Placenta components

Abstract

Uses of placental extracts and compositions comprising placental extracts to treat or prevent inflammation in diseases or conditions associated with, for example, TNFα, IL-1 beta, IL-6 and other pro-inflammatory cytokines, such as asthma, infection, acute respiratory distress syndrome (ARDS), or COVID-19, and their use as medicaments, for example, to treat or prevent inflammatory conditions, are described.

Description

[Technical field]

[0001] The present invention relates to the use of placental extracts and compositions comprising placental extracts to treat or prevent inflammation in diseases or conditions associated with TNFα, IL-1 beta, IL-6 and other pro-inflammatory cytokines, such as asthma, infection, acute respiratory distress syndrome (ARDS), or COVID-19. The present invention also relates to methods of treatment by administration of placental extracts and compositions comprising placental extracts, and the use of placental extracts in the manufacture of medicaments or compositions for treating such diseases or conditions. [Background technology]

[0002] Although the production of proinflammatory cytokines by cells of the innate immune system plays an important role in mediating the initial host defense against invading pathogens (O'Neill, LA et al., Immunol. Today, (2000), 21(5): 206-9), the inability of the host to regulate the nature or duration of the inflammatory response often mediates detrimental effects on the host, such as those observed in chronic inflammatory diseases. Furthermore, in the early stages of sepsis, the host inflammatory response is in an extremely active state, dominated by an increase in the production of proinflammatory cytokines, which is thought to mediate host tissue damage and lethal shock (Cohen, J., Nature, (2002), 420(6917): 885-91). In this regard, the ability to suppress pro-inflammatory cytokines and / or enhance anti-inflammatory cytokines has been shown to significantly reduce the toxic effects of endotoxin (Berg, DJ et al., J. Clin. Invest., (1995), 96(5):2339-47; and Howard, M. et al., J. Exp. Med., (1993), 177(4):1205-8).

[0003] COVID-19, caused by SARS-CoV-2, is also characterized by immune dysfunction rather than viral load, leading to abnormal production of pro-inflammatory cytokines (Ma WT,et al. The protective and pathogenic roles of IL-17 in viral infections:friend or foe? Open Biol. 2019;9(7):190109). In particular, COVID-19 can induce a cytokine storm in lung tissues through an overactive immune system and uncontrolled release of cytokines (Ye Q,Wang B,Mao J.Cytokine storm in COVID-19 and treatment. J Infect. 2020;80:607-13). Proinflammatory cytokines, such as interleukin-6 (IL-6), interleukin-1β (IL-1β), and tumor necrosis factor-alpha (TNF-α), play a crucial role in lung injury in COVID patients who present with acute respiratory distress syndrome (ARDS) due to dysfunction of the respiratory epithelium (Montazersaheb, S., Hosseiniyan Khatibi, SM, Hejazi, MS et al. COVID-19 infection: an overview on cytokine storm and related interventions. Virol J 19, 92 (2022)).

[0004] Other conditions associated with abnormal or unwanted inflammation, such as skin conditions, can also be difficult to manage. Treatment consists primarily of medications such as steroids, antihistamines and antibiotics. Steroids have anti-inflammatory and immunosuppressant effects and have shown good results, but are harmful to the intestinal tract, kidneys, liver, bones and brain, and the condition often recurs if treatment is discontinued.

[0005] The availability of improved or alternative formulations suitable for cosmetic, pharmaceutical, nutraceutical, supplement and beverage products is important for subjects suffering from conditions associated with abnormal or unwanted inflammation associated with TNFα, IL-1 beta, IL-6 and other pro-inflammatory cytokines.

[0006] It is an object of the present invention to provide improved or alternative compositions and / or at least to provide the public with a useful choice.

[0007] Where references are made in this specification to external sources of information, including patents and other documents, this is generally for the purpose of providing a context for discussing features of the invention. Unless specifically stated, the reference to such sources should not be construed as an admission in any jurisdiction that such sources are prior art or form part of the common general knowledge in the art. Summary of the Invention

[0008] Accordingly, in one aspect, the present invention relates to a method for treating, preventing, or ameliorating a disease, disorder, or condition associated with inflammation or immune modulation, comprising the step of administering to a subject in need thereof an effective amount of a placental extract, or a composition comprising an effective amount of a placental extract.

[0009] In another aspect, the present invention relates to a composition for use in the treatment, prevention, or amelioration of a disease, disorder, or condition associated with inflammation or immune modulation, the composition comprising a placental extract.

[0010] In another aspect, the present invention relates to the use of a placental extract in the manufacture of a medicament or composition for treating, preventing or ameliorating a disease, disorder or condition associated with inflammation or immune modulation.

[0011] Preferably, the composition or medicament is a food, beverage, food additive, beverage additive, food component, beverage component, dietary supplement, nutritional product, medical food, nutritional functional food, medicine or pharmaceutical product.

[0012] In one embodiment, the placental extract is administered at a concentration sufficient to inhibit one or more pro-inflammatory cytokines, including the cytokines IL-1α, β, IL-2, IL-3, IL-6, IL-7, IL-9, IL-12, IL-17, IL-18, TNF-α, LT, LIF, oncostatin, or IFNc1α, β, γ.

[0013] In one embodiment, the placental extract is administered at a concentration sufficient to stimulate expression of one or more anti-inflammatory cytokines, including IL-4, IL-10, IL-11, W-13, or TGFβ.

[0014] The following embodiments may relate to any of the above aspects.

[0015] In one embodiment, the placental extract is derived from deer, sheep, goat, horse, donkey, rabbit, or cow placenta. Preferably, the placental extract is derived from deer placenta. In one embodiment, the placental extract is CerviCenta®.

[0016] Preferred methods of extraction from placental material include solvent extraction, supercritical solvent extraction, including supercritical CO2 extraction, distillation, countercurrent extraction, decoction, percolation, maturation, molecular distillation, microwave extraction, ultrasonic extraction, and chromatographic separation.

[0017] The extraction method may include contacting the placental material with a solvent, separating the placental material from the solvent, and at least partially removing the solvent to obtain an extract.

[0018] In one embodiment, the placental material is freeze-dried and powdered prior to contact with the solvent. In one embodiment of the invention, the solvent is water. In another embodiment of the invention, any suitable organic solvent may be used. The solvent may be ethanol or a mixture of any such solvents. In one embodiment, the extract is prepared by solvent extraction, e.g., ethanol extraction, followed by distillation or supercritical extraction.

[0019] Preferably, the extract is prepared using aqueous extraction.

[0020] In one embodiment, the placental extract is or comprises dried or powdered placental material. In one embodiment, the extraction method comprises micronizing the placental material by wet grinding, followed by powdering or granulation, such as by freeze-drying, vacuum drying or spray drying, or fluidized bed drying. In an alternative embodiment, the placental material is first dried, for example by freeze-drying or vacuum drying, and then optionally ground into a powder.

[0021] In a preferred embodiment, the composition or medicament is provided in a delivery formulation selected from the group consisting of a tablet, a capsule, a liquid, an oil, a suspension, an emulsion, a paste, a jelly, a pudding, a solution, and a powder.

[0022] In one embodiment, the composition or medicament comprises, consists essentially of, or consists of a placental extract.

[0023] In one embodiment, the placental extract comprises, consists essentially of, or consists of placental material prepared by solvent extraction, dried or powdered placental material, and / or combinations thereof.

[0024] In one embodiment, the composition or medicament comprises at least about 0.1, 0.2, 0.5, 1, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 99, 99.5, 99.8 or 99.9% by weight of placental extract, with useful ranges ranging between any of these aforesaid values ​​(e.g., about 0.1 to about 50%, about 0.2 to about 50%, about 0.5 to about 50%, about 1 to about 50%, about 5 to about 50%, about 10 to about 50%, about 15 to about 50%, about 20 to about 25%, about 25 to about 25% by weight of placental extract. %, about 20 to about 50%, about 25 to about 50%, about 30 to about 50%, about 35 to about 50%, about 40 to about 50%, about 45 to about 50%, about 0.1 to about 60%, about 0.2 to about 60%, about 0.5 to about 60%, about 1 to about 60%, about 5 to about 60%, about 10 to about 60%, about 15 to about 60%, about 20 to about 60%, about 25 to about 60%, about 30 to about 60%, about 35 to about 60%, about 40 to about 60%, about 45 to about 60%, about 0.1 to about 70%, about 0.2 to about 70%, about 0.5 to about 70%, about 1 to about 70%, about 5 to about 70% , about 10 to about 70%, about 15 to about 70%, about 20 to about 70%, about 25 to about 70%, about 30 to about 70%, about 35 to about 70%, about 40 to about 70%, about 45 to about 70%, about 0.1 to about 80%, about 0.2 to about 80%, about 0.5 to about 80%, about 1 to about 80%, about 5 to about 80%, about 10 to about 80%, about 15 to about 80%, about 20 to about 80%, about 25 to about 80%, about 30 to about 80%, about 35 to about 80%, about 40 to about 80%, about 45 to about 80%, about 0.1 to about 90%, about 0.2 to about 90%, about 0.5 to about 90 %, about 1 to about 90%, about 5 to about 90%, about 10 to about 90%, about 15 to about 90%, about 20 to about 90%, about 25 to about 90%, about 30 to about 90%, about 35 to about 90%, about 40 to about 90%, about 45 to about 90%, about 0.1 to about 99%, about 0.2 to about 99%, about 0.5 to about 99%, about 1 to about 99%, about 5 to about 99%, about 10 to about 99%, about 15 to about 99%, about 20 to about 99%, about 25 to about 99%, about 30 to about 99%, about 35 to about 99%, about 40 to about 99%, and about 45 to about 99%).

