WO2001017523A1 - Compositions et methodes pour modifier le taux de lipides cutanes - Google Patents

Compositions et methodes pour modifier le taux de lipides cutanes Download PDF

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Publication number
WO2001017523A1
WO2001017523A1 PCT/US2000/024659 US0024659W WO0117523A1 WO 2001017523 A1 WO2001017523 A1 WO 2001017523A1 US 0024659 W US0024659 W US 0024659W WO 0117523 A1 WO0117523 A1 WO 0117523A1
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Prior art keywords
skin
acid compound
mammal
ursohc
content
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PCT/US2000/024659
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English (en)
Inventor
David A. Brown
Daniel B. Yarosh
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Applied Genetics Incorporated Dermatics
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Priority to JP2001521314A priority Critical patent/JP2003508486A/ja
Priority to KR1020027003107A priority patent/KR20020032577A/ko
Priority to US10/070,466 priority patent/US6982284B1/en
Priority to AU73588/00A priority patent/AU7358800A/en
Priority to EP00961668A priority patent/EP1210075A4/fr
Publication of WO2001017523A1 publication Critical patent/WO2001017523A1/fr

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
    • A61Q19/00—Preparations for care of the skin
    • A61Q19/007—Preparations for dry skin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/185—Acids; Anhydrides, halides or salts thereof, e.g. sulfur acids, imidic, hydrazonic or hydroximic acids
    • A61K31/19—Carboxylic acids, e.g. valproic acid
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K8/00—Cosmetics or similar toiletry preparations
    • A61K8/02—Cosmetics or similar toiletry preparations characterised by special physical form
    • A61K8/14—Liposomes; Vesicles
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K8/00—Cosmetics or similar toiletry preparations
    • A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
    • A61K8/30—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds
    • A61K8/63—Steroids; Derivatives thereof
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K8/00—Cosmetics or similar toiletry preparations
    • A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
    • A61K8/30—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds
    • A61K8/67—Vitamins
    • A61K8/671—Vitamin A; Derivatives thereof, e.g. ester of vitamin A acid, ester of retinol, retinol, retinal
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P17/00—Drugs for dermatological disorders
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
    • A61Q19/00—Preparations for care of the skin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
    • A61Q19/00—Preparations for care of the skin
    • A61Q19/08—Anti-ageing preparations

Definitions

  • the epidermis and dermis of mammalian skin contain different cell types, perform different functions, and have different chemical compositions. A particularly important difference between these layers is their lipid concentrations.
  • the dermis contains fibroblasts which produce collagen and other proteins, but very little lipid.
  • the epidermis contains keratinocytes which, among other things, produce lipids, but essentially no collagen.
  • the collagen produced by the fibroblasts provides tensile strength to the skin.
  • the lipids produced by the kerotinocytes provide a barrier between the living tissue and the outside world.
  • the present invention relates to modifying the lipid content of skin for purposes of altering appearance, improving function, improving vitality, reversing the effects of aging, reversing the effects of photodamage, or treating disease by topical administration of ursolic acid, ursolic acid analogs, derivatives of ursolic acid, derivatives of ursolic acid analogs, or combinations thereof. Because the skin lipids are located in the epidermis, this modification of the lipid content of the skin takes place in that layer.
  • ursolic acid, ursolic acid analogs, derivatives of ursolic acid, derivatives of ursolic acid analogs, or combinations thereof will be referred to herein as simply a "ursolic acid compound".
  • the ursolic acid compound can be encapsulated in liposomes or administered in other formulations suitable for topical administration.
  • U.S. Patent 4,857,554 Methods for Treatment of Psoriasis, is directed to treating psoriasis by applying an ointment containing ursolic acid and oleanolic acid dispersed in a petroleum jelly /lanolin carrier.
  • U.S. Patent 4,530,934 Pharmaceutically Active Ursolic Acid Derivatives, is directed to using active derivatives of ursolic acid to treat ulcers.
  • U.S. Patent 3,903,089 Ursolic Acid Derivatives, is directed to the synthesis of ursolic acid derivatives and analogs.
  • U.S. Patent 5,624,909 Derivatives of Triterpenoid Acids as Inhibitors of Cell-adhesion Molecules ELAM-1 (e-selectin) and LECAM-1 (I- selectin), is directed to alleviating inflammation by administration of triterpenoid acid derivatives.
  • U.S. Patent 5,314,877 Water-soluble Pentacyclic Triterpene
  • composition and Method for Producing the Same is directed to making ursolic acid, oleanolic acid, and related triterpenoids soluble in water by formulation in cyclodextrins.
  • U.S. Reissue Patent RE036068 Methods for Treatment of Sundamaged Human Skin with Retinoids, is directed to reversing the effects of photodamage by topical application of retinoids.
  • U.S. Patent 5,051,449 Treatment of Cellulite with Retinoids, is directed towards retarding or reversing cellulite accumulation in skin by topical application of retinoids.
