Peptides for preventing or treating hair loss and their uses

Abstract

The peptides according to aspects of the present invention, as well as pharmaceutical compositions and external skin preparations containing the peptides, promote the growth of individual hair follicle cells and increase the amount of hair growth-related cytokines, thereby having the effects of hair care, hair growth, or hair growth promotion, and thus effectively treating, improving or preventing hair loss.

Description

Technical Field

[0001] The present disclosure relates to peptides for preventing or treating hair loss and uses thereof. Background Art

[0002] Testosterone is involved in skeletal muscle growth, genital development, spermatogenesis, and sexual desire, which are normal aspects of male physical characteristics. On the other hand, when dihydrotestosterone acts on related tissues, negative aspects (such as acne, increased sebum, hair loss, and prostate hyperplasia) develop. Among them, the study of hair loss (mainly in men) has been carried out for a long time, but the mechanisms of hair loss and hair growth have not been accurately elucidated. However, in modern society where appearance is highly regarded, the need for its prevention and treatment has become urgent.

[0003] Recently, the number of cases of hair loss in women has also been increasing. Although an increase in androgens is not observed in most patients with female pattern hair loss, the causes are known to be thyroid diseases and deficiencies of iron and trace elements in the body. Although female pattern hair loss is largely influenced maternally (on the mother's side), the family history is not as clear as that of male pattern hair loss. Recently, with the increase in the number of female pattern hair loss cases, the demand for various treatments and procedures in the beauty industry and related fields has been increasing.

[0004] Hair is a living organism that regularly repeats cyclic changes, including the anagen phase, catagen phase, and telogen phase. In addition, each hair repeats independent generation and destruction, thereby maintaining a constant number of hairs. The number of hairs is determined at birth. However, after puberty, the process of terminal hair formation due to the action of sex hormones is often misinterpreted as an increase in the number of hairs. The anagen phase is the period when hair grows due to the activity of hair follicles and active cell division, and it lasts for about 2 - 6 years. Approximately 85% of all hairs are in this phase. The hair bulb cells in the anagen phase are surrounded by a hair papilla composed of matrix cells to form a round shape, and cell division for generating new hair is active. The catagen phase is the period when the hair follicle stops growing and rapidly shrinks. At this time, the shape of the hair resembles a stick. The catagen phase takes about 2 - 3 weeks, and approximately 5% of all hairs are in the catagen phase. The telogen phase is the period when the activity of hair follicle cells completely disappears. During a period of 2 - 3 months, the hair follicle is pushed out and shed by the hair in the anagen phase growing from the root of the hair in the telogen phase, or naturally shed by mechanical actions such as combing or shampooing. Generally, hair loss refers to an abnormal increase in the number of shed hairs due to a decrease in the proportion of hairs in the anagen phase in the growth cycle while the number of hairs in the catagen phase or telogen phase increases.

[0005] There are various causes of hair loss where hair falls out from the scalp, and they are roughly classified into the effects of male hormones, mental stress, accumulation of lipid peroxides in the scalp, side effects of drugs, chronic diseases (such as leukemia or tuberculosis), side effects caused by radiotherapy, and nutritional deficiencies. In addition, hair loss has been known to be a problem for men in recent years. However, the demand for preventing and treating female hair loss, especially hair loss in young people, is increasing.

[0006] Drugs currently used as hair growth and hair care agents include vasodilators for promoting sufficient blood circulation to the scalp, estrogen for suppressing the effects of male hormones, and anti-androgens for inhibiting 5α-reductase that converts testosterone into 5-dihydrotestosterone (5-DHT). Vasodilators include caprochlorin, minoxidil, and various plant extracts, estrogen includes estradiol and progesterone, and anti-androgens include finasteride and pentadecanoic acid.

[0007] However, due to insufficient effects or side effects of hair loss treatment, there is a need to develop more effective and safe treatments or hair care agents for hair loss. Minoxidil administered topically or orally causes irritation to the skin, such as pain, erythema, inflammation, infection, or irritation on the scalp, and has the effect of lowering blood pressure. Therefore, hypertensive patients (including those taking antihypertensive drugs) should be cautious when administered. In addition, finasteride has drawbacks such as erectile dysfunction and reduced libido due to its effect of inhibiting male hormone activity.

[0008] Therefore, different from existing chemical-based compositions, the present inventors solved the above problems by inventing an erythropoietin-derived peptide to reduce the risk of side effects. Summary of the Invention

[0009] Technical Objectives

[0010] Exemplary embodiments of the present disclosure provide a pharmaceutical composition for preventing and treating hair loss or promoting hair growth, the pharmaceutical composition comprising a peptide as an active ingredient, the peptide comprising any one of the amino acid sequences selected from the group consisting of SEQ ID NO: 1 to SEQ ID NO: 3.

[0011] In addition, exemplary embodiments of the present disclosure provide a topical skin preparation composition for preventing or improving hair loss, the composition comprising a peptide, the peptide comprising any one of the amino acid sequences selected from the group consisting of SEQ ID NO: 1 to SEQ ID NO: 3.

[0012] Advantageous Effects

[0013] The composition according to one aspect contains novel peptides. Through the peptides according to one aspect, the pharmaceutical composition containing the peptides, and the topical skin preparation, the growth of individual hair follicle cells is promoted, and the expression of hair cytokines is increased, so as to have the effects of hair care, hair growth or promoting hair growth, thereby ensuring the effects of effectively treating, improving or preventing hair loss. Description of the Drawings

[0014] Figure 1 The figure shows the analysis by MTT assay 3 days after treating dermal papilla cells with three types of peptides according to one aspect.

