WO2018024902A1

WO2018024902A1 - Uses of hydrophobically-modified hyaluronic acid through amide and/or hydrazide linkages in cosmetics and/or dermatology

Info

Publication number: WO2018024902A1
Application number: PCT/EP2017/069844
Authority: WO
Inventors: Manuel Silva
Original assignee: Biokawthar Technologies
Current assignee: Biokawthar Technologies
Priority date: 2016-08-04
Filing date: 2017-08-04
Publication date: 2018-02-08
Anticipated expiration: 2019-02-04

Abstract

The present invention relates to the field of cosmetics and pharmaceuticals such as dermatology. Especially, the present invention relates to particles made of hyaluronic acid modified by amidation with hydrophobic groups having an amine or a hydrazine function having general formula (I), wherein Z, R and n are as defined in the claims. In particular, the particles of the invention are useful for restoring the endogenic production of hyaluronic acid and provide a lasting effect thanks to their slow degradation in vivo. The present invention also refers to a cosmetic care method comprising applying the particles of the invention on the skin of a subject in need thereof.

Description

USES OF HYDROPHOBICALLY-MODI 1F 3)I IE MD » HYALURONIC ACID THROUGH AMIDE AND/OR HYDRAZIDE LINKAGES IN COSMETICS

AND/OR DERMATOLOGY

FIELD OF INVENTION

The present invention relates to the field of cosmetics and pharmaceuticals such as dermatology. Especially, the present invention relates to particles of hyaluronic acid hydrophobically modified by amidation with hydrophobic groups having an amine or a hydrazine function; and their use as a stimulating agent in the epidermis and/or in the dermis. In particular, the particles of the invention are useful for restoring the endogenic production of hyaluronic acid and providing a lasting effect thanks to their slow degradation in vivo. The present invention also refers to a skin improvement method comprising applying the particles of the invention.

BACKGROUND OF INVENTION

Both the largest and the heaviest organ in the human body, the skin plays many fundamental biologic functions such as thermal regulation, barrier against the external environment or hormonal synthesis.

In addition, the skin also ensures a psychosocial role that impacts the social interactions between individuals.

Its structure is organized in layers comprising the epidermis and the dermis.

The epidermis refers to the superficial layer of the skin made of keratinocyte cells, melanocytes cells or Langerhans cells. Its role is mainly directed to protection (immunity function). Located under the epidermis, the dermis is the resistant part of the skin and is made of water, glycosaminoglycans (such as hyaluronic acid) and fibers of proteins (such as, elastin and collagen). These proteins, and glycosaminoglycans (and particularly endogenic hyaluronic acid), allow ensuring good physical properties to the skin (i.e. toughness, elasticity and smoothness).

Over time, the physical properties of the skin decrease due to the lack of glycosaminoglycans, proteins and cells renewal. Consequently, the aging effects appear. For overcoming these drawbacks, many cosmetic and/or dermo-cosmetic products comprising hyaluronic acid chains have been developed and marketed.

However, these products, (1) either contain hyaluronic acid having high molar weight mat cannot diffuse through the deepest layer of the skin; or (2) contain shorter hyaluronic acid that induces inflammatory phenomenon after its application on the skin. Second, these products only aim to be used as filling agents and/or as hydrating agents, and they do not focuse on the provision of an efficient way to reactivate the production of endogenic hyaluronic acid.

Furthermore, after the in vivo administration of the product in a subject in need thereof, the hyaluronic acid chains are easily degraded by the human organism over time. This degradation implies a regular administration of the product by the subject that may be uncomfortable.

Thus, there still exists a need for providing efficient cosmetic, dermo-cosmetic and/or dermatological compounds for avoiding the drawbacks as described above.

Surprisingly, the Applicant has evidenced that using hyaluronic acid modified by hydrophobic groups through an amide and/or a hydrazide bond allows:

- diffusing both in the epidermis and the dermis;

- reactivating the endogenous hyaluronic acid production; and

- reducing in the admiiiistration frequency of the product by the subject in need thereof. Preferably, the hyaluronic acid modified by hydrophobic groups through an amide and/or a hydrazide bond, is under the form of particles. Advantageously, the hydrophobically modified hyaluronic acid particles of the invention are able to efficiently diffuse in the epidermis and the dermis; especially, in the papillary dermis, in the upper reticular dermis and in the lower reticular dermis. Furthermore, the particles of the invention do not provoke any inflammatory side-effects. Advantageously, the particles of the invention are able to increase the expression of collagen HI and/or elastin in the skin; preferably, in the epidermis and/or the dermis; more preferably, in the papillary dermis, in the upper reticular dermis and in the lower reticular dermis.

Advantageously, the amide and the hydrazide linkages are more resistant to degradation by hydrolysis of hyaluronidase enzymes in the skin compared to esters linkages. Consequently, the particles of the invention allow a lasting action, even after a sole application, contrary to the hyotaphobic-modified HA having ester linkage, hyaluronic acid or any salt thereof such as sodium hyahironate.

SUMMARY

The present invention thus concerns the use as a long lasting antiaging agent of a particle comprising at least one hyaluronic acid derivative of general formula (I):

or any salts and solvates thereof,

wherein

Z represents a single bond or -NH-; R represents a C1-C20 alkyl group; preferably, a C6-C18 alkyl group;

preferably, a C18 alkyl group; n represents the number of disaccnaride units of D- glucuronic acid and iV-acetyl-D- glucosamine and is a positive integer; and wherein the particle has a mean hydrodynamic diameter ranging from 200 nm to 500 nm; preferably, from 250 nm to 400 nm; more preferably, from 320 nm to

360 nm

According to one embodiment, the degree of substitution (DS) of the hyaluronic acid derivatives of formula (I) ranges from 0.1 % to 55 %. According to one embodiment, the molecular weight of the hyaluronic acid derivatives of formula (I) ranges from 100 kDa to 4 000 kDa; preferably, from 200 kDa to 3 000 kDa.

According to one embodiment, the long lasting antiaging agent is a long lasting skin plumping agent, a long lasting skin hydrating agent and/or a long lasting stimulating agent of the endogenic production of hyaluronic acid. The present invention also concerns a particle comprising at least one hyaluronic acid derivative of general formula (I):

or any salts and solvates thereof, wherein

Z represents -NH-; R represents a C1-C20 alkyl group; preferably, a C6-C18 alkyl group; more preferably, a C18 alkyl group; n represents the number of disaccharide units of D- glucuronic acid and iV-acetyl-D- glucosamine and is a positive integer; and wherein the particle has a mean hydrodynamic diameter ranging from 200 nm to

500 nm; preferably, from 250 nm to 400 nm; more preferably, from 320 nm to 360 nm

The present invention also concerns a particle as defined above, for use as a medicament

According to one embodiment, the particle as defined above is for use in the treatment and/or the prevention of dermatological conditions; preferably as a healing agent

The present invention also concerns a cosmetic and/or denno-cosmetic composition comprising at least one particle as defined above, and a cosmetically acceptable base.

According to one embodiment the cosmetic and/or dermo-cosmetic composition of the invention is under an injectable form, The present invention also concerns a skin improvement method comprising using the cosmetic and/or dermo-cosmetic composition of the invention in a subject in need thereof.

According to one embodiment the skin improvement method of the invention is for restoring the production of endogenic hyaluronic acid in the epidermis and/or the dermis.

According to one embodiment the skin improvement method of the invention is for limiting the shortcomings of the skin; preferably, for improving the hydration of the skin.

According to one embodiment the composition of the invention is administered to the skin of a subject in need thereof, once per week.

