FR3006897A1 - COSMETIC OR THERAPEUTIC USE, IN PARTICULAR DERMATOLOGY OF DERMICIDINE, ANALOGUES OR FRAGMENTS THEREOF FOR PREVENTING THE APPEARANCE OF IMPERFECTIONS, AS WELL AS FOR THE TREATMENT AND DIAGNOSIS OF ACNEIC OR ACNETIC TREND SKIN - Google Patents

Abstract

The subject of the present invention is, in particular, the use of a cosmetic and / or therapeutic composition, in particular a dermatological composition containing at least one amino acid sequence of Dermicidin, an analogue or a fragment thereof. , or at least one nucleic acid sequence encoding this sequence, to prevent the appearance of imperfections, as well as for the treatment and / or prevention and / or diagnosis of acne-prone or acne-prone skin. It also relates to a method for applying the aforementioned cosmetic composition for preventing the appearance of imperfections and the treatment of acne-prone skin. It further relates to a method of treating an acne skin comprising topically applying to said skin the pharmaceutical composition for the treatment of acne skin. The present invention also relates to the use of a biomarker for diagnosing acne-prone or acne-prone skin.

Description

The present invention relates to the use of an amino acid sequence of Dermicidin, an analogue or a fragment thereof to prevent the appearance of imperfections or for the treatment and / or prevention and / or diagnosis of acne or acne prone skin. Acne vulgaris is a multifactorial disease (face, scapular region, arms and intertriginous regions). It is the main cause of the most frequent dermatoses. It is important not to trivialize this disease and to treat it properly, because it can have disabling psychosocial consequences, in particular because of the formation of scars.

In the lightest form, it concerns almost every human being. It peaks at the age of puberty, but can occur for the first time as early as 7 to 9 years of age and up to 40 years of age. It is common to suffer from acne again after 25 years. Acne is a disease of the follicle of the sebaceous gland. The following pathogenic factors play a determining role in the constitution of acne: - genetic predisposition - androgen - abnormally increased keratinization (keratinization disorder) in the infundibular portion of the hair follicle. Indeed, in the deepest parts of the infundibulum, there is the formation of a higher amount of keratinocytes than normal. These cells differentiate into horny cells that gradually block the lumen of the follicular canal. The physiological process of continuous desquamation of the acro-infundibulum to the surface is disturbed by the increased adhesion of the horny cells produced. It forms a hyperkeratotic plug constituting the comedo, initial lesion of acne. - Bacterial infections responsible for the hydrolysis of free fatty acids and inflammatory phenomena (papules and pustules). The three predominant local organisms, Staphyloccus epidermidis, Malassezia furfur and Propionibacterium acnes, are found in the sebaceous follicle as an ideal nutrient medium. The clinical manifestations are characterized by a polymorphous table. We will focus here on the most common forms of acne namely comedian acne (juvenile acne), papulopustular and / or nodular acne.

Retention lesions may be open or closed comedo (microcyst, micro comedo, white head). Inflammatory lesions derived from retention lesions may be papules, pustules, with indurated nodules, abscesses, fistulas, scarring. ECLA GRID ASSESSMENT The clinical severity of acne was determined using the ECLA semi-quantitative rating grid. This grid consists of three factors: Table of the ECLA grid Factor i i pe isitensite of acne; counting, age, fairness Pyloyen I nt = 1 = 2 = 3 = Auc ', <5 5 to 9 -Io to a <5 5 to 10 to 19 20 to Is a and nu' v is None 4 5 in '1 Terisio factor, 1 t sort of acne, excluding F2 visa ica iaue Zo cen, Pe: UH re Dos Bris Score 2 = Factor - scars absent = o; present lr.flarr <arc: one inflammatory, loriatic- CI CN I Score .3 - Score Score. 3 = The s between 0 and 36. Jou-, In the sense of the invention, is meant by "skin" the entire epidermis of the human body. More particularly, the skin considered in the present invention is preferably the skin of the face, décolleté, back, and preferably the skin of the face. For the purposes of the invention, the term "acne-prone skin" means a skin presenting an acne with an ECLA score of less than 10, preferably between 1 and 9.