[0015] Figure 2 The figure shows the length measurements of human hair follicle organ cultures 3 days, 6 days and 9 days after treating with three types of peptides according to one aspect.

[0016] Figure 3 The figure shows the determination of the gene expression levels of important cytokines involved in the proliferation of hair growth-related cells by treating dermal papilla cells with three types of peptides according to one aspect.

[0017] Figure 4 The figure shows the determination of the gene expression levels of important cytokines involved in the proliferation of hair growth-related cells by treating outer root sheath cells with three types of peptides according to one aspect.

[0018] Figure 5 The figure shows the images of checking the length growth of hair by injecting three types of peptides according to one aspect into the skin of animals.

[0019] Figure 6 The figure shows the images of checking the length growth of hair by injecting three types of peptides according to one aspect into the skin of animals three times.

[0020] Figure 7 The figure shows the images of determining the length growth and growth cycle of hair by injecting three types of peptides according to one aspect into the skin of animals under a microscope.

[0021] Figure 8 The figure shows the images of determining the length growth and growth cycle of hair by injecting three types of peptides according to one aspect into the skin of animals three times under a microscope. Detailed Description of the Invention

[0022] One aspect provides a peptide that comprises any one amino acid sequence selected from the group consisting of SEQ ID NO: 1 to SEQ ID NO: 3. The peptide may be derived from the sequence of the erythropoietin protein. The peptide can bind to the erythropoietin receptor. For example, it can bind to the target site (site 1) or target site (site 2) of the receptor.

[0023] Another aspect provides a pharmaceutical composition for preventing and treating hair loss or promoting hair growth, the composition comprising a peptide as an active ingredient, the peptide comprising any one amino acid sequence selected from the group consisting of SEQ ID NO: 1 to SEQ ID NO: 3.

[0024] As used herein, the term "hair loss" refers to the state of having no hair in an area where hair should be present, or having a low hair density, or thin and weakened hair. Generally, if the number of hairs lost per day is about 100 or more, a high likelihood of hair loss is considered to exist.

[0025] According to an exemplary embodiment, the hair loss can be female pattern baldness or male pattern baldness. In addition, the hair loss can be one or more selected from the group consisting of: alopecia areata, hereditary androgenetic alopecia, telogen effluvium, traumatic alopecia, trichotillomania, stress-induced alopecia, anagen effluvium, alopecia pityroides, syphilitic alopecia, seborrheic alopecia, symptomatic alopecia, cicatricial alopecia, or congenital alopecia.

[0026] The peptide may be derived from the sequence of the erythropoietin protein, and the peptide may be derived from the erythropoietin binding site.

[0027] The peptide has a binding affinity for the erythropoietin receptor of 0.1 μM to 1 nM. Specifically, the erythropoietin receptor has two target sites and forms a conjugate with erythropoietin. Among the two binding target sites, it is known that target site 1 forms a strong binding (KD = ~1 nM), and target site 2 forms a weak binding (KD = ~1 μM). The peptide according to one aspect can regulate the effect of cell proliferation by forming a weak bond with target site 2 in the erythropoietin receptor.

[0028] The peptide can form an α-helical structure and also has cytoprotective activity. In addition, the peptide can promote the proliferation or differentiation of hematopoietic stem cells, adipocytes, pancreatic cells, muscle cells, vascular cells, hair follicle cells, or skin cells.

[0029] In an exemplary embodiment, the peptide can promote the growth of dermal papilla cells or matrix cells. In addition, the peptide can promote the growth of outer root sheath cells or inner root sheath cells.

[0030] In an exemplary embodiment, the peptide can promote the expression of cytokines related to hair growth. Specifically, the peptide can promote the expression of insulin-like growth factor 1 (IGF1) in dermal papilla cells. Since the proliferation of dermal papilla cells involves the proliferation of hair-generating stem cells, it can have the effect of promoting hair growth. Therefore, if the expression of differentiation markers is stimulated, it can have the effect of promoting hair growth to make the hair grow well.

[0031] In addition, in outer root sheath cells, the peptide can promote the expression of keratinocyte differentiation markers K6, K16, K17, or K75. The differentiation of outer root sheath cells involves the effect of promoting hair growth (i.e., thickening the hair), thereby ensuring a good hair care effect. Therefore, when the expression of differentiation markers is stimulated, due to the thickened hair, it can have the effect of helping the hair grow well.

[0032] In an exemplary embodiment, the peptide can be included in an amount of 10 ppm to 10,000 ppm. Specifically, the content can be 20 ppm to 10,000 ppm, 10 ppm to 8,000 ppm, 30 ppm to 6,000 ppm, 50 ppm to 5,000 ppm, 70 ppm to 4,000 ppm, 80 ppm to 3,000 ppm, 100 ppm to 2,500 ppm, 110 ppm to 2,000 ppm, 150 ppm to 1,800 ppm, 200 ppm to 1,500 ppm, 300 ppm to 1,400 ppm, 400 ppm to 1,300 ppm, 500 ppm to 1,250 ppm, 600 ppm to 1,200 ppm, 700 ppm to 1,200 ppm, 800 ppm to 1,230 ppm, 900 ppm to 1,100 ppm, or 750 ppm to 1,150 ppm. Preferably, 1,000 ppm can be included. When the peptide is present in the above-listed content, the best effects of improving and treating hair loss or promoting hair growth can be obtained.