  • U.S. Patent 5,556,844, Pharmaceutical or Cosmetic Composition Treatment of Cellulite with Retinoids, is directed towards retarding or reversing cellulite accumulation in skin by topical application of retinoids.
  • Containing a Combination of a Retinoid and a Sterol is directed towards treatment of disorders of epidermial keratinization, epithelial proliferation, or disorders of sebaceous function by topical application of retinoids.
  • U.S. Patent 5,075,340, Retinoic Acid Glucuronide Preparations for Application to the Skin, is directed towards treatment of acne or wrinkled skin and prevention of retinoid dermatitis by topical application of retinoic acid glucoronides
  • U.S. Patent 5,837,224 Method of Inhibiting Photoaging of Skin, is directed to reversing the effects of photodamage by topical application of agents that inhibit UVB-inducible matrix metalloproteinase.
  • Japanese Patent Publication No. 11-5727 published January 12, 1999, describes the use of ursolic acid in combination with retinols in a final cosmetic product to increase dermal collagen.
  • collagen is located and produced in the dermis by fibroblasts.
  • the present invention is concerned with modifying the content of lipids located and produced in the epidermis by kerotinocytes
  • Phospholipids Most common are phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine, phosphatidylinositol, and sphingomyelin. Phospholipids are the most abundant lipids in the basal layers of the epidermis, but decrease towards the surface of the skin so much so that they are one of the least abundant lipids in the cornified layer. Thus, the phospholipid content of keratinocytes decreases as they differentiate.
  • Glucosylceramide A also known as acylglucosylceramide is the major form, comprising 56 wt.% of this group of lipids.
  • the acyl group in glucosylceramide A is often linoleic acid which is bound to the hydroxyl group of the ⁇ -hydroxyacid (Abraham, Wertz and Downing, 1985, J. Lipid Res. 26:761-766).
  • Ceramides 1-7 Ceramides are the major lipids in the stratum corneum. They result from deglycosylation of glucosylceramides at the end of the epidermal differentiation process.
  • Ceramide 1 is derived from Glucosylceramide A
  • Ceramide 2 is derived from Glucosylceramide B
  • Ceramide 3 is derived from Glucosylceramide C-l
  • Ceramide 4 is derived from Glucosylceramide C-3
  • Ceramide 5 is derived from Glucosylceramide D-l
  • Ceramide 6-A is derived from Glucosylceramide D-2
  • Ceramide 6-B is derived from Glucosylceramide C-2.
  • Glucosylceramide A comprises 56 wt.% of the glucosyleramides
  • Ceramide 1 comprises only 8 wt.% of the extractable ceramides because most of it is converted to the ⁇ -hydroxyceramide which permits covalent binding to glutamates of the cornified cell envelope protein, thus, forming a protective barrier around each corneocyte.
  • Ceramide 2 comprises 42 wt.% of this group.
  • Ceramide 6 (phytosphingosine) and ceramide 7 (6-hydroxy-4- sphingenine) together comprise 20 wt.%.
  • Cholesterol This Hpid increases as keratinocytes differentiate so that it comprises 30 mol% of stratum corneum lipids (Schaefer and Redelmeier, 1996, Skin Barrier).
  • Cholesterol sulfate This Hpid increases ceU cohesiveness by forming interceUular cholesterol sulfate calcium bridges
  • Cholesterol esters During the latter stages of epidermal differentiation, phospholipids are degraded liberating fatty acids which are utilized to produce cholesterol esters. Examination of Hpids in serial sections through pig skin showed the foUowing (Cox and Squier, 1986, J. Invest. Dermatol.
  • Vitamin C (50 ug/ml) has been found to result in increases of glucosylceramides and ceramides, most notably ceramides 6 and 7 in Hving skin equivalents (Ponec et al., 1997, J. Invest. Dermatol. 109:348-355). These increases were accompanied by increased barrier function. Since Vitamin E had no effect on Hpid composition even though it is hydrophobic, it was concluded that the main role of Vitamin C is as a donor of hydroxyl groups to sphingoid bases and fatty acids for the formation of protein-bound hydroxyceramides (Ponec et al., 1997, J. Invest. Dermatol. 109:348-355).
  • UVA 50 J/cm 2
  • UV-B (0.15 J/cm 2 )
  • UV-B short-term exposure to UV-B also resulted in a marked (>2-fold) depletion of phosphoHpids (HoUeran et al, 1997, above).
  • Retinoic acid is weU known as an agent for treatment of photoaged skin.
  • Topical retinoic acid has been shown to restore coUagen I levels that are reduced in photodamaged skin (Griffiths et al., 1993, New Engl. J. Med. 329:530-5).
  • Restoration of coUagen I levels correlate with a reduction of fine wrinkles in skin (Griffiths et al., 1993, New Engl. J. Med. 329:530-5).
