Human Breast Topical Preparations

Abstract

Provided herein are compositions, such as topical preparations, that contain one or more fractions, parts, or components of human milk, as well as methods of use and manufacture thereof.In some embodiments, the compositions provided include human milk cream, or one or more fatty acids found in or derived from human milk.Methods for treating or preventing skin conditions, such as atopic dermatitis or psoriasis, are also included.

Description

[Technical field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims priority to U.S. Provisional Application No. 63 / 140,544, filed January 22, 2021, entitled "TOPICAL HUMAN MILK FORMULATIONS," and U.S. Provisional Application No. 63 / 195,392, filed June 1, 2021, entitled "TOPICAL HUMAN MILK FORMULATIONS," the contents of both of which are incorporated by reference in their entireties.

[0002] Provided herein are compositions, such as topical preparations, that contain one or more fractions, parts, or components of human milk, as well as methods of use and manufacture thereof.In some embodiments, the compositions provided include human milk cream, or one or more fatty acids found in or derived from human milk.Methods for treating or preventing skin conditions, such as atopic dermatitis or psoriasis, are also included. [Background technology]

[0003] The skin serves many functions, but its main function is as a protective layer or barrier. An important role of the skin for terrestrial animals is to protect the moist internal organs from the dry external environment. In addition, the skin protects the internal tissues from harmful chemical and physical forces as well as from the invasion of pathogens.

[0004] Existing creams, lotions, ointments, and other product formulations for skin care may improve the appearance of skin in some cases, but may not improve barrier function. In other cases, such treatments may be formulated to provide antioxidant, anti-inflammatory, or free radical scavenging effects, but may not provide such effects, for example, due to an inability to penetrate or integrate well into the barrier. Other skin treatments, such as corticosteroids, may actually weaken barrier function. Others, while claiming to be natural, are still derived from potentially allergenic plant extracts or animal proteins. There is a need for topical formulations that can treat and / or prevent skin conditions, promote wound healing, or reduce inflammation while avoiding undesirable secondary effects. Summary of the Invention

[0005] Provided herein is a topical formulation comprising one or more human milk fractions and one or more excipients, e.g., pharma- ceutically acceptable excipients, wherein the one or more human milk fractions comprise one or more of human milk cream or a subfraction thereof, human skim milk, human milk permeate, or human milk retentate, wherein the human milk permeate and human milk retentate are obtained from filtration of human skim milk. In some embodiments, the topical formulation comprises human milk cream. In certain embodiments, the topical formulation comprises a subfraction obtained from or derived from human milk cream. In some embodiments, the subfraction is butter oil. In certain embodiments, the subfraction is a milk fat globule membrane solid subfraction. In some embodiments, the topical formulation comprises human milk fatty acid.

[0006] In some embodiments, the topical formulation comprises one or more of oleic acid, palmitic acid, linoleic acid, omega-3 fatty acid, omega-6 fatty acid, omega-9 fatty acid, stearic acid, myristic acid, lauric acid, palmitoleic acid, vaccenic acid, myristoleic acid, heptadecenoic acid, gamma linoleic acid, docosapentaenoic acid, eicosapentaenoic acid, arachidonic acid, and docosahexaenoic acid.In some embodiments, the topical formulation comprises oleic acid, palmitic acid, linoleic acid, omega-3 fatty acid, omega-6 fatty acid, omega-9 fatty acid, stearic acid, myristic acid, lauric acid, palmitoleic acid, vaccenic acid, myristoleic acid, heptadecenoic acid, gamma linoleic acid, docosapentaenoic acid, eicosapentaenoic acid, arachidonic acid, and docosahexaenoic acid.

[0007] In certain embodiments, the topical formulation comprises human milk permeate, which is obtained from filtering human skim milk. In some embodiments, filtering is or includes ultrafiltration. In some embodiments, the topical formulation comprises human milk fatty acids and human milk permeate. In certain embodiments, the topical formulation comprises a mixture of human milk oligosaccharides, which mixture of human milk oligosaccharides comprises two or more different HMOs.

[0008] In certain embodiments, the topical formulation comprises 1%-30% or 1%-40% human milk fat, fatty acid, or lipid as a percentage of fat, fatty acid, or lipid weight relative to total weight (w / w). In certain embodiments, the topical formulation comprises 5%-25% human milk fat, fatty acid, or lipid (w / w). In some embodiments, the topical formulation comprises no more than 30%, 25%, or 10% human milk fat, fatty acid, or lipid (w / w). In some embodiments, the topical formulation comprises 1%-10% human milk fat, fatty acid, or lipid (w / w). In certain embodiments, the topical formulation comprises at least or about 10% human milk fat, fatty acid, or lipid (w / w). In certain embodiments, the topical formulation comprises at least or about 20% human milk fat, fatty acid, or lipid (w / w). In certain embodiments, the topical formulation comprises at least or about 30% human milk fat, fatty acid, or lipid (w / w). In some embodiments, the topical formulation comprises one or more human milk oligosaccharides.

[0009] Also provided herein are topical formulations comprising human milk cream and human milk permeate, the topical formulations comprising 1%-10%, or 1%-40%, or 10%-40% human milk fats, lipids, or fatty acids (w / w), the human milk permeate being obtained from ultrafiltration of human skim milk. In some embodiments, the topical formulations comprise at least 0.3% human milk oligosaccharides (w / w). In some embodiments, the topical formulations comprise at least 0.3% human milk oligosaccharides (w / w). In some embodiments, the topical formulations comprise 0.1%-0.5% human milk oligosaccharides (w / w).

[0010] In some embodiments, the topical formulation comprises a mixture of at least two human milk oligosaccharides. In certain embodiments, the topical formulation comprises a mixture of at least 25, 50, 75, or 100 human milk oligosaccharides. In some embodiments, the topical formulation comprises one or more of 2'-fucosyl-lactose, 3-fucosyl-lactose, 3'-sialyl-lactose, 6'-sialyl-lactose, lacto-N-tetraose, lacto-N-difucohexaose I, lactodifucotetraose, lacto-N-fucopentaose I, sialylacto-N-tetraose c, sialylacto-N-tetraose b, and disialylacto-N-tetraose. In some embodiments, the topical formulation comprises 2'-fucosyl-lactose, 3-fucosyl-lactose, 3'-sialyl-lactose, 6'-sialyl-lactose, lacto-N-tetraose, lacto-N-difucohexaose I, lactodifucotetraose, lacto-N-fucopentaose I, sialylacto-N-tetraose c, sialylacto-N-tetraose b, and disialyllacto-N-tetraose.

[0011] In some embodiments, the topical formulation comprises human milk retentate, which is obtained from filtration of human skim milk. In some embodiments, the filtration is or comprises ultrafiltration. In some embodiments, the topical formulation comprises one or more human milk proteins. In certain embodiments, the topical formulation comprises one or more of casein, alpha-lactalbumin, lactoferrin, secretory immunoglobulin IgA, lysozyme, serum albumin, lactadherin, dermcidin, immunoglobulin, cytokine, mucin, or growth factor.

[0012] In some embodiments, the topical formulation comprises 0.1%-0.5% human milk oligosaccharides (w / w) and 1%-30% human milk fat, lipid, or fatty acid (w / w). In certain embodiments, the topical formulation comprises about 0.3%% human milk oligosaccharides (w / w) and about 20% human milk fat, fatty acid, or lipid (w / w). In some embodiments, the topical formulation comprises about 0.3%% human milk oligosaccharides (w / w) and about 10% human milk fat, lipid, or fatty acid (w / w). In some embodiments, the one or more excipients comprise one or more of a hydrocolloid, a thickener, an antioxidant, a preservative, or a gelling agent.

[0013] In some embodiments, the topical formulation is or comprises an oil-in-water emulsion, a water-in-oil emulsion, a nanoemulsion, an emulsified gel, an oily ointment, a hydrocolloid system, or a cream preparation. In some embodiments, the topical formulation is or comprises an oil-in-water emulsion.

[0014] Provided herein is a method for treating a disorder or condition, comprising administering any of the topical formulations described herein to a subject in need thereof.In some embodiments, the subject exhibits or is at risk of exhibiting one or more symptoms associated with a skin condition or disorder.In some embodiments, the present application comprises topically applying the topical formulation to an area of ​​skin that exhibits or is at risk of exhibiting one or more symptoms.

[0015] In some embodiments, the skin condition or disorder comprises one or more of dermatitis, atopic dermatitis, contact dermatitis, hand dermatitis, allergic dermatitis, stasis dermatitis, asteatotic dermatitis, nummular dermatitis, seborrheic dermatitis, infectious dermatitis, acne, acne vulgaris, rosacea, epidermolysis bullosa simplex, chronic superficial scaly acrodermatitis, chondrodermatitis, perioral dermatitis, dermatomyositis, neutrophilic dermatosis, transient acantholytic dermatosis, eczema, ichthyosis, xerosis, asteatosis, psoriasis, lupus erythematosus, exfoliative keratolysis, impetigo, allergic inflammation of the skin, urticaria, lacerations, burns, or wounds resulting from surgical incision or biopsy.In some embodiments, the skin condition or disorder comprises psoriasis.In some embodiments, the skin condition or disorder comprises atopic dermatitis.

[0016] In certain embodiments, the one or more symptoms associated with a skin condition or disorder include one or more of dryness, lesions, cracks, fissures, scaling, discoloration, thickening, redness, bleeding, swelling, peeling, ulcers, pain, wounds, blistering, rashes, inflammation, infection, or loss of flesh. In some embodiments, administering the topical formulation includes applying the topical formulation topically or directly to an area of ​​skin having or at risk of having one or more symptoms.

[0017] Provided herein is a method for reducing or preventing inflammation in a subject in need thereof, comprising topically or directly applying a topical formulation described herein to an area of ​​the subject's skin or in the subject's mouth that is inflamed or at risk of becoming inflamed. In some embodiments, one or more symptoms associated with a skin condition or disorder include inflammation.

[0018] In some embodiments, provided herein is a method for treating a wound, comprising topically applying the topical formulation of any one of claims 1-29 to a subject in need thereof at the wound, optionally to promote healing or reduce the probability or extent of scarring.In some embodiments, the subject is experiencing or at risk of experiencing pain associated with breast feeding.In some embodiments, administering comprises applying the topical formulation to the subject's breast or nipple.

[0019] Provided herein is a method for reducing transepidermal water loss in a subject in need thereof, comprising applying a topical formulation as described herein to an area on the skin of the subject.In some embodiments, the subject exhibits or is at risk of exhibiting one or more symptoms associated with mucositis.In certain embodiments, the one or more symptoms associated with mucositis include one or more of pain, inflammation, or ulceration.In some embodiments, administering comprises oral administration of the topical formulation.

[0020] In some embodiments, provided herein is the use of a topical formulation in the treatment or prevention of one or more symptoms associated with a skin condition or disorder.In some embodiments, provided herein is the use of a topical formulation described herein in the manufacture of a medicament for the treatment or prevention of one or more symptoms associated with a skin condition or disorder.In some embodiments, provided herein is a method for improving the appearance of damaged or aged skin, comprising administering a topical formulation described herein to a subject in need thereof.

[0021] In some embodiments, administering the topical formulation comprises topically or directly applying the topical formulation to aged or damaged skin. In some embodiments, aged or damaged skin comprises one or more of skin inflammation, irritation, and erythema, wrinkles, skin blemishes, "marionette" lines, laugh lines, deep nasolabial folds, crow's feet, fine lines, vertical frown lines, horizontal forehead lines, loose thin / weak skin, dryness, dullness, loss of elasticity, lack of radiance, appearance of spider veins or red spots, exaggerated lines, and wrinkles.

[0022] Provided herein are methods of making a topical formulation that include formulating one or more human milk fractions with one or more excipients, e.g., pharma- ceutically acceptable excipients, where the one or more human milk fractions include one or more of human milk cream, human milk butter oil, human milk fat globule membrane solids, human skim milk, human milk permeate, and human milk retentate, where the human milk permeate and human milk retentate are obtained from filtration of human skim milk, thereby making the topical formulation.

[0023] In some embodiments, the human milk fraction is obtained from a pool of human milk collected from multiple human milk donors. In some embodiments, the pool of human milk comprises human milk collected from at least 5, 10, 25, 50, or 100 human milk donors. In some embodiments, one or more human milk fractions comprise human milk cream, or a subfraction obtained from or derived from human milk cream, such as human milk butter oil or milk fat globule membrane solids. In some embodiments, the topical formulation is formulated with about or at least 1%, 3%, 5%, 10%, or 20% human milk cream or human milk butter oil. In some embodiments, the topical formulation is formulated with less than 75%, 60%, or 50% human milk cream or human milk butter oil. In certain embodiments, the topical formulation is formulated with between 20% and 70% human milk cream or human milk butter oil. In certain embodiments, the human milk cream or butter oil is mixed in an amount sufficient to provide a final concentration of human milk fat, fatty acid, or lipid (w / w) of between 1% and 30%. In some embodiments, the amount of human milk cream or butter oil is sufficient to provide a final concentration of human milk fat, fatty acid, or lipid (w / w) of between 5% and 25%. In certain embodiments, the amount of human milk cream or butter oil is sufficient to provide a final concentration of human milk fat, fatty acid, or lipid (w / w) of about 20%. In certain embodiments, the amount of human milk cream or butter oil is sufficient to provide a final concentration of human milk fat, fatty acid, or lipid (w / w) of about 10%.

[0024] In some embodiments, the one or more human milk fractions include a human milk permeate obtained from filtration of human skim milk. In some embodiments, the filtration is or includes ultrafiltration. In some embodiments, the permeate includes at least 0.1%, 0.3%, 0.5%, or 1.0% human milk oligosaccharides (w / w). In some embodiments, prior to mixing the permeate with one or more excipients, the permeate undergoes one or more steps to concentrate the human milk oligosaccharide content to at least 0.5%, 1%, 3%, 5%, or 10% human oligosaccharides (w / w). In some embodiments, the one or more steps to concentrate the human milk oligosaccharide content include one or both of nanofiltration or reverse osmosis.

[0025] In some embodiments, prior to mixing the permeate with one or more excipients, the permeate undergoes one or more steps to reduce the lactose content. In some embodiments, the one or more steps to reduce the lactose content include (i) contacting the permeate with a lactase enzyme to produce a permeate / lactase mixture, (ii) incubating the permeate / lactase mixture at a temperature between 45° C. and 55° C., and (iii) filtering the permeate / lactase mixture to remove at least 90% of the lactase from the permeate, thereby reducing the lactose content of the permeate.

[0026] In some embodiments, the topical formulation is formulated with at least 5%, 10%, or 25% permeate obtained from ultrafiltration of human skim milk. In some embodiments, the topical formulation is formulated with less than 90%, 80%, or 75% permeate obtained from ultrafiltration of human skim milk. In some embodiments, the human milk permeate is mixed in an amount sufficient to provide a final concentration of at least 0.1% human milk oligosaccharides (w / w). In certain embodiments, the amount of human milk permeate is sufficient to provide a final concentration of at least 0.3% human milk oligosaccharides (w / w). In certain embodiments, the amount of human milk permeate is sufficient to provide a final concentration of human milk oligosaccharides (w / w) from 0.1% and 0.5%. In some embodiments, the amount of human milk permeate is sufficient to provide a final concentration of human milk oligosaccharides (w / w) from 0.3%. In certain embodiments, the topical formulation is formulated with 1%-60% human milk cream or human milk butter oil, and human milk permeate obtained from ultrafiltration of 25%-80% human skim milk. In certain embodiments, the topical formulation is formulated with 20%-60% human milk cream or butter oil, and human milk permeate obtained from ultrafiltration of 25%-80% human skim milk.

[0027] In some embodiments, the one or more excipients include one or more of a hydrocolloid, an emulsifier, a structuring agent, an antioxidant, a silicone-containing compound, a thickener, a preservative, an additional skin conditioning agent, an emollient, a humectant, or an essential oil. In some embodiments, the topical formulation is or includes an oil-in-water emulsion, a water-in-oil emulsion, a nanoemulsion, an emulsified gel, an oily ointment, a hydrocolloid system, or a cream preparation.

[0028] Provided herein are methods of making an oil-in-water emulsion comprising one or more human milk fractions, the methods comprising combining an aqueous phase and an oil phase to create an oil-in-water emulsion, where (i) the aqueous phase comprises water, human milk cream or a subfraction thereof, such as human milk butter oil, or a human milk fat globule membrane solid subfraction, and one or more excipients that are hydrophilic or soluble in water, and (ii) the oil phase comprises one or more excipients that are hydrophobic or soluble in oil, thereby creating an oil-in-water emulsion comprising one or more human milk fractions.

[0029] Provided herein are methods of making an oil-in-water emulsion comprising one or more human milk fractions, the methods comprising combining an aqueous phase and an oil phase to produce an oil-in-water emulsion, where (i) the aqueous phase comprises human milk permeate and one or more excipients that are hydrophilic or soluble in water, where the human milk permeate is obtained from ultrafiltration of human skim milk, and (ii) the oil phase comprises one or more excipients that are hydrophobic or soluble in oil, thereby producing an oil-in-water emulsion comprising one or more human milk fractions.

[0030] In some embodiments, the aqueous phase comprises human milk cream or a subfraction thereof, such as human milk butter oil. In some embodiments, one or more thickeners are added to the oil-in-water emulsion. In some embodiments, the human milk cream or a subfraction thereof is added to the oil-in-water emulsion after combining the aqueous phase and the oil phase. In some embodiments, the aqueous phase comprises one or more of an emulsifier, an antimicrobial agent, and a humectant. In some embodiments, the oil phase comprises one or more of an emollient, an emulsifier, and an antioxidant.

[0031] Provided herein is a method of making an oil-in-water emulsion comprising one or more human milk fractions, the method comprising combining an aqueous phase and an oil phase to produce an oil-in-water emulsion, where (i) the aqueous phase comprises an emulsifier, an antimicrobial agent, and a human milk permeate, the human milk permeate being obtained from ultrafiltration of human skim milk, and (ii) the oil phase comprises one or more excipients that are hydrophobic or soluble in oil, e.g., pharma- ceutically acceptable excipients, thereby producing an oil-in-water emulsion, adding human milk cream or a subfraction thereof to the oil-in-water emulsion, and adding a thickening agent to the oil-in-water emulsion, thereby producing an oil-in-water emulsion comprising one or more milk fractions. In some embodiments, the oil-in-water emulsion comprising one or more milk fractions comprises 3%-30% human milk fat, fatty acid, or lipid (w / v). In certain embodiments, the oil-in-water emulsion comprises about 20% human milk fat, fatty acid, or lipid (w / v).In some embodiments, the oil-in-water emulsion comprises about 10% human milk fat, fatty acid, or lipid (w / v). [Brief description of the drawings]

[0032] [Figure 1A] 1A and 1B illustrate a three-dimensional in vitro human skin model. A schematic of the cell culture process is shown in FIG. 1A. A stained section of the three-dimensional in vitro skin model with labels indicating the cell layers corresponding to human skin is shown in FIG. 1B. [Figure 1B] 1A and 1B illustrate a three-dimensional in vitro human skin model. A schematic of the cell culture process is shown in FIG. 1A. A stained section of the three-dimensional in vitro skin model with labels indicating the cell layers corresponding to human skin is shown in FIG. 1B. [Diagram 2] FIG. 2 shows skin barrier function as a percentage of transepithelial / transendothelial electrical resistance (TEER) measured in Ohms*cm2 at 1,000 Hz after incubation in a three-dimensional in vitro human skin model with vehicles containing 1% DMSO (D), human milk cream and permeate (PC), 30% cream (CI 30%), 1% human milk permeate (P 1%), or PBS with 30% human milk retentate (UF 30%). [Diagram 3] FIG. 3 shows the levels of IL-8 measured in the medium of three-dimensional human skin models after incubation in an untreated control group (NC) or after treatment with TNF-alpha and incubation in vehicles containing PBS with 1% DMSO (D), 1% human milk cream (CI 1%), 5% permeate (P 5%), or 1% human milk retentate (UF). [Figure 4] Figure 4 shows barrier function three-dimensional human skim model as a percentage of TEER compared to measurements collected from vehicle-only controls prior to mechanical disruption. Barrier function is shown before and immediately after, or 3, 5, or 8 days after mechanical disruption in cell culture models incubated in PBS containing 1% DMSO (D), human milk cream permeate combination (PC), or 30% skim milk (SM30%). [Diagram 5] FIG. 5 shows TEER measurements collected from three-dimensional skin models incubated in vehicle containing PBS with 1% DMSO (D) or 10% human milk retentate (10% UF) before, immediately after (After), and various time points thereafter following skin punch removal. [Figure 6A] Figures 6A-6C show levels of IL-8 measured in the medium of the three-dimensional human skin model in untreated controls (untreated) or after treatment with TNF-alpha and incubation for 24 hours (Figure 6A) and 72 hours (Figure 6B) with a vehicle oil-in-water emulsion or a positive control, an oil-in-water emulsion containing human milk cream, human milk permeate, or both human milk cream and permeate. IL-8 levels measured 48 hours after removal of the emulsion are shown in Figure 6C. [Figure 6B]Figures 6A-6C show levels of IL-8 measured in the medium of the three-dimensional human skin model in untreated controls (untreated) or after treatment with TNF-alpha and incubation for 24 hours (Figure 6A) and 72 hours (Figure 6B) with a vehicle oil-in-water emulsion or a positive control, an oil-in-water emulsion containing human milk cream, human milk permeate, or both human milk cream and permeate. IL-8 levels measured 48 hours after removal of the emulsion are shown in Figure 6C. [Figure 6C] Figures 6A-6C show levels of IL-8 measured in the medium of the three-dimensional human skin model in untreated controls (untreated) or after treatment with TNF-alpha and incubation for 24 hours (Figure 6A) and 72 hours (Figure 6B) with a vehicle oil-in-water emulsion or a positive control, an oil-in-water emulsion containing human milk cream, human milk permeate, or both human milk cream and permeate. IL-8 levels measured 48 hours after removal of the emulsion are shown in Figure 6C. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0033] Provided herein are compositions, such as topical formulations (such as ointments, skin creams, lotions, or emulsions) that contain or include milk (e.g., human milk, or fractions, portions, or components thereof). In some embodiments, the milk is human milk. In certain embodiments, the composition is or includes a topical formulation that contains a fraction of human milk. In certain embodiments, the composition is a topical formulation that is or includes one or more of human milk cream, human skim milk, or a permeate or retentate resulting from filtration of human skim milk (also referred to herein as human milk permeate or human milk retentate, respectively). Methods of use for the provided compositions to treat or prevent skin conditions or disorders are also provided, as well as methods of making compositions, such as topical formulations that contain one or more human milk fractions, portions, or components.

[0034] Currently, there are numerous treatments for skin conditions and disorders, ranging from special diets to emollients and immunosuppressive ointments, e.g., corticosteroid ointments. Corticosteroids such as hydrocortisone or clobetasol propionate (administered topically, orally, or intradermally) may provide improvement, but they often have side effects. Long-term use is thought to increase the risk of these side effects, the most common of which is thinning and fragility of the skin (atrophy). For this reason, when used on delicate skin, lower-strength steroids should be used or applied less frequently. In addition, high-strength steroids used over large areas or under occlusion may be absorbed into the body and may cause hypothalamic-pituitary-adrenal axis suppression (HPA axis suppression) and all the side effects normally associated with systemic steroid use.

[0035] Increased risk of skin infection by bacteria such as Staphylococcus aureus or fungi may result from skin conditions or disorders such as atopic dermatitis or eczema. Therefore, in more severe cases, dermatologists may also prescribe conventional antibiotics, either topically or orally, such as penicillin, streptomycin, and chloramphenicol. While the effectiveness of such treatments may vary from person to person, there are well-known disadvantages of conventional antibiotics, such as non-specificity, e.g., toxicity to non-pathogenic or beneficial bacteria, and the risk of resistance by not only the target bacteria but also other pathogenic bacteria. Moreover, conventional systemic antibiotic treatments may interact with other drugs, and certain antibiotics cannot be combined with the use of alcohol. As a result, there is a need for skin treatments that avoid these disadvantages while promoting effective wound healing and / or reducing inflammation.

[0036] The provided compositions address these needs. In some embodiments, the provided compositions improve barrier function and promote healing and regeneration and / or reduce inflammation in human skin with a high degree of efficacy. In certain embodiments, the provided compositions contain natural human lipids, oligosaccharides, or proteins, and thus reduce the possibility of sensitization, irritation, or allergy, for example, compared to skin treatments that contain active ingredients of non-human origin. In certain embodiments, the provided compositions reduce symptoms associated with inflammation while also strengthening the skin, such as by improving barrier function.

[0037] Human milk contains a complex mixture of bioactive components, including lipids, vitamins, amino acids, oligosaccharides, sugars, cofactors, nucleotides, amino acids, and energy intermediates, each of which may benefit aspects of skin health. The provided compositions include formulations that incorporate some or all of these components into a formulation, such as topical formulations that allow for topical and / or direct application to human skin. Unlike currently available skin treatments, the provided compositions may contain natural human lipids, proteins, and oligosaccharides that support skin health while minimizing adverse effects. For example, in some embodiments, the human origin of the active ingredients reduces the probability or likelihood of adverse effects resulting from sensitization or allergic reactions.

[0038] Provided compositions are or include topical formulations that can be applied to damaged skin, for example, areas that exhibit one or more symptoms associated with skin disease or disorder.In some embodiments, provided compositions are particularly useful for reducing skin inflammation while also strengthening skin barrier function.This effect is more beneficial than alternative skin treatments, such as corticosteroids, which may reduce inflammation but may weaken barrier function in some embodiments.

[0039] In addition to enhancing immune function in infants, human milk also contains antimicrobial and antibacterial properties. For example, in certain embodiments, human milk may contain antibodies and oligosaccharides that can kill or prevent the growth of pathogenic bacteria or otherwise promote a healthy skin microbiome. In some embodiments, antimicrobial human milk components are formulated into stable compositions that may be administered or applied, for example, topically, to a subject to treat or prevent infection without any of the undesirable side effects associated with acute or long-term treatment with antibiotics.

[0040] In certain aspects, sensitization may be a concern with some existing skin treatments. In some aspects, skin sensitization may manifest as an allergic response to a substance after skin contact, such as in the form of allergic contact dermatitis (ACD). By some estimates, up to about 20% of the general population is sensitive to at least one allergen (Thyssen et al., Contact Dermatitis 2007;57:287-299). ACD is a clinical condition caused by an allergic immune response after skin exposure to a large subset of small reactive chemicals (haptens) that can be found in the environment and in many household products. In certain embodiments, the compositions and formulations provided are formulated or composed of proteins, lipids, oligosaccharides, or other macromolecules of natural and human origin. In certain aspects, such formulations reduce the potential for sensitization and irritation to the skin compared to alternative skin treatments, especially those that contain natural plant extracts and / or non-human animal-based ingredients.