[0033] According to an exemplary embodiment, the peptide can have the effects of hair growth, promoting hair growth, or hair care.

[0034] As used herein, the term "hair growth" refers to the phenomenon of hair growth. That is to say, it broadly includes the growth of new hair. The terms "hair care" or "promoting hair growth" used herein refer to the effect of making the hair grow longer. That is to say, hair care or promoting hair growth refers to the effect of widely increasing the root, circumference, length, and strength of the hair by thickening and strengthening the hair.

[0035] According to an exemplary embodiment, the peptide may have all the functions of hair care, hair growth promotion, and hair growth. Therefore, in the composition according to one aspect, there may be an effect of hair regrowth, an effect of increasing the thickness of existing hair, and an effect of strengthening overall hair.

[0036] As used herein, the term "prevention" refers to any action of suppressing hair loss or delaying its occurrence by administering a pharmaceutical composition according to one aspect.

[0037] The term "amelioration" refers to any action of at least reducing a parameter (such as the degree of a symptom) related to an abnormal hair loss state by administering a pharmaceutical composition according to one aspect.

[0038] As used herein, the term "treatment" refers to any action of alleviating or beneficially altering the symptoms or disease of hair loss by administering a pharmaceutical composition according to one aspect. In this case, the composition may be used simultaneously with or separately from a drug for treatment before or after the onset of the disease to prevent or ameliorate hair loss.

[0039] Furthermore, as used herein, the term "active ingredient" or "pharmaceutically effective amount" refers to any amount of the composition used in the practice of the disclosure provided herein that is sufficient to alleviate, inhibit the progression of, or prevent a disease, disorder, or condition or one or more of their symptoms.

[0040] As used herein, the terms "administer", "introduce", and "implant" are used interchangeably and may refer to the method and route by which a composition according to an exemplary embodiment is placed into a subject's body such that the composition according to the exemplary embodiment is at least partially located at a desired site. The composition according to one aspect can be administered by any suitable route to deliver the composition to a desired site in a living individual.

[0041] As used herein, the term "pharmaceutically acceptable" relates to a composition that is suitable, within the scope of sound medical judgment, for contact with the tissues of a subject (such as a human) due to a reasonable benefit / risk ratio, without excessive toxicity, irritation, allergic response, or other problems or complications.

[0042] In addition to the active ingredient, the pharmaceutical composition may further include a pharmaceutically acceptable carrier. In this case, the pharmaceutically acceptable carriers are those commonly used in formulations, including lactose, dextrose, sucrose, sorbitol, mannitol, starch, gum arabic, calcium phosphate, alginate, gelatin, calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, water, syrup, methylcellulose, methylparaben, propylparaben, talc, magnesium stearate, and mineral oil, but are not limited thereto. In addition, in addition to the above components, lubricants, wetting agents, sweeteners, flavoring agents, emulsifying agents, suspending agents, and preservatives may also be included.

[0043] According to the desired method, the pharmaceutical composition can be administered orally (oral formulation) or parenterally (e.g., as an inhalant or injection for intravenous, subcutaneous, intraperitoneal, or topical application). For example, it can be administered subcutaneously (i.e., topical skin preparation) or orally (i.e., oral formulation), but is not limited thereto.

[0044] As the buffer to be added to the above various formulations, an isotonic, non-irritating solution with a pH of 4 - 9 and preferably a pH of 5 - 9 is used. The dosage varies according to the patient's condition and weight, the degree of the disease, the type of drug, the administration route, and the duration, but can also be appropriately selected by those skilled in the art.

[0045] According to an exemplary embodiment, the peptide or pharmaceutical composition can be injected by the following methods: a method of directly injecting under the skin of an individual; or a method of injecting the peptide or pharmaceutical composition into beads and then injecting the beads into the body. The subcutaneous injection method can be an injection type using a syringe or a patch injection type using a nano needle. The patch injection type can avoid the inconvenience that occurs in the direct injection type that requires frequent injections.

[0046] The pharmaceutical composition can be administered in a pharmaceutically effective amount. As used herein, the term "pharmaceutically effective amount" refers to an amount sufficient to treat a disease with a reasonable benefit / risk ratio applicable to medical treatment. The effective dose level can be determined by: the type of disease of the patient, the severity, the drug activity, the drug sensitivity, the administration time, the administration route and excretion rate, the treatment duration, factors including drugs used in combination, and other factors well known in the medical field. The other pharmaceutical compositions of the exemplary embodiments of the present disclosure can be administered as a separate therapeutic agent or in combination with other therapeutic agents, and can also be administered sequentially or simultaneously with conventional therapeutic agents. In addition, single or multiple administrations are allowed. Considering all the above factors, it is important to administer an amount that can achieve the best effect with the least amount without side effects, which can be easily determined by those skilled in the art.

[0047] The effective amount of the pharmaceutical composition may vary depending on the age, sex, condition, body weight of the patient, the absorption rate, inactivation rate and excretion rate of the active ingredient in the body, the type of disease, and the drugs used in combination, and may generally be in the range of 0.1 mg to 500 mg per 1 kg per day. Administration can be carried out daily or every other day in divided doses within the range of 1 to 5 times a day. However, since the dose may increase or decrease depending on the route of administration, severity of obesity, sex, body weight and age, the dose does not limit the scope in any way.

