WO2020145564A1 - Composition permettant d'augmenter le taux de réussite d'une greffe de cheveux, contenant des exosomes dérivés de cellules souches utilisés comme principe actif - Google Patents

Composition permettant d'augmenter le taux de réussite d'une greffe de cheveux, contenant des exosomes dérivés de cellules souches utilisés comme principe actif Download PDF

Info

Publication number
WO2020145564A1
WO2020145564A1 PCT/KR2020/000001 KR2020000001W WO2020145564A1 WO 2020145564 A1 WO2020145564 A1 WO 2020145564A1 KR 2020000001 W KR2020000001 W KR 2020000001W WO 2020145564 A1 WO2020145564 A1 WO 2020145564A1
Authority
WO
WIPO (PCT)
Prior art keywords
hair
composition
increasing
scalp
rate
Prior art date
Application number
PCT/KR2020/000001
Other languages
English (en)
Korean (ko)
Inventor
조병성
이용원
김광일
Original Assignee
주식회사 엑소코바이오
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by 주식회사 엑소코바이오 filed Critical 주식회사 엑소코바이오
Publication of WO2020145564A1 publication Critical patent/WO2020145564A1/fr

Links

Images

Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
    • A61K35/12—Materials from mammals; Compositions comprising non-specified tissues or cells; Compositions comprising non-embryonic stem cells; Genetically modified cells
    • A61K35/28—Bone marrow; Haematopoietic stem cells; Mesenchymal stem cells of any origin, e.g. adipose-derived stem cells
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
    • A61K47/08—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite containing oxygen, e.g. ethers, acetals, ketones, quinones, aldehydes, peroxides
    • A61K47/10—Alcohols; Phenols; Salts thereof, e.g. glycerol; Polyethylene glycols [PEG]; Poloxamers; PEG/POE alkyl ethers
    • 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
    • A61K47/20—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite containing sulfur, e.g. dimethyl sulfoxide [DMSO], docusate, sodium lauryl sulfate or aminosulfonic 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
    • A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
    • A61K47/22—Heterocyclic compounds, e.g. ascorbic acid, tocopherol or pyrrolidones
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
    • A61K47/26—Carbohydrates, e.g. sugar alcohols, amino sugars, nucleic acids, mono-, di- or oligo-saccharides; Derivatives thereof, e.g. polysorbates, sorbitan fatty acid esters or glycyrrhizin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • 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
    • A61K47/36—Polysaccharides; Derivatives thereof, e.g. gums, starch, alginate, dextrin, hyaluronic acid, chitosan, inulin, agar or pectin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K8/00—Cosmetics or similar toiletry preparations
    • A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
    • A61K8/96—Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution
    • A61K8/98—Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution of animal origin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K8/00—Cosmetics or similar toiletry preparations
    • A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
    • A61K8/96—Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution
    • A61K8/98—Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution of animal origin
    • A61K8/981—Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution of animal origin of mammals or bird
    • 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
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/0012—Galenical forms characterised by the site of application
    • A61K9/0014—Skin, i.e. galenical aspects of topical compositions
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/0012—Galenical forms characterised by the site of application
    • A61K9/0019—Injectable compositions; Intramuscular, intravenous, arterial, subcutaneous administration; Compositions to be administered through the skin in an invasive manner
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/0012—Galenical forms characterised by the site of application
    • A61K9/0019—Injectable compositions; Intramuscular, intravenous, arterial, subcutaneous administration; Compositions to be administered through the skin in an invasive manner
    • A61K9/0021—Intradermal administration, e.g. through microneedle arrays, needleless injectors
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/14—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles
    • A61K9/19—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles lyophilised, i.e. freeze-dried, solutions or dispersions
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
    • A61Q90/00—Cosmetics or similar toiletry preparations for specific uses not provided for in other groups of this subclass
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K2800/00—Properties of cosmetic compositions or active ingredients thereof or formulation aids used therein and process related aspects
    • A61K2800/80—Process related aspects concerning the preparation of the cosmetic composition or the storage or application thereof
    • A61K2800/91—Injection
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
    • A61L27/36—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix
    • A61L27/3683—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix subjected to a specific treatment prior to implantation, e.g. decellularising, demineralising, grinding, cellular disruption/non-collagenous protein removal, anti-calcification, crosslinking, supercritical fluid extraction, enzyme treatment
    • A61L27/3687—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix subjected to a specific treatment prior to implantation, e.g. decellularising, demineralising, grinding, cellular disruption/non-collagenous protein removal, anti-calcification, crosslinking, supercritical fluid extraction, enzyme treatment characterised by the use of chemical agents in the treatment, e.g. specific enzymes, detergents, capping agents, crosslinkers, anticalcification agents