[0041] In certain aspects, the topical formulations provided contain one or more components, constituents, parts, or fractions derived from human milk. In some aspects, the topical formulations have advantages over alternative topical formulations that may be present in human milk but contain certain compounds, such as lipids, fats, fatty acids, proteins, or oligosaccharides, that are synthetic or derived from non-human milk sources. Such advantages may include anti-inflammatory activity or promotion of skin repair to a greater extent than may be observed from these alternative topical formulations. In some such embodiments, combinations of compounds, such as (but not limited to) lipids, fats, fatty acids, proteins, and / or oligosaccharides, found in topical formulations derived from or formulated with human milk fractions or components contain various human milk compounds that may act synergistically to provide such advantages, e.g., anti-inflammatory or skin repair, over alternative topical formulations.

[0042] In some aspects, the provided compositions, e.g., topical formulations, overcome the difficulties that may be associated with formulating compositions containing creams derived from human milk. For example, in some aspects, compositions containing human milk creams may be prone to spoilage, phase separation, denaturation, or discoloration (e.g., discoloration indicating degradation of macromolecules, proteins, oligosaccharides, or lipids) when prepared in typical emulsions, e.g., those containing excipients that may be suitable for formulating creams or lipids from other sources, e.g., bovine or vegetable sources. In certain embodiments, the provided compositions and methods overcome these problems by using specific formulations, e.g., oil-in-water emulsions, lotions, creams, or ointments, etc., tailored to stabilize compositions containing human milk creams. Such formulations result in stable formulations that do not spoil, discolor, or phase separate over weeks, months, or even years.

[0043] All publications, including patent documents, scientific articles, and databases, referred to in this application are incorporated by reference in their entirety for all purposes to the same extent as if each individual publication was individually incorporated by reference. To the extent that the definitions set forth herein are contrary to or otherwise inconsistent with the definitions set forth in the patents, applications, published applications, and other publications incorporated herein by reference, the definitions set forth herein take precedence over the definitions incorporated herein by reference.

[0044] The section headings used herein are for organizational purposes only and are not to be construed as limiting the subject matter described. I. Topical Formulations Containing Human Milk Fractions, Portions, or Constituents

[0045] Provided herein are compositions that are or include topical formulations that may be applied topically and / or directly to the skin or mucosa. In some embodiments, the topical formulations may be applied to the skin for cosmetic purposes. In certain embodiments, the topical formulations may be applied topically and / or directly to the skin or mucosa to treat or prevent a disease or disorder. In certain embodiments, the topical formulations include at least one component derived from or from cow's milk. In some embodiments, the topical formulations include one or more of parts, fractions, or components derived from or from human milk. In certain embodiments, milk, or parts, fractions, or components thereof, are formulated into a topical formulation, such as by mixing, combining, or formulating with one or more pharma- ceutically acceptable excipients.

[0046] In some embodiments, the milk is mammalian milk. In certain embodiments, the milk is or comprises cow's milk, donkey's milk, goat's milk, sheep's milk, camel's milk, mare's milk, or buffalo's milk. In some embodiments, the milk is or comprises cow's milk. In certain embodiments, the milk is or comprises camel's milk. In various embodiments, the topical formulation comprises one or more components originating from or derived from human milk. In certain embodiments, the topical formulation comprises human milk, or a part, fraction, or component thereof.

[0047] In certain embodiments, the topical formulation is or comprises human whole milk. In some embodiments, the topical formulation is or comprises at least 1%, 5%, 10%, 20%, 25%, 30%, 40%, 50%, 60%, 70%, 75%, or 80% or more human whole milk. In certain embodiments, the topical formulation contains no more than 75%, 70%, 60%, 50%, 40%, 30%, 25%, 20%, or 10% human whole milk. In some embodiments, the topical formulation contains, for example, by weight / weight (w / w) or weight / volume (w / v), between 1% and 75%, 5% to 50%, 10% to 50%, 10% to 30%, 10% to 20%, 30% to 50%, 40% to 60%, or 50% to 75% or about 1% to 75%, 5% to 50%, 10% to 50%, 10% to 30%, 10% to 20%, 30% to 50%, 40% to 60%, or 50% to 75% human whole milk. Human milk cream

[0048] In some embodiments, the topical formulation is formulated with a cream, such as human milk cream. In some embodiments, the topical formulation is or comprises a cream (e.g., cream obtained from or isolated from whole milk). In certain embodiments, the topical formulation is or comprises a human milk cream (also referred to herein as human cream or cream derived from human milk). In some embodiments, the composition contains a portion or fraction of human milk cream (also referred to herein as a subfraction of human milk cream) (e.g., a portion or component of human milk cream isolated or enriched from human milk cream).

[0049] In some embodiments, the cream, e.g., human milk cream, is obtained from a method or technique for separating human milk cream from human skim milk. Certain embodiments contemplate that the method for separating cream from skim may be performed routinely by any method known in the art. In some embodiments, the cream is separated from the skim by a method or technique that is or includes creaming separation or centrifugation, such as any of those described herein, e.g., in Section III. In some embodiments, the human milk cream is or includes a human milk fraction containing 20%-60% fat w / v or a high concentration of fat w / w, e.g., separated from a human skim milk fraction of human whole milk.

[0050] In certain embodiments, human milk cream contains only human milk components, such as lipids, fatty acids, fats, carbohydrates, sugars, oligosaccharides, or proteins (concentrated human milk, isolated human milk, or derived from, produced from, or found in human milk). In certain embodiments, human milk cream is free or essentially free of non-human lipids, fatty acids, fats, carbohydrates, sugars, oligosaccharides, or proteins (e.g., lipids, fatty acids, fats, carbohydrates, sugars, oligosaccharides, or proteins that are not concentrated human milk, not isolated human milk, or not derived from, produced from, or found in human milk). In certain embodiments, human milk cream is free or essentially free of components or compounds, such as, but not limited to, xenogenous lipids, fatty acids, fats, carbohydrates, sugars, oligosaccharides, or proteins.

[0051] In some embodiments, human milk cream is or comprises, e.g., as a weight / weight (w / w) percentage, about 20%-60% human milk lipids, fats, or fatty acids. In various embodiments, human milk cream is a milk fraction separated from whole milk by or during a process to produce human skim milk from whole human milk, e.g., by centrifugation, and in certain embodiments contains at or about 20%-60% human milk lipids, fats, or fatty acids (e.g., w / w). In some embodiments, human milk cream is or comprises 30%-70%, 40%-65%, 45%-65%, 48%-62%, or 50%-60% lipids, fats, or fatty acids (e.g., w / w).

[0052] In certain embodiments, the topical formulation is, comprises, or is formulated with at least 1%, 5%, 10%, 20%, 25%, 30%, 40%, 50%, 60%, 70%, 75%, or 80% or more human milk cream (e.g., w / w or v / w). In certain embodiments, the topical formulation is, or comprises, no more than 75%, 70%, 60%, 50%, 40%, 30%, 25%, 20%, or 10% human milk cream. In some embodiments, the topical formulation contains at or about 1%-75%, 5%-50%, 10%-50%, 10%-30%, 10%-20%, 30%-50%, 40%-60%, or 50%-75% (inclusive) human milk cream.

[0053] In certain embodiments, the topical formulation contains 10%-30%, or about 10%-30% human milk cream (e.g., w / v or v / v). In certain embodiments, the topical formulation contains 50%-75%, or about 50%-75% human milk cream. In certain embodiments, the topical formulation contains 10%, or about 10% human milk cream. In some embodiments, the topical formulation contains 30%, or about 30% human milk cream. In certain embodiments, the topical formulation contains 50%, or about 50% human milk cream.

[0054] In certain embodiments, an amount of human milk cream is mixed, added, or incorporated into the topical formulation. In certain embodiments, the amount is sufficient to achieve a target amount or content of human milk lipids, fats, or fatty acids. In certain embodiments, the human milk cream is mixed, added, or incorporated into the topical formulation in an amount sufficient to achieve a topical formulation containing at least 1%, 2%, 3%, 4%, 5%, 10%, 15%, 20%, 25%, or 30%, or 1%-40%, 5%-30%, 10%-30%, 5%-15%, or 7.5%-12.5% ​​(each inclusive, e.g., w / w or w / v) of human milk lipid, fat, or fatty acid content. In some embodiments, an amount of human milk cream is mixed, added, or incorporated into the topical formulation to achieve a topical formulation containing about 10% human milk fat (e.g., w / w or w / v). In certain embodiments, an amount of human milk cream is mixed, added, or incorporated to achieve a topical formulation containing about 20% human milk fat (e.g., w / w or w / v). In certain embodiments, an amount of human milk cream is mixed, added, or incorporated to achieve a topical formulation containing about 30% human milk fat (e.g., w / w or w / v).

[0055] In some embodiments, the topical formulation is free or essentially free of non-human lipids, hi certain embodiments, the topical formulation is free or essentially free of mammalian milk lipids that are not human milk lipids.

[0056] In certain embodiments, the topical formulation contains or comprises one or more milk lipids, fats, or fatty acids. In certain embodiments, the topical formulation contains or comprises at least 1%, 2%, 3%, 4%, 5%, 10%, 15%, 20%, 25%, 30%, 35%, or 40% human milk lipids (e.g., w / w or w / v). In certain embodiments, the topical formulation contains or comprises at least 1%, 2%, 3%, 4%, 5%, 10%, 15%, 20%, 25%, 30%, 35%, or 40% human milk fats (e.g., w / w or w / v). In certain embodiments, the topical formulation contains or comprises at least 1%, 2%, 3%, 4%, 5%, 10%, 15%, 20%, 25%, 30%, 35%, or 40% human milk fatty acids (e.g., w / w or w / v). In certain embodiments, the topical formulation contains 1%-60%, 1%-40%, 25%-50%, 5%-30%, 10%-30%, 5%-15%, or 7.5%-12.5% ​​(each inclusive, e.g., w / w or w / v) human milk lipids, lipids, or fatty acids. In some embodiments, the topical formulation contains 1%-60%, 25%-50%, 5%-30%, 5%-15%, or 7.5%-12.5% ​​(each inclusive, e.g., w / w or w / v) human milk fats, lipids, or fatty acids.

[0057] In certain embodiments, the topical formulation contains one or more human milk lipids, such as lipids found in or derived from human milk. In some embodiments, the topical formulation contains one or more neutral complex lipids (e.g., free fatty acids, cholesteryl esters, diacylglycerols, triacylglycerols, monoacylglycerols), sphingolipids (e.g., ceramides, dihydroceramides, hexosylceramides, lactosylceramides, sphingomyelins), phospholipids (e.g., phosphatidylcholine, lysophosphatidylcholine, phosphatidylethanolamines, lysophophatidylethanolamines, phosphatidylinositol), or a combination of any or all of the foregoing, which are found in or derived from human milk.

[0058] In certain embodiments, the topical formulation contains one or more human lipids found in or derived from the human stratum corneum. In some embodiments, the composition, e.g., the topical composition, contains one or more neutral complex lipids (e.g., free fatty acids, cholesteryl esters, diacylglycerols, triacylglycerols, monoacylglycerols), sphingolipids (e.g., ceramides, dihydroceramides, hexosylceramides, lactosylceramides, sphingomyelins), phospholipids (e.g., phosphatidylcholine, lysophosphatidylcholine, phosphatidylethanolamines, lysophophatidylethanolamines, phosphatidylinositol), or a combination of any or all of the foregoing, which are found in the human stratum corneum or derived from human milk.

[0059] In certain embodiments, topical formulations contain one or more fatty acids that are concentrated from human milk, isolated from human milk, or derived from human milk, or are found in human milk or human milk cream.In some embodiments, human milk oligosaccharides include, but are not limited to, oleic acid, palmitic acid, linoleic acid, omega-3 fatty acid, omega-6 fatty acid, omega-9 fatty acid, stearic acid, myristic acid, lauric acid, palmitoleic acid, vaccenic acid, myristolic acid, heptadecenoic acid, gamma-linolenic acid, docosapentaenoic acid, eicosapentaenoic acid (EPA), arachidonic acid (AA), and docosahexaenoic acid (DHA).

[0060] In certain embodiments, the topical formulation is or comprises at least 1%, 2%, 3%, 4%, 5%, 10%, 15%, 20%, 25%, or 30% human milk lipids, fats, or fatty acids. In some embodiments, the topical formulation is or comprises at least 1%, 2%, 3%, 4%, 5%, 10%, 15%, 20%, 25%, or 30% human milk fat. In certain embodiments, the topical formulation is or comprises 20%-50%, 1%-40%, 5%-30%, 10%-30%, 5%-15%, or 7.5%-12.5% ​​(inclusive, e.g., w / w or w / v) human milk lipids, fats, or fatty acids. In some embodiments, the topical formulation is or comprises 5%-30%, 5%-15%, or 7.5%-12.5% ​​(e.g., w / w) human milk fat. In certain embodiments, the topical formulation is or comprises about 10% (e.g., w / w) human milk fat. In certain embodiments, the topical formulation is or comprises about 20% (e.g., w / w) human milk fat. In certain embodiments, the topical formulation is or comprises about 30% (e.g., w / w) human milk fat.

[0061] In some embodiments, the topical formulation is formulated with a subfraction obtained from or derived from human milk cream. In certain embodiments, the topical formulation is formulated with human milk butter oil. In certain embodiments, the human milk butter oil results from removing some or all of the milk solids and water from human milk cream. In certain embodiments, the butter oil is anhydrous milk fat. In some embodiments, the human milk butter oil is a fraction of human milk that contains at least 80%, 85%, 90%, 95%, 98%, 99%, or 99.8% human milk fat, fatty acid, or lipid. Certain embodiments contemplate that the butter oil subfraction of human milk cream may be obtained from or derived from human milk cream by any known suitable method as a routine procedure.

[0062] In certain embodiments, the topical formulation is, comprises, or is formulated with at least 1%, 5%, 10%, 20%, 25%, 30%, 40%, 50%, 60%, 70%, 75%, or 80% or more human milk butter oil (e.g., w / w or w / v). In certain embodiments, the topical formulation is, comprises, or contains no more than 75%, 70%, 60%, 50%, 40%, 30%, 25%, 20%, or 10% human milk butter oil. In some embodiments, the topical formulation contains 1%-60%, 10%-30%, 30%-60%, or 20%-40% (w / w or w / v) human milk butter oil.

[0063] In some embodiments, an amount of human milk butter oil is mixed, added, or incorporated into the topical formulation. In certain embodiments, the amount is sufficient to achieve a target amount or content of human milk lipids, fats, or fatty acids. In certain embodiments, butter oil is mixed, added, or incorporated into the topical formulation in an amount sufficient to achieve a final concentration of about or at least 1%, 5%, 10%, 20%, 30%, or 40%, or 5%-50% or 10%-30% (e.g., w / w or w / v) human milk lipids, fats, or fatty acids. In certain embodiments, human milk butter oil is added to achieve a topical formulation containing about 10% human milk fat. In certain embodiments, human milk butter oil is added to achieve a topical formulation containing about 20% human milk fat. In some embodiments, human milk butter oil is added to achieve a topical formulation containing about 30% human milk fat.

[0064] In some embodiments, the topical formulation is formulated with a sub-fraction of human milk cream that is or comprises human milk fat globule membrane solids. In some embodiments, the human milk fat globule membrane solids sub-fraction is obtained by any suitable known means, including, but not limited to, the methods described herein, for example, in Section III and / or Example 17.

[0065] In certain embodiments, the topical formulation comprises or is formulated with at least 1%, 5%, 10%, 20%, 25%, 30%, 40%, 50%, 60%, 70%, 75%, or 80% (w / w or w / v) or more of human milk fat globule membrane solids obtained from or derived from human milk cream. In certain embodiments, the topical formulation is or comprises no more than 75%, 70%, 60%, 50%, 40%, 30%, 25%, 20%, or 10% of human milk fat globule membrane solids obtained from or derived from human milk cream. In some embodiments, the topical formulation contains 1%-60%, 1-40%, 10%-30%, 30%-60%, or 20%-40% (w / w or w / v).

[0066] In some embodiments, an amount of human milk fat globule membrane solids derived or obtained from human milk cream, for example by centrifugation, is mixed, added, or incorporated into a topical formulation. In certain embodiments, the amount is sufficient to achieve a target amount or content of human milk lipids, fats, or fatty acids. In certain embodiments, human milk fat globule membrane solids are mixed, added, or incorporated into a topical formulation in an amount sufficient to achieve a final concentration of about or at least 1%, 5%, 10%, 20%, 30%, or 40%, or 5%-50% or 10%-30% (e.g., w / w or w / v) of human milk lipids, fats, or fatty acids. Human skim milk

[0067] In some embodiments, topical formulations are formulated with skim milk.In certain embodiments, topical formulations are formulated with human skim milk.In certain embodiments, skim milk is obtained from a method or technique that is or includes centrifugation or creaming separation.In some embodiments, skim can be further processed, for example, by additional centrifugation, to further remove fat globules, lipids, and / or fatty acids from skim.

[0068] In some embodiments, the topical formulation contains human skim milk. In certain embodiments, the topical formulation contains 1%, 5%, 10%, 20%, 25%, 30%, 40%, 50%, 60%, or 70% (e.g., w / w or w / v) or more of human skim milk, or contains about 1%, 5%, 10%, 20%, 25%, 30%, 40%, 50%, 60%, or 70% (e.g., w / w or w / v) or more of human skim milk, or contains at least 1%, 5%, 10%, 20%, 25%, 30%, 40%, 50%, 60%, or 70% (e.g., w / w or w / v) or more of human skim milk. In certain embodiments, the topical formulation contains 75%, 70%, 60%, 50%, 40%, 30%, 25%, 20%, or 10% or less of human skim milk. In some embodiments, the topical formulation contains at or about 1%-75%, 5%-50%, or 10%-50% (each of the extremes inclusive, e.g., w / w or w / v) human skim milk.

[0069] In certain embodiments, the topical formulation is formulated as a standard formulation, e.g., a typical, standard, or art-known formulation such as an emulsion, gel, or lotion, in which all or a portion of the water or water-containing buffer solution (such as saline or PBS) typically used to formulate topical formulations is replaced with skim milk. Human milk permeate

[0070] In some embodiments, the topical formulation contains a permeate fraction of milk. In certain embodiments, the permeate is derived from the filtration, e.g., ultrafiltration, of skim milk. In some embodiments, the permeate is or comprises the portion of skim milk that passes through a filter, e.g., an ultrafilter. In some embodiments, the permeate is enriched in milk oligosaccharides, e.g., enriched relative to whole milk.

[0071] In certain embodiments, the topical formulation contains a permeate fraction of human milk (also referred to herein as "human milk permeate" or "permeate"). In some embodiments, the human milk permeate results from filtration, e.g., ultrafiltration, of human skim milk. In certain embodiments, the permeate is or includes the portion of skim milk that passes through a filter, e.g., an ultrafilter. In some embodiments, the permeate is enriched in human milk oligosaccharides (HMOs), e.g., enriched relative to whole human milk.

[0072] In some embodiments, the topical formulation comprises human milk permeate. In certain embodiments, the topical formulation comprises or comprises about 1%, 5%, 10%, 20%, 25%, 30%, 40%, 50%, 60%, or 70% (e.g., w / w or w / v) or more human milk permeate, or comprises at least 1%, 5%, 10%, 20%, 25%, 30%, 40%, 50%, 60%, or 70% (e.g., w / w or w / v) or more human milk permeate. In some embodiments, the topical formulation comprises, contains, or is formulated with 50%-95%, 60%-90%, or 60%-80% human milk permeate. In certain embodiments, the topical formulation comprises, contains, or is formulated with about or at least 50%, 60%, or 70%, or 50%-75%, 60%-80%, or 65%-75% (each of the extremes inclusive, e.g., w / w or w / v) human milk permeate.

[0073] In certain embodiments, the topical formulation contains a fragment or portion derived from or derived from human milk permeate. In some embodiments, the permeate, or a fragment or portion thereof, is, contains, or is enriched for one or more HMOs. In certain embodiments, the human milk permeate, or a fraction or portion thereof, is, or contains one or more cofactors, vitamins, amino acids, carbohydrates, sugars, and / or nucleosides present in or derived from human milk. In some embodiments, the permeate, or a fraction or portion thereof, is enriched for one or more compounds present in human milk or human milk permeate. Such compounds may include, but are not limited to, proteins (e.g., low molecular weight proteins), cofactors, vitamins, amino acids, carbohydrates, sugars, polynucleotides, oligonucleotides (e.g., human milk oligosaccharides).

[0074] In certain embodiments, the human milk permeate, or a fraction or portion thereof, is or comprises a concentration of at least 0.1%, 0.2%, 0.3%, 0.4%, 0.5%, 0.75%, or 1% (e.g., w / w) human milk oligosaccharides. In certain embodiments, the human milk permeate is added to, mixed with, or formulated with, or arrives in a topical formulation that comprises or contains at least 0.1%, 0.2%, 0.3%, 0.4%, 0.5%, 0.75%, or 1% (e.g., w / w) human milk oligosaccharides. In certain embodiments, the permeate comprises or contains at least 0.5% (e.g., w / w) HMOs prior to incorporation into the topical formulation.

[0075] In certain embodiments, topical formulations contain, include, or are formulated with relatively small amounts of human milk permeate.In certain embodiments, topical formulations contain, include, or are formulated with at least 0.001%, 0.001%, 0.01%, 0.1%, 0.5%, 1.0%, 2.0%, 2.5%, 5.0%, 7.5%, or 10%, or human milk permeate, or a fraction or part thereof.In certain embodiments, topical formulations contain 20% or less, 10% or less, 5.0% or less, 2.5% or less, 1.0% or less, or 0.5% or less of human milk permeate, or a fraction or part thereof. In some embodiments, the topical formulation contains between 0.001% and 20%, between 0.01% and 5%, or between 0.1% and 2% (each inclusive) of human milk permeate at or about 0.001% and 20%, between 0.01% and 5%, or between 0.1% and 2% (each inclusive).

[0076] In certain embodiments, topical formulations are formulated with permeate instead of water.Thus, in some embodiments, formulations such as emulsions, gels, ointments, etc. that are typically formulated with water are formulated with human milk permeate instead of water.Thus, in some embodiments, topical formulations are or comprise at least 30%, 40%, 50%, 60%, 70%, 75%, 80%, or 90% permeate.

[0077] In some embodiments, the human milk permeate, or a fraction or portion thereof, is further treated or processed before being added, mixed, or formulated into a topical formulation. In certain embodiments, the permeate is treated to remove one or more substances, such as by filtration and / or by the addition of an enzyme, e.g., lactase, to remove or digest undesirable compounds, e.g., sugars, such as lactose. In some embodiments, the permeate is further treated to increase the concentration of HMOs, such as by filtration, e.g., ultrafiltration or nanofiltration, before being added, mixed, or formulated into a topical formulation.

[0078] In certain embodiments, human milk permeate is processed before its incorporation into topical formulation. Such processing steps may function to enrich or increase the concentration of human milk oligosaccharides and / or to reduce the content or concentration of one or more sugars, such as lactose. In some embodiments, human milk permeate is processed into a fraction or part of human milk permeate that is or contains an increased concentration of HMOs, for example, relative to human whole milk or unprocessed human milk permeate, and / or into a reduced content or concentration of one or more sugars, such as lactose, for example, relative to human whole milk or unprocessed human milk permeate.

[0079] In some embodiments, the permeate is processed prior to its incorporation into the topical formulation to contain less than about 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, or 1% w / w lactose. In certain embodiments, the permeate is processed to contain less than 2% or 1% lactose. In certain embodiments, the permeate is processed to contain an HMO concentration or total HMO concentration of at least 0.5%, 1.0%, 2.0%, 3.0%, 4.0%, 5.0%, 7.5%, or 10%, 15%, or 20%, or 1.0%-2.0%, 2.0%-4.0%, 4.0%-5.0%, 1.0%-5.0%, 5.0%-10.0%, or 5.0%-7.5% w / v HMO (each inclusive).

[0080] In some embodiments, the permeate has or comprises an HMO profile that is substantially similar, both structurally and functionally, to the profile of HMOs observed across a population of whole human milk, i.e., because in some aspects the permeate may be obtained from a source of human milk derived from a pool of donors rather than an individual donor, the array of HMOs will be more diverse than in any one typical individual and will represent or more closely represent the spectrum of HMOs found in human milk as opposed to the spectrum of HMOs found or typically found in human milk produced by any particular individual.

[0081] In some embodiments, the permeate contains an amount of different individual HMOs that is greater than the number of different individual HMOs found in human milk from an individual donor. In certain embodiments, the permeate contains at least 1, 2, 3, 4, 5, 10, 15, 20, 25, 30, 35, 40, 45, or 50 more different individual HMOs than the number of different individual HMOs found in human milk from an individual donor. In certain embodiments, the permeate contains an amount of different individual HMOs that is greater than the average or median number of different individual HMOs found in multiple human milk samples from individual donors.

[0082] In some embodiments, one of the largest variables of HMO diversity comes from the mother's Lewis blood type, specifically whether she has an active fucosyltransferase 2 (FUT2) and / or fucosyltransferase 3 (FUT3) gene. When an active FUT2 gene is present, α1-2 linked fucose is generated, whereas fucose residues are α1-4 linked when the FUT3 gene is active. The result of this "secretor status" is that "secretors" (i.e., those with an active FUT2 gene) generally produce a much more diverse profile of HMOs dominated by α1-2 linked oligosaccharides, while "non-secretors" (i.e., those without an active FUT2 gene) may, for example, contain a more diverse array of α1,-4 linked oligosaccharides (compared to secretors), but may contain an overall reduction in diversity due to the inability of non-secretors to synthesize major components of the secretor's HMO repertoire. In some embodiments, the human milk permeate comprises human milk oligosaccharides comprising alpha 1-2 linked fucose and human milk oligosaccharides comprising alpha 1-4 linked fucose. In certain embodiments, the topical formulation comprises human milk oligosaccharides comprising alpha 1-2 linked fucose and human milk oligosaccharides comprising alpha 1-4 linked fucose.

[0083] In some embodiments, the topical formulation contains one or more HMOs. In certain embodiments, the topical formulation is or comprises a plurality of HMOs, for example, a plurality of about 10, 25, 50, 75, 100, or 150, or at least 10, 25, 50, 75, 100, or 150 HMOs. In some embodiments, the topical formulation comprises some or all of 2'-fucosyl-lactose, 3-fucosyl-lactose, 3'-sialyl-lactose, 6'-sialyl-lactose, lacto-N-tetraose, lacto-N-difucohexaose I, lactodifucotetraose, lacto-N-fucopentaose I, sialylacto-N-tetraose c, sialylacto-N-tetraose b, and disialylacto-N-tetraose. In certain embodiments, the topical formulation comprises all of 2'-fucosyl-lactose, 3-fucosyl-lactose, 3'-sialyl-lactose, 6'-sialyl-lactose, lacto-N-tetraose, lacto-N-difucohexaose I, lactodifucotetraose, lacto-N-fucopentaose I, sialyllacto-N-tetraose c, sialyllacto-N-tetraose b, and disialyllacto-N-tetraose.

