Methods and compositions for treating skin and hair disorders

Abstract

To provide a composition for use as a pharmaceutical for treating skin disorders.SOLUTION: The composition comprises extracellular vesicles and includes Neuregulin-1 B1 (NRG1-B1) and urokinase-type plasminogen activator receptor (uPAR). The composition may further comprise at least one selected from the group consisting of epidermal growth factor (EGF), hepatocyte growth factor (HGF), vascular endothelial growth factor (VEGF), and interleukin 6 (IL-6), and the composition may further comprise bone morphogenetic protein 1 (BMP-1).SELECTED DRAWING: Figure 2A

Description

Background Art

[0001] People share 99.9% of exactly the same genes. That is, the differences between people are only contained in 0.1% of an individual's genes. That small difference is contained in single nucleotide polymorphisms (SNPs). Six to twenty of these SNPs are presumed to be the cause of all the differences observed regarding skin, eye, and hair color, as well as hair loss. Therefore, the epidermis and dermis of the skin are anatomically, physiologically, and biologically different depending on skin color and ethnicity, which is the reason why some people get gray hair at a young age and why some people never get gray hair. Similarly, male pattern baldness is hereditary and appears to be related to an abnormal reaction of hair follicles to by-products of testosterone. These small differences are contained in single nucleotide polymorphisms (SNPs). Recent research has discovered that from a very specific SNP profile, it is possible to accurately predict who will develop hair loss and who will not. This applies to both men and women. Hair loss begins at puberty and affects two-thirds of men by the age of 35 and approximately 85% of men by the age of 50. This is also a common problem in women. Similarly, certain SNPs have been identified in erectile dysfunction.

[0002] Currently, even if there are any, there are few skin products specific to a particular skin color or ethnicity, and there are also few hair products that explain the genetic factors causing gray hair or hair loss. Similarly, there are no products that address the genetic basis associated with erectile dysfunction. Therefore, conventional skin products, hair products, and the treatment of erectile dysfunction do not appropriately address the biological differences found in the dermis of each skin or hair color.

[0003] Accordingly, what is needed are new dermatological and cosmetic treatments specific to human skin color, hair color, alopecia, erectile dysfunction, and specific countries or ethnic groups, as well as methods of making such treatments, and treatment methods, including target SNPs for specific skin color, specific hair color, alopecia, and / or erectile dysfunction found in the DNA of representative donor-derived mesenchymal stem cells (MSCs), keratinocytes, or melanocytes.

Summary of the Invention

[0004] Disclosed are mesenchymal stem cell growth factor compositions for treating skin disorders, gray hair, alopecia, and erectile dysfunction, as well as methods of using them, and methods and compositions for their use in skin color / ethnicity-specific beauty and skin treatments.

[0005] In one aspect, a method of making a therapeutic composition for treating, suppressing, reducing, preventing, and / or restoring hair (e.g., white hair, alopecia (including, but not limited to, alopecia caused by male pattern baldness, androgenetic alopecia, alopecia areata, cicatricial alopecia, telogen effluvium, and / or female pattern baldness), etc.), and / or skin discoloration or skin disorders (e.g., vitiligo, melasma, nevus flammeus, and / or rosacea, etc.), and / or erectile dysfunction is disclosed. The method includes identifying the end-user's physical characteristics (e.g., skin color, skin type, ethnicity, race, hair type, and / or hair color, etc.), obtaining mesenchymal skin cells (MSCs) from a target donor having the same physical characteristics as the end-user (e.g., skin color, skin type, ethnicity, race, hair type, and / or hair color), but who has never experienced hair disorders or skin disorders and / or erectile dysfunction, and who may be specific to gender, race, and ethnicity, and having a single nucleotide polymorphism (SNP) profile; preparing a powder preparation of growth factors for MSCs, keratinocytes, and melanocytes from the obtained MSCs, wherein the MSC preparation comprises at least one member selected from the group consisting of cells cultured under normal hypoxic culture conditions or cell conditioned medium, and cells cultured under harsh wound healing conditions; adding the powder preparation to a base; and the therapeutic composition comprises a member selected from the group consisting of topical compositions and injectable compositions. In one aspect, the therapeutic composition corrects skin discoloration, stimulates hair follicle activation, promotes hair growth, stimulates the production of hair pigmentation, or promotes appropriate erectile function.In one aspect, the SNP comprises one or more SNPs present in one or more genes encoding interleukin 18 (IL-18), platelet-derived growth factor (PDGF) receptor (PDGFR) B (PDGFR-B), tissue metalloprotease inhibitor (TIMP) 1 (TIMP-1), TIMP-2, IL-23, activin A, intracellular adhesion molecule (ICAM-2), plasminogen activator inhibitor 1 (PAI-1), osteopontin (OPN), insulin, insulin growth factor binding protein 4 (IGF-BP4), tumor necrosis factor (TNF) receptor (TNF-R), neuregulin-1 β1 (NRG1-β1), urokinase-type plasminogen activator receptor (uPAR), ectodysplasin A2 receptor (XEDAR), follistatin, and / or activated leukocyte cell adhesion molecule (ALCAM). In one aspect, the SNP comprises the SNPs rs1946518 (-607C>A) and / or rs187238 (-137G>C) of IL-18.

[0006] Also disclosed herein are therapeutic compositions for treating, suppressing, reducing, preventing, or reversing hair disorders, skin disorders, and / or erectile dysfunction, the compositions comprising a composition base and a powder preparation of growth factors of mesenchymal stem cells (MSCs), keratinocytes, and / or melanocytes derived from a donor having the same physical characteristics as the end user (e.g., skin color, skin type, ethnicity, race, hair type, and / or hair color, etc.), but having a different single nucleotide polymorphism (SNP) profile indicative of the donor never having had a hair disorder, skin disorder, and / or erectile dysfunction and being specific to gender, race, and ethnicity, wherein the MSC preparation comprises at least one member selected from the group consisting of cells cultured under normal oxygen culture conditions or cell conditioned medium and cells cultured under harsh wound healing conditions.

[0007] In one aspect, the SNPs of the therapeutic composition are one or more SNPs present in one or more genes encoding interleukin 18 (IL-18), platelet-derived growth factor (PDGF) receptor (PDGFR) B (PDGFR-B), tissue metalloprotease inhibitor (TIMP) 1 (TIMP-1), TIMP-2, IL-23, activin A, intracellular adhesion molecule (ICAM-2), plasminogen activator inhibitor 1 (PAI-1), osteopontin (OPN), insulin, insulin growth factor binding protein 4 (IGF-BP4), tumor necrosis factor (TNF-R) receptor (TNF-R), neuregulin-1 B1 (NRG1-B1), urokinase-type plasminogen activator receptor (uPAR), ectodysplasin A2 receptor (XEDAR), follistatin, and / or activated leukocyte cell adhesion molecule (ALCAM). In one aspect, the SNPs include the SNPs rs1946518 (-607C>A) and / or rs187238 (-137G>C) of IL-18.

[0008] Also disclosed herein is a method of treating a skin disorder, erectile dysfunction, or hair disorder in a subject, which comprises administering to the subject any of the therapeutic compositions described in any of the preceding aspects.

[0009] In one aspect, any of the therapeutic compositions described in any of the preceding aspects is disclosed herein, and the hair disorder includes alopecia (e.g., alopecia includes male pattern baldness, androgenetic alopecia, alopecia areata, cicatricial alopecia, telogen effluvium, and / or female pattern baldness) or gray hair.

[0010] Methods for making skin treatment compositions for a specific skin color, race, type, or ethnicity are also disclosed herein, the methods comprising identifying the skin color, type, race, and ethnicity of an end user and obtaining mesenchymal skin cells (MSCs) from a target donor having the same skin color, type, race, and ethnicity as the end user based on an appropriate single nucleotide polymorphism panel of donors, and preparing a powder preparation of growth factors for MSCs, keratinocytes, and melanocytes from the obtained MSCs, wherein the MSC preparation comprises at least one member selected from the group consisting of cells cultured under normal oxygen culture conditions or cell conditioned media and cells cultured under harsh wound healing conditions, preparing, and adding the powder preparation to a cosmetic base.

[0011] In one aspect, a skin treatment composition for a specific skin color, race, type, or ethnicity is disclosed herein, the composition comprising a composition base and a powder preparation of growth factors for mesenchymal stem cells (MSCs), keratinocytes, and / or melanocytes derived from a donor having the same skin color, type, race, and ethnicity as the end user, wherein the MSC preparation comprises at least one member selected from the group consisting of cells cultured under normal oxygen culture conditions or cell conditioned media and cells cultured under harsh wound healing conditions.

Brief Description of the Drawings

[0012] The accompanying drawings, which are incorporated herein and constitute a part of this specification, illustrate several embodiments and, together with the description, serve to exemplify the disclosed compositions and methods.

[0013]

Figure 1

Figure 2A

Figure 2B

Figure 2C

Figure 2D

Figure 2E

DETAILED DESCRIPTION OF THE INVENTION

[0014] Before the compounds, compositions, articles, devices, and / or methods disclosed and described herein, since they can naturally vary, unless otherwise specified, they are not limited to a particular synthetic method or a particular recombinant biotechnological method, and unless otherwise specified, they are not limited to particular reagents. It should also be understood that the technical terms used herein are for the sole purpose of describing particular embodiments and are not intended to be limiting.

[0015] A. Definitions As used in this specification and the appended claims, the singular forms "a", "an", and "the" include plural referents unless the context clearly dictates otherwise. Thus, for example, reference to "a pharmaceutical carrier" includes mixtures of two or more such carriers.

[0016] A range can be expressed in this specification as from "about" one particular value and / or to "about" another particular value. When such a range is expressed, another embodiment includes from one particular value and / or to another particular value. Similarly, when a value is expressed as an approximation, it will be understood that the use of the antecedent "about" causes that particular value to form another embodiment. It will further be understood that each endpoint of a range is important both in relation to the other endpoint and independently of the other endpoint. It is also understood that several values disclosed in this specification exist, and each value, in addition to the value itself, is disclosed in this specification as "about" that particular value. For example, if the value "10" is disclosed, "about 10" is also disclosed. Also, as will be appropriately understood by those skilled in the art, when a value is disclosed as "less than" that value, it is understood that that value "greater than or equal to" and the possible ranges between that value are disclosed. For example, if the value "10" is disclosed, "10 or less" as well as "10 or more" are disclosed. Also, throughout this application, data is provided in several different formats, and it is understood that this data represents ranges of endpoints and start points, as well as any combination of data points. For example, if a particular data point "10" and a particular data point "15" are disclosed, greater than 10 and greater than 15, 10 or more and 15 or more, less than 10 and less than 15, 10 or less and 15 or less, as well as 10 and 15 are understood to be disclosed as being the same as 10 - 15. It is also understood that each unit between two particular units is also disclosed. For example, if 10 and 15 are disclosed, then 11, 12, 13, and 14 are also disclosed.

[0017] The term "subject" is defined herein to include, but not be limited to, animals such as mammals, including primates (e.g., humans), cows, horses, pigs, sheep, goats, dogs, cats, rabbits, rats, mice, etc. In some embodiments, the subject is a human.

[0018] "Administration" to a subject includes any route by which a drug is introduced or delivered to the subject. Administration can be effected by any suitable route, including oral, topical, intravenous, subcutaneous, transdermal, percutaneous, intramuscular, intra-articular, parenteral, intra-arterial, intradermal, intraventricular, intracranial, intraperitoneal, intralesional, intranasal, rectal, vaginal, by inhalation, via an implantable reservoir, parenteral (e.g., subcutaneous, intravenous, intramuscular, intra-articular, intrasynovial, intrasternal, intramedullary, intraperitoneal, intrahepatic, intralesional, and intracranial injection or infusion techniques), and the like. As used herein, "concurrent administration," "administration in combination," "simultaneous administration," or "administered simultaneously" means that the compounds are administered at the same time or essentially immediately after each other. In the latter case, the two compounds are administered at a sufficiently close time such that the observed results are indistinguishable from those achieved when the compounds are administered at the same time. "Systemic administration" refers to introducing or delivering a drug to a subject via a route by which the drug is introduced or delivered to a substantial portion (e.g., more than 50% of the body volume) of the subject's body, such as via an entry into the circulatory or lymphatic system. In contrast, "local administration" refers to introducing or delivering a drug to a subject via a route by which the drug is introduced or delivered directly adjacent to the area of administration or the point of administration and not systemically introduced in a therapeutically significant amount. For example, a drug administered locally is readily detectable in the immediate vicinity of the point of administration but is undetectable or present in negligible amounts at distal portions of the subject's body. Administration includes self-administration and administration by another person.

[0019] "Biocompatible" generally refers to a substance, as well as any of its metabolic or degradation products, that is generally non-toxic to a recipient and does not cause a significant adverse effect on the subject.

[0020] "Comprising" is intended to mean that a composition, method, etc. includes the recited elements but does not exclude other elements. When used to define a composition and method, "consisting essentially of" shall mean including the recited elements but excluding other elements that are important to the combination. Thus, a composition consisting essentially of the elements defined herein does not exclude trace contaminants from isolation and purification methods and pharmaceutically acceptable carriers (e.g., phosphate buffered saline, preservatives, etc.). "Consisting of" shall mean excluding other components for administering the composition of the present invention and trace elements of substantial method steps. Embodiments defined by each of these transitional terms are within the scope of the present invention.

[0021] "Control" is an alternative subject or sample used in an experiment for comparison purposes. The control can be "positive" or "negative".

[0022] "Controlled release" or "sustained release" refers to the release of a drug from a given dosage form in a controlled manner to achieve desired pharmacokinetic characteristics in vivo. One aspect of "controlled release" drug delivery is the ability to manipulate the formulation and / or dosage form to establish the desired kinetics of drug release.