According to one embodiment the composition of the invention is administered to the epidermis and/or the dermis of a subject in need thereof, by a topical application or by injection. DEFINITIONS

In the present invention, the following terms have the following meanings:

"amidatkra reaction": relates to the reaction between the carboxylic function of the hyaluronic acid with (1) either the amine function of a hydrophobic compound or (2) the hydrazine function of a hydrophobic compound. In the present invention, the resulting linkage may be either an amide linkage or a hydrazide linkage;

- "antiaging agent": relates to a compound mat limits the effects of the age on the skin; preferably, it relates to skin plumping agent and/or skin hydrating agent;

- "alkyl": relates to any saturated linear or branched hydrocarbon chain, with 1 to 20 carbon atoms, preferably 6 to 18 carbon atoms, preferably hexyl, heptyl, octyl, nonyl, decyl, undecyl, dodecyl, tridecyl, tetradecyl, pentadecyl, hexadecyl, heptadecyl, octadecyl; more preferably, octadecyl;

- "alkenyl": relates to any unsaturated linear or branched hydrocarbon chain, with 1 to 20 carbon atoms, preferably 6 to 18 carbon atoms, preferably hexenyl, heptenyl, octenyl, nonenyl, decenyl, undecenyl, dodecenyl, tridecenyl, tetradecenyl, pentadecyl, hexadecyl, heptadecyl, octadecenyl; more preferably, tetradecenyl, octadecenyl;

- "cydoalkyT: relates to a substituted or not substituted cyclic alkyl substituent such as cyclopropyl, cyclopentyl, or cyclohexyl;

"cosmetically acceptable base": relates to any compound for use in contact with the epidermis and/or the dermis, and which does not provoke any side effects such as toxicity, irritation, inflammation or allergic response. In the present invention, a cosmetically acceptable base includes base for topical administration and/or injection;

- "cosmetic use" refers to the use in order to improve the esthetic appearance of the body;

"degree of substitution (DS)": relates to me number of me carboxylic functions of the hyaluronic acid that have been modified after amidation with at least one hydrophobic group having an amine or a hydrazine function; - "dermis": relates to the conjunctive tissue forming with the epidermis and the hypodermis, the skin. The dermis is constituted of the papillary dermis and the reticular dermis;

"endogenic hyaluronic add": relates to hyaluronic acid synthetized by the human body;

- "epidermis": relates to the superficial layer of the skin made of keratinized dead cells;

- "healing agent": relates to a compound able to favor the decrease of wounds; preferably, of superficial wounds; more preferably, of the superficial wounds of the skin or the mucous;

- "long-lasting agent": relates to a compound able to provide its effects for a long time.

For example, a 'long-lasting antiaging agent" refers to a compound having antiaging properties, said properties being maintained or improved for a long time compared to the effects of a non-long-lasting antiaging agent;

- "particle": relates to an assembly of the hyaluronic acid derivatives of the invention organized by weak bonds, i.e. an assembly of hyaluronic acid modified by amidation with at least one hydrophobic group having an amine or a hydrazine function. In the present invention, the term "particle" comprises nanoparticles and/or microparticles obtained from hydrophobically modified hyaluronic acid of the invention. According to one embodiment, the term "particles" does not comprise polymersomes or liposomes. According to one embodiment, the term "particles" does not comprise aggregates;

''pharmaceutically acceptable base": relates to any ingredients of a pharmaceutical composition that are compatible with each other and not deleterious to the subject to which it is adrninistered;

- "hydrophobic": relates to a compound or a chemical group that has no affinity for water or aqueous medium;

"skin plumping agent": relates to a compound able to provide a measurable increase of the skin density; especially, of the epidermis and/or the dermis. The "skin plumping effect" may be analyzed by echography or by the implementation of techniques suitable of the human body area for which the effect is analyzed;

"skin hydrating agent": relates to a compound able to overcome the dry skin; especially, to overcome the dry of the epidermis and/or the dermis and/or the mucous.

DETAILED DESCRIPTION

This invention relates to a hyaluronic acid derivative; especially, a hyaluronic acid modified by amidation with at least one hydrophobic group through an amine or a hydrazine function, of general formula (I):

or any salts and solvates thereof,

wherein

Z represents a single bond or -NH-;

R represents a C1-C20 alkyl or C1-C20 alkenyl group; preferably, a C6-C18 alkyl group; more preferably, a C18 alkyl group; and

n represents the number of disaccharide units of D- glucuronic acid and iV-acetyl-D- glucosamine and is a positive integer.

According to one embodiment, the hyaluronic acid salt is a hyaluronate, preferably sodium hyaluronate. According to one embodiment, hydrophobic groups are grafted on sodium hyaluronate. According to one embodiment, R represents a linear CI -20 alkyl group or a linear Cl- C20 alkenyl group, preferably a linear C6-C18 alkyl group, more preferably a linear C18 alkyl group. According to one embodiment, R represents a C1-C20 alkyl group. According to one embodiment, R represents a C6, C7, C8, C9, CIO, CI 1, C12, C13, C14, C15, C16, C17 or C18 alkyl group, preferably a linear C6, C7, C8, C9, CIO, CI 1, C12, C13, C14, C15, C16, C17 or C18 alkyl group.

According to one embodiment, preferred compounds are of formula (la):

or any salts and solvates thereof,

wherein

n represents the number of disaccharide units of D- glucuronic acid and iV-acetyl-D- glucosamine and is a positive integer. According to another embodiment, preferred compounds are of formula (lb):

or any salts and solvates thereof, wherein

Especially, the present invention relates to a hyaluronic acid modified by amidation with at least one hydrophobic group through an amine or a hydrazine function, of general formula (Π):

or any salts and solvates thereof,

wherein

Z represents a single bond or -NH-; and

n represents the number of disaccharide units of D- glucuronic acid and iV-acetyl-D- glucosamine and is a positive integer. According to one embodiment, when Z is a single bond, R is not a C1-, CIS-, C16- or C20-alkyl group. According to one embodiment, when Z is a single bond, R is not a C8- , C12- or C16-alkyl group. According to one embodiment, R does not represent a C8- alkyl group. According to one embodiment, R does not represent a C12-alkyl group. According to one embodiment, R does not represent a C16-alkyl group. According to one embodiment, hyaluronic acid is not partially deacetylated. According to one embodiment, hyaluronic acid is not deacetylated. According to one embodiment, hyaluronic acid derivatives of formula (I) or (II) nave a molecular weight ranging from 3 kDa to 4000 kDa; preferably from 200 kDa to 3 000 kDa. According to one embodiment, hyaluronic acid derivatives of formula (I) or (II) have a molecular weight ranging from 100 kDa to 4000 kDa; preferably from 200 kDa to 3 000 kDa. According to one embodiment, hyaluronic acid derivatives of formula (I) or (II) have a molecular weight of about 200 kDa or of about 3000 kDa. According to one embodiment, hyaluronic acid derivatives of formula (I) or (II) have a molecular weight of about 100, 200, 300, 400, 500, 600, 700, 800, 900, 1000, 1100, 1200, 1300, 1400, 1500, 1600, 1700, 1800, 1900, 2000, 2100, 2200, 2300, 2400, 2500, 2600, 2700, 2800, 2900, 3000, 3100, 3200, 3300, 3400, 3500, 3600, 3700, 3800, 3900 or 4000 kDa.