[0048] On the other hand, a method for preventing or treating hair loss is provided, the method comprising administering a pharmaceutically effective amount of the composition to an individual in need, contacting with cells in vitro, or administering to a laboratory animal in vivo. The specific details regarding the peptide and hair loss are the same as those described above.

[0049] The composition according to an exemplary embodiment may comprise any one or more peptides selected from the group consisting of SEQ ID NO: 1 to SEQ ID NO: 3, and the amount of the peptide is 0.001 wt% to 80 wt% based on the total weight of the composition.

[0050] In an exemplary embodiment, the peptide may be included in an amount of 10 ppm to 10000 ppm. Specifically, the content may be 20 ppm to 10000 ppm, 10 ppm to 8000 ppm, 30 ppm to 6000 ppm, 50 ppm to 5000 ppm, 70 ppm to 4000 ppm, 80 ppm to 3000 ppm, 100 ppm to 2500 ppm, 110 ppm to 2000 ppm, 150 ppm to 1800 ppm, 200 ppm to 1500 ppm, 300 ppm to 1400 ppm, 400 ppm to 1300 ppm, 500 ppm to 1250 ppm, 600 ppm to 1200 ppm, 700 ppm to 1200 ppm, 800 ppm to 1230 ppm, 900 ppm to 1100 ppm or 750 ppm to 1150 ppm. Preferably, the content may be about 1000 ppm. If the peptide is present in the amounts listed above, the most significant effects of improving and treating hair loss or promoting hair growth can be obtained. In addition, the peptide or composition can be administered 1 to 6 times a day. Specifically, within the range of clinically acceptable side effects, the number of administrations can be 1 or more than 2 times. Regarding the administration site, administration can be carried out at 1 site, or at 2 or more sites. Preferably, it can be administered 1 to 3 times.

[0051] In addition, the dosage of the composition can be from 0.01 mg to 10,000 mg, from 0.1 mg to 1000 mg, from 1 mg to 100 mg, from 0.01 mg to 1000 mg, from 0.01 mg to 100 mg, from 0.01 mg to 10 mg, or from 0.01 mg to 1 mg. However, the dosage can be prescribed in different ways according to factors such as the preparation method, type of administration, age, weight, gender, pathological condition, food, administration time, administration route, excretion rate, and response sensitivity of the patient, and those skilled in the art can appropriately adjust the dosage considering these influencing factors. For animals other than humans, the same dosage as that per kilogram of human can be applied, or an amount obtained, for example, by converting the above dosage according to the volume ratio (such as the average value) of the organs (such as the heart) of the target animal and humans.

[0052] Possible administration routes may include oral, sublingual, parenteral (such as subcutaneous, intramuscular, intra-arterial, intraperitoneal, intrathecal, or intravenous), rectal, topical (including transdermal), inhalation, and injection or insertion of an implantable device or substance.

[0053] According to an exemplary embodiment, the peptide or pharmaceutical composition can be injected by the following methods: directly injecting under the skin of an individual; or injecting the peptide or pharmaceutical composition into beads and then injecting the beads into the body. The subcutaneous injection method can be a patch injection type using nano-needles. The patch injection type can avoid the inconvenience that occurs in the direct injection type that requires frequent injections.

[0054] Subjects according to an exemplary embodiment include humans and other targets, such as mammals, and specifically include humans, monkeys, mice, rats, rabbits, sheep, cows, dogs, horses, and pigs.

[0055] The pharmaceutical composition according to an exemplary embodiment may include a pharmaceutically acceptable carrier and / or additive. For example, sterile water, physiological saline, conventional buffers (phosphoric acid, citric acid, and other organic acids), stabilizers, salts, antioxidants (ascorbic acid), surfactants, suspending agents, isotonic agents, or preservatives. For topical administration, it also includes combinations with organic materials (such as biopolymers) and inorganic materials (such as hydroxyapatite), especially collagen matrices, polylactic acid polymers or copolymers, polyethylene glycol polymers or copolymers, and their chemical derivatives. In the case where the pharmaceutical composition according to an exemplary embodiment is prepared in a form suitable for injection, the peptide can be dissolved in a pharmaceutically acceptable carrier or frozen in the dissolution solution.

[0056] The pharmaceutical composition according to an exemplary embodiment may suitably include (if required according to the type of administration or dosage form) a suspending agent, a solubilizing agent, a stabilizing agent, an isotonic agent, a preservative, an adsorption inhibitor, a surfactant, a diluent, an excipient, a pH adjuster, an analgesic, a buffer, a reducing agent, and an antioxidant. Reference [Remington’s Pharmaceutical Sciences, 19th Edition, 1995] describes in detail pharmaceutically acceptable carriers and dosage forms applicable to the present disclosure (including those exemplified above). The pharmaceutical composition according to an exemplary embodiment may be formulated into a unit dosage form using pharmaceutically acceptable carriers and / or excipients according to methods that can be easily implemented by those of ordinary skill in the art to which the present disclosure pertains, or may be prepared by placing it in a multi-dose container. In this case, the dosage form may be in the form of an emulsion, solution, or suspension in an oily or aqueous medium, or in the form of a powder, granule, tablet, or capsule.

[0057] On the other hand, there is provided a functional health food composition for preventing or improving hair loss, the composition containing a peptide as an active ingredient, the peptide consisting of any one amino acid sequence selected from the group consisting of SEQ ID NO: 1 to SEQ ID NO: 3.