Definitions

  • compositions for increasing the success rate of hair transplant comprising an exosome derived from stem cells as an active ingredient.
  • the present invention relates to a method for increasing the success rate of hair transplantation using the composition for increasing the success rate of hair transplantation.
  • hair loss The exact mechanism of hair growth and hair loss has not been fully understood to date, but the causes of hair loss include decreased hair follicle function due to male hormone involvement, reduced hair follicle and hairball metabolic function, local blood flow disorder caused by scalp tension, malnutrition, It is very diverse due to stress, drug side effects, genetic factors, autoimmunity, local infection, chemicals, leukemia, tuberculosis and other diseases and abuse of hair products.
  • a prescription drug such as minoxidil or a drug to eat, such as finasteride, is prescribed, but if it is not effective by these drugs, a wig or a hair transplant is performed.
  • Hair transplants have less heterogeneity than wigs.
  • hair transplantation is actually the only alternative if the effect of a hair loss treatment is minimal or has side effects.
  • Artificial hair transplantation is a method of planting artificial hair made of polyethylene on the scalp one by one.
  • the transplanted artificial hair is a foreign substance, if the body rejects it, there is a problem in that the success rate of the hair transplant is lowered because the transplanted artificial hair falls out while the scalp area where the artificial hair is implanted exhibits redness.
  • the incision method has a short operation time and can transplant many hairs at once, but has side effects such as pain and scarring.
  • the non-incision method has the advantage that there is almost no swelling, pain, and scar after surgery.
  • hair follicles need to be quickly collected and transplanted individually, but the quality of the hair follicles varies depending on the skill of the person who collects the hair follicles. There is a problem that the transplantation success rate is lowered.
  • Extracellular vesicles are also called cell membrane-derived endoplasmic reticulum, ectomes, shedding vesicles, microparticles, exosomes, and in some cases, they are used separately from exosomes.
  • the exosome is a endoplasmic reticulum of several tens to hundreds of nanometers made of a double phospholipid membrane having the same structure as a cell membrane, and contains protein, nucleic acid (mRNA, miRNA, etc.) called exosome cargo.
  • Exosomal cargoes contain a wide range of signaling factors, which are known to be cell type specific and differently regulated according to the secretory cell environment.
  • Exosomes are intercellular signaling mediators secreted by cells, and various cellular signals transmitted through them regulate cell behavior including target cell activation, growth, migration, differentiation, dedifferentiation, apoptosis, and necrosis. Is known.
  • Exosomes contain specific genetic material and bioactive factors depending on the nature and condition of the derived cells. In the case of proliferating stem cell-derived exosomes, cell behavior such as cell migration, proliferation and differentiation is regulated and characteristics of stem cells related to tissue regeneration are reflected (Nature Review Immunology 2002 (2) 569-579).
  • the present inventors have been carrying out earnest research on new applications of stem cell-derived exosomes and grafting with medical or cosmetic techniques, and when treated with stem cell-derived exosomes in the hair transplantation process, the transplanted hairs will be eliminated.
  • the present invention was completed by confirming that the success rate of hair transplantation is reduced by reducing the phenomenon.
  • An object of the present invention is to provide a composition for increasing the success rate of hair transplantation, which includes exosomes derived from stem cells as an active ingredient.
  • Another object of the present invention is to provide a method for increasing the success rate of hair transplantation by using the composition for increasing the rate of successful hair transplantation.
  • the present invention provides a composition for increasing the success rate of hair transplantation comprising exosomes derived from stem cells as an active ingredient and a method for increasing the success rate of hair transplantation using the same.
  • exosomes refers to a endoplasmic reticulum of several tens to hundreds of nanometers (preferably approximately 30 to 200 nm) composed of a double phospholipid membrane having the same structure as a cell membrane (however, the object to be separated is The particle size of exosomes may vary depending on the cell type, separation method, and measurement method (Vasiliy S. Chernyshev et al., "Size and shape characterization of hydrated and desiccated exosomes", Anal Bioanal Chem, (2015) DOI 10.1007/s00216-015-8535-3).
  • Exosomes include proteins called exosomes cargo, nucleic acids (mRNA, miRNA, etc.). Exosomal cargoes contain a wide range of signaling factors, which are known to be cell type specific and differently regulated according to the secretory cell environment. Exosomes are intercellular signaling mediators secreted by cells, and various cellular signals transmitted through them regulate cell behavior including target cell activation, growth, migration, differentiation, dedifferentiation, apoptosis, and necrosis. Is known.
  • exosome has a nano-sized vesicle structure secreted from stem cells and released into the extracellular space, and has a composition similar to that of exosomes (eg, exosome- It is meant to include all the similar vegetables).