[0023] The "effective amount" of a drug refers to an amount of the drug sufficient to produce the desired effect. The amount of an "effective" drug will vary for each subject depending on many factors such as the age and general condition of the subject, the particular drug(s), etc. Thus, it is not always possible to specify a quantified "effective amount". However, the appropriate "effective amount" for any given subject case can be determined by one of ordinary skill in the art using routine experimentation. Also, as used herein, unless otherwise specified, the "effective amount" of a drug can also refer to an amount that encompasses both a therapeutically effective amount and a prophylactically effective amount. The "effective amount" of a drug required to achieve a therapeutic effect can vary depending on factors such as the age, sex, and weight of the subject. The dosing regimen can be adjusted to provide an optimal therapeutic response. For example, several divided doses can be administered daily, or the dose can be proportionally decreased as indicated by the urgency of the therapeutic situation.

[0024] "Decrease" can refer to any change that results in a lesser amount of gene expression, protein production, symptoms, disease, composition, condition, or activity. A substance is also understood to decrease the genetic output of a gene if the genetic output of the gene product associated with the substance is less compared to the output of the gene product not associated with the substance. Also, for example, a decrease can be a change in the symptoms of a disorder such that the symptoms are less than those previously observed. A decrease can be a statistically significant decrease in the amount of a state, symptom, activity, composition for any individual, median, or average. Thus, a decrease can be a 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100% decrease as long as the decrease is statistically significant.

[0025] "Inhibit", "inhibiting", and "inhibition" mean reducing an activity, response, state, disease, or other biological parameter. This can include, but is not limited to, complete disappearance of the activity, response, state, or disease. This can also include, for example, a 10% reduction in an activity, response, state, or disease compared to a native level or a control level. Thus, the reduction can be a 10, 20, 30, 40, 50, 60, 70, 80, 90, 100%, or any amount of reduction in between compared to a native level or a control level.

[0026] "Prevent", "preventing", "prevention", and their grammatical variations, as used herein, refer to a method of partially or completely delaying or eliminating the onset or recurrence of one or more of a disease and / or its attendant symptoms, or preventing a subject from acquiring or reacquiring a disease, or reducing the risk that a subject will acquire or reacquire one or more of a disease or its attendant symptoms.

[0027] A "pharmaceutically acceptable" component is a component that is not biologically or otherwise undesirable, i.e., a component that can be incorporated into a pharmaceutical formulation of the invention as described herein and administered to a subject without causing a significant undesirable biological effect or interacting in a harmful manner with any of the other components of the formulation in which it is contained. When used with respect to administration to humans, this term generally means that the component meets the required criteria of toxicity tests and manufacturing tests or that it is included in the Inactive Ingredients Guide prepared by the U.S. Food and Drug Administration.

[0028] A "pharmaceutically acceptable carrier" (sometimes referred to as a "carrier") generally means a carrier or excipient useful in the preparation of a pharmaceutical composition or therapeutic composition that is generally safe and non-toxic and includes carriers acceptable for veterinary and / or human pharmaceutical or therapeutic use. The term "carrier" or "pharmaceutically acceptable carrier" may include, but is not limited to, phosphate buffered saline, water, emulsions (e.g., oil / water or water / oil emulsions) and / or various types of wetting agents. As used herein, the term "carrier" includes, but is not limited to, any excipient, diluent, filler, salt, buffer, stabilizer, solubilizer, lipid, stabilizer, or other material well known in the art for use in pharmaceutical formulations and further described herein.

[0029] "Pharmacological activity" (or simply "activity") can refer to derivatives or analogs, such as "pharmacologically active" derivatives or analogs, that have the same type of pharmacological activity as the parent compound and are of approximately equal degree (e.g., salts, esters, amides, conjugates, metabolites, isomers, fragments, etc.).

[0030] A "therapeutic agent" refers to any composition having a beneficial biological effect. Beneficial biological effects include both therapeutic effects, e.g., treatment of a disorder or other undesirable physiological condition, and prophylactic effects, e.g., prevention of a disorder or other undesirable physiological condition (e.g., non-immunogenic cancer). These terms also include pharmaceutically acceptable and pharmacologically active derivatives of the beneficial agents specifically mentioned herein, including, but not limited to, salts, esters, amides, proagents, active metabolites, isomers, fragments, analogs, etc. When the term "therapeutic agent" is used, or when a particular agent is specifically identified, it should be understood that the term includes the agent itself, as well as pharmaceutically acceptable and pharmacologically active salts, esters, amides, proagents, conjugates, active metabolites, isomers, fragments, analogs, etc.

[0031] "Polymer" refers to a relatively high molecular weight natural or synthetic organic compound whose structure can be represented by monomers, which are repeating subunits. Non-limiting examples of polymers include polyethylene, rubber, and cellulose. Synthetic polymers are typically formed by the addition or condensation polymerization of monomers. The term "copolymer" refers to a polymer formed from two or more different repeating units (monomer residues). By way of example, and without limitation, copolymers can be alternating copolymers, random copolymers, block copolymers, or graft copolymers. In certain embodiments, it is also contemplated that the various block segments of a block copolymer can themselves include copolymers. The term "polymer" encompasses all forms of polymers and includes, but is not limited to, natural polymers, synthetic polymers, homopolymers, heteropolymers or copolymers, addition polymers, and the like.

[0032] A "therapeutically effective amount" or "therapeutically effective dose" of a composition (e.g., a composition comprising an agent) refers to an amount effective to achieve a desired therapeutic result. In some embodiments, the desired therapeutic result is the control of type I diabetes. In some embodiments, the desired therapeutic result is the control of obesity. The therapeutically effective amount of a given therapeutic agent typically varies with respect to factors such as the type and severity of the disorder or disease being treated, as well as the age, sex, and weight of the subject. This term can also refer to the amount of a therapeutic agent effective to promote a desired therapeutic effect such as relief of pain (i.e., nociception), or the rate of delivery of the therapeutic agent (e.g., the amount over time). The exact desired therapeutic effect will vary depending on the condition being treated, the subject's tolerance, the agent and / or pharmaceutical formulation being administered (e.g., the potency of the therapeutic agent, the concentration of the agent in the formulation, etc.), as well as various other factors understood by those of skill in the art. In some cases, the desired biological or medical response is achieved after administering multiple doses of the composition to the subject over a period of days, weeks, or years.

[0033] As used herein and in the following claims, several terms are referenced and shall be defined to have the following meanings.

[0034] "Optional" or "optionally" means that the event or situation described below may or may not occur, and the description means that it includes cases where the event or situation occurs and cases where it does not occur.

[0035] Throughout this application, various publications are referenced. The disclosures of these publications are hereby incorporated by reference in their entirety into this application to more fully describe the state of the art relevant hereto. The disclosed reference documents are also incorporated herein individually and specifically with respect to the materials contained therein discussed in the passages that rely on the reference documents.

[0036] B. Composition The components used to prepare the disclosed compositions, as well as the compositions themselves used in the methods disclosed herein, are disclosed. These and other materials are disclosed herein, and when combinations, subsets, interactions, groups, etc. of these materials are disclosed, specific references to each and every individual and collective combination and permutation of these compounds may not be explicitly disclosed, but each is specifically contemplated and described herein. For example, if a particular therapeutic composition (such as a composition containing a growth factor, exosome, or protein derived from MSC, etc.) is disclosed and discussed, and some modifications that can be made to several molecules including the therapeutic composition (such as a composition containing a growth factor, exosome, or protein derived from MSC, etc.) are discussed, specifically, each and every combination and permutation of the therapeutic composition (such as a composition containing a growth factor, exosome, or protein derived from MSC, etc.), and unless otherwise specified, possible modifications are contemplated. Thus, if classes of molecules A, B, and C, and classes of molecules D, E, and F are disclosed, and A-D is disclosed as an example of a combined molecule, each is individually and collectively contemplated, i.e., the combinations A-E, A-F, B-D, B-E, B-F, C-D, C-E, and C-F are considered disclosed. Similarly, any subset or combination of these is also disclosed. Thus, for example, the subgroups of A-E, B-F, and C-E would be considered disclosed. This concept applies to all aspects of this application, including but not limited to the steps in the methods of making and using the disclosed compositions. Thus, if there are various additional steps that can be performed, it is understood that each of these additional steps can be performed by any particular embodiment or combination of embodiments of the disclosed method.

[0037] In one aspect, a therapeutic composition for treating, suppressing, reducing, preventing, and / or restoring alopecia, gray hair, erectile dysfunction, and / or skin disorders is disclosed herein that is specific to a subject's physical characteristics (e.g., skin color, skin type, ethnicity, race, hair type, and / or hair color). As disclosed herein, the therapeutic composition can include concentrated growth factors, exosomes, extracellular proteins, proteoglycans, cytokines, chemokines, proteins, and peptides derived from MSCs or similar fibroblasts, keratinocytes, or melanocytes, and the cells can be obtained from bone marrow, adipose (fat), stromal vascular fraction (SVF), bone, or other tissue sources before and after cell proliferation. In some aspects, the therapeutic composition can further include stem cell factor (SCF).

[0038] In embodiments, the therapeutic composition can include a base and a powder preparation of growth factors of MSCs, keratinocytes, and / or melanocytes, and the MSC preparation (MSC / K / M / Prep) can include at least one member selected from the group consisting of cells cultured under normal normoxic culture conditions or conditioned cell media, and cells cultured under harsh wound healing conditions. The normoxic culture conditions can be defined as about 21%, where about 21% can be 21% ± 5% oxygen (including serum supplementation and oxygen). On the other hand, the wound healing conditions can be defined as about 1 to about 5% oxygen in the presence of inflammatory cytokines, angiogenic factors, and / or low glucose.

[0039] MSC / K / M / PREP can include either conditioned media or lysates derived from cultured proliferating MSCs, keratinocytes, or melanocytes. In some embodiments, the composition can further include about 0.01 to about 10 wt% of cell-free media conditioned by the proliferation of MSC / K / M / PREP or lineage cells, and the cells are cultured under normal normoxic culture conditions or wound healing conditions.

[0040] The MSC / K / M / PREP conditioned medium, lysate, and induced products, or combinations thereof, can optionally be dissolved, mixed, or suspended, together with other active ingredients, in a mixture of emulsified lanolin alcohol, wax, and oil, or a mixture of petrolatum or mineral oil, a quaternary ammonium compound, an aliphatic alcohol, and a fatty acid ester softener, or in a lotion having a substantially similar composition.

[0041] The base of the compositions of the present disclosure can be any suitable or desired base such as a lotion, cream, pigment, serum, oil, gel, hydrogel, powder, foundation, facial mask, lip care product, hair care product, hair care product, skin cleanser, peeling agent, ointment, etc. Alternatively, the base can include materials suitable for direct injection into the dermis.

[0042] In embodiments, the base may include a lotion and may include a mixture of emulsified lanolin alcohol, wax, and oil, or a mixture of petrolatum or mineral oil, a quaternary ammonium compound, an aliphatic alcohol, and a fatty acid ester softener. Alternatively, the base may include a cream and may include a mixture of emulsified lanolin alcohol, water, petrolatum, glycerin, isostearyl palmitate, butylene glycol, glyceryl stearate, or mixtures thereof.

[0043] In some embodiments, the cosmetic base may be a carrier containing, for example, about 1 to about 20 wt% humectant, about 0.1 to about 10 wt% thickener and water. Alternatively, the carrier may include about 70 to about 99 wt% surfactant and about 0 to about 20 wt% fat. Alternatively, the carrier may include about 80 to 99.9% thickener, about 5 to about 15% surfactant, about 2 to about 15% humectant, about 0 to about 80% oil, a very small amount (less than 2%) of preservative, colorant and / or fragrance, and water as needed.

[0044] In an embodiment, the composition may further comprise a penetration enhancer for improving the epidermal penetration of the bioactive substance. Suitable penetration enhancers may include dimethyl sulfoxide (DMSO), DMSO-like compounds, ethanol-based compounds, pyroglutamic acid esters, and the like. The composition may also include a sunscreen, an anti-acne agent, an anti-cellulite agent, and other additional ingredients.

[0045] The composition may be filter sterilized or concentrated. Further, the composition may not contain non-human animal products or may be derived from an animal source.

[0046] The composition has been described above as including MSCs, but the use of other fibroblast-like cells is also contemplated. The product may include keratinocytes or melanocytes. MSCs may be derived from multiple sources such as bone marrow stroma, adipose, blood, dermis, periosteum, bone, and other tissues. In an embodiment, the MSCs may be derived from the patient (autologous) to whom the composition is applied or from another individual (allogeneic). The MSC / K / M / PREP may be expanded in culture for the purpose of recovering conditioned media, or increasing the amount of cells for lysates, or for fresh use prior to incorporation into the compositions of the present disclosure.

[0047] Producing the therapeutic composition of the present disclosure may include first identifying the hair characteristics of the target consumer, procuring MSCs from an individual having similar hair characteristics, and using the procured MSCs to create a topical and injectable treatment that is applied to the scalp or dermis in an area affected by miniaturization, the treatment stimulating hair follicles to generate hair growth.