[0025] In one embodiment, the composition or medicament comprises at least about 0.001, 0.01, 0.05, 0.1, 0.15, 0.2, 0.3, 0.4, 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 grams or more of placental extract, and useful ranges may be selected between any of these values ​​aforesaid (e.g., about 0.01 to about 1 gram, about 0.01 to about 10 grams, about 0.01 to about 19 grams, about 0.1 to about 1 gram, about 0.1 to about 10 grams, about 0.1 to about 19 grams, about 1 to about 5 grams, about 1 to about 10 grams, about 1 to about 19 grams, about 5 to about 10 grams, and about 5 to about 19 grams).

[0026] Preferably, the composition or medicament comprises at least about 1, 10, 50, 100, 125, 150, 200, 250, 300, 350, 400, 450, 500, 600, 700, 800, 900, or 1000 mg or more of placental extract, and a useful range may be selected between any of these aforementioned values ​​(e.g., about 10 to about 1000 mg, about 50 to 1000 mg, about 75 to about 800 mg, about 75 to about 500 mg, about 75 to about 350 mg, about 75 to about 300 mg, about 75 to about 250 mg, about 100 to about 350 mg, about 100 to about 300 mg, about 100 to about 250 mg, about 100 to about 200 mg, about 100 to about 150 mg, more preferably about 100 to about 125 mg).

[0027] In one embodiment, the composition, medicament, or method comprises administering placental extract in an amount of about 125 mg suitable for twice daily administration, or in an amount of 250 mg suitable for once daily administration.

[0028] In one embodiment, the composition, medicament, or method comprises administering a placental extract in an amount of about 100 mg or 125 mg suitable for twice daily administration, or in an amount of 200 mg or 250 mg suitable for once daily administration.

[0029] In one embodiment, the composition, medicament, or method comprises administering about 40% to 90% by weight of placental extract, for example, about 50% to 80% by weight, or about 60% to 75% by weight of placental extract.

[0030] In one embodiment, the composition or medicament further comprises about 0.1, 0.5, 1, 5, 10, 15, 20, 25, 30, 35, 40, 45, or 50% by weight of another anti-inflammatory agent, and a useful range may be selected between any of these aforementioned values ​​(e.g., about 0.1 to about 50%, about 0.2 to about 50%, about 0.5 to about 50%, about 1 to about 50%, about 5 to about 50%, about 10 to about 50%, about 15 to about 50%, about 20 to about 50%, about 25 to about 50%, about 30 to about 50%, about 35 to about 50%, about 40 to about 50%, and about 45 to about 50%).

[0031] In one embodiment, the composition or medicament further comprises a carrier, eg, a pharma- ceutically acceptable carrier.

[0032] In one embodiment, the composition or medicament is in the form of a tablet, caplet, pill, hard or soft capsule, or lozenge. In one embodiment, the composition or medicament is in the form of a cachet, dispensable powder, granule, suspension, elixir, liquid, or other form that can be added to food or beverages, including, for example, water or fruit juice. In one embodiment, the composition or medicament further comprises one or more components (e.g., antioxidants) that prevent or reduce degradation of the composition during storage or after administration.

[0033] In one embodiment, the composition or medicament is or is formulated as a food, beverage, food additive, beverage additive, dietary supplement, nutritional product, medical food, nutritional functional food, medicine or pharmaceutical product. Preferably, the composition or medicament is formulated as a powder, liquid, food bar, spread, sauce, paste, jelly, pudding, soup base, ointment, tablet or capsule.

[0034] In one embodiment, the composition or medicament is formulated for oral, nasal, or parenteral (including topical, subcutaneous, intramuscular, and intravenous) administration. In one embodiment, the composition or medicament is formulated for ingestion, inhalation, or topical application. When the composition or medicament is formulated for inhalation, it is preferably formulated as an inhalable powder, solution, or aerosol. When the composition or medicament is formulated for topical application, it is preferably formulated as an ointment, cream, or lotion.

[0035] As will be appreciated, the dose of the composition or medicament administered, the duration of administration, and the general administration regimen may vary between subjects depending on such variables as the severity of the subject's symptoms, the type of disorder to be treated, the mode of administration selected, and the age, sex, and / or general health of the subject. However, as a general example, the inventors contemplate administering about 1 mg to about 1000 mg per kg of body weight per day, preferably about 50 to about 500 mg per kg of body weight per day, of the composition or medicament of the present invention. In one embodiment, the inventors contemplate administering about 0.05 mg to about 250 mg per kg of body weight of the pharmaceutical composition or medicament according to the present invention. It should be understood that administration may include a single daily dose, or multiple individually divided doses may be administered as needed.

[0036] In this specification, where reference is made to patent specifications, other external documents, or other sources of information, this is generally for the purpose of providing a context for discussing the features of the present invention. Unless specifically stated otherwise, the reference to such external documents should not be construed as an admission in any jurisdiction that such documents or such sources are prior art or form part of the common general knowledge in the art.

[0037] Reference to various numbers disclosed herein (e.g., 1-10) also incorporates reference to all rational numbers within that range (e.g., 1, 1.1, 2, 3, 3.9, 4, 5, 6, 6.5, 7, 8, 9, and 10), as well as any range of rational numbers within that range (e.g., 2-8, 1.5-5.5, and 3.1-4.7), and thus all subranges of all ranges explicitly disclosed herein are intended to be hereby expressly disclosed. These are merely examples of what is specifically intended, and all possible combinations of numerical values ​​between the highest and lowest values ​​recited should be considered to be expressly defined in this application in a similar manner.

[0038] The present invention may also be broadly described as consisting of the parts, elements and features referred to or indicated in the specification of this application, either individually or collectively, in any or all combinations of two or more of said parts, elements or features, and where specific integers having known equivalents in the art to which the invention pertains are referred to herein, such known equivalents are deemed to be incorporated herein as if individually set forth. The present invention will now be described, by way of example only, with reference to the drawings in which: [Brief description of the drawings]

[0039] [Figure 1] Figure 1 provides a determination of protein integrity of placental extracts: As indicated, protein extracts were separated on a 4-20% BioRad gradient gel and then stained with Coomassie Brilliant Blue. The following order can be observed: Lanes 1-10 on the left or right: BioRad Precision Plus Protein Standard; from top to bottom: Mw (kDa): 250 / 150 / 100 / 75 / 50 / 37 / 25 / 20 / 15; Lane 2: culture medium; Lane 3: 0.9% NaCl; Lane 4: PBS / 1% TritonX100; Lane 5: PBS / 1% SDS; Lane 6: overflow from lane 7; Lane 7: distilled water; Lane 8 / 9: PBS. [Diagram 2]Figure 2 provides the evaluation of IL-1 beta. PBMCs were treated with DPE and stimulants as described. Supernatants were isolated and transferred to the MSDU-Plex multiplexing platform at different dilutions (1 / 5, 1 / 20 and 1 / 50) to define the optimal range of reactivity. The highest reactivity was achieved at a 1 / 5 dilution of the extract tested. These data are illustrated here. Zymosan 2 (1 μg / mL), Endotoxin 1 (0.05 EU / mL). HSKA (108 cells). DPE 1 / 5 (12.33 mg / mL), DPE 1 / 20 (3.08 mg / mL and DPE 1 / 50 (1.23 mg / mL). X-axis: reactivity (EC units), Y-axis: stimulation [Diagram 3] Figure 3 provides the evaluation of IL-6. PBMCs were treated with DPE and stimulants as described. Supernatants were isolated and transferred to the MSDU-Plex multiplexing platform at different dilutions (1 / 5, 1 / 20 and 1 / 50) to define the optimal range of reactivity. The highest reactivity was achieved at a 1 / 5 dilution of the extract tested. These data are illustrated here. Zymosan 3 (0.2 μg / mL), Endotoxin 1 (0.05 EU / mL). HSKA (108 cells). DPE1 / 5 (12.33 mg / mL), DPE1 / 20 (3.08 mg / mL and DPE1 / 50 (1.23 mg / mL). X-axis: reactivity (EC units), Y-axis: stimulation. [Figure 4] Figure 4 provides DPE interference in the induction of IL-1 beta. Human PBMCs were pretreated with DPE as described. Then established stimuli were given at the following concentrations: zymosan (1 μg / mL), HKSA (1×108), endotoxin (0.05 EU / mL). Supernatants were collected and analyzed on the Mesoscale U-PLEX platform. Data shown here are similar to a 1 / 10 dilution on the Mesoscale platform. [Diagram 5]Figure 5 provides the results of a dose study of DPE in BEAS-2B cells 24 hours after addition. PC = positive control (ZDBC polyurethane film (SPU-ZDBC); Lot number: B-172K; Hatano Research Institute, Japan). Values ​​are given as mean ± SD, n = 6. [Figure 6] Figure 6 provides IL-10 release in BEAS-2B cells 24 hours after addition of three concentrations of DPE followed by stimulation with activated lymphocytes (0.5x106 cells / mL, 24 hours). Values ​​are given as mean ± SD, n=3. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0040] The present invention is based on the discovery that placental extracts have useful properties, including anti-inflammatory activity.