  • the type of the stem cells is not limited, but as an example of not limiting the present invention, preferably, mesenchymal stem cells may be, for example, fat, bone marrow, umbilical cord or cord blood-derived stem cells, more preferably Fat-derived stem cells.
  • the type of fat-derived stem cells is not limited as long as there is no risk of infection by the pathogen and does not cause an immune rejection reaction, but may preferably be human fat-derived stem cells.
  • the stem cell-derived exosome used in the present invention is effective in increasing the success rate of hair transplantation, and if it does not cause adverse effects on the human body, it is possible to use various stem cell-derived exosomes that can be used or used in the future.
  • the stem cell-derived exosomes isolated according to the separation method of the embodiments described below should be understood as an example of the exosomes that can be used in the present invention, and the present invention is clearly not limited thereto.
  • the term “hair” includes artificial or natural hair made of artificial materials such as polyethylene. Therefore, as used herein, the term “hair transplantation” refers to a site in which artificial hair or natural hair is individually planted at a desired site, for example, a site where a hair follicle collected from a patient having a hair loss is planted in an incision or non-incision method. For example, it covers transplantation to the hair loss site.
  • the term “success in hair transplantation” means that “attachment of the transplanted hair to the scalp (which may be referred to as engraftment or implantation)” is successful, especially when the scalp in which the hair is transplanted is redness. It indicates that the phenomenon that the transplanted hair falls out is reduced, indicating that the hair adheres well to the scalp.
  • the expression "increased the rate of hair transplantation success” does not show or decrease redness compared to the case of untreated hair transplantation in the case of the treated hair transplantation composition for increasing the hair transplantation success rate of the present invention. It means that the phenomenon of hair falling off is reduced.
  • composition for increasing the hair transplantation success rate of one embodiment of the present invention includes exosomes derived from stem cells as an active ingredient.
  • composition for increasing the rate of successful hair transplantation of one embodiment of the present invention is treated on a transplanted hair or scalp, or both, the redness of the scalp site where the hair is transplanted may not appear or be reduced.
  • Composition for increasing the rate of hair transplantation success rate of one embodiment of the present invention may include a lyophilized formulation comprising the stem cell-derived exosome, methionine, mannitol and trehalose as active ingredients.
  • the weight ratio of methionine, mannitol and trehalose is 1:1:1.
  • the lyophilized formulation may further contain ascorbic acid and retinol.
  • the weight ratio of methionine, mannitol, trehalose, ascorbic acid and retinol is 9:9:9:0.5:0.5.
  • the composition for increasing the rate of hair transplant success in one embodiment of the present invention may include the lyophilized preparation and a diluent.
  • the diluent may be water for injection, physiological saline, phosphate buffer solution, purified water, or deionized water.
  • the diluent may further include hyaluronic acid or a hyaluronic acid salt (eg, sodium hyaluronate).
  • the composition may be a suspension.
  • composition for increasing the success rate of hair transplantation in one embodiment of the present invention may be a pharmaceutical composition.
  • the pharmaceutical composition can be prepared as an injection.
  • composition for increasing the success rate of hair transplantation in one embodiment of the present invention may be administered to the scalp in which the hair is transplanted by microneedling, iontophoresis or injection.
  • composition for increasing the rate of successful transplantation of hair in one embodiment of the present invention can be effectively used to increase the success rate of hair transplantation when planting artificial hair or natural hair in a desired area or treating a hair follicle collected from a hair loss patient to a desired site. have.
  • the pharmaceutical composition of one embodiment of the present invention may include a pharmaceutically acceptable carrier, excipient or diluent.
  • the carrier, excipients, and diluents include lactose, dextrose, trehalose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia rubber, alginate, gelatin, calcium phosphate, calcium carbonate, calcium silicate, and cellulose , Methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil.
  • an effective amount of the pharmaceutical composition of an embodiment of the present invention means an amount required for administration in order to expect an effect of increasing the success rate of hair transplantation.
  • the blending ratio of the pharmaceutical composition of one embodiment of the present invention can be appropriately selected according to the type, amount, and form of the additional components as described above.
  • the pharmaceutical composition of the present invention may contain about 0.1 to 99% by weight, preferably about 10 to 90% by weight.