  • retinoids have been shown to alter Hpids in cultured skin equivalents (Ponec and Weerheim, 1990, Meth. Enzymol. 190:30-41), there are no reports indicating that retinoids reverse aging or photodamage by altering Hpid levels.
  • retinoids reduce ceramide levels in skin equivalents (Ponec and Weerheim, 1990, Meth. Enzymol. 190:30-41), and reduce the thickness of the stratum corneum when applied topicaUy to human skin (KHgman and Leyden, 1993, Skin Pharmacol. 6, Suppl.1:78-82), which could exacerbate the depletion of ceramides and barrier function that occurs in the aged.
  • UrsoHc acid is pentacyclic triterpene compound known to have a number of pharmacological effects (reviewed in Liu, 1995, J. Ethanopharmacol. 49:57-68). Ursolic acid is closely related to steroids since both are derived from the cycHzation of squalene (Suh et al., 1998, Cancer Res. 58:717-723). It is found in the waxy coating of fruit and in the leaves of many plants, such as heather and rosemary. It is insoluble in most common solvents and as a result it is not widely used. In fact, commercial extraction processes for plant leaves fail to recover measurable levels of ursoHc acid.
  • UrsoHc acid has been characterized as an inhibitor of Hpoxygenase and cyclooxygenase in inflammatory ceUs (Najid et al., 1992, FEBS 299:213-217; Suh et al., 1998, Cancer Res. 58:717-723). As such, ursoHc acid is expected to have usefulness as an anti-inflammatory agent. UrsoHc acid has been shown to inhibit chronic dermal inflammation induced by phorbal esters in an animal model (Manez et al., 1997, Eur. J. Pharmacol. 334:103- 105).
  • UrsoHc acid has also been shown to inhibit induction of inducible nitric oxide synthase in macrophages (Suh et al., 1998, Cancer Res. 58:717- 723), which may contribute to its anti-inflammatory activity. UrsoHc acid has also been shown to induce differentiation and growth arrest of several types of ceUs, suggesting that it may be useful as a chemotherapeutic differentiation agent (Es-Saady et al., 1996, Cancer Lett. 106:193-197; Hsu et al., 1997, Cancer Lett. 111:7-13; Es-Saady et al., 1996, Anticancer Res.
  • UrsoHc acid has also been shown to induce apoptosis in tumor ceUs (Baek et al., 1997, Int. J. Cancer 73:725-728). Both ursoHc acid and oleanoHc acid, a closely related structural analog of ursolic acid, have been shown to inhibit tumor promotion induced in mouse skin by phorbal esters (Tokuda et al., 1986, Cancer Lett. 33:279-285; Huang et al., 1994, Cancer Res. 54:701-708). Both compounds have also been shown to prevent Hpid peroxidation, which may inhibit free radical damage during cancer initiation and promotion (Balanehru and Nagarajan, 1991, Biochem. Int. 24:981-990).
  • Ursolic acid also downregulates matrix metalloproteinases (Cha et al., 1998, Oncogene 16:771-778) and elastase (Ying et al., 1991, Biochem. J. 277:521-526) which may provide a mechanism for preventing tumor invasion (Cha et al., 1996, Cancer Res. 56:2281-84), and, inflammation related damage in skin (Ying et al., 1991, Biochem. J. 277:521-526).
  • UrsoHc acid and a number of triterpenoid derivatives have been shown to have hypolipidemic and anti-atherosclerotic properties (reviewed in Liu, 1995, J. Ethanopharmacol. 49:57-68).
  • UrsoHc acid and oleanoHc acid lowered blood cholesterol and ⁇ -lipoprotein levels 40-50% in animal models of atherosclerosis (reviewed in Liu, 1995, J. Ethanopharmacol. 49:57-68).
  • topical ursolic acid has been proposed for use in the treatment of psoriasis, a condition characterized by hyperproliferation and inflammation of the epidermis (US patent
  • the present invention provides a method for altering the Hpid content of mammalian skin by administering an effective amount of a ursolic acid compound to the skin of a mammal (e.g., a human) in need of such a treatment, e.g., to skin which is aged, photoaged, atrophied, etc.
  • a mammal e.g., a human
  • the alteration of the lipid content of the skin takes place in that layer.
  • the present invention provides a method for reversing certain aspects of the photoaging or aging process in mammalian skin and, in yet another aspect, the present invention provides a method for improving function, increasing barrier function, improving vitaHty, or treating lipid deficient diseases of mammalian skin, which comprises topical appHcation of: (a) an effective amount of a ursoHc acid compound in a suitable medium for topical administration, e.g., a lotion, gel, or the like;
  • ursoHc acid compound encapsulated in Hposomes and incorporated into a suitable medium for topical administration, e.g., a lotion, gel, or the like.
  • ursolic acid compounds are highly insoluble in many solvents, including water, administration of such compounds in Hposomes is a particularly preferred embodiment of the invention.