[0048] Accordingly, the final treatment composition may be customized for the end - user based on hair characteristics. For using the hair and scalp treatment composition of the present disclosure, the user can simply topically apply the composition to the scalp. Alternatively, the composition can be directly injected into the dermis in areas affected by miniaturization. The administration can be from 0.1 mL to a maximum of 100 mL, suitable for a given indication. For example, the dose administered can be, in a single administration, a total of 0.1, 0.2, 0.25, 0.3, 0.4, 0.5, 0.6, 0.7, 0.75, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 3.0, 3.5, 4.0, 4.5, 5.0, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100 mL, or it may be divided equally over multiple injections. For example, a single 1 - mL amount can be dispersed into 0.1 - mL injections. When injecting multiple times, the amount of each injection can be 0.1, 0.2, 0.25, 0.3, 0.4, 0.5, 0.6, 0.7, 0.75, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 3.0, 3.5, 4.0, 4.5, 5.0, 6, 7, 8, 9, 10 mL. The injection can be made as a single injection site or as 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 250, 300, 350, 400, 450, 500, 600, 700, 800, 900, or 1000 separate injections.When multiple injections are used in the affected area, the distance between injections may be at intervals of 0.1, 0.2, 0.25, 0.3, 0.4, 0.5, 0.6, 0.7, 0.75, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 3.0, 3.5, 4.0, 4.5, 5.0, 6, 7, 8, 9, or 10 cm. It is understood and contemplated herein that, although desired, a single administration of the therapeutic agent may not be sufficient to achieve the desired therapeutic result. Thus, administration may be one time, or separately 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 250, 300, 350, 400, 450, 500, 600, 700, 800, 900, or 1000 times during the host's lifetime or treatment period. When administered multiple times, it may be administered once every 6, 12, 18, 24, 36, 48, 60, 72 hours, every 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 35, 36, 42, 49, 56, 58, 59, 60, 61, 62 days, every 9, 10, 11, 12, 13, 14, 15, 16 weeks, every 5, 6, 7, 8, 9, 10, 11, 12, 18, 24 months, or every 3, 4, 5, 6, 7, 8, 9, or 10 years.

[0049] It is understood and contemplated herein that the MSC or similar fibroblast-like cells, keratinocytes or melanocytes-derived concentrated growth factors, exosomes, extracellular proteins, proteoglycans, cytokines, chemokines, proteins, and peptides used in the therapeutic compositions of the present disclosure can be diluted to a dosage for administration. The diluent can be any suitable substance including, but not limited to, physiological saline, or any pharmaceutical-based carrier or excipient disclosed herein. The dilution of the MSC or similar fibroblast-like cells, keratinocytes or melanocytes-derived growth factors, exosomes, extracellular proteins, proteoglycans, cytokines, chemokines, proteins, and peptides can be 10:1, 9:1, 8:1, 7:1, 6:1, 5:1, 4:1, 3:1, 2:1, 1:1, 1:2, 1:3, 1:4, 1:5, 1:6, 1:7, 1:8, 1:9, 1:10, 1:15, 1:20, 1:25, 1:30, 1:35, 1:40, 1:45, 1:50, or 1:100.

[0050] 1. Skin color Currently, there are few skin products specific to a particular skin color or ethnicity. Thus, conventional skin products do not adequately address the biological differences found in the dermis of each skin color or particular race.

[0051] As described above, the base can include a lotion containing a mixture of emulsified lanolin alcohol, wax, and oil, or a mixture of petrolatum or mineral oil, quaternary ammonium compounds, aliphatic alcohols, and a fatty acid ester softener. Alternatively, the base may include a cream and include a mixture of emulsified lanolin alcohol, water, petrolatum, glycerin, isostearyl palmitate, butylene glycol, glyceryl stearate, or a mixture thereof.

[0052] In yet a further embodiment, the composition of the present disclosure may include eyelash treatment, and thus, the cosmetic base may include a mixture of mineral oil, linseed oil, castor oil, eucalyptus oil, lanolin, beeswax, or a mixture thereof.

[0053] In some embodiments, the base can be a cosmetic base and can include any known ingredients typically found in the fields of cosmetics and pharmaceutical cosmetics, such as oils, waxes or other standard fatty substances, or conventional gelling agents and / or thickeners, emulsifiers, humectants, softeners, sunscreens, hydrophilic or lipophilic active agents (such as ceramides), agents for dealing with free radicals, bactericides, occlusives, preservatives, basifying or acidifying agents, fragrances, surfactants, fillers, natural products or extracts of natural products (such as aloe or green tea extract), vitamins, or colorants. The amounts of the components of the cosmetic base can depend on the desired effects.

[0054] The cosmetic compositions of the present disclosure can provide skin tissue rejuvenation, enhancement, and improved or restored skin tissue. This composition can also be used for treating burns or other wounds as a wound healing enhancer. In multiple embodiments, the composition may be used for treating erectile dysfunction and vaginal atrophy. Some embodiments do not require the use of commonly used skin fillers and carriers (such as and hyaluronic acid, etc.). However, these components may be included. Further, embodiments of the present composition may not need to include growth factors (such as insulin, insulin-like growth factors, thyroid hormones, fibroblast growth factors, estrogen, retinoic acid, etc.). The present composition may also not need to include adipocytes.

[0055] The cosmetic compositions of the present disclosure can be applied topically to the skin and can treat any area of the skin, such as the face, neck, hands, or any other desired part of the body. In some embodiments, the compositions of the present disclosure can be pre-loaded into bandages or other dressings that can be applied to the skin.

[0056] As described above, to produce the skin treatment composition of the present disclosure, it may first include identifying the desired skin type, color, ethnicity, or race, and obtaining MSCs from a target donor having the same skin type, color, ethnicity, and race. Thus, the final skin treatment composition can be customized for the end user based on skin type, color, ethnicity, and race.

[0057] For example, as shown in FIG. 1, skin wrinkles can be treated by injecting the composition disclosed herein into the wrinkle site. Thus, MSCs containing the composition were diluted at a ratio of 1:10 in physiological saline (1 mL of the composition in 9 mL of physiological saline), and 0.5 mL was injected into each wrinkle. Within 10 days, the patient already showed improvement.

[0058] Accordingly, a method for preparing a therapeutic composition for treating, suppressing, reducing, preventing, and / or restoring skin discoloration or skin disorders (such as vitiligo, melasma, pityriasis rosea, and / or rosacea, etc.) is disclosed herein. The method includes identifying the physical characteristics of the end user (such as skin color, skin type, ethnicity, race, hair type, and / or hair color, etc.), and obtaining mesenchymal skin cells (MSCs) from a target donor having the same physical characteristics as the end user (such as skin color, skin type, ethnicity, race, hair type, and / or hair color), but the donor has never experienced skin disorders or skin discoloration and may be specific to gender, race, and ethnicity, and having a single nucleotide polymorphism (SNP) profile, preparing a powder preparation of growth factors for MSCs, keratinocytes, and melanocytes from the obtained MSCs, wherein the MSC preparation includes at least one member selected from the group consisting of cells cultured under normal hypoxic culture conditions or cell conditioned medium, and cells cultured under harsh wound healing conditions, adding the powder preparation to a base, and the therapeutic composition includes a member selected from the group consisting of topical compositions and injectable compositions.

[0059] As used herein, there is also disclosed a therapeutic composition for treating, suppressing, reducing, preventing, or reversing a skin disorder or discoloration (e.g., vitiligo, melasma, pityriasis rosea, and / or rosacea), comprising a composition base and a powder preparation of growth factors of mesenchymal stem cells (MSCs), keratinocytes, and / or melanocytes derived from a donor having a single nucleotide polymorphism (SNP) profile that indicates that the donor never has a skin disorder and can be gender, race, and ethnicity specific, but having the same physical characteristics (e.g., skin color, skin type, ethnicity, race, hair type, and / or hair color, etc.) as the end user. The MSC preparation comprises at least one member selected from the group consisting of cells cultured under normal oxygen culture conditions or cell conditioned medium, and cells cultured under harsh wound healing conditions.

[0060] Also disclosed herein is a method of treating a skin disorder in a subject, comprising administering to the subject any of the therapeutic compositions described herein.

[0061] Also disclosed herein is a method of making a skin therapeutic composition for a particular skin color, race, type, or ethnicity. The method comprises identifying the physical characteristics of the end user (e.g., skin color, skin type, ethnicity, race, hair type, and / or hair color, etc.), obtaining mesenchymal skin cells (MSCs) from a target donor having the same physical characteristics as the end user based on an appropriate single nucleotide polymorphism panel of the donor, and preparing a powder preparation of growth factors of MSCs, keratinocytes, and melanocytes from the obtained MSCs, wherein the MSC preparation comprises at least one member selected from the group consisting of cells cultured under normal oxygen culture conditions or cell conditioned medium, and cells cultured under harsh wound healing conditions, preparing, adding the powder preparation to a cosmetic base agent.

[0062] In one aspect, a skin treatment composition for a specific skin color, race, type, or ethnicity is disclosed herein, the composition comprising a composition base and a powder preparation of growth factors of mesenchymal stem cells (MSCs), keratinocytes, and / or melanocytes derived from a donor having the same skin color, type, race, and ethnicity as the end user, wherein the MSC preparation comprises at least one member selected from the group consisting of cells cultured under normal hypoxic culture conditions or cell conditioned medium and cells cultured under harsh wound healing conditions.

[0063] In one aspect, the SNPs of the therapeutic composition comprise one or more SNPs present in one or more genes encoding interleukin 18 (IL-18), platelet-derived growth factor (PDGF) receptor (PDGFR) B (PDGFR-B), tissue metalloprotease inhibitor 1 (TIMP) 1 (TIMP-1), TIMP-2, IL-23, activin A, intracellular adhesion molecule (ICAM-2), plasminogen activator inhibitor 1 (PAI-1), osteopontin (OPN), insulin, insulin growth factor binding protein 4 (IGF-BP4), tumor necrosis factor (TNF-R) receptor (TNF-R), neuregulin-1 B1 (NRG1-B1), urokinase-type plasminogen activator receptor (uPAR), ectodysplasin A2 receptor (XEDAR), follistatin, and / or activated leukocyte cell adhesion molecule (ALCAM). In one aspect, the SNPs comprise the SNPs rs1946518 (-607C>A) and / or rs187238 (-137G>C) of IL-18.

[0064] 2. Gray hair Stem cell factor (SCF) expression in the hair matrix identifies the immediate progenitor cells of the hair shaft structural cells, and the transcription factor KROX20 characterizes a sub-lineage of epithelial cells that differentiates into hair shaft progenitor cells. Stem cell factor (also known as SCF, KIT ligand, or hematopoietic stem cell factor) is a growth factor that regulates multiple physiological homeostasis events, including the maintenance of hematopoietic stem cells, mast cells, and melanocytes. The important role of SCF and its receptor KIT in hair pigmentation is supported by the absence of hair pigment in several SCF- and KIT-deficient animals. It is clear that melanocytes are the target cells involved in this type of reduced pigmentation, because they are melanin-producing cells and the main KIT-expressing cells in the hair follicle (HF). The source of SCF that supports melanocyte activity in the HF appears to be skin fibroblasts.

[0065] KROX20 (also known as EGR2) is a zinc finger transcription factor expressed in a subpopulation of hair follicle (HF) cells. However, the role of KROX20-expressing cells in HF development is not known. The transcription factor KROX20 has been reported to identify a sub-lineage of HF epithelial cells that are directed towards hair shaft differentiation during HF morphogenesis. The findings indicate that skin fibroblasts are the source of SCF necessary to maintain hair matrix melanocytes and generate hair pigmentation, as demonstrated by the complete loss of pigment when SCF is deleted in KROX20-lineage cells.

[0066] Studies have shown that depletion of SCF in hair shaft progenitor cells results in loss of hair pigmentation. In addition, melanocytes are specialized melanin-producing cells and the main KIT-expressing cells of the HF. Thus, melanocytes mediate SCF signaling to hair pigmentation, and SCF is required for melanocyte activity and hair pigmentation. This finding demonstrates that loss of SCF in hair shaft progenitor cells affects only melanocytes, highlighting the upper HF hair matrix as an important niche for regulating hair pigmentation through non-cell-autonomous SCF / KIT signaling. This also points to an important role of dermal papilla SCF in controlling the fate of melanocytes in terminal maturation without affecting previous differentiation and migration. SCF production in fibroblasts and keratinocytes appears to regulate the generation of hair pigmentation by melanocytes.

[0067] Fibroblasts secrete a number of melanogenic factors such as stem cell factor (SCF), hepatocyte growth factor (HGF), keratinocyte growth factor (KGF), corticotropin-releasing hormone (CRH), endothelin-I (ET-I), interferon-y (IFN-y), and interleukin-I (IL-I). Melanocytes are activated through multiple paracrine factors secreted by adjacent cells such as keratinocytes and fibroblasts. Paracrine factors bind to specific receptors expressed on melanocytes and then accelerate signaling to initiate melanin production.

[0068] The above-mentioned association with hair pigmentation is known, but currently there are no existing products for preventing or treating gray hair at the cellular level. Rather, gray hair has traditionally been addressed by dyeing gray hair to match the desired hair color.