[0058] Specific details regarding the peptide and hair loss are the same as those described above.

[0059] For the food composition, there are no particular restrictions on other added components except for the active ingredient contained as an essential component in a specified proportion. Various flavoring agents or natural carbohydrates may be included as additional components, such as in a conventional beverage. Examples of the above natural carbohydrates include monosaccharides (such as glucose and fructose); disaccharides (such as maltose and sucrose); and polysaccharides (such as conventional sugars, including dextrin and cyclodextrin) and sugar alcohols (such as xylitol, sorbitol, and erythritol). Natural flavoring agents (thaumatin, stevia extracts such as rebaudioside A and glycyrrhizin) and synthetic flavoring agents (saccharin, aspartame) may be advantageously used as flavoring agents other than those described above. The proportion of natural carbohydrates may be appropriately determined by the choice of those skilled in the art.

[0060] In addition to the above, the food composition may further include various nutrients, vitamins, minerals (electrolytes), flavoring agents (such as synthetic flavoring agents and natural flavoring agents), coloring agents, and thickening agents (cheese, chocolate), pectic acid and its salts, alginic acid and its salts, organic acids, protective colloid thickening agents, pH adjusters, stabilizing agents, preservatives, glycerol, alcohols, and carbonating agents used in carbonated beverages. These components may be used alone or in combination. The proportion of additives may also be appropriately selected by those skilled in the art.

[0061] On the other hand, there is provided a topical skin preparation composition for preventing or improving hair loss, the composition comprising a peptide containing any one amino acid sequence selected from the group consisting of SEQ ID NO: 1 to SEQ ID NO: 3.

[0062] Specific details regarding the peptide and hair loss are the same as those described above.

[0063] The composition has a hair growth promoting, hair nourishing or hair growth effect. Therefore, hair loss can be prevented or improved by: promoting hair growth to cause hair to grow, increasing the activity of hair-related keratin and cytokines in the body, and increasing the expression of hair-related cells, thereby enhancing the overall hair growth activity of an individual.

[0064] According to an exemplary embodiment, prevention or improvement of hair loss may include promoting hair growth or increasing hair thickness.

[0065] According to an exemplary embodiment, hair loss may be female pattern baldness or male pattern baldness. In addition, hair loss may be one or more selected from the group consisting of: alopecia areata, hereditary androgenetic alopecia, telogen effluvium, traumatic alopecia, trichotillomania, stress-induced alopecia, anagen effluvium, pityriasis alopecia, syphilitic alopecia, seborrheic alopecia, symptomatic alopecia, cicatricial alopecia or congenital alopecia.

[0066] A topical skin preparation composition refers to a composition of all dosage forms that can be applied to the skin surface except for quasi-drugs. According to an exemplary embodiment, the topical skin preparation composition may be a quasi-drug or a cosmetic composition.

[0067] According to an exemplary embodiment, the topical skin preparation may be one or more selected from the group consisting of: hair growth agents, scalp clinic agents, scalp scaling agents, scalp massage agents, scalp care agents, cleansers, shampoos, nourishing agents, hair conditioners, hair lotions, gels, packs, creams, essences, hair powders, sprays, hair oils, soaps, ointments, hair styling agents, hair dyes, perms, scalp nourishing agents, hair lotions for the scalp, scalp milks, scalp serums, hair mousses, hair gels, eyebrow growth promoters, eyelash growth promoters, eyelash nutrients and pet shampoos or pet conditioners.

[0068] As used herein, the term "quasi-drug" refers to a composition having a lower pharmaceutical effect compared to a therapeutic composition or a pharmaceutical composition. As used herein, the term "cosmetic composition" refers to a composition used on the human body for cleansing and caring for the human body to increase more charm, brighten the appearance, or maintain or improve the condition of the skin and hair.

[0069] For example, if a peptide of an aspect is included in a high amount, it can be classified as a therapeutic composition, and if there is a pharmaceutical effect but lower than that of a therapeutic composition, it can be classified as a quasi-drug for the skin. If the pharmaceutical effect is mild and intended for skin care, it can be classified as a cosmetic composition. Therefore, the quasi-drugs and cosmetic compositions for the skin described above can be collectively referred to as topical preparation compositions for the skin.

[0070] According to an exemplary embodiment, a hair growth promoter, a scalp care agent, a gel, a hair mask, a hair milk, a hair brightener, a hair powder, a spray, a hair oil, a soap, or an ointment containing a peptide of an aspect as an active ingredient can be a quasi-drug of a topical preparation for the skin.

[0071] According to an exemplary embodiment, a scalp treatment agent, a scalp scaler, a scalp massager, a scalp care agent, a cleanser, a shampoo, a conditioner, a hair conditioner, a hair wash, a gel, a hair mask, a hair milk, a hair brightener, a hair powder, a spray, a hair oil, a soap, an ointment, a hair styling agent, a hair dye, a hair perming agent, a scalp conditioner, a scalp wash, a scalp milk, a scalp essence, a hair mousse, a hair gel, an eyebrow growth promoter, an eyelash growth promoter, an eyelash nutrient, or a pet shampoo or a pet hair conditioner containing a peptide of an aspect can be a cosmetic composition.

[0072] When a peptide is added to a quasi-drug composition for the purpose of preventing or improving hair loss, the peptide of the above serial numbers can be added as it is or used in combination with other quasi-drug components, and can be appropriately used according to conventional methods. The mixing amount of the active ingredient can be appropriately determined according to the purpose of use (preventive, health care, or therapeutic treatment).

