WO2025137844A1

WO2025137844A1 - Cosmetic compositions comprising robinin

Info

Publication number: WO2025137844A1
Application number: PCT/CN2023/141750
Authority: WO
Inventors: Sweelin CHEW; Marie-Céline FRANTZ; Richard Betts; Wagner Magalhaes; Edson KATEKAWA
Original assignee: LOreal SA
Current assignee: LOreal SA
Priority date: 2023-12-26
Filing date: 2023-12-26
Publication date: 2025-07-03
Anticipated expiration: 2026-06-26
Also published as: FR3157204A3

Abstract

A cosmetic composition comprises a compound of formula (I). The cosmetic composition is used for promoting and/or speeding up skin regeneration, for preventing and/or treating signs of aging, and for promoting and/or speeding up wound healing. The cosmetic method comprises the application of the composition.

Description

COSMETIC COMPOSITIONS COMPRISING ROBININ

The present invention relates to cosmetic compositions comprising a compound of formula (I) as described hereafter or an optical isomer thereof, or a cis–trans isomer thereof, or a tautomer thereof or a salt thereof, or a hydrate thereof, and their use for promoting and/or speeding up skin regeneration, for preventing and/or treating signs of aging, and for promoting and/or speeding up wound healing. The invention also concerns cosmetic methods comprising the application of such compositions.

Human skin is made up of two compartments: a deep compartment, the dermis, and a superficial compartment, the epidermis. The epidermis is in contact with the external environment, and its role is to protect the body from dehydration and external chemical or mechanical aggression.

Accelerated wound closure is an area of intense scientific research to improve healing that follows skin injury, or within skin that has compromised barrier function. For example, the cutaneous barrier can be unbalanced for example after a cosmetic procedure such as laser treatments and chemical peels, or in the presence of external aggressors such as irritant agents (detergents, acids, bases, oxidants, reducing agents, concentrated solvents, toxic gases or fumes) , mechanical stresses (friction, shocks, abrasion, tearing of the surface, projection of dust, particles, shaving or hair removal) , thermal or climatic imbalances (cold, dryness, radiation) , xenobiotics (undesirable microorganisms, allergens) or internal attacks such as psychological stress. Following such external aggressions, a healing process is triggered, aimed at rapidly and completely restoring this barrier. This physiological process is dependent on complex biological mechanisms involving numerous molecules and enzymes.

There is a need for compositions that promote skin healing, and in particular, that accelerate cutaneous healing.

The aim of the invention is therefore to provide a cosmetic composition that meets all these requirements.

The inventors made the surprising discovery that the natural molecules of formula (I) are able to accelerate wound healing and thus are potentially of utility for skin regeneration.

The present application provides compositions to accelerate skin regeneration following skin wounding or following cosmetic procedure, to promote recovery following said skin injury and/or improve the efficacy of cosmetic skin procedures. Use of the compositions will result in a faster healing time and recovery from the procedure, resulting in greater willingness of the patient to undergo cosmetic procedures.

Thus, the present invention relates to a cosmetic composition, comprising in a cosmetically acceptable medium:

- at least one compound of formula (I) or an optical isomer thereof, or a cis–trans isomer thereof, or a tautomer thereof or a salt thereof, or a hydrate thereof, and

- at least one compound chosen among thickeners, preservatives, perfumes, bactericides, pigments, dyes, carbon and/or silicone oils of inorganic, waxes, fillers, emulsifiers, co-emulsifiers, UVA and/or UVB light protection agents also called UV filters, polymers, hydrophilic and lipophilic gelling agents.