[0069] Accordingly, a method for preparing a therapeutic composition for treating, suppressing, reducing, preventing, and / or restoring gray hair is disclosed herein, the method comprising identifying an end user's physical characteristics (e.g., skin color, skin type, ethnicity, race, hair type, and / or hair color, etc.), obtaining mesenchymal skin cells (MSCs) from a target donor having the same physical characteristics (e.g., skin color, skin type, ethnicity, race, hair type, and / or hair color) as the end user but who has never experienced gray hair and having a single nucleotide polymorphism (SNP) profile that may be specific to gender, race, and ethnicity, preparing a powder preparation of growth factors for MSCs, keratinocytes, and melanocytes from the obtained MSCs, wherein the MSC preparation comprises at least one member selected from the group consisting of cells cultured under normal hypoxic culture conditions or cell conditioned medium and cells cultured under harsh wound healing conditions, adding the powder preparation to a base, and the therapeutic composition comprises a member selected from the group consisting of topical compositions and injectable compositions. In one aspect, the therapeutic composition modifies and / or stimulates the production of hair pigmentation. In one aspect, the SNPs of the method for preparing the therapeutic composition comprise one or more SNPs present in one or more genes encoding interleukin 18 (IL-18), platelet-derived growth factor (PDGF) receptor (PDGFR) B (PDGFR-B), tissue metalloprotease inhibitor (TIMP) 1 (TIMP-1), TIMP-2, IL-23, activin A, intracellular adhesion molecule (ICAM-2), plasminogen activator inhibitor 1 (PAI-1), osteopontin (OPN), insulin, insulin growth factor binding protein 4 (IGF-BP4), tumor necrosis factor (TNF-R) receptor (TNF-R), neuregulin-1 B1 (NRG1-B1), urokinase-type plasminogen activator receptor (uPAR), ectodysplasin A2 receptor (XEDAR), follistatin, and / or activated leukocyte cell adhesion molecule (ALCAM). In one aspect, the SNPs comprise the SNPs rs1946518 (-607C>A) and / or rs187238 (-137G>C) of IL-18.

[0070] Also disclosed herein are therapeutic compositions for treating, suppressing, reducing, preventing, or reversing gray hair, the compositions comprising a composition base and a powder preparation of growth factors of mesenchymal stem cells (MSCs), keratinocytes, and / or melanocytes derived from a donor having the same physical characteristics as the end user (such as skin color, skin type, ethnicity, race, hair type, and / or hair color, etc.), but who never has gray hair and has a different single nucleotide polymorphism (SNP) profile that may be specific to gender, race, and ethnicity, wherein the MSC preparation comprises at least one member selected from the group consisting of cells cultured under normal oxygen culture conditions or cell conditioned medium, and cells cultured under harsh wound healing conditions.

[0071] In one aspect, the SNPs of the therapeutic composition comprise one or more SNPs present in one or more genes encoding interleukin 18 (IL-18), platelet-derived growth factor (PDGF) receptor (PDGFR) B (PDGFR-B), tissue metalloprotease inhibitor 1 (TIMP-1), TIMP-2, IL-23, activin A, intracellular adhesion molecule (ICAM-2), plasminogen activator inhibitor 1 (PAI-1), osteopontin (OPN), insulin, insulin growth factor binding protein 4 (IGF-BP4), tumor necrosis factor (TNF-R) receptor (TNF-R), neuregulin-1 B1 (NRG1-B1), urokinase-type plasminogen activator receptor (uPAR), ectodysplasin A2 receptor (XEDAR), follistatin, and / or activated leukocyte cell adhesion molecule (ALCAM). In one aspect, the SNPs comprise the SNPs rs1946518 (-607C>A) and / or rs187238 (-137G>C) of IL-18.

[0072] Also disclosed herein is a method of treating gray hair in a subject, comprising administering to the subject any of the therapeutic compositions disclosed herein.

[0073] 3. Alopecia Hair is made of a protein called keratin. Hair sits in hair follicles, and at the base of the hair follicle, there are stem cells called dermal papilla stem cells that are stimulated to generate new hair. The average head has over 100,000 hair follicles, and each hair follicle repeats a cycle of generating hair and then resting. The cycle of hair generation and rest is normal, and one will always have hair all over the head.

[0074] Hair loss occurs because more and more hair follicles enter the resting phase or shrink until no new hair grows. Due to genetics, men destined to have hair loss have hair follicles that are overly sensitive to the effects of dihydrotestosterone (DHT), a byproduct of testosterone. DHT binds to the hair follicles and causes them to shrink. More and more hair follicles shrink until no hair grows, resulting in thinning hair and ultimately hair loss. This process is called miniaturization.

[0075] Currently, the only drugs approved by the FDA to treat hair loss are minoxidil and finasteride. Minoxidil is available as an over-the-counter medication, but most experts agree that minoxidil is a relatively ineffective drug in dealing with hair loss and has no effect on the process of miniaturization. Therefore, its benefits are temporary. Finasteride works by inhibiting the conversion of testosterone to DHT, but its side effects include erectile dysfunction, decreased libido, and ejaculatory disorders. Low-level laser therapy has been shown to stimulate hair growth in both men and women, but it has no effect on miniaturization.

[0076] Hair implantation is another method for treating alopecia, involving harvesting hair follicles from the back of the head that are resistant to DHT and transplanting them to the alopecia sites. However, this only redistributes them more evenly across the entire scalp since the amount of the patient's hair remains the same and does not stimulate new hair growth. What is needed are compositions and methods for preventing and treating alopecia, and the present methods and compositions activate hair follicles to produce hair and prevent miniaturization by promoting the stimulation of hair follicle epithelial stem cells and mesenchymal stem cells in the hair follicles.

[0077] Some embodiments of the present disclosure include a method of manufacturing a composition for preventing and treating alopecia, comprising preparing a concentrate of mesenchymal stem cells (MSCs), exosomes, and secretome from a target donor having hair characteristics of certain SNPs indicating that the donor will never experience hair loss. The donor can be sex-specific. The method can include identifying the target donor by the hair characteristics of the SNPs, culturing MSCs from the target donor to produce a culture medium under normal oxygen culture conditions or harsh wound healing hypoxic conditions, producing a powder from the culture medium, and combining the powder with a cream base or lotion base, and the final product can be applied to the dermis of areas affected by miniaturization. The application can be topical or by injection, and in particular, in areas affected by miniaturization or by direct injection into the dermis, it can stimulate the activation of hair follicles and promote hair growth.

[0078] When injected into the affected scalp of a subject with male pattern alopecia or androgenetic alopecia, successful regrowth was observed as early as 2 weeks after treatment (see Figures 2A - 2E).

[0079] The proteomic evaluation of the exosome suspension generated using the methods and parameters described in this specification was performed to characterize the molecular composition contributing to the clinically significant efficacy of the described invention. The proteomic evaluation utilized an antibody array manufactured by commercially available RayBio Tech (Norcross, GA, USA). The concentrations of 230 different proteins known to either be secreted, transported, or present on the outer surface of the cell membrane (within the extracellular microenvironment) were measured. Proteins found to be present at physiologically appropriate concentrations in duplicate test samples and supported as being related to hair repair and coloring are listed in Table 1. Physiologically appropriate concentrations were considered to be those with an average concentration of 1 pg / mL or higher. A literature search of proteins with an average concentration exceeding 50 μg / mL was conducted to identify the most likely candidate molecules supporting the clinical effect. The list in Table 1 should not be considered an exhaustive or comprehensive list of the individual proteins found in the present invention related to an effective effect.

[0080]

Table 1

[0081] The proteins are arranged from the highest average concentration to the lowest. Proteins expressed at a concentration exceeding any value of 50 ng / mL were investigated in the medical literature using the PubMed search engine to identify studies providing evidence of the potential effects of the present invention. In the scientific literature, the proteins of the present invention supported as playing a role in hair follicle regeneration are listed and described below.

[0082] IL18 was the most concentrated protein present in the exosome suspension sample. Its main function is involved in the regulation of innate immune responses in the skin, specifically stimulating interferon-γ production and activating dermal natural killer cells and TH1 T cells. Excessive IL18 is associated with the onset of autoimmune diseases. Literature research provides evidence regarding its role in hair growth. Two single nucleotide polymorphisms (SNPs) found within the IL18 gene sequence are associated with the onset of alopecia areata (AA), an organ-specific autoimmune disease. Two specific SNPs, rs1946518 (-607C>A) and rs187238 (-137G>C) polymorphisms, are associated with the alopecia areata disease. Celik et al. concluded that the SNPs of rs187238 and rs1946518 of IL-18 could be the cause of AA susceptibility. 1 IL18 may play a role in the irregular interactions observed between mast cells and CD8+ cells around the hair follicle. These interactions may disrupt normal interactions within the hair follicle. By histological analysis, IL18 and its receptor are found within keratinocytes of the skin and within the outer root sheath cells of the hair follicle. Two possible effects of the action of IL18 that initiate hair growth observed when the present invention is applied to the dermis are that the sequence of this donor-derived IL18 does not have a disease-related sequence and changes the level of IL18 signal transduction to a non-pathological level. Alternatively, there is evidence of an IL18 isoform that includes the main form in serum that can function as an autocrine inhibitor. The form of IL18 in the present invention may be an inhibitory isoform that reduces IL18 signal transduction and restores an appropriate innate immune response to an appropriate level.

[0083] The induction of the growth phase using conditioned medium from a population of mesenchymal stem cells / mesenchymal stromal cells derived from hypoxic fat has been demonstrated. PDGF receptor B is the major protein detected, and PDGF can stimulate the proliferation of dermal papillae via the PDGF receptor. Exosome introduction of PDGF receptor B into the membrane of resting dermal papilla cells may enable PDGF-mediated signaling that initiates hair growth in the growth phase. Genetic variants of PDGFR-B are associated with hair loss in Penthinnen syndrome.

[0084] Insulin growth factors are agonists of hair growth. IGF-binding proteins regulate IGF activity by binding to IGF. Binding to various IGF-BPs can modify IGF activity and provide further specificity of IGF activity. IGF-BP4, together with IGF-BP3 and IGF-BP5, is expressed in non-human hair follicle dermal papillae and plays a role in regulating IGF activity within the hair follicle.

[0085] Normal hair growth is cyclic. Each hair follicle undergoes extracellular remodeling throughout each cycle. TIMP-1 and TIMP-2 play important roles in regulating the proteolytic activity of collagenase and other proteases, and these are associated with the remodeling of the extracellular matrix within and around the hair follicle. The additional TIMP provided by the present invention can restore the balance to protease levels altered by the inflammatory disease state associated with alopecia.

[0086] Interleukin 23 has been found to be at a higher level in hair follicles during alopecia areata, but its exact function within the hair follicle remains unclear. Similar to other cytokines, IL23 can have multiple functions depending on the isoform, and the receptors are expressed in different cell types. The additional IL23 provided by the present invention can provide an autocrine inhibitory signal that helps reduce the inflammatory state that causes hair growth inhibition.

[0087] Activin is a member of the TGF-β signaling pathway, is important for the initial formation of the hair follicle during differentiation, and plays an important role in the epidermal / mesenchymal interactions required during hair organogenesis. Subsequently, in combination with follistatin, activin is an important regulator of the hair cycle.

[0088] ICAM functions to connect cells and form a barrier. The hair follicle is an immune-privileged organ. ICAM establishes a physical barrier that creates its immune-privileged environment. In alopecia areata, the immune-privileged environment is disrupted, triggering an autoimmune response against antigens within melanocytes that provide color to the hair. By providing ICAM-2, the present invention may enable the re-establishment of a microenvironment with immune privilege.

[0089] Osteopontin is expressed in the outer root sheath cells of the hair follicle. Osteopontin is proteolytically cleaved in vivo to generate a peptidic signaling molecule that regulates FGF-7 production by outer root sheath keratinocytes. Peptides derived from osteopontin appear to suppress the synthesis of FGF-7, thereby delaying hair growth. Thus, osteopontin may be an important regulator of the hair growth cycle.

[0090] EDAR and XEDAR bind to Eda A1 and Eda A2, which are family members of ectodysplasin. XEDAR activates the NFκB signaling pathway and is involved in signaling during hair follicle morphogenesis. Studies of mouse gene knockout have shown that this signaling pathway results in abnormal hair follicle formation.

[0091] During hair follicle regeneration after skin wound healing, the presence of macrophage signaling is important for initiating new hair follicle formation. Lgr5+ cells are the first stem cells to be activated during the telogen-anagen transition in the hair cycle. Macrophages have been shown to activate these cells to proliferate via TNF signaling. Introduction of TNFR-1 via uptake of the present invention into Lgr5+ cells of the hair follicle bulb increases the sensitivity of these cells to respond to TNF signals and initiate hair follicle formation.

[0092] In summary, the protein characterization data provided by the present invention demonstrate the possibility that prominent protein elements can function to resume quiescent hair follicles via multiple modes of action (including, but not limited to, inhibition of diseases and modification or resolution of inflammatory conditions, activation of dermal cell, melanocyte, and keratinocyte cell proliferation, and induction of hair follicle cell differentiation and extracellular matrix production).

[0093] Accordingly, a method for preparing a therapeutic composition for treating, suppressing, reducing, preventing, and / or restoring alopecia (including, but not limited to, alopecia caused by male pattern baldness, androgenetic alopecia, alopecia areata, cicatricial alopecia, telogen effluvium, and / or female pattern baldness) is disclosed herein, the method comprising identifying an end-user's physical characteristics (such as skin color, skin type, ethnicity, race, hair type, and / or hair color, etc.), obtaining mesenchymal skin cells (MSCs) from a target donor having the same physical characteristics (such as skin color, skin type, ethnicity, race, hair type, and / or hair color) as the end-user, but who has never experienced alopecia and may be specific to gender, race, and ethnicity, and having a single nucleotide polymorphism (SNP) profile indicating this, preparing a powder preparation of growth factors for MSCs, keratinocytes, and melanocytes from the obtained MSCs, wherein the MSC preparation comprises at least one member selected from the group consisting of cells cultured under normal hypoxic culture conditions or cell conditioned medium, and cells cultured under harsh wound healing conditions, adding the powder preparation to a base, and the therapeutic composition comprises a member selected from the group consisting of topical compositions and injectable compositions. In one aspect, the therapeutic composition stimulates the activation of hair follicles to promote hair growth. In one aspect, the SNPs of the method for preparing the therapeutic composition comprise one or more SNPs present in one or more genes encoding interleukin 18 (IL-18), platelet-derived growth factor (PDGF) receptor (PDGFR) B (PDGFR-B), tissue metalloprotease inhibitor (TIMP) 1 (TIMP-1), TIMP-2, IL-23, activin A, intracellular adhesion molecule (ICAM-2), plasminogen activator inhibitor 1 (PAI-1), osteopontin (OPN), insulin, insulin growth factor binding protein 4 (IGF-BP4), tumor necrosis factor (TNF-R) receptor (TNF-R), neuregulin-1 B1 (NRG1-B1), urokinase-type plasminogen activator receptor (uPAR), ectodysplasin A2 receptor (XEDAR), follistatin, and / or activated leukocyte cell adhesion molecule (ALCAM).In one aspect, the SNPs include the SNPs rs1946518 (-607C>A) and / or rs187238 (-137G>C) of IL-18.