[0073] According to an exemplary embodiment, the peptide may be included in an amount of 10 ppm to 10,000 ppm. Specifically, the content may be 20 ppm to 10,000 ppm, 10 ppm to 8,000 ppm, 30 ppm to 6,000 ppm, 50 ppm to 5,000 ppm, 70 ppm to 4,000 ppm, 80 ppm to 3,000 ppm, 100 ppm to 2,500 ppm, 110 ppm to 2,000 ppm, 150 ppm to 1,800 ppm, 200 ppm to 1,500 ppm, 300 ppm to 1,400 ppm, 400 ppm to 1,300 ppm, 500 ppm to 1,250 ppm, 600 ppm to 1,200 ppm, 700 ppm to 1,200 ppm, 800 ppm to 1,230 ppm, 900 ppm to 1,100 ppm, or 750 ppm to 1,150 ppm. Preferably, the content may be about 1,000 ppm. If the peptide is present in the content listed above, the most excellent effects of improving and treating hair loss or promoting hair growth can be obtained.

[0074] Example

[0075] Hereinafter, preferred embodiments will be presented to assist in understanding the present disclosure. However, the following embodiments are provided only to more easily understand the present disclosure, and the content of the present disclosure is not limited by the following embodiments.

[0076] Example

[0077] Example 1. Synthesis of a Peptide for Hair Loss Treatment

[0078] According to a conventionally known solid-phase peptide synthesis technique (Peptron, Daejeon, Korea), an erythropoietin-derived peptide according to one aspect was synthesized in monomer form. Specifically, an erythropoietin-derived peptide having an important amino acid sequence (Arg103, Ser104, Leu105, Leu108, and Arg110) in a sequence that can bind to a target site (site 2) of a natural erythropoietin receptor was synthesized, and the concentration of the synthesized peptide was measured using liquid chromatography / mass selective detector (HP1100 series). Purity measurement (>95% purity) was performed by high performance liquid chromatography (SHIMADZU prominence HPLC) analysis.

[0079] Example 2. Method for Preparing a Hair Cell Model

[0080] 2.1 Isolation of Hair Follicles in the Anagen Phase

[0081] To test the effect of the peptide of Example 1 on hair, the present inventors prepared a hair cell model. Specifically, healthy, normal adult scalp tissues were obtained from hair transplant patients to isolate hair follicles in the anagen phase.

[0082] Specifically, a hair follicle cell isolation medium was prepared to isolate dermal papilla cells and outer root sheath cells from hair follicles in the anagen phase. The hair follicle cell isolation medium was prepared by adding 20% FBS (Hyclone) to a basal medium containing DMEM / low glucose (Hyclone).

[0083] As a result, hair follicles in the anagen phase could be isolated from adult scalp tissues.

[0084] 2.2 Isolation of hair follicle constituent cells from the isolated hair follicle organs

[0085] Using the hair follicle cells isolated in Example 2.1, outer root sheath cells and dermal papilla cells as hair follicle organs were isolated. Specifically, hair follicle cells in the anagen phase were cultured using a hair follicle organ medium.

[0086] After adding 1000 μL of the hair follicle organ medium, the hair follicle cells in the anagen phase were cultured in an incubator at 37 °C under conditions of 5% CO 2 / 95% air. The medium was changed every 4 days of culture, and the second passage was mainly used for experiments.

[0087] Specifically, William E medium (Gibco) was used as the basal medium of the hair follicle organ medium, and 100 u / mL penicillin / streptomycin, 2 mM L-glutamine, 10 μg / mL insulin, and 10 ng / mL hydrocortisone were added.

[0088] As a result, outer root sheath cells and dermal papilla cells as hair follicle organs could be isolated from hair follicle cells in the anagen phase.

[0089] 2.3 Culture of hair follicle constituent cells

[0090] To prepare a cell model for testing the effect of the peptide on hair loss treatment, the hair follicle organs isolated in Example 2.2 were cultured.

[0091] Specifically, two hair follicles per well were placed in the hair follicle organ medium and cultured at 37 °C under conditions of 5% CO 2 / 95% air. The medium was changed every 3 days.

[0092] For the follicular organ culture medium, different culture media are used for the follicular organs. The outer root sheath cells are cultured in EpiLife medium (Cascade Biologics), and the dermal papilla cells are cultured in a basal medium based on DMEM (Hyclone) supplemented with penicillin (100 U / mL), streptomycin (100 μg / mL), and 20% FBS. EpiLife medium is a sterile liquid medium containing 60 μM calcium chloride.

[0093] As a result, 15 isolated dermal papilla cells (DP cell isolation) and 30 outer root sheath cells (ORS cell isolation) were obtained.

[0094] Dermal papilla cells (DP cells) are a group of skin mesenchymal cells that not only regulate hair follicle development and growth but are also known as a reservoir of multipotent stem cells and are thus important for hair loss treatment.

[0095] In addition, outer root sheath cells (ORS cells) are the cells surrounding the hair fiber and the inner root sheath. The outer root sheath cells are composed of basal cells, spinous cells, and granular layers on the skin surface. It may also include cells that perform specific follicular functions and represents an alternative source of keratinocytes suitable for skin replacement.