[0041] [Definition] The term "comprising" as used in this specification means "consisting at least in part of." When interpreting descriptions in this specification that include that term, all features preceded by that term in each description or claim must be present, although other features may also be present. The related terms "comprises" and "comprised" should be interpreted in the same manner.

[0042] As used herein, the term "and / or" means "and" or "or," or both.

[0043] An "effective amount" is the amount required to produce a therapeutic effect. The interrelationship of dosages for animals and humans (based on milligrams per square meter of body surface) is described by Freireich et al. (1966). Body surface area may be approximately determined from the subject's height and weight. See, for example, Scientific Tables, Geigy Pharmaceuticals, Ardley, New York, 1970, 537. Effective doses also vary as recognized by those skilled in the art depending on the route of administration, carrier usage, and the like.

[0044] As used herein, the term "extract" refers to a preparation from a raw material that is in a different form than the raw material from which it is derived. An extract can be as simple as mechanically comminuted cellular material, in which case the preparation can be dehydrated to remove water, or it can be a preparation produced by contacting the raw material with one or more solvents. The term "extract" also encompasses preparations that have undergone one or more separation and / or purification steps to enrich the content of active agent(s), as well as preparations that include a partially or substantially purified fraction from placental material.

[0045] The term "pharmaceutically acceptable carrier" is intended to refer to a carrier, including but not limited to an excipient, diluent or auxiliary, that can be administered to a subject as a component of a composition of the present invention, without diminishing the activity of the composition and that is non-toxic when administered in a dose sufficient to deliver an effective amount of one or more active agents. The formulation may be administered orally, nasally, or parenterally (including topically, intramuscularly, intraperitoneally, subcutaneously, and intravenously).

[0046] The term "cosmetic composition" as used herein refers to a composition containing a compound and having any type of formulation.Non-limiting examples of cosmetic formulations prepared using the composition include creams, such as skin creams, nourishing creams, eye creams, massage creams, and cleansing creams; packs; lotions, such as nourishing lotions; essences; serums, poultices, ointments; tonics, such as softening and nourishing tonics; powders; foundations, and makeup bases.To achieve the technical goal of the present disclosure, the cosmetic composition may be prepared into any formulation selected from the above-listed formulations that will be commercialized, and the present disclosure is not limited to the above examples.Furthermore, the cosmetic composition according to the present disclosure may be formulated using a general cosmetic preparation method.

[0047] The term "health functional food", as used herein, refers to food prepared and processed into the form of tablets, capsules, pastes, jellies, puddings, soft gels, powders, granules, liquids, pills, etc., by using raw materials or ingredients that have useful functionality in the human body. The term "functionality", as used herein, refers to controlling the structural nutrients of the functions of the human body, or providing useful effects for hygiene purposes, such as psychological effects. The health functional food of the present disclosure may be prepared using methods commonly used in the art, or may be prepared by adding raw materials and ingredients commonly added in the art. Furthermore, the health functional food formulation is not particularly limited as long as it is recognized as a health functional food. Unlike generic drugs, the health functional food composition of the present disclosure uses food as a raw material, and therefore has no side effects that may occur during their long-term administration, is highly portable, and may be administered as an adjuvant for enhancing skin moisture, exfoliating the skin, improving skin elasticity, promoting skin healing, preventing erythema, improving skin wrinkles, and / or reducing skin photoaging.

[0048] The term "steroid sparing" is intended to mean that the dose of a steroid medication administered to a subject can be reduced to a level below the level administered before the subject began taking the composition or using the method of the invention. Preferably, the daily or weekly or monthly dose of steroid can be reduced by at least 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95 or 99%.

[0049] A "subject" according to the present invention is an animal, preferably a mammal, more preferably a companion animal to a mammal or a human. Preferred companion animals include cats, dogs and horses.

[0050] The term "synergy," as used herein, means that the effect of the compositions useful herein is superior to the use of an individual agent alone, e.g., as measured by the extent of the effect in vitro or in vivo or both. For example, the effect of a combination of a placental extract with another agent, e.g., another anti-inflammatory extract, is synergistic if the effect is superior to the effect achievable with the placental extract alone or with the other agent or extract alone.

[0051] Furthermore, the effect of the combination is synergistic if a beneficial effect is obtained in a group of subjects that are unresponsive (or less responsive) to the placenta extract or other drug or extract alone. Furthermore, the effect of the combination is synergistic if one of the components is used at its conventional dose and the other component is used at a reduced dose, and the effect, e.g., measured in a range of effects in vitro or in vivo or both, is equivalent to or better than the effect achievable with conventional amounts of any one of the components of the combination treatment. Related terms, such as "synergistic," should be interpreted similarly.

[0052] The term "treat" and its derivatives should be interpreted in its broadest possible context. The term should not be interpreted to suggest that a subject is treated until complete recovery. Thus, "treat" broadly includes amelioration and / or prevention of the onset of symptoms or severity of a particular condition.

[0053] [Treatment or prevention methods] The data in the examples herein demonstrate that placental extracts and compositions comprising placental extracts as described herein are useful in methods for treating or preventing a variety of conditions, including diseases, disorders, or conditions associated with inflammation or immune modulation.

[0054] In one embodiment, the extracts and compositions herein provide immune modulating effects useful, for example, in the treatment or prevention of conditions associated with inflammation.

[0055] In a preferred embodiment, the placental extract is administered in an amount sufficient to treat inflammation-related diseases by inhibiting expression of pro-inflammatory cytokines and / or stimulating anti-inflammatory cytokines.

[0056] It should be understood that the use may include, but is not limited to, treating inflammation-associated diseases by modulating cytokines involved in pathological progression by preventing inflammation associated with the disease, thus preventing the onset of inflammation-associated diseases.

[0057] In one embodiment, the inflammation-related disease is selected from the group consisting of type I diabetes; Sjogren's syndrome; uveitis; celiac disease; allergic conjunctivitis; and non-specific colitis.

[0058] Other inflammation-related diseases disclosed but not claimed herein include: arthritis, rheumatoid arthritis, inflammatory bowel disease; psoriasis; multiple sclerosis; neurodegenerative disorders; congestive heart failure; stroke; aortic stenosis; renal failure; lupus; pancreatitis; allergies; fibrosis; anemia; atherosclerosis; metabolic diseases; bone diseases; cardiovascular diseases, chemotherapy / radiation-related complications; type II diabetes; liver diseases; gastrointestinal disorders; ophthalmological diseases; diabetic retinopathy; pulmonary disorders, kidney diseases; dermatitis; HIV-associated cachexia; cerebral malaria; ankylosing spondylitis; leprosy; anemia; and fibromyalgia.

[0059] Disclosed herein are neurodegenerative disorders including: Alzheimer's and Parkinson's disease, geroscience, osteoporosis, macular degeneration, healthspan extension; the inflammatory bowel disease is selected from the group consisting of Crohn's disease or ulcerative colitis; the gastrointestinal complication is diarrhea; the liver disease is selected from the group consisting of autoimmune hepatitis, hepatitis C, primary biliary cirrhosis, primary sclerosing cholangitis, or fulminant hepatic failure; the bone disease is osteoporosis; the pulmonary disorder is selected from the group consisting of allergic rhinitis, asthma, chronic obstructive pulmonary disease, chronic granulomatous inflammation, cystic fibrosis, and sarcoidosis; the cardiovascular disease is selected from the group consisting of atherosclerotic heart disease, congestive heart failure and restenosis, thrombosis (atrial fibrillation, AF), hemodynamic stress, metabolic syndrome (MetS), heart rate variability (HRV), vagal anti-inflammatory activity (VAI), and the like. anti-inflammatory), arterial sclerosis, heart failure, congenital heart disease, vascular disease; renal disease is selected from the group consisting of glomerulonephritis and vasculitis.