[0094] Also disclosed herein are therapeutic compositions for treating, suppressing, reducing, preventing, or restoring alopecia (including, but not limited to, alopecia caused by male pattern alopecia, androgenetic alopecia, alopecia areata, cicatricial alopecia, telogen effluvium, and / or female pattern alopecia). The compositions include a composition base and a powder preparation of a growth factor of mesenchymal stem cells (MSCs), keratinocytes, and / or melanocytes derived from a donor having the same physical characteristics as the end user (such as skin color, skin type, ethnicity, race, hair type, and / or hair color), but the donor never has alopecia and has a different single nucleotide polymorphism (SNP) profile that can be specific to gender, race, and ethnicity. The MSC preparation includes at least one member selected from the group consisting of cells cultured under normal hypoxic culture conditions or cell conditioned medium and cells cultured under severe wound healing conditions.

[0095] In one aspect, the SNPs of the therapeutic composition include one or more SNPs present in one or more genes encoding interleukin 18 (IL-18), platelet-derived growth factor (PDGF) receptor (PDGFR) B (PDGFR-B), tissue metalloprotease inhibitor 1 (TIMP) 1 (TIMP-1), TIMP-2, IL-23, activin A, intracellular adhesion molecule (ICAM-2), plasminogen activator inhibitor 1 (PAI-1), osteopontin (OPN), insulin, insulin growth factor binding protein 4 (IGF-BP4), tumor necrosis factor (TNF-R) receptor (TNF-R), neuregulin-1 B1 (NRG1-B1), urokinase-type plasminogen activator receptor (uPAR), ectodysplasin A2 receptor (XEDAR), follistatin, and / or activated leukocyte cell adhesion molecule (ALCAM). In one aspect, the SNPs include the SNPs rs1946518 (-607C>A) and / or rs187238 (-137G>C) of IL-18.

[0096] Also disclosed herein are methods of treating alopecia in a subject (including, but not limited to, alopecia caused by androgenetic alopecia, male pattern baldness, alopecia areata, cicatricial alopecia, telogen effluvium, and / or female pattern baldness), which comprise administering to the subject any of the therapeutic compositions disclosed herein.

[0097] 4. Erectile Dysfunction The proteomic evaluation of the exosome suspension generated using the methods and parameters described herein was performed to characterize the molecular composition contributing to the clinically significant efficacy of the described invention. The proteomic evaluation utilized a commercially available antibody array from RayBio Tech (Norcross, GA, USA). The concentrations of 230 different proteins known to either be secreted, transported, or present on the outer surface of the cell membrane (within the extracellular microenvironment) were measured. Proteins present at physiologically appropriate concentrations in duplicate test samples and supported by the scientific literature as potentially effective in the treatment of erectile dysfunction are listed in Table 2. Physiologically appropriate concentrations were considered to be those with an average concentration of 1 pg / mL or greater. A literature search of proteins with an average concentration exceeding 50 μg / mL was performed to identify the most likely candidate molecules supporting a clinical effect.

[0098]

Table 2

[0099] The protein content in the present invention having the ability to positively affect angiogenesis and function, somatic or autonomic innervation and function, or smooth muscle cell physiology may have a beneficial effect on erectile dysfunction. The research literature supports the potential effects of various protein components of the present invention that improve or resolve elements of the erectile dysfunction pathology.

[0100] IL18 was the most concentrated protein present in the exosome suspension sample. Its main function is involved in regulating the innate immune response in the skin. Specifically, it stimulates interferon-γ production and activates natural killer cells and TH1 T cells in the dermis. Excessive IL18 is associated with the onset of autoimmune diseases. An increase in IL-18 levels was observed in the obese male population with erectile dysfunction. There is evidence of IL18 isoforms, including the main form in serum that can function as an autocrine inhibitor. The form of IL18 in the present invention may be an inhibitory isoform that reduces the pro-inflammatory signaling of IL18 and regulates the immune response to an appropriate level. In one aspect, the SNP includes the SNPs rs1946518 (-607C>A) and / or rs187238 (-137G>C) of IL-18.

[0101] Platelet-rich plasma has been used in the treatment of ED. Platelet-derived growth factor (PDGF) receptor (PDGFR) signaling has been shown to be important in regulating the differentiation and contractility of cavernous smooth muscle cells and remodeling of connective tissue. Incorporation of PDGFR-B into the CCSMC cell membrane can increase the sensitivity or response rate to PDGF signals that improve the function of penile smooth muscle cells.

[0102] A common cause of erectile dysfunction is the accumulation of collagenous fibrous tissue in the penis. Tissue inhibitor of metalloproteinase (TIMP)-1 (TIMP-1) and TIMP-2 inhibitors play an important role in regulating the proteolytic activity of collagenase and other proteases. In a rat model of erectile dysfunction, TIMP-1 derived from mesenchymal stem cells has been shown to restore erectile function.

[0103] The smooth muscle-endothelial junction is important for establishing the anatomical compartmentalization necessary for penile erection. In a PAI-1 (plasminogen activator inhibitor-1) knockout mouse model of erectile dysfunction, recombinant PAI-1 restored the number of smooth muscle-endothelial junctions in the corpus cavernosum tissue and also induced a significant decrease in the time to maximal corpus cavernosum pressure. Smooth muscle-endothelial junctions were similarly identified in human corpus cavernosum tissue. These results suggest an important role for smooth muscle-endothelial junctions in erectile pathophysiology, and therapies such as the present invention may restore the number of smooth muscle-endothelial junctions in the corpus cavernosum tissue and provide a novel therapy for erectile dysfunction.

[0104] Osteopontin (OPN) is generally associated with bone matrix formation because it was first characterized in bone, but osteopontin has been shown to have additional functionality in other cell types. For example, it has been shown to have anti-apoptotic activity in cell types such as oligodendrocyte progenitor cells. Increased amounts of osteopontin were observed in a corpus cavernosum injury model of ED in rats. In this scenario, osteopontin is involved in regulating apoptosis of myelin-producing cells near nerves and may play a role in enabling cell survival and rapid repair of damaged tissue.

[0105] TNF has functions regulating cell proliferation and cell death in a number of different tissue types, and TNFR-1 (tumor necrosis factor receptor-1) has typically been described as an initiator of apoptosis signaling when complexed with TNF-α as a homodimer, but TNF receptors can interact with other members of the protein TNF receptor superfamily and can act as decoy molecules that downregulate the activity of TNF. In this capacity, TNFR-1 can function as an inhibitor of chronic inflammation signaling involved in the etiology of some forms of erectile dysfunction.

[0106] Other neuregulin family molecules (e.g., GGF2) have been shown to have a neuroprotective effect in prostatectomy-related nerve injury that results in erectile dysfunction. Similarly, NRG1-B1 (neuregulin-1 B1) has been shown to alter nerve function and significantly reduce pain in a rat model of neuropathic pain, and thus may support the growth of neurons and the recovery of damaged nerves resulting from the development of acute injury-related erectile dysfunction.

[0107] Neurons release uPAR (urokinase-type plasminogen activator receptor) during the recovery phase from ischemic injury, and astrocytes, axon terminal buttons, and dendritic spines mobilize uPAR to their plasma membranes. The binding of uPA to uPAR promotes the repair of synapses damaged by ischemic injury. The uptake of the uPA receptor by recipient neurons can increase the synaptic connectivity of the parasympathetic and sympathetic neural circuitry that is dysfunctional in some forms of erectile dysfunction.

[0108] ALCAM plays a role in immune, endothelial, and neuronal physiology. It has been shown to be important in maintaining the blood-brain barrier and in posterior root ganglion neuron axon elongation and cell proliferation. ALCAM has been identified by one study as the gene closest to two different SNPs associated with erectile dysfunction related to type I diabetes. The ALCAM of the present invention may serve to provide signals to endothelial and neuronal cells or to modify the immune state of penile tissue in erectile dysfunction that results in the restoration of tissue integrity and neural connectivity.

[0109] In summary, the protein characterization data provided by the present invention demonstrate the feasibility that prominent protein elements can function to initiate the repair or reduction of negative signaling events that cause erectile dysfunction. The proteins of the present invention can act directly on neurons and smooth muscle cells involved in the regulation of vascular pressure and fluid inflow and outflow, and can indirectly address external disease states that affect the anatomy and physiology of the penis.

[0110] Accordingly, a method of making a therapeutic composition for treating, suppressing, reducing, preventing, and / or restoring erectile dysfunction is disclosed herein, the method comprising identifying the end user's physical characteristics (e.g., skin color, skin type, ethnicity, race, hair type, and / or hair color, etc.), obtaining mesenchymal skin cells (MSCs) from a target donor having the same physical characteristics (e.g., skin color, skin type, ethnicity, race, hair type, and / or hair color) as the end user, but who has never experienced erectile dysfunction and may be specific to gender, race, and ethnicity, and having a single nucleotide polymorphism (SNP) profile indicating this, preparing a powder preparation of growth factors for MSCs, keratinocytes, and melanocytes from the obtained MSCs, wherein the MSC preparation comprises at least one member selected from the group consisting of cells cultured under normal oxygen culture conditions or cell conditioned medium, and cells cultured under harsh wound healing conditions, adding the powder preparation to a base, and the therapeutic composition comprises a member selected from the group consisting of topical compositions and injectable compositions. In one aspect, the therapeutic composition promotes appropriate erectile function.

[0111] Also disclosed herein are therapeutic compositions for treating, suppressing, reducing, preventing, or reversing erectile dysfunction, the compositions comprising a composition base and a powder preparation of growth factors of mesenchymal stem cells (MSCs), keratinocytes, and / or melanocytes derived from a donor having the same physical characteristics as the end user (e.g., skin color, skin type, ethnicity, race, hair type, and / or hair color, etc.), but who never has erectile dysfunction and has a different single nucleotide polymorphism (SNP) profile that may be specific to gender, race, and ethnicity, wherein the MSC preparation comprises at least one member selected from the group consisting of cells cultured under normal oxygen culture conditions or cell conditioned media and cells cultured under harsh wound healing conditions. In one aspect, the SNPs of the method of making the therapeutic composition comprise one or more SNPs present in one or more genes encoding interleukin 18 (IL-18), platelet-derived growth factor (PDGF) receptor (PDGFR) B (PDGFR-B), tissue metalloprotease inhibitor 1 (TIMP) 1 (TIMP-1), TIMP-2, IL-23, activin A, intracellular adhesion molecule (ICAM-2), plasminogen activator inhibitor 1 (PAI-1), osteopontin (OPN), insulin, insulin growth factor binding protein 4 (IGF-BP4), tumor necrosis factor (TNF-R) receptor (TNF-R), neuregulin-1 B1 (NRG1-B1), urokinase-type plasminogen activator receptor (uPAR), ectodysplasin A2 receptor (XEDAR), follistatin, and / or activated leukocyte cell adhesion molecule (ALCAM). In one aspect, the SNPs comprise the SNPs rs1946518 (-607C>A) and / or rs187238 (-137G>C) of IL-18.

[0112] In one aspect, the SNPs of the therapeutic composition comprise one or more SNPs present in one or more genes encoding interleukin 18 (IL-18), platelet-derived growth factor (PDGF) receptor (PDGFR) B (PDGFR-B), tissue metalloprotease inhibitor 1 (TIMP) 1 (TIMP-1), TIMP-2, IL-23, activin A, intracellular adhesion molecule (ICAM-2), plasminogen activator inhibitor 1 (PAI-1), osteopontin (OPN), insulin, insulin growth factor binding protein 4 (IGF-BP4), tumor necrosis factor (TNF-R) receptor (TNF-R), neuregulin-1 B1 (NRG1-B1), urokinase-type plasminogen activator receptor (uPAR), ectodysplasin A2 receptor (XEDAR), follistatin, and / or activated leukocyte cell adhesion molecule (ALCAM). In one aspect, the SNPs comprise the SNPs rs1946518 (-607C>A) and / or rs187238 (-137G>C) of IL-18.

[0113] Also disclosed herein is a method of treating erectile dysfunction in a subject, which comprises administering to the subject any of the therapeutic compositions disclosed herein.