Within the meaning of the present invention, and unless a different indication is given:

- a "sugar" radical is a monosaccharide or a disaccharide radical. Examples of sugar radicals are sucrose (or saccharose) , glucose, galactose, rhamnose, galactose, ribose, fucose, maltose, fructose, mannose, arabinose, xylose or lactose;

- a “monosaccharide" means a monosidic sugar comprising at least 5 carbon atoms of formula C_x (H₂O) _x, with x being an integer greater than or equal to 5, preferably x is greater than or equal to 6, in particular x is from 5 to 7, preferably x = 6; They may be of D or L configuration, and of alpha or beta anomer, as well as one of its salts or solvates such as one of its hydrates;

- "disaccharide" means a di-osidic sugar which is a compound consisting of two oses linked together by O-osidic bonds, the said compounds consisting of two monosaccharides (also called mono-osidic) as defined above, the said monosaccharide units comprising at least 5 carbon atoms, preferably 6, in particular the monosaccharide units are linked to one another in 1, 4 or 1, 6, of alpha or beta anomer, each osidic unit being of L or D configuration, as well as one of its salts or solvates such as hydrates. A disaccharide is more particularly a polymer formed from two oses (or monosaccharides) having the general formula: [C_x (H₂O) _y) ] ₂ or [ (CH₂O) _x] ₂, with x being an integer greater than or equal to 5, preferably x is greater than or equal to 6, in particular x is from 5 to 7, preferably x = 6, and y is an integer which represents x-1;

- "salt" means a cosmetically acceptable salt, i.e. one that is compatible with application to the skin, mucous membranes and/or appendages.

- in the following, and unless otherwise indicated, the limits of a range of values are included in this range, in particular in the expressions "between" and "ranging from ... to ..." ; .

- the expression "at least one" used in the present description is equivalent to the expression "one or more" .

Another object of the invention is the use of said compound or an optical isomer thereof, or a cis–trans isomer thereof (such as an alpha or beta anomer) , or a tautomer thereof or a salt thereof, or a hydrate thereof, or a composition according to the invention, to promote and/or speed up skin regeneration.

Another object of the invention is the use of said compound or an optical isomer thereof, or a cis–trans isomer thereof (such as an alpha or beta anomer) , or a tautomer thereof or a salt thereof, or a hydrate thereof, or a composition according to the invention, to prevent and/or treat signs of aging.

Another object of the invention is the use of said compound or an optical isomer thereof, or a cis–trans isomer thereof (such as an alpha or beta anomer) , or a tautomer thereof or a salt thereof, or a hydrate thereof, or a composition according to the invention, to promote and/or speed up wound healing.

The present invention also relates to a non-therapeutic method of treatment of the skin comprising the step of applying said at least one compound, or an optical isomer thereof, or a cis–trans isomer thereof (such as an alpha or beta anomer) , or a tautomer thereof, or a salt thereof, or a hydrate thereof, or the composition according to the invention, on the skin.

Another object of the invention is a method of treating skin to promote and/or speed up wound healing after aesthetic procedures, the method comprising:

(a) treating the skin with at least one skin modification stimulus, and

(b) after the step (a) applying a composition according to the invention, to the skin.

Other subjects and characteristics, aspects and advantages of the invention will emerge even more clearly on reading the description and the examples that follows.

Compositions

The composition according to the invention comprises at least one compound of formula (I) or an optical isomer thereof, or a cis–trans isomer thereof (such as an alpha or beta anomer) , or a tautomer thereof or a salt thereof, or a hydrate thereof:

wherein:

- R' and R” are identical or different, preferably different, and represent a hydrogen, or a (C1-C4) alkyl such as a methyl, an ethyl, a propyl, an isopropyl, a butyl, and an isobutyl, preferably a methyl, or a monosaccharide or a polysaccharide comprising up to 20 sugar units, preferably up to 6 sugar units, in pyranose and/or furanose form and of L and/or D series, the said mono-or polysaccharide may be substituted by an obligatorily free hydroxyl group, and/or optionally one or more amine function (s) , optionally protected amine function (s) , and the said monosaccharide or polysaccharide optionally being substituted on its -CH₂OH group by a -C (O) -CO₂H group and the said monosaccharide or polysaccharide optionally being substituted on one or more of its hydroxy groups by a (C1-C4) acyl group, preferably an acetyl group, or by a hydroxycinnamyl moiety, chosen among coumaroyl, caffeoyl, feruloyl, or sinapoyl ;

- R”’ and R”” are identical or different, and represent a hydrogen or a (C1-C4) alkyl, such as a methyl, an ethyl, a propyl, an isopropyl, a butyl, and an isobutyl, preferably a methyl,

being understood that R’ , R” , R”’ , and R”” cannot represent a hydrogen at the same time.