  • a suitable dosage of the pharmaceutical composition of one embodiment of the present invention depends on the severity of the patient's hair loss symptoms, the type of formulation, the formulation method, the patient's age, gender, weight, health status, diet, excretion rate, administration time and administration method. Can be adjusted accordingly. For example, when administering the pharmaceutical composition of one embodiment of the present invention to an adult, it can be divided into 1 to several times in a dose of 0.001 mg/kg to 100 mg/kg per day.
  • Another embodiment of the present invention is to provide a cosmetic method excluding treatment for increasing the hair transplant success rate by using the composition for increasing the hair transplant success rate.
  • a cosmetic method of increasing the success rate of hair transplantation in one embodiment of the present invention is a step of coating the composition for increasing the rate of hair transplantation success before hair transplantation, or for increasing the rate of success of the hair transplantation on the scalp with hair transplantation after hair transplantation And applying at least one of the steps of applying the composition.
  • redness of the scalp area where the hair is transplanted may not appear or may be reduced.
  • the composition for increasing the hair transplantation success rate may be administered to the scalp in which the hair is implanted by microneedling, iontophoresis or injection.
  • the composition for increasing the rate of hair transplantation success including the stem cell-derived exosome as an active ingredient of the present invention is treated in the hair transplantation process, the redness of the scalp site where the hair is transplanted does not appear or is reduced and the hair transplanted into the scalp This phenomenon of falling off can be reduced. Therefore, the composition for increasing the rate of successful transplantation of the hair of the present invention can increase the success rate of hair transplantation by reducing the phenomenon in which the hair transplanted into the scalp falls off while the scalp area where the hair is transplanted exhibits redness.
  • FIG. 1 shows the results of analyzing physical properties of exosomes obtained according to one embodiment of the present invention.
  • FIG. 1A shows the particle size distribution and number of particles obtained by nanoparticle tracking analysis (NTA).
  • FIG. 1B is a particle image photograph taken by TEM (transmitted electron microscopy).
  • FIG. 1C shows Western blot results for positive markers of exosomes obtained according to one embodiment of the present invention.
  • FIG. 1D shows Western blot results for negative markers of exosomes obtained according to one embodiment of the present invention.
  • FIG. 1E shows the results of flow cytometry for CD9, CD63 and CD81 in marker analysis for exosomes obtained according to one embodiment of the present invention.
  • Figure 2 shows the results confirming that there is no cytotoxicity after treating the exosomes obtained according to one embodiment of the present invention to human skin fibroblasts HS68 cells.
  • FIG. 3 is a photograph showing good properties of lyophilized exosomes according to the method of one embodiment of the present invention.
  • 4A to 4G are photographs of the properties of the lyophilized exosomes for each combination of the lyophilized protective agent components after different combinations of the lyophilized protective agent components and lyophilization.
  • FIG. 5A is a photograph of a hair transplantation site (scalp region) treated with the composition for increasing the rate of hair transplantation success of the present invention
  • FIG. 5B is a hair transplantation site of a negative control group not treated with the composition for increasing the rate of success of the invention. This is a picture of (scalp area).
  • Figure 6A is a graph for quantifying a photographic image of a hair transplantation site (scalp area) treated with the composition for increasing the rate of success of the hair transplantation of the present invention with respect to a red value
  • FIG. 6B is a composition for increasing the rate of success of the hair transplantation of the present invention This is a graph quantifying the red value of a photographic image of a hair transplantation site (scalp area) of an untreated negative control.
  • FIG. 7 is a comparison graph combining the graphs of FIGS. 6A and 6B, compared with the hair transplant scalp region (dashed line) treated with the composition for increasing the rate of successful hair transplantation of the present invention, compared to the untreated hair transplant scalp region (solid line). It represents this small thing (the small red one).
  • HS68 cells human dermal fibroblasts, are purchased from ATCC and contain 10% fetal bovine serum (fetal bovine serum: purchased from ThermoFisher Scientific) and 1% antibiotic-antimycotic (antibiotics-antimycotics: purchased from ThermoFisher Scientific).
  • the medium containing DMEM (purchased from ThermoFisher Scientific) medium was cultured at 5% CO 2 at 37°C.
  • Fat-derived stem cells were cultured at 5% CO 2 and 37° C. according to a cell culture method known in the art. Then, washed with phosphate-buffered saline (purchased from ThermoFisher Scientific), replaced with serum-free, phenol-free red medium, cultured for 1 to 10 days, and the supernatant (hereinafter, culture solution) was recovered. .
  • 2% by weight of trehalose was added to the culture medium in order to obtain exosomes having a uniform particle size distribution and high purity.
  • the culture was filtered through a 0.22 ⁇ m filter to remove impurities such as cell debris, waste products, and large particles.
  • the filtered culture medium was immediately separated exosomes through a separation process.