According to one embodiment, the degree of substitution (DS) of the hyaluronic acid by the hydrophobic groups ranges from 0.01 % to 55 %; preferably, from 1 % to 15%; more preferably, from 1 to 5 %; more preferably, from 10 to 15 %. According to one embodiment, the degree of substitution (DS) of the hyaluronic acid by the hydrophobic groups ranges from 0.1 % to 55 %; preferably, from 1 % to 15 %; more preferably, from 1 to 5 %; more preferably, from 10 to 15 %. According to one embodiment, the degree of substitution (DS) is about 1.5 %. According to one embodiment, the degree of substitution (DS) is about 12 %. The degree of substitution (DS) of the hyaluronic acid by the hydrophobic groups was detemiined by NMR (Nuclear Magnetic Resonance) analysis by comparing the area of cyclic protons of the hyaluronic acid or the sodium hyaluronate (10 protons between 3.5 and 54.3 ppm), with the terminal CH3 of the aliphatic graft (3 protons at around 1.57 ppm for 100 % of substitution). The calculation may be also done by comparing the areas of the terminal CH3 of the aliphatic graft (3 protons around 1.57 ppm for 100 % substitution) and the CH3 of N-acetyl glucosamine (3 protons around 1.47 ppm). According to one embodiment, the degree of substitution (DS) of the hyaluronic acid by the hydrophobic groups ranges from 0.1 % to 5 %; preferably from 0.1 % to 4 % preferably from 0.1 % to 3 % preferably from 0.1 % to 2 % preferably from 0.1 % to 1 %. According to one embodiment, the degree of substitution (DS) of the hyaluronic acid by the hydrophobic groups ranges from 0.1 % to 10 %. According to one embodiment, the degree of substitution (DS) of the hyaluronic acid by the hydrophobic groups is about 1 %, 5 %, 10 %, IS %, 20 %, 25 %, 30 %, 35 %, 40 %, 45 %, 50 % or 55 %. According to one embodiment, the degree of substitution (DS) of the hyaluronic acid by the hydrophobic groups is about 1 %, 2 %, 3 %, 4 %, 5 %, 6 %, 7 %, 8 %, 9 %, 10 %, 11 %, 12 %, 13 %, 14 % or 15 %. According to one embodiment, the hydrophobic groups are grafted to the hyaluronic acid by a covalent linkage; especially, by an amide linkage or a hydrazide linkage. According to one embodiment, the hydrophobic groups are grafted to the hyaluronic acid or any salts and solvates thereof, by a covalent linkage; especially, by an amide linkage or a hydrazide linkage. According to one embodiment, the hyaluronic acid is modified by an amidation between the carboxylic acid function of hyaluronic acid and the amine function of the hydrophobic compounds to be grafted. According to one embodiment, the hyaluronic acid salt which is a hyaluronate, is modified by an amidation between the carboxylate function of said hyahironate and the amine function of the hydrophobic compounds to be grafted. According to one embodiment, the hydrophobic compound is selected from the organic compounds having at least one amine function or having at least one hydrazine function, i.e. a function having the following structure:

wherein

Z represents a single bond or -NH-; and

R represents a C1-C20 alkyl or C1-C20 alkenyl group; preferably, a C6-C18 alkyl group; more preferably, a C18 alkyl group.

According to one embodiment, the hydrophobic compound is not hydrazine molecule, i.e. compound of formula: NH2-NH2. According to one embodiment, the modified hyaluronic acid does not comprise free- hydrazido groups, i.e. - NH-NH2 or -NR'-NIfc pendent groups wherein R' is an organic functional group as for example an alkyl, an aryl, or an alkene. According to one embodiment, me modified hyaluronic acid does not comprise oxalic hydrazido groups.

According to one embodiment, the hyaluronic acid is modified by an alkylamine; preferably, methylamine, ethylamine, propylamine, butylamine, pentylamine, hexylamine, heptylamine, octylamine, nonylamine, decylamine, undecylamine, dodecylamine, tridecylamine, tetradecylamine, pentadecylamine, hexadecylamine, heptadecylamine, octadexylamine; nonadecylamine, eicosylamine; more preferably, hexylamine, heptylamine, octylamine, nonylamine, decylamine, undecylamine, dodecylamine, tridecylamine, tetradecylamine, pentadecylamine, hexadecylamine, heptadecylamine, octadecylamine. According to one embodiment, the hyaluronic acid is not modified by octylamine. According to one embodiment, the hyaluronic acid is not modified by dodecylamine. According to one embodiment, the hyaluronic acid is not modified by hexadecylamine.

According to one embodiment, the hyaluronic acid is modified by alkenylamine; preferably, ethylenylamine, propylenylamine, butylenylamine, pentenylamine, hexenylamine, heptenylamine, octenylamine, nonenylamine, decenylamine, undecenylamine, dodecenylamine, tridecenylamine, tetradecenylamine, pentadecenylamine, hexadecenylamine, heptadecenylamine, octadecenylamine; nonadecenylamine, eicosenylamine; more preferably, hexenylamine, heptenylamine, octenylamine, nonenylamine, decenylamine, undecenylamine, dodecenylamine, tridecenylamine, tetradecenylamine, pentadecenylamine, hexadecenylamine, heptadecenylamine, octadecenylamine.

According to one embodiment, the hyaluronic acid is modified by octadecylamine. According to one embodiment, the amine is not octylamine, hexadecylamine or octadecylamine. According to one embodiment, the amine compound does not comprise more than one amine function. According to one embodiment, the amine compound does not comprise emylarnine. According to one embodiment, the amine compound does not comprise any amino- acids or derivatives. According to one embodiment, the amine compound does not comprise any amino- acid ester derivatives. According to one embodiment, the amine compound does not comprise a hydroxyl group. According to one embodiment, me amine compound does comprise other chemical functional groups than amino group.

According to one embodiment, the hyaluronic acid is modified by an alkylhydrazine; preferably, methylhydrazine, ethylhydrazine, propylhydrazine, butylhydrazine, pentylhydrazine, hexylhydrazine, heptylhydrazine, octylhydrazine, nonylhydrazine, decylhydrazine, undecylhydrazine, dodecylhydrazine, tridecylhydrazine, tetradecylhydrazine, pentadecylhydrazine, hexadecylhydrazine, heptadecylhydrazine, octadexylhydrazine; nanodecylhydrazine, eicosylhydrazine; more preferably, hexylhydrazine, heptylhydrazine, octylhydrazine, nonylhydrazine, decylhydrazine, undecylhydrazine, dodecylhydrazine, tridecylhydrazine, tetradecylhydrazine, pentadecylhydrazine, hexadecylhydrazine, heptadecylhydrazine, octadecylhydrazine. According to one embodiment, the hyaluronic acid is modified by octadecylhydrazine. According to one embodiment, the hyaluronic acid is modified by octadecylhydrazine (also called stearylhydrazine).

According to one embodiment, the hyaluronic acid is modified by an alkenylhydrazine; preferably, ethylenylhydrazine, propylenylhydrazine, butylenylhydrazine, pentylenylhydrazine, hexylenylhydrazine, heptylenylhydrazine, octylenylhydrazine, nonylenylhydrazine, decylenylhydrazine, undecylenylhydrazine, dodecylenylhydrazine, tridecylenylhydrazine, tetradecylenylhydrazine, pentadecylenylhydrazine, hexadecylenylhydrazine, heptadecylenylhydrazine, octadecylenylhydrazine; nonadecylenylhydrazine, eicosylenylhydrazine; more preferably, hexylenylhydrazine, heptylenylhydrazine, octylenylhydrazine, nonylenylhydrazine, decylenylhydrazine, undecylenylhydrazine, dodecylenylhydrazine, tridecylenylhydrazine, tetradecylenylhydrazine, pentadecylenylhydrazine, hexadecylenylhydrazine, heptadecylenylhydrazine, octadecylenylhydrazine.

