、DIPEA，溶解于二氯甲烷20ml中，加入合成管中。室温N 2鼓泡震荡1～3小时，直接向反应液中加入甲醇2ml封闭30min。然后分别用二甲基甲酰胺洗涤4次，25ml/次，抽干树脂。
(3)脱除Fmoc保护基：向反应器中加入20％哌啶-DMF(v/v)溶液20ml，N 2鼓泡反应20min，抽干；然后用二甲基甲酰胺洗涤6次，25ml/次，3分钟/次，抽干，茚三酮法检测Fmoc脱除结果。
(4)氨基酸预活化：在250ml圆底烧瓶中加入4.38mmol的Fmoc保护的氨基酸、5.26mmol HOBt、4.60mmol DIC，用1：1的DCM-DMF(v/v)20ml溶解，-5-0℃冰浴搅拌下预活化30～60min。
(5)氨基酸连接：将已活化的保护氨基酸溶液倒入反应器中，补加适量的DCM清洗用具。室温下N 2鼓泡反应1～3小时，茚三酮法检测氨基酸连接是否完全，若完全，抽干。树脂用二甲基甲酰胺洗涤4次，25ml/次，3min/次，抽干。每一种氨基酸、缩合剂的用量及具体反应时间见表2。
(6)第一个氨基酸缩合完成后，重复步骤(3)～(5)，按照氨基酸顺序延长肽链至最后一个氨基酸偶联完毕。
(7)树脂肽用二氯甲烷洗涤6次，25ml/次，3min/次，抽干。
表2氨基酸、缩合剂的用量
氨基酸名称 AA/eq 氨基酸用量/g HOBt/g DIPEA/g DIC/g
Fmoc-L-Asp(OtBu)-OH 2.19 0.90 0 0.31 0
Fmoc-L-Gln(Trt)-OH 4.38 2.67 0.71 0.57 0.58
Fmoc-L-Lys(Boc)-OH 4.38 2.05 0.71 0.57 0.58
Fmoc-L-Val-OH 4.38 1.49 0.71 0.57 0.58
Fmoc-L-Leu-OH 4.38 1.55 0.71 0.57 0.58
Fmoc-L-Pro-OH 4.38 1.48 0.71 0.57 1.16
Fmoc-L-Val-OH 4.38 1.49 0.71 0.57 0.58
Fmoc-L-Pro-OH 4.38 1.48 0.71 0.57 1.16
Fmoc-L-Glu(OtBu)-OH .H 2O 4.38 1.94 0.71 0.57 1.16
Fmoc-L-Ala-OH .H 2O 4.38 1.44 0.71 0.57 1.16
Fmoc-L-Ala-OH .H 2O 4.38 1.44 0.71 0.57 1.16
Fmoc-L-Pro-OH 4.38 1.48 0.71 0.57 1.16
步骤2、切割、脱保护
(1)在步骤1的合成管中，加入切割剂(TFA:TIPS:H2O＝95:2.5:2.5，v/v)50ml，N2鼓泡反应1.5-3小时。
(2)切割反应完成后，将切割剂抽滤至250ml圆底烧瓶中。真空浓缩至原切割剂体积的三分之一后，加入10倍现有体积的甲基叔丁基醚，搅拌30min。将所得混合溶剂过滤，并用30ml甲基叔丁基醚分别清洗3次后，将所得粗肽产品置于砂芯漏斗在通风橱中N2吹干，使溶剂挥发至粗肽为粉末状。得到粗肽1.87g，粗产率85.1％。
步骤3、纯化(换盐)和冻干
采用以下色谱参数条件A，对步骤2中获得的粗肽进行HPLC纯化。具体而言，用水和/或乙腈溶解步骤2中获得的粗肽，并经0.45μm滤膜过滤；进样；用乙腈-水流动相梯度洗脱；收集目的肽洗脱液；最后进行旋蒸浓缩。
色谱参数条件A：
色谱柱：YMC-Actus Triart C18 30*250mm；
洗脱液A：0.1％(v/v)TFA/H 2O；
洗脱液B：乙腈；
流速25ml/min；
紫外检测波长220nm；
表3梯度洗脱条件
时间min 洗脱液A(％) 洗脱液B(％)
0 90 10
30 75 25
接着采用以下色谱参数条件B利用HPLC法对上述步骤所获得的产品进行换盐，得到最终的肽化合物1。具体而言，使用95％A1+5％B平衡色谱柱；然后进样；接着采用95％A2+5％B平衡色谱柱；用A1和B梯度洗脱；收集目的肽洗脱液；最后旋蒸浓缩并且冷冻干燥，得到化合物1(纯化收率73.3％，纯度100％)。化合物1的结构经MS、 1H-NMR确认。
色谱参数条件B：
色谱柱：YMC-Actus Triart C18 30*250mm
洗脱液A1：0.1M乙酸
洗脱液A2:0.025M乙酸+0.1M乙酸铵
洗脱液B：乙腈
流速25ml/min
紫外检测波长220nm
表4梯度洗脱条件
时间min 洗脱液A1(％) 洗脱液B(％)
0 95 5
5 95 5
35 70 30
以与合成化合物1类似的方式，合成其他化合物。结果参见表5和说明书其他部分。
表5合成的化合物
Figure PCTCN2021103943-appb-000001
Figure PCTCN2021103943-appb-000002
Figure PCTCN2021103943-appb-000003
注释：双电荷代表质谱出1/2目标峰，三电荷代表质谱出1/3目标峰；N/A代表称量有难度，未计实际重量。
化合物1:Pro-Ala-Ala-Glu-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp醋酸盐
1H NMR(600MHz,DMSO)δ8.27(d,J＝7.6Hz,1H),8.21(d,J＝7.8Hz,1H),8.09(d,J＝7.5Hz,1H),8.03(d,J＝7.9Hz,1H),7.99(d,J＝7.4Hz,1H),7.94(dd,J＝16.7,8.0Hz,2H),7.58(d,J＝6.3Hz,1H),7.51(d,J＝8.6Hz,1H),7.16(s,1H),6.70(s,1H),4.48(dd,J＝13.3,7.9Hz,1H),4.39(dd,J＝8.2,4.2Hz,1H),4.35–4.21(m,6H),4.18–4.03(m,3H),3.71–3.47(m,5H),2.93–2.69(m,4H),2.45–2.32(m,2H),2.25(t,J＝7.7Hz,2H),2.08(t,J＝7.9Hz,2H),2.01–1.93(m,3H),1.93–1.84(m,14H,AcOH),1.84–1.76(m,3H),1.75–1.57(m,8H),1.54–1.39(m,5H),1.36–1.27(m,2H),1.21–1.10(m,6H),0.91–0.74(m,18H).
化合物3:Pro-Ala-Ala-Glu-Pro-Val-Pro-Leu-Val-Lys醋酸盐
1H NMR(600MHz,DMSO)δ8.22–8.13(m,2H),8.10(s,1H),7.98(d,J＝7.6Hz,1H),7.92(d,J＝8.4Hz,1H),7.72(s,1H),7.59(s,1H),4.52–4.43(m,1H),4.39–4.19(m,6H),4.12–4.04(m,1H),3.89(d,J＝5.9Hz,1H),3.67–3.46(m,6H),2.85(dt,J＝10.2,6.8Hz,1H),2.80–2.73(m,1H),2.69(s,2H),2.21(t,J＝7.4Hz,2H),2.03–1.76(m,14H,AcOH),1.73(d,J＝5.5Hz,1H),1.70–1.61(m,4H),1.61–1.52(m,3H),1.47(s,4H),1.26(s,2H),1.20–1.12(m,6H),0.91–0.83(m,9H),0.83–0.77(m,9H).
化合物4:Pro-Ala-Ala-Glu-Gly-Val-Pro-Leu-Val-Lys-Gln-Asp醋酸盐
1H NMR(600MHz,D 2O)δ4.45–4.16(m,10H),4.04(d,J＝8.1Hz,1H),3.93–3.76(m,3H),3.67–3.58(m,1H),3.42–3.28(m,2H),2.94(t,J＝7.4Hz,2H),2.72(qd,J＝16.4,6.0Hz,2H),2.45–2.27(m,5H),2.27–2.16(m,1H),2.11–1.86(m,17H,AcOH),1.84–1.47(m,8H),1.45–1.25(m,8H),0.91(d,J＝6.8Hz,3H),0.89–0.73(m,15H).