Advantageously, the said compound of formula (I) or an optical isomer thereof, or a cis–trans isomer thereof (such as an alpha or beta anomer) , or a tautomer thereof or a salt thereof, or a hydrate thereof, is in the form of a plant extract.

Advantageously, R' and R” are identical or different, preferably different and represent a monosaccharide or a polysaccharide containing up to 6 sugar units, in pyranose and/or furanose form and of L and/or D series, said mono-or polysaccharide having at least one obligatorily free hydroxyl function and/or optionally one or more obligatorily protected amine function (s) .

Advantageously, the monosaccharide is chosen from D-glucose, D-allose, D-galactose, D-mannose, D-xylose, D-lyxose, L-fucose, L-arabinose, L-rhamnose, D- glucuronic acid, D-galacturonic acid, D-iduronic acid, N-acetyl-D-glucosamine, N-acetyl-D-galactosamine and advantageously designates D-glucose, L-rhamnose, D-xylose, N-acetyl-D-glucosamine or L-fucose and more preferably L-rhamnose.

Advantageously, the polysaccharide containing up to 6 sugar units is chosen from D-maltose, D-lactose, D-cellobiose, D-maltotriose, a disaccharide combining a uronic acid chosen from D-iduronic acid or D-glucuronic acid with a hexosamine chosen from D-galactosamine, D-glucosamine, N-acetyl-D-galactosamine, N-acetyl-D-glucosamine, a disaccharide combining L-rhamnose and D-galactose, an oligosaccharide containing at least one xylose which may advantageously be chosen from xylobiose, methyl-β-xylobioside, xylotriose, xylotetraose, xylopentaose and xylohexaose and in particular xylobiose which is composed of two xylose molecules linked by a 1-4 bond.

More preferably, R' and R” are different and represent a monosaccharide or a polysaccharide chosen from D-glucose, D-allose, D-xylose, L-fucose, L-rhamnose, D-galactose, and D-maltose, preferably L-rhamnose and D-galactose as a monosaccharide or a disaccharide. More preferably, R' represents a monosaccharide such as L-rhamnose and R” represents a disaccharide such as a disaccharide combining L-rhamnose and D-galactose.

More preferably, R”’ and R”” represent a hydrogen.

The monosaccharide or polysaccharide of R' and/or R” may be substituted on one or more hydroxymethyl residue (s) by a -C (O) -COOH group.

The salt of the compound of formula (I) can be an alkali or alkaline-earth metal salt, preferably a calcium, magnesium, sodium or potassium salt.

Preferably, the compound of formula (I) or an optical isomer thereof, or a cis–trans isomer thereof (such as an alpha or beta anomer) , or a tautomer thereof, or a salt thereof, or a hydrate thereof, is chosen from robinin and its salts.

The compound of formula (I) or an optical isomer thereof, or a cis–trans isomer thereof (such as an alpha or beta anomer) , or a tautomer thereof or a salt thereof, or a hydrate thereof, preferably robinin or one of its salts, is present in solubilized form in the composition. By "in solubilised form" is meant that the compound does not form crystals visible to the naked eye in the composition.

Robinin (C₃₃H₄₀O₁₉) (or Kaempferol-3-o-gal-rham-7-o-rham, or 3- [ [6-o- (6-deoxy-α-l-mannopyranosyl) -β-d-galactopyranosyl] oxy] -7- [ (6-deoxy-α-l-mannopyranosyl) oxy] -5-hydroxy-2- (4-hydroxyphenyl) -4h-1-benzopyran-4-one ; IUPAC name 4′, 5-Dihydroxy-3- [α-L- rhamnopyranosyl- (1→6) -β-D-galactopyranosyloxy] -7- (α-L-rhamnopyranosyloxy) flavone) is the compound of the following formula (II) :

Robinin can exist in two anomers forms, α and β. The compound of formula (II) is the β anomer of robinin.

Preferably, the compound of formula (I) or one of its optical, cis–trans isomers or tautomeric isomers, or one of its salts or hydrates is chosen from robinin and its salts, more preferably in β-form.