According to one embodiment, the hydrazine compound does not comprise more than one hydrazine function. According to one embodiment, the hydrazine compound is a linear hydrophobic compound having one hydrazine function. According to one embodiment, the hydrophobically-modified HA derivative is a HA of 200 kDa grafted with a C8 alkyl group via a hydrazide linkage. According to one embodiment, the hydrophobically-modified HA derivative is a HA of 200 kDa grafted with a C6 alkyl group via an amide linkage. According to one embodiment, the hydrophobically-modified HA derivative is a HA of 3000 kDa grafted with a C8 alkyl group via a hydrazide linkage. According to one embodiment, the hydrophobically- modified HA derivative is a HA of 3000 kDa grafted with a C6 alkyl group via an amide linkage. According to one embodiment, the hydrophobically-modified HA derivative is a HA of 3000 kDa grafted with a C18 alkyl group via an amide linkage. According to one embodiment, the hydrophobically-modified HA derivative is a HA of 3000 kDa grafted with a C18 alkyl group via a hydrazide linkage. According to one embodiment, the hydrophobically-modified HA derivative is a HA of 200 kDa grafted with a C18 alkyl group via an amide linkage. According to one embodiment, the hydrophobically-modified HA derivative is a HA of 200 kDa grafted with a C18 alkyl group via a hydrazide linkage. According to one embodiment, the hydrophobically-modified HA derivative is a HA of 3000 kDa grafted with a C12 alkyl group via an amide linkage. According to one embodiment, the hydrophobically-modified HA derivative is a HA of 200 kDa grafted with a C12 alkyl group via an amide linkage. According to one embodiment, the hydrophobically-modified HA derivative is a HA of 200 kDa grafted with a C18 alkyl group via an amide linkage. According to one embodiment, the hydrophobically-modified HA derivative is a HA of 200 kDa grafted with a C18 alkyl group via a hydrazide linkage. According to one embodiment, the hydrophobically-modified HA derivative is a HA of 3000 kDa grafted with a C18 alkyl group via a hydrazide linkage. According to one embodiment, the hydrophobically-modified HA derivative is a HA of 3000 kDa grafted with a C18 alkyl group via an amide linkage.

According to one embodiment, the hydrazine compound does not comprise a hydroxy 1 group. According to one embodiment, the hydrazine compound does comprise other chemical functional groups than hydrazine group. According to one embodiment, the hydrophobic compound does not comprise more man one amine or hydrazine function. According to one embodiment, the hydrophobic compound does not comprise aryl compounds.

According to one embodiment, the hyaluronic acid derivative is N- stearylamine-modified hyaluronic acid, i.e. a hyaluronic acid modified by amidation reaction with stearylamine (also called octadecylamine) by amide linkage.

According to one embodiment, the hyaluronic acid derivative is -V-stearyl hydrazine- modified hyaluronic acid, i.e. a hyaluronic acid modified by amidation reaction with stearylhydrazine (also called octadecylhydrazine) by hydrazide linkage. According to one embodiment, the hyaluronic acid is not crosslinked. According to one embodiment, the modified hyaluronic acid is not crosslinked.

The invention also relates to a particle comprising at least one hyaluronic acid derivative of formula (I) and/or (II), and salts and/or solvates thereof.

According to one embodiment, the particle of the invention has a size (i.e. a mean hydrodynamic diameter) ranging from 200 nm to 500 nm; preferably, from 250 nm to 400 nm; more preferably, from 320 nm to 360 nm. The mean hydrodynamic diameter is measured in a mixture of water/phosphate buffer (S0/S0, v/v) by using a nanosizer apparatus. According to one embodiment, the size of the particle of the invention is about 327 nm. According to one embodiment, the size of the particle of the invention is about 357 nm. According to one embodiment, the particle of the invention has a size of about 200 nm, 250 nm, 300 nm, 350 nm, 400 nm, 450 nm or 500 nm. According to one embodiment, the particle of the invention has a size of about 300 nm, 310 nm, 320 nm, 330 nm, 340 nm, 350 nm or 360 nm.

According to one embodiment, the particle of the invention does not comprise cyclodextrin. According to one embodiment, the particle of the invention does not comprise alpha-cyclodextrin. According to one embodiment, the particle of the invention does not comprise beta-cyclodextrin. According to one embodiment, the particle is constituted of hyaluronic acid derivatives of formula (I) and/or (Π), and salts and/or derivatives thereof. According to one embodiment, the particle of the invention does not comprise other kind of particle such as for example but not limited to, inorganic particles, hybrid particles and/or metallic particles. According to one embodiment, the particle of the invention does not comprise other polymer than hyaluronic acid. According to one embodiment, the particle of the invention does not comprise other polymer than the modified hyaluronic acid of the invention.

According to one embodiment, hyaluronic acid derivatives of formula (I) and/or (Π), and salts and/or derivatives thereof are not associated with cyclodextrin. According to one embodiment, hyaluronic acid derivatives of formula (I) and/or (Π), and salts and/or derivatives thereof are not associated with alpha-cyclodextrin. According to one embodiment, hyaluronic acid derivatives of formula (I) and/or (Π), and salts and/or derivatives thereof are not associated with beta-cyclodextrin. In the present invention, the hyaluronic acid derivatives of the invention have been characterized by spectroscopy and/or by Nuclear Magnetic Resonance (NMR). Especially, the hyaluronic acid derivatives of the invention have been characterized by Fourier Transform Infrared (FT-IR) spectroscopy.

The invention also relates to a composition comprising at least one hyaluronic acid derivative of formula (I) and/or (Π), and salts and/or derivatives thereof; and at least one cosmetically acceptable base.

Especially, the invention also relates to a cosmetic and/or dermo-cosmetic composition comprising at least one particle of the invention as defined above, and at least one cosmetically acceptable base. According to one embodiment, the composition of the invention comprises at least one particle having a size (i.e. a mean hydrodynamic diameter) ranging from 200 nm to 500 nm; preferably, from 250 nm to 400 nm; more preferably, from 320 nm to 360 nm. The mean hydrodynamic diameter is measured in a mixture of water/phosphate buffer (S0/S0, v/v) by using a nanosizer apparatus. According to one embodiment, the size of the particle of the invention is about 327 nm. According to one embodiment, the size of the particle of the invention is about 357 nm.

The invention also relates to a pharmaceutical composition, preferably a dermatological composition, comprising at least one hyaluronic acid derivative of formula (I) and/or (Π), and salts and/or derivatives thereof; and at least one pharmaceutically acceptable carrier.

Especially, the invention also relates to a pharmaceutical composition comprising at least one particle of the invention as defined above, and at least one pharmaceutically acceptable carrier.

According to one embodiment, the composition does not comprise cyclodextrin. According to one embodiment, the composition does not comprise alpha-cyclodextrin. According to one embodiment, the composition does not comprise beta-cyclodextrin.

According to one embodiment, the composition comprises the particles of the invention in an amount ranging from 0.01 % to 100 % by weight to the total weight of the composition; preferably from 0.1 to 20 %; more preferably, from 0.5 % to 1 %. According to one embodiment, the composition comprises the particles of the invention in an amount ranging from 1 % to 100 %, preferably from 10 % to 100 %, preferably from 20 % to 100 %, preferably from 30 % to 100 %, preferably from 40 % to 100 %, preferably from SO % to 100 %, by weight to the total weight of the composition. According to one embodiment, the composition comprises the particles of the invention in an amount ranging from 1 % to 100 %, preferably from 1 % to 20 %, preferably from 5 % to 20 %, preferably from 10 % to 20 %, preferably from 15 % to 20 % by weight to the total weight of the composition. According to one embodiment, the composition comprises the particles of the invention in an amount of about 1 %, 2 %, 3 %, 4 %, 5 %, 6 %, 7 %, 8 %, 9 %, 10 %, 11 %, 12 %, 13 %, 14 %, 15 %, 16 %, 17 %, 18 %, 19 % or 20 % by weight to the total weight of the composition.

According to one embodiment, the composition comprises the particles of the invention in an amount of about 0.5 %, 0.6 %, 0.7 %, 0.8 %, 0.9 %, or 1 % by weight to the total weight of the composition. According to one embodiment, the composition is under the form of a solid, a solution, a suspension or an emulsion. According to one embodiment, the composition is selected from oil-in-water emulsion (O/W) and water-in-oil emulsion (W/O), wherein the oil may be sourced from vegetal oil or mineral oil. According to one embodiment, the composition further comprises a solvent; preferably, a biocompatible solvent According to one embodiment, the composition further comprises an aqueous solution or a hydro-alcoholic solution.