化合物5:Pro-Ala-Ala-Glu-Pro-Val-Gly-Leu-Val-Lys-Gln-Asp醋酸盐
1H NMR(600MHz,D 2O+D 3COD)δ4.66–4.62(m,2H),4.40(dd,J＝8.1,5.9Hz,1H),4.35–4.21(m,6H),4.03–3.98(m,2H),3.92–3.81(m,2H),3.78–3.71(m,1H),3.62(dt,J＝10.3,7.1Hz,1H),3.39–3.30(m,2H),2.96–2.87(m,4H),2.51–2.35(m,3H),2.32(t,J＝7.6Hz,2H),2.26–2.19(m,1H),2.10–1.91(m,14H,AcOH),1.91–1.78(m,2H),1.78–1.71(m,1H),1.70–1.58(m,3H),1.53(d,J＝3.6Hz,3H),1.42–1.34(m,2H),1.33(d,J＝7.2Hz,3H),1.30(d,J＝7.2Hz,3H),0.94–0.88(m,6H),0.88–0.83(m,9H),0.82–0.79(m,3H).
化合物8:Pro-Ala-Ala-Glu-Gly-Val-Gly-Leu-Val-Lys-Gln-Asp醋酸盐
1H NMR(600MHz,DMSO)δ8.30–8.24(m,3H),8.15(d,J＝7.3Hz,1H),8.11–8.03(m,2H),7.94(d,J＝7.4Hz,1H),7.88(d,J＝8.0Hz,1H),7.80(t,J＝9.4Hz,2H),7.59(d,J＝6.4Hz,1H),7.17(s,1H),6.70(s,1H),4.44–3.99(m,10H),3.82–3.57(m,7H),2.94–2.69(m,5H),2.47–2.41(m,1H),2.41–2.32(m,1H),2.21(t,J＝7.9Hz,2H),2.08(t,J＝7.9Hz,2H),2.03–1.79(m,11H,AcOH),1.78–1.26(m,14H),1.19(dd,J＝6.9,4.7Hz,6H),0.87–0.76(m,18H).
化合物9:Pro-Ala-Ala-Glu-Gly-Val-Gly-Leu
1H NMR(600MHz,DMSO)δ8.43(d,J＝7.6Hz,1H),8.29–8.23(m,2H),8.22–8.14(m,2H),7.81(d,J＝8.5Hz,1H),7.75(d,J＝7.9Hz,1H),4.32–4.08(m,5H),3.78–3.69(m,4H),3.63(dd,J＝16.4,5.6Hz,1H),2.98–2.86(m,2H),2.22(t,J＝7.9Hz,2H),2.07–1.92(m,2H),1.92–1.86(m,1H),1.79–1.64(m,4H),1.62–1.54(m,1H),1.50–1.40(m,2H),1.19(dd,J＝7.0,1.3Hz,6H),0.86–0.78(m,12H).
化合物10:Pro-Ala-Ala-Gln-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp醋酸盐
1H NMR(600MHz,DMSO)δ8.28(d,J＝7.7Hz,1H),8.14(d,J＝7.8Hz,1H),8.07(d,J＝7.4Hz,1H),8.04(d,J＝8.0Hz,1H),8.00(d,J＝7.3Hz,1H),7.95(t,2H),7.58(d,J＝6.3Hz,1H),7.50(d,J＝8.7Hz,1H),7.17(d,J＝15.3Hz,2H),6.76(s,1H),6.70(s,1H),4.45(q,J＝13.6,7.9Hz,1H),4.39(dd,J＝8.3,4.1Hz,1H),4.33–4.20(m,6H),4.20–4.09(m,2H),4.09–4.02(m,1H),3.69–3.47(m,5H),2.86(dt,J＝10.2,6.7Hz,1H),2.80–2.62(m,3H),2.46–2.40(m,1H),2.37(dd,J＝15.5,2.7Hz,1H),2.14–2.03(m,4H),2.02–1.76(m,16H,AcOH),1.75–1.39(m,13H),1.39–1.22(m,2H),1.22–1.11(m,6H),1.00–0.68(m,18H).
化合物11:Pro-Ala-Ala-Asp-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp醋酸盐
1H NMR(600MHz,DMSO)δ8.26(d,J＝7.6Hz,1H),8.19(d,J＝7.7Hz,1H),8.09(d,J＝7.7Hz,1H),8.04(d,J＝7.5Hz,1H),7.96(t,J＝7.4Hz,2H),7.87(d,J＝7.0Hz,1H),7.60(d,J＝6.4Hz,1H),7.37(d,J＝8.9Hz,1H),7.17(s,1H),6.70(s,1H),4.70(q,J＝14.2,7.0Hz,1H),4.37–4.21(m,7H),4.16–4.10(m,2H),4.09–4.03(m,1H),3.74–3.67(m,2H),3.68–3.59(m,4H),3.49–3.45(m,1H),2.99–2.84(m,3H),2.80–2.71(m,2H),2.63(dd,J＝16.5,7.5Hz,1H),2.46–2.25(m,4H),2.07(t,J＝7.9Hz,2H),2.05–1.76(m,20H,AcOH),1.74–1.58(m,6H),1.56–1.42(m,5H),1.32(d,J＝8.0Hz,2H),1.18(d,J＝7.0Hz,3H),1.14(d,J＝7.1Hz,3H),0.88–0.74(m,18H).
化合物12:Pro-Ala-Ala-Glu-Pro-Val-Pro-Phe-Val-Lys-Gln-Asp醋酸盐
1H NMR(600MHz,DMSO)δ8.29(d,J＝7.5Hz,1H),8.17(d,J＝7.7Hz,1H),8.09(d,J＝7.4Hz,1H),8.03(d,J＝7.2Hz,1H),7.96(d,J＝7.6Hz,1H),7.90(d,J＝8.7Hz,1H),7.86(d,J＝7.9Hz,1H),7.73(d,J＝8.5Hz,1H),7.58(d,J＝6.4Hz,1H),7.25–7.19(m,4H),7.18–7.12(m,2H),6.71(s,1H),4.54–4.44(m,2H),4.37(q,J＝8.3Hz,1H),4.33–4.21(m,5H),4.19–4.10(m,2H),4.09–4.02(m,1H),3.64–3.53(m,5H),3.51–3.45(m,2H),3.00(dd,J＝9.6Hz,1H),2.89–2.82(m,2H),2.80–2.71(m,3H),2.46–2.36(m,2H),2.24(t,J＝7.4Hz,2H),2.08(t,J＝8.0Hz,2H),2.00–1.70(m,18H,AcOH),1.69–1.64(m,2H),1.63–1.56(m,2H),1.55–1.47(m,3H),1.34(s,2H),1.16(t,J＝6.9Hz,6H),0.87–0.76(m,12H).
化合物26:Pro-Ala-Ala-Glu-Pro-Val-Pro-Leu醋酸盐
1H NMR(600MHz,DMSO)δ8.25(s,1H),8.09(d,J＝7.5Hz,1H),7.94(d,J＝7.6Hz,1H),7.89(d,J＝8.3Hz,2H),4.53–4.46(m,1H),4.39(dd,J＝8.3,4.2Hz,1H),4.34(dd,J＝8.4,3.8Hz,1H),4.31–4.19(m,3H),4.13(dd,J＝15.1,7.7Hz,1H),3.71–3.49(m,5H),2.94–2.77(m,2H),2.33–2.20(m,2H),2.06–1.77(m,13H,AcOH),1.77–1.56(m,6H),1.46(t,J＝7.3Hz,2H),1.25–1.11(m,6H),0.95–0.76(m,12H).
化合物27:Ala-Glu-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp醋酸盐
1H NMR(600MHz,D 2O)δ4.61(dd,J＝9.6,4.4Hz,1H),4.44–4.22(m,8H),4.09–3.99(m,2H),3.85–3.72(m,2H),3.69–3.57(m,2H),2.94(t,J＝7.4Hz,2H),2.79–2.63(m,2H),2.43– 2.16(m,7H),2.09–1.74(m,19H,AcOH),1.73–1.47(m,7H),1.48–1.13(m,6H),1.00–0.79(m,21H).