Robinin is a molecule derived from kaempferol. It is a flavone in the form of a yellow-orange powder. This molecule is found, for example, in the leaves and seeds of the black locust (Robinia pseudoacacia) .

Robinin, for example, is marketed by INTERCHIM under the name or by SIGMA under the name

Preferably, the composition comprises between 0.001%and 5%by weight relative to the total weight of the composition, more preferably between 0.01 and 4%, even more preferably between 0.02 and 3%, or between 0.03 and 1%by weight relative to the total weight of the composition, of the at least one compound of formula (I) or an optical isomer thereof, or a cis–trans isomer thereof (such as an alpha or beta anomer) , or a tautomer thereof or a salt thereof, or a hydrate thereof.

Preferably, the composition comprises between 0.001%and 5%by weight relative to the total weight of the composition, more preferably between 0.01 and 4%, even more preferably between 0.02 and 3%, or between 0.03 and 1%by weight relative to the total weight of the composition, of the total amount of compound of formula (I) and optical isomer thereof, and cis–trans isomer thereof, and tautomer thereof and salt thereof, and hydrate thereof.

Preferably, the composition comprises between 0.001 and 5%by weight of robinin or one of its salts relative to the total weight of the composition, preferably between 0.01 and 4%by weight, more preferably between 0.02 and 3%by weight, most preferably between 0.03 and 1%by weight.

Unless mention of the contrary, the percentages are expressed by weight of the total weight of the composition.

Eventually, the composition according to the invention comprises ingredients customary in the cosmetic field.

The composition according to the invention comprises at least one compound chosen among thickeners, preservatives, perfumes, bactericides, pigments, dyes, carbon and/or silicone oils of inorganic, waxes, fillers, emulsifiers, co-emulsifiers, UVA and/or UVB light protection agents also called UV filters, polymers, hydrophilic and lipophilic gelling agents. A person skilled in the art can adjust the type and amount of these ingredients in the compositions according to the invention by means of routine operations, so that the desired cosmetic properties and stability properties for these compositions are not negatively affected. The quantities of these various ingredients are conventionally the quantities used in the particular domain, for example from 0.01%to 20%of the total weight of the composition.

In an embodiment, the composition of the invention further comprises one or more skin active agents chosen among anti-aging agents and wound healing agents.

The term "anti-aging agents" denotes any cosmetic agents known to those skilled in the art suitable for reducing and/or slowing the progression of the signs of aging of the skin such as marked microrelief of the skin, fine lines, appearance of wrinkles, loss of tonicity, loss of density loss of firmness, loss of elasticity, decrease in the thickness of the dermis and/or epidermis, dryness, withered skin, loss of the radiance of the skin complexion, appearance of a wizened appearance of the skin, sagging of the skin, withering of the skin and loss of the capacity of the skin to regenerate.

The term "wound healing agents" denotes any cosmetic agents known to those skilled in the art suitable for promoting and/or speeding up wound healing.

The one or more skin active agents may be included in the cosmetic composition in an amount ranging from 0.001%to 5%by weight relative to the total weight of the composition, more preferably between 0.01 and 2%, even more preferably between 0.02 and 1%, or between 0.05 and 1%by weight relative to the total weight of the composition.

In another embodiment, the composition of the invention only comprises the said at least one compound of formula (I) or an optical isomer thereof, or a cis–trans isomer thereof (such as an alpha or beta anomer) , or a tautomer thereof or a salt thereof or a hydrate thereof, as skin active agent.

The composition according to the invention comprises a cosmetically acceptable medium.

Here, "cosmetically acceptable medium" means a non-toxic support that can be applied on keratin materials such as the skin and mucous membranes. Thus a cosmetically acceptable medium is compatible with the skin, teguments and/or mucous membranes, it does not induce any sensations of discomfort, or more generally disorders able to lead the user to suspend or stop the administration of the cosmetic composition.

More particularly, said cosmetically acceptable medium can comprise water and/or one or more water-miscible organic solvents. Said solvent may be chosen from polyols and alcohols, such as glycerin, sorbitol, glycols such as butylene glycol, propylene glycol, isoprene glycol, dipropylene glycol, hexylene glycol, polypropylene glycol, ethanol or propanediol.