For the purposes of the invention, the term "acne skin" means a skin presenting an acne with an ECLA score of between 10 and 36. For "imperfections" within the meaning of the invention, it is meant the cutaneous marks or temporary scars or definitive due to acne lesions that are perceived by the subject as unsightly. For the purposes of the invention, the term "prevent" means reducing the risk of occurrence or slowing down the occurrence of a given phenomenon, namely in the present invention, cutaneous marks and scars induced by pimples. , open or closed comedon-like retention lesions (microcysts, micro comedones, white heads), inflammatory lesions derived from retention lesions such as papules, pustules, with indurated nodules, abscesses, fistulas, cicatricial states.

The cosmetic treatment of this type of skin generally involves the topical application to the skin of a mixture of compounds making it possible to act on the various causes of imperfections and in particular to promote desquamation and / or to limit microbial growth.

For the control of pathogens such as Propionibacterium acnes, active agents such as retinoids, azelaic acid, benzole peroxide, clindamycin, erythromycin and sodium sulfacetamide are commonly used. But the use of these assets is not devoid of side effects. Thus, retinoids and azelic acid are only used in the case of mild acne because they are not very effective on inflammatory lesions and can induce dryness of the skin. Benzole peroxide is likely to cause skin irritation, hair whitening and photoallergic reactions, so it has a significant toxicity (Research in humans phototoxic power of 5% henzoyl peroxide. M Jeanmougin, J Pedreiro, J Bouchet, J Civate - Dermatology, 1983. Drug-induced photoallergic and phototoxic reactions, July 2007, Vol 6, No. 4, Pages 431-443 Kevin Stein & Noah S Scheinfeld). Clindamycin, erythromycin and sodium sulphonamide in the form of salts are sensitizing substances, likely to cause allergies, and leading to resistance (D Sasseville - Progress in Dermato-Allergology Volume XVI: Strasbourg 2010 There is therefore still a need for topically active agents having an effect on the pathologies related to the genus Propionibacterium, and particularly Propionibacterium acnes, while not having the disadvantages of the compounds known for this use. The present invention reveals the need to identify new molecules in order to maintain an effective action on the growth inhibition of P. acnes over time, and the Applicant has discovered that Dermicidine, a natural antimicrobial, makes it possible to achieve this goal. Unexpectedly, the inventors have discovered that the antimicrobial peptide DCD-1L, derived from the cleavage of Dermic idine, known to be present in sweat, was active in the growth of the P. acnes pathogen. The Applicant has indeed found that Dermicidine and the peptides derived therefrom according to the invention have a strong inhibitory activity on the growth of Propionibacterium acnes, and could therefore be used in a cosmetic or pharmaceutical composition, especially a dermatological composition, in particular for prevent the appearance of imperfections or for the treatment and diagnosis of acne or acne prone skin.

The subject of the invention is therefore the use of a cosmetic composition containing at least one amino acid sequence of Dermicidin, an analogue or a fragment thereof to treat and / or prevent the skin from occurring. acne tendency and prevent the appearance of imperfections related to the genus Propionibacterium, including Propionibacterium acnes.

Another subject of the invention is the use of a pharmaceutical composition containing at least one amino acid sequence of Dermicidin, an analogue or a fragment thereof for preventing and treating skins. acne and imperfections associated with the genus Propionibacterium, including Propionibacterium acnes.

Another subject of the invention is the use of a pharmaceutical composition containing at least one nucleic acid sequence coding for an amino acid sequence according to the invention, or at least one expression modulating agent or the activity, especially biological, of said amino acid sequence to prevent the appearance of imperfections and to treat and / or prevent acne skin. The invention also relates to a cosmetic treatment method for the treatment of acne-prone skin and the prevention of imperfections, in which a cosmetic composition comprising an amino acid sequence of Dermicidin, an analogue, is applied to the skin. or a fragment thereof as defined hereinafter. The invention relates to a pharmaceutical treatment method, especially a dermatological method, in which a pharmaceutical composition comprising Dermicidin, an analogue or a fragment thereof or at least one amino acid sequence according to the invention is applied to the skin, at least one nucleic acid sequence coding for an amino acid sequence according to the invention, or at least one agent that modulates the expression or the activity, in particular the biological activity, of said amino acid sequence.