According to one embodiment, the composition further comprises at least one cosmetically acceptable base under the form of a gel, a paste, a cream, a lotion, a milk, a stick, a shampoo, a powder, an aerosol, a film or a patch.

According to the present invention, the compositions of the invention may be used in combination of at least one cosmetic agent selected from fatty acids, organic solvents, gelling agents, softening agents, surfactants, detergents, gelling agents, fragrances, emulsifying agents, opacifying agents, stabilizing agents, foaming agents, chelating agents, preservative agents, sunscreens, essential oils, dyes, mineral loads, or any compound used in cosmetics.

According to the present invention, the compositions of the invention may be used in combination of at least one active ingredient

According to one embodiment the compositions of the invention are topically administrated. According to one embodiment the compositions of the invention are administrated by a direct topical mean. According to one embodiment, the compositions of the invention are administrated by an indirect topical mean. In the present invention, "indirect topical mean" refers to the use of a textile support or a material for administering the composition of the invention. According to another embodiment the composition of the invention is injected; preferably, by transdermal injection.

The invention also relates to a process for preparing hyaluronic acid derivatives of formula (I):

or any salts and solvates thereof,

wherein

Z represents a single bond or -NH-;

n represents the number of disaccharide units of D- glucuronic acid and iV-acetyl-D- glucosamine and is a positive integer;

comprising:

(i) providing a solution comprising a carbodiirnide compound, N- hydrosuccinimide (NHS) and a hyaluronic acid, in a buffer;

(ii) adding a solution comprising at least one hydrophobic compound having an amine or hydrazine function.

According to one embodiment, the cart>odiirnide compound relates to any organic compound having a carbodiirnide function (i.e. a function -N=C=N-); preferably, symmetric carbodiirnide. According to one embodiment, a suitable symmetric carbodiirnide compound has the following formula:

R"-N=C=N-R" wherein R" is an alkyl or arylalkyl group; preferably, R" is a cycloalkyl group; more preferably, R" is cyclohexyl group.

According to one embodiment, the caibodiimide compound is ethyl carbodiirnide. According to one embodiment, the process for preparing hyaluronic acid derivatives of formula (I) is carried out at a temperature ranging from 0°C to 40°C; preferably, from 10°C to 30°C; more preferably, at a temperature ranging from 20°C to 2S°C. According to one embodiment, the process for preparing hyaluronic acid derivatives of formula (I) is carried out at room temperature.

According to one embodiment, the process for preparing hyaluronic acid derivatives of formula (I) is carried out at atmospheric pressure.

According to one embodiment, the concentration of the hyaluronic acid, ranges from 10 mg/mL to 70 mg/mL; preferably, from 20 mg/mL to 60 mg/mL; more preferably, ranges from 30 mg/mL to SO mg/mL.

According to one embodiment, the process is carried out for a period ranging more than 0 to 12 hours; preferably, from 1 hour to 6 hours; more preferably, the time of the process is carried out about 5 hours.

The invention also relates to a process for preparing particles of hyaluronic acid derivatives of formula (I):

or any salts and solvates thereof,

wherein

Z represents a single bond or -NH-;

R represents a C1-C20 alkyl or C1-C20 alkenyl group; preferably, a C6-C18 alkyl group; more preferably, a C18 alkyl group; n represents the number of disaccnaride units of D- glucuronic acid and JV-acetyl-D- glucosamine and is a positive integer; and comprising solubilizing hyaluronic acid derivatives or formula (I) in an aqueous dissolution medium at a concentration ranging from 0.25 to 2 % wt Especially, the invention relates to a process for preparing particles of hyaluronic acid derivatives of formula (II):

or any salts and solvates thereof,

wherein

Z represents a single bond or -NH-;

n represents the number of disaccnaride units of D- glucuronic acid and JV-acetyl-D- glucosamine and is a positive integer; and comprising solubilizing hyaluronic acid derivatives of formula (II) in an aqueous dissolution medium at a concentration ranging from 0.25 to 2 % wt According to one embodiment, the particle has a size (i.e. a mean hydrodynamic diameter) ranging from 100 nm to 2000 nm; preferably, from 200 nm to 500 nm; preferably, from 250 nm to 400 nm; more preferably, from 320 nm to 360 nm The mean hydrodynamic diameter is measured in a mixture of water/phosphate buffer (S0/S0, v/v) by using a nanosizer apparatus. According to one embodiment, the size of the particle of the invention is about 327 nm. According to one embodiment, the size of the particle of the invention is about 3S7 nm.

According to one embodiment, the particle has a size (i.e. a mean hydrodynamic diameter) depending on the substitution degree of modified hyaluronic acid. According to one embodiment, the aqueous dissolution medium is selected from MilliQ* water, phosphate buffer or mixture of MilliQ* water and phosphate buffer (SO/SO; v/v).

According to one embodiment, the phosphate buffer comprises monosodium phosphate, dis odium phosphate and sodium chloride. According to one embodiment, the phosphate buffer comprises monosodium phosphate at the concentration of 0.045 g/L. According to one embodiment, the phosphate buffer comprises disodium phosphate at the concentration of 0.S63 g/L. According to one embodiment, the phosphate buffer comprises sodium chloride at the concentration of 8.5 g/L.

According to one embodiment, the aqueous dissolution medium has a pH ranging from 5 to 8; preferably from 5.6 to 6.9. According to one embodiment, the aqueous dissolution medium has a pH ranging from about 4 to about 8; preferably is about 4.S.

According to one embodiment, the concentration in the aqueous dissolution medium of hyaluronic acid derivatives is about 0.25 %, 0.S % or 1 %wt According to one embodiment, the concentration in the aqueous dissolution medium of hyaluronic acid derivatives wherein Z is single bond, is about 1 %wt According to one embodiment, the concentration in the aqueous dissolution medium of hyaluronic acid derivatives wherein Z is -NH-, is about 025 %wt or about 5 %wt.

According to one embodiment, the hyaluronic acid derivatives of formula (I) are completely solubilized in the aqueous dissolution medium.

In the present invention, the hyaluronic acid modified with a hydrophobic compound having (1) at least one amine function or at least one hydrazine function, and (2) a C18- alkyl chain, allows improving the association of hyaluronic acid derivatives in particulate form. The invention also relates to die use of hyaluronic acid derivatives or die particles of the invention, as described above. Especially, the invention relates to the use of hyaluronic acid derivatives of formula (I) or particles thereof, in the field of cosmetics and/or pharmaceuticals . More especially, the invention relates to the use of hyaluronic acid derivatives of formula (I) or particles thereof, as a long-lasting anti-aging agent

According to one embodiment, the hyaluronic acid derivatives or particles thereof are useful in the field of cosmetics and/or dennocosmetics. According to one embodiment, the hyaluronic acid derivatives or particles thereof are useful as cosmetic and/or dermocosmetic agents, preferably as long lasting cosmetic and/or dermocosmetic agent; more preferably as long lasting anti-aging agents. According to one embodiment, the long lasting anti-aging effect is observed more than 3 days, preferably more man 5 days, more preferably more than 7 days after the application of the hyaluronic acid derivatives of the invention or particles thereof on the skin of a subject in need thereof.