[0114] 5. Mesenchymal stem cells As pointed out throughout, the therapeutic compositions disclosed herein can utilize exosomes and / or growth factors derived from mesenchymal stem cells (MSCs). Existing autologous and allogeneic MSCs contained within bone marrow concentrates or adipose-derived stromal vascular fraction (SVF), or various postpartum products derived from umbilical cord, placenta, or amnion, but expanded MSC cultures are currently used to treat wounds, orthopedic pathologies, and spinal pathologies. Existing treatments do not contain a large amount of MSC secretome (including, but not limited to, growth factors, cytokines, chemokines, exosomes, extracellular vesicles, and / or extracts). Further, while there is evidence in the art that treatments containing stem cells (including injectable treatments) can be useful in preventing aging and treating scars and uneven pigmentation, existing skin products (creams, lotions, serums, cosmetics, etc.) contain ingredients that potentially can be useful in treating and strengthening the skin, while other topical products do not penetrate the epidermis and, more importantly, do not contain human MSCs, or MSC-derived growth factors and proteins. Indeed, prior to the present disclosure, no active MSC growth factor products have been developed that can be used for these applications. Accordingly, in one aspect, an MSC secretome composition (including, but not limited to, MSC growth factors, MSC exosomes, MSC extracts, and / or extracellular vesicles containing the composition) for use in treating wounds, orthopedic disorders, orthopedic injuries, ophthalmology, spinal cord injuries, or spinal cord disorders is disclosed herein, the therapeutic composition comprising (i) a powdered additive of growth factors comprising a preparation derived from mesenchymal stem cells (MSCs), and (ii) a pharmaceutically acceptable carrier.

[0115] As described above, MSC are multipotent cells with the ability to differentiate into a number of cell types, including myocytes, chondrocytes, adipocytes, and osteoblasts. Typically, these cells can be found in amniotic fluid, including placenta, umbilical cord blood, adipose tissue, bone marrow, or perivascular tissue. As used herein, "MSC" refers to non-terminally differentiated cells and includes, but is not limited to, multipotent stem cells, multipotent stromal cells, stromal vascular cells, pericytes, perivascular cells, stromal cells, multipotent cells, multipotent cells, adipose-derived fibroblast-like cells, adipose-derived stromal vascular cell populations, adipose-derived MSC, bone marrow-derived fibroblast-like cells, bone marrow-derived stromal vascular cell populations, bone marrow-derived MSC, tissue-derived fibroblast-like cells, adult stem cells, adult stromal cells, keratinocytes, and / or melanocytes.

[0116] In addition to their differentiation potential, MSC have long been recognized to have immunomodulatory capabilities that result in the expression of many different cytokines and growth factors. As used herein, "MSC preparation" or "MSC secretome composition" refers to a composition comprising MSC growth factors, MSC exosomes, extracellular vesicles, or cell-free extracts or lysates of MSC obtained from human MSC, fibroblast-like cells, and non-human animal MSC (including, but not limited to, MSC from horse, cow, pig, sheep, non-human primate, dog, cat, rabbit, rat, and mouse). In embodiments, the MSC can be derived from the patient (autologous) to whom the composition is applied or from another individual (allogeneic). The MSC may be grown in culture to recover conditioned media, or to increase the amount of cells in the lysate, or for fresh use prior to incorporation into the compositions of the disclosure.

[0117] An MSC secretome composition (including, but not limited to, MSC growth factors, MSC exosomes, MSC extracts, and / or extracellular vesicles including the composition) may contain mesenchymal stem cell (MSC) extracts, MSC exosomes, or MSC growth factor preparations between about 0.00001% and about 20% by weight (e.g., between about 0.01% and about 10% by weight). The MSC preparation may include either MSC conditioned medium or an MSC lysate derived from cell culture-expanded MSCs. In some embodiments, the composition may further include a cell-free medium conditioned by the growth of about 0.01% to about 10% by weight of MSCs or MSC lineage cells, and the cells are cultured under normal hypoxic culture conditions or artificial wound healing conditions.

[0118] As disclosed herein, the MSCs used to produce the disclosed MSC additives (including growth factor secretome compositions, either as frozen additives or powder additives) can be selectively stimulated to produce MSC growth factors, secretomes, cytokines, chemokines, mesenchymal stem cell proteins, peptides, glycosaminoglycans, extracellular matrix (ECM), proteoglycans, secretomes, and exosomes. As used herein, MSC growth factors include, but are not limited to, prostaglandin E2 (PGE2), transforming growth factor 1 (TGF-β1), hepatocyte growth factor (HGF), stromal cell-derived factor-1 (SDF-1), nitric oxide, indoleamine 2,3-dioxygenase, interleukin-4 (IL-4), IL-6, interleukin-10 (IL-10), IL-1 receptor antagonist and soluble TNF-α receptor, insulin-like growth factor, fibroblast growth factor (FGF) 1-23 (especially FGF1 and FGF2), bone morphogenetic protein (BMP) 1-15, epidermal growth factor (EGF), transforming growth factor-α (TGF-α), macrophage stimulating protein (MSP), platelet-derived growth factor (PLGF), vascular endothelial growth factor (VEGF), macrophage colony stimulating factor (M-CSF), insulin, granulocyte colony stimulating factor (G-CSF), granulocyte macrophage colony stimulating factor (GM-CSF), and hormones including estrogen and thyroid hormone.

[0119] In one aspect, the MSC preparation (e.g., an MSC secretome composition, etc.) comprises MSC growth factors, MSC exosomes, and / or cell extracts or lysates of MSCs obtained from MSCs cultured under standard hyperoxic culture conditions (e.g., 21% oxygen) or MSCs cultured under artificial wound healing conditions (e.g., 0.1% - about 5% oxygen in the presence of inflammatory cytokines, angiogenic factors, and low glucose).

[0120] As disclosed herein, the artificial wound healing state simulates the proliferative state in an actual wound with reduced nutrient supply and reduced waste removal (usually caused by disruption of local blood circulation). This creates a harsh environment for cells until new blood vessels are formed and blood circulation is restored. Thus, the artificial wound healing conditions used to culture MSCs can include one or more of the following growth conditions: reduced glucose availability, reduced oxygen partial pressure, reduced pH, and increased temperature.

[0121] In one aspect, glucose availability can be reduced compared to a normal control. A modified culture medium that reduces glucose but does not damage the cells can be a reduction of 0 - 50% of glucose, more preferably a reduction of about 5% - 40% of glucose. For example, the artificial wound healing culture conditions for MSCs can include a glucose reduction of about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, or 50% glucose reduction (e.g., about 5% - about 15%, about 10% - about 20%, about 15% - about 25%, about 20% - about 30%, or about 25% - about 35%).

[0122] In one aspect, the oxygen partial pressure can be reduced to a hypoxic state. Normal atmospheric oxygen is about 21%, and any reduction is considered hypoxic. Thus, in one aspect, the MSCs can be cultured under artificial wound healing conditions with 0.0% to 20.9% oxygen, about 0.1% to about 0.5% oxygen, about 0.1% to about 2.0% oxygen, about 0.1% to about 5.0% oxygen, about 0.5% to 5.0% oxygen, about 1.0% to about 10% oxygen, about 5.0% to about 10.0% oxygen, and about 10.0% to about 15.0% oxygen. Preferably, in the wound healing culture conditions of the MSCs, the oxygen partial pressure is about 0.5% to 20.5% oxygen, for example, 0, 0.05, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.7, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, 4.0, 4.1, 4.2, 4.3, 4.4, 4.5, 4.6, 4.7, 4.8, 4.9, 5.0, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9, 6.0, 6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, 7.0, 7.1, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9, 8.0, 8.1, 8.2, 8.3, 8.4, 8.5, 8.6, 8.7, 8.8, 8.9, 9.0, 9.1, 9.2, 9.3, 9.4, 9.5, 9.6, 9.7, 9.8, 9.9, 10, 10.5, 11, 11.5, 12, 12.5, 13, 13.5, 14, 14.5, 15, 15.5, 16, 16.5, 17, 17.5, 18, 18.5, 19, 19.5, 20, or 20.5% oxygen.

[0123] The pH can also be reduced under artificial wound healing conditions. Physiological pH is very tightly maintained and is usually very close to neutral pH = 7.2 ± 0.2 (7.0 to 7.4). However, in a wound, the acidic environment can have a pH = 6.2 ± 0.2 (i.e., a pH of 6.0 to about 6.4). Thus, under artificial wound healing culture conditions, the pH can be from about 6.0 to about 7.4, for example, 6.0 to about 6.4, about 6.2 to about 6.4, about 6.2 to about 6.6, about 6.4 to about 6.6, about 6.4 to about 6.8, or about 6.6 to about 7.0 (such as 6.0, 6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, 7.0, 7.1, 7.2, 7.3, or 7.4).

[0124] Under artificial wound healing culture conditions, the temperature of the culture environment can be increased to simulate the temperature increase at the wound site. Physiological homeostatic temperature is maintained at 37 °C (98.6 o F). Even a slight increase or decrease can cause significant changes in cell metabolism. By increasing the temperature to any temperature up to about 40 °C (104 o F) above 37 °C, a "pyrogenic" environment can be created. Thus, in one aspect, the artificial wound healing culture conditions for MSCs can include from about 35 °C to about 39 °C, from about 35 °C to about 36 °C, from about 36 °C to about 37 °C, from about 37 °C to about 38 °C, from about 38 °C to about 39 °C, from about 39 °C to about 40 °C. In one aspect, the temperature of the artificial wound healing culture can be 35.0, 35.1, 35.2, 35.3, 36.4, 35.5, 35.6, 35.7, 35.8, 35.9, 36.0, 36.1, 36.2, 36.3, 36.4, 36.5, 36.6, 36.7, 36.8, 36.9, 37.0, 37.1, 37.2, 37.3, 37.4, 37.5, 37.6, 37.7, 37.8, 37.9, 38.0, 38.1, 38.2, 38.3, 38.4, 38.5, 38.6, 38.7, 38.8, 38.9, 39.0, 39.1, 39.2, 39.3, 39.4, 39.5, 39.6, 39.7, 39.8, 39.9, or 40.0 °C.

[0125] In one aspect, the MSC secretome composition (including, but not limited to, MSC growth factors, MSC exosomes, MSC extracts, and / or extracellular vesicles including the composition) can further include a protective coating agent (e.g., cryoprotective oligosaccharides and protein solutions, etc.) to reduce the degradation of growth factors. It is understood that the protective coating agent can be manipulated as a polymer, which is contemplated herein. "Polymer" refers to a relatively high molecular weight natural or synthetic organic compound whose structure can be represented by monomers which are repeating small units. Non-limiting examples of polymers include polyethylene, rubber, and cellulose. Synthetic polymers are typically formed by the addition or condensation polymerization of monomers. The term "copolymer" refers to a polymer formed from two or more different repeating units (monomer residues). By way of example, and not limitation, the copolymer can be an alternating copolymer, a random copolymer, a block copolymer, or a graft copolymer. In certain aspects, it is also contemplated that the various block segments of a block copolymer can themselves include copolymers. The term "polymer" encompasses all forms of polymers, including, but not limited to, natural polymers, synthetic polymers, homopolymers, heteropolymers, or copolymers, addition polymers, etc. In one aspect, the gel matrix can include copolymers, block copolymers, diblock copolymers, and / or triblock copolymers.

[0126] In one aspect, the protective coating agent may include a biocompatible polymer. In one aspect, the biocompatible polymer may be crosslinkable. Such a polymer may also serve to gradually release a fat browning agent and / or a fat regulator into the tissue. Biocompatible polymers used herein include polysaccharides, hydrophilic polypeptides, poly(amino acids) (e.g., poly-L-glutamic acid (PGS), gamma-polyglutamic acid, poly-L-aspartic acid, poly-L-serine, or poly-L-lysine), polyalkylene glycols and polyalkylene oxides (e.g., polyethylene glycol (PEG), polypropylene glycol (PPG), and poly(ethylene oxide) (PEO)), poly(oxyethylated polyols), poly(olefin alcohols), polyvinylpyrrolidone, poly(hydroxyalkyl methacrylamide), poly(hydroxyalkyl methacrylate), poly(saccharides), poly(hydroxy acids), poly(vinyl alcohol), polyhydroxy acids (e.g., poly(lactic acid), poly(glycolic acid), and poly(lactic-co-glycolic acid)), polyhydroxyalkanoic acids (e.g., poly-3-hydroxybutyric acid or poly-4-hydroxybutyric acid), polycaprolactone, poly(orthoesters), polyanhydrides, poly(phosphazenes), poly(lactide-co-caprolactone), polycarbonates (e.g., tyrosine polycarbonate), polyamides (including synthetic and natural polyamides), polypeptides, and poly(amino acids)), polyester amides, polyesters, poly(dioxanone), poly(alkylene alkylate), hydrophobic polyesters, polyurethanes, polyether esters, polyacetals, polycyanoacrylates, polyacrylates, polymethyl methacrylate, polysiloxanes, poly(oxyethylene) / poly(oxypropylene) copolymers, polyketals, polyphosphoric acid, polyhydroxyvaleric acid, polyalkylene succinic acid, polyalkylene adipic acid, poly(maleic acid), and copolymers thereof, but are not limited thereto.In addition, biocompatible polymers may include polyamides, polycarbonates, polyalkylenes, polyalkylene glycols, polyalkylene oxides, polyalkylene terephthalates, polyvinyl alcohol (PVA), methacrylate PVA (m-PVA), polyvinyl ethers, polyvinyl esters, halogenated polyvinyls, polyvinyl pyrrolidone, polyglycolide, polysiloxanes, polyurethanes and their copolymers, alkyl celluloses, hydroxyalkyl celluloses, cellulose ethers, nitrocellulose, polymers of acrylic acid esters and methacrylic acid esters, methyl cellulose, ethyl cellulose, hydroxypropyl cellulose, hydroxypropyl methyl cellulose, hydroxybutyl methyl cellulose, cellulose acetate, cellulose propionate, cellulose acetate butyrate, cellulose acetate phthalate, carboxyethyl cellulose, cellulose triacetate, sodium salt of cellulose sulfate, poly(methyl methacrylate), poly(ethyl methacrylate), poly(butyl methacrylate), poly(isobutyl methacrylate), poly(hexyl methacrylate), poly(isodecyl methacrylate), poly(lauryl methacrylate), poly(phenyl methacrylate), poly(methyl acrylate), poly(isopropyl acrylate), poly(isobutyl acrylate), poly(octadecyl acrylate), polyethylene, polypropylene, poly(ethylene glycol), poly(ethylene oxide), poly(ethylene terephthalate), poly(vinyl alcohol), poly(vinyl alcohol), poly(vinyl acetate), polyvinyl chloride, polystyrene and polyvinyl pyrrolidone, their derivatives, linear and branched copolymers, as well as their block copolymers, and blends thereof. Exemplary biodegradable polymers include polyesters, poly(orthoesters), poly(ethylene amines), poly(caprolactone), poly(hydroxybutyric acid), poly(hydroxyvaleric acid), polyanhydrides, poly(acrylic acid), polyglycolide, poly(urethane), polycarbonate, polyphosphate esters, polyphosphazenes, their derivatives, linear and branched copolymers and their block copolymers, and blends thereof.