[0096] Experimental Examples

[0097] Experimental Example 1. Determination of the effect of peptides in cell experiments on hair loss treatment

[0098] 1.1 Peptide treatment according to concentration in follicular organs

[0099] Using the peptide of Example 1, the effect of treating or preventing hair loss in the cell model of Example 2 was determined. The cell model was treated with peptides at different concentrations of 1 pM, 100 pM, and 1000 pM, respectively. Treatment with Scr peptide was used as a negative control group. The purity of all peptides used in the experiment was 95% or higher.

[0100] 1.2 Cell viability assay using MTT

[0101] The dermal papilla cells and outer root sheath cells in Example 2 were cultured in 96-well plates respectively, and then treated with peptides according to type and concentration in serum-free medium. Next, after 3 days, the cells were measured at 570 nm after treatment with MTT solution. At the same time, the untreated negative control group and the control group treated with 10 U erythropoietin were also compared by performing MTT analysis.

[0102] Compared with the 1000 pM peptide, 10 U has a binding degree similar to EPOR.

[0103] 1.3 Cell viability measurement using real-time PCR

[0104] To detect the gene expression levels of cytokines crucial for dermal papilla cell proliferation, the dermal papilla cells and outer root sheath cells in Example 2 were treated with each peptide, and then total RNA was isolated after 24 hours. Subsequently, cDNA was synthesized from the isolated total RNA, and the synthesized cDNA was subjected to PCR using primers for the gene to be analyzed.

[0105] 1.4 Determination of hair follicle growth during the incubation time

[0106] Human hair follicle cells in Example 2 were treated with the peptides of the present disclosure at concentrations, and then cultured for 3 days, 6 days, and 9 days. After culturing, the grown hair follicle cells were photographed under a microscope for comparison. The length of the grown hair was measured and expressed in mm. Meanwhile, in the untreated negative control group and the control group treated with 10 U of erythropoietin, the length of the grown hair was measured and compared. 10 U has a binding degree similar to that of EPOR compared to the 1000 pM peptide.

[0107] 1.5 Determination of the role of the peptide in the cell model

[0108] As a result, as Figure 1 shown, all peptides of SEQ ID NO: 1 to SEQ ID NO: 3 showed a high degree of cell differentiation compared to the negative control group. In addition, compared to the control group treated with erythropoietin, it was found that the three types of peptides of the present disclosure showed excellent effects even when treated with an amount that caused less binding to the receptor. In this case, since 10 U has a binding degree similar to that of EPOR compared to the 1000 pM peptide, and since there are no specific side effects due to the non-use of erythropoietin, the peptide in one aspect can be used in an amount smaller than that of erythropoietin to show an alopecia treatment effect.

[0109] In addition, as Figure 2 shown, when all peptides of SEQ ID NO: 1 to SEQ ID NO: 3 were compared with the negative control group, hair growth was promoted. In addition, compared to the control group treated with erythropoietin, there are no specific side effects due to the non-use of erythropoietin. Meanwhile, an alopecia treatment effect was obtained with an amount less than that of erythropoietin.

[0110] Next, as Figure 3 shown, as a result of determining the gene expression levels of cytokines important for dermal papilla cell proliferation, the peptides of SEQ ID NO: 1 to SEQ ID NO: 3 increased the gene expression levels of hair growth-related factors, keratinocytes, and cytokines.

[0111] In addition, as Figure 4 shown, as a result of determining the gene expression levels of cytokines important for the differentiation of outer root sheath cells, it was found that all cytokines involved in the differentiation of outer root sheath cells increased. The results showed the effect of thickening hair, thus having a beneficial effect on hair growth.

[0112] Particularly, compared with the control group treated with erythropoietin, since erythropoietin was not used, there were no specific side effects. And at the same time, although in a smaller amount than erythropoietin, the degree of increase in expression was better.

[0113] Since the proliferation of dermal papilla cells means the proliferation of stem cells that ultimately produce hair, this indicates a significant hair growth effect, and the experimental results indicate that IGF is a cytokine involved in proliferation. It was determined that this effect was more significant than that of EPO.

[0114] Experimental Example 2. Determination of the effect of the peptide on hair loss treatment in animal experiments

[0115] 2.1 Peptide treatment according to concentration in experimental animals

[0116] Using the peptide in Example 1, the effect of treating or preventing hair loss was determined in experimental animals. Specifically, in order to improve the accuracy in animal experiments, two methods were used for the animal experiments, which were carried out as follows: after injecting the peptide into beads, the beads were subcutaneously injected into experimental animals; or by repeatedly injecting the peptide. The experimental animals were divided into a bead - peptide injection model experimental group and a peptide injection model experimental group according to the injection method. The experimental animals in both experimental groups were treated with 1000 pM of the peptide.

[0117] The Bio - rad Bead product (#153 - 7302) was used for the experimental group of the bead - peptide injection model. Specifically, the impurities on the beads were removed and washed, and then each peptide was combined with the beads and incubated at 37°C. Next, the finally combined bead - peptide was injected into the subcutaneous fat layer of experimental animals using a needle. In addition, for the experimental group of the peptide injection model, the peptide in Example 1 was injected into the subcutaneous fat layer of experimental animals using an injection.

[0118] 2.2 H and E staining analysis method

[0119] After completing the animal experiment, skin tissue was cut around the injection area of the peptide. The tissue was fixed in a fixative and cryopreserved after processing the tissue in the form of frozen sections. The frozen tissue was cut into 10 - μm - thick slices using a microtome, and hematoxylin and eosin staining was performed.