  • Figure 1 shows micrographs of cultured normal human epidermal keratinocytes (NHEK) that were: (a) Untreated, (b) treated with 1%
  • Figure 2 shows the amount of phosphatidylchoHne per cell relative to Untreated cultured normal human epidermal keratinocytes (NHEK), cultured NHEK treated with 1% (volume/volume) Empty Liposomes, or cultured NHEK treated with 1% 4 mM Ursolic Acid Liposomes. Details are described in Example 1.
  • Figure 3 shows the amount of phosphatidylethanolamine per cell relative to Untreated cultured normal human epidermal keratinocytes (NHEK), cultured NHEK treated with 1% (volume/volume) Empty Liposomes, or cultured NHEK treated with 1% 4 mM Ursolic Acid Liposomes. Details are described in Example 1.
  • Figure 4 shows the amount of free fatty acid per ceU relative to Untreated cultured normal human epidermal keratinocytes (NHEK), cultured NHEK treated with 1% (volume/volume) Empty Liposomes, or cultured NHEK treated with 1% 4 M UrsoHc Acid Liposomes. DetaUs are described in Example 1.
  • Figure 5 shows the amount of total ceramides per ceU relative to Untreated cultured normal human epidermal keratinocytes (NHEK), cultured NHEK treated with 1% (volume/volume) Empty Liposomes, or cultured NHEK treated with 1% 4 mM UrsoHc Acid Liposomes. Details are described in Example 1
  • Figure 6 shows the amount of total glycosylceramides per ceU relative to Untreated cultured normal human epidermal keratinocytes (NHEK), cultured NHEK treated with 1% (volume/volume) Empty Liposomes, or cultured NHEK treated with 1% 4 mM UrsoHc Acid Liposomes. Details are described in Example 1.
  • Figure 7 shows the amount of cholesterol per ceU relative to Untreated cultured normal human epidermal keratinocytes (NHEK), cultured NHEK treated with 1% (volume/volume) Empty Liposomes, or cultured NHEK treated with 1% 4 mM UrsoHc Acid Liposomes. Details are described in Example 1.
  • Figure 8 shows the amount of extractable ceramides in human skin foUowing treatment with 0.3% or 1% UrsoHc Acid Liposomes in a hydrogel Lotion for 3 and 11 days, relative to adjacent areas treated with Empty Liposome Lotion. These concentrations of UrsoHc Acid Liposomes resulted in a final concentration of ursoHc acid in the hydrogel Lotion of lO ⁇ M and 30 ⁇ M, respectively.
  • the compounds and compositions of the present invention effectively and efficiently increase the phospholipid content of normal human epidermal keratinocytes. Furthermore, the compounds and compositions of the present invention increase the free fatty acid content and the ceramide and glycosylceramide content of normal human epidermal keratinocytes. Increases of phosphoHpid, free fatty acid, ceramide and glycosylceramide content are due to the effects of ursoHc acid and are over and above any effects of the lipid carrier. Increasing the lipid content of epidermal keratinocytes has several consequences. First, increasing the phospholipid content prevents ceU senescence and stimulates proliferation, which is manifested as increased ceUular viabiHty and increased epidermal thickening.
  • increasing the free fatty acid content relates to increases of several Hpid fractions including phospholipids, triglycerides, glucoceramides, ceramides and acylceramides.
  • Increasing the Hpid content of epidermal keratinocytes reverses the Hpid reductions associated with aging.
  • Third, increasing the glucoceramide, ceramide and acylceramide content of keratinocytes results in an improved barrier function. Improved barrier function, in turn, reduces atopic dermatitis and protects the skin and body from the effects of many agents including ultraviolet irradiation, toxic chemicals, toxins, and irritants.
  • Fourth, preventing ceU senescence and increasing barrier function reverses the effects of aging.
  • Fifth, increases of phospholipids directly reverse the depleting effect of UV-B on phosphoHpids in skin, resulting in increased viabiHty as manifested by cellular proliferation, and reducing the effects of photoaging.
  • the users of this invention who benefit from its discovery include, among others, those suffering from ichthyosis and ichthyosiform dermatoses, those with acute dry skin, those with skin atrophy and retinoid dermatoses, those with acne and those with aging and photoaging skin.
  • the active compound according to the present invention is a ursoHc acid compound.
  • Examples of such compounds are set forth in Table 1, it being understood that the invention is not Hmited to the examples of this table but includes aU ursolic acid compounds which achieve the beneficial, lipid altering effects of the invention when appHed to mammalian skin.
  • the table shows the chemical structure common to this famfly of compounds.
  • the reference compound, ursoHc acid contains the substituents at the indicated sites as shown in the second Hne of the table.
  • Each of the analogs/derivatives Hsted below ursoHc acid differs from the ursolic acid structure only where indicated. Blank cells indicate that the substituents at those sites are identical to ursoHc acid.
  • these compounds share great similarity in the A, B, and C rings of the pentacycHc structure.
  • the methods and compositions of the present invention employ a ursoHc acid compound (which, as defined above, can be a combination (mixture) of compounds) as an active ingredient for various uses.