[0060] Pro-inflammatory cytokines have also been associated with a number of other diseases, disorders or conditions, all of which are amenable to treatment, amelioration or prevention by the extracts, compositions and methods of the present invention, including: Autoimmune - autoimmune and inflammatory diseases (hypertension, fatigue, autism spectrum disorders, bipolar disorder, cancer, kidney transplants, Kawasaki disease); Brain Health - cognitive dysfunction, vascular and neurodegenerative disorders, Alzheimer's disease, brain morphology, tumor necrosis; Lung / Respiratory Health - Macrophages, chronic respiratory disease, pulmonary fibrosis, autophagic, hyperactive and dysregulated inflammation, interferon (type I IFN) response, cystic fibrosis (CF) lung disease, acute respiratory distress syndrome, chronic bronchitis, chronic obstructive pulmonary disease (COPD), respiratory syncytial virus infection (RSV), mucus obstruction and neutrophilic inflammation; Heart health - coronary artery disease (HIV+ individuals), cardiovascular disease (CVD), atherosclerosis, thrombosis (atrial fibrillation, AF), hemodynamic stress, metabolic syndrome (MetS), heart rate variability (HRV) - anti-inflammatory action of the vagus nerve, arterial stiffening, heart failure, congenital heart disease, vascular disease; Skin health - characterizing erythematotelangiectatic rosacea (ETR), psoriasis-like skin inflammation, autoimmune skin diseases (psoriasis), skin diseases (skin cancer), skin aging / oxidative stress, natural skin aging, photoaging, skin barrier dysfunction, extracellular matrix dysfunction, dendritic cells (DC), skin pigmentation, wrinkles; Depression-Central nervous system disorders, chronic stress, depression, dementia, major depressive disorder (MDD); Obesity-high fat diet (HFD), type 2 diabetes mellitus (T2DM), low-grade inflammation; Organ-asthmatic chronic obstructive pulmonary disease (ACOS), kidney disease, inflammatory bowel disease, organ disease (tissue damage to the heart, pancreas, liver, kidneys, lungs, brain, intestinal tract, reproductive system); Cancer – liver cancer, breast cancer, stomach cancer, intestinal cancer, lung cancer, arterial disease; Others - stroke, myocardial infarction, diabetes, psychomotor degeneration.

[0061] The compound is in an amount to inhibit the expression of a pro-inflammatory cytokine and / or stimulate the expression of an anti-inflammatory cytokine. In one embodiment, the compound is preferably in an amount to inhibit at least 30% expression of one or more of the pro-inflammatory cytokines selected from the group consisting of IL-1α, β, IL-2, IL-3, IL-6, IL-7, IL-9, IL-12, IL-17, IL-18, TNF-α, LT, LIF, oncostatin, and IFNc1α, β, γ; more preferably, at least 40% expression of the cytokine is inhibited, and most preferably, 50% or more expression is inhibited. In another embodiment, the compound is preferably in an amount to stimulate the expression of an anti-inflammatory cytokine. In this embodiment, the compound is preferably in an amount to increase the anti-inflammatory cytokine selected from the group consisting of the cytokines IL-4, IL-10, IL-11, W-13, or TGFβ by at least 25%, more preferably, by at least 50%, and most preferably, by at least 75%.

[0062] In one embodiment, the condition is arthritis, myositis, tendinitis, ligamentitis, joint injury, joint sprain or strain, muscle sprain, muscle strain, cartilage injury, osteoarthritis, rheumatoid arthritis, atopic condition, allergy, arteriosclerosis, atherosclerosis, heart disease, hypertension, blood clot, hypotension, vasoconstriction, cancer, or depression. In one embodiment, the condition is arthritis. In one embodiment, the condition is myositis, tendinitis, ligamentitis, joint injury, joint sprain or strain, muscle sprain or strain, or cartilage injury. In another embodiment, the condition is osteoarthritis or rheumatoid arthritis. In another embodiment, the condition is an atopic condition. In another embodiment, the condition is an allergy. In another embodiment, the condition is arteriosclerosis, atherosclerosis, heart disease, hypertension, blood clot, hypotension, vasoconstriction, cancer, or depression. In various embodiments, the treatment is associated with a steroid-sparing effect.

[0063] In another embodiment, the combinations described herein are useful in reducing inflammation caused by skin / tissue damage and are useful in atopic dermatitis, psoriasis, actinic keratosis, rosacea, skin tissue healing and other chronic skin disorders.

[0064] In another embodiment, the combination described herein is useful for lung disease, disorder or condition.In certain embodiments, the disease, disorder or condition is (a) associated with or caused by immune response; (b) interstitial lung disease; (c) obstructive pulmonary disease; (d) acute lung injury; or (e) lung injury caused by neoplastic or paraneoplastic disease, pneumonia, or cystic fibrosis.

[0065] In certain embodiments, the disease, disorder, or condition associated with or caused by an immune response is an autoimmune disease or graft-versus-host disease. In yet another embodiment, the autoimmune disease is rheumatoid arthritis, scleroderma, inflammatory bowel disease, or systemic lupus erythematosus. In another embodiment, the interstitial lung disease is interstitial pulmonary fibrosis. In another embodiment, the obstructive pulmonary disease is asthma, bronchitis, acute respiratory distress syndrome, or chronic obstructive pulmonary disease.

[0066] In a particular embodiment, the lung disease, disorder or condition is acute lung injury.In a more particular embodiment, the acute lung injury is one or more of physical trauma, chemical injury, such as chemical burn, smoke inhalation, or exposure to toxic substances.In another particular embodiment, the lung disease, disorder or condition is injury caused by neoplastic or paraneoplastic disease.

[0067] In certain embodiments, the disease, disorder or condition is one or more of pulmonary fibrotic disease, acute respiratory distress syndrome (ARDS), COVID-19, chronic obstructive pulmonary disease (COPD), emphysema, asthma, lung viral or bacterial infection, pneumonia (including chemically induced pneumonia), or cystic fibrosis. In particular embodiments, the pulmonary fibrotic disease is interstitial lung disease (diffuse parenchymal lung disease). In more particular embodiments, the interstitial lung disease is silicosis, asbestosis, beryllium disease, systemic sclerosis, polymyositis, or dermatomyositis. In other more particular embodiments, the interstitial lung disease is caused by antibiotics, chemotherapy drugs, antiarrhythmic drugs, or infections.

[0068] In certain embodiments, the pulmonary disease, disorder or condition is associated with or caused by a harmful, toxic, inappropriate or unwanted immune response, such as inflammation, and the disease, disorder or condition affects or manifests in the lungs. In particular embodiments, the disease, disorder or condition is one or more of lupus, such as lupus erythematosus, scleroderma, or a rheumatic disease (e.g., rheumatoid arthritis).

[0069] In another particular embodiment, the disease, disorder or condition is a rheumatic pulmonary disease (RLD), e.g., a rheumatic pulmonary disease associated with rheumatoid arthritis. In another particular embodiment, administration is sufficient to cause a detectable improvement in one or more symptoms of RLD, or to detectably reduce or slow the progression of one or more symptoms of RLD, e.g., in the lungs of an individual. In a more particular embodiment, the symptom of RLD is a condition associated with RLD. In a more particular embodiment, the condition associated with RLD is an infection, e.g., a viral infection of the lungs, or pulmonary fibrosis (e.g., as a result of methotrexate therapy).

[0070] In another particular embodiment, the disease, disorder or condition is lupus erythematosus, e.g., systemic lupus erythematosus (SLE). In a more particular embodiment, said symptoms of lupus erythematosus are one or more of lung and / or pleural inflammation, pleurisy, pleuritis, pleural effusion, lupus pneumonitis, or chronic diffuse interstitial lung disease.

[0071] In another embodiment, the combination described herein is useful for the prevention or treatment of a viral disease and / or inhibition of viral activation, wherein the virus is selected from the group consisting of a herpes virus, such as a cytomegalovirus (CMV); an influenza virus, such as an H1N1, H3N2, H5N1 or H5N7 virus; a paramyxovirus, such as a measles virus; a respiratory syncytial virus; a coronavirus, such as SARS or SARS-CoV-2 (Covid-19); an HIV virus; a hepatitis virus; or a rotavirus.

[0072] The compositions, medicaments and therapeutic or prophylactic methods useful herein may employ compositions as described below.

[0073] [Cosmetic applications] Placental extracts and compositions comprising placental extracts as described herein are useful in methods for treatment, including prevention, reduction, amelioration, and / or elimination of the signs and consequences of dermatological aging of the skin, particularly wrinkles and fine lines, age spots, red skin, dark spots, and / or for improving the aesthetic appearance of the skin, stimulating collagen production, elasticity, and / or increasing skin tone and hydration. The combination provides a treatment for wrinkles, fine lines, and other signs of dermatological aging (i.e., intrinsic aging) or exposure of the skin to sunlight (i.e., extrinsic aging).