化合物28:Pro-Val-Pro-Leu-Val-Lys-Gln-Asp醋酸盐
1H NMR(600MHz,DMSO)δ8.28(d,J＝7.8Hz,1H),8.13–7.87(m,3H),7.57(d,J＝6.4Hz,1H),7.50(d,J＝8.7Hz,1H),7.15(s,1H),6.69(s,1H),4.40–4.31(m,2H),4.29–4.21(m,2H),4.19–4.02(m,3H),3.67–3.60(m,1H),3.59–3.53(m,2H),2.92–2.85(m,1H),2.80–2.70(m,3H),2.47–2.38(m,1H),2.38–2.31(m,1H),2.08(t,J＝7.9Hz,2H),2.04–1.76(m,14H,AcOH),1.74–1.41(m,11H),1.41–1.14(m,2H),0.91–0.74(m,18H).
化合物30:Ala-Glu-Pro-Val-Pro-Leu醋酸盐
1H NMR(600MHz,DMSO)δ8.29(d,J＝7.1Hz,1H),7.89(d,J＝8.6Hz,1H),7.79(d,J＝8.0Hz,1H),4.54(d,J＝5.4Hz,1H),4.42–4.38(m,1H),4.37–4.28(m,2H),4.09(dd,J＝14.2,8.5Hz,1H),3.66–3.46(m,5H),2.28(t,J＝7.4Hz,2H),2.13–1.71(m,16H,AcOH),1.71–1.58(m,2H),1.53–1.32(m,2H),1.18(d,J＝6.9Hz,3H),0.90–0.83(m,9H),0.80(d,J＝6.6Hz,3H).
化合物31:Glu-Pro-Val-Pro-Leu醋酸盐
1H NMR(600MHz,DMSO)δ7.94–7.86(m,2H),4.44(dd,J＝8.3,4.5Hz,1H),4.37–4.27(m,2H),4.13(q,J＝7.6Hz,1H),3.74–3.69(m,1H),3.66–3.42(m,4H),2.40–2.25(m,2H),2.06–1.58(m,17H,AcOH),1.46(t,J＝7.3Hz,2H),0.89–0.84(m,9H),0.81(d,J＝6.5Hz,3H).
化合物32:Pro-Val-Pro-Leu醋酸盐
1H NMR(600MHz,DMSO)δ8.15(d,J＝9.0Hz,1H),7.91(d,J＝8.0Hz,1H),4.35(dt,J＝10.1,5.2Hz,2H),4.22–4.02(m,1H),3.67–3.59(m,2H),3.56(q,J＝13.2,9.3Hz,1H),2.96–2.88(m,1H),2.86–2.76(m,1H),2.05–1.59(m,11H,AcOH),1.59–1.42(m,2H),0.91–0.72(m,12H).
化合物34:Pro-Ala-Ala-Glu-Pro-Val醋酸盐
1H NMR(600MHz,DMSO)δ8.01(d,J＝7.3Hz,1H),7.14(d,J＝7.8Hz,1H),4.49(q,J＝6.8Hz,1H),4.31–4.22(m,3H),3.75(dd,J＝7.8,4.7Hz,1H),3.70–3.61(m,2H),2.88–2.81(m,1H),2.81–2.75(m,1H),2.27–2.13(m,2H),2.01–1.64(m,18H,AcOH),1.58(p,J＝6.9Hz,2H),1.17(dd,J＝10.2,7.1Hz,6H),0.78(dd,J＝6.9,3.5Hz,6H).
化合物35:Pro-Ala-Ala-Glu-Pro醋酸盐
1H NMR(600MHz,DMSO)δ8.28(s,1H),8.12(d,J＝7.5Hz,1H),7.90(d,J＝7.7Hz,1H),4.49(dd,J＝13.7,7.7Hz,1H),4.35–4.13(m,3H),3.76–3.50(m,3H),2.97–2.79(m,2H),2.33–2.18(m,2H),2.16–1.77(m,10H,AcOH),1.73–1.58(m,4H),1.26–1.13(m,6H).
化合物45:Ala-Glu-Pro-Val醋酸盐
1H NMR(600MHz,DMSO)δ8.36(d,J＝7.4Hz,1H),7.46(d,J＝8.0Hz,1H),4.65–4.53(m,1H),4.35(dd,J＝8.6,3.4Hz,1H),3.89(dd,J＝7.9,5.4Hz,1H),3.74–3.53(m,3H),2.38–2.19(m,2H),2.05–1.64(m,10H,AcOH),1.27–1.17(m,3H),0.83(d,J＝7.1Hz,6H).
化合物48:Pro-Ala-Gly-Glu-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp醋酸盐
1H NMR(600MHz,DMSO)δ8.33(d,J＝7.6Hz,1H),8.27(d,1H),8.21(t,J＝5.8Hz,1H),8.10 (d,J＝7.5Hz,1H),8.05–7.97(m,2H),7.93(d,J＝8.5Hz,1H),7.57(d,J＝6.4Hz,1H),7.51(d,J＝8.8Hz,1H),7.16(s,1H),6.70(s,1H),4.53–4.47(m,1H),4.38(dd,J＝8.2,4.2Hz,1H),4.35–4.20(m,5H),4.18–4.09(m,2H),4.06(s,1H),3.75–3.50(m,8H),2.93–2.69(m,5H),2.45–2.41(m,1H),2.41–2.30(m,1H),2.25(t,J＝7.3Hz,2H),2.08(t,J＝7.9Hz,2H),2.01–1.76(m,15H,AcOH),1.74–1.53(m,8H),1.53–1.40(m,4H),1.33(d,J＝7.2Hz,2H),1.20(t,J＝9.0Hz,3H),0.91–0.73(m,18H).
化合物50:Pro-Ala-Ala-Glu-Pro-Val-Pro-leu-Val-Lys-Gln-Asn醋酸盐
1H NMR(600MHz,DMSO)δ8.49(d,J＝7.8Hz,1H),8.17–8.06(m,2H),8.05–7.99(m,2H),7.97(d,J＝7.7Hz,1H),7.92(d,J＝8.7Hz,1H),7.59(s,1H),7.54(d,J＝8.8Hz,1H),7.37(d,J＝6.9Hz,1H),7.10(s,1H),6.68(d,J＝15.7Hz,2H),4.48(d,J＝5.1Hz,1H),4.39(dd,J＝8.1,4.3Hz,1H),4.35–4.19(m,6H),4.17–4.10(m,1H),4.05–3.97(m,2H),3.67–3.48(m,6H),2.92–2.78(m,2H),2.78–2.66(m,3H),2.44–2.34(m,1H),2.33–2.20(m,3H),2.07(t,J＝7.8Hz,2H),2.01–1.76(m,15H,AcOH),1.75–1.62(m,6H),1.62–1.53(m,3H),1.49–1.29(m,6H),1.19–1.12(m,6H),0.91–0.73(m,18H).
化合物51:Pro-Ala-Ala-Glu-Pro-Val-Pro-leu-Val-Val-Gln-Asp醋酸盐
1H NMR(600MHz,DMSO)δ8.51(d,J＝7.6Hz,1H),8.08(d,J＝7.3Hz,1H),8.03–7.96(m,2H),7.95–7.89(m,2H),7.78(d,J＝8.9Hz,1H),7.75–7.69(m,1H),7.61(d,J＝8.8Hz,1H),7.18(s,1H),6.68(s,1H),4.51(d,J＝5.4Hz,1H),4.40(dd,J＝8.2,4.2Hz,1H),4.35–4.12(m,10H),4.01–3.95(m,1H),3.69–3.51(m,4H),3.09(d,J＝6.4Hz,2H),2.45–2.40(m,1H),2.39–2.34(m,1H),2.27(dd,J＝13.4,6.7Hz,2H),2.21–2.13(m,1H),2.06(t,J＝8.0Hz,2H),2.02–1.75(m,18H,AcOH),1.75–1.58(m,5H),1.44(t,J＝7.3Hz,2H),1.22(d,J＝7.1Hz,3H),1.16(d,J＝7.0Hz,3H),0.88(d,J＝6.7Hz,3H),0.85(dd,J＝6.5,2.7Hz,6H),0.83–0.77(m,15H).