[0084] In certain embodiments, the topical formulation is selected from the group consisting of 2'-fucosyl-lactose, 3-fucosyl-lactose, 3'-sialyl-lactose, 6'-sialyl-lactose, lacto-N-tetraose, lacto-N-neo-tetraose, lacto-N-fucopentaose I, lacto-N-fucopentaose II, lacto-N-fucopentaose III, sialyl-lacto-N-tetraose a, sialyl-lacto-N-tetraose b, sialyl-lacto-N-tetraose c, lacto-N-difuco-hexaose I, lacto-N-difuco-hexaose II, lacto-N-hexaose, para-lacto-N-hexaose, disialyllacto-N-tetraose, fucosyl-lacto-N-hexaose, difucosyl-lacto-N-hexaose a, difucosyl-lacto-N-hexaose b, lactodifucotetraose (LD), 6'galactosyllactose, 3'galactosyllactose, 3-sialyl-3-fucosyllactose, sialylfucosyllactose-N-tetraose, sialyllacto-N-fucopentaose V, disialyl-lacto-N-fucopentaose II, disialyl-lacto-N-fucopentaose V, lacto-N-neo-difucohexaose II, 3-fucosyl-sialylacto-N-tetraose c, para-lacto-N-neohexaose, lacto-N-octaose, lacto-N-neooctaose, lacto-N-neohexaose, lacto-N-fucopentaose V, iso-lacto-N-octaose, para-lacto-N-octaose, lactodecaose, and sialyl-lacto-N-fucopentaose I.In certain embodiments, the topical formulation is selected from the group consisting of 2'-fucosyl-lactose, 3-fucosyl-lactose, 3'-sialyl-lactose, 6'-sialyl-lactose, lacto-N-tetraose, lacto-N-neo-tetraose, lacto-N-fucopentaose I, lacto-N-fucopentaose II, lacto-N-fucopentaose III, sialyl-lacto-N-tetraose a, sialyl-lacto-N-tetraose b, sialyl-lacto-N-tetraose c, lacto-N-difuco-hexaose I, lacto-N-difuco-hexaose II, lacto-N-hexaose, para-lacto-N-hexaose, disialyllacto-N-tetraose, fucosyl-lacto-N-hexaose, difucosyl-lacto-N-hexaose a, difucosyl-lacto-N-hexaose and all of sialyl-3-fucosyllactose, sialylfucosyllactose-N-tetraose, sialyllacto-N-fucopentaose V, disialyl-lacto-N-fucopentaose II, disialyl-lacto-N-fucopentaose V, lacto-N-neo-difucohexaose II, 3-fucosyl-sialylacto-N-tetraose c, para-lacto-N-neohexaose, lacto-N-octaose, lacto-N-neooctaose, lacto-N-neohexaose, lacto-N-fucopentaose V, iso-lacto-N-octaose, para-lacto-N-octaose, lactodecaose, and sialyl-lacto-N-fucopentaose I.

[0085] In certain embodiments, the topical formulation comprises or contains at least 10, 25, 50, 100, or 150 HMOs, including all of 2'-fucosyl-lactose, 3-fucosyl-lactose, 3'-sialyl-lactose, 6'-sialyl-lactose, lacto-N-tetraose, lacto-N-difucohexaose I, lactodifucotetraose, lacto-N-fucopentaose I, sialylacto-N-tetraose c, sialylacto-N-tetraose b, and disialyllacto-N-tetraose. In certain embodiments, the topical formulation comprises 2'-fucosyl-lactose, 3-fucosyl-lactose, 3'-sialyl-lactose, 6'-sialyl-lactose, lacto-N-tetraose, lacto-N-neo-tetraose, lacto-N-fucopentaose I, lacto-N-fucopentaose II, lacto-N-fucopentaose III, sialyl-lacto-N-tetraose a, sialyl-lacto-N-tetrose ... Laose b, Sialyl-lacto-N-tetraose c, Lacto-N-difuco-hexaose I, Lacto-N-difuco-hexaose II, Lacto-N-hexaose, Para-lacto-N-hexaose, Disialyllacto-N-tetraose, Fucosyl-lacto-N-hexaose, Difucosyl-lacto-N-hexaose a, Difucosyl-lacto-N-hexaose b, Lactodifucotetraose (LD) , 6'galactosyl lactose, 3'galactosyl lactose, 3-sialyl-3-fucosyllactose, sialylfucosyllactose-N-tetraose, sialyl lacto-N-fucopentaose V, disialyl-lacto-N-fucopentaose II, disialyl-lacto-N-fucopentaose V, lacto-N-neo-difucohexaose II, 3-fucosyl-sialyllacto-N-tetraose c, para- It comprises or contains at least 25, 50, 100, or 150 HMOs, including all of lacto-N-neohexaose, lacto-N-octaose, lacto-N-neooctaose, lacto-N-neohexaose, lacto-N-fucopentaose V, iso-lacto-N-octaose, para-lacto-N-octaose, lactodecaose, and sialyl-lacto-N-fucopentaose I.

[0086] In certain embodiments, the topical formulation is or comprises at least 10, 25, 50, 75, 100, 125, 150 different HMOs found, present, or detected in pooled human milk (e.g., pooled from the milk of at least 10, 25, 50, or 100 individual donors) or in the permeate obtained from the pooled human milk (e.g., the permeate resulting from ultrafiltered human milk skim). In some embodiments, the topical formulation comprises at least 50%, 60%, 70%, 80%, 85%, 90%, 95%, 97%, 99%, or 99.9% different HMOs found, present, or detected in the pooled human milk or in the permeate obtained from the pooled human milk. In certain embodiments, the topical formulation comprises at least 50%, 60%, 70%, 80%, 85%, 90%, 95%, 97%, 99%, or 99.9% of the individual HMOs that may be found, present, or detected across a sample of human milk. In some embodiments, the topical formulation comprises the same, or substantially the same HMOs that are found, present, or detected in pooled human milk or in the permeate obtained from pooled human milk.

[0087] In certain embodiments, the topical formulation contains or comprises a permeate resulting from ultrafiltration of pooled human whole milk or skim milk from milk collected from at least 10, 25, 50, or 100 individual human milk donors, and further concentrated, for example, by nanofiltration or reverse osmosis, to increase the concentration (e.g., by w / w) of total HMOs. In some embodiments, the concentration of total HMOs is increased to at least 0.5%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, or 10%. In certain embodiments, the concentration of total HMOs is increased to at least 5% (w / w). In certain embodiments, the concentration of total HMOs is increased to 8%-12% (w / w), inclusive.

[0088] In certain embodiments, topical formulations contain one or more cofactors or vitamins found in or derived from human milk.In certain embodiments, topical formulations contain one or more cofactors or vitamins selected from threonate, pantothenante (vitamin B5), nicotinamide, N1-methyl-2-pyridone-5-carboxamide, pyridoxal dehydroascorbate, nicotinamide ribonucleotide, and pyridoxamine.In some embodiments, topical formulations threonate, pantothenante (vitamin B5), nicotinamide, N1-methyl-2-pyridone-5-carboxamide, pyridoxal dehydroascorbate, nicotinamide ribonucleotide, and pyridoxamine.

[0089] In some embodiments, the topical formulation contains one or more amino acids derived from or found in human milk. In certain embodiments, the topical formulation contains at least one of creatine, creatinine, or arginine. In some embodiments, the topical formulation contains creatine, creatinine, and arginine.

[0090] In some embodiments, topical formulations contain one or more sugars, such as lactose, or energy sources derived from or found in human milk.In certain embodiments, topical formulations contain one or more of citrate, alpha-ketoglutarate, succinate, or phosphate.In certain embodiments, topical formulations contain citrate, alpha-ketoglutarate, succinate, and phosphate. Human Milk Retentate

[0091] In certain embodiments, the topical formulation contains or comprises a retentate fraction of human milk, or a portion or fraction thereof. In some embodiments, the retentate results from filtration, e.g., ultrafiltration, of human skim milk. In certain embodiments, the retentate, e.g., human milk retentate, is or comprises a fraction of skim milk that is retained by a filter, e.g., an ultrafilter. In some embodiments, the retentate fraction, e.g., human milk retentate, contains enriched or concentrated human milk proteins, e.g., enriched or more concentrated relative to whole human milk. In some embodiments, the retentate, e.g., human milk retentate, contains enriched or concentrated human milk proteins that have a molecular weight greater than that which can pass through a filter, e.g., an ultrafilter. In some embodiments, the retentate fraction contains enriched or concentrated human milk proteins having a molecular weight of at least, or greater than, 1 kDa, 5 kDa, 10 kDa, 20 kDa, 25 kDa, 40 kDa, 50 kDa, 100 kDa, 250 kDa, or 500 kDa. In some embodiments, the retentate fraction contains enriched or concentrated human milk proteins having a molecular weight of at least, or greater than, 1 kDa to 40 kDa, or at least about, or greater than about, 1 kDa to 40 kDa (inclusive).

[0092] In certain embodiments, topical formulations contain retentate, such as human milk retentate, or a portion or fraction thereof.In certain embodiments, topical formulations contain, comprise, or are formulated with at least 1%, 5%, 10%, 20%, 25%, 30%, 40%, 50%, 60%, or 70% or more retentate (w / w or w / v), such as human milk retentate, or a portion or fraction thereof.In certain embodiments, topical formulations contain 30%, 25%, 20%, 10%, 5%, 1%, 0.5%, 0.1%, 0.01%, or 0.001% or less retentate (w / w or w / v), such as human milk retentate, or a portion or fraction thereof. In some embodiments, the topical formulation contains at or about 0%-75%, 5%-50%, or 10%-50% retentate, e.g., human milk retentate (w / w or w / v, inclusive).

[0093] In certain embodiments, the topical formulation contains about or at least 0.001%, 0.001%, 0.01%, 0.1%, 0.5%, 1.0%, 2.0%, 2.5%, 5.0%, 7.5%, or 10% or more of a retentate, e.g., human milk retentate (w / w or w / v). In certain embodiments, the topical formulation contains no more than 20%, 10%, 5.0%, 2.5%, 1.0%, or 0.5% retentate, e.g., human milk retentate (w / w or w / v). In some embodiments, the topical formulation contains 0.1%-20%, 1%-5%, or 0.1%-2%, or about 0.1%-20%, 1%-5%, or 0.1%-2% retentate, e.g., human milk retentate (w / w or w / v, inclusive).

[0094] In certain embodiments, topical formulations contain or include one or more proteins found in or derived from human milk.In some aspects, human milk protein comprises or can be divided into whey fraction and casein fraction or complex.In certain embodiments, human milk protein includes, but is not limited to, casein, α-lactalbumin, lactoferrin, secretory immunoglobulin IgA, lysozyme, and serum albumin. In some embodiments, the topical formulation comprises one or more human milk proteins, such as, but not limited to, immunoglobulins, e.g., IgA, sIgA, IgG, and IgM; cytokines, e.g., IL-6, IL-7, IL-8, IL-10, IFN-gamma, TGF-beta, TNF-alpha; chemokines, e.g., G-CSF, and MIF; growth factors, e.g., EGF, VEGF, NGF, IGF, TNFRI, and TNFRII, and erythropoietin; antimicrobials, e.g., lactoferrin, lactadherin; and peptide hormones, e.g., somatostatin, calcitonin, adiponectin, leptin, and ghrelin. Exemplary Topical Formulations

[0095] In some embodiments, the topical formulation is or comprises one or more human milk fractions, parts, or components and one or more pharma- ceutically acceptable excipients. In certain embodiments, the topical formulation is or comprises one or more human milk fractions, parts, or components that are formulated into a pharma- ceutically acceptable topical formulation, such as an oil-in-water emulsion, a water-in-oil emulsion, a nanoemulsion, an emulsified gel, an oil-based ointment, a hydrocolloid system, or a cream preparation.

[0096] In certain embodiments, the topical formulation is or comprises two or more human milk fractions, parts, or components. In certain embodiments, the topical formulation is or comprises human milk cream, or a fraction, subfraction, or portion thereof. In certain embodiments, the topical formulation is or comprises human milk permeate, or a portion or fraction thereof. In some embodiments, the topical formulation is or comprises human milk retentate, or a portion or fraction thereof. In certain embodiments, the topical formulation is or comprises two or more of human milk cream, skim, permeate, or retentate. In various embodiments, the topical formulation is or comprises human milk cream and human milk permeate.

[0097] In certain embodiments, the topical formulation is or comprises human milk cream or a subfraction thereof, and human milk permeate. In certain embodiments, the topical formulation is or comprises human milk cream and human milk permeate. In some embodiments, the topical formulation is or comprises human milk butter oil and human milk permeate. In some embodiments, the topical formulation is or comprises 20%-30% human milk fat (w / w or w / v).

[0098] In some embodiments, the topical formulation is or comprises one or more human milk lipids, fats, or fatty acids. In certain embodiments, the topical formulation is or comprises one or more human milk fatty acids. In certain embodiments, the topical formulation comprises a plurality of human milk fatty acids. In some embodiments, the topical formulation is or comprises one or more human milk oligosaccharides. In certain embodiments, the topical formulation is or comprises a plurality of human milk oligosaccharides. In certain embodiments, the topical formulation is or comprises one or more human milk proteins. In certain embodiments, the topical formulation comprises one or more human milk fatty acids and one or more human milk oligosaccharides. In certain embodiments, the human milk formulation is or comprises a plurality of human milk fatty acids and a plurality of human milk oligosaccharides.

[0099] In certain embodiments, the human milk formulation is or comprises at least 1%, 2.5%, 5%, 7.5%, 10%, 15%, or 20%, or 1%-50%, 10-40%, 1%-40%, 10%-30%, 1%-25%, 5%-20%, or 5%-15% (e.g., w / v or w / w) human milk lipids, fats, or fatty acids, and at least 0.01%, 0.1%, or 0.25%, or 0.1%-1% (e.g., w / v or w / w), inclusive, of human milk oligosaccharides. In certain embodiments, the topical formulation is or comprises 5%-15% human milk lipids, fats, or fatty acids, and 0.1%-1% human milk oligosaccharides (e.g., w / v or w / w). In certain embodiments, the topical formulation is or comprises about 10% human milk lipids, fats, or fatty acids, and 0.1%-0.5% human milk oligosaccharides (e.g., w / v or w / w). In some embodiments, the topical formulation is or comprises 15%-30% human milk fat, and 0.1%-1% human milk oligosaccharides (e.g., w / w). In certain embodiments, the topical formulation is or comprises about 20% human milk fat, and 0.1%-0.5% human milk oligosaccharides (e.g., w / w). In various embodiments, the topical formulation is or comprises about 30% human milk fat, and 0.1%-0.5% human milk oligosaccharides (e.g., w / w, inclusive).

[0100] In some embodiments, the topical formulation is or comprises a plurality of human milk lipids and one or more human milk oligosaccharides. In various embodiments, the topical formulation contains a plurality of human milk lipids that is or comprises one or more neutral complex lipids (e.g., free fatty acids, cholesteryl esters, diacylglycerols, triacylglycerols, monoacylglycerols), sphingolipids (e.g., ceramides, dihydroceramides, hexosylceramides, lactosylceramides, sphingomyelins), phospholipids (e.g., phosphatidylcholine, lysophosphatidylcholine, phosphatidylethanolamines, lysophophatidylethanolamines, phosphatidylinositol), or any or all of the above in combination with one or HMOs.

[0101] In certain embodiments, topical formulation is or comprises a plurality of human milk fatty acids and one or more HMOs.In certain embodiments, topical formulation contains or comprises a plurality of human milk fatty acids, including some or all of oleic acid, palmitic acid, linoleic acid, omega-3 fatty acid, omega-6 fatty acid, omega-9 fatty acid, stearic acid, myristic acid, lauric acid, palmitoleic acid, vaccenic acid, heptadecanoic acid, myristoleic acid, heptadecene, docosahexaenoic acid, gamma-linolenic acid, docosapentaenoic acid, eicosapentaenoic acid (EPA), arachidonic acid (AA), and docosahexaenoic acid (DHA), and one or more HMOs.

[0102] In certain embodiments, the topical formulation is or comprises a plurality of human milk oligosaccharides and one or more human milk lipids, fats, or fatty acids. In certain embodiments, the topical formulation is or comprises a plurality of at least 10, 25, 50, or 100 human milk oligosaccharides, including some or all of 2'-fucosyl-lactose, 3-fucosyl-lactose, 3'-sialyl-lactose, 6'-sialyl-lactose, lacto-N-tetraose, lacto-N-difucohexaose I, lactodifucotetraose, lacto-N-fucopentaose I, sialyllacto-N-tetraose c, sialyllacto-N-tetraose b, and disialyllacto-N-tetraose, as well as one or more human milk lipids, fats, or fatty acids. In certain embodiments, the topical formulation is or comprises a plurality of at least 10, 25, 50, or 100 human milk oligosaccharides, including some or all of 2'-fucosyl-lactose, 3-fucosyl-lactose, 3'-sialyl-lactose, 6'-sialyl-lactose, lacto-N-tetraose, lacto-N-difucohexaose I, lactodifucotetraose, lacto-N-fucopentaose I, sialyllacto-N-tetraose c, sialyllacto-N-tetraose b, and disialyllacto-N-tetraose, as well as one or more human milk lipids, fats, or fatty acids.

[0103] In various embodiments, the topical formulation is or comprises a plurality of human milk oligosaccharides and a plurality of at least 10, 25, 50, or 100 human milk fatty acids. In certain embodiments, the topical formulation comprises (i) oleic acid, palmitic acid, linoleic acid, omega-3 fatty acids, omega-6 fatty acids, omega-9 fatty acids, stearic acid, myristic acid, lauric acid, palmitoleic acid, vaccenic acid, heptadecanoic acid, myristoleic acid, and (ii) a plurality of human milk oligosaccharides including some or all of 2'-fucosyl-lactose, 3-fucosyl-lactose, 3'-sialyl-lactose, 6'-sialyl-lactose, lacto-N-tetraose, lacto-N-difucohexaose I, lactodifucotetraose, lacto-N-fucopentaose I, sialylacto-N-tetraose c, sialylacto-N-tetraose b, and disialyllacto-N-tetraose.

[0104] In certain embodiments, the topical formulation is stable, e.g., can be stored for a period of time without spoilage or loss of efficacy or biological activity. In some embodiments, the topical formulation can be stored, e.g., refrigerated, at 2°C and 8°C, or at 2°C to 8°C, for at least 6 weeks, 8 weeks, 10 weeks, 12 weeks, or at least 2 months, 3 months, 6 months, 9 months, or 12 months or more, e.g., without spoilage and / or without detectable loss of efficacy or biological activity. In certain embodiments, the topical formulation can be stored at room temperature and / or ambient temperature (such as 16°C and 24°C, or at 16°C to 24°C, or about 20°C), for at least 6 weeks, 8 weeks, 10 weeks, 12 weeks, or at least 2 months, 3 months, 6 months, 9 months, or 12 months or more, e.g., without spoilage and / or without detectable loss of efficacy or biological activity. Certain embodiments contemplate that changes in efficacy or bioactivity may be determined by one of skill in the art as a matter of routine, including, for example, by any of the methods or assays described herein in the Examples. In some embodiments, stability is or includes a low degree of change over time in the composition of human milk components present in the topical formulation, such as, for example, a change in the degree of the human milk fatty acid composition of the topical formulation. In some embodiments, a low degree of change is or includes less than a 10%, 5%, or 1% relative difference in the concentration or amount of 10, 15, or 20 or more distinct human milk components, such as structurally distinct human milk fatty acids and / or human milk oligosaccharides, between two time points. II. Uses of the Provided Compositions and Formulations

[0105] Also provided herein are uses and methods for using topical formulations containing one or more parts or fractions of human milk, such as any one of the compositions described herein, such as Section I. In some embodiments, the topical formulations are applied topically and / or directly to the skin and / or mucous membranes. In some embodiments, the topical formulations are applied topically and / or directly to the skin to promote, improve, or enhance hydration or barrier function. In some embodiments, the topical formulations are applied topically and / or directly to the skin to improve appearance (e.g., reduce wrinkles or eliminate redness). In some embodiments, the topical formulations may be applied topically and / or directly to the skin to eliminate, improve, or reduce pain, irritation, or inflammation.

[0106] In certain embodiments, topical formulations may be applied or administered to subjects to treat or prevent disease or pathology.In certain embodiments, the method is or includes administering or applying an effective amount of the provided topical formulation to a subject that has or is at risk of having disease or pathology, or that exhibits symptoms and / or clinical signs of disease or pathology.In certain embodiments, the disease or pathology is a skin disorder or pathology.In some embodiments, at least one symptom and / or clinical sign of disease or pathology, for example, skin disorder or pathology, is changed, reduced, alleviated, or improved after administration or application of the topical formulation.

[0107] In certain embodiments, the skin disorder or condition is or includes acne, chronic skin ulcers, dermatitis, skin sunburn, infection, nipple cracks, psoriasis, rosacea, side effects of skin cancer therapy, vitiligo, wound care, or xerosis. In some embodiments, the skin disorder or condition is or includes acne, signs of aging, cellulite, melasma, scars, stretch marks, sun damage. In certain embodiments, the topical formulation may be applied as an anti-aging treatment, an anti-cellulate treatment, a cleanser treatment, a face mask or body compress lip care, a makeup remover, melasma reduction, a moisturizer, scar reduction, skin care before or after a cosmetic procedure, a stretch mark treatment, sun care, or a toner.

[0108] In some embodiments, the skin disorder or condition is or includes dermatitis, such as, but not limited to, atopic dermatitis, contact dermatitis, hand dermatitis, allergic dermatitis, stasis dermatitis, asteatotic dermatitis, nummular dermatitis, seborrheic dermatitis, infectious dermatitis, chronic superficial scaly acrodermatitis, chondrodermatitis, and perioral dermatitis. In certain embodiments, the skin disorder or condition is acne, such as acne vulgaris, severe necrotizing acne, necrotizing acne or necrotizing lymphocytic folliculitis, and / or variolous. In certain embodiments, the skin disorder or condition is rosacea, such as erythermatotelangietatic rosacea, papulopustular rosacea, phymatous rosacea, or ocular rosacea. In some embodiments, the skin disorder or condition is an autoimmune disorder or condition. In some embodiments, the skin condition or disorder is a drug-induced skin condition or disorder. In certain embodiments, the skin condition or disorder is autoimmune or drug-induced dermatomyositis, or similar conditions associated with internal pathology, such as neutrophilic dermatosis, dermatomyositis, or transient acantholytic dermatosis. In some embodiments, the skin condition or disorder is eczema and / or is associated with a genetic or metabolic condition, including but not limited to ichthyosis, xerosis, or asteatosis. In some embodiments, the skin condition or disorder is a dry skin condition, such as psoriasis, lupus erythematosus, exfoliative keratolysis, or any type of allergic inflammation of the skin, for example, urticaria.

[0109] In certain embodiments, the skin condition or disorder is epidermolysis bullosa or epidermolysis bullosa simplex. In certain embodiments, epidermolysis bullosa simplex may include, but is not limited to, epidermolysis bullosa simplex Weber-Cockayne type, epidermolysis bullosa simplex Koebner type, epidermolysis bullosa simplex with patchy pigmentation, epidermolysis bullosa simplex Dowling-Meara type, or epidermolysis bullosa simplex with muscular dystrophy.

[0110] In some embodiments, the skin condition or disorder is contagious, such as those associated with or triggered by viruses or bacteria. In some embodiments, the skin condition or disorder is or includes impetigo. In certain embodiments, the skin condition or disorder is or includes necrotizing fasciitis.

[0111] In some embodiments, the skin condition or disease is epidermolytic or non-epidermolytic keratin-based skin disease.Exemplary types of keratin-based skin disease include, but are not limited to, epidermolytic hyperkeratosis, ichtyosis bullosa of Siemens, pachyonychia congenita, epidermolytic or non-epidermolytic palmoplantar keratoderma (diffuse or focal), multiple sebaceous cysts, Naegeli-Franceschetti-Jadassohn syndrome, and reticular pigmented dermatosis.

[0112] In some embodiments, the skin condition or disorder is or results from mechanical disruption, injury, wound, or damage. In some embodiments, the skin condition or disorder is or results from medical or surgical treatment, such as surgical cut or incision. In some embodiments, the skin condition or disorder is or results from a biopsy, such as, but not limited to, a biopsy performed using a needle or trocar. In some embodiments, the biopsy is or includes an incision biopsy. In certain embodiments, the biopsy is or includes a punch biopsy. In some embodiments, the skin condition or disorder is or results from a burn, such as from radiation therapy.

[0113] In some embodiments, the skin condition or disorder is or includes contact dermatitis or contact sensitivity. In certain embodiments, the skin condition or disorder is or includes contact sensitivity to mercury compounds (such as those found in ointments), chromates, nickel, turpentine, varnishes, resins, cosmetics, or dyes. In certain embodiments, the skin disorder or condition is or includes nickel hypersensitivity. In some embodiments, the skin condition or disorder is or includes keratosis pilaris, keratoderma,

[0114] In certain embodiments, the skin disorder or condition is the result of exposure to plants such as, for example, Toxicodendron diversilobum (sumac), Toxicodendron radicans (poison ivy), Toxicodendron rydbergii (Rocky Mountain poison oak), Toxicodendron vernix (poison sumac or poison sumac), Toxicodendron vernicifluum (Japanese or Asian lacquer tree), Magnifera indica (mango tree), Anacardium occidentale (cashew nut tree), Gluta renghas (rengas tree), Melanorrhoea usitata (Burmese sumac tree), Metopium toxiferum or Comocladia dodnaea (both Caribbean shrubs), Semecarpus anacardium (Indian marking nut tree), or Ginkgo biloba. In some embodiments, the skin condition is, or results from, exposure to or a bite from an arachnid or insect, such as a mosquito, a blackfly, a sand fly, a flea, a stable fly, a sand flea, an ant, a spider, a tick other than an ixodid, or a tick.

[0115] In some embodiments, the skin disorder or condition is atopic dermatitis. In certain embodiments, the pathogenesis of atopic dermatitis (and eczema in general) involves or is associated with immunologically mediated leukocyte infiltration, such as infiltration of mononuclear cells, lymphocytes, neutrophils, and eosinophils into the skin or dermis. In some aspects, chronic eczema is also associated with significant hyperproliferation of the epidermis. In some aspects, atopic dermatitis and eczema can be fatal if severe enough. Less severe, but unpleasant and often painful symptoms associated with atopic dermatitis may include, but are not limited to, itching, swelling, redness, blisters, crusting, ulceration, pain, scaling, cracking, hair loss, scarring, or fluid exudation involving the skin, eyes, or mucous membranes. In certain embodiments, the topical formulation is administered or applied to the skin of a subject with atopic dermatitis. In certain embodiments, the topical formulation is administered or applied to a subject to treat, prevent, reduce, improve, or alleviate atopic dermatitis. In certain embodiments, the topical formulation is administered or applied to a subject to treat, prevent, reduce, improve, or alleviate one or more symptoms associated with atopic dermatitis.