According to one embodiment, the hyaluronic acid derivatives or particles thereof increase the surface occupied by glycosaminoglycans (GAGs) in the skin (epidermis and/or dermis). According to one embodiment, the hyaluronic acid derivatives or particles thereof increase the surface occupied by glycosaminoglycans (GAGs) in the skin (epidermis and/or dermis). According to one embodiment, the hyaluronic acid derivatives or particles thereof increase the surface occupied by glycosaminoglycans (GAGs) in the skin (epidermis and/or dermis) in papillary dermis. According to one embodiment, the hyaluronic acid derivatives or particles thereof are useful for increasing the surface occupied by glycosaminoglycans (GAGs) in the skin (epidermis and/or dermis), preferably in a range from 0.1 % to ISO %, preferably 1 % to ISO %, preferably from 10% a ISO %, preferably from 20 % to 150 %, preferably from 30 % to ISO %, preferably from 40 % to 150 %, preferably from 50 % to ISO %, preferably from 60 % to 150 %, preferably from 70 % to 150 %, preferably from 80 % to ISO %, preferably from 90 % to 150 %, preferably from 100 % to 150 % compared to the surface occupied by glycosaminoglycans (GAGs) in an untreated skin.

According to one embodiment, the hyaluronic acid derivatives or particles thereof are useful for increasing the surface occupied by glycosaminoglycans (GAGs) in the skin (epidermis and/or dermis) of a subject of about 3 %, 15 %, 30 % or 119 %, compared to the surface occupied by glycosaminoglycans (GAGs) in an untreated skin.

According to one embodiment, the hyaluronic acid derivatives or particles thereof are useful for increasing the surface occupied by glycosaminoglycans (GAGs) in the skin of a subject (epidermis and/or dermis) of about 1 %, 5 %, 10 %, 15 %, 20 %, 25 %, 30 %, 35 %, 40 %, 45 %, 50 %, 55 %, 60 %, 65 %, 70 %, 75 %, 80 %, 85 %, 90 %, 95 %, 10 %, 105 %, 110%, 115 %, 120 %, 125 %, 130 %, 135 %, 140 %, 145 %, 150 %, compared to the surface occupied by glycosaminoglycans (GAGs) in an untreated skin.

According to one embodiment, the particle has a size (i.e. a mean hydrodynamic diameter) ranging from 100 nm to 2000 nm; preferably, from 200 nm to 500 nm; preferably, from 250 nm to 400 nm; more preferably, from 320 nm to 360 nm. The mean hydrodynamic diameter is measured in a mixture of water/phosphate buffer (50/50, v/v) by using a nanosizer apparatus. According to one embodiment, the size of the particle of the invention is about 327 nm. According to one embodiment, the size of the particle of the invention is about 357 nm.

According to one embodiment, the hyaluronic acid derivatives of the invention and/or the particles thereof are for use as a medicament

According to one embodiment, the present invention relates to the use of the hyaluronic acid derivatives of the invention and/or the particles thereof for the manufacture of a medicament

According to one embodiment, the hyaluronic acid derivatives and/or the particles of the invention are for use in the treatment and/or the prevention of dermatological conditions.

According to one embodiment the hyaluronic acid derivatives and/or the particles of the invention are for use as a healing agent According to one embodiment the hyaluronic acid derivatives and/or the particles of the invention are for use as a long-lasting healing agent

According to one embodiment the present invention relates to a method of treatment of dermatological diseases and/or conditions comprising the administration to a patient in need thereof, of the hyaluronic acid derivatives of the invention and/or the particles thereof.

Especially, the invention concerns the use as a cosmetic agent of a particle comprising at least one hyaluronic acid derivative of general formula (I) as defined above. According to one embodiment, the invention concerns the use as a cosmetic agent of a particle comprising at least one hyaluronic acid derivative of general formula (la), (lb) or (II) as defined above.

According to one embodiment, the hyaluronic acid derivatives or the invention and/or the particles thereof, are useful as cosmetic agents; preferably; as an anti-aging agent; more preferably, as a skin plumping agent, a skin hydrating agent and/or a stimulating agent of the endogenic production of hyaluronic acid. According to one embodiment, the hyaluronic acid derivatives or the invention and/or the particles thereof, are useful as long-lasting cosmetic agents; preferably as a long-lasting anti-aging agent; more preferably, as a long-lasting skin plumping agent, a long-lasting skin hydrating agent and/or a long-lasting stimulating agent of the endogenic production of hyaluronic acid.

According to one embodiment, the hyaluronic acid derivatives or the invention and/or the particles thereof, are useful for stimulating the expression of endogen glycosaminoglycans (GAGs), preferably for stimulating the expression of endogen elastin, collagen and/or hyaluronic acid. According to one embodiment, the invention concerns a method for stimulating the expression of endogen glycosaminoglycans (GAGs), preferably for stimulating the expression of endogen elastin, collagen and/or hyaluronic acid, said method comprising applying at least one hyaluronic acid derivative of the invention and/or the particles thereof, on the skin of a subject in need thereof. According to one embodiment, the expression of glycosaminoglycans (GAGs) is stimulated at least during 2 days, preferably during 5 days, more preferably during 7 days, after the application of hyaluronic acid derivatives of the invention and/or the particles thereof, on the skin of a subject in need thereof. According to one embodiment, the expression of glycosaminoglycans (GAGs) is improved in a range from 0.1 % to 150 %, preferably 1 % to 150 %, preferably from 10 % a 150 %, preferably from 20 % to 150 %, preferably from 30 % to 150 %, preferably from 40 % to 150 %, preferably from 50 % to 150 %, preferably from 60 % to 150 %, preferably from 70 % to 150 %, preferably from 80 % to 150 %, preferably from 90 % to 150 %, preferably from 100 % to 150 %, compared to the expression of glycosaminoglycans (GAGs) in an untreated skin.

According to one embodiment, the expression of glycosaminoglycans (GAGs) in the skin of a subject treated with the hyaluronic acid derivatives or particles of the invention, is improved of about 3 %, 15 %, 30 % or 119 %, compared to the expression of glycosaminoglycans in an untreated skin.

According to one embodiment, the expression of glycosaminoglycans (GAGs) in the skin of a subject treated with the hyaluronic acid derivatives or particles of the invention, is

(GAGs) in an untreated skin.

The invention also relates to a skin improvement method comprising administering the composition as defined above, to the skin of a subject in need thereof. In the present invention, "administering" refers to a topical application or injection means.

According to one embodiment, the skin improvement method comprises applying the composition as defined above, on the skin of a subject in need thereof. According to one embodiment, the skin improvement method comprises applying in a sole application the composition as defined above, on the skin of a subject in need thereof. According to one embodiment, the composition is applied via a device comprising said composition; preferably, said device is a syringe, a patch, a film, a stick, a plaster, a stupe or a bandage. According to one embodiment, the skin improvement method comprises applying the composition as defined above, on the skin of a subject in need thereof, once a week. According to one embodiment, the skin improvement method comprises applying the composition as defined above, on the skin of a subject in need thereof, once a month. According to one embodiment, the skin improvement method comprises applying the composition as defined above, on the skin of a subject in need thereof, twice a month.

According to one embodiment, the skin improvement method comprises the injection of the composition as defined above to a subject in need thereof. According to one embodiment, the injection is a transdermal injection.

According to one embodiment, the skin improvement method is for restoring the production of endogenic hyaluronic acid in the epidermis and/or the dermis.

According to one embodiment, the skin improvement method is for limiting the shortcomings of the skin; preferably, for improving the hydration of the skin.

According to one embodiment, the composition is applied on the skin of a subject in need thereof, in a sole application. According to one embodiment, the composition is applied on the skin of a subject in need thereof, at least once a month, preferably, twice a month; more preferably, once a week; more preferably, the composition of the invention is applied only once a week.

The invention also relates to a device comprising at least one hyaluronic acid derivative of formula (I) and/or (Π); or comprising at least one particle of the invention.

According to one embodiment, the device is selected from a syringe, a patch, a film, a stick, a plaster, a stupe or a bandage.

BRIEF DESCRIPTION OF THE DRAWINGS

Figure 1 is FT-IR spectra of N- stearylarnine-modified hyaluronic acid (Figure 1A) and iV-steaiymydrazme-modified hyaluronic acid (Figure IB).

Figure 2 is a histogram showing the percentage of the surface occupied glycosaminoglycans (GAGs) in papillary dermis. EXAMPLES

The present invention is further illustrated by the following examples.