[0074] As used herein, the terms treating and treatment are understood to include and encompass preventing, reducing, ameliorating, improving, alleviating, and / or eliminating the dermatological effects of aging and sun exposure, particularly with respect to skin wrinkles, fine lines, folds, deep lines, deep creases, and the like. The present invention further encompasses the treatment of "marionette" lines on either side of the mouth, as well as forehead wrinkles and vertical frown lines. The present compositions and methods are also suitable for use in treating dermatological conditions of the skin, as defined above, in many areas of the body, including, but not limited to, the face, forehead, neck, arms, hands, legs, knees, feet, chest, back, groin, buttocks, and the like.

[0075] It is another aspect of the present invention to provide compositions, formulations and methods comprising newly determined materials useful in treating dermatological aging of the skin, particularly wrinkles, fine lines, folds, deep wrinkles and other signs of aged skin, age spots, red skin, dark spots and / or improving the aesthetic appearance of the skin, stimulating collagen production, elasticity and / or increasing skin tone and hydration.

[0076] Furthermore, it is understood that contraction or hypercontraction of certain muscles, particularly facial muscles, is associated with the appearance of wrinkles, fine lines, and the like, and therefore relaxation of such muscles and / or control or modulation of the contraction of such muscles by the newly determined action of the placental extracts of the present invention can serve an important function in the treatment, prevention, reduction, amelioration, or elimination of wrinkles, fine lines, folds, deep wrinkles, and the like.

[0077] In accordance with the present invention, the disclosed combinations include, but are not limited to, compositions including topically applied sunscreens, antioxidants, anti-inflammatories, cosmetics including makeup, anti-aging formulations such as topical fine line and / or wrinkle creams, skin penetrating antiperspirants, deodorants, etc. Also encompassed in accordance with the present invention are ingredients, components, or compounds formulated in such compositions in various product forms, such as transdermal, e.g., patches, etc., particularly for topical administration.

[0078] Another aspect of the present invention provides compositions comprising the disclosed combinations, preferably for topical administration, without inducing significant irritation.Furthermore, such compositions are preferably delivered by using targeted delivery systems, such as, but not limited to, liposomes, microspheres, transdermal patches, etc., so that the active agent can more easily reach and affect the muscle layer of the application area, such as the face or neck, or the dermal layer of the skin.Compositions comprising the disclosed combinations, including liposomal formulations, may be administered by direct injection subcutaneously, intradermally, or by iontophoresis, to deposit the active agent at the site requiring muscle relaxation or decontraction.

[0079] In another of its aspects, the present invention provides the disclosed combinations and methods thereof that can improve the aesthetic appearance of the skin by treating, including preventing, ameliorating and / or reducing at least one of the following: dermatological aging, in particular natural, photogenic or hormonal aging, the improvement preferably occurring after topical application of a product or formulation comprising one or more of the disclosed combinations as described herein.

[0080] [Composition] The composition described herein may be formulated as food, beverage, food additive, beverage additive, dietary supplement, nutritional product, medical food, nutritional functional food, medicine or pharmaceutical product.Preferably, the composition of the present invention is formulated as powder, liquid, food bar, spread, sauce, ointment, tablet or capsule.Suitable formulations may be prepared by those skilled in the art with reference to the technology and teachings of this specification.

[0081] The compositions described herein may be formulated to be administered to a subject by any selected route, including, but not limited to, oral, nasal, or parenteral (including topical, subcutaneous, intramuscular, and intravenous) administration.

[0082] Thus, pharmaceutical compositions useful in the present invention may be formulated with suitable pharma- ceutically acceptable carriers (including excipients and diluents) selected with respect to the intended route of administration and standard pharmaceutical practice. For example, the compositions of the present invention may be administered orally as a powder, liquid, tablet or capsule, or topically as an ointment, cream or lotion. Suitable formulations may optionally contain additional agents, including emulsifiers, antioxidants, flavorings or colorings, and may be adapted for immediate, delayed, modified, sustained, pulsatile or controlled release.

[0083] The compositions useful herein may be used alone or in combination with one or more other therapeutic agents. The therapeutic agent may be a food, beverage, food additive, beverage additive, food component, beverage component, dietary supplement, nutritional product, medical food, nutritional functional food, medicine or pharmaceutical product.

[0084] When used in combination with another therapeutic agent, administration of the composition of the present invention and the other therapeutic agent may be simultaneous or sequential. Simultaneous administration includes administration of a single dosage form containing all components, as well as substantially simultaneous administration of the composition of the present invention and the other therapeutic agent as separate dosage forms. Sequential administration includes administration of the composition of the present invention and the other therapeutic agent according to different schedules, preferably such that there is an overlap in the periods during which the composition of the present invention and the other therapeutic agent are provided.

[0085] Suitable agents that may be found for use with the compositions of the present invention may be co-administered, including antihistamines, anti-inflammatory agents, antirheumatic agents, corticosteroids, nonsteroidal anti-inflammatory drugs (NSAIDs) including cyclooxygenase-2 selective inhibitors, muscle relaxants, combinations of any two or more of these, and other suitable agents known in the art.

[0086] As will be appreciated, the dose of the composition administered, the duration of administration, and the general administration regimen may vary between subjects depending on such variables as the severity of the subject's symptoms, the type of disorder to be treated, the mode of administration selected, and the age, sex, and / or general health of the subject. However, as a general example, the inventors contemplate administration of about 1 mg to about 1000 mg per kg of body weight or more of the composition of the invention per day, preferably about 50 to about 500 mg per kg per day. In one embodiment, the inventors contemplate administration of about 0.05 mg to about 250 mg per kg of body weight of the pharmaceutical composition according to the invention. It should be understood that administration may include a single daily dose, or multiple individually divided doses may be administered as needed.

[0087] The compositions described herein also include cosmetic compositions that have the effect of enhancing skin moisture, exfoliating the skin, enhancing skin elasticity, promoting skin healing, preventing erythema, improving skin wrinkles, and / or reducing photoaging of the skin.

[0088] In another exemplary embodiment, the cosmetic composition is prepared into any one formulation selected from the group consisting of skin lotion, skin softener, skin toner, astringent, lotion, milk lotion, moisturizing lotion, nourishing lotion, massage cream, nourishing cream, moisturizing cream, hand cream, essence, pack, mask pack, mask sheet, peeling agent, soap, shampoo, cleansing foam, cleansing lotion, cleansing cream, body lotion, body cleanser, emulsion, pressed powder, loose powder, and eye shadow.

[0089] In one exemplary embodiment, the amount of placental extract is in the range of about 0.05 to about 1 wt.% based on the total weight of the cosmetic composition. Preferably, the amount of placental extract is in the range of about 0.05 to about 0.5 wt.%, about 0.05 to about 0.4 wt.%, about 0.05 to about 0.35 wt.%, about 0.1 to about 0.5 wt.%, about 0.1 to about 0.4 wt.%, about 0.135 to about 0.35 wt.%, or about 0.15 to about 0.2 wt.%. In one embodiment, the composition is for application to the skin, such as a face mask, and comprises 0.13 wt.% of placental extract. In one embodiment, the composition is a cream, gel, or lotion for anti-aging applications, and comprises 0.135% to 0.35 wt.% of placental extract. In one embodiment, the composition is a cream, gel, or lotion for skin whitening, and comprises 0.15 to 0.2 wt.% of placental extract.

[0090] In addition to the placenta extract, the cosmetic composition of the present disclosure may further contain other additives, such as excipients, carriers, etc., and common ingredients added to common skin cosmetics may be applied to the cosmetic composition in the required amount and mixed therein.

[0091] In particular, the cosmetic composition of the present disclosure may further comprise a percutaneous penetration enhancer. The term "percutaneous penetration enhancer" as used herein refers to a composition that allows desired components to penetrate into the vascular cells of the skin at a high absorption rate. Non-limiting examples of percutaneous penetration enhancers may include lecithin, other phospholipid components such as those used in cosmetics, liposome components, etc.

[0092] Furthermore, the oil that may be mainly used as the oil component may be at least one selected from vegetable oil, mineral oil, silicone oil, and synthetic oil.More specifically, mineral oil, cyclomethicone, squalane, octyldodecyl myristate, olive oil, European grape seed oil, macadamia nut oil, glyceryl octanoate, castor oil, ethylhexyl isononanoate, dimethicone, cyclopentasiloxane, sunflower seed oil, etc. may be used.