[0127] In some embodiments, the protective coating agent includes carbohydrate constructs of carbohydrate polymers such as monosaccharides, as well as disaccharides or polysaccharides (including, but not limited to, non-reducing polysaccharides or disaccharides) and any combination thereof. Examples of carbohydrates that can be used in the protective coating agent include glucose, aldoses (such as D-allose, D-altrose, D-mannose), glucopyranose, pentahydroxyhexanal, α-D-glucopyranosyl-D-glucose, α-D-glucopyranosyl dihydrate, polymers of β-D-glycopyranosyl units, β-D-fructofuranosyl α-D-glucopyranoside (anhydride / dihydrate), β-D-galactopyranosyl-D-glucose, β-D-glucopyranosyl-α-D-glucopyranoside (anhydride / dihydrate), galactose, pentoses (such as ribose, xylose, lyxose), dextrose, dodecacarboxylic monodecahydrate, fructose, sucrose, lactose, maltose, trehalose, agarose, D-galactosyl-β-(1-4)-anhydro-L-galactosyl, cellulose, polymers of β-D-glycopyranosyl units, and starch, as well as polyhydric alcohols, polyalcohols, alditols, erythritol, glycitol, glycerol, xylitol, and sorbitol.

[0128] In some embodiments, the protective coating agent contains biocompatible and / or biodegradable polyesters or polyanhydrides such as poly(lactic acid), poly(glycolic acid), and poly(lactic-co-glycolic acid). The particles may include one or more of the following polyesters: homopolymers containing glycolic acid units (referred to herein as "PGA"), as well as homopolymers containing lactic acid units (such as poly-L-lactic acid, poly-D-lactic acid, poly-D,L-lactic acid, poly-L-lactide, poly-D-lactide, and poly-D,L-lactide 5, collectively referred to herein as "PLA"), as well as homopolymers containing caprolactone units (such as poly(ε-caprolactone), collectively referred to herein as "PCL"), as well as copolymers containing units of lactic acid and glycolic acid (such as various forms of poly(lactic-co-glycolic acid) and poly(lactide-co-glycolide) characterized by the ratio of lactic acid to glycolic acid, collectively referred to herein as "PCL"), as well as their derivatives. Exemplary polymers also include copolymers of polyethylene glycol (PEG) and the aforementioned polyesters (such as various forms of PLGA-PEG or PLA-PEG copolymers, collectively referred to herein as "PEGylated polymers"). In certain embodiments, the PEG region can be covalently bonded to the polymer to obtain a "PEGylated polymer" with a cleavable linker. In one aspect, the polymer contains at least 60, 65, 70, 75, 80, 85, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, or 99 percent acetal pendant groups.

[0129] The triblock copolymers disclosed in this specification include, for example, copolymers such as polyethylene glycol (PEG), polyvinyl acetate, polyvinyl alcohol, polyvinyl pyrrolidone (PVP), polyethylene oxide (PEO), poly(vinyl pyrrolidone-co-vinyl acetate), polymethacrylate, polyoxyethylene alkyl ether, polyoxyethylene castor oil, polycaprolactam, polylactic acid, polyglycolic acid, poly(lactic-glycolic) acid, poly(lactic-co-glycolic) acid (PLGA), cellulose derivatives (such as hydroxymethyl cellulose, hydroxypropyl cellulose, etc.).

[0130] Examples of diblock copolymers that can be used in the protective coating agents disclosed in this specification include, for example, polymers such as polyethylene glycol (PEG), polyvinyl acetate, polyvinyl alcohol (PVA), polyvinyl pyrrolidone (PVP), polyethylene oxide (PEO), poly(vinyl pyrrolidone-co-vinyl acetate), polymethacrylate, polyoxyethylene alkyl ether, polyoxyethylene castor oil, polycaprolactam, polylactic acid, polyglycolic acid, poly(lactic-glycolic) acid, poly(lactic-co-glycolic) acid (PLGA).

[0131] In one aspect, the protective coating agent may further include lecithin or hydrolyzed lecithin as a carrier or encapsulating material (i.e., encapsulated, encapsulated composition). As used herein, lecithin and / or hydrolyzed lecithin coating agents include coatings containing phosphatidylcholine, phosphatidylinositol, phosphatidylethanolamine, phosphatidylserine, and phosphatidic acid. The source of lecithin can be a plant or animal source.

[0132] In one aspect, any of the polymers, monosaccharides, disaccharides, or polysaccharides used to form the protective coating agent, which is formed by placing the MSC additive in the encapsulation solution, can be at a concentration suitable for forming the protective coating agent. For example, the polymer, monosaccharide, disaccharide, or polysaccharide can be at any concentration from 0.01 mM to 10.0 M (e.g., from about 0.01 M to about 0.1 M, from about 0.1 mM to about 1.0 M, from about 1.0 M to about 10.0 M). Exemplary concentrations include 0.01, 0.02, 0.03, 0.04, 0.05, 0.06, 0.07, 0.08, 0.09, 0.1, 0.2, 0.3, 0.4, 0.4, 0.6, 0.7, 0.8, 0.9, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 225, 250, 275, 300, 325, 350, 375, 400, 450, 500, 600, 700, 800, 900 mM, 1, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2, 3, 4, 5, 6, 7, 8, 9, 10 M.

[0133] As shown in FIGS. 1 and 2, exosomes and extracellular vesicles in the disclosed MSC secretome composition were produced.

[0134] In one aspect, one way to treat a wound is by administering an MSC secretome composition (including, but not limited to, MSC growth factors, MSC exosomes, MSC extracts, and / or extracellular vesicles comprising the composition) subcutaneously, intramuscularly, intravenously, locally (e.g., through the use of salves, creams, and / or ointments), although it is also understood and contemplated herein that it is by impregnating a stent, sponge, matrix, scaffold, bandage, dressing, suture, graft, surgical drape, surgical adhesive, and / or staple with the MSC secretome composition. Thus, in one aspect, a medicated stent, scaffold, sponge, matrix, adhesive bandage, wound dressing, graft, surgical drape, suture, salve, cream, or wound adhesive comprising a therapeutically effective amount of an MSC secretome composition is disclosed herein. As described above, the MSC secretome composition (including, but not limited to, MSC growth factors, MSC exosomes, MSC extracts, and / or extracellular vesicles comprising the composition) can be administered locally and applied to the face, neck, hands, or any other desired part of the body. When applied to an adhesive bandage, wound dressing, graft, surgical drape, suture, scaffold, scaffold, matrix, sponge, or stent, the MSC secretome composition can be applied as a powder.

[0135] In one aspect, the MSC secretome compositions disclosed herein (including, but not limited to, MSC growth factors, MSC exosomes, MSC extracts, and / or extracellular vesicles comprising the compositions) may include any known components typically found in the field of wound healing (e.g., oils, waxes, or other standard lipid substances), or conventional gelling agents and / or thickening agents, emulsifiers, humectants, emollients, sunscreens, hydrophilic or lipophilic activators (e.g., ceramides), agents for dealing with free radicals, bactericides, occlusives, preservatives, basifying or acidifying agents, fragrances, surfactants, fillers, natural products or extracts of natural products (e.g., aloe or green tea extract), vitamins, or colorants. Other components that can be combined with the powder may include antioxidants that can be selected from various antioxidants. Suitable antioxidants include vitamin C (L-ascorbic acid, magnesium ascorbyl-2-phosphate, ascorbyl palmitate, tetrahexyldecyl ascorbate), vitamin E (tocotrienols), vitamin A (retinol, retinal, retinoic acid, provitamin A carotenoids (e.g., β-carotene)), N-acetylglucosamine, or other derivatives of glucosamine. Other components may include at least one essential fatty acid such as omega-3, omega-6, and omega-9 polyunsaturated fatty acids (e.g., linoleic acid (LA), gamma-linolenic acid (GLA), alpha-linolenic acid (ALA), dihomo-gamma-linolenic acid (DGLA), arachidonic acid (ARA), and others). The fatty acids may be derived from various sources including evening primrose oil, black currant oil, borage oil, or GLA-modified safflower seed. Other components may include at least one component that supports the production of platelet-rich fibrin matrix, ECM, and hyaluronic acid (e.g., N-acetylglucosamine or other derivatives of glucosamine, ultra-low molecular weight (ULMW) hyaluronic acid, chondroitin sulfate, or keratin sulfate).

[0136] The MSC secretome compositions disclosed herein are understood to and contemplated herein to be capable of providing wound healing, rejuvenation, enhancement, and improved or restored skin tissue. This composition can also be used as an injectable for the treatment of arthritis and degenerated intervertebral discs. Further, embodiments of the composition may not require the inclusion of additional growth factors or hormones such as insulin, insulin-like growth factors, thyroid hormones, fibroblast growth factors, estrogen, retinoic acid. In some aspects, the disclosed stem cell growth factor compositions can include additional active ingredients, including, but not limited to, antibiotics, anti-acne agents, liposomes, antioxidants, platelet-rich fibrin matrices, analgesics, anti-inflammatory agents, and additional growth factors (e.g., insulin, insulin-like growth factors, thyroid hormones, fibroblast growth factors, estrogen, retinoic acid). Such additional active ingredients are mixed with the stem cell growth factors and extracellular vesicle compositions disclosed herein, as well as MSC conditioned media, MSC lysates, and MSC-derived products, and then thawed or dissolved, mixed, or suspended in a mixture of emulsified lanolin alcohol, wax, and oil, or a mixture of petrolatum or mineral oil, quaternary ammonium compounds, aliphatic alcohols, and fatty acid ester softeners, or a lotion having a substantially similar composition.

[0137] 6. Pharmaceutical Carrier / Pharmaceutical Product Delivery As described above, the composition can be administered in vivo in a pharmaceutically acceptable carrier. "Pharmaceutically acceptable" means a material that is not biologically or otherwise undesirable, i.e., a material that can be administered to a subject with a nucleic acid or vector without causing any undesirable biological effects or interacting in a harmful manner with any of the other components of the pharmaceutical composition in which it is contained. Carriers will be known to those of skill in the art and will be naturally selected to minimize any degradation of the active ingredient and to minimize any harmful side effects in the subject.

[0138] The composition can be administered orally, parenterally (e.g., intravenously), by intramuscular injection, by intraperitoneal injection, transdermally, ex vivo, locally, etc. (including administration by topical intranasal administration or inhalation). As used herein, "topical intranasal administration" means delivery of the composition to the nose and nasal passages through one or both nostrils and can include delivery by a spray or droplet mechanism or by aerosolization of the nucleic acid or vector. Administration of the composition by inhalation can be effected through the nose or mouth via delivery by a spray or droplet mechanism. Delivery can also be directly to any area of the respiratory system (e.g., the lungs) via intubation. The exact amount of the composition required will vary depending on the species, age, weight, and general condition of the subject, the severity of the allergic disorder being treated, the particular nucleic acid or vector used, its mode of administration, etc. Thus, it is not possible to specify an exact amount for all compositions. However, an appropriate amount can be determined by one of ordinary skill in the art using only routine experimentation as provided in the teachings herein.

[0139] Parenteral administration of the composition, when used, is generally characterized by injection. Injectables can be prepared in conventional forms, either as liquid solutions or suspensions, solid forms suitable for solution of the suspension in a liquid prior to injection, or as emulsions. More recently revised approaches for parenteral administration involve the use of sustained release or controlled release systems so as to maintain a constant dosage. See, e.g., U.S. Patent No. 3,610,795, which is incorporated herein by reference.

[0140] The materials can be in solution or suspension (e.g., incorporated into microparticles, liposomes, or cells). These can target specific cell types via antibodies, receptors, or receptor ligands. The following references are examples of the use of this technology for targeting specific proteins in tumor tissue (Senter, et al., Bioconjugate Chem., 2:447-451, (1991), Bagshawe, K.D., Br. J. Cancer, 60:275-281, (1989), Bagshawe, et al., Br. J. Cancer, 58:700-703, (1988), Senter, et al., Bioconjugate Chem., 4:3-9, (1993), Battelli, et al., Cancer Immunol Immunother., 35:421-425, (1992), Pietersz and McKenzie, Immunolog. Reviews, 129:57-80, (1992), and Roffler, et al., Biochem. Pharmacol, 42:2062-2065, (1991)). "Stealth" and other antibody-conjugated liposomes (including lipid-mediated drug targeting to colon cancer), receptor-mediated targeting of DNA via cell-specific ligands, lymphocyte-specific tumor targeting, and highly specific therapeutic retroviral targeting of mouse glioma cells in vivo, etc. vehicles. The following references are examples of the use of this technology for targeting specific proteins in tumor tissue (Hughes et al., Cancer Research, 49:6214-6220, (1989), and Litzinger and Huang, Biochimica et Biophysica Acta, 1104:179-187, (1992)). Generally, receptors are involved in the endocytosis pathway either constitutively or ligand-induced. These receptors cluster within clathrin-coated pits, enter the cell via clathrin-coated vesicles, pass through acidified endosomes where the receptors are sorted, and then are either recycled to the cell surface, stored intracellularly, or degraded in lysosomes.Internal trafficking pathways serve various functions such as nutrient uptake, removal of activating proteins, clearance of macromolecules, opportunistic entry of viruses and toxins, dissociation and degradation of ligands, and regulation of receptor levels. Many receptors follow two or more intracellular pathways depending on cell type, receptor concentration, ligand type, ligand valency, and ligand concentration. The molecular and cellular mechanisms of receptor-mediated endocytosis have been outlined (Brown and Greene, DNA and Cell Biology 10:6, 399-409 (1991)).