Preferably, the composition according to the invention has a water content ranging from 20%to 95%by weight, more preferably from 30%to 70%by weight in relation to the total weight of the composition.

A composition according to the invention may comprise an oily phase.

The compositions according to the invention can have all the dosage forms conventionally used for a topical application and in particular in the form of aqueous solutions, hydroalcoholic solutions, oil-in-water (O/W) or water-in-oil (W/O) or multiple (triple: W/O/W or O/W/O) emulsions, aqueous gels, or dispersions of an oily phase in an aqueous phase using spheres, these spheres potentially being lipid vesicles of the ionic and/or non-ionic type (liposomes, niosomes, oleosomes) . These compositions are prepared using routine methods.

The cosmetic compositions of the instant disclosure are preferably stable. The term “stable” as used herein means that the cosmetic composition does not visually phase separate or crystallize and does not completely degrade.

In a preferred embodiment, compositions used in the framework of the invention are intended for topical administration. In a preferred embodiment, compositions used in the framework of the invention are intended for application on the skin.

In the framework of the invention, the term "skin" designates any cutaneous surface of the body, preferably the skin of the face or the neck or the legs, and more particularly the skin of the face or the neck.

Advantageously, the compositions according to the invention have the form of a gel, or an emulsion, powder or paste. Furthermore, the composition according to the invention can be more or less fluid and have the appearance of a white or colored cream, an ointment, a milk, a lotion, a serum, a paste, a foaming gel, a peeling, a mask, a care, a tonic or a foam.

The compositions according to the invention can be applied directly on the skin or, alternatively, on cosmetic supports of the occlusive or non-occlusive type, intended to be applied locally on the skin. As non-limiting examples of cosmetic supports, mention can be made of a patch, a wipe, a mask on a support. The composition can be rinsed or not after having been applied to the skin, preferably it is not rinsed.

Uses

The present invention also relates to the use of at least one compound of formula (I) as described herein or an optical isomer thereof, or a cis–trans isomer thereof (such as an alpha or beta anomer) , or a tautomer thereof or a salt thereof, or a hydrate thereof, to promote and/or speed up skin regeneration.

Preferably, the compound of formula (I) or an optical isomer thereof, or a cis–trans isomer thereof (such as an alpha or beta anomer) , or a tautomer thereof or a salt thereof, or a hydrate thereof, is chosen from robinin and its salts, preferably it is chosen from robinin and its salts in β-form.

The present invention also relates to the use of a composition according to the invention, to promote and/or speed up skin regeneration.

Skin regeneration is the innate capacity of the living organisms to repair their skin, by a healing process. In the context of the invention, skin regeneration can follows skin injury, skin that has compromised barrier function, for example, during or after cosmetic surgery procedures.

Here, the term "promote" refers to increasing the effects of the phenomenon. For example, promoting skin regeneration can lead to greater skin generation than without the use of according to the invention.

Here, the term "speed up" refers to the acceleration of the phenomenon. For example, speeding up skin regeneration can lead to same amount of regenerated skin but in less time than without the use of according to the invention.

The present invention relates to use of at least one compound of formula (I) as described herein or an optical isomer thereof, or a cis–trans isomer thereof (such as an alpha or beta anomer) , or a tautomer thereof, or a salt thereof, or a hydrate thereof, according to the invention, to prevent and/or treat signs of aging.

The present invention relates to the use of a composition according to the invention, to prevent and/or treat signs of aging.

Here, the term "prevent" or "prevention" designates any action that aims to prevent the appearance of a discomfort or uneasiness of an individual. This terms therefore covers the stoppage or limitation of the symptoms, but is limited to the cosmetic aspects.

Here, the term "treat" or "treatment" designates any action that aims to improve the comfort or the well-being of an individual. This terms therefore covers the attenuation, slowing down, easing or suppression of the symptoms, but is limited to a cosmetic treatment.

The term "signs of aging” means here that all the modifications that take place in a subject with age. Preferably, the signs of aging according to the invention are signs of aging of the skin.