Abbreviations

°C: Celsius degree;

DS: degree of substitution;

HA: hyaluronic acid;

FT-IR: Fourier-Transform Infra-Red spectroscopy;

kDa: kiloDalton;

M: mol/L;

MES: 2-(iV-morpholino)ethanesulfonic acid;

mg: milligram(s);

ml: milliliter(s);

Mw: molecular weight;

NHS: iV-hydroxysuccinimide;

N2: dinitrogen;

NMR: Nuclear Magnetic Resonance.

Materials and Methods

Reagents

All chemicals and solvents have been purchased from commercial sources (Aldrich, Acros) and were used as such. The solvents of spectroscopy quality were purchased from Aldrich.

ATR FTIR spectroscopy

Spectra were recorded in function of time under N2 flow using infrared spectrometer FT/IR-4100 JASCO. The principle deals with contacting a crystal (diamond) and the sample before to be through by the infrared light Quasi-elastic tight scattering

The mean hydrodynamic diameter of the particles of the invention was determined at 2S°C using a Zetasizer Nanoseries Nano-ZS90 (Malvern Instruments, France).

Image analysis All the micro scopial observations were realized using a Leica DMLB or Olympus BX43 microscope. Pictures were digitalized using an Olympus DP72 camera and the Cell^AD data storing software.

Collagen III immunostaining

Colagen III immunostaining has been realized on frozen sections with a polyclonal anti- collagen III antibody (SBA, Ref: 1330-01) diluted at 1:20 in PBSBSA 0.3 %-Tween 20, 0.0S % during lh at room temperature using a Vectastain Kit Vector amplifier system avidin/biotin, and revealed by VIP (Vector laboratories, Ref: SK-4600). The immunostaining was assessed by microscopic observation.

Elastin immunostaining Elastin immunostaining has been realized on paraffinized sections with a polyclonal anti- eslatin antibody (Noviotec, Ref.:25011) diluted at 1:400 during lh at room temperature, and revealed by AlexaFluor 488 (Lif etechnologies , Ref: A11008). Nuclei were post stained with propidium iodide. The immunostaining was assessed by microscopic observation. PART 1: CHFMISTPV

Example 1 ; Synthesis of the hyaluronic acid derivatives from hydrophobic compounds having at least one amine or hydrazine function

General protocol

Typical procedures for grafting hydrophobic groups onto sodium hyaluronate are described hereinafter from an amine compound (point 1.1) or from a hydrazine compound (point 1.2).

1.1 From an amine compound

About 1 equiv. of ethyl carbodiimide and 0.6 equiv. of N-hydroxysuccinirnide (NHS) are dissolved in a MES buffer (pH=4.5). Then, 1 equiv. of sodium hyaluronate (Mw=200 kDa) is slowly dissolved as well. After complete dissolution, 0.2 equivalent of an amine compound such as for example octadecylamine, is added so the final hyaluronic acid concentration ranges from 30 and SO mg/mL, and the mixture is let at rest for 5 hours at room temperature. The grafted HA is diluted with a solution of sodium chloride (0.5 M) to exchange cations. Then, the grafted HA is precipitated using a water/ethanol mixture (2/3; v/v) and can be further purified by washing it with more hydrophobic water/ethanol mixtures (up to 19/1; v/v). Drying is performed under vacuum between at around 30°C. A white powder is obtained.

The product was then characterized by FT-IR spectroscopy. This technique is an efficient and easy method for identifying the chemical functions of the HA derivatives by comparison with native HA. For example, the spectrum of HA grafted with octadecylamine is presented Figure 1 A. This analysis evidenced the presence of the bands corresponding to the grafting of alkyl chains compared to non-modified HA. The substitution degree of sodium hyaluronate (Mw=200 kDa) modified by amidation reaction with octadecylamine is 12 %. 1.2. Frnm a hydrazine cnmpound

About 0.2S equiv. of ethyl carbodiimide are dissolved in a MES buffer (pH=4.5). Then, 1 equiv. of sodium hyaluronate (Mw=3000 kDa) is slowly dissolved as well. After complete dissolution, 0.5 equivalent of a hydrazine compound such as for example octadecylhydrazine, is added so the final hyaluronic acid concentration ranges from 10 and 25 mg/mL, and the mixture is let at rest for 5 hours at room temperature. The grafted HA is diluted with a solution of sodium chloride (0.5 M) to exchange cations. Then, the grafted HA is precipitated using a water/ethanol mixture (2/3; v/v) and can be further purified by washing it with more hydrophobic water/ethanol mixtures (up to 19/1; v/v). Drying is performed under vacuum between at around 30°C. A white powder is obtained.

The product was characterized by FT-IR spectroscopy. The spectrum of HA grafted with octadecylhydrazine is presented Figure IB. This analysis evidenced the presence of the bands corresponding to the grafting of the alkyl chain compared to non-modified HA. The substitution degree of sodium hyaluronate (Mw=3000 kDa) modified by amidation reaction with octadecylhydrazide is 1.5 %.

1 3 Examples of hvdrophoMcallv-modified HA derivatives

Several hydrophobically-modified HA derivatives have been synthesized by one of the protocols described in paragraph 1.1 or 1.2. The characteristics of such compounds are presented in Table 1.

Example 2· Synthesis of particles comprising hyaluronic acid derivatives

General protocol

Hydrophobically-modified hyaluronic acids of example 1 were dispersed into a mixture of MilliQ* water and phosphate buffer (S0/S0 %: v/v) at room temperature under magnetic stirring. The formulation of die phosphate buffer is presented Table 2:

The final concentration of hydrophobically-modified hyaluronic acids was modified from 0.25 to 2 %wt depending on the solubility of HA derivatives and to keep the formulation liquid for size measurement The particles were characterized by quasi-elastic light scattering in order to determine their mean hydrodynarnic diameter. For example, the mean hydrodynamic diameter is 327 nm for .V-stearylamine modified HA having 200 kDa and is 357 nm for N- stearylamine modified HA (3000 kDa).

Examples of particles of hydrophobically-modified HA Several particles of hydrophobically-modified HA have been synthesized by the protocol of the invention described above. The characteristics of such compounds are presented in Table 3, including the degree of substitution calculated by NMR (DS NMR) and the molecular weight of hyaluronic acid (Mw of HA).

PART 2: BIOLOGY

Example 3; Lasting effect of the particles of the invention in the papillary dermis

The aim is to show that the particles of the invention are able to deliver in the papillary dermis by a sole application, a large amount of hyaluronic acid derivatives while limiting the degradation of the product in the skin over time.

Three formulations were studied during 7 days on ex vivo human skin abdomen explants:

PI 1 : particles of .V-stearylamine modified HA having 200 kDa (amide linkage); P14: particles of .V-stearylamine modified HA having 3000 kDa (amide linkage); and

- PO: particles of 0-stearoyle modified HA which are HA modified with stearic acid (ester linkage).

PI 1 and P14 are the particles of the invention whereas PO are particles for comparison, said particles PO comprising hydrophobically-modified HA through ester bond.

The skin human explants have an average diameter of 11 nm (± 1 mm) and were collected after abdominoplasty from a 44-year-old woman. The explants were kept in survival in a culture medium at 37°C in a humid, 5 % CO2 atmosphere.

For each of these formulations, the content of glycosaminoglycan (GAG) in papillary dermis has been quantified over time (day 3, day 5 and day 7) compared to a witness sample (no treated sample, noted "C")- The general morphology is observed over time via histological processing.

The beginning of this experiment (day 0) corresponds to the day where a topical administration of the formulation has been carried out on the explant (4 mg/cm²).

The results are presented Figure 2.

The histogram shows that:

- After 3 days and 5 days, the amount of particles of PO (ester linkage) is slightly higher than those of PI 1 and P14 (amide linkage); After 7 days, the amount of particles of PO (ester linkage) has dramatically decreased contrary to those of PI 1 and P14 (amide linkage).