[0141] a) A pharmaceutically acceptable carrier Compositions containing antibodies can be used for treatment in combination with a pharmaceutically acceptable carrier.

[0142] Suitable carriers and their formulations are described in Remington: The Science and Practice of Pharmacy (19th ed.) ed. A.R. Gennaro, Mack Publishing Company, Easton, PA 1995. Typically, appropriate amounts of pharmaceutically acceptable salts are used in the formulation to render it isotonic. Examples of pharmaceutically acceptable carriers include, but are not limited to, physiological saline, Ringer's solution, and dextrose solutions. The pH of the solution is preferably from about 5 to about 8, more preferably from about 7 to about 7.5. Further carriers include sustained release preparations such as a semipermeable matrix of a solid hydrophobic polymer containing an antibody where the matrix is in the form of a shaped article (e.g., film, liposome, or microparticle). For example, it will be apparent to those skilled in the art that certain carriers may be more preferred depending on the route of administration and the concentration of the composition being administered.

[0143] Pharmaceutical carriers are known to those skilled in the art. These are most typically standard carriers for drug administration to humans, including solutions such as sterile water, physiological saline, and buffer solutions at physiological pH. The composition can be administered intramuscularly or subcutaneously. Other compounds will be administered according to standard procedures used by those skilled in the art.

[0144] In addition to the selected molecule, the pharmaceutical composition can include a carrier, thickening agent, diluent, buffer, preservative, surfactant, etc. The pharmaceutical composition can also include one or more active ingredients (such as antibacterial agents, anti-inflammatory agents, anesthetics, etc.).

[0145] The pharmaceutical composition can be administered in several ways depending on whether local or systemic treatment is desired and on the area to be treated. Administration can be local (including eye, vagina, rectum, intranasal), oral, by inhalation, or parenterally (e.g., intravenous drip, subcutaneous, intraperitoneal, or intramuscular injection). The disclosed antibodies can be administered intravenously, intraperitoneally, intramuscularly, subcutaneously, intracoelomically, or transdermally.

[0146] Preparations for parenteral administration include sterile aqueous or non-aqueous solutions, suspensions, and emulsions. Examples of non-aqueous solvents are vegetable oils such as propylene glycol, polyethylene glycol, olive oil, and organic esters for injection such as ethyl oleate. Aqueous carriers include water, alcohol / aqueous solutions, emulsions, or suspensions containing physiological saline and buffer media. Parenteral vehicles include sodium chloride solution, Ringer's dextrose, dextrose and sodium chloride, lactated Ringer's, or fixed oils. Intravenous vehicles include fluids and nutrient replenishers, electrolyte replenishers (such as those based on Ringer's dextrose), etc. Also, preservatives and other additives, such as antibacterial agents, antioxidants, chelating agents, and inert gases, may be present.

[0147] Formulations for topical administration may include ointments, lotions, creams, gels, drops, suppositories, sprays, liquids, and powders. Conventional pharmaceutical carriers, aqueous, powder or oily bases, thickeners, etc. may be necessary or desirable if needed or desired.

[0148] Compositions for oral administration include powders or granules, suspensions or solutions in water or non-aqueous media, capsules, sachets, or tablets. Thickeners, flavorings, diluents, emulsifiers, dispersion aids or binders may be desirable if needed or desired.

[0149] Some compositions may potentially be administered as pharmaceutically acceptable acid or base addition salts formed by the reaction of inorganic acids (e.g., hydrochloric acid, hydrobromic acid, perchloric acid, nitric acid, thiocyanic acid, sulfuric acid, and phosphoric acid) with organic acids (e.g., formic acid, acetic acid, propionic acid, glycolic acid, lactic acid, pyruvic acid, oxalic acid, malonic acid, succinic acid, maleic acid, and fumaric acid), or by the reaction of inorganic bases (e.g., sodium hydroxide, ammonium hydroxide, potassium hydroxide) with organic bases (e.g., mono-, di-, tri-alkylamines, and arylamines, and substituted ethanolamines).

[0150] b) Therapeutic use Effective dosages and schedules for administering the compositions are determined empirically and making such determinations is within the skill of the art. The dosage range for administration of the compositions is large enough to produce the desired effect on the symptoms of the disorder. The dosage should not be so large as to cause side effects such as unwanted cross-reactions, anaphylactic reactions, etc. Generally, the dosage will vary depending on the patient's age, condition, sex, and degree of the disease, the route of administration, or whether other drugs are included in the regimen and can be determined by one of ordinary skill in the art. The dosage can be adjusted by the individual physician in the event of any contraindications. The dosage may vary and may be administered in doses of more than once a day. Guidance can be found in the literature for the appropriate dosage of a given class of pharmaceuticals. For example, guidance for selecting an appropriate dosage of an antibody can be found in the literature regarding the therapeutic use of antibodies, e.g., Handbook of Monoclonal Antibodies,Ferrone et al.,eds.,Noges Publications,Park Ridge,N.J.,(1985)ch.22 and pp.303-357, Smith et al.,Antibodies in Human Diagnosis and Therapy,Haber et al.,eds.,Raven Press,New York(1977)pp.365-389. The typical daily dosage of an antibody used alone can range from about 1 μg / kg body weight to a maximum of 100 mg / kg body weight, or more, depending on the above factors.

[0151] The foregoing examples are presented to provide a complete disclosure and description of the methods of making and evaluating the compounds, compositions, articles, devices, and / or methods claimed herein to one of ordinary skill in the art and are intended to be purely exemplary and not limiting of the disclosure. Although efforts have been made to ensure accuracy with respect to numerical values (e.g., amounts, temperatures, etc.), some error and deviation should be accounted for. Unless otherwise indicated, parts are by weight, temperature is in °C or ambient temperature, and pressure is at or near atmospheric pressure.

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Recombinant PAI-1 therapy restores myoendothelial junctions and erectile function in PAI-1-deficient mice. Andrologia. 2015 Dec;47(10):1147-52. Kawabe TT, Rea TJ, Flenniken AM, Williams BR, Groppi VE, Buhl AE. Localization of TIMP in cycling mouse hair. Development. 1991 Apr;111(4):877-9 Kim SK, Park HJ, Chung JH, et al. Association between interleukin 18 polymorphisms and alopecia areata in Koreans. J Interferon Cytokine Res. 2014;34:349-353. Kim SK, Park HJ, Chung JH, et al. Association between interleukin 18 polymorphisms and alopecia areata in Koreans. J Interferon Cytokine Res. 2014;34:349-353. Koizumi H,Sato-Matsumura KC,Nakamura H,Shida K,Kikkawa S,Matsumoto M,Toyoshima K,Seya T.Arch Dermatol Res.2001 Jul;293(7):325-33.PubMed PMID:11550805. 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Claims

**Claim 1** A method for producing a therapeutic composition for treating, inhibiting, reducing, preventing, or restoring hair disorders or skin disorders, or a therapeutic composition for treating erectile dysfunction, comprising: identifying the hair, skin color, skin type, race, and ethnicity of an end user; obtaining mesenchymal skin cells (MSCs) from a target donor who has the same hair type, color, and / or ethnicity as the end user or is not sensitive to erectile dysfunction, provided that the donor has never experienced hair disorders, skin disorders, or erectile dysfunction and may be specific to gender, race, and ethnicity, and has a single nucleotide polymorphism (SNP) profile; preparing a powder preparation of growth factors for MSCs, keratinocytes, and melanocytes from the obtained MSCs, wherein the MSC preparation comprises at least one member selected from the group consisting of cells cultured under normal oxygen culture conditions or cell conditioned medium, and cells cultured under harsh wound healing conditions; adding the powder preparation to a base; and wherein the therapeutic composition comprises a member selected from the group consisting of topical compositions and injectable compositions. **Claim 2** The method according to claim 1, wherein the hair disorder or skin disorder includes gray hair, alopecia, or skin discoloration. **Claim 3** The method according to claim 2, wherein the skin discoloration is caused by vitiligo, melasma, nevus flammeus, or rosacea. **Claim 4** The method according to claim 3, wherein the therapeutic composition corrects skin discoloration. **Claim 5** The method according to claim 2, wherein the alopecia includes male pattern baldness, androgenetic alopecia, alopecia areata, cicatricial alopecia, telogen effluvium, or female pattern baldness. **Claim 6** The method according to claim 5, wherein the therapeutic composition stimulates the activation of hair follicles to promote hair growth. **Claim 7** The method according to claim 2, wherein the therapeutic composition stimulates the production of hair pigmentation. **Claim 8** The method according to claim 1, wherein said SNP comprises one or more SNPs present in one or more genes encoding interleukin 18 (IL-18), platelet-derived growth factor (PDGF) receptor (PDGFR) B (PDGFR-B), tissue metalloprotease inhibitor (TIMP) 1 (TIMP-1), TIMP-2, IL-23, activin A, intracellular adhesion molecule (ICAM-2), plasminogen activator inhibitor 1 (PAI-1), osteopontin (OPN), insulin, insulin growth factor binding protein 4 (IGF-BP4), tumor necrosis factor (TNF) receptor (TNF-R), neuregulin-1 β1 (NRG1-β1), urokinase-type plasminogen activator receptor (uPAR), ectodysplasin A2 receptor (XEDAR), follistatin, and / or activated leukocyte cell adhesion molecule (ALCAM).

9. The method according to claim 1, wherein said SNP comprises the SNP rs1946518 (-607C>A) and / or rs187238 (-137G>C) of said IL-18.

10. A therapeutic composition for treating, inhibiting, alleviating, preventing, or restoring hair damage or skin damage, comprising a composition base, a powder preparation of growth factors of mesenchymal stem cells (MSC), keratinocytes, and / or melanocytes derived from a donor having the same hair type, hair color, hair type, and / or hair color as the end user, but the donor never has hair damage and / or skin damage and may be specific to gender, race, and ethnicity, and having a single nucleotide polymorphism (SNP) profile, wherein said MSC preparation comprises at least one member selected from the group consisting of cells cultured under normal hypoxic culture conditions or cell conditioned medium and cells cultured under harsh wound healing conditions.

11. The therapeutic composition according to claim 10, wherein the SNP comprises one or more SNPs present in one or more genes encoding interleukin 18 (IL-18), platelet-derived growth factor (PDGF) receptor (PDGFR) B (PDGFR-B), tissue metalloprotease inhibitor (TIMP) 1 (TIMP-1), TIMP-2, IL-23, activin A, intracellular adhesion molecule (ICAM-2), plasminogen activator inhibitor 1 (PAI-1), osteopontin (OPN), insulin, insulin growth factor binding protein 4 (IGF-BP4), tumor necrosis factor (TNF) receptor (TNF-R), neuregulin-1 B1 (NRG1-B1), urokinase-type plasminogen activator receptor (uPAR), ectodysplasin A2 receptor (XEDAR), follistatin, and / or activated leukocyte cell adhesion molecule (ALCAM).

12. The therapeutic composition according to claim 10, wherein the SNP comprises the SNP rs1946518 (-607C>A) and / or rs187238 (-137G>C) of the IL-18.

13. The therapeutic composition according to claim 10, wherein the hair disorder comprises alopecia or gray hair.

14. The therapeutic composition according to claim 10, wherein the alopecia comprises male pattern alopecia, androgenetic alopecia, alopecia areata, cicatricial alopecia, telogen effluvium, or female pattern alopecia.

15. A method for treating a hair disorder, comprising administering any one of the therapeutic compositions according to any one of claims 10 to 14 to a subject.

16. The therapeutic composition according to claim 10, wherein the skin disorder comprises vitiligo, chloasma, pityriasis rosea, or skin discoloration caused by acne rosacea.

17. A method for treating a skin disorder, comprising administering any one of the therapeutic compositions according to any one of claims 10 to 12 or 16 to a subject.

18. A method for preparing a skin therapeutic composition for a specific skin color, race, type, or ethnicity, comprising: identifying the skin color, type, race, and ethnicity of an end user; obtaining mesenchymal skin cells (MSCs) from a target donor having the same skin color, type, race, and ethnicity as the end user based on the appropriate single nucleotide polymorphism panel of the donor; Preparing a powder preparation of growth factors for MSC, keratinocytes, and melanocytes from the aforementioned obtained MSC, wherein the MSC preparation comprises at least one member selected from the group consisting of cells cultured under normal hypoxic culture conditions or cell conditioned medium, and cells cultured under harsh wound healing conditions; Adding the powder preparation to a cosmetic base; A method comprising:

19. A skin treatment composition for a specific skin color, race, type, or ethnicity, comprising: A composition base; A powder preparation of growth factors for mesenchymal stem cells (MSC), keratinocytes, and / or melanocytes derived from a donor having the same skin color, type, race, and ethnicity as the end user, wherein the MSC preparation comprises at least one member selected from the group consisting of cells cultured under normal hypoxic culture conditions or cell conditioned medium, and cells cultured under harsh wound healing conditions; A skin treatment composition comprising:

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