[0093] Furthermore, in order to enhance the emulsifying ability, about 0.1 wt% to about 5 wt% of a surfactant, higher alcohol, etc. may be added. The surfactant may be a general surfactant, for example, a nonionic surfactant, an anionic surfactant, a cationic surfactant, an amphoteric surfactant, a phospholipid, etc., such as sorbitan sesquinolate, polysorbate 60, glyceryl stearate, lipophilic glyceryl stearate, sorbitan oleate, sorbitan stearate, diacetyl phosphate, sorbitan stearate / sucrose cocoate, glyceryl stearate / polyethylene glycol-100 stearate, ceteareth-6 olivate, arachidyl alcohol / behenyl alcohol / arachidyl glucoside, polypropylene glycol-26-buteth-26 / polyethylene glycol-40 hydrogenated castor oil, etc. The higher alcohol may be a C12 to C20 alcohol, such as cetyl alcohol, stearyl alcohol, octyldodecanol, isostearyl alcohol, etc., and these higher alcohols may be used alone or in combination of at least two of them.

[0094] To adjust the viscosity or hardness of the aqueous phase component, about 0.001 wt % to about 5 wt % of at least one thickening agent selected from carbomer, xanthan gum, bentonite, magnesium aluminum silicate, cellulose gum, dextrin palmitate, and the like, may be further added.

[0095] Furthermore, the cosmetic composition according to the present disclosure may further contain, as necessary, components such as medicinal ingredients, for example, higher fatty acids, vitamins, etc.; UV screening agents; antioxidants (butyl hydroxyanisole, propyl gallate, erythorbic acid, tocopherol acetate, butylated hydroxytoluene, etc.); preservatives (methylparaben, butylparaben, propylparaben, phenoxyethanol, imidazolidinyl urea, chlorphenetin, etc.); colorants, pH adjusters (triethanolamine, citric acid, sodium citrate, malic acid, sodium malate, fumaric acid, sodium fumarate, succinic acid, sodium succinate, sodium hydroxide, sodium monohydrogen phosphate, etc.); moisturizers (glycerin, sorbitol, propylene glycol, butylene glycol, hexylene glycol, diglycerin, betaine, glycereth-26, methyl gluceth-20, etc.); lubricants, etc.

[0096] Additionally, the cosmetic compositions of the present disclosure may further include adjuvants for providing essential nutrients to the skin, such as adjuvants including natural or cosmetic fragrances or medicinal herbs, but are not limited to these examples.

[0097] The compositions described herein also include health functional food compositions that have the effects of enhancing skin moisture, exfoliating the skin, promoting skin elasticity, preventing erythema, improving skin wrinkles, and / or reducing photoaging of the skin.

[0098] In an exemplary embodiment, the health functional food composition is prepared into any one formulation selected from the group consisting of tablets, granules, powders, soft gels, pastes, jellies, puddings, capsules, liquid solutions (such as beverages), and pills.

[0099] In one exemplary embodiment, the amount of placenta extract is in the range of about 50 to about 500 mg when the health functional food composition is in the form of a beverage. Preferably, the amount of placenta extract is in the range of about 50 to about 350 mg, about 55 to about 350 mg, about 60 to about 300 mg, about 65 to about 250 mg, about 65 to about 200 mg, about 65 to about 180 mg, about 65 to about 160 mg, or about 65 to about 155 mg. In one embodiment, the health functional food composition is in the form of a shot and contains about 10 to about 100 mg, about 10 to about 80 mg, about 20 to about 70 mg, or about 25 to about 50 mg of placenta extract.

[0100] In another exemplary embodiment, the cosmetic composition or health functional food composition may further comprise an ingredient that improves skin wrinkles.

[0101] In another exemplary embodiment, the ingredient for improving skin wrinkles includes one or more selected from the group consisting of vitamin C, retinoic acid, transforming growth factor (TGF), animal placenta-derived protein, betulinic acid, and chlorella extract.

[0102] Various aspects of the present invention will now be illustrated, in a non-limiting manner, by reference to the following examples. EXAMPLES

[0103] [Example 1] Evaluating the immune modulation effects of deer placenta Materials and Methods 1. Generation of placental extract (DPE) Freeze-dried deer placenta powder was dissolved in various solvents to solubilize the major portion of the base material and generate the final extracts tested that are physiologically compliant. Protein concentration of the extracts (DPE) was determined by absorbance at 280 nm. Protein integrity was checked by SDS-PAGE / Coomassie Brilliant Blue staining.

[0104] 2. Determination of cytotoxicity Cytotoxicity was determined according to the requirements of ISO 10993-5, 2009. The test performance will be described below.

[0105] 3. Assessment of acute systemic oral toxicity Acute oral toxicity determinations were performed in an accredited animal testing facility under GLP according to OECD 423.

[0106] 4. Evaluation of immune modulation effects Human immune cells (PBMCs: peripheral blood mononuclear cells) were exposed to various concentrations of additional immunological stimuli as controls for a defined time period (16 h, 37°C), after which they were isolated from the supernatant and cellular and cytokine expression patterns were analyzed on a multiplexing platform (Mesoscale Discoveries U-Plex).

[0107] Experimental setup:

[0108] [Table 1]

[0109] [Table 2]

[0110] [Table 3]

[0111] [Table 4]

[0112] [Table 5]

[0113] result Preparation of placental extract: Freeze-dried deer placenta powder was dissolved in various solvents to solubilize the major portion of the base material and ensure that the final tested extracts were physiologically compliant. Protein concentration of the extracts was determined by absorbance at 280 nm. Protein integrity was checked by SDS-PAGE / Coomassie Brilliant Blue staining (Figure 1).

[0114] [Table 6]

[0115] conclusion Concerning the generation of protein extracts, it could be observed that after 24 hours of extraction at 37 °C, a maximum of about 1 / 3 of the initially applied material yielded a soluble protein fraction. In this context, no relevant differences were observed between distilled water, PBS and cell culture medium (RPMI 1640 w / o phenol red), giving the option of sticking to cell culture medium for the following physiological tests. Concerning protein integrity, a similar banding pattern could be observed within the range of the different extracts, this was no exception even for extracts without detergent. The high molecular weight bands migrating above an apparent molecular weight of 150 kDa are typical of cytokeratins. Concerning these results, the following setup was performed in cell culture medium (RPMI 1640 w / o phenol red supplemented with FBS (10% fc) and gentamicin (50 μg / mL) prior to the physiological tests).

[0116] Cytotoxicity determination Cytotoxicity was determined according to the requirements of ISO 10993-5, 2009. In this particular context, various dilutions of the test extracts ranging from 1 / 2 to 1 / 1000 dilution were subjected to an evaluation of their cytotoxic potential against the test cell line L929. The readout was generated by conversion of the vital dye XTT (ISO 10993-5. Annex D) combined with visual inspection of the cultures.

[0117] Control: Negative control (NC): Test cells in complete culture medium (ccm) Positive control (PC): Test cells in complete culture medium and 1% SDS Solvent control (SC): extraction medium w / o test substance

[0118] [Table 7]

[0119] [Table 8]

[0120] Optical assessment (grading): 0: cells with optimal shape; 1: cells with good shape, some detached cells may be seen; 2-3: cells partially to definitively detached and showing signs of apoptosis; 4: culture is completely damaged, no viable cells are seen.

[0121] Conclusion: Regarding in vitro cytotoxicity determination, no cytotoxic effects were observed at dilutions of 1:10 and protein concentrations below 3.49 mg / mL.

[0122] Acute oral toxicity assessment Acute oral toxicity determinations were performed as described above.

[0123] [Table 9]

[0124] Determining the immune modulation effect For the evaluation of the immunomodulation effect, human PBMCs were treated directly with the indicated extracts and stimuli (16 h, 37° C., 5% CO2) or pre-incubated for the same time period (1.23 mg / mL) and then 3 h later. Three different concentrations were selected for the evaluation of the immunomodulation effect. Starting with the starting extract (61.6 mg / mL), the following dilutions were tested: 1 / 5 dilution: 12.32mg / mL → Extract within cytotoxic range 1 / 20 dilution: 3.08mg / mL → Extract just below the border of cytotoxicity 1 / 50 dilution: 1.23 mg / mL → extract in the non-cytotoxic range. This dilution was also used for pretreatment of PBMCs before addition of known immunologically active stimuli (endotoxin, zymosan, heat-inactivated Staphylococcus aureus).

[0125] After treatment, supernatants from each test culture were isolated and analyzed on a multiplexing platform (Mesoscale U-Plex). The following parameters / cytokines were analyzed: IL-1beta, IL-6, IL-10, TNF-alpha.

[0126] Immunomodulation stimuli: endotoxin (0.1 / 0.05 / 0.01EU / mL fc); zymosan (5 / 1 / 0.2mg / mL fc); heat-inactivated Staphylococcus aureus (1×10 8 / 1×10 7 cell fc)

[0127] All parameters were analyzed in at least two individual settings. Mean values ​​of signal intensity are shown in the graphs and tables below.