  • the active ingredient is given topically in an acceptable formulation.
  • a particularly preferred formulation is the incorporation of the ursoHc acid compound into liposomes.
  • a variety of different types of lipids at various concentrations can be used to form the liposomes, examples of which can be found in Liposome Technology, ed. Gregory Gregoriadis, CRC Press Inc., Boca Raton, Florida 1984.
  • the present invention also relates to the incorporation of the ursoHc acid compound, either alone or incorporated in Hposomes, into lotions, gels, creams or other acceptable formulations conducive to uptake of active ingredients into the epidermis.
  • Liposomal formulations are preferred because ursoHc acid is highly insoluble in many solvents, particularly water, and common emulsifiers such as LECINOL S-10 have Httle effect.
  • this insolubility problem is addressed by taking advantage of the flat, planar structure of ursoHc acid to stack it between the lipid tails in the phosphoHpid bflayer membranes of liposomes. Due to the charged head- group of the phospholipids, Hposomes containing ursolic acid are readily soluble in water.
  • ursolic acid only a Hmited number of sites within the tails of the lipid bflayer of the Hposome membrane are available for stacking ursolic acid.
  • the preferred ratio of ursolic acid to lipid components must be determined by experimentation. For example, the preferred ratio of ursoHc acid to phosphatidylcholine and cholesterol is approximately 1.5 (range 1.0 to 3.0):10:1.9 (w/v).
  • concentrations of ursoHc acid the effects on lipid production by the epidermal layer of mammalian skin are not easfly evoked.
  • other compounds that partition into the liposome membrane cannot be included in the preparation, because they displace the ursoHc acid and lead to its precipitation.
  • the preferred preservatives that are compatible with ursoHc acid Hposomes are water-soluble preservatives that do not partition into the Hposome membrane.
  • One such water-soluble preservative that can be used in the practice of the invention is potassium sorbate.
  • Other water soluble preservatives can be found in: Cosmetic and Drug Preservation. Principles and Practices. Ed. J. J. Kabara. Marcel Dekker, Inc. New York, 1984.
  • additives to a liposomal ursolic acid composition should not displace the ursoHc acid from the liposome, and the preferred form of such additives should be water soluble or otherwise sequestered from the Hposomes.
  • the dose regimen wiU depend on a number of factors which may be readfly determined, such as severity and responsiveness of the condition to be treated, but will normaUy be one or more treatments per day, with treatment lasting from several days to several months, or untfl the desired response is obtained, or a cure is effected, or a remediation of a condition or a diminution of a disease state is achieved.
  • factors which may be readfly determined such as severity and responsiveness of the condition to be treated, but will normaUy be one or more treatments per day, with treatment lasting from several days to several months, or untfl the desired response is obtained, or a cure is effected, or a remediation of a condition or a diminution of a disease state is achieved.
  • One of ordinary skill may readfly determine optimum dosages, dosing methodologies and repetition rates.
  • the unit dosage for compositions according to the present invention will contain from 0.1 mM to 10 mM of the ursoHc acid compound in Hposomes or an alternative carrier, with said Hposomes or carrier comprising from 1 wt.% to 90 wt.% of a lotion or alternative topical formulation.
  • said Hposomes or alternative formulations full strength to skin.
  • the ursoHc acid compound can be combined with other active ingredients in the formulation.
  • retinoids and topical steroids have the undesirable side effect of skin atrophy. This side effect can be ameHorated by co-administration of a ursoHc acid compound with these agents.
  • the co-administration can be performed using a single vehicle, e.g., a lotion, containing both active ingredients or by means of separate vehicles, e.g., separate lotions, which can be administered either simultaneously or sequentiaUy, with either agent being administered first.
  • a ursolic acid compound can be combined with a sunblock to form a sunscreen product.
  • Ursolic acid-containing Hposomes and empty liposomes containing no ursolic acid were prepared as foUows: 0.393 g phosphatidylchoHne (DOOSAN) and 0.077 g cholesterol (AVANTI) were dissolved in 20 ml ethanol, and split into two 10 ml aHquots. Sixty mg ursolic acid (SIGMA- ALDRICH) was then dissolved in one aHquot which was then used to make UrsoHc Acid Liposomes. No additions were made to the aliquot designated as Empty Liposomes. Seven ml of each mixture were injected through a 3O 2G needle into 10 ml cold 1XPBS.
  • DOOSAN phosphatidylchoHne
  • AVANTI 0.077 g cholesterol
  • CLONETICS BIOWHITTAKER and cultured in KGM-2 media (CLONETICS-BIOWHITTAKER) according to the manufacturer's instructions.
  • 10 5 NHEK were plated in each of six 10 cm 2 CORNING cell culture dishes in 10 ml KGM-2 media.
  • media was replaced with 10 ml fresh media.
  • 100 ul Empty Liposomes were added to each of two dishes, and 100 ul 4 mM Ursolic Acid Liposomes were added to each of two dishes.