化合物52:Pro-Ala-Ala-Glu-Pro-Val-Pro-Val-Val-Lys-Gln-Asp醋酸盐
1H NMR(600MHz,DMSO)δ8.28(s,1H),8.17(d,J＝8.0Hz,1H),8.09(d,J＝7.5Hz,1H),7.99–7.92(m,2H),7.87(dd,J＝16.4,8.8Hz,2H),7.68(d,J＝8.5Hz,1H),7.57(d,J＝6.1Hz,1H),7.17(s,1H),6.70(s,1H),4.50–4.45(m,1H),4.44–4.35(m,2H),4.32–4.21(m,4H),4.18–4.10(m,3H),4.08–3.99(m,1H),3.66–3.49(m,6H),2.90–2.82(m,1H),2.82–2.72(m,3H),2.46–2.30(m,2H),2.27–2.23(m,1H),2.07(t,J＝7.9Hz,2H),2.01–1.76(m,19H,AcOH),1.75–1.60(m,5H),1.60–1.45(m,5H),1.39–1.25(m,2H),1.20–1.10(m,6H),0.88(d,J＝6.6Hz,3H),0.86–0.76(m,15H).
化合物56:Pro-Ile-Ala-Glu-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp醋酸盐
1H NMR(600MHz,DMSO)δ8.27(d,J＝7.8Hz,1H),8.17(d,J＝7.3Hz,1H),8.10(d,J＝9.4Hz,1H),8.05–7.97(m,2H),7.92(q,J＝13.5,8.1Hz,2H),7.57(d,J＝6.3Hz,1H),7.51(d,J＝8.8Hz,1H),7.16(s,1H),6.70(s,1H),4.54–4.45(m,1H),4.39(dd,J＝8.3,4.2Hz,1H),4.35–4.22(m,5H),4.21–4.09(m,3H),4.10–4.00(m,1H),3.67–3.48(m,6H),3.41–3.16(m,4H),2.95–2.86(m,1H),2.81–2.68(m,3H),2.47–2.41(m,1H),2.41–2.32(m,1H),2.25(t,J＝7.6Hz,2H),2.08(t,J＝7.9Hz,2H),2.02–1.75(m,19H,AcOH),1.75–1.55(m,9H),1.54–1.36(m,5H),1.33(d,J＝7.0Hz,2H),1.15(d,J＝7.1Hz,3H),0.89–0.784(m,8H),0.82–0.76(m,14H).
化合物57:Pro-Ala-Ala-Glu-Pro-Val-Pro-Leu-Val-Lys-Glu-Asp醋酸盐
1H NMR(600MHz,DMSO)δ8.31(d,J＝7.8Hz,1H),8.20(d,J＝7.8Hz,1H),8.09(d,J＝7.5Hz,1H),8.03(t,J＝7.7Hz,2H),7.96–7.87(m,2H),7.58(d,J＝6.3Hz,1H),7.49(d,J＝8.8Hz, 1H),4.49(q,J＝5.3Hz,1H),4.39(dd,J＝8.2,4.4Hz,1H),4.35–4.20(m,7H),4.19–4.12(m,2H),4.10–4.00(m,1H),3.67–3.49(m,7H),2.91–2.84(m,2H),2.84–2.70(m,4H),2.46–2.38(m,1H),2.38–2.33(m,1H),2.29–2.18(m,4H),2.04–1.76(m,19H,AcOH),1.76–1.55(m,8H),1.55–1.41(m,5H),1.32(s,2H),1.21–1.11(m,6H),0.94–0.73(m,18H).
化合物58:Pro-Ala-Ala-Glu-Pro-Val-Pro-Leu-Val-Lys-Asn-Asp醋酸盐
1H NMR(600MHz,DMSO)δ8.28(d,J＝7.1Hz,1H),8.19(d,J＝7.7Hz,1H),8.09(d,J＝7.5Hz,1H),8.04(d,J＝7.7Hz,1H),8.00–7.89(m,3H),7.50(d,J＝8.6Hz,1H),7.44(d,J＝6.0Hz,1H),7.30(s,1H),6.80(s,1H),4.56–4.46(m,2H),4.38(dd,J＝8.1,4.3Hz,1H),4.35–4.20(m,6H),4.18–4.12(m,1H),4.04–4.00(m,1H),3.66–3.49(m,6H),2.93–2.84(m,1H),2.83–2.71(m,3H),2.43–2.31(m,3H),2.25(t,J＝8.0Hz,2H),2.04–1.57(m,26H,AcOH),1.56–1.46(m,3H),1.46–1.43(m,2H),1.35–1.27(m,2H),1.20–1.10(m,6H),0.86(dd,J＝14.8,7.9Hz,9H),0.81–0.76(m,9H).
化合物60:Pro-Ala-Ala-Glu-Pro-Val-Pro-Leu-Val-Lys-Gln-Glu醋酸盐
1H NMR(600MHz,DMSO)δ8.51(d,J＝8.0Hz,1H),8.17(d,J＝7.8Hz,1H),8.10(d,J＝7.4Hz,1H),8.04(d,J＝6.2Hz,2H),7.96(d,J＝7.7Hz,1H),7.93(d,J＝8.5Hz,1H),7.52(d,J＝8.8Hz,1H),7.34(d,J＝6.6Hz,1H),7.11(s,1H),6.71(s,1H),4.48(d,J＝5.4Hz,1H),4.39(dd,J＝8.1,4.3Hz,1H),4.34–4.18(m,7H),4.18–4.10(m,1H),4.07–3.99(m,1H),3.83(q,J＝12.8,6.5Hz,2H),3.69–3.48(m,7H),2.92–2.83(m,1H),2.81–2.62(m,4H),2.31–2.02(m,7H),2.01–1.75(m,22H,AcOH),1.74–1.25(m,16H),1.20–1.11(m,6H),0.91–0.83(m,9H),0.81–0.76(m,9H).
化合物62:Pro-Ala-Ile-Glu-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp醋酸盐
1H NMR(600MHz,DMSO)δ8.31–8.23(m,1H),8.20(d,J＝7.5Hz,1H),8.03(d,J＝7.7Hz,2H),7.99(d,J＝7.6Hz,1H),7.93–7.85(m,2H),7.57(d,J＝6.6Hz,1H),7.51(d,J＝8.1Hz,1H),7.16(s,1H),6.70(s,1H),4.49(d,J＝5.5Hz,1H),4.41–4.20(m,6H),4.19–4.00(m,5H),3.71–3.61(m,2H),3.61–3.47(m,4H),2.91–2.73(m,4H),2.45–2.39(m,1H),2.39–2.29(m,1H),2.24(t,J＝7.1Hz,2H),2.07(t,J＝7.9Hz,2H),2.02–1.76(m,20H,AcOH),1.74–1.47(m,12H),1.47–1.29(m,5H),1.16(d,J＝6.9Hz,3H),1.04(d,J＝8.4Hz,1H),0.89–0.84(m,8H),0.83–0.72(m,16H).
化合物66:Pro-Ala-Ala-Glu-Pro-Val-Pro-Leu-Val-Arg-Gln-Asp醋酸盐
1H NMR(600MHz,DMSO)δ9.30(s,1H),8.39(d,J＝8.7Hz,1H),8.19(d,J＝7.9Hz,1H),8.09(d,J＝7.4Hz,1H),8.03(d,J＝8.1Hz,1H),7.98(d,J＝7.0Hz,1H),7.96–7.89(m,3H),7.52(d,J＝8.9Hz,1H),7.11(s,1H),6.69(s,1H),4.49(d,J＝5.3Hz,1H),4.39(dd,J＝8.2,4.3Hz,1H),4.34(d,J＝4.3Hz,1H),4.31–4.21(m,5H),4.20–4.10(m,3H),3.68–3.49(m,6H),3.13(d,J＝6.1Hz,1H),3.00–2.93(m,1H),2.92–2.85(m,1H),2.83–2.76(m,1H),2.41–2.32(m,1H),2.25(t,J＝8.2Hz,2H),2.09–2.03(m,2H),2.02–1.76(m,20H,AcOH),1.74–1.70(m,1H),1.69–1.56(m,7H),1.55–1.39(m,4H),1.22–1.11(m,6H),0.93–0.83(m,9H),0.83–0.77(m,9H).