  • the filtered culture medium was stored in a refrigerator (up to 10°C of the image) and then used for exosome separation.
  • the filtered culture was cryopreserved in a cryogenic freezer below -60°C and thawed to perform exosome separation. Thereafter, the exosomes were separated from the culture medium using a Tangential Flow Filtration (TFF) device.
  • TTFF Tangential Flow Filtration
  • exosomes were separated from the culture medium filtered with a 0.22 ⁇ m filter, and a TFF (Tangential Flow Filtration) method was used for concentration, desalting, and diafiltration.
  • a filter for the TFF method a cartridge filter (cartridge filter, aka hollow fiber filter; purchased from GE Healthcare) or a cassette filter (purchased from Pall or Sartorius or Merck Millipore) was used.
  • TFF filters can be selected by various molecular weight cutoff (MWCO). The exosomes were selectively separated and concentrated by the selected MWCO, and particles, proteins, lipids, nucleic acids, and low molecular compounds smaller than MWCO were removed.
  • TFF filter of MWCO 100,000 Da (Dalton), 300,000 Da, or 500,000 Da was used.
  • the culture medium was concentrated to a volume of about 1/100 to 1/25 using the TFF method, and exosomes were separated by removing substances smaller than MWCO.
  • the separated and concentrated exosome solution was further subjected to desalination and diafiltration using the TFF method.
  • desalination and buffer exchange were continuously or continuously performed (discontinuous diafiltration), and at least 4 times, preferably 6 times to 10 times or more, more preferably, relative to the starting volume It was performed using a buffer solution having a volume of 12 times or more. 2% by weight of trehalose dissolved in PBS was added to the buffer solution to obtain exosomes having a uniform particle size distribution and high purity.
  • the separated exosomes were measured for particle size and concentration by nanoparticle tracking analysis (NTA; purchased from Malvern).
  • NTA nanoparticle tracking analysis
  • TEM transmitted electron microscopy
  • FIG. 1C shows the presence of CD63, CD9, CD81, Alix, and TSG101 markers as a result of performing Western blot on positive markers of exosomes isolated according to the separation method of an embodiment of the present invention.
  • Anti-CD63, anti-CD9, anti-CD81, anti-Alix and anti-TSG101 were used as antibodies to each marker, respectively.
  • 1D is a result of performing Western blot on a negative marker of exosomes separated according to the separation method of an embodiment of the present invention.
  • Anti-GM130 and anti-Calnexin were used as antibodies to each marker, respectively.
  • GM130 and Calnexin are negative markers that should not be present in exosomes when characterizing exosomes.
  • Figure 1D GM130 and calnexin were confirmed to be present in the lysate of adipose stem cells, but were not found in the exosomes separated according to the separation method of one embodiment of the present invention. Accordingly, when synthesizing the results of FIGS. 1C and 1D, it can be seen that the exosomes separated according to the separation method of one embodiment of the present invention are exosomes satisfying the positive marker and negative marker characterization.
  • Figure 1E confirmed the presence of CD9, CD63 and CD81 markers as a result of analysis using a flow cytometer for exosomes isolated according to the separation method of an embodiment of the present invention.
  • an exosome-human CD81 separation/detection kit purchased from ThermoFisher Scientific
  • PE-mouse anti-human CD9 PE-Mouse anti -human CD9
  • PE-Mouse anti-human CD63 purchasedd from BD
  • BD PE-mouse anti-human CD81 Markers
  • the exosome used in the present invention is not limited to the exosomes of the embodiments as described above, it is of course possible to use a variety of exosomes used in the art or may be used in the future.
  • the exosomes isolated according to the above embodiments should be understood as an example of exosomes that can be used in the present invention, and it is clearly revealed that the present invention is not limited thereto.
  • HS68 cells which are human skin fibroblasts
  • the cells were treated with exosomes by concentration and the cell proliferation rate was confirmed.
  • HS68 cells were suspended in DMEM containing 10% FBS, then dispensed to have 80 to 90% confluency and cultured for 24 hours in a 37°C, 5% CO 2 incubator. After 24 hours, the culture solution was removed and the exosomes prepared in Example 2 were treated by concentration to evaluate cell viability while culturing for 24 to 72 hours.
  • WST-1 reagent (WST-1 reagent) (purchased from Takara), MTT reagent (purchased from Sigma), CellTiter-Glo reagent (purchased from Promega), or Amarmar blue reagent ( Measurements were made using alamarBlue reagent (purchased from ThermoFisher Scientific) and a microplate reader (purchased from Molecular Devices).
  • the comparison group was based on the number of cells cultured in a normal cell culture medium in which exosomes were not treated, and it was confirmed that the cytotoxicity by the exosomes of the present invention was not observed within the tested concentration range (FIG. 2).
  • a lyophilized protective agent comprising methionine, mannitol and trehalose.
  • a lyophilized protective agent was added to a solution containing ascorbic acid and retinol, but an aqueous solution may be prepared by adding a lyophilized protective agent to water for injection, purified water, physiological saline, or deionized water.
  • concentrations of methionine, mannitol, and trehalose in the aqueous solution were respectively 9 mg/mL.