[0116] In certain embodiments, the disease state or disorder is psoriasis. In some aspects, psoriasis is one of the most common autoimmune diseases characterized by abnormal skin patches. Affected skin patches are red, show scaling, and are itchy and stinging. In certain aspects, there are five main types of psoriasis: patchy, guttate, inverse, pustular, and erythrodermic, with patchy psoriasis being the most common. In certain embodiments, the topical formulation is administered or applied to a subject to treat, prevent, reduce, improve, or alleviate psoriasis, such as patchy psoriasis, guttate psoriasis, inverse psoriasis, pustular psoriasis, or erythrodermic psoriasis. In certain embodiments, the topical formulation is administered or applied to a subject to treat, prevent, reduce, improve, or alleviate one or more symptoms associated with psoriasis, such as patchy psoriasis, guttate psoriasis, inverse psoriasis, pustular psoriasis, or erythrodermic psoriasis.

[0117] In some embodiments, the topical formulation is administered and / or applied topically. In some embodiments, the topical formulation is applied topically and / or directly to an area, for example, an area of ​​skin, that has or is at risk of having one or more symptoms associated with a skin disorder or condition. In some embodiments, the topical formulation is administered and / or applied, for example, directly and / or topically, to an area of ​​skin that has or is at risk of having one or more symptoms associated with a skin disorder or condition. In some embodiments, symptoms associated with a skin disorder or condition include, but are not limited to, dryness, lesions, cracks, fissures, scaling, discoloration, thickening, redness, bleeding, swelling, peeling, ulcers, pain, wounds, blistering, rash, infection, or flesh loss. In certain embodiments, the topical formulation is administered and / or applied to treat, prevent, reduce, improve, or alleviate one or more symptoms associated with a skin disorder or condition.

[0118] In some embodiments, the topical formulation is applied to improve the appearance of the skin, such as to reduce or prevent wrinkles or blemishes due to skin damage and / or aging. In certain embodiments, application of the topical formulation to the skin may reduce inflammation, irritation, and erythema of the skin, along with increasing skin elasticity and flexibility. In certain embodiments, application of the topical formulation reduces the presence or severity of wrinkles and skin blemishes, including but not limited to "marionette" lines, laugh lines, deep nasolabial folds, crow's feet, fine lines / wrinkles, vertical frown lines, horizontal forehead lines, loose thin / weak skin, redness and dullness of the skin. In some aspects, improvement of barrier function is expected to improve the appearance of the skin, and thus increasing barrier function with the provided topical formulation prevents or reduces skin aging, dryness, dullness, loss of elasticity, and lack of radiance. In some aspects, topical application or administration prevents, reduces, or delays the appearance of spider veins or red spots, exaggerated lines, and wrinkles.

[0119] In certain embodiments, the topical formulation is a moisturizer. In certain embodiments, the topical formulation is administered or applied to moisturize the skin or to treat or prevent the appearance of dry skin, flaky skin, or rough skin. In some embodiments, the topical formulation is administered or applied to treat (e.g., reduce the severity, frequency, or appearance of) or prevent skin conditions ranging from pruritus to spider veins, lentigines, age spots, senile purpura, keratosis, melasma, plaques, fine lines or wrinkles, nodules, sun-damaged skin, dermatitis (including but not limited to seborrheic dermatitis, nummular dermatitis, contact dermatitis, atopic dermatitis, exfoliative dermatitis, perioral dermatitis, and stasis dermatitis), psoriasis, folliculitis, rosacea, acne, impetigo, erysipelas, erythrasma, eczema, and other inflammatory skin conditions. In certain non-limiting embodiments, the skin condition may be caused by exposure to UV light, age, irradiation, chronic sun exposure, environmental pollutants, air pollution, wind, cold, heat, chemicals, disease pathology, smoking, or nutritional deficiency. The skin may be facial skin or non-facial skin (e.g., arms, legs, hands, chest, back, feet, etc.).

[0120] In certain embodiments, the topical formulation is applied or administered to increase the stratum corneum turnover rate of the skin, increase collagen synthesis in fibroblasts, increase cellular antioxidant defense mechanisms (e.g., to reduce or prevent oxidative damage to skin, cells, proteins, and lipids, to enhance, replenish, and / or prevent loss of cellular antioxidants such as catalase and glutathione in skin cells (e.g., keratinocytes, melanocytes, Langerhans cells, etc.), inhibit melanin production in melanocytes, and / or reduce or prevent oxidative damage to the skin (including reducing the amount of lipid peroxides and / or protein oxidation in the skin). In certain embodiments, the topical formulation is applied and / or administered to reduce the amount of internal oxidation and / or external oxidative damage in cells. In certain embodiments, the topical formulation is administered or applied to increase collagen synthesis in cells. In some embodiments, the topical formulation reduces skin inflammation, such as by reducing inflammatory cytokine production in cells. Non-limiting examples of such cells include human epidermal keratinocytes, human fibroblast dermal cells, human melanocytes, and three-dimensional human cell-derived in vitro tissue equivalents comprising human keratinocytes, human fibroblasts, or human melanocytes, or any combination thereof (e.g., a combination of human keratinocytes and human fibroblasts, or a combination of human keratinocytes and human melanocytes).

[0121] In some embodiments, the topical formulation is a cosmetic formulation or composition.Non-limiting examples of cosmetic compositions or formulations include, but are not limited to, moisturizers, creams, lotions, emollients, foundations, night creams, lipsticks, cleansers, toners, sunscreens, masks, or anti-aging products.

[0122] In some embodiments, the topical formulation is or comprises a cosmetic preparation that protects, moisturizes, and / or lubricates the skin. In some embodiments, the topical formulation is or comprises an emollient. In certain embodiments, the topical formulation is or comprises a moisturizer. In some embodiments, the topical formulation is applied or administered to the skin to perform one or more functions typically performed by sebum produced by healthy skin. In some embodiments, the topical formulation is administered or applied to the skin to prevent, reduce, or improve transepidermal water loss (TEWL).

[0123] In some embodiments, the TEWL is typically about 4-8 g / m 2 In certain embodiments, application or administration of the topical formulation reduces or decreases TEWL by at least 25%, 30%, 40%, 50%, 60%, 70%, 75%, 80%, or 90%, for at least 30 minutes, 1 hour, 2 hours, 3 hours, 4 hours, 6 hours, 8 hours, 12 hours, or 24 hours, e.g., compared to immediately before the topical formulation is applied or administered and / or compared to untreated skin.

[0124] In some embodiments, the subject is a vertebrate. In some embodiments, the subject is a mammal, such as a human. In some embodiments, the subject is a human subject. In certain embodiments, the subject is an infant, toddler, child, adolescent, teen, or adult.

[0125] Certain embodiments contemplate that application of a topical formulation containing lipids, oligosaccharides, or proteins (or combinations thereof) derived from or produced by human milk is more effective in preventing, reducing, improving, or alleviating one or more symptoms in a human subject than topical formulations containing lipids, oligosaccharides, or proteins derived from or produced by non-human sources (e.g., bovine milk). In some embodiments, a "more effective" prevention, reduction, improvement, or alleviation of one or more symptoms is or includes a higher degree of prevention, reduction, improvement, or alleviation of one or more symptoms. In certain embodiments, a "more effective" prevention, reduction, improvement, or alleviation of one or more symptoms is or includes a shorter duration of time required to observe prevention, reduction, improvement, or alleviation after application or administration of the topical formulation. In some aspects, a person skilled in the art can identify whether a given topical formulation is more effective for preventing, reducing, improving, or alleviating one or more symptoms as a routine procedure.

[0126] Certain embodiments contemplate that application of a topical formulation containing lipids, oligosaccharides, or proteins (or combinations thereof) derived from or produced by human milk is less likely to trigger or cause sensitization (e.g., contact dermatitis), skin irritation, inflammation, or immune responses in a human subject, e.g., on human skin, than a topical formulation containing lipids, oligosaccharides, or proteins derived from or produced by a non-human source (e.g., bovine milk). In some embodiments, provided topical formulations are 30%, 40%, 50%, 60%, 70%, 80%, or 90% less likely to trigger or cause sensitization, skin irritation, inflammation, allergy, or immune responses in a human subject, e.g., on human skin, than topical formulations containing lipids, oligosaccharides, or proteins derived or produced from a non-human source (e.g., bovine milk), or are about 30%, 40%, 50%, 60%, 70%, 80%, or 90% less likely, or at least 30%, 40%, 50%, 60%, 70%, 80%, or 90% less likely. In some embodiments, the likelihood or probability that a topical formulation will trigger or cause sensitization, skin irritation, inflammation, allergy, or immune response is less than or about 20%, 10%, 5%, 1%, 0.1%, or 0.01%.

[0127] In certain embodiments, the topical formulation is applied or administered to a subject to improve or increase the barrier function of the skin. In certain embodiments, the subject has healthy skin. In some embodiments, the subject is experiencing one or more symptoms associated with a skin disease or condition. In certain embodiments, the topical formulation, e.g., topical formulation, improves or increases barrier function by at least 5%, 10%, 20%, 25%, 30%, 40%, 50%, 60%, 70%, 75%, 80%, 90%, 100% 150%, 200%, or 1-fold, 2-fold, 5-fold, 10-fold, 20-fold, or 50-fold, e.g., compared to the barrier function before administration or application of the topical formulation and / or compared to untreated skin. In some aspects, barrier function may be measured or assessed by any suitable routine known technique (e.g., non-invasive methods that measure water content and loss through the skin surface, such as corneometry or measuring transepidermal water loss (TEWL)).

[0128] In some embodiments, the barrier function of a subject's skin may be measured or assessed by corneometry, which in some aspects indirectly measures barrier function by determining the capacitance of the skin, e.g., 10-20 μm thickness of the stratum corneum, due to its behavior as a dielectric medium. In some aspects, this measurement relates to the degree of hydration under various physiological conditions in response to injury, metabolic events, or topical therapy. For example, in some embodiments, changes in individual or average skin hydration scores in a group of human subjects may serve as an indicator of skin health, with higher values ​​typically being considered more desirable. In some embodiments, the topical formulation increases skin hydration, as measured by corneometry, by a detectable amount or by at least 5%, 10%, 20%, 25%, 30%, 40%, 50%, 60%, 70%, 75%, 80%, 90%, 100% 150%, 200%, or 1-fold, 2-fold, 5-fold, 10-fold, 20-fold, or 50-fold, for example, as compared to barrier function prior to administration or application of the topical formulation and / or as compared to untreated skin.

[0129] In some embodiments, for example, the integrity of the barrier function of the subject's skin is evaluated by measuring TEWL through the epidermal surface.In some aspects, the TEWL value is a measure of the rate of water loss (g / h-m2) through the skin, and is an estimate of the skin's ability to retain water.The method is based on the measurement of the water vapor density gradient established in the layer 10 mm above the skin surface.This is an indication of the degree of possible damage to the water barrier function of the skin.Since water loss through the skin usually occurs by passive diffusion through the epidermis, a higher TEWL value indicates a greater water loss and is consistent with increased damage to the barrier function of the stratum corneum, such as that which may occur during irritant exposure or atopic dermatitis. In certain embodiments, the topical formulation increases TEWL by a detectable amount or by at least 10%, 20%, 25%, 30%, 40%, 50%, 60%, 70%, 75%, 80%, 90%, 95% 100%, for example, as compared to barrier function prior to administration or application of the topical formulation.

[0130] In certain embodiments, the topical formulation is applied or administered to a subject to treat, prevent, reduce, alleviate or improve inflammation, for example, inflammation of the skin. In some embodiments, inflammation is associated with skin disorders or pathologies. In certain embodiments, application or administration of the topical formulation reduces inflammation, for example, chronic inflammation or acute inflammation. In certain embodiments, application or administration of the topical formulation reduces the amount or presence of one or more inflammatory cytokines, for example, in or near the area where the topical formulation is administered or applied. In some embodiments, the inflammatory cytokine is or includes IL-8.

[0131] In some embodiments, the topical formulation is administered or applied to a wound, such as a cut, abrasion, lesion, laceration, or scar, to promote or improve healing or repair.In certain embodiments, the wound heals or repairs in less time, such as at least 10%, 25%, or 50% less time than a similar wound that has not been administered or applied with the topical formulation.In certain embodiments, application of the topical formulation to the wound reduces or decreases the probability or likelihood of scarring, such as in contrast to a similar wound that has not been applied with the topical formulation.

[0132] In certain embodiments, the topical formulation is a nipple cream or nipple balm. In certain embodiments, the topical formulation is applied to treat or prevent nipple cracking. In some embodiments, the topical formulation is applied or administered to a subject to treat or prevent pain, skin irritation, or skin damage caused by or associated with breastfeeding. In certain embodiments, a concern with nipple creams and nipple balms is that breast-fed infants may ingest the nipple cream or nipple balm while breastfeeding, for example, inadvertently. For example, many nipple creams or nipple balms contain lanolin, for example, as an active ingredient. Lanolin is a by-product extract from the sebum of sheep's wool after shearing. In addition to potential allergy risks of sheep-derived products, sheep are commonly treated with pesticides and insecticides, thereby increasing the risk of exposing infants to pesticide residues (discussed in Heikes and Craun, J. Agric. Food Chem. 1992, 40 (9), pp 1586-1590, incorporated herein by reference in its entirety). In contrast, in some embodiments, provided topical formulations contain natural ingredients derived or derived from human breast milk, and thus are safe for infants, and in some embodiments may promote health benefits in infants as well. In certain embodiments, the topical formulations are alcohol-free.

[0133] In certain embodiments, the topical formulation is administered or applied to a subject to treat, prevent, reduce, improve, or alleviate stomatitis or mucositis. In certain embodiments, the topical formulation is administered or applied to a subject as an oral formulation, for example, by rinsing, swabbing, or swallowing. In some embodiments, the topical formulation is administered or applied to a subject to reduce one or more symptoms associated with stomatitis. In certain embodiments, the topical formulation is administered or applied to a subject to treat, prevent, reduce, alleviate, or alleviate pain or inflammation anywhere in the mouth, such as the inside of the cheek, gums, tongue, lips, or palate. In some embodiments, the topical formulation is administered or applied to reduce or reduce the severity or amount of aphthous stomatitis or cold sores.

[0134] In certain embodiments, topical formulations are administered or applied to subjects to reduce one or more symptoms associated with mucositis.In certain embodiments, topical formulations are administered or applied to subjects to treat, prevent, reduce, alleviate or improve pain, inflammation or ulcer anywhere along the mucous membrane that lines the digestive tract.In some embodiments, mucositis is associated with or occurs as a result of cancer treatment, for example, chemotherapy, radiation therapy or hematopoietic stem cell transplantation.

[0135] In some embodiments, the topical formulation is administered or applied to a subject to reduce one or more symptoms of a disease or condition that is localized to the esophagus. In some embodiments, the topical formulation is administered or applied to a subject to treat, prevent, reduce, ameliorate, or alleviate one or more symptoms associated with eosinophilic esophagus. In certain embodiments, the topical formulation is administered or applied to a subject to reduce or decrease the severity or occurrence of dysphagia, food impaction, vomiting, heartburn, or acid reflux. III. Preparation of Compositions Containing Parts or Fractions of Human Milk

[0136] In certain embodiments, the topical formulations provided are formulated, produced, or produced using one or more fractions, parts, or components of human milk. In some embodiments, one or more fractions or parts of human milk, such as cream, butter oil, milk fat globule membrane solids, skim milk, permeate, or retentate, are collected, isolated, or obtained from human whole milk, such as for preparing, producing, or manufacturing the topical formulation. In certain embodiments, the parts or fractions from human whole milk are obtained by methods similar to or identical to those described in U.S. Patent No. 8,545,920, U.S. Patent No. 8,377,445, U.S. Patent No. 9,149,052, U.S. Patent No. 8,927,027, or WO2014158911 and WO2018053535 (herein incorporated by reference in their entirety). In certain embodiments, one or more of the fractions, portions, or components of human milk are mixed, formulated, or incorporated into a topical formulation, such as a pharma- ceutically acceptable topical formulation described herein.

[0137] In some embodiments, provided herein is a method for producing a topical formulation containing human milk, or one or more parts, fractions, or components thereof. In certain embodiments, the method is or includes isolating, separating, purifying, or obtaining a part, fraction, or component of human milk, and formulating or incorporating the part, fraction, or component into a topical formulation, such as an emulsion, cream, ointment, or lotion. In some embodiments, the part, fraction, or component is or includes human milk cream, skim, retentate, or permeate, or a part or fraction derived from or derived from cream, skim milk, retentate, or permeate. In some embodiments, the method is or includes formulating or incorporating two or more parts, fractions, or components of human milk (e.g., cream, skim milk, permeate, or retentate) into a topical formulation, such as a pharma- ceutically acceptable topical formulation.

[0138] In certain embodiments, the method of making a topical formulation is or includes isolating, separating, purifying, or obtaining a fraction, portion, or component from human whole milk. In certain embodiments, the fraction or portion is or includes human milk cream, butteroil, skim, retentate, or permeate (or a fraction or portion derived from or from a cream, skim milk, retentate, or permeate). In some embodiments, the method is or includes formulating or incorporating human milk cream, skim milk, retentate, or permeate (or a subfraction, fraction, or portion derived from or from a cream, such as butteroil or milk fat globule membrane solids, skim milk, retentate, or permeate) into a topical formulation, such as a cream, ointment, or lotion. In certain embodiments, the isolated, separated, purified, or obtained fraction may be further processed, such as to further purify or enrich one or more components of the fraction (e.g., to purify or enrich lipids, human milk oligosaccharides, or proteins above or below a certain molecular weight) prior to formulation or incorporation into a topical formulation.

[0139] In certain embodiments, the method of making a topical formulation is or includes measuring the content, level, or concentration of one or more macronutrients, compounds, vitamins, proteins, growth factors, fats, fatty acids, or lipids of a fraction, part, or component from human whole milk. In some embodiments, a fraction, part, or component from human whole milk is added to the formulation to achieve a target amount, level, or concentration of one or more macronutrients, compounds, vitamins, proteins, growth factors, fats, fatty acids, or lipids of the fraction, part, or component from human whole milk. In certain aspects, the content, level, or concentration of one or more macronutrients, compounds, vitamins, proteins, growth factors, fats, fatty acids, or lipids present in a fraction, part, or component from human whole milk may be measured as a routine procedure by those skilled in the art, including by any of the means described herein, such as by the examples.

[0140] In certain embodiments, production is or includes isolating, separating, refining, or obtaining the cream, skim milk, retentate, or permeate, and then further processing the cream, skim milk, retentate, or permeate to enrich or purify one or more fractions, portions, or substances within the cream, skim milk, retentate, or permeate prior to incorporation or formulation into a topical formulation. In certain embodiments, human milk cream is processed to produce subfractions, such as butter oil and / or milk fat globule membrane solids.

[0141] In certain embodiments, topical formulations are provided herein that contain one or more fractions of human milk.In certain aspects, topical formulations contain fractions or portions of human milk that are obtained or derived from human whole milk sources.In some embodiments, the human whole milk source is human whole milk obtained from one or more individual human donors.In certain embodiments, the source is human whole milk pooled from multiple individual donors.

[0142] In certain embodiments, the method of manufacture is or includes isolating, separating, purifying, or obtaining one or more fractions from human milk (such as from a human whole milk source, e.g., from human whole milk from an individual donor or pooled from multiple donors). In certain embodiments, a sample is collected from the human milk source, e.g., pooled human milk, and tested for contamination. Testing for contamination may include, but is not limited to, PCR testing for viruses such as HIV, HBV, and HCV.

[0143] In some embodiments, human milk, or a portion, fraction, or component thereof, is treated to reduce bioburden. In some embodiments, the process of reducing bioburden may include one or both of filtration and pasteurization (or any other suitable known method of reducing bioburden). In certain embodiments, human milk, or a fraction or portion thereof, may be treated (e.g., filtered or pasteurized) to reduce bioburden at any stage of the process for producing the topical formulation. In some embodiments, human milk, or a fraction or portion thereof, may undergo more than one process to reduce bioburden. In certain embodiments, human milk, or a fraction or portion thereof, undergoes a process to reduce bioburden more than once. For example, a first filtration and / or pasteurization of human whole milk, followed by a second filtration and / or pasteurization of a fraction or portion of human milk that has been purified, separated, or removed from the human whole milk.

[0144] In some embodiments, the temperature of the human milk, and any fractions or portions thereof, is continuously monitored during the process for manufacturing the topical formulation, for example to ensure consistency between batches and to verify that the human milk, or fractions or portions thereof, does not reach temperatures outside of threshold temperatures that may be present during each phase of the process.

[0145] In some embodiments, human milk from multiple individual donors is pooled and filtered through a large micron filter or screen, such as to remove clumps or large particulate matter. In some embodiments, the pooled whole human milk is filtered through a filter or screen. In certain embodiments, the filter or screen contains a pore size, e.g., an average or median pore size, of 100 μm to 1,000 μm. In certain embodiments, the screen contains a pore size, e.g., an average or median pore size, of at least or about 100 μm, 200 μm, 250 μm, 500 μm, or 1,000 μm. In certain embodiments, the whole human milk is filtered through a screen having a median or average pore size of 200 μm or about 200 μm.

[0146] In certain embodiments, the process of reducing bioburden is or includes heat treatment, e.g., pasteurization. In some embodiments, human whole milk is pasteurized or heat treated by any suitable means, including, but not limited to, techniques such as vat or batch pasteurization, high temperature short time (HTST), very high temperature very short time (HHST), ultra pasteurization (UP), or ultra high temperature (UHT). In some embodiments, portions or fractions of milk having a high fat or lipid content, e.g., greater than 10%, are pasteurized at a higher temperature, e.g., at least 3° C. higher, than whole milk or low-fat fractions.

[0147] In some embodiments, pooled human milk from individual donors is separated into one or more fractions, such as cream, permeate, and retentate. In some embodiments, the fractions are separated by a process that includes: (a) filtering the pooled human whole milk through a 200 μm filter; (b) pasteurizing the milk at or about 63° C. for 30 minutes or about 30 minutes; (c) producing a cream fraction and a skim milk fraction from the human whole milk by centrifugation; and (d) ultrafiltering some or all of the skim milk to produce a permeate and a retentate fraction. In some embodiments, one or more of the human milk cream, skim, permeate, or retentate fractions are formulated into a topical formulation. In some embodiments, the cream, skim, permeate, or retentate fractions are further processed, such as by filtration or centrifugation to enrich for one or more compounds, before being incorporated into a topical formulation.

[0148] In certain embodiments, the method of making a topical formulation is or includes isolating, separating, purifying, or obtaining from human whole milk one or more of the cream (or a fragment or portion derived from or derived from the cream) and the retentate or permeate (or a portion or fraction derived from or derived from the retentate or permeate) for incorporation into a topical formulation. In some embodiments, the method of making a composition is or includes isolating, separating, purifying, or obtaining from human whole milk the cream (or a fragment or portion derived from or derived from the cream) and the permeate (or a portion or fraction derived from or derived from the permeate) for incorporation into a topical formulation.

[0149] In various embodiments, the method of making the composition is or includes isolating, separating, purifying, or obtaining the cream and permeate (or portions or fractions derived or derived from the cream and / or permeate) from human whole milk, and formulating or incorporating the cream and permeate (or fragments or portions derived or derived from the cream and / or permeate) into a composition, e.g., a topical formulation. In some embodiments, the method of making the composition is or includes formulating or incorporating one or more of the cream (or subfractions or portions derived from or derived from the cream, such as butter oil or milk fat globule membrane solids), and the retentate or permeate (or portions or fractions derived from or derived from the permeate) isolated, separated, purified, or obtained from human whole milk into a composition, e.g., a topical formulation. Cream and Skim Fractions

[0150] In some embodiments, the cream and skim milk fractions are obtained by removing fat or cream from human whole milk. In some embodiments, human whole milk is separated into skim milk fractions and cream fractions, such as by known or routine methods, including but not limited to centrifugation. In some embodiments, human whole milk is centrifuged into separated cream and skim milk fractions. In certain embodiments, the topical formulation contains cream obtained by centrifuging human whole milk. In certain embodiments, the composition, e.g., topical formulation, contains skim milk obtained by removing cream by centrifugation or creaming separation.

[0151] Certain embodiments contemplate that methods and techniques for removing cream from whole milk, e.g., human whole milk or breast milk, are known and can be performed as a routine procedure. In some embodiments, the method of making the composition is or includes isolating, separating, purifying, or obtaining cream (or a fragment or portion derived from or derived from cream, such as butter oil or milk fat globule membrane solid subfraction) from human whole milk, and formulating or incorporating the cream or a subfraction thereof into a composition, e.g., a topical formulation. In some embodiments, the method of making the composition is or includes formulating or incorporating the cream or a subfraction thereof isolated, separated, purified, or obtained from human whole milk into a topical formulation.

[0152] In certain embodiments, the cream, e.g., human milk cream, is obtained from a method or technique that is or includes creaming separation or centrifugation. In some aspects, creaming separation allows for the combination of fat globules over time and therefore the accumulation of fat globules in the cream layer towards the surface of the milk. In some aspects, the whole milk may be heated, e.g., to 20°C to 50°C (68°F to 122°F), to increase the rate of fat globule combination and accumulation in the cream layer. In certain embodiments, the topical formulation is or includes a cream, e.g., human milk cream, obtained or collected by creaming separation. In some embodiments, the cream is obtained by centrifugation of whole milk. In certain aspects, the centrifugation relies on spinning the whole milk in a centrifuge, e.g., at 5,000 to 10,000 RCF, to produce a cream layer and a skim layer. In certain aspects, the cream fraction may be further centrifuged, e.g., to enrich or concentrate the lipid, fat, or fatty acid content. In certain embodiments, the topical formulation contains a cream, for example, human milk cream, separated or obtained by centrifugation.

[0153] In some embodiments, the human milk cream is homogenized prior to formulation into a topical formulation. In some aspects, homogenization may be performed by any suitable means known as routine, including but not limited to mechanical homogenization, pressure homogenization, or ultrasonic homogenization. In certain embodiments, homogenization of the human milk cream results in an average fat globule size of about 3 μm, 2 μm, 1 μm, 0.5 μm, or 0.1 μm in diameter, or less than 3 μm, 2 μm, 1 μm, 0.5 μm, or 0.1 μm in diameter. In certain embodiments, homogenization of the human milk cream results in an average fat globule size of about 0.5 μm in diameter, or less than 0.5 μm in diameter. In some embodiments, about or at least 60%, 70%, 75%, 80%, or 95% of the milk fat globules of the homogenized cream have a diameter of 0.2 μm to 2 μm.