化合物73:Val-Pro-Leu-Val醋酸盐
1H NMR(600MHz,MeOD)δ4.54–4.49(m,1H),4.40(dd,J＝9.2,6.0Hz,1H),4.18(d,J＝5.0Hz,1H),3.99(d,J＝5.3Hz,1H),3.69(dd,J＝10.7,5.0Hz,1H),3.65–3.58(m,1H),2.32–2.20(m,2H),2.19–2.01(m,3H),2.00–1.86(m,4H,AcOH),1.85–1.75(m,1H),1.69–1.54(m,2H), 1.12(d,J＝7.0Hz,3H),1.03–1.01(m,3H),0.95(d,J＝6.6Hz,3H),0.93–0.89(m,9H).
实施例2：多肽(化合物1)对乙醇诱导的小鼠胃溃疡模型的抗溃疡作用
1、实验动物：SPF级C57BL/6小鼠，成都药康生物科技有限公司，动物许可证号：
SCXK(川)2020-034
2、方法：实验动物适应性喂养后，小鼠按随机分组法分成4组，每组10只，即：对照组(空白组)、模型组(无水乙醇造模)、阳性药物组(替普瑞酮160mg/kg)，化合物1组(1mg/kg)。动物分组后，对照组及模型组给予纯水治疗外，其它给药组给予相应药物治疗，每天一次，连续8天。第7天给药后所有动物禁食不禁水24h，于第8天给药30min后对各组(除对照组外)小鼠经口灌胃给予0.15mL的无水乙醇造模，1h后CO 2过量吸入处死动物，结扎胃贲门和夹闭幽门，摘取全胃。向胃体内注入1％甲醛溶液1mL，结扎贲门，取出胃后即放入1％甲醛溶液中固定25min。沿胃大弯剪开，生理盐水清洗干净胃内容物，平铺后计算溃疡指数。溃疡指数计算方法：条索状损伤长度大于1mm者，测量其长度，每毫米计1分；若其宽度大于1mm，将计分按宽度的毫米数加倍；长度小于1mm计0.5分，将计分相加得出该动物的溃疡指数。
3、结果：
在小鼠乙醇诱导的胃溃疡模型中，化合物1每天一次连续8天经口灌胃可显著使小鼠胃溃疡指数降低，且效果优于阳性药物组(表6)，结果表明化合物1治疗胃溃疡作用显著。实验结果参见图2。
表6.受试药对乙醇致胃溃疡小鼠溃疡指数的影响(n＝10，
Figure PCTCN2021103943-appb-000004
)
Figure PCTCN2021103943-appb-000005
注：与对照组相比， ##P<0.01；与模型组相比，*P<0.05，**P<0.01
实施例3：实施例1获得的部分多肽样品对乙醇诱导的小鼠胃溃疡模型的抗溃疡作用
1、实验动物：SPF级C57BL/6小鼠，成都药康生物科技有限公司，动物许可证号：
SCXK(川)2020-034
2、方法：
实验动物适应性喂养后，实验前1天给药后所有动物开始禁食不禁水24h。造模前实验 小鼠随机分组：空白组5只、模型组10只、各给药组每组10只，除空白组及模型组灌胃给予纯化水以外，给药组均按照0.2mg/kg的给药剂量灌胃给予不同受试样品，给药1小时后，各组小鼠经口灌胃给予0.9ml/kg的无水乙醇造模，1h后利用脱颈法处死动物，结扎胃贲门和夹闭幽门，摘取全胃。向胃体内注入1％甲醛溶液1mL，结扎贲门，取出胃后即放入1％甲醛溶液中。浸泡30min后取出胃组织，沿胃大弯剪开，使用生理盐水冲洗干净胃内容物，平铺后观察并测定小鼠胃黏膜的损伤，计算溃疡指数和溃疡抑制率。
溃疡指数计算方法：条索状损伤长度大于1mm者，测量其长度，每毫米计1分；若其宽度大于1mm，将计分按宽度的毫米数加倍；长度小于1mm计0.5分，将计分相加得出该动物的溃疡指数。
溃疡抑制率＝(模型组溃疡指数-给药组溃疡指数)/模型组溃疡指数×100％；
相对溃疡抑制率＝(测试化合物溃疡抑制率)/(化合物1溃疡抑制率)。
3、结果：表7示出了本发明化合物的相对溃疡抑制率
表7小鼠乙醇造模单次给药抗溃疡活性
Figure PCTCN2021103943-appb-000006
*注释：
每个化合物的抗溃疡作用经多批试验完成。为便于比较，抗溃疡活性以相对溃疡抑制率(每批试验都以化合物1作为对照组)的平均值表示。
相对溃疡抑制率＝(测试化合物溃疡抑制率)/(化合物1溃疡抑制率)
相对溃疡抑制率>1.20，表示为“++++”；
相对溃疡抑制率为0.9-1.20，表示为“+++”；
相对溃疡抑制率为0.6–0.9，表示为“++”；
相对溃疡抑制率为0.3–0.6，表示为“+”；
0<相对溃疡抑制率<0.3，表示为“/”(极低活性)；
相对溃疡抑制率<0，表示为“-”；
ND表示未与化合物1进行对比。
实施例4：化合物1对小鼠胃类器官的影响
方法：用健康野生型小鼠的胃窦上皮组织进行类器官(organoid)培养，体视镜下将小鼠胃窦上皮层与肌肉层分离，剪成0.5mm左右大小的小块，放入到2.5mM的EDTA/DPBS中4℃条件下进行消化，约1h。将消化后的胃腺上皮细胞过滤离心并弃掉上清，所得的细胞沉淀重悬于基质胶中，在培养皿中加入含生长因子的类器官培养基中进行培养，对照组除生长因子外不加入受试药物，实验组在与对照组相同培养条件加入化合物1，用倒置显微镜观察拍照其在Day1、Day3、Day5和Day7时成球和生长状况，观察化合物1对类器官生长的影响。
结果：与对照组相比，随着时间的推移，在含有化合物1的培养基中类器官的生长速率明显加快，通过对类器官直径进行定量分析，结果表明化合物1处理的类器官体积明显大于对照组(参见图3，图3中与对照组相比，*代表p<0.05)，可以显著促进类器官的增殖(统计数据也均有显著性差异)。提取化合物1处理的类器官RNA，通过实时荧光定量PCR检测分化相关的基因表达发现，内分泌细胞标志物SST和Gastrin、颈部粘液细胞标志物TFF2的mRNA水平明显上升(参见图4)。这些结果表明多肽化合物1能促进胃类器官的增殖和分化，说明化合物1参与并调控胃上皮干细胞的增殖和分化。
实施例5：化合物1对小鼠慢性萎缩性胃炎模型的治疗作用
方法：小鼠(Lgr5-GFP-CreERT小鼠)慢性萎缩性胃炎采用MNNG(N-甲基-N-硝基-N-亚硝基胍)联合雷尼替丁造模法：小鼠自由饮用含有MNNG(100mg/ml)的水溶液，同时每日固定时间给小鼠按照150mg/kg的剂量灌胃雷尼替丁(8mg/ml)水溶液，持续20周。造模20周后在小鼠饮用普通蒸馏水基础上，每日灌胃化合物1(5mg/kg)，给药2周后观察化合物1对慢性萎缩性胃炎的治疗效果。
结果：组织染色结果显示，模型组小鼠的胃体部和胃窦部腺体结构紊乱，伴随着胃酸细胞(H +-K +-ATPase阳性)细胞数目的减少，并且胃窦部的粘膜上皮高度下降。通过2周的化合物1治疗，与模型组相比，化合物1组小鼠的胃腺结构恢复正常，胃酸细胞数目显著增多，胃窦部粘膜高度基本恢复至正常状态，结果参见图5。这些结果表明化合物1能够促进小鼠慢性萎缩性胃炎的修复。
在慢性萎缩性胃炎中，上皮的修复过程需要干细胞的参与。我们通过免疫荧光染色技术检测胃上皮干细胞(Lgr5 +和AQP5 +)的改变。实验结果表明，与对照组相比，模型组的干细胞数目显著减少；与模型组相比，化合物1治疗组的增殖活跃的干细胞数目(PCNA阳性的Lgr5 +细胞和Ki67阳性的AQP5 +细胞)也存在显著增加，统计结果均存在显著性差异，结果参见参看图6。
实施例6：化合物1对慢性萎缩性胃炎大鼠的治疗作用
1、实验动物
SPF级SD大鼠85只，雌雄各半，体重160-280g，由斯贝福(北京)生物技术有限公司提供，许可证编号：SCXK(京)2016-0002。
2、实验方法
SD大鼠85只，适应性喂养1周后，随机分为空白对照组10只和慢性萎缩性胃炎造模组(简称CAG组)75只。空白对照组给予每日灌胃去离子水5ml/kg，正常饮食，自由饮水。CAG组给予每日灌胃120μg/mL的MNNG(N-甲基-N-硝基-N-亚硝基胍)水溶液，5ml/kg，自由食用0.03％雷尼替丁饲料，自由饮用2％水杨酸钠自由饮水。第15周起每周禁食18小时后灌胃高热淀粉糊(5ml/kg，60-70℃)，且当日不再灌胃MNNG，余处理同前。第18周起，每2周分别随机取非空白组雌雄大鼠各2只，取胃粘膜，病理切片观察至模型成功(固有腺体减少+肠化)，造模过程死亡19只大鼠。造模第25周将成模大鼠随机分为3组，模型组、化合物1高剂量组(3mg/kg)、化合物1低剂量组(1mg/kg)。正常对照组和模型组每日灌胃等量生理盐水，其它各组灌胃给予相应药物，给药8周。
试验期间每周定期检测大鼠体重变化，测定摄食量与饮水量，观察大鼠活动状态等。给药8周后，大鼠禁食不禁水24小时，10％水合氯醛3.5ml/kg腹腔注射麻醉。摘离全胃，迅速沿大弯侧剪开，生理盐水漂洗后，取全小弯侧及近大弯侧上至食管端下至十二指肠端的胃黏膜组织，于10％中性福尔马林液固定，常规石蜡包埋切片，常规HE染色，进行病理结果分析。