  • Example 2 After mixing the exosomes (5 ⁇ 10 8 particles) prepared in Example 2 with an aqueous solution containing a lyophilization protecting agent, they were frozen using the freeze-drying equipment (manufacturer: VIRTIS, ITME No.: 34424) under the conditions of Table 1 below. Drying was performed. Lyophilization was performed in the order of conditions 1, 2, 3, 4, 5, 6, 7 and 8 in Table 1 below.
  • Lyophilization conditions Total time (min) 4320 Condition Time (min) Temperature (°C) Pressure (mmHg) One 700 -50 760 2 60 -50 760 3 999 -50 0 4 999 -50 0 5 999 -50 0 6 370 -50 0 7 120 -20 0 8 73 10 0
  • Example 5-2 Comparison of properties of exosome products according to lyophilized protective agent components
  • Example 5-1 the properties of exosomes when exosomes were lyophilized using various freeze-dried protective agents including at least one of methionine, mannitol and trehalose (hereinafter referred to as a lyophilized protective agent component) were compared.
  • a lyophilized protective agent component a lyophilized protective agent component alone, a combination of two components, or a combination of three components.
  • the concentration of each lyophilized protective agent component in the aqueous solution was adjusted to 9 mg/mL.
  • the exosomes prepared in Example 2 (5 ⁇ 10 8 particles) were mixed in an aqueous solution containing each combination of lyophilization protection agent components, followed by lyophilization. .
  • Example 6 Treatment of composition for increasing hair transplant success rate
  • Biofibre HAIR-STRAIGHT an artificial hair product manufactured by Biofiber, Italy
  • a 5 mL aqueous solution was prepared by mixing the lyophilized product (lyophilized exosome prepared in Example 5-1) containing stem cell-derived exosomes at a concentration of 5 ⁇ 10 8 particles/vial with purified water.
  • the prepared exosome-containing aqueous solution was immersed in the root portion of the artificial hair to a depth of approximately 2 to 3 mm so that the aqueous solution of the exosome was coated on the root of the artificial hair.
  • the prepared artificial hairs are transplanted into the scalp of the alopecia part of a male subject in their 50s using an automatic biofiber hair implant system (manufactured by Biofiber, Italy). Did. Then, using a very thin needle equipped with an MTS (Microneedle Therapy System), for example, a general derma roller, an aqueous solution containing approximately 2 to 3 mL of exosomes remaining after artificial hair coating is applied to the scalp (hair transplantation site) of the subject. Penetrated. The remaining artificial hair that was not treated with the aqueous solution containing the exosomes was implanted into the scalp of the remaining hair loss part of the test subject as a negative control.
  • MTS Microcroneedle Therapy System
  • the lyophilized exosome prepared in Example 5-1 was mixed with an aqueous solution and used as a composition for increasing the rate of hair transplant success, but alternatively, the exosome prepared in Example 2 (for example, 1 ⁇ 10 Of course, it can be used as a composition for increasing the success rate of hair transplantation after suspending exosomes having a concentration of 8 particles/mL or more in water for injection.
  • natural hair can be used in place of artificial hair.
  • the scalp in which the hair is transplanted exhibits redness as a prognostic symptom. Therefore, if less redness appears on the scalp after hair transplantation, it can be evaluated that the hair transplantation was successful. For example, by quantifying the red value of the hair transplant scalp region treated with the composition for increasing the rate of successful hair transplantation of the present invention, the success rate of the hair transplantation of the present invention can be quantitatively compared with the negative control.
  • ImageJ is an open source Java image processing program based on NIH's image analysis tool.
  • ImageJ is a program that can be run on a computer equipped with a Jave 1.8 or higher virtual machine and can be downloaded from https://imagej.net/ImageJ, and is widely used for image analysis (references: Schneider, CA; Rasband, WS & Eliceiri, KW (2012), "NIH Image to ImageJ: 25 years of image analysis", Nature methods 9(7): 671-675; Schindelin, J.; Rueden, CT & Hiner, MC et al. (2015), "The ImageJ ecosystem: An open platform for biomedical image analysis", Molecular Reproduction and Development).
  • the red value (dashed line) of the hair transplant scalp region treated with the composition for increasing the rate of successful hair transplantation of the present invention is not treated with a negative control. It was confirmed that the hair transplantation was smaller than the red value (solid line) of the scalp area (see FIG. 7 ).
  • 7 is a comparison graph combining the graphs of FIGS. 6A and 6B. As shown in Figure 7, it can be seen that the peak value shifted to the left when the composition for increasing the rate of hair transplantation success of the present invention was treated. This indicates that the hair transplant scalp site treated with the composition for increasing the rate of successful hair transplantation of the present invention has a smaller red value (less redness) compared to the untreated hair transplant scalp site.
  • the composition for increasing the rate of hair transplantation success of the present invention is treated on the hair and/or the scalp during the hair transplantation process, the redness of the scalp area of the hair transplantation rarely occurs, and accordingly, the phenomenon that the hair falls out is reduced, thereby increasing the hair transplantation success rate. have.