[0154] In some embodiments, the method includes measuring the fat, lipid, or fatty acid content of the human milk cream. In certain embodiments, the cream fraction may be further processed, such as by centrifugation, to achieve a target fat, lipid, or fatty acid content. In some embodiments, the human milk cream initially contains 20%-60% fat, lipid, or fatty acid content and is further processed, e.g., centrifuged, to increase the fat, lipid, or fatty acid content to a target concentration, e.g., 50%-60%, or at least 70%, 80%, 90%, 95% or more fat, lipid, or fatty acid content (e.g., w / w or w / v).

[0155] In certain embodiments, the production includes one or more steps to isolate, separate, or obtain a fraction (also referred to herein as a subfraction), part, or component of the cream. For example, in some embodiments, the human milk cream is further processed, e.g., centrifuged, to remove additional or residual skim milk present in the cream. In certain embodiments, the human milk cream is centrifuged to produce a fraction or part of the cream that is or contains enriched or concentrated human milk lipids, e.g., compared to unprocessed cream. In some embodiments, centrifugation of the cream produces or results in human milk butter or butter oil. In some embodiments, the human milk butter or butter oil is incorporated into a composition, e.g., a topical formulation. In certain embodiments, the human butter is or comprises at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 99% or more human milk lipids or human milk fats.

[0156] In some embodiments, one or more lipids or a subset of lipids are purified, isolated, separated, or obtained from human milk cream, butter oil, or butter. In certain embodiments, at least one, a plurality, or all or substantially all neutral complex lipids (e.g., free fatty acids, cholesteryl esters, diacylglycerols, triacylglycerols, monoacylglycerols), sphingolipids (e.g., ceramides, dihydroceramides, hexosylceramides, lactosylceramides, sphingomyelins), phospholipids (e.g., phosphatidylcholine, lysophosphatidylcholine, phosphatidylethanolamines, lysophophatidylethanolamines, phosphatidylinositol), or any or all combinations of the foregoing are purified, isolated, separated, or obtained from human milk cream, milk fat, butter, or butter oil, and formulated into a topical formulation.

[0157] In some embodiments, one or more lipids found in or derived from human milk are synthesized, isolated or purified from non-human milk sources and formulated into compositions, such as topical preparations. In some embodiments, non-human milk sources may include, but are not limited to, milk from non-human mammals, such as cows, goats, or camels, or lipids found in or produced by plants. In some embodiments, one or more lipids found in human milk are chemically synthesized from chemicals or lipid precursors and formulated into topical preparations.

[0158] In some embodiments, one or more fatty acids are refined, isolated, separated or obtained from human milk cream, butter or butter oil.In certain embodiments, at least one, a plurality, or all or substantially all of oleic acid, palmitic acid, linoleic acid, omega-3 fatty acid, omega-6 fatty acid, omega-9 fatty acid, stearic acid, myristic acid, lauric acid, palmitoleic acid, vaccenic acid, heptadecanoic acid, myristoleic acid, heptadecene, docosahexaenoic acid, gamma-linolenic acid, docosapentaenoic acid, eicosapentaenoic acid (EPA), arachidonic acid (AA) and docosahexaenoic acid (DHA), or any or all combinations of the above, are refined, isolated, separated or obtained from human milk cream or butter, and formulated into topical preparations.

[0159] In some embodiments, one or more fatty acids found in or derived from human milk are synthesized, isolated or purified from non-human milk sources and formulated into compositions, such as topical preparations.In some embodiments, non-human milk sources may include, but are not limited to, milk from non-human mammals, such as cows, goats, or camels, or lipids found in or produced by plants.In some embodiments, one or more lipids found in human milk are chemically synthesized from chemicals or lipid precursors and formulated into topical preparations. Cream Subfraction

[0160] In certain embodiments, the human milk cream is separated into one or more subfractions. In certain embodiments, the human milk cream subfraction is obtained by removing water and / or non-fat solids. In certain embodiments, the human milk cream subfraction is produced or produced by a process that is or includes centrifugation of the human milk cream. In certain embodiments, the human milk cream is homogenized prior to centrifugation. In some aspects, homogenization may be performed as a routine operation, including, but not limited to, mechanical homogenization, pressure homogenization, or ultrasonic homogenization.

[0161] In certain embodiments, the human milk cream is homogenized prior to centrifugation to produce an average fat globule size of about or less than 3 μm, 2 μm, 1 μm, 0.5 μm, or 0.1 μm in diameter. In some embodiments, the human milk cream is homogenized to produce an average fat globule size of about or less than 0.5 μm in diameter. In some embodiments, about or at least 60%, 70%, 75%, 80%, or 95% of the milk fat globules in the homogenized cream have a diameter between 0.2 μm and 2 μm.

[0162] In some embodiments, the human milk cream is pressure homogenized and the homogenized human milk cream is centrifuged to produce a human milk cream sub-fraction. In some embodiments, the human milk cream sub-fraction is produced or generated by pressure homogenizing the human milk cream at about or at least 1,000 psi, 2,500 psi, 5,000 psi, 10,000 psi, 15,000 psi, 20,000 psi, 25,000 psi, 30,000 psi, 40,000 psi, or 50,000 psi pounds per square inch or pounds force (psi) at a rate of at least 1 mL / min, 10 mL / min, 25 mL / min, 50 mL / min, 60 mL / min, 70 mL / min, 80 mL / min, 90 mL / min, 100 mL / min, 150 mL / min, 200 mL / min, 300 mL / min, 400 mL / min, 500 mL / min, or 1,000 mL / min, and then centrifuging the homogenized human milk cream. In certain embodiments, the human milk cream may be pressure homogenized for one cycle or recirculation. In certain embodiments, the human milk cream may be pressure homogenized for multiple cycles or recirculations, for example, at least 2, 3, 4, 5, or 10 recirculations.

[0163] In certain embodiments, the human milk cream sub-fraction is produced or generated by pressure homogenizing the human milk cream at 10,000 psi to 25,000 psi at a rate of at least 10 mL / min to 100 mL / min for at least one cycle or circulation, and then centrifuging the homogenized human milk cream. In certain embodiments, the human milk cream is pressure homogenized at a pressure of at least or about 20,000 psi at a rate of at least or about 60 mL / min for at least one cycle or circulation.

[0164] In certain embodiments, the human milk cream sub-fraction is produced or generated by homogenizing human milk cream and then centrifuging the homogenized human milk cream at about or at least 100g, 500g, 1,000g, 1,500g, 2,000g, 2,500g, 3,000g, 4,000g, 5,000g, or 10,000g for about or at least 1, 5, 10, 15, 30, 45, 60, 75, or 90 minutes, or for about or at least 1, 2, 3, 6, 12, 18, or 24 hours. In certain embodiments, the homogenized human milk cream is centrifuged at 3500g for 30 minutes, or with an alternative protocol to achieve equivalent sedimentation. Certain embodiments contemplate that one of skill in the art may calculate alternative centrifugation settings or protocols to achieve the same or similar sedimentation results resulting from a given g-force and time.

[0165] In certain embodiments, human milk cream, e.g., homogenized human milk cream, is centrifuged, e.g., at 3500 g for 30 minutes or an equivalent protocol, resulting in separate layers. In certain embodiments, the top layer after centrifugation of human milk cream is the butter oil subfraction of human milk cream. In some embodiments, human milk cream is centrifuged into three layers, including a top layer that is or comprises butter oil, a middle layer that is or comprises human milk fat globule membrane solids, and a bottom layer that is or comprises human skim milk. In certain embodiments, the butter oil fraction contains at least 80%, 90%, 95%, 99%, or 99.8% human milk fatty acids by weight / weight. In certain embodiments, the relative centrifugal force or g force and / or centrifugation time may be increased, such as to increase the fatty acid content and reduce the water or skim milk content of the butter oil layer.

[0166] In some embodiments, the butter oil is obtained or derived from human milk. In some embodiments, the butter oil is obtained or derived from human milk by the Alfa-Laval process. In certain embodiments, the Alfa-Laval process is or includes heating or warming human milk cream, e.g., human milk cream having a fat or fatty acid content of 30-40%, to a temperature such as 55-58°C, and then passing the warmed or heated human milk cream through a pre-concentrator, such as an airtight separator, to concentrate the fat or fatty acid concentration to about 70% to about 75% or more. In some embodiments, the resulting concentrated cream composition is then homogenized, e.g., using a centrifixator, and then passed through a paring chamber, e.g., for phase inversion to a water-in-oil emulsion. In certain embodiments, the fat is separated from the water-in-oil emulsion, e.g., by passing the oil and water emulsion through a concentrator to separate and set aside the fat and / or fatty acids. In some embodiments, remaining residual water may be removed through the use of a vacuum chamber. In some embodiments, butter oil is obtained or derived from human milk by the Westfalia process. In certain embodiments, the Westfalia process is or includes a process for removing non-fat milk solids from human milk cream, such as by placing the human milk cream into a concentrator that removes the non-fat milk solids portion. In certain embodiments, the Westfalia process also includes a process for homogenization, such as breaking the membranes around the fat globules and then centrifuging to remove the aqueous phase during centrifugation. Permeate fraction

[0167] In some embodiments, human skim milk is filtered to produce or result in a retentate fraction and a permeate fraction, such as a human milk retentate and a human milk permeate fraction. In certain embodiments, human skim milk is filtered, such as by ultrafiltration, using a membrane or filter having a pore size capable of filtering substances having a molecular weight of 1 to 10,000 kDa. In some embodiments, skim milk is filtered to obtain a human milk retentate and a human milk permeate. Details of this process can be found, for example, in U.S. Pat. Nos. 8,545,920, 7,914,822, 7,943,315, 8,278,046, 8,628,921, and 9,149,052, as well as WO 2018 / 053535, each of which is incorporated herein by reference in its entirety.

[0168] In certain embodiments, the retentate, e.g., human milk retentate, is a fraction captured during or by ultrafiltration of human skim milk. In certain embodiments, the retentate contains concentrated or enriched milk proteins, e.g., having a high molecular weight relative to the filter. In certain embodiments, the permeate fraction is or is obtained from the liquid that passes through the filter during filtration of human skim milk. In certain embodiments, the permeate contains human milk oligosaccharides. In some embodiments, the human milk oligosaccharides of the permeate fraction are concentrated or enriched relative to the skim fraction or the retentate fraction.

[0169] In some embodiments, the human skim milk is filtered with an ultrafiltration membrane(s). In certain embodiments, the membrane is sized to prevent the passage of any substance having a molecular weight of greater than or about 0.1 kDa, 1 kDa, 5 kDa, 10 kDa, 20 kDa, 25 kDa, 30 kDa, 40 kDa, or 50 kDa. In certain embodiments, the membrane is sized to prevent the passage of substances having a molecular weight of greater than 40 kDa. In certain embodiments, the membrane prevents or captures substances having a molecular weight greater than a molecular weight in the range of 1 kDa to 40 kDa. In some embodiments, the filter is or includes an ultrafilter. In certain embodiments, the filter is or includes a 10 kDa filter. In certain embodiments, the filter is or comprises a 25 kDa filter. In some embodiments, the filter is or comprises a 40 kDa filter. In some embodiments, the membrane has an average or median pore size of less than 5 μm, 1 μm, 0.5 μm, 0.25 μm, 0.2 μm, 0.1 μm, or less. In certain embodiments, the filter is a 0.45 μm filter. In certain embodiments, the filter is a 0.3 μm filter. In certain embodiments, the filter is a 0.2 μm filter. In some embodiments, stepwise filtration can be used (e.g., a first filtration at 0.45 μm, and a second filtration at 0.3 μm, and a third filtration at 0.2 μm, or any combination thereof).

[0170] Other suitable membranes may include ultrafiltration membranes with a molecular weight cut-off of 3.5 kDa or less, such as those from Dow Denmark (Naskov, Denmark). Ultrafiltration membranes made of ceramic materials may also be used. Ceramic filters have an advantage over synthetic filters because they may be sterilized with live steam.

[0171] In some embodiments, the method includes measuring the human milk oligosaccharide content of the human milk permeate. Suitable methods for measuring human milk oligosaccharide content are known and include, but are not limited to, liquid chromatography mass spectrometry (LC-MS) techniques. In certain embodiments, the permeate fraction may be further processed, such as by filtration, e.g., nanofiltration, to achieve a target human milk oligosaccharide content. In some embodiments, the human milk permeate initially contains 0.1%-1.0%, or about 0.1%-1.0% (e.g., w / w or w / v) human milk oligosaccharide content. In some embodiments, the human milk permeate is processed into an HMO composition, such as described herein, to enrich the HMO content and / or to reduce the content of one or more sugars, e.g., lactose.

[0172] In some embodiments, the permeate fraction may be filtered a second time, such as to produce a second retentate fraction. In certain embodiments, the additional filtration further removes proteins and residual lipids or other macromolecules from the permeate. In certain embodiments, the second retentate fraction is added or pooled with the first retentate fraction, such as to increase the biological agent concentration (e.g., protein concentration).

[0173] In certain embodiments, the permeate fraction is measured for HMO content before the step of formulation with one or more excipients. In certain embodiments, the permeate is evaluated to ensure that the HMO content is above a minimum concentration before adding to one or more excipients. In certain embodiments, only the permeate fraction containing at least 0.01%, 0.05%, 0.1%, 0.5%, or 1% HMO (w / w) is formulated with one or more excipients for the manufacture of a topical formulation. In some embodiments, only the permeate fraction containing at least 0.1% HMO (w / w) is formulated with one or more excipients for the manufacture of a topical formulation. In certain embodiments, only the permeate fraction containing at least 0.5% HMO (w / w) is formulated with one or more excipients for the manufacture of a topical formulation. In certain embodiments, only the permeate fraction containing at least 1.0% HMO (w / w) is formulated with one or more excipients for the manufacture of a topical formulation.

[0174] In some embodiments, the permeate is further or additionally processed before mixing, formulating, or adding to one or more excipients. In certain embodiments, the permeate is further processed to enrich or purify HMOs, for example, compared to human whole milk, human skim milk, or a permeate fraction before the additional processing step. In some embodiments, the further or additional processing of the permeate reduces one or more components of the permeate, for example, lactose or minerals. In certain embodiments, the permeate is further or additionally processed to reduce, for example, substantially reduce, the level of lactose, for example, compared to human whole milk, human skim milk, or a permeate fraction before the additional processing step.

[0175] In certain embodiments, suitable additional or further steps for the permeate are described in WO2018053535, which is hereby incorporated in its entirety. In some embodiments, the further or additional processing step is or includes biochemical and / or enzymatic reduction or removal of lactose from the permeate fraction, such as with no or minimal loss of yield or alteration of the molecular profile of the HMO content of the human permeate. In certain embodiments, the permeate is treated to reduce or remove lactose and / or enrich or increase the HMO concentration prior to its incorporation into the topical formulation.

[0176] In certain embodiments, the permeate may be treated to further enrich or purify HMOs, for example, as compared to whole milk, skim milk, or the untreated permeate, and in certain embodiments, such treatment may include additional filtration or chromatographic techniques.

[0177] In some embodiments, the permeate undergoes one or more steps to concentrate or increase the HMO content. In some embodiments, the permeate undergoes filtration, such as using nanofiltration or reverse osmosis, to concentrate or increase the HMO content. In some embodiments, the nanofiltration or reverse osmosis is performed using a nanofilter, such as a filter having a pore size of 1-10 nanometers or about 1-10 nanometers. In certain embodiments, the filter is sized between 100 Da and 1,000 Da, or about 100 Da and 1,000 Da. In certain embodiments, the filter is sized between 400 Da and 600 Da, or about 400 Da and 600 Da. In some embodiments, the filter is sized between 400 Da and 500 Da, or about 400 Da and 500 Da. In certain embodiments, the filter is a 400 kDa filter, or about a 400 kDa filter. In certain embodiments, the filter is a 500 kDa filter or about a 500 kDa filter. In certain embodiments, the permeate is passed through one or more filters, for example, by nanofiltration or reverse osmosis, which collects or prevents the flow of human milk oligosaccharides therethrough. In some such embodiments, the material that does not pass through the filter is enriched for human milk oligosaccharides and collected as a processed permeate for use in the formulation or manufacture of a topical formulation.

[0178] In certain embodiments, the HMO content of the permeate is concentrated or increased to contain at least 0.5%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, or 10% HMOs (w / w or w / v). In some embodiments, the HMO content of the permeate is concentrated to 1%-10%, 2%-12%, or 4%-8%. In certain embodiments, the HMO concentration of the permeate is concentrated to at least 5%.

[0179] In some embodiments, the permeate undergoes one or more treatment steps to remove or reduce lactose content. In some embodiments, lactase enzyme is contacted or added to the permeate, and the resulting permeate / lactase mixture is incubated under conditions that allow for enzymatic digestion of lactate. In some embodiments, such conditions may include, but are not limited to, pH, lactase concentration, temperature, mixing conditions, and duration of time. In certain embodiments, the permeate / lactase mixture is incubated at a pH of 4.3-4.7 or about 4.3-4.7, such as a pH of 4.5, and at a temperature of 48°C-55°C or about 48°C-55°C, such as a temperature of 50°C or about 50°C. In some embodiments, the permeate / lactase mixture contains 0.01%-1.0% or about 0.01%-1.0% lactase (w / w or w / v), such as 0.1% lactase. In certain embodiments, the mixture may be incubated for at least 5, 15, 30, 60, 90, 120, or 180 minutes. In some embodiments, the mixture is incubated for 60 minutes or about 60 minutes. After incubation, in some embodiments, the mixture may be cooled to, for example, 25° C. or about 25° C., such as to 20° C. to about 30° C. or about 20° C. to about 30° C.

[0180] In some embodiments, lactase is removed from the mixture via filtration. In some embodiments, lactase is removed from the mixture using a filter with a pore size that collects or prevents flow through of substances with molecular weights greater than 50 kDa. In some such embodiments, the material that flows through the filter is collected as a processed permeate that is formulated into a topical formulation with one or more excipients. In certain embodiments, filtration removes at least 50%, 75%, 90%, 95%, 99%, 99.9%, or 99.99% of lactase from the mixture.

[0181] In certain embodiments, the treated permeate is further filtered through one or more additional filters, such as an additional filter having a pore size of 2,000 to about 3,000 Daltons or about 2,000 to about 3,000 Daltons. In some embodiments, the one or more additional filters comprise a membrane having a pore size of <600 Daltons. In some embodiments, the composition is filtered through both filters, including a membrane of about 2,000 to about 3,000 Daltons followed by filtration through a membrane of <600 Daltons.

[0182] In some embodiments, the permeate fraction or a portion thereof is or comprises a purified HMO composition, e.g., an HMO composition having enriched or purified HMOs compared to human whole milk or human permeate without additional processing or filtration. In some embodiments, a purified HMO composition has reduced levels of lactose and minerals, e.g., compared to human whole milk or human permeate without additional processing or filtration.

[0183] In certain embodiments, the permeate or a fraction or portion thereof, e.g., an HMO composition, is stored below 37° C., below 25° C., below 10° C., or at or about 2° C. to 8° C., e.g., at or about 4° C. In some embodiments, the permeate or a fraction or portion thereof is frozen and stored. In some embodiments, the permeate or a fraction or portion thereof is stored until further use, e.g., incorporation into a topical formulation.

[0184] In some embodiments, the pH of the human milk permeate, or a fraction or portion thereof, is adjusted prior to incorporation into a topical formulation. In certain embodiments, the pH of the human milk permeate, or a fraction or portion thereof, is adjusted to an acid pH. In certain embodiments, the pH of the human milk permeate, or a fraction or portion thereof, is adjusted to a pH of 5.0-7.0, 5.0-6.0, or 5.3-5.7. In certain embodiments, the pH of the human milk permeate is adjusted to 5.2 or about 5.2. In some embodiments, the pH of the human milk permeate is adjusted to 5.5 or about 5.5. Retentate fraction

[0185] In certain embodiments, the retentate fraction is or contains proteins and compounds having a molecular weight greater than a value between 1 kDa and 10,000 kDa, such as between 1 kDa and 200 kDa or between about 1 kDa and 200 kDa. In certain embodiments, the retentate fraction is enriched for proteins and compounds having a molecular weight greater than a value between 1 kDa and 10,000 kDa, such as between 1 kDa and 200 kDa or between about 1 kDa and 200 kDa, for example, compared to the skim or permeate fractions. In some embodiments, the retentate is about 0.5 kDa, 1 kDa, 2 kDa, 3 kDa, 4 kDa, 5 kDa, 10 kDa, 15 kDa, 20 kDa, 25 kDa, 30 kDa, 40 kDa, 50 kDa, 60 kDa, 70 kDa, 75 kDa, or 100 kDa. Da, 30 kDa, 40 kDa, 50 kDa, 60 kDa, 70 kDa, 75 kDa, or 100 kDa, or greater than 0.5 kDa, 1 kDa, 2 kDa, 3 kDa, 4 kDa, 5 kDa, 10 kDa, 15 kDa, 20 kDa, 25 kDa, 30 kDa, 40 kDa, 50 kDa, 60 kDa, 70 kDa, 75 kDa, or 100 kDa. In some embodiments, the retentate comprises human milk substances, e.g., proteins, having a molecular weight greater than 10 kDa, 5 kDa, 3.5 kDa, 2.5 kDa, or 1 kDa, or less than 10 kDa, 5 kDa, 3.5 kDa, 2.5 kDa, or 1 kDa. In certain embodiments, the retentate comprises human milk substances, e.g., proteins, having a molecular weight of 40 kDa, about 40 kDa, or at least 40 kDa. In some embodiments, the retentate comprises human milk substances, e.g., proteins, having a molecular weight of less than 3.5 kDa.

[0186] In some embodiments, the human milk proteins collected or captured by the ultrafilter include one or more of lactalbumin (14 kDa), casein (23 kDa), lactoglobulin (37 kDa), albumin (65 kDa), and immunoglobulins (>100 kDa) (with approximate molecular weights listed in parentheses). In some embodiments, the retentate includes or contains one or more of human lactalbumin, human casein, human lactoglobulin, human albumin, and human immunoglobulin. In certain embodiments, the human retentate is or includes enriched or purified human milk proteins, such as human lactalbumin, human casein, human lactoglobulin, human albumin, or human immunoglobulin, as compared to whole human milk.

[0187] In certain embodiments, the retentate, e.g., human milk retentate fraction, is stored at 2° C. to 8° C. or about 2° C. to 8° C., such as below 37° C., below 25° C., below 10° C., or at or about 4° C. In some embodiments, the human retentate is frozen and stored. In certain aspects, the human retentate is stored until subsequent use, e.g., incorporation into a topical formulation, or further fractionation to generate a permeate fraction and a retentate fraction. Exemplary Methods

[0188] In some embodiments, the method of making a topical formulation is or includes isolating, separating, purifying, or obtaining one or more fractions, parts, or components from human milk, for example, whole human milk pooled from the milk of multiple individual donors. In some embodiments, the whole milk is processed, such as by centrifugation or creaming separation to collect, separate, or derive human milk cream and human skim milk. In some embodiments, the lipid, fat, or fatty acid content of the cream is measured and optionally further centrifuged, such as to increase the lipid, fat, or fatty acid content to a target value or range of values ​​for lipid, fat, or fatty acid content, for example, 50%-60% w / v or w / w. In certain embodiments, the fat content of the cream is measured and optionally further centrifuged, such as to increase the fat content to a target value or range, for example, at least 60%, 70%, 80%, 90% w / w or more. In certain embodiments, the skim milk may optionally be further centrifuged to further remove fat, lipid, or fatty acid content, such as to produce skim having a total fat, lipid, or fatty acid content of less than 1%, 0.9%, 0.8%, 0.7%, 0.6%, 0.5%, 0.4%, 0.3%, 0.2%, or 0.1%, e.g., w / w or w / v.

[0189] In some embodiments, the method of making a topical formulation is or includes isolating, separating, purifying, or obtaining one or more fractions from whole human milk pooled from the milk of multiple individual donors. In some embodiments, the human milk is pooled from the milk of at least 10, 20, 30, 40, 50, 75, or 100 individual donors. In certain embodiments, the human milk is pooled from milk from 50-200 donors.

[0190] In certain embodiments, human skim milk is filtered, e.g., ultrafiltered, to produce a human milk permeate fraction and a human milk retentate fraction. In certain embodiments, the oligosaccharide (e.g., HMO) content of the human milk permeate may be measured, e.g., to ensure or confirm that the permeate contains a threshold HMO content, e.g., of at least 0.1%, 0.5%, 1%, or 5% HMO (w / w or w / v), and in some cases may be optionally processed, e.g., by an additional filtration step, to increase the HMO concentration, e.g., to achieve a target value or range of HMO content of at least 0.5%, 1%, or 5% HMO (w / w or w / v).

[0191] In certain embodiments, the protein content of the human milk retentate may be measured, such as to ensure or confirm that the retentate contains a threshold protein content, e.g., at least 1%, 3%, 5%, 6%, 7%, 8%, 9%, or 10%, or 5% to 8% or 6% to 7% total protein content (e.g., w / w or w / v), and in some cases the human milk retentate may optionally be processed, e.g., by an additional filtration step, to increase the protein concentration to achieve a target value or range of protein content, e.g., a total protein content of at least 6%, 7%, 8%, 9%, or 10% w / w or w / v.

[0192] In some embodiments, human whole milk or one or more human milk fractions, portions, or components thereof are added to, mixed with, or combined with one or more pharma- ceutically acceptable excipients to produce, formulate, or produce a topical formulation. In some embodiments, human whole milk or one or more human milk fractions, portions, or components thereof are formulated into a topical formulation that is an oil-in-water emulsion, a water-in-oil emulsion, a nanoemulsion, an emulsified gel, an oily ointment, a hydrocolloid system, a cream preparation, or any other pharma- ceutically acceptable formulation suitable for use as a topical formulation.

[0193] In some embodiments, human milk cream is added to or incorporated into a topical formulation to contain a target human milk cream content. In some embodiments, human milk cream is added to a topical formulation in an amount sufficient to contain or include at least 5%, 10%, 20%, 25%, 30%, 40%, or 50%, or 1%-75%, 5%-50%, 10%-50%, 10%-30%, 10%-20%, 30%-50%, 40%-60%, or 50%-75% (e.g., w / w or w / v, inclusive) of human milk cream. In certain embodiments, human milk cream is added to or incorporated into a topical formulation to contain, for example, at least or about, 10%-30% (inclusive) of human milk cream, w / v or w / w. In certain embodiments, human milk cream is added or incorporated into a topical formulation, for example, to contain about 20% human milk cream, w / v or w / w.