  • the Empty Liposome and 4 M Ursolic Acid Liposome treatments each received 1% Hposomes, and the final concentration of ursoHc acid in the UrsoHc Acid Liposome treatment was 40 uM. All treatments received fresh media and Hposomes on the fourth day foUowing the initiation of treatments.
  • CeUs were harvested 8 days foUowing the initiation of treatments. Media was removed and ceUs were washed once with 10 ml CLONETICS- BIOWHITTAKER Hank's Buffered SaHne Solution (HBSS), then 5 ml
  • HBSS was added, and ceUs were photographed using a NIKON microscope equipped with camera Hnked to a NORTHERN EXPOSURE computer imaging system. FoUowing photograph, HBSS was removed, 6 ml CLONETICS Trypsin/EDTA was added, and ceUs were incubated at 37°C for 6 min untfl they detached from dishes. Then, 6 ml CLONETICS Trypsin NeutraHzation Solution was added, and the ceUs were mixed thoroughly and transferred to a 15 ml conical tube. One-half ml of suspended ceUs were added to 19.5 ml Isoton II and counted on a model ZBI COULTER counter. The remainder of cells were then pelleted by centrifugation at 178 X g for 5 min. Supernatant was removed and ceUs were then resuspended in 5 ml
  • Lipids were extracted using procedures developed by Ponec and Weerheim (1990, Meth. Enzymol. 190:30-41), that were a modification of procedures developed by BHgh and Dyer (1959, Canad. J. Biochem. Physiol. 37:911-917).
  • Pelleted ceUs were extracted by mixing with 2 ml chlorform: methanol (2:1) for 60 min at room temperature (RT) on a LABQUAKE rotary mixer.
  • Both the upper aqueous layer and the bottom extract layer were transferred to clean tubes, and the aqueous layer was re-extracted with 4 ml chloroform by mixing for 10 min at RT.
  • This chloroform extract was combined with the previous extracts.
  • the combined extracts were placed in a 50°C water bath and dried under a stream of nitrogen.
  • the peUet was dissolved in 500 ul chloroform(2):methanol(l) and stored in a teflon lined vial under nitrogen at -20°C.
  • Hpids were extracted from subjects using a modification of the protocol described by Bonte, F., A. Saunois, P. Pinguet, and A. Meybeck, in Arch. Dermatol. Res. 289:78-82, 1997. FoUowing treatments, the area on the forearm to be extracted was first rinsed with tap water, dried thoroughly and then tape-stripped once with SCOTCHTM 810 MAGICTM tape. The excised top 1-inch of 50 ml polypropylene conical tubes were used as reservoirs for the solvents during extraction of subjects. The reservoirs were placed and held firmly on the arm and 1 ml of cyclohexane:ethanol (4:1) was added and stirred gently for one minute.
  • Solvent was then removed and placed into a pyrex tube with a teflon-Hned Hd.
  • One ml of cyclohexane:ethanol (1:1) was then added to the reservoir, stirred for one minute, and then removed and added to the tube containing the first extract.
  • the tubes were then dried at 50°C under nitrogen gas as described above.
  • the dried extracts were dissolved in 200 ⁇ l of chloroform:methanol (2:1).
  • the Hpid solution was then placed into a smaU storage tube with a teflon-Hned lid, purged with nitrogen gas and stored at -20°C.
  • HPTLC plates were developed using the foUowing sequence of mobile phases for FFA and CH: (i) chloroform run 15 mm, (ii) chloroform-acetone- methanol (76:8:16) run 10 mm, (iii) chloroform -hexyl acetate-acetone- methanol (86:1:10:4) run 70 mm, (iv) chloroform-acetone-methanol (76:4:20) run 20 mm, (v) chloroform-diethyl ether-hexyl acetate-ethyl acetate- acetone -methanol (72:4:1:4:16:4) run 75 mm, and, (vi) hexane-diethyl ether- ethyl acetate (80:16:4) run 90 mm (Ponec and Weerheim, 1990, Meth.
  • Enzymol. 190:30-41 the foUowing sequence of mobile phases for Cer and GlyCer: (i) chloroform-methanol-water (40:10:1) run 30 mm, (n) chloroform-methanol-glacial acetic acid (190:9:1) run 75 mm, (iii) hexane- diethyl ether-glacial acetic acid (80:20:10) run 75 mm, and (iv) petroleum ether run 85 mm (Kennedy et al., 1996, Pharmaceut. Res. 13:1162-1167). TLC plates were dried, stained with iodine, photographed, and analyzed using a QUANTASCAN computer imaging system.
  • HPTLC plates were dried, sprayed with phosphomolybdic acid, baked at 120°C for 2 min for FFA and CH, or, sprayed with 10% copper sulfate in 8% phosphoric acid for Cer and GlyCer, and analyzed similar to TLC plates. Intensity of standard staining was found to be linear up to at least 0.625 ug/ul, and this was therefore used as a standard on chromatographic runs with samples.