The term "signs of aging of the skin" or “signs of skin aging” means here that all the modifications of the skin that take place with age and which are sometimes not visible in the early stages. Signs of aging of the skin include marked microrelief of the skin, fine lines, appearance of wrinkles, loss of tonicity, loss of density loss of firmness, loss of elasticity, decrease in the thickness of the dermis and/or epidermis, dryness, withered skin, loss of the radiance of the skin complexion, appearance of a wizened appearance of the skin, sagging of the skin, withering of the skin and loss of the capacity of the skin to regenerate.

The present invention relates to the use at least one compound of formula (I) as described herein or an optical isomer thereof, or a cis–trans isomer thereof (such as an alpha or beta anomer) , or a tautomer thereof or a salt thereof, or a hydrate thereof, to promote and/or speed up wound healing.

The present invention relates to the use of a composition according to the invention, to promote and/or speed up wound healing and in particular, skin wound healing.

Wound healing is the innate capacity of the living organisms to recover from wound, by a healing process. In the context of the invention, skin regeneration can follow skin injury, or skin wound.

Preferably, uses according to the invention are topical uses. Preferably, the composition is applied on the skin, more preferably on the face, the legs or the body, more preferably on the face.

Methods

The invention relates to a non-therapeutic, preferably cosmetic, method of treatment of the skin comprising the step of applying the composition according to the invention on the skin.

Preferably, the step of applying the composition is done by the topical application of said composition, more preferably on the face, the legs or the body, more preferably on the face or the neck.

Preferably, the skin displays at least one sign of skin aging as defined here above.

Preferably, the application is carried out on the skin of a subject that needs it and in an effective quantity of the composition. Here, the term "subject" means a human being.

Preferentially, the subject does not suffer from dermatologic lesion and/or from dermatologic disease, more preferentially the subject does not suffer from any disease.

Here, the term "effective quantity" refers to the quantity of composition according to the invention, that, as a whole, makes it possible to:

- promote and/or speed up skin regeneration, and/or

- prevent and/or treat signs of aging, and/or

- promote and/or speed up wound healing.

According to the present invention, the methods are non-therapeutic methods.

The methods of the invention can comprise a single administration. In another embodiment, the administration is repeated for example 2 to 3 times per day for one day or more and generally during an extended period of at least 4 weeks or 4 to 15 weeks, or as long as necessary.

The invention also relates to a non-therapeutic, preferably cosmetic method of treating skin to promote and/or speed up wound healing after aesthetic procedures, the method comprising:

(a) treating the skin with at least one skin modification stimulus, and

(b) after the step (a) applying at least one compound of formula (I) , or an optical isomer thereof, or a cis–trans isomer thereof (such as an alpha or beta anomer) , or a tautomer thereof or a salt thereof, or a hydrate thereof according to the invention to the skin.

By “skin modification stimulus” it is meant an action on skin that will induce a modification of the skin, such stimulus may temporarily compromise the barrier function of the skin. For example skin modification comprises a peeling agent, a laser, radiofrequency, an electrical, heat, a low-level light, a needle, or an abrasive instrument.

(a) treating the skin with at least one skin modification stimulus, and

(b) after the step (a) applying a composition according to the invention to the skin.

In still further embodiments, the non-therapeutic, preferably cosmetic method according to the invention comprises:

(a) treating skin with at least one type of skin modification stimulus, for example a peeling agent, a laser, radiofrequency, an electrical, heat, a low-level light, a needle, or an abrasive instrument; and

(b) after the step (a) applying at least one compound of formula (I) , or an optical isomer thereof, or a cis–trans isomer thereof (such as an alpha or beta anomer) , or a tautomer thereof or a salt thereof, or a hydrate thereof, according to the invention to the skin, in particular within about 6 hours after step (a) .

In still further embodiments, the method according to the invention comprise:

(b) after the step (a) applying a composition according to the invention to the skin, in particular within about 6 hours after step (a) .