[0194] In some embodiments, human milk cream is added to or incorporated into a topical formulation to contain the target fatty acid, fat, or lipid content. In some embodiments, human milk cream is added to a topical formulation in an amount sufficient to contain or include at least 5%, 10%, 20%, 25%, or 30%, or 5%-25%, 10%-20%, 1%-20%, or 5%-15% human milk lipid, fat, or fatty acid content, for example, w / w or w / v. In certain embodiments, human milk cream is added to or incorporated into a topical formulation to contain or include about 20% human milk lipid, fat, or fatty acid. In certain embodiments, human milk cream is added to or incorporated into a topical formulation to contain or include about 10% human milk lipid, fat, or fatty acid.

[0195] In certain embodiments, human milk permeate is added or incorporated into a topical formulation to contain a target human milk permeate content. In some embodiments, human milk permeate is added to a topical formulation in an amount sufficient to contain or include at least 1%, 10%, 20%, 30%, 40%, 50%, 60%, or 70% human milk permeate, for example, w / w or w / v. In certain embodiments, human milk permeate is formulated into a topical formulation by replacing an amount of human milk permeate with the amount of water typically added to a standard topical formulation, such as an oil-in-water emulsion, a water-in-oil emulsion, a nanoemulsion, an emulsified gel, an oily ointment, a hydrocolloid system, a cream preparation, or any other pharma- ceutically acceptable formulation suitable for use as a topical formulation, typical or known formulation. Thus, where water is typically added to produce a topical formulation, such as an oil-in-water emulsion, water-in-oil emulsion, nanoemulsion, emulsified gel, oil-based ointment, hydrocolloid system, or cream preparation, that amount of human milk permeate is added in place of water to produce the topical formulation. In certain embodiments, the permeate contains at least 0.5% HMO (w / v or w / w).

[0196] In certain embodiments, HMO compositions, such as HMO compositions derived from human milk permeate as described herein, are added or incorporated into topical formulations to contain a target HMO composition content. In some embodiments, the HMO composition is added in an amount sufficient for the topical formulation to contain or include at least 1%, 10%, 20%, 30%, 40%, 50%, 60%, or 70% (e.g., w / w or w / v) HMO composition. In certain embodiments, the HMO composition is formulated into a topical formulation by substituting an amount of human milk permeate for the amount of water typically added to standard topical formulations, such as oil-in-water emulsions, water-in-oil emulsions, nanoemulsions, emulsified gels, oily ointments, hydrocolloid systems, cream preparations, or any other pharma- ceutically acceptable formulations suitable for use as topical formulations, typical or known formulations. In certain embodiments, the HMO composition contains at least 5% HMO (w / v or w / w).

[0197] In some embodiments, human milk retentate is added or incorporated into a topical formulation to contain a target human milk protein content. In some embodiments, human milk retentate is added to a topical formulation in an amount sufficient to contain or include, for example, at least 1%, 5%, 10%, 20%, 25%, 30%, 40%, 50%, 60%, or 70% or more human milk retentate, w / w or w / v. In certain embodiments, human milk retentate is added or incorporated into a topical formulation to contain, for example, at least or about, 10%-30% human milk retentate, w / v or w / w. In certain embodiments, human milk retentate is added or incorporated into a topical formulation to contain, for example, 0.1%-20%, 1%-5%, or 0.1%-2% human milk retentate, w / v or w / w.

[0198] In some embodiments, human milk retentate is added to or incorporated into a topical formulation to contain a target protein content. In some embodiments, human milk retentate is added to a topical formulation in an amount sufficient to contain or include at least 0.1%, 0.5%, 1%, 2%, 3%, 4%, or 5% human milk protein content, for example, w / w or w / v.

[0199] In some embodiments, the topical formulation is formulated with human milk cream or a subfraction thereof, such as a butter oil fraction or a milk fat globule membrane solid fraction. In certain embodiments, the topical formulation is formulated with human milk cream or a subfraction thereof, and human milk permeate or a portion or fraction thereof. In some embodiments, the topical formulation is formulated with human milk cream or a subfraction thereof to contain a fat, fatty acid, or lipid content of 10%-30% w / w. In certain embodiments, the topical formulation is formulated with one or more excipients, such as one or more emulsifiers, one or more preservatives, one or more thickeners, one or more surfactants, one or more antioxidants, and / or one or more hydrocolloids. In certain embodiments, the topical formulation is formulated with human milk cream or a subfraction thereof, in an amount sufficient for a final concentration of fat, fatty acid, or lipid of 10%-30% w / w, mixing the contents, and then adding a hydrocolloid or gum while mixing. In certain embodiments, the contents are mixed with a mechanical disruptor at a speed of at least 500 rpm, 1,000 rpm, 5,000 rpm, 10,000 rpm, 25000 rpm, 50,000 rpm or more for at least 5 minutes, 15 minutes, 30 minutes, 60 minutes, 90 minutes, 120 minutes, 150 minutes, 180 minutes, 2 hours, 3 hours, 4 hours, 6 hours, 8 hours, or 12 hours. In certain embodiments, xanthan gum is added while mixing at a low speed, for example, 5,000 rpm, 2,000 rpm, 1,000 rpm, or 500 rpm or less.

[0200] In certain embodiments, topical formulations are formulated with human milk cream or a subfraction thereof, such as a butter oil fraction or a milk fat globule membrane solid subfraction, human milk permeate or a fraction or portion thereof, and one or more excipients that are or include an emulsifier, a preservative, a thickener, a non-ionic surfactant, an antioxidant, and / or a hydrocolloid. In certain embodiments, topical formulations are formulated by adding human milk cream or a subfraction thereof to the human milk permeate and the emulsifier, the preservative, the thickener, the non-ionic surfactant, the antioxidant in an amount sufficient for a final concentration of fat, fatty acid, or lipid of 10%-30% w / w, mixing the contents, and then adding a hydrocolloid while mixing. In certain embodiments, the emulsifier is or includes TWEEN 80 or polysorbate 80. In some embodiments, the preservative is or includes phenoxyethanol. In certain embodiments, the thickening agent is or includes liquid paraffin. In some embodiments, the antioxidant is or includes alpha tocopherol. In certain embodiments, the hydrocolloid is or includes xanthan gum. In certain embodiments, the topical formulation is formulated by adding human milk cream or a subfraction thereof to human milk permeate and Tween 80, phenoxyethanol, liquid paraffin, Span 83, and alpha tocopherol in an amount sufficient for a final concentration of fat, fatty acid, or lipid of 10% to 30% w / w, mixing the contents, and then adding xanthan gum while mixing. In certain embodiments, the contents are mixed with a mechanical disruptor at a speed of at least 500 rpm, 1,000 rpm, 5,000 rpm, 10,000 rpm, 25000 rpm, 50,000 rpm or more for at least 5 minutes, 15 minutes, 30 minutes, 60 minutes, 90 minutes, 120 minutes, 150 minutes, 180 minutes, 2 hours, 3 hours, 4 hours, 6 hours, 8 hours, or 12 hours. In certain embodiments, xanthan gum is added while mixing at a low speed, for example, 5,000 rpm, 2,000 rpm, 1,000 rpm, or 500 rpm or less.

[0201] In certain embodiments, topical formulations are formulated with human milk cream and human milk permeate. In some embodiments, human milk cream is added or incorporated into the topical formulation to contain a target fat, fatty acid, or lipid content in an amount sufficient for the topical formulation to contain or include at least 5%, 10%, 20%, 25%, or 30%, or 5%-25%, 10%-20%, 1%-20%, or 5%-15% human milk lipid, fat, or fatty acid content, for example, w / w or w / v. In certain embodiments, human milk cream is added or incorporated into the topical formulation to contain at least 5%, 10%, 20%, 25%, or 30%, or 5%-25%, 10%-20%, 1%-20%, or 5%-15% human milk fat target human milk fat content, for example, w / w. In certain embodiments, the human milk permeate is formulated into a topical formulation by substituting an amount of human milk permeate for the amount of water typically added to a typical or known formulation for a standard topical formulation, such as an oil-in-water emulsion, a water-in-oil emulsion, a nanoemulsion, an emulsified gel, an oily ointment, a hydrocolloid system, a cream preparation, or any other pharma- ceutically acceptable formulation suitable for use as a topical formulation. In certain embodiments, the topical formulation is formulated with an amount of human milk cream sufficient for the topical formulation to contain about 10% human milk lipids, fats, or fatty acids, and with the human milk permeate in place of water, for example, in an amount sufficient for the topical formulation to contain 0.1%-0.5% HMOs w / w or w / v. In some embodiments, the topical formulation is formulated using an amount of human milk cream sufficient for the topical formulation to contain about 20% human milk fat, and using human milk permeate in place of water, e.g., in an amount sufficient for the topical formulation to contain 0.1%-0.5% HMO (e.g., w / w). IV. I. Preparations Containing Human Milk Fractions, Portions, or Components

[0202] In some embodiments, the topical formulation provided may be formulated as a cosmetic or pharmaceutical composition suitable for topical administration to a subject, for example, topically and / or directly to the skin.In certain embodiments, the topical formulation may be formulated into one or more of a lotion, gel, emulsion, oil-in-water emulsion, water-in-oil emulsion, nanoemulsion, emulsified gel, oily ointment, hydrocolloid system, cream preparation, serum, nanoemulsion, or powder.In certain embodiments, the topical formulation may be formulated as an oil-in-water emulsion, nanoemulsion, emulsified gel, oily ointment, hydrocolloid system, or cream preparation.

[0203] In some embodiments, the topical formulation is stable or remains stable, e.g., does not undergo phase separation, changes in chemical composition, oxidation of fatty acids, changes in color, odor, or appearance, or changes in viscosity over a period of time. In some embodiments, the topical formulation is stable or remains stable when refrigerated, e.g., stored below 10°C, or at or about 2°C to 8°C, at about 4°C, etc. In certain embodiments, the topical formulation is stable for at least 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, 8 weeks, 9 weeks, 10 weeks, 12 weeks, 14 weeks, 2 months, 3 months, 4 months, 6 months, 12 months, 18 months, 1 year, 2 years, or 5 years when refrigerated, e.g., stored below 10°C, or at or about 2°C to 8°C, at about 4°C, etc. In some embodiments, the topical formulation is stable for at least 3 months when refrigerated. In certain embodiments, the topical formulation is stable for at least six months when refrigerated. In some embodiments, the topical formulation is stable for at least one year when refrigerated.

[0204] In certain embodiments, the topical formulation is stable or remains stable when stored at room temperature, e.g., at or about 20°C to 30°C, at or about 20°C to 30°C, at or about 23°C to 27°C, or at or about 25°C. In some embodiments, the topical formulation is stable at room temperature, e.g., at or about 20°C to 30°C, at or about 23°C to 27°C, or at or about 25°C, for at least 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, 8 weeks, 9 weeks, 10 weeks, 12 weeks, 14 weeks, 2 months, 3 months, 4 months, 6 months, 12 months, 18 months, 1 year, 2 years, or 5 years. In some embodiments, the topical formulation is stable at room temperature for at least 3 months. In some embodiments, the topical formulation is stable at room temperature for at least 6 months. In certain embodiments, the topical formulation is stable at room temperature for at least 1 year.

[0205] In some embodiments, the topical formulation contains one or more parts, fractions, or components of human milk and is formulated or configured as a cream preparation, an oil-in-water emulsion, a water-in-oil emulsion, a nanoemulsion, a hydrocolloid system, an emulsified gel, or an oily ointment. In some embodiments, the topical formulation comprises human milk cream, or a part or component thereof. In certain embodiments, the topical formulation comprises human milk permeate, for example, from ultrafiltration of human milk skim, or a part or component thereof. In certain embodiments, the topical formulation comprises human milk cream and human milk permeate, and is formulated or configured as a cream preparation, an oil-in-water emulsion, a water-in-oil emulsion, a nanoemulsion, a hydrocolloid system, an emulsified gel, or an oily ointment.

[0206] In some embodiments, topical formulations may be delivered by various applicators suitable for topical and general application. Such applicators may include a dropper, applicator wand, cotton swab, or any other suitable device. It should be understood that the compositions herein may also be applied topically and / or directly by using a finger or in other conventional or routine manners. In some embodiments, topical formulations are administered topically, for example, to all or an area of ​​the skin or mucous membrane. In certain embodiments, topical formulations are administered directly, for example, to all or an area of ​​the skin or mucous membrane.

[0207] In certain embodiments, the topical formulation comprises one or more excipients, such as, but not limited to, hydrocolloids, emulsifiers, structuring agents, antioxidants, silicone-containing compounds, thickening agents, preservatives, additional skin conditioning agents, emollients, humectants, or essential oils.

[0208] In some embodiments, topical formulations include one or more excipients that are or include one or more hydrocolloids.Hydrocolloids, also referred to herein and in the art as gums, are in some aspects hydrophilic polymers of plant, animal, microbial, or synthetic origin, which generally contain numerous hydroxyl groups.In some embodiments, hydrocolloids are or include polyelectrolytes.Hydrocolloids are in certain aspects well known in the art, and include, but are not limited to, xanthan gum, cellulose, carboxymethylcellulose, hydroxyethylcellulose, hydroxypropylcellulose, methyl and ethylcellulose, natural gums, natural starch, and deionized starch (e.g., starch octenyl succinate). In certain embodiments, non-limiting examples of gums that can be used with the provided compositions and formulations include acacia, agar, algin, alginic acid, ammonium alginate, amylopectin, calcium alginate, calcium carrageenan, carnitine, carrageenan, dextrin, gelatin, gellan gum, guar gum, guar hydroxypropyltrimonium chloride, hectorite, hyaluroinic acid, silicic acid, hydroxypropyl chitosan, hydroxypropyl guar, karaya gum, kelp, locust bean gum, natto gum, potassium alginate, potassium carrageenan, propylene glycol alginate, sclerotium gum, sodium carboxymethyl dextran, sodium carrageenan, tragacanth gum, xanthan gum, and mixtures thereof.

[0209] In certain embodiments, the topical formulation provided comprises one or more excipients that are or contain emulsifiers.In certain embodiments, the addition of emulsifiers reduces the interfacial tension between phases, improving the formulation and stability of emulsions.In certain embodiments, emulsifiers include nonionic, cationic, anionic, and zwitterionic emulsifiers (see McCutcheon in "Detergents and Emulsifiers" North American Ed. 1986, U.S. Patent Nos. 5,011,681, 4,421,769, and 3,755,560). Suitable emulsifiers include, but are not limited to, esters of glycerin, esters of propylene glycol, fatty acid esters of polyethylene glycol, fatty acid esters of polypropylene glycol, esters of sorbitol, esters of sorbitan anhydride, carboxylic acid copolymers, esters and ethers of glucose, ethoxylated ethers, ethoxylated alcohols, alkyl phosphates, polyoxyethylene fatty ether phosphates, fatty acid amides, acyl lactylates, soaps, TEA stearate, DEA oleth-3 phosphate, polyethylene glycol 20 sorbitan monolaurate (also known as polysorbate 20, Tween 20), polyethylene glycol 5 soy sterol, steareth-2, steareth-20, steareth-21, ceteareth-20, PPG-2 methyl glucose ether distearate, ceteth-10, polysorbate 80, cetyl phosphate, potassium cetyl phosphate, diethanolamine cetyl phosphate, polysorbate 60, glyceryl stearate, PEG-100 stearate, and mixtures thereof. In some embodiments, the emulsifier is or includes paraffin, such as soft white paraffin, liquid paraffin, cetomacrogol 1000, Tween 60, Span 60, Brij S2 (Brij 72), Brij S721 (Brij 721), and / or ascorbyl palmitate.

[0210] In some embodiments, the topical formulation comprises and / or is formulated with a surfactant. In some embodiments, the surfactant is or comprises a non-ionic surfactant. The non-ionic surfactant may include, but is not necessarily limited to, alkyl polyglycosides, sorbitan esters, methyl glucoside esters, alcohol ethoxylates, or polyglycol esters. In some embodiments, the non-ionic surfactant is Span 83 or Span 60. In some embodiments, the surfactant is or comprises a cationic surfactant. The cationic surfactant may include, but is not limited to, arginine methyl esters, alkanolamines, and alkylenediamides. In certain embodiments, the surfactant is an anionic surfactant. Anionic surfactants may include, but are not limited to, alkali metal alkyl sulfates, alkyl or alkylaryl sulfonates, linear or branched alkyl ether sulfates and sulfonates, alcohol polypropoxylated and / or polyethoxylated sulfates, alkyl or alkylaryl disulfonates, alkyl disulfates, alkyl sulfosuccinates, alkyl ether sulfates, linear and branched ether sulfates, and mixtures thereof.

[0211] In certain embodiments, the topical formulation contains one or more excipients that are or include a structuring agent. In certain embodiments, the structuring agent helps provide rheological properties to the composition and / or contributes to the stability of the composition. In some embodiments, the structuring agent functions as an emulsifier or surfactant. Suitable structuring agents for use as excipients include, but are not limited to, stearic acid, palmitic acid, stearyl alcohol, cetyl alcohol, behenyl alcohol, stearic acid, palmitic acid, polyethylene glycol ether of stearyl alcohol having an average of about 1 to about 21 ethylene oxide units, polyethylene glycol ether of cetyl alcohol having an average of about 1 to about 5 ethylene oxide units, and mixtures thereof.

[0212] In some embodiments, the topical formulation contains one or more excipients that are or include antioxidants. In some aspects, the antioxidants include one or more of acetylcysteine, ascorbic acid polypeptide, ascorbyl dipalmitate, ascorbyl methylsilanol pectate, ascorbyl palmitate, ascorbyl stearate, BHA, BHT, t-butylhydroquinone, cysteine, cysteine ​​HCl, diamylhydroquinone, di-t-butylhydroquinone, dicetyl thiodipropionate, dioleyl tocopheryl methylsilanol, disodium ascorbyl sulfate, distearyl thiodipropionate, ditridecyl thiodipropionate, dodecyl gallate, erythorbic acid, esters of ascorbic acid, ethyl ferulate, ferulic acid, gallic acid esters, hydroquinone, isooctyl thioglycolate, kojic acid, magnesium ascorbate, magnesium ascorbyl phosphate, methylsilanol ascorbate, green tea. or natural botanical antioxidants such as grape seed extract, nordihydroguaiaretic acid, octyl gallate, phenylthioglycolic acid, potassium ascorbyl tocopheryl phosphate, potassium sulfite, propyl gallate, quinones, rosmarinic acid, sodium ascorbate, sodium bisulfite, sodium erythrobate, sodium metabisulfite, sodium sulfite, superoxide dismutase, sodium thioglycolate, sorbityl furfural, thiodiglycol, thiodiglycolamide, thiodiglycolic acid, thioglycolic acid, thiolactic acid, thiosalicylic acid, tocophereth-5, tocophereth-10, tocophereth-12, tocophereth-18, tocophereth-50, tocopherol, tocophersolan, tocopheryl acetate, tocopheryl linoleate, tocopheryl nicotinate, tocopheryl succinate, and tris(nonylphenyl) phosphite.

[0213] In some embodiments, topical formulations contain one or more excipients that are or contain silicone-containing compounds.In some aspects, silicone-containing compounds include any member of the polymeric product family whose molecular backbone consists of alternating silicon and oxygen atoms with side groups bonded to the silicon atoms.In some aspects, depending on the -Si-O- chain length, side groups, and crosslinking of silicone-containing compounds, silicones can be synthesized into a wide variety of materials.Their consistency can vary from liquid to gel to solid.

[0214] In certain embodiments, the silicone-containing compound includes, but is not limited to, silicone oils (e.g., volatile and non-volatile oils), gels, and solids.In certain embodiments, the silicon-containing compound is or includes silicone oils, such as polyorganosiloxanes.In certain embodiments, the polyorganosiloxane is or includes dimethicone, cyclomethicone, polysilicone-11, phenyltrimethicone, trimethylsilyl amodimethicone, stearoxytrimethylsilane, or mixtures of these and other organosiloxane materials.In certain embodiments, the polyoranosiloxanes and organosiloxane materials may be mixed in any given ratio to achieve the desired consistency and application properties, depending on the intended application (e.g., on a particular area, such as skin, hair, or eyes). In some embodiments, "volatile silicone oil" is or includes silicone oils that have low heat of vaporization, i.e., typically less than about 50 calories per gram of silicone oil. In some aspects, suitable silicone oils may include, but are not limited to, cyclomethicones such as Dow Corning 344 Fluid, Dow Corning 345 Fluid, Dow Corning 244 Fluid, and Dow Corning 245 Fluid, Volatile Silicon 7207 (Union Carbide Corp., (Danbury, Connecticut, USA)), low viscosity dimethicones, i.e., dimethicones with a viscosity of about 50 cst or less (e.g., dimethicones such as Dow Corning 200-0.5 cst Fluid). Dow Corning Fluid is available from Dow Corning Corporation (Midland, Michigan, USA).Cyclomethicone and dimethicone are described in the CTFA Cosmetic Ingredient Dictionary, Third Edition (incorporated by reference) as a mixture of cyclic dimethylpolysiloxane compounds and fully methylated linear siloxane polymers, respectively, endblocked with trimethylsiloxy units. In certain embodiments, suitable volatile silicone oils also include those available from General Electric Co., Silicone Products Div., (Waterford, New York, USA) and Stauffer Chemical Co., SWS Silicones Div. (Adrian, Michigan, USA).

[0215] In some embodiments, topical formulations are or contain one or more thickening agents.In certain embodiments, thickening agents may include, but are not limited to, carboxylic acid polymers, crosslinked polyacrylate polymers, polyacrylamide polymers, polysaccharides, and gums.Examples of carboxylic acid polymers include crosslinked compounds that contain one or more monomers derived from acrylic acid, substituted acrylic acid, and the salts and esters of these acrylic acids, and substituted acrylic acids, where the crosslinker contains two or more carbon-carbon double bonds and is derived from polyhydric alcohols (see U.S. Patent Nos. 5,087,445, 4,509,949, 2,798,053, CTFA International Cosmetic Ingredient Dictionary, Fourth edition, 1991, pp.12 and 80). Examples of commercially available carboxylic acid polymers include carbomers (e.g., Carbopol™ 900 series from BFGoodrich), which are homopolymers of acrylic acid crosslinked with allyl ethers of sucrose or pentaerythritol. In certain embodiments, the thickener is or includes carabopol, carrageenan, HPC, paraffin wax, PEG4000, sodium alginate, and Avacel CL-611.

[0216] Non-limiting examples of crosslinked polyacrylate polymers include cationic and nonionic polymers, examples of which are described in U.S. Patent Nos. 5,100,660, 4,849,484, 4,835,206, 4,628,078, and 4,599,379.

[0217] Non-limiting examples of polyacrylamide polymers (including non-ionic polyacrylamide polymers, including substituted branched or unbranched polymers) include polyacrylamide, isoparaffin and laureth-7, multiblock copolymers of acrylamide, and substituted acrylamides with acrylic acid and substituted acrylic acid.

[0218] Non-limiting examples of polysaccharides include cellulose, carboxymethyl hydroxyethyl cellulose, cellulose acetate propionate carboxylate, hydroxyethyl cellulose, hydroxyethyl ethyl cellulose, hydroxypropyl cellulose, hydroxypropyl methyl cellulose, methyl hydroxyethyl cellulose, microcrystalline cellulose, sodium cellulose sulfate, and mixtures thereof. Another example is alkyl-substituted cellulose in which the hydroxy groups of the cellulose polymer are hydroxyalkylated (preferably hydroxyethylated or hydroxypropylated) to form hydroxyalkylated cellulose, which is then further modified with C10-C30 linear or branched alkyl groups via ether linkages. Typically, these polymers are ethers of hydroxyalkyl cellulose and C10-C30 linear or branched alcohols. Other useful polysaccharides include scleroglucan, which contains linear chains of (1-3) linked glucose units and (1-6) linked glucose units every 3 units.

[0219] In certain embodiments, topical formulations contain one or more preservatives.Non-limiting examples of preservatives that can be used in the formulation of provided topical formulations include quaternary ammonium preservatives such as polyquaternium-1 and benzalkonium halides (e.g., benzalkonium chloride ("BAC") and benzalkonium bromide), parabens (e.g., methylparaben and propylparaben), phenoxyethanol, benzyl alcohol, chlorobutanol, phenol, sorbic acid, thimerosal, or combinations thereof.

[0220] In certain embodiments, topical formulations are formulated with additional skin conditioning agents. In some aspects, such agents may include materials that may adhere to the skin to reduce flaking, restore softness, and generally improve the appearance of the skin, as well as substances that enhance the appearance of dry or damaged skin.Specific examples of skin conditioning agents include acetylcysteine, N-acetyldihydrosphingosine, acrylates / behenyl acrylate / dimethicone acrylate copolymer, adenosine, adenosine cyclic phosphate, adenosine phosphate, adenosine triphosphate, alanine, egg white, algae extract, allantoin and derivatives, aloe barbadensis extract, amyloglucosidase, arbutin, arginine, bromelain, buttermilk powder, butylene glycol, calcium gluconate, carbocysteine, carnosine, beta Carotene, Casein, Catalase, Cephalin, Ceramide, Chamomilla Recutita (Matricaria) Flower Extract, Cholecalciferol, Cholesteryl Esters, Coco-Betaine, Modified Corn Starch, Crystallin, Cycloethoxymethicone, DNA Cysteine, Cytochrome C, Darutoside, Dextran Sulfate, Dimethicone Copolyol, Dimethylsilanol Hyaluronate, Elastin, Elastin Amino Acids, Ergocalciferol, Ergosterol, Fibronectin, Folic Acid, Gelatin, Gliadin, Beta-Glucan, Glucose Glycine, glycogen, glycolipids, glycoproteins, glycosaminoglycans, sphingoglycolipids, horseradish peroxidase, hydrogenated proteins, hydrolyzed proteins, jojoba oil, keratin, keratin amino acids, kinetin, lactoferrin, lanosterol, lecithin, linoleic acid, linolenic acid, lipase, lysine, lysozyme, malt extract, maltodextrin, melanin, methionine, niacin, niacinamide, oat amino acids, oryzanol, palmitoyl hydrolyzed protein, pancreatin, papain, polyethylene The active ingredients in the extract may include, but are not limited to, glycerin, pepsin, phospholipids, phytosterols, placental enzymes, placental lipids, pyridoxal 5-phosphate, quercetin, resorcinol acetate, riboflavin, Saccharomyces lysate extract, silk amino acids, sphingolipids, stearamidopropyl betaine, stearyl palmitate, tocopherol, tocopheryl acetate, tocopheryl linoleate, ubiquinone, Vitis vinifera (grape) seed oil, wheat amino acids, xanthan gum, zinc gluconate, or combinations thereof.In some embodiments, skin conditioning agents other than those listed above may be incorporated or formulated into the provided topical formulations, as would be readily understood by one of ordinary skill in the art.