Abstract

La présente invention concerne une composition permettant d'augmenter le taux de réussite d'une greffe de cheveux, contenant des exosomes dérivés de cellules souches utilisés comme principe actif, et une méthode permettant d'augmenter le taux de réussite d'une greffe de cheveux en faisant appel à celle-ci. Lorsqu'une composition, selon la présente invention, permettant d'augmenter le taux de réussite d'une greffe de cheveux, contenant des exosomes dérivés de cellules souches utilisés comme principe actif, est utilisée pendant un processus de greffe de cheveux, la rougeur de la zone du cuir chevelu au niveau de laquelle sont greffés les cheveux n'apparaît pas ou est réduite, et le phénomène dans lequel les cheveux greffés dans le cuir chevelu tombent peut être réduit. Par conséquent, la composition permettant d'augmenter le taux de réussite d'une greffe de cheveux, selon la présente invention, peut augmenter le taux de réussite d'une greffe de cheveux en réduisant le phénomène dans lequel les cheveux greffés dans le cuir chevelu tombent alors que la zone du cuir chevelu au niveau de laquelle sont greffés les cheveux présente une rougeur.
PCT/KR2020/000001 2019-01-11 2020-01-01 Composition permettant d'augmenter le taux de réussite d'une greffe de cheveux, contenant des exosomes dérivés de cellules souches utilisés comme principe actif WO2020145564A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
KR1020190003615A KR20200087898A (ko) 2019-01-11 2019-01-11 줄기세포 유래의 엑소좀을 유효성분으로 포함하는 모발이식 성공률 증가용 조성물
KR10-2019-0003615 2019-01-11

Publications (1)

Publication Number Publication Date
WO2020145564A1 true WO2020145564A1 (fr) 2020-07-16

Family

ID=71520785

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/KR2020/000001 WO2020145564A1 (fr) 2019-01-11 2020-01-01 Composition permettant d'augmenter le taux de réussite d'une greffe de cheveux, contenant des exosomes dérivés de cellules souches utilisés comme principe actif

Country Status (2)

Country Link
KR (1) KR20200087898A (fr)
WO (1) WO2020145564A1 (fr)

Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
KR20170044999A (ko) * 2015-10-16 2017-04-26 (주)프로스테믹스 줄기세포 유래 엑소좀을 포함하는 상처 치료, 피부 개선 및 탈모 방지 또는 치료용 조성물 및 그 제조방법
KR20170085010A (ko) * 2016-01-12 2017-07-21 주식회사 강스템바이오텍 고함량의 성장인자를 함유한 줄기세포 유래 엑소좀
KR20180003322A (ko) * 2016-06-30 2018-01-09 (주)아모레퍼시픽 성체줄기세포 유래의 엑소좀-모사 나노베지클을 포함하는 혈관 신생 촉진용 조성물
KR20180015994A (ko) * 2016-08-05 2018-02-14 한양대학교 에리카산학협력단 인간발모세포 및 지방줄기세포유래 엑소좀을 포함하는 발모유도 조성물
WO2018150440A1 (fr) * 2017-02-16 2018-08-23 OCT Therapies and Research Private Limited Milieux conditionnés de cellules souches pour des applications cliniques et cosmétiques

Family Cites Families (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
KR102305747B1 (ko) 2015-03-03 2021-09-27 김두희 모발 임플란트 및 이를 이식하는 방법
KR101800365B1 (ko) 2017-08-09 2017-11-22 김태희 자동 모낭분리장치

Patent Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
KR20170044999A (ko) * 2015-10-16 2017-04-26 (주)프로스테믹스 줄기세포 유래 엑소좀을 포함하는 상처 치료, 피부 개선 및 탈모 방지 또는 치료용 조성물 및 그 제조방법
KR20170085010A (ko) * 2016-01-12 2017-07-21 주식회사 강스템바이오텍 고함량의 성장인자를 함유한 줄기세포 유래 엑소좀
KR20180003322A (ko) * 2016-06-30 2018-01-09 (주)아모레퍼시픽 성체줄기세포 유래의 엑소좀-모사 나노베지클을 포함하는 혈관 신생 촉진용 조성물
KR20180015994A (ko) * 2016-08-05 2018-02-14 한양대학교 에리카산학협력단 인간발모세포 및 지방줄기세포유래 엑소좀을 포함하는 발모유도 조성물
WO2018150440A1 (fr) * 2017-02-16 2018-08-23 OCT Therapies and Research Private Limited Milieux conditionnés de cellules souches pour des applications cliniques et cosmétiques