Figures:

Figure 1: Figure 1. Both Keratinocyte Growth Factor (KGF) and Robinin promote proliferation in (A) primary human keratinocytes as determined by manual cell counts following treatment, and (B) within HaCaT keratinocyte cell line as determined by BrdU assay. **denotes p, 0.01, ****p<0.0001, 1-way ANOVA, Tukey post-hoc comparison. n = 3-8.

Figure 2: Robinin increases primary keratinocyte proliferation as determined by total cell counts, while Kaempferol reduces cell numbers. ***denotes p, 0.001, ****p<0.0001, 1-way ANOVA, Tukey post-hoc comparison. n = 3.

Figure 3: Robinin promotes keratinocyte survival after DUVR as evaluated with MTT assay. Viability is normalized to untreated keratinocytes which were not exposed to DUVR. N=10 well/group. One-way ANOVA, Tukey post-hoc comparison. *p<0.05, ****p<0.0001. n=8.

Example:

Materials & Methods

Cell proliferation assays

Cell culture and treatment

Primary adult human keratinocytes cells were obtained from biopsies. HaCaT keratinocytes were obtained from ATCC (Manassas, Virginia, USA) . For primary keratinocyte proliferation assays, cells were plated on a 6 well plate at 1 × 10³ cells/cm² and grown overnight in 1 X Keratinocyte serum-free medium at 37℃, 5%CO₂. The cells were then treated at days 1 and 3 post-seeding, with either vehicle (1: 1000 DMSO) , Keratinocyte Growth Factor (KGF) , Robinin, or Kaempferol. At day 5 the cells were trypsinized, washed, and manually counted manually using a Trypan Blue exclusion assay. HaCaT cells used in proliferation assays were seeded at 8 × 10³ cells/well in a 96 well plate and starved overnight in serum-free DMEM. The cells were then treated as indicated in for 48 hours, after which they were treated using commercial BrdU kit as per the manufacturer’s instructions. Data represents mean ± standard deviation of 6 technical replicates per condition.

Cell survival assay

Primary human keratinocytes were plated at 3 × 10⁴ cells/well in a 96 well plate for 24 hours, after which the cells were washed 3 times with PBS, with 100μl of PBS added to the well after the final wash. The cells were then exposed to daily UV radiation (DUVR) at 10mJ/cm². Following exposure the PBS was replaced with serum-free Keratinocyte Serum-Free Growth Medium (KSFM) with the indicated treatments for 24 hours, after which cell viability was determined using MTT Cell Growth Assay kit.

Results

Promotion of keratinocyte proliferation is a key mechanism through which many anti-aging and skin regenerative molecules improve clinical signs of skin aging, including growth factors such as KGF. To examine the ability of Robinin to drive proliferation within skin cells, primary normal human keratinocytes (NEHK) or HaCaT keratinocytes were treated with either KGF or Robinin as indicated. Treatment with KGF resulted in increased cell numbers consistent with previous reports as did treatment with Robinin (50μM i.e. 0,037%) (Figure 1A) . Both KGF and Robinin also promoted cell proliferation with HaCaT keratinocytes (Figure 1B) . These data collectively demonstrate Robinin promotes this key skin cell regenerative endpoint consistent with KGF.

Kaempferol, has been reported to possess anti-oxidative and photoprotective properties consistent with a skin anti-aging effect but has considerable issues around in vitro toxicity and poor water solubility. To compare the activity of Robinin and Kaemperfol, a proliferation assay was conducted within primary keratinocytes using the same concentrations of both molecules. While Robinin increased cell numbers as anticipated, cells treated with Kaempferol demonstrated dramatically reduced cell numbers (Figure 2) , a likely result of Kaempferol’s toxicity in keratinocytes. These data demonstrate the biological difference between Kaemperfol and Robinin.

One of the primary biological properties of KGF is that it can promote cell survival. Inventors therefore examined the ability of Robinin to promote survival within primary keratinocytes exposed to toxic levels of UV light. Exposure of keratinocytes to daily ultraviolet radiation (DUVR) at 10J/cm² resulted in significant cell mortality, which was ameliorated by KGF treatment (Figure 3) . Keratinocytes treated with 5μM (0,0037%) of robinin for 24 hours post-DUVR did not alter cell survival. However, treatment with 50μM (0, 037%) increased survival to similar level as KGF. This data demonstrates that Robinin acts on keratinocytes in a similar manner to KGF.