[0221] In certain embodiments, the topical formulation contains one or more emollients. In some aspects, emollients are ingredients that may help the skin maintain a soft, smooth, and supple appearance. In some embodiments, emollients include acetyl arginine, acetylated lanolin, algae extract, apricot kernel oil polyethylene glycol-6 ester, avocado oil polyethylene glycol-11 ester, bis-polyethylene glycol-4 dimethicone, butoxyethyl stearate, C 18 C 36 Acid glycol ester, C 12 C 13 Alkyl lactate, caprylyl glycol, cetyl esters, cetyl laurate, coconut oil polyethylene glycol-10 ester, C 12 C 13Dialkyl tartrate, diethyl sebacate, dihydrocholesteryl butyrate, dimethiconol, dimyristyl tartrate, disteareth-5 lauroyl glutamate, ethyl avocado fatty acid, ethylhexyl myristate, glyceryl isostearate, glyceryl oleate, hexyldecyl stearate, hexyl isostearate, hydrogenated palm glycerides, hydrogenated soy glycerides, hydrogenated tallow glycerides, isostearyl neopentanoate, isostearyl palmitate, isotridecyl isononanoate, laureth-2 acetate, lauryl polyglyceryl-6 cetearyl glycol ether, methyl gluceth-20 benzoate, mineral oil, These may include, but are not limited to, myreth-3 palmitate, octyl decanol, octyldodecanol, odontella aurita oil, 2-oleamido-1,3 octadecanediol, palm glyceride, polyethylene glycol avocado glyceride, polyethylene glycol castor oil, polyethylene glycol-22 / dodecyl glycol copolymer, polyethylene glycol shea butter glyceride, phytol, raffinose, stearyl citrate, sunflower seed oil glyceride, tocopheryl glucoside, paraffin (e.g., soft white paraffin or liquid paraffin), or combinations thereof.Those skilled in the art will readily appreciate that emollients other than those listed above may also be formulated or incorporated into the topical formulations provided.

[0222] In some embodiments, topical formulations are formulated with one or more humectants.In certain aspects, humectants are ingredients that may help maintain the moisture level of skin.Specific examples of humectants include, but are not limited to, acetylarginine, algae extract, aloe barbadensis leaf extract, 2,3-butanediol, chitosan lauroyl glycinate, diglycereth-7 malate, diglycerin, diglycol guanidine succinate, erythritol, fructose, glucose, glycerin, honey, hydrolyzed wheat protein / polyethylene glycol-20 acetate copolymer, hydroxypropyltrimonium hyaluronate, inositol, lactitol, maltitol, maltose, mannitol, mannose, methoxypolyethylene glycol, myristamidobutylguanidine acetate, polyglyceryl sorbitol, potassium PCA, propylene glycol, sodium PCA, sorbitol, sucrose, urea, or combinations thereof.

[0223] In certain embodiments, the topical formulation may include one or more essential oils. In certain embodiments, essential oils include oils derived from herbs, flowers, trees, and other plants. Such oils are typically present as small droplets between the cells of the plant and can be extracted by several methods known to those skilled in the art, such as steam distillation, enfleurage (i.e., extraction using fats), maceration, solvent extraction, or mechanical expression. In some aspects, these types of oils tend to evaporate when exposed to air (i.e., volatile oils). In certain embodiments, essential oils are insoluble in water, but may be soluble in alcohol, ether, fixed oils (vegetable), or other organic solvents. In certain embodiments, typical physical properties found in essential oils include boiling points that vary from about 160° to 240° C., and densities ranging from about 0.759 to about 1.096.

[0224] The topical formulations herein may include one or more sunscreen actives (or sunscreen agents) and / or ultraviolet light absorbers. As used herein, "sunscreen active" includes sunscreen actives, sunscreens, and / or ultraviolet light absorbers as a whole. Sunscreen actives include both sunscreens and physical sunscreens. Sunscreen actives may be organic or inorganic. Examples of suitable sunscreen actives are disclosed in the Personal Care Product Council's International Cosmetic Ingredient Dictionary and Handbook, Thirteenth Edition, as "sunscreen agents." Particularly suitable sunscreen actives are 2-ethylhexyl p-methoxycinnamate (commercially available as PARSOL™ MCX), 4,4'-t-butylmethoxydibenzoyl-methane (commercially available as PARSOL™ 1789), 2-hydroxy-4-methoxybenzophenone, octyldimethyl-p-aminobenzoic acid, digalloyl trioleate, 2,2-dihydroxy-4-methoxybenzophenone, ethyl 4-(bis(hydroxypropyl))aminobenzoate, 2-ethylhexyl-2-cyano-3,3-diphenoxyethanol, and the like. nyl acrylate, 2-ethylhexyl salicylate, glyceryl p-aminobenzoate, 3,3,5-tri-methylcyclohexyl salicylate, menthyl anthranilate, p-dimethyl-aminobenzoic acid or aminobenzoates, p-dimethyl-aminobenzoate-2-ethylhexyl, 2-phenylbenzimidazole-5-sulfonic acid, 2-(p-dimethylaminophenyl)-5-sulfonic benzoxazonic acid, octocrylene, zinc oxide, benzylidene camphor and its derivatives, titanium dioxide, and mixtures thereof.

[0225] In some embodiments, the topical formulation is adjusted to a target pH or apparent pH. In certain embodiments, the topical formulation contains a neutral pH or apparent pH. In some embodiments, the topical formulation contains a pH or apparent pH of 5.0-8.0 or about 5.0-8.0, inclusive. In certain embodiments, the topical formulation contains a pH or nn apparent pH of 6.6-6.9 or about 6.6-6.9, inclusive. In some embodiments, the composition contains an acidic pH or apparent pH, such as when the composition is formulated as a topical cream or cream preparation. In certain embodiments, the pH or apparent pH is 4.0-5.0 or 4.4-4.7, or about 4.0-5.0 or 4.4-4.7, inclusive. In some embodiments, the pH or apparent pH of the topical formulation is 4.0-5.5. In certain embodiments, the pH or apparent pH of the topical formulation is 5.5 or is about 5.5. In certain embodiments, the pH or apparent pH of the topical formulation is 5.2 or about 5.2.

[0226] In some embodiments, the topical formulation can be filtered, for example, to remove or reduce bioburden.In some embodiments, the topical formulation can be filtered through a filter with a size of 0.1 μm to 1.0 μm or about 0.1 μm to 1.0 μm, for example, a filter with a size of 0.45 μm or 0.2 μm or about 0.45 μm or 0.2 μm.In some embodiments, the composition can be filtered through a filter with a size of 0.2 μm or about 0.2 μm.

[0227] In certain embodiments, the topical formulation is or comprises an emulsion, e.g., an oil-in-water emulsion. In some aspects, the oil-in-water emulsion contains an oil phase dispersed into an aqueous phase. In some embodiments, the aqueous phase is or contains human milk permeate, or a portion, fraction, or component thereof. In certain embodiments, the aqueous phase is or contains human milk oligosaccharides. In certain embodiments, the oil phase contains human milk cream. In certain embodiments, the oil phase contains human milk globules. In certain embodiments, the oil phase contains human milk lipids. In certain embodiments, the oil-in-water emulsion contains oil droplets (e.g., 1-1,000 nm in diameter). In some embodiments, the composition contains oil droplets, e.g., droplets containing or consisting of human milk cream, lipids, and / or globules, or fractions or portions thereof. In certain embodiments, the droplets are relatively uniform in size, e.g., at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, or 95% of the droplets fall within 2, 1.5, 1, or 0.5 standard deviations from the mean or median droplet size. In some embodiments, the mean or median droplet size is between 100 nm and 2 μm. In certain embodiments, the mean or median droplet size is between 250 nm and 1,000 nm, between 500 nm and 2,000 nm, or between 500 nm and 1,000 nm.

[0228] In certain embodiments, the topical formulation is or comprises an emulsion, e.g., a nanoemulsion, such as an emulsion having nanometer-sized droplets (e.g., 1-1,000 nm in diameter). In some embodiments, the composition contains droplets that contain or consist of oil droplets, e.g., human milk lipids, or human milk globules, or portions or fractions thereof. In certain embodiments, the droplets are relatively uniform in size, e.g., at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, or 95% of the droplets fall within 2, 1.5, 1, or 0.5 standard deviations from the mean or median droplet size. In some embodiments, the average or median droplet size is less than or about 1,000 nm, 750 nm, 500 nm, 400 nm, 300 nm, 200 nm, or 100 nm in diameter. In certain embodiments, the average or median droplet size is between or about 100 nm and 250 nm in diameter. In certain embodiments, the average or mean droplet size is between or about 200 nm in diameter.

[0229] In certain embodiments, the topical formulation is or comprises a hydrocolloid system, for example, containing human milk cream or lipids. In some embodiments, the hydrocolloid system is formulated with or contains 50%-75% or about 50%-75% (inclusive) human milk cream, or a portion or fraction derived or derived from human milk cream. In some embodiments, the hydrocolloid system comprises one or more hydrocolloids. In certain embodiments, the hydrocolloid system comprises one or more thickening agents. In certain embodiments, the hydrocolloid system comprises one or more antioxidants. In certain embodiments, the hydrocolloid system comprises one or both of xanthan gum and HPMC hydrocolloids, and one or more thickening agents selected from carabopol, carrageenan, HPC, paraffin wax, PEG4000, sodium alginate, and Avacel CL-611. In some embodiments, the hydrocolloid system is or comprises a formulation containing 50% human milk cream and one or more hydrocolloids selected from xanthan gum, sodium alginate, Protanal HF120PBS, and Avicel CL-611.In some embodiments, benzyl alcohol is added and is a preservative, and ascorbyl palmitate and alpha-tocopherol are added to the hydrocolloid system as antioxidants.

[0230] In some embodiments, the topical formulation provided is or comprises a cream preparation. In some embodiments, the cream preparation contains 5%-20% or about 5%-20% (inclusive) human milk cream and is formulated from an aqueous phase and an oil phase. In some embodiments, the aqueous phase consists of or comprises one or more of xanthan gum, cetostearyl alcohol, phenoxyethanol, benzyl alcohol, or steric acid. In certain embodiments, the lipid phase contains or comprises one or more of paraffin, such as soft white paraffin, liquid paraffin, cetomacrogol 1000, Tween 60, Span 60, Brij S2 (Brij 72), Brij S721 (Brij 721), ascorbyl palmitate, or ascorbic acid. In some embodiments, the cream preparation comprises one or more antioxidants.

[0231] In certain embodiments, the compositions provided are or include emulsion gel compositions. In certain embodiments, the emulsion gel compositions contain 5%-20% or about 5%-20% human milk cream. In certain embodiments, the emulsion gel contains an aqueous phase and an oil phase. In certain embodiments, the aqueous phase includes one or both of Tween 80 and benzyl alcohol. In certain embodiments, the oil phase includes human milk cream and one or both of liquid paraffin and SPAN 83. In some embodiments, the emulsion gel also includes one or more antioxidants. In certain embodiments, the emulsion gel includes xanthan gum as a gelling agent.

[0232] In some embodiments, the composition is formulated as an oily ointment composition. In some embodiments, the oily ointment contains 5% to 20% or about 5% to 20% (inclusive) human milk cream. In some embodiments, the oily ointment contains one or more of liquid paraffin, IPM, crodamole GRCC, castor oil, white soft paraffin, cetomacrogol 1000, and cetostearyl alcohol.

[0233] In certain embodiments, the topical formulation is or comprises an oil-in-water emulsion. In some embodiments, the oil-in-water emulsion contains an aqueous phase and an oil phase. In some embodiments, the oil-in-water emulsion contains human milk cream. In certain embodiments, the oil-in-water emulsion contains homogenized human milk cream. In certain embodiments, the oil-in-water emulsion contains human milk lipids.

[0234] In certain embodiments, the aqueous phase contains a human milk permeate, e.g., a permeate resulting from ultrafiltration of human skim milk. In certain embodiments, the aqueous phase contains human milk oligosaccharides. In some embodiments, the aqueous phase contains human milk cream. In certain embodiments, the aqueous phase contains human milk fat, fatty acids, and / or lipids. In certain embodiments, the aqueous phase contains one or more excipients, including an emulsifier. In some embodiments, the emulsifier is a polysorbate, e.g., polysorbate 80 (Tween 80). In certain embodiments, the aqueous phase contains one or more excipients that are or include a preservative. In certain embodiments, the preservative is phenoxyethanol. In certain embodiments, the oil-in-water emulsion contains an aqueous phase that is or includes a human milk permeate, e.g., resulting from ultrafiltration of human skim milk, an emulsifier, and a preservative. In some embodiments, the oil-in-water emulsion contains an aqueous phase that is or includes a human milk oligosaccharide, an emulsifier, and a preservative.

[0235] In certain embodiments, the oil phase includes one or more excipients that are or include an emulsifier. In certain embodiments, the emulsifier is or includes Span 83. In some embodiments, the oil phase includes one or more excipients that are or include an emollient. In some embodiments, the emollient is or includes paraffin, e.g., liquid paraffin. In various embodiments, the oil phase is or includes an antioxidant. In certain embodiments, the oil phase includes alpha tocopherol. In some embodiments, the oil phase contains liquid paraffin, Span 83, and alpha tocopherol.

[0236] In some embodiments, the oil-in-water emulsion is formulated by combining the aqueous phase and oil phase mixture, adding human milk cream, and adding a thickening agent. In some embodiments, the human milk cream is homogenized prior to addition. In certain embodiments, the human milk cream is added to achieve a target concentration of human milk fatty acids, fats, or lipids. In some embodiments, the target concentration is 1%-20%, 5%-15%, 7.5%-12.5%, or 8%-12% total fats, fatty acids, or lipids by weight per volume of the oil-in-water emulsion (w / v). In some embodiments, the thickening agent is a gum. In certain embodiments, the gum is xanthan gum.

[0237] In some embodiments, the topical formulation is formulated with human milk cream or a subfraction thereof, such as a butter oil fraction or a milk fat globule membrane solid subfraction, human milk permeate or a fraction or portion thereof, and one or more excipients. In certain embodiments, the topical formulation is formulated with human milk cream, human milk permeate, and one or more excipients. In certain embodiments, the topical formulation is formulated with human milk butter oil, human milk permeate, and one or more excipients. In certain embodiments, the topical formulation contains a human milk fat content of 10% to 30% w / w. In certain embodiments, the topical formulation contains at least 0.1%, 0.3%, 0.5%, 1%, or 3% w / w human milk oligosaccharides.

[0238] In certain embodiments, the one or more excipients are or include one or more of an emulsifier, a preservative, a thickener, a non-ionic surfactant, an antioxidant, and / or a hydrocolloid. In certain embodiments, the topical formulation is formulated with human milk cream or a subfraction thereof, human milk permeate or a fraction or portion thereof, and an excipient that includes an emulsifier, a preservative, a thickener, a non-ionic surfactant, an antioxidant, and a hydrocolloid. In certain embodiments, the emulsifier is or includes Tween 80 or polysorbate 80. In some embodiments, the preservative is or includes phenoxyethanol. In certain embodiments, the thickener is or includes liquid paraffin and / or glycerin. In some embodiments, the antioxidant is or includes alpha tocopherol. In certain embodiments, the hydrocolloid is or includes xanthan gum. V. Definition

[0239] Unless otherwise defined, all technical terms, notations, and other technical and scientific terms or terminology used herein are intended to have the same meaning as commonly understood by those skilled in the art to which the subject matter of the claims pertains. In some cases, terms having commonly understood meanings are defined herein for clarity and / or ease of reference, and the inclusion of such definitions herein should not necessarily be construed as representing a substantial difference to what is commonly understood in the art.

[0240] As used herein, the singular forms "a," "an," and "the" include plural references unless the context clearly dictates otherwise. For example, "a" or "an" means "at least one" or "one or more." It is understood that the aspects and variations described herein include "consisting of" and / or "consisting essentially of" aspects and variations.

[0241] Throughout this disclosure, various aspects of the claimed subject matter are presented in the form of a range. It should be understood that descriptions in the form of a range are merely for convenience and brevity, and should not be construed as inflexible limitations on the scope of the claimed subject matter. As a result, descriptions of ranges should be considered to have specifically disclosed all possible subranges within that range, as well as individual numerical values. For example, when a range of values ​​is provided, it is understood that each intervening value between the upper and lower limits of that range, and any other stated or intervening value within that stated range, is encompassed within the claimed subject matter. The upper and lower limits of these smaller ranges may be independently included within the smaller ranges, and are also encompassed within the claimed subject matter, depending on any specifically excluded limits within the stated range. When a stated range includes one or both of the limits, ranges excluding either or both of those included limits are also included in the stated or claimed subject matter. This applies regardless of the breadth of the range.

[0242] Throughout this disclosure, ranges presented or expressed as "between" two endpoints, e.g., "between A and B," are understood to include the endpoints, e.g., "A" and "B," unless otherwise indicated.

[0243] As used herein, the term "about" refers to a normal error range for the respective value, which is readily known to those skilled in the art. Reference herein to "about" a value or parameter includes (and describes) embodiments directed to the value or parameter itself. For example, a statement referring to "about X" includes a statement of "X". In some aspects, "about" a value means within ±25%, ±10%, ±5%, ±1%, ±0.1%, or ±0.01% of the value.

[0244] Unless otherwise indicated, the term "topical formulation" is used herein generally to include compositions or formulations suitable for topical delivery or administration, e.g., to the skin or mucosa.

[0245] As used herein, the term "pharmaceutical composition" refers to a mixture or formulation containing a particular amount, e.g., a therapeutically effective amount, of an active ingredient, such as a human milk fraction, in a pharma- ceutically acceptable carrier for administration to a mammal, e.g., a human.

[0246] As used herein, the term "pharmacologically acceptable" refers to compounds, materials, compositions, and / or dosage forms that are suitable for contact with the tissues of mammals, particularly humans, without undue toxicity, irritation, allergic response, and other problematic complications, within the scope of sound medical judgment, commensurate with a reasonable benefit / risk ratio, which may be routinely determined by one of ordinary skill in the art.

[0247] "Human milk oligosaccharides" (also referred to herein as "HMOs") refer to a family of structurally diverse non-conjugated glycans found in human breast milk. Human milk oligosaccharides are carbohydrates that contain lactose at the reducing end and typically fucose, sialic acid, or N-acetylglucosamine at the non-reducing end (Morrow et al., J. Nutri. 2005 135:1304-1307).

[0248] As used herein, the terms "mammalian milk lipids," "mammalian milk proteins," or "mammalian milk oligosaccharides" refer to lipids, proteins, or oligosaccharides, respectively, that are naturally found in or derived from mammalian milk. Similarly, the terms "human milk lipids" or "human milk proteins" as used herein refer to lipids or proteins, respectively, that are naturally found in or derived from human milk.

[0249] "Permeate" refers to the portion of milk (e.g., pooled human milk) obtained as the product flowing through the filter during ultrafiltration. Specifically, it is the liquid that flows through after retention of proteins and lipids during ultrafiltration (e.g., through a filter of about 1-10,000 kDa). The liquid that passes through this ultrafiltration process is called the permeate. The retentate of this process concentrates human milk proteins, which may then be used to generate other life-saving formulations, for example, to make human milk fortifier compositions such as those described in U.S. Pat. No. 8,377,455. Thus, in contrast to methods that rely on protein precipitation with solvents, which may contaminate the HMO product, the use of ultrafiltration to obtain a substantially protein-free starting material, as used herein, preserves the remaining portion of the valuable macronutrients in human milk while avoiding the use of organic solvents.

[0250] "Milk" means the fluid produced by the mammary glands and expressed by the breasts of a mammal. Milk includes all lactation products, including but not limited to colostrum, whole milk, and skim milk, taken after birth or at any time during the lactation cycle, in the production of hindmilk and foremilk. Unless otherwise specified, milk as used herein typically refers to human whole milk.

[0251] "Whole milk" means milk that has not had any fat removed (eg, pooled human milk).

[0252] "Skim milk" means milk from which at least 75% of the fat has been removed (eg, pooled human milk), or alternatively, milk that has been centrifuged to remove fat.

[0253] The term "essentially free" of a substance, e.g., non-human milk lipids or non-human milk oligosaccharides, means that no substance, e.g., any non-human milk lipids or non-human milk oligosaccharides, has been intentionally added or that no more than trace amounts of a substance, e.g., non-human milk lipids or non-human milk oligosaccharides, are present.

[0254] Unless otherwise indicated, "human milk cream" or "human cream" means that portion of human whole milk that is separated from human skim milk. Typically, cream contains higher concentrations of long-, medium-, and short-chain fatty acids than those in skim milk obtained from the same preparation.

[0255] "Substantially," as in "substantially reduced lactose content and / or mineral content," means that the reduction in mineral and / or lactose levels represents a statistical difference when compared to concentrated permeate that has not been subjected to current methods. By way of example, in some embodiments, HMO compositions having substantially reduced lactose include lactose levels of 5% or less.

[0256] As used herein, "consists essentially of" refers to a composition that contains the specified recited components, excluding other major bioactive factors.For example, a composition that consists essentially of HMOs excludes intact proteins, fats, or exogenously added materials, but may contain other inactive or trace substances, such as water, acceptable levels of certain salts, microRNA, or exosomes. Working Example

[0257] The following examples are included for illustrative purposes only and are not intended to limit the scope of the invention. Example 1: Production of cream, permeate, and retentate fractions of human milk

[0258] Pooled human milk from individual donors was separated into cream, permeate, and retentate fractions. Pooled whole human milk was filtered through a 200 μm filter, pasteurized at a temperature of at least 63° C. for 30 minutes, and then cooled to 2° C.-8° C. The milk was then transferred to a centrifuge to separate the cream from the skim. The cream was collected and stored at −20° C., and the skim milk was ultrafiltered with a 10 kDa membrane. The material that did not pass through the filter was collected as the retentate fraction, and the material that passed through the filter was the permeate fraction. Example 2: Effects of human milk fractions on a three-dimensional in vitro healthy human skin model

[0259] The effect of human milk fractions produced as described in Example 1 on skin was evaluated using a three-dimensional in vitro human skin model. The in vitro skin model was similar to that described in WO2001092477, which is incorporated by reference in its entirety. A human skin model preparation was produced by embedding human dermal fibroblasts into a biomatrix containing matrix proteins and submerged in culture medium, as illustrated in FIG. 1A. After approximately one day, the medium volume was reduced to convert the preparation into an air-liquid interface culture. Keratinocytes were then seeded on top of the fibroblast layer, which acted as a scaffold. The cells were cultured for 2-3 weeks under conditions that allowed the keratinocytes to differentiate into a multi-level epidermis with layers of basal, spinous, granular, and stratum corneum, as well as a functional basement membrane composed of matrix proteins, as shown in FIG. 1B.

[0260] Human milk cream fraction, retentate fraction, and permeate fraction produced as described in Example 1 were formulated to concentrations of 1%, 5%, 10%, and 30% in PBS vehicle containing 1% DMSO. Vehicle-only condition served as a negative control, and a plant extract previously shown to increase barrier function in in vitro skin preparations was used as a control. The formulations were applied to the in vitro skin preparations once a day for three days. The transepithelial / transendothelial electrical resistance (TEER) of the in vitro human skin preparations was measured once a day by impedance spectroscopy at a reference frequency of 1,000 Hz to assess barrier function. Culture medium was also collected daily and assessed for levels of IL-8 by ELISA, glucose consumption using a photometric hexokinase assay, and lactate dehydrogenase (LDH) using an enzymatic photometric assay. After 72 hours, the in vitro human skin preparations were sectioned and stained with hematoxylin and eosin (H&E) or assessed using the MTT assay to assess cell viability.

[0261] As shown in Figure 2, application of formulations containing each human milk fraction increased the TEER measurements observed in in vitro human skin preparations. Formulations containing human milk cream or retentate showed a trend towards a dose-dependent increase in TEER (Ohm*cm at 1000 Hz) compared to the vehicle only control. 2 Formulations containing 1% permeate increased TEER compared to the vehicle-only control, while decreases in TEER were observed at higher concentrations of permeate. The observation of increased TEER is consistent with increased barrier function following application of the human milk fraction.

[0262] No differences were observed in any of the formulations in the medium concentrations of IL-8, glucose, or LDH, or in the MTT assay results after application of the human milk fraction to skin preparations. These results are consistent with the absence of adverse effects of the human milk fraction on skin inflammation, cell metabolism, membrane integrity, and cell viability. Consistent with these observations, no damage was observed in the H&E stained cross-sections of the skin preparations after application of any of the formulations. Example 3: Effect of human milk fractions on a three-dimensional in vitro model of skin inflammation

[0263] Human milk fractions produced as described in Example 1 were evaluated for their effect on skin inflammation. Three-dimensional in vitro skin preparations produced as described in Example 2 were incubated with 50 ng / ml TNF-alpha to model skin inflammation. After 3 hours of incubation, a vehicle-only control or a formulation containing a human milk fraction similar to that described in Example 2 was applied to the in vitro skin preparations. Additional controls that were not treated with TNF-alpha were also tested. After 24 hours, media was collected from the skin preparations and assessed for IL-8 levels by ELISA.

[0264] As shown in Figure 3, a trend towards reduced IL-8 was observed in the culture medium following treatment with formulations containing human milk cream, permeate, and retentate compared to vehicle-only controls. Treatment with a formulation containing 5% permeate reduced IL-8 levels by approximately 30%. The reduced levels of IL-8 were consistent with the anti-inflammatory effects of human milk fractions on human skin. Example 4: Effects of human milk fractions on a three-dimensional in vitro model of damaged skin

[0265] Human milk fractions produced as described in Example 1 were evaluated using a three-dimensional in vitro model of wounded human skin.

[0266] In one experiment, three-dimensional in vitro skin preparations generated as described in Example 2 were mechanically disrupted using friction stress, resulting in a reduction in barrier function. Approximately 24 hours after mechanical disruption, a formulation containing vehicle alone (1% DMSO in PBS), a combination of human milk cream and permeate, or 30% skim milk in vehicle was applied to the in vitro skin preparations for 7 days. TEER was measured in in vitro skin preparations similar to those described in Example 2 approximately 3, 5, and 8 days after mechanical disruption and expressed as a percentage of the TEER measurements collected from vehicle-only controls prior to mechanical disruption.

[0267] As shown in Figure 4, increased TEER measurements were observed 5 and 8 days after mechanical disruption in cream / permeate and skim milk treated skin preparations compared to vehicle only controls. These results are consistent with the combined cream / permeate and skim milk fractions improving repair and barrier function in human skin, as elevated TEER measurements indicate increased barrier function.

[0268] In the second experiment, skin punches were removed from in vitro skin preparations to evaluate re-epithelialization. Immediately after skin punches were collected, vehicle alone or formulations containing human milk retentate were applied to the in vitro skin preparations. TEER was measured before and immediately after removing the skin punch, and approximately 24 hours, 72 hours, 144 hours, 192 hours, and 240 hours later. As shown in Figure 5, at 144 hours, increased TEER measurements were observed in the in vitro skin preparations treated with formulations containing human milk retentate compared to vehicle alone controls, consistent with increased re-epithelialization and barrier function recovery. Example 5: Characterization of human milk cream

[0269] The stability of the human milk cream obtained as described in Example 1 was evaluated by exposing samples of the cream to accelerated physical and thermal stresses. The rheological properties of the human milk cream were also investigated.