  • vacuolation observed here is specific for UrsoHc Acid Liposome treated ceUs, indicating that ursoHc acid is responsible for induction of vacuolation.
  • photographs in Figure 1 were taken after 8 days of treatment, visual observation indicated that significant vacuolation of Ursolic Acid Liposome treated ceUs was apparent as soon as 2 days after the initiation of treatments.
  • Hposomal ursolic acid was a more optimal concentration for induction of ceramides in human skin. However, it should be noted that whereas cell culture treatments were done only once every third day, treatment of human skin was done twice daily. Thus, the ability of Hposomal ursolic acid to induce ceramides in human skin may have been saturated by this treatment regimen.
  • the compounds of the present invention result in increased levels of phosphoHpids in ceUs.
  • incorporation of the compounds of the present invention into liposomes results in accumulation of additional Hpids, including total ceramides and glycosylceramide s which are extremely important for barrier function.
  • the barrier function of skin wiU not be compromised by the compositions of the present invention.
  • the compositions will stimulate phospholipid synthesis and thereby increase ceU viability.
  • the topical administration of a ursoHc acid compound in accordance with the invention will serve to counteract the effects of aging and photoaging and to treat diseases of dry skin related to impaired barrier function and dysfunctional stratum corneum.
  • Ursolic acid is the reference compound. Blank cells indicate that the site is identical to ursolic acid.

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Abstract

L'invention se rapporte à l'usage topique de composés d'acide ursolique pouvant modifier le taux de lipides cutanés chez un mammifère. Les composés peuvent être encapsulés dans des liposomes et administrés comme tels à la peau, par exemple, sous forme de lotion ou de gel. Les composés de l'invention sont efficaces en ce qu'ils ont, entre autres qualités, celles de diminuer les effets du vieillissement, du photovieillissement, et de l'atrophie cutanée, notamment l'atrophie cutanée résultant de l'usage topique de rétinoïdes et/ou de stéroïdes. L'invention se rapporte en outre à des compositions comprenant des composés d'acide ursolique associés à d'autres composés topiques ayant une action thérapeutique, tels que, par exemple, un rétinoïde et/ou un stéroïde.
PCT/US2000/024659 1999-09-10 2000-09-08 Compositions et methodes pour modifier le taux de lipides cutanes WO2001017523A1 (fr)

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JP2001521314A JP2003508486A (ja) 1999-09-10 2000-09-08 皮膚脂質含量の改良のための組成物及び方法
KR1020027003107A KR20020032577A (ko) 1999-09-10 2000-09-08 피부 지질 함량 조절을 위한 조성물 및 방법
US10/070,466 US6982284B1 (en) 1999-09-10 2000-09-08 Compositions and methods for modification of skin lipid content
AU73588/00A AU7358800A (en) 1999-09-10 2000-09-08 Compositions and methods for modification of skin lipid content
EP00961668A EP1210075A4 (fr) 1999-09-10 2000-09-08 Compositions et methodes pour modifier le taux de lipides cutanes

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FR2830195A1 (fr) * 2001-10-03 2003-04-04 Sederma Sa Compositions cosmetiques et dermopharmaceutiques pour les peaux a tendance acneique
EP1410790A1 (fr) * 2002-10-09 2004-04-21 Amorepacific Corporation Submicro-Liposomes comprenant des triterpenoides et la méthode de préparation
KR100427777B1 (ko) * 2001-09-25 2004-04-28 엔프라니 주식회사 나노캡슐화된 울솔산 및 이의 제조방법
WO2006132033A1 (fr) 2005-06-10 2006-12-14 Pola Chemical Industries Inc. Nouveau dérivé d’acide triterpénique et préparation comprenant celui-ci pour une application externe sur la peau
WO2009010687A2 (fr) * 2007-07-06 2009-01-22 Galderma Research & Development Enzyme cible dans le traitement de l'acne
WO2012024395A3 (fr) * 2010-08-19 2013-05-02 Johnson & Johnson Consumer Companies Inc. Compositions comprenant de la paulownine et/ou des extraits de paulownia et utilisations de celles-ci
US9161958B2 (en) 2010-08-19 2015-10-20 Johnson & Johnson Consumer Inc. Methods of treating cellulite