[0128] [Table 10]

[0129] Conclusion: IL-1β Concerning the induction of IL-1 beta upon stimulation of human PBMCs, it could be observed that the cytotoxic concentration (12.33 mg / mL) of DPE led to an induction / release of IL-1 beta from PBMCs within the range of the positive control HKSA, even higher than the stimulants zymosan and endotoxin. For DPE concentrations below the cytotoxic border (1:20:3.08 mg / mL; 1:50:1.23 mg / mL), a strong reduction in the induction of IL-1 beta could be observed (Figure 2).

[0130] IL-6 Regarding the induction of IL-6 when stimulating human PBMC, DPE at a concentration (12.33 mg / mL) that induces cytotoxicity induced / released IL-6 from PBMC within the range of the positive control endotoxin, but zymosan (0.2 μg / mL) and HKSA (10 8 It could be observed that the induction of IL-6 was clearly lower than that of stimulation with 1:20: 3.08 mg / mL; 1:50: 1.23 mg / mL DPE concentrations below the cytotoxicity border (Figure 3).

[0131] Interference with cytokine induction To determine the potential interference of DPE with the induction of inflammatory cytokines upon stimulation, human PBMCs were pretreated for 3 h with DPE at a concentration clearly below the borderline cytotoxicity (1 / 50 dilution of the initial extract: 1.23 mg / mL). Then, the established PBMC stimuli endotoxin, zymosan and heat-killed Staphylococcus aureus (HKSA) were added to the cultures and pretreated for another 16 h. In a final step, the supernatants were removed and analyzed on the Mesoscale U-Plex platform.

[0132] [Table 11]

[0133] Conclusion: Interleukin 1 beta (IL-1β) is considered a proinflammatory cytokine and is a key mediator of the inflammatory response.

[0134] Regarding the interference of DPE treatment of human PBMCs at a concentration of 1.23 mg / mL before the addition of the indicated stimuli, it can be observed that the IL-1beta expression induced by endotoxin was reduced by 23.7%, by HSKA by 37.2% and by zymosan by 49.5% (Figure 4).

[0135] [Table 12]

[0136] Interleukin 6 (IL-6) is an interleukin that acts as a proinflammatory cytokine, and there is some evidence that IL-6 can be used as an inflammatory marker for severe COVID-19 infection associated with poor prognosis.

[0137] With regard to the intervention of DPE treatment of human PBMC at a concentration of 1.23 mg / mL prior to the addition of the indicated stimuli, a DPE-dependent inhibition (-24.9%) of IL-6 expression by zymosan can be observed.

[0138] [Table 13]

[0139] Conclusion: Interleukin-10 (IL-10) is considered an anti-inflammatory cytokine with many functions in regulating the immune system.

[0140] With regard to the intervention of DPE treatment of human PBMC at a concentration of 1.23 mg / mL before the addition of the indicated stimuli, it can be observed that IL-10 expression by HKSA was induced by DPE (19.3%).

[0141] [Table 14]

[0142] Human PBMCs were pretreated with DPE as described. Then, established stimuli were administered at the following concentrations: Zymosan 2 (1 μg / mL), HKSA1 (1 × 10 8 ), and endotoxin 1 (0.05EU / mL). Supernatants were collected and analyzed on the Mesoscale U-PLEX platform. Data shown here is similar to a 1 / 10 dilution on the Mesoscale platform.

[0143] Conclusion: In general, DPE treatment by itself did not induce expression of TNF-alpha, which was neither suppressed nor induced by DPE pretreatment of human PBMCs in the context of endotoxin reactivity as a stimulator.

[0144] Regarding the interference of DPE treatment at a concentration of 1.23 mg / mL on human PBMCs prior to the addition of the indicated stimuli, it can be observed that TNF-alpha expression by HKSA was slightly inhibited by endotoxin (9%) and by DPE (11.8%). IL-10 expression by zymosan was clearly inhibited (60.3%) by pretreatment with DPE.

[0145] Final conclusion: Cytotoxicity determination: Cytotoxic effects of DPE were observed at concentrations above 3.5 mg / mL. Dilutions below 3.5 mg / mL did not demonstrate cytotoxic effects.

[0146] Assessment of acute systemic oral toxicity: No acute systemic oral toxicity was observed

[0147] Determining immune modulation effects: Concerning the direct effect on human PBMCs at concentrations in the cytotoxic range (12.33 mg / mL), induction of the proinflammatory cytokines IL-1 beta and IL-6 was observed. At concentrations below 3.08 mg / mL, these effects were reduced. Concerning the immunomodulatory effect of DPE, by pretreating human PBMCs with a non-cytotoxic concentration (1.23 mg / mL) of DPE before stimulation with potent activators such as endotoxin, zymosan and heat-killed S. aureus, it could be observed that DPE reduces the zymosan-dependent induction of IL-1 beta, IL-6 and TNF-alpha. A general effect on the expression of IL-1 beta could be observed. DPE was also shown to induce the expression of the anti-inflammatory cytokine IL-10 in response to heat-killed S. aureus.

[0148] As discussed above, COVID-19 caused by SARS-CoV-2 is characterized by immune dysfunction rather than viral load, which leads to the abnormal production of pro-inflammatory cytokines. In particular, COVID-19 can induce a cytokine storm in lung tissues via a hyperactive immune system and uncontrolled release of cytokines. Pro-inflammatory cytokines, such as interleukin-6 (IL-6), interleukin-1β (IL-1β), and tumor necrosis factor-alpha (TNF-α), play a crucial role in lung injury in COVID patients who present with acute respiratory distress syndrome (ARDS) due to dysfunction of the respiratory epithelium.

[0149] The findings that DPE can suppress the induction of IL-6, IL-1β, and TNF-α, respectively, as well as induce expression of the anti-inflammatory cytokine IL-10, point to the potential of DPE as a potential therapeutic to suppress the inflammatory response associated with COVID-19.

[0150] [Example 2] [method] DPE Generation Test product: DPE (Deer Placenta Powder, Lot No. NPR00 / 65#2 / 2R008). Stored refrigerated at 4°C.

[0151] Deer placenta powder (100 mg / mL) was dissolved in additive-free cell culture medium (KGM2, Promocell) and incubated for 24 ± 2 hours at 37 ± 1 °C. The extract (DPE) was centrifuged (2 min, 14000 rpm, room temperature) and filtered (0.4 μm). The protein concentration of the extract was determined by absorbance measurement at 280 nm.

[0152] Dose study results The working concentration of DPE was evaluated based on its effect on cell viability. A neutral red uptake assay was performed.

[0153] For the following experiments, according to the results of this viability test, the extract was produced in one concentration (non-toxic) or three concentrations (non-toxic / borderline / toxic).

[0154] Lymphocyte stimulation As an in vitro test system, the BEAS-2B immortalized human bronchial epithelial cell line was used. The BEAS-2B cell line was used as an in-vitro model for various diseases based on inflammatory processes, such as asthma, infection, ARDS or COVID-19.

[0155] BEAS-2B cells were treated with or without (control) three different concentrations of DPE for 24 hours. Final concentrations of DPE were: 3.6 mg / mL; 1.8 mg / mL; 0.9 mg / mL.

[0156] T lymphocytes were isolated from freshly drawn human blood samples using the Pan T cell isolation kit (Miltenyi Biotec) and activated using T cell transAct (Miltenyi Biotec).

[0157] Activated T-lymphocytes were then added to the pre-loaded cells for 1 h to trigger the inflammatory process, and after a further 24 h, supernatant samples were taken for cytokine analysis.

[0158] Cells were harvested and total cell protein was determined as a reference value.

[0159] Cytokine analysis Human interleukin 10 (IL-10) concentrations in the supernatants were determined by ELISA technique (manufacturer: RnD Systems D1000B; lot / expiry date: P32207118.10.2022).

[0160] [result] Preparation of extracts Protein content was determined spectrophotometrically at 280 nm using a NanoQuant Plate and an Infitite 200Pro plate reader (Tecan).

[0161] Protein content in deer placenta powder extract: 25.33mg / mL

[0162] Dose study results The working concentration of DPE was evaluated based on its effect on cell viability. Figure 5 shows the results of the dose study.

[0163] Based on the results of the viability assay, further investigations were carried out using the concentrations of DPE shown in Table 15:

[0164] [Table 15]

[0165] Interleukin 10 after stimulation with activated lymphocytes For the evaluation of the effect of DPE on IL-10 release in BEAS-2B cells, the cells were pre-treated with or without (control) three different concentrations of deer placenta extract (C1: 3.6 mg / mL; C2: 1.8 mg / mL; C3: 0.9 mg / mL) for 24 hours. The cells were then co-cultured with freshly isolated activated lymphocytes or with non-activated lymphocytes as a trigger for the inflammatory process. After 24 hours, supernatant samples were taken for cytokine analysis. Activated lymphocytes were chosen as representative for COVID-19 as a trigger for the inflammatory process in an in vitro test system.

[0166] Cells were harvested and total cell protein was determined as a reference value, and the results are shown in Figure 6.