In conclusion, these results show that the modified HA having an amide linkage, independently of the molecular weight, is less degraded man modified HA having an ester linkage after a long time. Consequently, a sole application on the skin of the particles of the invention allows keeping the benefits of these compounds over time.

Example 4; Penetration depth of the particles of the invention in the dermis

The aim is to show that the particles of the invention are able to be diffused until the deepest layer of the skin. For this purpose, three formulations were studied during 7 days on ex vivo human skin abdomen explants:

PI 1 : particles of .V-stearylamine modified HA having 200 kDa (amide linkage); P14: particles of -V-stearylamine modified HA having 3000 kDa (amide linkage); and

- PO: particles of O-stearoyle modified HA which are HA modified with stearic acid (ester linkage).

For each of these formulations, the total content of glycosaminoglycan (GAG) has been quantified over time (day 3, day 5 and day 7) through 3 different compartments of dermis: the papillary dermis, the upper reticular dermis and the lower reticular dermis. These results have been compared to a witness sample (no treated sample, noted "W").

The results show mat:

After 3 days and 5 days, the amount of particles of PO (ester linkage) is higher than those of PI 1 and P14 (amide linkage) in the three compartments of the dermis; After 7 days, the amount of particles of PO (ester linkage) has dramatically decreases in the three compartments of the dermis contrary to those of PI 1 and P14

(amide linkage). In conclusion, these results evidenced that die particles of the invention are able to keep the benefits of a sole topical application until the deepest layer of the skin.

Example 5; Action of the particles of the invention on collagen HI and elastin

The action of the particles on the expression of collagen HI and elastin were evaluated ex vivo in the dermis after a sole application on human skin abdomen explants.

The studied formulations are:

R: sodium hyaluronate;

PI 1 : particles of -V-stearylamine modified HA having 200 kDa (amide linkage); P14: particles of -V-stearylamine modified HA having 3000 kDa (amide linkage); and

PO: particles of 0-stearoyle modified HA which are HA modified with stearic acid (ester linkage).

For each of these formulations, the collagen Π and elastin contents have been quantified in the papillary dermis after 7 days compared to a witness sample (no treated sample, noted "C").

The quantification has been carried out by an image analysis of collagen Π and elastin after specific immunological labelling of samples.

After 7 days, the results show that: for elastin

the topical application of sodium hyaluronate (R) does not induce any variations in the expression of elastin in the papillary dermis;

the topical application of 0-stearoyle modified HA (PO) slightly increases the expression of elastin in the papillary dermis;

the topical application of -V-stearylamine modified HA (PI lor P14) significantly increases the expression of elastin in the papillary dermis. for collagen III

for all the formulations, die expression of collagen HI is similar. These results show that the particles of the invention keep a good efficiency compared to hydrophobized HA comprising ester linkages. In conclusion, these results show that the modified HA having an amide linkage, independently of the molecular weight, allows efficiently stimulating the expression of elastin compared to sodium hyaluronate and modified HA having ester linkage.

F.xample fr Lasting effect of particles of modified HA having hydrazide linkage

The aim is to show that the particles of modified-HA comprising hydrazide linkages, are able to deliver in the papillary dermis by a sole application, a large amount of hyaluronic acid derivatives while limiting the degradation of the product in the skin over time. The aim is also to compare the efficiency as a long lasting anti-aging agent of particles made of modified-HA comprising hydrazide linkages and of particles made of modified-HA comprising amide linkages. The following formulations were studied during 7 days on ex vivo human skin abdomen explants:

P13: particles of N- steary lhydrazine modified HA having 3000 kDa (hydrazide linkage); and

PI 1 : particles of .V-stearylamine modified HA having 200 kDa (amide linkage). The skin human explants have an average diameter of 11 nm (± 1 mm) and were collected after abdominoplasty from a 51-year-old woman. The explants were kept in survival in a culture medium at 37°C in a humid, 5 % CO2 atmosphere.

For each of these formulations, the content of glycosaminoglycan (GAG) in papillary dermis has been quantified over time (day 2 and day 7) compared to a witness sample (no treated sample, noted "C"). The general morphology is observed over time via histological processing. The beginning of this experiment (day 0) corresponds to the day where a topical administration of the formulation has been carried out on the explant (4 mg/cm²).

The results are presented in the following Table 4:

These results show that:

- a sole application of particles Pll (amide linkage) or of particles P13 (hydrazide linkage) on the skin increases the surface of GAGs in the papillary dermis even after 7 days; and

the application on the skin of particles P13 (hydrazide linkage) provides an improved effect compare to particles PI 1 (amide linkage). In conclusion, these results show mat the modified HA having a hydrazide linkage has an improved long lasting anti-aging effect than modified HA having an amide linkage.

Consequently, a sole application on the skin of the particles of modified HA having hydrazide linkages, allows achieving improved effects of these compounds over time.

Claims

Use as a long lasting antiaging agent of a particle comprising at least one hyaluronic acid derivative of general formula (I):

or any salts and solvates thereof, wherein

Z represents a single bond or -NH-;

R represents a C1-C20 alkyl group; preferably, a C6-C18 alkyl group; more preferably, a C18 alkyl group; n represents the number of disaccharide units of D- glucuronic acid and iV-acetyl-D- glucosamine and is a positive integer; and wherein the particle has a mean hydrodynamic diameter ranging from 200 nm to 500 nm; preferably, from 250 nm to 400 nm; more preferably, from 320 nm to 360 nm.

2. The use of a particle according to claim 1, wherein the degree of substitution (DS) of the hyaluronic acid derivatives of formula (I) ranges from 0.1 % to 55 %. 3. The use of a particle according to claim 1 or claim 2, wherein the molecular weight of the hyaluronic acid derivatives of formula (I) ranges from 100 kDa to 4000 kDa; preferably, from 200 kDa to 3 000 kDa. Hie use according to anyone of claims 1 to 3, wherein the long lasting antiaging agent is a long lasting skin plumping agent, a long lasting skin hydrating agent and/or a long lasting stimulating agent of the endogenic production of hyaluronic acid.

A particle comprising at least one hyaluronic acid derivative of general formula (I):

or any salts and solvates thereof,

wherein

Z represents -NH-;

R represents a C1-C20 alkyl group; preferably, a C6-C18 alkyl group; more preferably, a C18 alkyl group;

n represents the number of disaccharide units of D- glucuronic acid and -V-acetyl-D- glucosamine and is a positive integer; and wherein the particle has a mean hydrodynamic diameter ranging from 200 nm to 500 nm; preferably, from 250 nm to 400 nm; more preferably, from 320 nm to 360 nm.

6. A particle acconimg to claim 5, for use as a medicament

7. A particle for use according to claim 6, in the treatment and/or the prevention of dermatological conditions; preferably, as a healing agent

8. A cosmetic and/or dermo-cosmetic composition comprising at least one particle according to claim 5, and a cosmetically acceptable base.

9. Hie cosmetic and/or dermo-cosmetic composition according to claim 8, wherein said composition is under an injectable form. 10. A skin improvement method comprising using the cosmetic and/or dermo-cosmetic composition according to claim 8 or claim 9, in a subject in need thereof.

11. Ihe skin improvement method according to claim 10, for restoring the production of endogenic hyaluronic acid in the epidermis and/or the dermis.

12. The skin improvement method according to claim 10, for limiting the shortcomings of the skin; preferably, for improving the hydration of the skin.

13. The skin improvement method according to anyone of claims 10 to 12, wherein the composition is administered to the skin of a subject in need thereof, once per week.

14. Ihe skin improvement method according to anyone of claims 10 to 13, wherein the composition is administered to the epidermis and/or the dermis of a subject in need thereof, by a topical application or by injection.