US9168279B2 (en) 2010-08-19 2015-10-27 Johnson & Johnson Consumer Inc. Compositions comprising paulownin and/or Paulownia extracts and uses thereof
US9168207B2 (en) 2010-08-19 2015-10-27 Johnson & Johnson Consumer Inc. Compositions comprising Paulownia tomentosa wood extracts and uses thereof
US9168219B2 (en) 2010-08-19 2015-10-27 Johnson & Johnson Consumer Inc. Compositions comprising Paulownia tomentosa wood extracts and uses thereof
US9173913B2 (en) 2010-08-19 2015-11-03 Johnson & Johnson Consumer Inc. Compositions comprising Paulownia tomentosa wood extracts and uses thereof
US9387349B2 (en) 2010-08-19 2016-07-12 Johnson & Johnson Consumer Inc. Compositions comprising Paulownia tomentosa wood extracts and uses thereof
US9962326B2 (en) 2010-08-19 2018-05-08 Johnson & Johnson Consumer Inc. Compositions comprising paulownia tomentosa wood extracts and uses thereof
CN112168785A (zh) * 2019-07-03 2021-01-05 上海交通大学 熊果酸脂质体制剂及其制备方法和用途

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JP4813917B2 (ja) * 2005-02-22 2011-11-09 丸善製薬株式会社 皮膚化粧料
JP2012171932A (ja) * 2011-02-23 2012-09-10 Kao Corp アロマターゼ活性化剤
CN107922962A (zh) * 2015-04-10 2018-04-17 俄勒冈州立大学 皮肤脂质组学测定
JP2018058800A (ja) * 2016-10-07 2018-04-12 国立大学法人九州大学 メラニン抑制剤
TW202216160A (zh) * 2020-07-16 2022-05-01 學校法人立命館 皮脂產生調節劑

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KR100427777B1 (ko) * 2001-09-25 2004-04-28 엔프라니 주식회사 나노캡슐화된 울솔산 및 이의 제조방법
US7182963B2 (en) 2001-10-03 2007-02-27 Sederma Cosmetic and dermopharmaceutical compositions for skin prone to acne
WO2003028692A2 (fr) * 2001-10-03 2003-04-10 Sederma Traitement de l'acne a l'aide d'acide oleanolique
WO2003028692A3 (fr) * 2001-10-03 2003-10-02 Sederma Sa Traitement de l'acne a l'aide d'acide oleanolique
FR2830195A1 (fr) * 2001-10-03 2003-04-04 Sederma Sa Compositions cosmetiques et dermopharmaceutiques pour les peaux a tendance acneique
EP1410790A1 (fr) * 2002-10-09 2004-04-21 Amorepacific Corporation Submicro-Liposomes comprenant des triterpenoides et la méthode de préparation
EP1889850A4 (fr) * 2005-06-10 2011-08-03 Pola Chem Ind Inc Nouveau dérivé d acide triterpénique et préparation comprenant celui-ci pour une application externe sur la peau
EP1889850A1 (fr) * 2005-06-10 2008-02-20 Pola Chemical Industries Inc. Nouveau dérivé d acide triterpénique et préparation comprenant celui-ci pour une application externe sur la peau
WO2006132033A1 (fr) 2005-06-10 2006-12-14 Pola Chemical Industries Inc. Nouveau dérivé d’acide triterpénique et préparation comprenant celui-ci pour une application externe sur la peau
KR101254938B1 (ko) * 2005-06-10 2013-04-16 포라가세이고오교오가부시끼가이샤 신규 트리테르펜산 유도체 및 이를 포함하는 피부 외용제
WO2009010687A2 (fr) * 2007-07-06 2009-01-22 Galderma Research & Development Enzyme cible dans le traitement de l'acne
WO2009010687A3 (fr) * 2007-07-06 2009-05-22 Galderma Res & Dev Enzyme cible dans le traitement de l'acne
US9161958B2 (en) 2010-08-19 2015-10-20 Johnson & Johnson Consumer Inc. Methods of treating cellulite
WO2012024395A3 (fr) * 2010-08-19 2013-05-02 Johnson & Johnson Consumer Companies Inc. Compositions comprenant de la paulownine et/ou des extraits de paulownia et utilisations de celles-ci
US9168279B2 (en) 2010-08-19 2015-10-27 Johnson & Johnson Consumer Inc. Compositions comprising paulownin and/or Paulownia extracts and uses thereof
US9168207B2 (en) 2010-08-19 2015-10-27 Johnson & Johnson Consumer Inc. Compositions comprising Paulownia tomentosa wood extracts and uses thereof
US9168219B2 (en) 2010-08-19 2015-10-27 Johnson & Johnson Consumer Inc. Compositions comprising Paulownia tomentosa wood extracts and uses thereof
US9173913B2 (en) 2010-08-19 2015-11-03 Johnson & Johnson Consumer Inc. Compositions comprising Paulownia tomentosa wood extracts and uses thereof
US9387349B2 (en) 2010-08-19 2016-07-12 Johnson & Johnson Consumer Inc. Compositions comprising Paulownia tomentosa wood extracts and uses thereof
US9962326B2 (en) 2010-08-19 2018-05-08 Johnson & Johnson Consumer Inc. Compositions comprising paulownia tomentosa wood extracts and uses thereof
CN112168785A (zh) * 2019-07-03 2021-01-05 上海交通大学 熊果酸脂质体制剂及其制备方法和用途
CN112168785B (zh) * 2019-07-03 2022-08-30 上海交通大学 熊果酸脂质体制剂及其制备方法和用途

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