[0270] The response to accelerated physical stress was assessed by centrifuging samples of human milk cream at 13,000 rpm for 2 minutes in a tabletop centrifuge, which separated the cream into three distinct layers: a clear bottom layer, a creamy middle layer, and a yellow liquid top layer.

[0271] The response to heat stress was evaluated by storing samples of human milk cream at temperatures of 2-8°C, 25°C, 40°C, 50°C, or 70°C. Samples were visually inspected after 3, 6, and 24 hours of storage at each temperature. Above 25°C, phase separation was observed at all time points. A direct relationship was observed between heat and the level of phase separation. Above 40°C, sedimentation and color change were observed.

[0272] Rheology was evaluated in samples containing either 100% human milk cream or 50% cream and 25% hydrocolloid. Oscillatory stress sweeps were performed to characterize the mechanical properties of the samples. Results showed that the addition of hydrocolloid increased the complex elasticity and yield stress of the samples, consistent with the possibility that the hydrocolloid reduces the probability of phase separation in formulations containing human milk cream. The rheology of samples containing 100% cream was also evaluated using frequency sweeps. At low frequencies, the loss modulus dominated the storage modulus, consistent with a relatively weak structure and a high probability of phase separation in the cream. These observations were consistent with a cream having the properties of a viscous solution and not being solid at rest.

[0273] Taken together, the results of the stress test and rheological evaluation were consistent with a human milk cream having a low density and a high probability of phase separation. Example 6: Preparation of a hydrocolloid system containing human milk cream

[0274] Test hydrocolloid systems containing human milk cream obtained as described in Example 1 were formulated and evaluated for stability. Hydrocolloid systems were prepared from various formulations containing 50% to 75% (inclusive) human milk cream, one or both of xanthan gum and HPMC hydrocolloids, and one or more thickeners selected from carabopol, carrageenan, HPC, paraffin wax, PEG 4000, sodium alginate, and Avacel CL-611. Samples prepared from each formulation were stored at 2-8°C, 25°C, 40°C, or 50°C. Visual and odor tests were performed immediately after formulation (day 0) and after approximately 2 weeks of storage at each temperature. At day 0, all of the hydrocolloid systems had a slight dairy odor. After 2 weeks, all formulations had a strong rancid odor regardless of physical appearance. The results of the visual inspection are summarized in Table E1. [Table 1]

[0275] All formulations appeared to improve stability compared to human milk cream alone. Test Formula 8 appeared to be the most stable formulation as it increased in viscosity after two weeks at 25° C. and 40° C., but no phase separation was observed at any of the temperatures tested. None of the other formulations prevented phase separation from occurring after two weeks of storage at temperatures above 25° C.

[0276] These results are consistent with the suitability of the human skin cream to be formulated in a stable hydrocolloid system, with the degree of stability depending on the particular formulation. Example 7: Production and characterization of different compositions containing human milk cream

[0277] Human milk cream was formulated into skin cream, hydrocolloid system, emulsified gel, and oil-based ointment compositions. Samples from each composition were evaluated immediately after formulation (day 0) and after 2 and 4 weeks of storage at 2-8°C, 25°C, 40°C, or 50°C. At each time point, samples were evaluated for apparent pH, macroscopic and microscopic appearance, and rheology. Rheology was evaluated as described in Example 6. For all test compositions, an increase in complex elasticity and apparent yield stress was observed after 2 weeks at 50°C, so rheology was not evaluated at the 4 week time point for the 50°C condition.

[0278] No rancid odor was detected in any of the compositions prepared from any of the formulations after 2 weeks of storage at any temperature, or after 4 weeks of storage at 25°C and 40°C. The apparent pH was measured to be 6.6 to 6.9 for the hydrocolloid-based, emulsified gel, and oil-based ointment compositions, and 4.4 to 4.7 for the cream preparation. The pH value did not appear to change over time in any of the compositions. A) Skin Cream Preparations

[0279] Test skin cream preparations containing either 5% or 20% human milk cream obtained as described in Example 1 were formulated from an aqueous phase and an oil phase. The aqueous phase consisted of xanthan gum, cetostearyl alcohol, and the presence or absence of phenoxyethanol, benzyl alcohol, and steric acid. The lipid phase contained white soft paraffin, and the presence or absence of liquid paraffin, cetomacrogol 1000, Tween 60, span 60, Brij S2 (Brij 72), Brij S721 (Brij 721), ascorbyl palmitate, and ascorbic acid. The formulations also contained the presence or absence of antioxidants.

[0280] At day 0, the skin cream preparations had a white color, with one exception having a light yellow color that became off-white after 4 weeks of storage at 25°C or 40°C. The skin cream preparations had a high viscosity and were not pourable. No rancid odor was detected in the cream preparations after 2 weeks or after 4 weeks at 25°C and 40°C from any of the formulations. Aeration was observed after 2 weeks in all cream preparations at all temperatures tested.

[0281] Small homogeneous oil droplets could be observed under a microscope in all cream formulations at day 0. An increase in droplet size was observed after 2 weeks of storage at 50° C., but no observable increase was observed in any of the formulations at lower temperatures. Only one of the skin cream formulations underwent phase separation, which occurred after 2 weeks at 50° C.

[0282] The measured values ​​of complex elasticity and apparent yield stress of the cream formulations were similar at day 0 and after 4 weeks of storage at 25°C and 40°C. B) Hydrocolloid system

[0283] Test hydrocolloid systems were prepared from formulations containing 50% human milk cream and one or more hydrocolloids selected from xanthan gum, sodium alginate, Protanal HF120PBS, and Avicel CL-611. Benzyl alcohol was added as a preservative, and ascorbyl palmitate and alpha-tocopherol were added to each formulation as antioxidants. Samples prepared from each formulation were stored at 2-8°C, 25°C, 40°C, or 50°C. The hydrocolloid systems were evaluated for pH, visual appearance, and odor after 0 days, approximately 2 weeks, and approximately 4 weeks of storage at each temperature.

[0284] At day 0, the hydrocolloid system had a light yellow color. Orange particles were found within the hydrocolloid system after 2 weeks at 50° C. and after 4 weeks at 25° C. and 40° C. Half of the formulations tested had an increase in viscosity after 4 weeks of storage at 25° C. and 40° C.

[0285] Heterogeneous oil droplets were observed microscopically within the hydrocolloid systems at day 0 and after 2 and 4 weeks of storage at 25° C., 40° C., and 50° C. No excipient crystals were observed at any condition.

[0286] Rheological measurements collected from the test hydrocolloid systems at 0 days, 2 weeks, and 4 weeks were consistent with greater instability in the hydrocolloid systems compared to the other cream preparations and emulsion gels. The complex elasticity of the test hydrocolloid systems increased upon storage for 2 weeks at 40° C. or 50° C. However, some formulations maintained similar values ​​for complex elasticity over 2 and 4 weeks of storage at 25° C. C) Emulsion gel composition

[0287] Emulsion gel compositions were produced from formulations containing either 5% or 20% human milk cream, an aqueous phase, and an oil phase. The aqueous phase contained polysorbate 80 and benzyl alcohol. The oil phase contained cream, liquid paraffin, and Span 83. The formulations also contained antioxidants and xanthan gum as a gelling agent.

[0288] At day 0, the emulsified gel had a slight yellow color, low viscosity, and was pourable. After storage at 40° C. for 2 weeks and at 25° C. for 4 weeks, moderate viscosity and slow pourability was observed. After storage at 50° C. for 2 weeks and at 40° C. for 4 weeks, high viscosity and lack of pourability was observed. After storage at 40° C. for several weeks, a slight translucent layer was observed in the emulsified gel containing 20% ​​cream.

[0289] As is typical of emulsion gel compositions, small oil droplets were observed under a microscope at 0 days and after 2 weeks of storage at 40°C and 50°C, and after 4 weeks of storage at 25°C and 40°C.

[0290] Rheological measurements indicated that among the different types of compositions tested, the emulsion gels showed the least change in mechanical behavior during storage. D) Oily ointment

[0291] Oil-based ointment compositions were generated from formulations containing either 5% or 20% human milk cream. Excipients for each formulation included liquid paraffin, IPM, crodamole GRCC, castor oil, white soft paraffin, cetomacrogol 1000, and cetostearyl alcohol.

[0292] At day 0, the oily ointment composition containing 5% human milk cream had an off-white color. The composition containing 20% ​​cream appeared yellow. The oily ointment composition initially had a medium viscosity and was not pourable. After storage for 2 and 4 weeks under all temperature conditions, an increase in viscosity was observed.

[0293] Large solid fluorescent particles (likely due to the cetostearyl alcohol excipient) were observed at all temperatures at day 0 and after 2 and 4 weeks of storage.

[0294] The complex elasticity and apparent yield stress measurements from the oily ointment were variable at all time points and at all temperatures examined, consistent with greater instability of the oily ointment compared to the other test compositions. Example 8: Production of nanoemulsions containing human milk cream

[0295] Human milk cream produced similarly to that described in Example 1 was formulated into different nanoemulsion formulations. Nanoemulsions were prepared from 27 different test formulations containing either 10 wt%, 20 wt%, or 30 wt% human milk cream with different combinations of excipients. Nanoemulsions were prepared in 1 mL volumes using either one or two rounds of high speed bead beading followed by centrifugation to remove bubbles. The resulting nanoemulsions were evaluated for appearance and clarity. Translucent and uniform samples were examined by microscopy for the presence of oil droplets. Samples were placed on a rotator at room temperature for 2-3 days and examined for appearance and particle size via laser diffraction analysis to identify nanoemulsions that remained translucent and uniform.

[0296] Based on the results of the initial testing, four test formulations were selected for further evaluation. Forty grams of nanoemulsion was generated for each of the four test formulations. The formulations contained 10% by weight human milk cream in a solution containing EDTA and glycerin. The formulations also contained various amounts of soy lecithin, as well as the presence or absence of histidine and benzalkonium chloride. Three of the formulations were then pH adjusted to 7.5±0.5 and the fourth formulation was adjusted to 11.5±0.5. The formulations were processed using a Microfluidizer and the resulting nanoemulsions were each pH adjusted to 7.0±0.5 and filtered through a 0.22 μm membrane into a sterile container. The filtrates were stored at 2-8° C., 25° C., or 40° C. and inspected twice weekly for appearance and particle size via laser diffraction analysis. The results are summarized in Tables E1-E4 below. [Table 2] [Table 3] [Table 4] [Table 5]

[0297] The translucent samples appeared clear under the microscope, whereas small particles could be detected in the opaque samples. The nanoemulsions appeared homogeneous at all time points and were stable over time at the temperatures and mean globule sizes examined. Taken together, these observations are consistent with the generation of stable nanoemulsions containing human milk cream. Example 9: Preparation of oil-in-water emulsions containing human milk cream

[0298] Various oil-in-water emulsions were produced containing either or both of the human milk cream and permeate fractions produced as described in Example 1. Emulsions were produced by mixing various aqueous and oil phases for use as test emulsions. Aqueous phases included those formulated by combining water or permeate in the absence or presence of various levels of human milk cream, Tween 80, and phenoxyethanol. Oil phases were formulated with liquid paraffin and Span 83. Alpha tocopherol was added as an antioxidant and xanthan gum was added as a thickener.

[0299] The oil-in-water emulsions were analyzed using a particle size analyzer (Malvern 2000, Malvern Panalytical, Malvern, UK). The emulsions containing Tween 80 had a lower average particle size than the corresponding emulsions lacking Tween 80. In addition, it was observed that the viscosity of the emulsions increased with Tween 80 concentration. For the emulsions containing human milk cream, those formulated with homogenized human milk cream generally had a narrower particle size distribution than similar emulsions formulated with non-homogenized cream. Example 10: Effect of oil-in-water emulsions containing human milk cream and permeate on TNF-induced IL-8 secretion in an in vitro model of human skin

[0300] Oil-in-water emulsions containing either or both of the human milk cream and permeate fractions produced as described in Example 1 were produced as test emulsions. The test emulsions were evaluated on an in vitro model of human skin similar to that described in Example 2. The test emulsions were produced by combining (i) an aqueous phase produced by combining water or permeate with Tween 80 and phenoxyethanol, with or without human milk cream, and (ii) an oil phase formulated with liquid paraffin and Span 83. Alpha tocopherol was added as an antioxidant and xanthan gum was added as a thickener. The emulsion lacking human milk permeate and cream served as a vehicle control, and a commercial topical formulation served as a positive control.

[0301] Test emulsions and controls were applied to in vitro human skin models. TNF (10 ng / ml) was applied to human skin models in parallel with additional experimental controls of untreated skin models incubated with or without TNF. Human skin models were incubated in the presence of control and test emulsions for 72 hours, followed by 48 hours post-treatment incubation in the absence of control and test emulsions. Human skin models were evaluated for IL-8 secretion at 24 hours and 72 hours, and after 48 hours of incubation, by solid-phase sandwich ELISA.

[0302] As shown in Figures 6A-C, TNF exposure was associated with increased levels of extracellular IL-8. Application of test emulsions containing human milk permeate or both human milk permeate and cream resulted in a significant decrease in TNF-stimulated IL-8 secretion at the 24-hour (Figure 6A) and 72-hour (Figure 6B) time points. Following a 48-hour post-treatment incubation, no effect on TNF-stimulated IL-8 secretion was observed (Figure 6C). These results are consistent with the anti-inflammatory properties of human milk permeate and human milk cream. Example 11: Preparation of oil-in-water emulsions containing human cream and permeate fractions

[0303] An oil-in-water emulsion was produced containing human cream and human milk permeate fraction, produced similarly as described in Example 1. The cream fraction was pressure homogenized using a Lactoscope FT-A (Perkin Elmer) and the total fat content of the cream was measured after homogenization. The emulsion was formulated by first gently combining the separate aqueous and oil phase mixtures, adding homogenized cream, and then adding xanthan gum as a thickening agent. The aqueous phase was formulated from the permeate fraction containing Tween 80, phenoxyethanol, and at least 0.5% HMO (w / v). The oil phase contained liquid paraffin, Span 83, and alpha tocopherol. The homogenized cream was added in an amount sufficient for the final emulsion to contain 10% human milk fat (w / v).

[0304] Samples from the resulting oil-in-water emulsions were analyzed using a particle size analyzer (Malvern 2000, Malvern Panalytical, Malvern, UK). The average particle size of the emulsions was determined to be 0.5-1.0 microns in diameter.

[0305] Antimicrobial efficacy testing (AET) was performed on the emulsion samples. Samples were stored in a refrigerator for 4 weeks prior to AET testing. A controlled inoculum of challenge organisms, including S. aureus, P. aeruginosa, E. coli, C. albicans, and A. brasiliensis, was placed in suspension with the emulsion samples, and viable challenge organism counts were assessed at baseline (day 0) and days 7, 14, and 28. At days 7, 14, and 28, counts were below the threshold of detection, consistent with the preservative activity of the emulsion with the ability to prevent microbial growth.

[0306] Fatty acid profiles of samples collected from the emulsions were generated by gas chromatography-mass spectrometry (GC-MS). Samples were collected at day 0 and at various time points after formulation. Fatty acid profiles were generated by calculating the weight percent values ​​of individual fatty acid species compared to the total fatty acid content. Comparison of fatty acid profiles was performed by determining the correlation between measurements of individual fatty acid species for each profile. Comparison of fatty acid profiles measured from samples of the same manufacturing lot stored at 2°C-8°C or room temperature and collected at day 0, 3 months, 6 months, 9 months, and 12 months showed little change in fatty acid profile from day 0 between these time points, consistent with stability of the emulsion for at least one year when stored refrigerated or at room temperature. Example 12: Treatment of Psoriasis with a Topical Formulation Containing Human Milk Ingredients

[0307] The safety and tolerability of a topical formulation containing human milk components formulated similarly as described in Example 11 will be evaluated for the treatment of psoriasis in an open-label, single-center study. The study will enroll male and female subjects aged 18 years or older who meet the following criteria: i) a clinical diagnosis of chronic plaque psoriasis and stable disease for at least 6 months prior to the study, and ii) 2%-20% body surface area involvement (excluding scalp, palms, fingernails, toenails, and soles).

[0308] Subjects will apply the topical formulation to the affected area twice daily, approximately 12 hours apart, for 8 weeks. The ingredients of the topical formulation are summarized in Table E5. [Table 6]

[0309] Lesions will be photographed at baseline (day 1), weeks 2, 4, and 8. Treatment evaluations may include peak itch numeric rating scales and surveys to assess tolerability and satisfaction with treatment. Safety evaluations will be performed for the occurrence, frequency, and duration of any treatment-related adverse events (TRAEs), serious adverse events (SAEs), or adverse events (AEs) leading to discontinuation of the study.

[0310] Less severe and / or total lesions or affected areas may be observed during treatment compared to baseline. Additional observations may be consistent with a decrease in severity of psoriasis, a decrease in skin irritation or itching sensation, and / or an improvement in skin appearance during treatment compared to baseline. Results may also indicate that the topical formulation is well tolerated. A low incidence, frequency, and / or duration of TRAEs, SAEs, and / or AEs, if present, is observed during treatment. Example 13: Treatment of psoriasis with a topical formulation containing human milk components

[0311] A clinical study evaluating the safety and tolerability of topical formulations for the treatment of psoriasis was conducted similar to that described in Example 12. The topical formulations were made similar to that described in Example 11 and contained the ingredients summarized in Table E5. Eleven subjects completed the study.

[0312] The topical formulation was generally well tolerated by patients. Subjects maintained a patient tolerability score of "0" throughout the study, except for one subject who reported a tolerability score of "1" at week 2, defined as "slight sensation, but no discomfort, no intervention required." This subject reported a score of "0" for all subsequent visits. No evidence of irritation or intolerance of the topical formulation was observed in the investigator's assessments at weeks 2, 4, and 8. No adverse events or serious adverse events were reported.

[0313] Subjects rated itch using a 10-point Peak Itch Numeric Rating Scale, with higher numbers indicating more severe itch sensations. Mean peak itch scores are shown in Table E6. [Table 7]

[0314] Over the course of the study, five of the subjects reported lower scores than baseline, and only one subject reported an increase in score versus baseline at week 8. Review of serial photographs revealed a trend toward improvement of the underlying psoriasis condition, as evidenced by a decrease in erythema, scaling, peeling, and lichenification.

[0315] Taken together, these data are consistent with a topical formulation derived from human milk as a safe remedy for the symptoms of plaque psoriasis. Example 14: Treatment of atopic dermatitis with a topical formulation containing human milk components

[0316] The safety and tolerability of two topical formulations containing human milk ingredients are evaluated for the treatment of psoriasis in an open-label, single-center study. The two topical formulations were formulated similarly as described in Example 11, with one formulation having 10% w / w human milk fat and the other having 20% ​​w / w human milk fat. The ingredients are summarized in Table E7. [Table 8]

[0317] The study will enroll male and female subjects aged 18 years or older who meet the following criteria: i) a clinical diagnosis of chronic atopic dermatitis and stable disease for at least 6 months prior to the study, and ii) ≤35% body surface area involvement (excluding scalp, palms, fingernails, toenails, and soles).

[0318] Each subject will apply one of the topical formulations to the affected area twice daily, approximately 12 hours apart, for eight weeks.

[0319] Lesions will be photographed at baseline (day 1), weeks 2, 4, and 8. Evaluation of treatment may include surveys to assess tolerability and satisfaction with treatment, as well as the investigator's assessment of the presence and overall degree of irritation at the application site according to a 5-point scale, such as that shown by Table E7. [Table 9] Safety assessments will be performed for the occurrence, frequency, and duration of either adverse events (AEs), serious adverse events (SAEs), or any adverse events leading to discontinuation of the study.

[0320] Less severe and / or total lesions or affected areas may be observed during treatment compared to baseline. Additional observations may be consistent with a decrease in severity of psoriasis, a decrease in skin irritation or itching sensation, and / or an improvement in skin appearance during treatment compared to baseline. Results may also indicate that the topical formulation is well tolerated, with a low incidence, frequency, and / or duration of SAEs and / or AEs (if any) observed during treatment. Example 15: Treatment of atopic dermatitis with a topical formulation containing human milk components

[0321] The safety and tolerability of a topical formulation containing a human milk-derived cream was evaluated for the treatment of atopic dermatitis in a clinical study conducted similarly to that described in Example 12. The topical formulation was made similarly to that described in Example 11 and contained the ingredients summarized in Table E7. The first six subjects enrolled received a topical formulation containing 10% human milk fat, while the remaining eight subjects received a topical formulation with 20% human milk fat.

[0322] The topical formulations were generally well tolerated by subjects. No subjects reported any irritation or sensitivity to any of the topical formulations, and no evidence of irritation or sensitivity to the topical formulations was observed in subjects at any of the 2, 4, or 8 week study visits.

[0323] Subjects rated itch using a 10-point peak itch numerical rating scale, with higher numbers indicating more severe itch sensation. Mean peak itch scores for subjects receiving topical formulations containing 10% or 20% human milk fat are shown in Table E9. A trend toward reduction in peak itch scores over the 8 weeks of treatment was observed in both study groups. [Table 10]

[0324] Taken together, these data are consistent with a human milk-derived topical formulation as a safe remedy for the symptoms of atopic dermatitis. Example 16: Preparation of a topical formulation containing human milk components

[0325] A topical formulation containing human milk cream and human milk permeate fraction was produced similarly as described in Example 1. The total fat content of the cream was measured using a Lactoscope FT-A (Perkin Elmer). The ingredients of the topical formulation are shown in Table E10. The topical formulation contains 20% fat and human milk permeate with Tween 80, phenoxyethanol, liquid paraffin, Span 83, and alpha tocopherol, and is mixed at room temperature for about 30 minutes at 25000 rpm using an IKA T25 dispersion unit. The final topical formulation was produced by slowly adding xanthan gum and mixing for about 15 minutes at 700 rpm. [Table 11] Example 17: Production of human milk cream subfraction

[0326] Human milk cream was separated from human whole milk as described in Example 1. The resulting human milk cream fraction was then homogenized at 20,000 psi for one period at a rate of 60 ml / min. The homogenized human milk cream was then centrifuged at 3,500 g for 30 minutes at ambient temperature. The homogenized cream was separated into three subfractions: an upper layer composed of butter oil, a middle layer composed of milk fat globule membrane solids, and a lower layer containing skim milk. Topical formulations containing 20% ​​fat (w / w) were produced from the butter oil subfraction and the milk fat globule membrane solids subfraction by replacing the human milk cream with the butter oil subfraction or the milk fat globule membrane solids subfraction in the procedure described in Example 16.

[0327] Additional topical formulations are produced by substituting the butter oil sub-fraction or the milk fat globule membrane solids sub-fraction for human milk cream in any of the methods, processes, or recipes for producing the topical formulations described in Examples 6-9, 11, 12, or 14.

Claims

1. A topical formulation comprising homogenized human milk cream, human milk permeate obtained by ultrafiltration of human skim milk, and one or more pharma- ceutically acceptable excipients, wherein the one or more pharma-ceutically acceptable excipients include one or more of an antioxidant, an emulsifier, or a hydrocolloid.

2. The homogenized human milk cream contains human milk fat globules, and the human milk fat globules at least 60%, 70%, 75%, 80%, or 95% of the particles have a diameter between 0.2 μm and 2 μm; 2. The topical formulation of claim 1.

3. 3. The topical formulation of claim 1 or 2, wherein the topical formulation comprises one or more of oleic acid, palmitic acid, linoleic acid, omega-3 fatty acids, omega-6 fatty acids, omega-9 fatty acids, stearic acid, myristic acid, lauric acid, palmitoleic acid, vaccenic acid, myristolic acid, heptadecenoic acid, gamma-linolenic acid, docosapentaenoic acid, eicosapentaenoic acid, arachidonic acid, and docosahexaenoic acid.

4. 4. A topical formulation according to any of claims 1 to 3, wherein the topical formulation comprises at least 1% human milk fat (wt / wt), such as from 1% to 40% or from 10% to 30% human milk fat (wt / wt).

5. The topical formulation of any of claims 1 to 4, wherein the topical formulation comprises at least 0.1% human milk oligosaccharides (weight / weight).

6. 6. The topical formulation of claim 5, wherein the human milk oligosaccharides comprise 2'-fucosyl-lactose, 3-fucosyl-lactose, 3'-sialyl-lactose, 6'-sialyl-lactose, lacto-N-tetraose, lacto-N-difucohexaose I, lactodifucotetraose, lacto-N-fucopentaose I, sialylacto-N-tetraose c, sialylacto-N-tetraose b, and disialyllacto-N-tetraose.

7. The topical formulation of any of claims 1 to 6, wherein the topical formulation comprises at least 0.1% human milk oligosaccharides (weight / weight) and 1% to 40% human milk fat (weight / weight).

8. The topical formulation of any of claims 1 to 7 for use in treating one or more symptoms associated with a skin condition or disorder in a subject in need thereof.

9. 10. The topical formulation of claim 8, wherein said use comprises applying said topical formulation directly to an area of ​​the subject's skin exhibiting or at risk of exhibiting said one or more symptoms.

10. 10. The topical formulation of claim 8 or 9, wherein the skin condition or disorder comprises one or more of dermatitis, atopic dermatitis, contact dermatitis, hand dermatitis, allergic dermatitis, stasis dermatitis, asteatotic dermatitis, nummular dermatitis, seborrheic dermatitis, infectious dermatitis, acne, acne vulgaris, rosacea, epidermolysis bullosa simplex, chronic superficial scaly acrodermatitis, chondrodermatitis, perioral dermatitis, dermatomyositis, neutrophilic dermatosis, transient acantholytic dermatosis, eczema, ichthyosis, xerosis, asteatosis, psoriasis, lupus erythematosus, exfoliative keratolysis, impetigo, allergic inflammation of the skin, hives, lacerations, burns, or wounds resulting from a surgical incision or biopsy.

11. The topical formulation of any of claims 8 to 10, wherein the skin condition or disorder comprises atopic dermatitis or psoriasis.

12. 12. The topical formulation of any of claims 8-11, wherein the one or more symptoms associated with the skin condition or disorder comprise one or more of dryness, lesions, cracks, fissures, scaling, discoloration, thickening, redness, bleeding, swelling, peeling, ulcers, pain, wounds, blistering, rash, inflammation, infection, or loss of flesh.

13. The topical formulation of claim 12, wherein the one or more symptoms comprises inflammation.

14. The topical formulation of any of claims 8 to 13, wherein the subject is experiencing or at risk of experiencing pain associated with breast feeding.

15. The topical formulation of claim 14, wherein said use comprises applying said topical formulation to the breast or nipple of the subject.

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