所有数据均采用SPSS23软件进行处理。计量资料以Shapiro-Wilk检验进行正态性检验，若符合正态分布，以均数加减标准差
Figure PCTCN2021103943-appb-000007
表示，多组间均值比较，采用单因素方差分析(One-way ANOVA)，方差齐者用LSD法进行组间比较，方差不齐者采用Dunnett’s T3法进行组间比较，P<0.05表示差异具有统计学意义。若不符合正态分布，进行秩和检验，多组间比较采用Kruskal-Wallis检验，组间两两比较采用Mann-Whitney检验。等级资料同非正态分布检验方法。P<0.05表示差异具有统计学意义。
3、结果
3.1大鼠一般情况
正常组大鼠体毛顺滑浓密，毛色洁白有光泽。活动度较高，对饲养笼搬动、投喂食水等活动以及声响反应度高。精神状态佳，灌胃、称重等操作时情绪稳定。模型组大鼠体毛枯槁稀疏，易脱落，毛色晦暗偏米黄。活动度低，喜蜷卧，对饲养笼搬动、投喂食水等活动以及声响反应度低，精神状态萎靡，灌胃、称重等操作时容易出现情绪波动以及抓咬撕挠实验操作者行为，化合物1低、高剂量组均有不同程度改善大鼠活动度、反应性等整体状态。
3.2大鼠体重情况
给药8周后，各组雌鼠体重无明显差异。与正常组相比，模型组雄鼠体重明显减轻，有统计学意义(P＜0.05)；与模型组相比，化合物1低剂量组、高剂量组,雄鼠体重明显升高，有统计学意义(P＜0.05)，其余各给药组体重差异无统计学意义，见表8。
表8.各组给药8周后体重(g)
组别 动物数量 雌鼠体重(均值) 雄鼠体重(均值)
空白对照组 10 364 602
模型组 12 369 446 #
化合物1高 14 396 637 *
化合物1低 14 374 532 *
注： #与正常组相比有显著性差异(P＜0.05)；*与模型组相比有显著性差异(P＜0.05)。
3.3病理结果
大鼠胃黏膜病理切片显微镜下观察：正常组大鼠黏膜各层结构清晰，腺体排列紧密、整齐，黏膜层未见慢性炎症；模型组大鼠胃黏膜见炎细胞浸润，黏膜肌增生，固有层腺体不同程度减少，排列稀疏不规则，部分腺体扩张，未见肠上皮化生，少数见异型增生。与正常组相比，模型组炎症及萎缩评分明显增高(P＜0.05)；与模型组相比，化合物1低、高剂量组可显著改善大鼠胃黏膜炎症、萎缩程度(P<0.05)。结果参见图7。
实施例7：化合物1对HaCAT细胞增殖的促进作用
方法：将HaCAT细胞浓度调整为1.0×10 5～5.0×10 5/mL进行传代培养，于37℃，5％CO 2条件下培养24～36小时用于生物学活性检测。用胰酶消化并收集细胞，用无血清培养基配成浓度2.5×10 4/mL接种于96孔细胞培养板中，每孔100μL，即2500个细胞/孔，于37℃，5％CO 2条件下培养过夜。再加入50μL用无血清培养基配制化合物溶液，使化合物1终浓度为0.4ug/mL；同时设置EGF对照组，即加入50μL用无血清培养基配制的重组人表皮生长因子(EGF)溶液，终浓度为100ng/mL；模型对照组，即加入等体积的无血清培养基。于37℃，5％CO 2条件下培养72小时，采用
Figure PCTCN2021103943-appb-000008
试剂盒检测HaCAT细胞株增殖情况。
结果：如图8所示，0.4μg/mL化合物1对HaCAT细胞具有显著的促增殖作用，表明化合物1对于表皮生长和皮肤损伤修复有较好的作用。图8中，与模型对照组相比，*代表p<0.05，**代表p<0.01。
实施例8：化合物1及化合物26对斑马鱼血管损伤的修复作用
方法：转基因血管绿色荧光斑马鱼(Fli-1)的血管被绿色荧光蛋白标记，在荧光显微镜下清晰可见(图9，图9中虚线框(原图为黄色)为分析部位肠下血管，箭头(原图为白色)指向为肠下血管分支)，成为观察血管变化的模式生物。Fli-1斑马鱼，以自然成对交配繁殖方式进行。随机选取受精后1天的Fli-1斑马鱼于6孔板中，每孔(实验组)30尾。正常对照组用标准稀释水处理斑马鱼，其余各实验组水溶给予辛伐他汀诱导3小时建立斑马鱼微血管缺失模型。3小时后，所有组别水溶液替换为标准稀释水，终止辛伐他汀诱导。受试药物组分别静脉注射给予化合物1(500ng/尾)或化合物26(500ng/尾)，28℃处理2天。每组随机选取10尾斑马鱼置于荧光显微镜下拍照，用NIS-Elements D 3.20高级图像处理软件分析并采集数据，分析肠下血管分支数。
结果：模型组可见肠下血管分支数量减少。化合物1和化合物26静脉注射可以显著逆转辛伐他汀造成的肠下微血管缺失，恢复斑马鱼肠下血管的分支数量。表明化合物1和化合物26可以促进血管的损伤后修复(图10和11)。图11中，与模型对照组比较，**p<0.01，***p<0.001。
实施例9：部分多肽样品的胃、肠稳定性试验
方法：取待测样品(化合物1、化合物26、化合物27、化合物28及EGF)各1mg，加水1ml溶解。取样品溶液100ul，加水900ul，混匀，作为对照品溶液。取样品溶液各100ul，分别加人工胃液(W)、人工肠液(X)、聚维酮碘溶液(I)、过氧化氢溶液(O)900ul，37℃恒温水浴1小时，放冷，滤过，作为供试品溶液，用高效液相色谱法分别检测处理前及处理后的样品峰面积,试验结果以样品的峰面积进行对比计算。以水稀释后不做任何处理的原液作为对照，对比统计其他供试品溶液相应位置的峰面积(含量)变化情况。
表9多肽样品的胃、肠稳定性试验
编号 W保留％ X保留％ I保留％ O保留％
化合物1 96 0 101 100
化合物26 100 103 101 102
化合物27 97 0 100 101
化合物28 96 0 99 99
EGF 0 0 0 0
注：W表示人工胃液，X表示人工肠液，I表示聚维酮碘溶液，O表示过氧化氢溶液
结果：如表9所示，4个供试品(化合物1、26、27和28)对人工胃液(W)、聚维酮碘溶液(I)和过氧化氢溶液(O)都100％保留，说明很稳定；化合物26对人工肠液(X)也极其的稳定；EGF在胃液和肠液里都没有保留，说明在胃液和肠液中不稳定，聚维酮碘溶液和过氧化氢溶液消毒后涂抹EGF外用也会被破坏。
实施例10：化合物1对阿司匹林诱导的大鼠胃溃疡的影响
方法：SD大鼠适应性喂养后，按Excel完全随机分组法分成3组(每组10只)，即：对照组、模型组、化合物1(0.3mg/kg)组。动物分组后，开始给予相应处理(对照组及模型组给予同体积纯化水，化合物1组给予相应药物治疗)，每天一次，连续8天。第7天给药后，所有动物禁食不禁水24h。第8天除对照组外，化合物1组及模型组大鼠给药(或 水)30min后经口灌胃给予250mg/kg阿司匹林溶液造模。造模4h后，处死动物，结扎贲门和夹闭幽门，摘取全胃。向胃体内注入1％甲醛溶液8mL，结扎幽门，取出胃后即放入1％甲醛溶液中固定，30min后沿胃大弯剪开，生理盐水清洗干净胃内容物，将胃平铺后，观察并拍摄胃全景照片，测定溃疡面积。
结果：每天1次连续8天经口灌胃给予化合物1(0.3mg/kg)对阿司匹林诱导胃溃疡模型大鼠的体重变化无明显影响，化合物1减少了阿司匹林诱导的胃溃疡模型大鼠胃部出血点，显著降低大鼠胃溃疡面积。表10示出了化合物1对阿司匹林诱导的大鼠胃溃疡的影响。
表10化合物1对阿司匹林诱导的大鼠胃溃疡的影响。
组别/给药 溃疡面积(mm 2)
对照组： 7.42±7.32
模型组： 140.36±82.40 ##
化合物1组： 20.55±10.45**
注： ##与对照组相比，p<0.01；**与模型组相比，p<0.01
尽管本发明披露了上述实施例，但本发明的实施方式并不限于上述实施例，其他的任何未背离本发明的改变、修饰、替代、组合、简化，均应为等效的置换方式均包含在本发明的保护范围之内。

Claims (24)

  1. 式(I)的化合物或其生理学上相容的盐，其中所述式(I)的化合物如下：
    H-Xaa 1-Xaa 2-Xaa 3-Xaa 4-Xaa 5-Val-Xaa 6-Xaa 7-Xaa 8-Xaa 9-Xaa 10-Xaa 11-OH(I)
    其中
    Xaa 1为Pro、Gly、Ala或缺失；
    Xaa 2为Ala、Leu、Ile、Gly、Cys、Ser或缺失；
    Xaa 3为Ala、Pro、Gly、Leu、Ile、Cys、Ser或缺失；
    Xaa 4为Glu、Gln、Asp、Asn、Leu、Ile、Val或缺失；