Also Published As

Publication number Publication date
KR20200087898A (ko) 2020-07-22

Similar Documents

Publication Publication Date Title
WO2019088656A1 (fr) Composition de charge contenant des exosomes stabilisés
WO2021033899A1 (fr) Composition pour la cicatrisation ou l'accélération de la cicatrisation, contenant, en tant que principe actif, une combinaison d'acide hyaluronique et d'exosomes lyophilisés dérivés de cellules souches
KR102170387B1 (ko) 줄기세포 유래 엑소좀의 동결건조제제 및 이를 유효성분으로 포함하는 여드름의 예방, 억제, 완화, 개선 또는 치료용 조성물
WO2020027466A1 (fr) Procédé pour lyophilisation d'exosomes
KR102646145B1 (ko) 줄기세포 유래의 엑소좀을 유효성분으로 포함하는 모공 축소용 조성물
CN115645388A (zh) 氨基酸组合物及其用途
WO2019151611A1 (fr) Composition d'encre biologique pour feuille de régénération du derme, procédé de fabrication de feuille de régénération du derme personnalisée faisant appel à celle-ci, et feuille de régénération du derme personnalisée fabriquée en faisant appel au procédé de fabrication
KR20190064745A (ko) 간엽줄기세포 유래 인공나노좀을 포함하는 폐질환 예방 또는 치료용 약학적 조성물
US20210145797A1 (en) Compositions for treating skin
KR20240037208A (ko) 줄기세포 유래의 엑소좀을 유효성분으로 포함하는 안면 홍조 개선용 조성물
WO2020027467A1 (fr) Préparation lyophilisée d'exosomes issus de cellules souches et composition anti-inflammatoire la comprenant en tant que principe actif
WO2020145564A1 (fr) Composition permettant d'augmenter le taux de réussite d'une greffe de cheveux, contenant des exosomes dérivés de cellules souches utilisés comme principe actif
WO2018215414A1 (fr) Neurotrophines à utiliser dans le traitement de la perte auditive
KR20220011861A (ko) 줄기세포 유래의 엑소좀 또는 이의 동결건조제제를 유효성분으로 포함하는 항암제나 방사선 조사로 인한 피부특이반응의 예방, 억제, 완화, 개선 또는 치료용 조성물
WO2019112238A2 (fr) Composition cosmétique pour hydrater la peau comprenant un exosome dérivé de cellules souches en tant qu'ingrédient actif
KR102209294B1 (ko) 줄기세포 유래 엑소좀의 동결건조제제 및 이를 유효성분으로 포함하는 발모 촉진 또는 탈모 방지용 조성물
WO2022050373A1 (fr) Concentré purifié de surnageant de culture de cellule souche mésenchymateuse ou de cellule progénitrice dérivée de ladite cellule souche mésenchymateuse, et son procédé de production
WO2021096089A1 (fr) Nouvelle composition comprenant des exosomes dérivés de cellules souches et du polydésoxyribonucléotide en tant que principes actifs
WO2019235789A1 (fr) Composition comprenant un milieu de régulation de cellules souches mésenchymateuses dérivées des amygdales pour la prévention ou le traitement de la maladie du greffon contre l'hôte
WO2023167484A1 (fr) Composition pour prévenir ou traiter une maladie dégénérative de la rétine contenant des exosomes extraits de cellules souches mésenchymateuses
WO2021125558A1 (fr) Composition pour le noircissement des cheveux, la prévention du blanchiment des cheveux, ou la teinture des cheveux, comprenant un exosome dérivé de cellules souches en tant que principe actif
WO2023136538A1 (fr) Matrice dermique acellulaire destinée à être implantée dans le corps, utilisant un tensioactif naturel complexe végétal, et son procédé de préparation
WO2021010702A1 (fr) Composition pour la régénération d'une plaque de croissance
WO2019112229A2 (fr) Nouvelle utilisation d'un kit d'exosomes comprenant un exosome dérivé d'une cellule souche
WO2019190218A1 (fr) Composition, comprenant un milieu de régulation de cellules souches mésenchymateuses dérivées d'amygdale, pour la prévention ou le traitement d'une maladie hépatique

Legal Events

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

Ref document number: 20738810

Country of ref document: EP

Kind code of ref document: A1

NENP Non-entry into the national phase

Ref country code: DE

122 Ep: pct application non-entry in european phase

Ref document number: 20738810

Country of ref document: EP

Kind code of ref document: A1