[0167] IL-10 was not detectable in the supernatants of BEAS-2B cultures containing non-activated lymphocytes. When stress was induced by activated lymphocytes, IL-10 was detectable in all groups.

[0168] The release of IL-10 was enhanced by 1.0 mg / mL DPE compared to the control.

[0169] The above methods should in no way be considered as limiting, and suitable variations or alternatives will be apparent to those skilled in the art. [Industrial Applicability]

[0170] The present invention has utility in the treatment or prevention of inflammatory conditions. The described compositions and methods of the invention may be used to treat or prevent one or more of the conditions discussed above.

[0171] Those skilled in the art will appreciate that the above description is provided by way of example only and that the present invention is not limited thereto.

Claims

1. 13. A composition for use in the treatment, prevention or amelioration of a disease, disorder or condition associated with inflammation or immune modulation, the composition comprising a placenta extract.

2. Use of a placental extract in the manufacture of a medicament or composition for treating, preventing or ameliorating a disease, disorder or condition associated with inflammation or immune modulation.

3. The composition according to claim 1 or the use according to claim 2, wherein the composition or medicament is a food, a beverage, a food additive, a beverage additive, a food component, a beverage component, a nutritional supplement, a nutritional product, a medical food, a nutritional functional food, a medicine or a pharmaceutical product.

4. The composition of claim 1 or the use of claim 2, wherein the composition or medicament is used such that the placental extract is administered at a concentration sufficient to inhibit one or more pro-inflammatory cytokines, including the cytokines IL-1α, β, IL-2, IL-3, IL-6, IL-7, IL-9, IL-12, IL-17, IL-18, TNF-α, LT, LIF, oncostatin, or IFNc1α, β, γ.

5. The composition of claim 1 or the use of claim 2, characterized in that the composition or medicament is used such that the placental extract is administered at a concentration sufficient to stimulate expression of one or more anti-inflammatory cytokines, including IL-4, IL-10, IL-11, W-13 or TGFβ.

6. 3. The composition according to claim 1 or the use according to claim 2, wherein the placental extract is derived from the placenta of a deer, sheep, goat, horse, donkey, rabbit or cow.

7. 3. The composition of claim 1 or the use of claim 2, wherein the placental extract is a placental extract extracted from placental material by an extraction method selected from the group consisting of solvent extraction, supercritical solvent extraction including supercritical CO2 extraction, distillation, countercurrent extraction, decoction, percolation, maturation, molecular distillation, microwave extraction, ultrasonic extraction, and chromatographic separation.

8. 3. The composition of claim 1 or the use of claim 2, wherein the placental extract is a placental extract extracted from placental material by solvent extraction.

9. 8. The composition or use of claim 7, wherein the extraction process comprises contacting the placental material with a solvent, separating the placental material from the solvent, and at least partially removing the solvent to obtain an extract.

10. 10. The composition or use of claim 9, wherein the placental material is freeze-dried and powdered prior to contacting with the solvent.

11. 10. The composition or use according to claim 9, wherein the solvent is water.

12. 10. The composition or use according to claim 9, wherein the solvent is an organic solvent.

13. 13. The composition or use according to claim 12, wherein the solvent is ethanol.

14. 3. The composition of claim 1 or the use of claim 2, wherein the placental extract is a placental extract prepared by solvent extraction followed by distillation or supercritical extraction.

15. The composition described in claim 1 or the use described in claim 2, wherein the placental extract is a placental extract prepared using aqueous extraction.

16. 3. The composition of claim 1 or the use of claim 2, wherein the placental extract is or comprises dried or powdered placental material.

17. 17. The composition or use of claim 16, wherein the placental material is powdered or granulated placental material that has been micronized by wet grinding followed by freeze drying, vacuum drying or spray drying, or fluidized bed drying.

18. 17. The composition or use of claim 16, wherein the placental material is dried placental material.

19. 3. The composition of claim 1 or the use of claim 2, wherein the composition or medicament is provided as a delivery formulation selected from the group consisting of a tablet, a capsule, a liquid, an oil, a suspension, an emulsion, a paste, a jelly, a pudding, a solution, and a powder.

20. The composition or the medicament is a. at least 0.1, 0.2, 0.5, 1, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 99, 99.5, 99.8, or 99.9% by weight of said placental extract; b. at least 0.001, 0.01, 0.05, 0.1, 0.15, 0.2, 0.3, 0.4, 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 grams or more of the placental extract; or c. at least 1, 10, 50, 100, 125, 150, 200, 250, 300, 350, 400, 450, 500, 600, 700, 800, 900, or 1000 mg or more of the placental extract; 3. The composition according to claim 1 or the use according to claim 2, comprising:

21. The composition or the medicament, a. the placental extract is used to be administered in an amount of 125 mg, suitable for administration twice daily, or in an amount of 250 mg, suitable for administration once daily; b. the placental extract is administered in an amount of 100 mg or 125 mg, suitable for administration twice daily, or in an amount of 200 mg or 250 mg, suitable for administration once daily; or c. 40% to 90% by weight of the placenta extract is administered.

3. The composition according to claim 1 or the use according to claim 2.

22. 3. The composition of claim 1 or the use of claim 2, wherein the composition or medicament further comprises 0.1, 0.5, 1, 5, 10, 15, 20, 25, 30, 35, 40, 45 or 50% by weight of another anti-inflammatory agent.

23. The composition of claim 1 or the use of claim 2, wherein the composition or medicament further comprises a carrier.

24. 3. The composition of claim 1 or the use of claim 2, wherein the composition or medicament is in the form of a tablet, caplet, pill, hard or soft capsule, lozenge, cachet, sachet, powder, granule, suspension, elixir, liquid, or other form that can be added to a food or beverage.

25. 3. The composition of claim 1 or the use of claim 2, wherein the composition or medicament further comprises one or more components that prevent or reduce degradation of the composition during storage or after administration.

26. 3. The composition of claim 1 or the use of claim 2, wherein the composition or medicament is or is formulated as a food, beverage, food additive, beverage additive, dietary supplement, nutritional product, medical food, nutritional functional food, medicine or pharmaceutical product.

27. The composition or medicament is formulated for: a. Oral, nasal or parenteral administration; or b. Ingestion, c. Inhalation; or d. topical application; A composition according to claim 1 or a use according to claim 2.

28. 3. The composition of claim 1 or the use of claim 2, wherein the placental extract or composition is formulated for administration in an amount sufficient to treat inflammation-related diseases by inhibiting expression of pro-inflammatory cytokines and / or stimulating anti-inflammatory cytokines.

29. The disease, disorder or condition is selected from the group consisting of type I diabetes; Sjogren's syndrome; uveitis; celiac disease; allergic conjunctivitis; non-specific colitis, arthritis, rheumatoid arthritis, inflammatory bowel disease; psoriasis; multiple sclerosis; neurodegenerative disorders; congestive heart failure; stroke; aortic stenosis; renal failure; lupus; pancreatitis; allergies; fibrosis; anemia; Atherosclerosis; metabolic diseases; bone diseases; The composition of claim 1 or the use of claim 2, wherein the inflammation-related disease is selected from the group consisting of cardiovascular disease, chemotherapy / radiation-related complications; type II diabetes; liver disease; gastrointestinal disorders; ophthalmological diseases; diabetic retinopathy; pulmonary disorders, kidney diseases; dermatitis; HIV-associated cachexia; cerebral malaria; ankylosing spondylitis; leprosy; anemia; and fibromyalgia.

30. 3. The composition of claim 1 or the use of claim 2, wherein the disease, disorder or condition is a pulmonary disease, disorder or condition.

31. The disease, disorder or condition is a) a disease, disorder or condition associated with or caused by an immune response, which is an autoimmune disease or graft-versus-host disease; b) an autoimmune disease selected from rheumatoid arthritis, scleroderma, inflammatory bowel disease, or systemic lupus erythematosus; c) interstitial lung disease; d) obstructive pulmonary disease selected from asthma, bronchitis, acute respiratory distress syndrome or chronic obstructive pulmonary disease; e) acute lung injury selected from one or more of physical trauma, chemical injury, smoke inhalation, or exposure to a toxic substance; 3. The composition according to claim 1 or the use according to claim 2,

32. 3. The composition of claim 1 or the use of claim 2, wherein the disease, disorder or condition is one or more of pulmonary fibrotic disease, acute respiratory distress syndrome (ARDS), COVID-19, chronic obstructive pulmonary disease (COPD), emphysema, asthma, viral or bacterial infection of the lungs, pneumonia (including chemically induced pneumonia), or cystic fibrosis.

33. The composition of claim 1 or the use of claim 2, wherein the composition or medicament is for treating, preventing or ameliorating a viral disease and / or for inhibiting activation of the virus, and the virus is selected from the group consisting of herpes virus; influenza virus; paramyxovirus; respiratory syncytial virus; coronavirus; HIV virus; hepatitis virus; or rotavirus.

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