    Xaa 5为Pro、Gly、Ala、Val或缺失；
    Xaa 6为Pro、Gly或Ala；
    Xaa 7为Leu、Phe、Val、Ala、Tyr、Glu、Lys、Asp、Ile、Met或缺失；
    Xaa 8为Val、Leu、Gln、Ile、Met或缺失；
    Xaa 9为Lys、Arg、His、Asp、Val或缺失；
    Xaa 10为Glu、Gln、Asp、Asn或缺失；以及
    Xaa 11为Glu、Asp、Asn、Gln或缺失，
    条件是：Xaa 1、Xaa 2、Xaa 3、Xaa 4、Xaa 5、Xaa 7、Xaa 8、Xaa 9、Xaa 10和Xaa 11中至多有9个是缺失的。
  2. 权利要求1的化合物或其生理学上相容的盐，其中X aa5为Pro。
  3. 权利要求1的化合物或其生理学上相容的盐，其中X aa6为Pro。
  4. 权利要求1的化合物或其生理学上相容的盐，其中X aa7为Leu。
  5. 权利要求1-5中任一项的化合物或其生理学上相容的盐，其中式(I)化合物具有以下式(II)结构，
    H-Xaa 1-Xaa 2-Xaa 3-Xaa 4-Pro-Val-Pro-Leu-Xaa 8-Xaa 9-Xaa 10-Xaa 11-OH(II)，
    其中
    Xaa 1为Pro、Gly、Ala或缺失；
    Xaa 2为Ala、Leu、Ile、Gly、Cys、Ser或缺失；
    Xaa 3为Ala、Pro、Gly、Leu、Ile、Cys、Ser或缺失；
    Xaa 4为Glu、Gln、Asp、Asn、Leu、Ile、Val或缺失；
    Xaa 8为Val、Leu、Gln、Ile、Met或缺失；
    Xaa 9为Lys、Arg、His、Asp、Val或缺失；
    Xaa 10为Glu、Gln、Asp、Asn或缺失；以及
    Xaa 11为Glu、Asp、Asn、Gln或缺失。
  6. 权利要求1-5中任一项的化合物或其生理学上相容的盐，其中式(I)或(II)中的Xaa 1为Pro或缺失；优选为Pro。
  7. 权利要求1-6中任一项的化合物或其生理学上相容的盐，其中式(I)或(II)中的Xaa 2为Ala或缺失；优选为Ala。
  8. 权利要求1-7中任一项的化合物或其生理学上相容的盐，其中式(I)或(II)中的Xaa 3为Ala、Gly或缺失；优选为Ala。
  9. 权利要求1-8中任一项的化合物或其生理学上相容的盐，其中式(I)或(II)中的Xaa 4为Glu、Gln、Asp或缺失；优选为Glu或Gln。
  10. 权利要求1-9中任一项的化合物或其生理学上相容的盐，其中式(I)或(II)中的Xaa 8为Val或缺失；优选为Val。
  11. 权利要求1-10中任一项的化合物或其生理学上相容的盐，其中式(I)或(II)中的Xaa 9为Lys、Arg或缺失；优选为Lys或缺失。
  12. 权利要求1-11中任一项的化合物或其生理学上相容的盐，其中式(I)或(II)中的Xaa 10为Glu、Gln或缺失；优选为Gln。
  13. 权利要求1-12中任一项的化合物或其生理学上相容的盐，其中式(I)或(II)中的Xaa 11为Asp或缺失；优选为Asp。
  14. 权利要求1-13中任一项的化合物或其生理学上相容的盐，其中式(I)或(II)中的Xaa 1、Xaa 2、Xaa 3和Xaa 4中有一个是缺失的；或者有两个是缺失的；或者有三个是缺失的；或者都是缺失的。
  15. 权利要求1-14中任一项的化合物或其生理学上相容的盐，其中式(I)或(II)中的Xaa 8、Xaa 9、Xaa 10和Xaa 11中有一个是缺失的；或者有两个是缺失的；或者有三个是缺失的；或者都是缺失的。
  16. 权利要求1的化合物或其生理学上相容的盐，其中所述化合物选自：
    Pro-Ala-Ala-Glu-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物1)
    Gly-Ala-Ala-Glu-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物2)
    Pro-Ala-Ala-Glu-Pro-Val-Pro-Leu-Val-Lys(化合物3)
    Pro-Ala-Ala-Glu-Gly-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物4)
    Pro-Ala-Ala-Glu-Pro-Val-Gly-Leu-Val-Lys-Gln-Asp(化合物5)
    Pro-Ala-Ala-Glu-Pro-Val-Gly-Ala-Val-Lys-Gln-Asp(化合物6)
    Pro-Ala-Ala-Glu-Pro-Val-Gly-Val-Val-Lys-Gln-Asp(化合物7)
    Pro-Ala-Ala-Glu-Gly-Val-Gly-Leu-Val-Lys-Gln-Asp(化合物8)
    Pro-Ala-Ala-Glu-Gly-Val-Gly-Leu(化合物9)
    Pro-Ala-Ala-Gln-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物10)
    Pro-Ala-Ala-Asp-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物11)
    Pro-Ala-Ala-Glu-Pro-Val-Pro-Phe-Val-Lys-Gln-Asp(化合物12)
    Pro-Ala-Ala-Glu-Pro-Val-Pro-Tyr-Val-Lys-Gln-Asp(化合物13)
    Pro-Ala-Ala-Glu-Pro-Val-Gly-Leu-Val-Lys(化合物14)
    Pro-Ala-Ala-Glu-Pro-Val-Gly-Val-Val-Lys(化合物15)
    Pro-Ala-Ala-Glu-Pro-Val-Ala-Leu-Val-Lys(化合物16)
    Pro-Ala-Ala-Glu-Pro-Val-Ala-Val-Val-Lys(化合物17)
    Pro-Ala-Ala-Glu-Ala-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物18)
    Pro-Ala-Ala-Glu-Pro-Val-Ala-Leu-Val-Lys-Gln-Asp(化合物19)
    Pro-Ala-Ala-Glu-Ala-Val-Ala-Leu-Val-Lys-Gln-Asp(化合物20)
    Ala-Ala-Ala-Glu-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物21)
    Pro-Ala-Ala-Glu-Pro-Val-Pro-Phe-Val-Lys(化合物22)
    Pro-Ala-Ala-Glu-Pro-Val-Pro-Tyr-Val-Lys(化合物23)
    Pro-Ala-Ala-Glu-Pro-Val-Gly-Phe-Val-Lys(化合物24)
    Pro-Ala-Ala-Glu-Pro-Val-Ala-Phe-Val-Lys(化合物25)
    Pro-Ala-Ala-Glu-Pro-Val-Pro-Leu(化合物26)
    Ala-Glu-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物27)
    Pro-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物28)
    Pro-Ala-Ala-Glu-Pro-Val-Pro(化合物29)
    Ala-Glu-Pro-Val-Pro-Leu(化合物30)
    Glu-Pro-Val-Pro-Leu(化合物31)
    Pro-Val-Pro-Leu(化合物32)
    Ala-Ala-Glu-Pro-Val-Pro-Leu(化合物33)
    Pro-Ala-Ala-Glu-Pro-Val(化合物34)
    Pro-Ala-Ala-Glu-Pro(化合物35)
    Pro-Ala-Ala-Glu(化合物36)
    Ala-Ala-Glu-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物37)