Conclusion

These data demonstrate that Robinin promotes keratinocyte proliferation in both primary normal human keratinocytes and HaCaT cells, consistent with the pro-proliferative effects of key skin regenerative growth factor KGF and is functionally different from Kaempferol. Robinin similarly promotes cellular survival following high dose UV exposure, consistent with the cellular protective effects imparted in the skin and skin cells by KGF.

Claims

A cosmetic composition, comprising in a cosmetically acceptable medium:

- at least one compound of formula (I) or an optical isomer thereof, or a cis–trans isomer thereof, or a tautomer thereof or a salt thereof, or a hydrate thereof,

wherein:

- R' and R” are identical or different, preferably different, and represent a hydrogen, or a (C1-C4) alkyl such as a methyl, an ethyl, a propyl, an isopropyl, a butyl, and an isobutyl, preferably a methyl, or a monosaccharide or a polysaccharide comprising up to 20 sugar units, preferably up to 6 sugar units, in pyranose and/or furanose form and of L and/or D series, the said mono-or polysaccharide may be substituted by an obligatorily free hydroxyl group, and/or optionally one or more amine function (s) , optionally protected amine function (s) , and the said monosaccharide or polysaccharide optionally being substituted on its -CH₂OH group by a -C (O) -CO₂H group and the said monosaccharide or polysaccharide optionally being substituted on one or more of its hydroxy groups by a (C1-C4) acyl group, preferably an acetyl group, or by a hydroxycinnamyl moiety, chosen among coumaroyl, caffeoyl, feruloyl, or sinapoyl;

- R”’ and R”” are identical or different, and represent a hydrogen or a (C1-C4) alkyl, such as a methyl, an ethyl, a propyl, an isopropyl, a butyl, and an isobutyl, preferably a methyl,

- being understood that R’, R”, R”’, and R”” cannot represent a hydrogen at the same time;

and;

- at least one compound chosen among thickeners, preservatives, perfumes, bactericides, pigments, dyes, carbon and/or silicone oils of inorganic, waxes, fillers, emulsifiers, co-emulsifiers, UVA and/or UVB light protection agents also called UV filters, polymers, hydrophilic and lipophilic gelling agents.
The composition according to claim 1, wherein said compound is robinin or one of its salts.
The composition according to claim 2, wherein said robinin or one of its salts is in the β-form.
The composition according to any one of claims 1-3 further comprising one or more skin active agents chosen among anti-aging agents and wound healing agents.
The composition according to any one of claims 1 to 4, characterized in that concentration of said least one compound of formula (I) is between 0.001%and 5%by weight relative to the total weight of the composition.
Use of at least one compound of formula (I) as described in claim 1, or an optical isomer thereof, or a cis–trans isomer thereof, or a tautomer thereof or a salt thereof, or a hydrate thereof, or a composition according to any one of claims 1 to 5, to promote and/or speed up skin regeneration.
The use of at least one compound of formula (I) as described in claim 1, or an optical isomer thereof, or a cis–trans isomer thereof, or a tautomer thereof or a salt thereof, or a hydrate thereof, or a composition according to any one of claims 1 to 5, to prevent and/or treat signs of aging.
The use according to claim 7, wherein sign of aging are signs of skin aging.
The use of at least one compound of formula (I) as described in claim 1, or an optical isomer thereof, or a cis–trans isomer thereof, or a tautomer thereof or a salt thereof, or a hydrate thereof, or a composition according to any one of claims 1 to 5, to promote and/or speed up wound healing.
A non-therapeutic method of treatment of the skin comprising the step of applying the composition according to any one of claims 1 to 5 on the skin.
The method according to claim 10, wherein the skin displays at least one sign of skin aging.
A non-therapeutic method of treating skin to promote and/or speed up wound healing after aesthetic procedures, the method comprising:

(a) treating the skin with at least one skin modification stimulus, and

(b) after the step (a) applying a composition according to any one of claims 1-5 to the skin.