[0120] 2.3 Determination of the role of the peptide in experimental animals

[0121] As a result, as Figure 5 and Figure 6 shown, in both methods, the peptides in one aspect effectively induced hair growth.

[0122] In particular, as Figure 7 and Figure 8 shown, it was observed that MLPH had a superior effect compared to any other peptide. When the peptide according to one aspect was injected into experimental animals, effective hair growth promotion was observed in both the bead injection method and the peptide injection method. In particular, in the case of animal experiments, it was found that the peptide in one aspect had a more significant hair growth effect than erythropoietin.

[0123] The above results indicate that the peptide in one aspect can effectively function as a composition for treating hair loss. <110> Daegu Gyeongbuk Institute of Science and Technology <120> Peptides for Preventing or Treating Hair Loss and Their Uses <130> PN131497 <160> 4 <170> KoPatentIn 3.0 <210> 1 <211> 19 <212> PRT <213> Artificial Sequence <220> <223> MLP Amino Acid Peptide <400> 1 Leu His Val Asp Lys Ala Val Ser Gly Leu Arg Ser Leu Thr Thr Leu 1 5 10 15 Leu Arg Ala <210> 2 <211> 19 <212> PRT <213> Artificial Sequence <220> <223> MLP-C Amino Acid Peptide <400> 2 Arg His Val Lys Lys Arg Val Lys Gly Leu Lys Ser Leu Thr Thr Leu 1 5 10 15 Leu Arg Ala <210> 3 <211> 19 <212> PRT <213> Artificial sequence <220> <223> MLP-H amino acid peptide <400> 3 Leu His Val Leu Lys Ala Val Ser Gly Leu Leu Thr Leu Thr Met Ile 1 5 10 15 Arg Arg Ala <210> 4 <211> 19 <212> PRT <213> Artificial sequence <220> <223> Scr amino acid peptide (negative control) <400> 4 Met Glu Leu Ser Gly Arg Thr Leu Gly Leu Ile Leu Lys Leu Arg Gln 1 5 10 15 Ser Ala Thr

Claims

1. A pharmaceutical composition for preventing and treating hair loss or promoting hair growth, the composition comprising a peptide as an active ingredient, the peptide consisting of any one of the following amino acid sequences selected from the group consisting of SEQ ID NO: 1 to SEQ ID NO:

3.

2. The composition according to claim 1, wherein, the peptide promotes the growth of dermal papilla (DP) cells or hair matrix cells.

3. The composition according to claim 1, wherein, the peptide promotes the growth of outer root sheath (ORS) cells.

4. The composition according to claim 1, wherein, the peptide promotes the expression of hair growth-related cytokines.

5. The composition according to claim 1, wherein, the peptide is contained in an amount of 10 ppm to 10,000 ppm.

6. The composition according to claim 1, wherein, the peptide has the effect of hair growth, promoting hair growth or hair care.

7. The composition according to claim 1, wherein, the hair loss is one or more selected from the group consisting of: telogen effluvium, anagen effluvium, symptomatic alopecia or congenital alopecia.

8. The composition according to claim 1, wherein, the hair loss is one or more selected from the group consisting of: alopecia areata, hereditary androgenetic alopecia, traumatic alopecia, stress alopecia, pityriasis alopecia or syphilitic alopecia.

9. The composition according to claim 1, wherein, the hair loss is one or more selected from the group consisting of: seborrheic alopecia or cicatricial alopecia.

10. A topical skin preparation composition for preventing or improving hair loss, the composition comprising a peptide consisting of any one of the following amino acid sequences selected from the group consisting of SEQ ID NO: 1 to SEQ ID NO:

3.

11. The composition according to claim 10, wherein, the prevention or improvement of hair loss is to promote hair growth or thicken hair.

12. The composition according to claim 10, wherein, the topical skin preparation composition is a quasi-drug or cosmetic composition.

13. The composition according to claim 10, wherein, the topical skin preparation composition is one or more selected from the group consisting of: cleanser, conditioner, gel, hair powder, spray, hair oil, ointment, hair dye and hair perming agent.

14. The composition according to claim 10, wherein, the topical skin preparation composition is one or more selected from the group consisting of: scalp treatment agent, scalp care agent, hair styling agent, eyebrow growth promoter, eyelash growth promoter and eyelash nutrient.

15. The composition according to claim 10, wherein, the topical skin preparation composition is one or more selected from the group consisting of: hair growth agent, scalp scale remover, hair conditioner, hair shampoo, hair mask, hair milk, hair brightener, soap, scalp lotion, hair mousse, hair gel and pet hair conditioner.

16. The composition according to claim 10, wherein, The topical skin preparation composition is one or more selected from the group consisting of: scalp massage agent, shampoo, scalp conditioner, scalp milk, and scalp essence.

17. The composition according to claim 16, wherein, the shampoo is a pet shampoo.

18. The composition according to claim 10, wherein, the peptide is contained in an amount of 10 ppm to 10,000 ppm.

19. The composition according to claim 10, wherein, the hair loss is one or more selected from the group consisting of: telogen effluvium, anagen effluvium, symptomatic hair loss, or congenital hair loss.

20. The composition according to claim 10, wherein, the hair loss is one or more selected from the group consisting of: alopecia areata, hereditary androgenetic alopecia, traumatic alopecia, stress alopecia, pityrosporum alopecia, or syphilitic alopecia.

21. The composition according to claim 10, wherein, the hair loss is one or more selected from the group consisting of: seborrheic alopecia or cicatricial alopecia.

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