    Glu-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物38)
    Val-Pro-Leu-Val-Lys-Gln-Asp(化合物39)
    Pro-Leu-Val-Lys-Gln-Asp(化合物40)
    Pro-Ala-Ala-Glu-Pro-Val-Pro-Ile-Val-Lys(化合物41)
    Pro-Ala-Ala-Glu-Pro-Val-Pro-Val-Val-Lys(化合物42)
    Pro-Ala-Ala-Glu-Pro-Val-Pro-Met-Val-Lys(化合物43)
    Ala-Glu-Pro-Val-Pro(化合物44)
    Ala-Glu-Pro-Val(化合物45)
    Pro-Ala-Ala-Asn-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物46)
    Pro-Ala-Ala-Leu-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物47)
    Pro-Ala-Gly-Glu-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物48)
    Pro-Ala-Pro-Glu-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物49)
    Pro-Ala-Ala-Glu-Pro-Val-Pro-leu-Val-Lys-Gln-Asn(化合物50)
    Pro-Ala-Ala-Glu-Pro-Val-Pro-leu-Val-Val-Gln-Asp(化合物51)
    Pro-Ala-Ala-Glu-Pro-Val-Pro-Val-Val-Lys-Gln-Asp(化合物52)
    Pro-Ala-Ala-Glu-Pro-Val-Pro-Ile-Val-Lys-Gln-Asp(化合物53)
    Pro-Ala-Ala-Glu-Pro-Val-Pro-Met-Val-Lys-Gln-Asp(化合物54)
    Pro-Leu-Ala-Glu-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物55)
    Pro-Ile-Ala-Glu-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物56)
    Pro-Ala-Ala-Glu-Pro-Val-Pro-Leu-Val-Lys-Glu-Asp(化合物57)
    Pro-Ala-Ala-Glu-Pro-Val-Pro-Leu-Val-Lys-Asn-Asp(化合物58)
    Pro-Ala-Ala-Glu-Pro-Val-Pro-Leu-Val-Lys-Asp-Asp(化合物59)
    Pro-Ala-Ala-Glu-Pro-Val-Pro-Leu-Val-Lys-Gln-Glu(化合物60)
    Pro-Ala-Leu-Glu-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物61)
    Pro-Ala-Ile-Glu-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物62)
    Pro-Ala-Ala-Glu-Pro-Val-Pro-Leu-Leu-Lys-Gln-Asp(化合物63)
    Pro-Ala-Ala-Glu-Pro-Val-Pro-Leu-Ile-Lys-Gln-Asp(化合物64)
    Pro-Ala-Ala-Glu-Pro-Val-Pro-Leu-Met-Lys-Gln-Asp(化合物65)
    Pro-Ala-Ala-Glu-Pro-Val-Pro-Leu-Val-Arg-Gln-Asp(化合物66)
    Pro-Ala-Ala-Val-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物67)
    Val-Pro-Leu-Val-Lys-Gln-Asp(化合物68)
    Gly-Ala-Ala-Val-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物69)
    Gly-Ala-Gly-Val-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物70)
    Gly-Ala-Gly-Val-Gly-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物71)
    Pro-Ala-Ala-Glu-Pro-Val-Ala-Phe-Val-Lys-Gln-Asp(化合物72)
    Val-Pro-Leu-Val(化合物73)。
  17. 权利要求6的化合物或其生理学上相容的盐，其中所述化合物选自
    Pro-Ala-Ala-Glu-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物1)；
    Pro-Ala-Ala-Glu-Pro-Val-Pro-Leu-Val-Lys(化合物3)；
    Pro-Ala-Ala-Gln-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物10)；
    Pro-Ala-Ala-Glu-Pro-Val-Pro-Leu(化合物26)；
    Ala-Glu-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物27)；
    Ala-Glu-Pro-Val-Pro-Leu(化合物30)；
    Glu-Pro-Val-Pro-Leu(化合物31)；
    Pro-Val-Pro-Leu(化合物32)；
    Pro-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物28)；
    Pro-Ala-Ala-Asp-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物11)；
    Pro-Ala-Gly-Glu-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物48)；
    Pro-Ala-Ala-Glu-Pro-Val-Pro-Leu-Val-Lys-Glu-Asp(化合物57)；或
    Glu-Pro-Val-Pro-Leu-Val-Lys-Gln-Asp(化合物38)。
  18. 权利要求1-17中任一项的化合物或其生理学上相容的盐在制备用于修复皮肤创伤或黏膜损伤的药物中的用途。
  19. 权利要求1-17中任一项的化合物或其生理学上相容的盐在制备用于调控干细胞增殖和分化的药物中的用途。
  20. 根据权利要求18的用途，其中所述的黏膜损伤是消化系统、呼吸系统等腔道内黏膜损伤。
  21. 权利要求20的用途，其中所述消化系统黏膜损伤与口腔、食道、胃肠疾病相关，所述 口腔疾病包括口腔溃疡、口腔炎、牙龈炎、牙周炎等；所述食道疾病包括食管炎、食管溃疡等；所述胃肠疾病包括慢性胃炎、慢性萎缩性胃炎、急性胃炎、胃十二指肠溃疡、功能性胃肠道疾病、消化不良、癌前病变、消化系统肿瘤、胃肠道出血、胃食管返流疾病、急慢性肠炎、溃疡性结肠炎、克罗恩病和放化疗引起的黏膜损伤；所述皮肤创伤与表皮炎症、机械及手术创面、烧伤及烫伤、溃疡、瘘管、褥疮、放化疗引起的皮肤损伤等疾病相关。
  22. 权利要求20的用途，其中所述消化系统黏膜损伤是由刺激性物质或药物引起的或应激状态引起的黏膜损伤。
  23. 权利要求1-17中任一项的化合物或其生理学上相容的盐在制备用于预防、减轻或治疗胃肠疾病或消除炎症水肿的药物中的用途。
  24. 一种药物、食品、保健品或化妆品、日化品组合物，所述组合物包括权利要求1-17中任一项的化合物或其生理学上相容的盐以及生理学上可接受的载体。
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