Another aspect of the invention is the use of Dermicidin or an amino acid sequence derived therefrom, an analog or fragment thereof, or a nucleic acid sequence encoding a DNA sequence. amino acid according to the invention, especially as a biomarker for the diagnosis of acne skin or acne prone.

Dermicidin Dermicidin / Protease Inducing Factor (DCD / PIF) is a 110 amino acid (aa) protein with a signal peptide of 19aa whose gene is located on chromosome 12, locus 12q13.1 and which is constitutively expressed in the sweat gland (Rieg, Garbe et al., 2004). In addition, two isoforms are found in the UniProt base, one shorter (isoform 1) and the other longer isoform 2. It is secreted in sweat (Schittek, Hipfel et al., 2001). After proteolytic post-translational processing (mainly by cathepsin D) in sweat (Baechle, Flad et al., 2006), the protein precursor gives rise to many peptides ranging in length from 25 to 48 aa (Flad et al. Bogumil et al., 2002). Some of these peptides, such as LL37, show antimicrobial activities against S. aureus, E. coli, Enterococcus faecalis, Candida albicans and P. acnes microorganisms. The antimicrobial peptides are derived from the C-terminal portion of the protein. The activity of these peptides is independent of their charge (Steffen, Rieg et al., 2006) and would go through a step of interaction with bacterial phospholipids (Li, Rigby et al., 2009). The N-terminal portion of the protein consists of a biologically active factor promoting cell survival (Cunningham, Hodge et al., 1998). The decrease in DCD observed in atopic sweat may partly explain the microbial susceptibilities of this type of skin (Rieg, Steffen et al., 2005). A recent publication reports on the use of Der) as a particularly sensitive specific marker, in which the range of sweat traces is determined by the use of R 1 or by L for use in rheumatoid arthritis. (Sakurada, Akutsu et al., 2009), 11 It has been shown that DCD associates with bacterial phospholipids and self-organizes by forming ibni channels in bacterial raertibrene (Paulniann, Arnold et al., 2012). The concentration of amino acids and urethane acids Dermidin (DCD) associated with its signal peptide is a protein of 110 amino acids (I), Q ID NO: I. having a signal peptide of 19 aelds (SEQ ID NO: 14) whose gene is located on chromosome 12, locus 12q13.1, two forms of this identified protein, one shorter, isoform 1 (SEQ ID NO: 1.23, the other longer, isof) me 2 (S.FQ ID NO: 13) After cleavage of the signal peptide, Dermicidin (DCD) is a 90 amino acid protein according to US Pat. Equetice SEQ ID: 16, A. After a proteolytic post-translational matteation essentially per cuthepsin takes place in sweat (13aechle, Flad and 2006), this precursor protein gives ilahiSaTICC to fsSEQ ID NO: 19) and to 1) CD Thus, many peptides, the length of which varies from 25 to 48 years, are of the same magnitude as those of Bouumil et al. (P) Unless otherwise indicated; the term Dermieidine is intended to include in the present invention the amino acid seques- ters represented by: SEQ ID # -EQ ID: NO:: 12, SEQ ID M "): SEQ ID N (1: 16, SEQ -NO: tr: SEQ: - 1r) NO: 20. a, wherein has undergone a: post-translational maturation # SOon: of embodiment of the title: Meid deigriff Dar Dcn-nkidirw Sequencing of amino acid depicted as SEQ ID NO: 16, According to a still more preferred embodiment, it is understood, in particular, that the amino acid sequence represented by SEQ ID NO: 1.0, the N-terminal part of the protease, is more particularly understood by mieidine. (SEQ ID NO: 15 consists of a biologically active 1z-guard promoting eelluTzlire survival (Ciamingbrum, I lodge et al., By "analog of an amino acid sequence" according to the invention is meant any amino acid sequence having a sequence identity of at least 85%, preferably at least 90%, and more preferably at least 95%, and still more preferred of at least 99% with said sequence, and a biological activity of the same kind. By "biological activity of the same nature" with respect to an amino acid sequence according to the invention is meant the antimicrobial or stimulation properties of the survival and / or cell proliferation usually attributed to Dermicidine.

Sequence identity can be determined by visual comparison or by any computer tool commonly used in the domain, such as BLAST programs available at www.ncbi.nlm.nih.gov and used with default settings. An analogue according to the invention may be a peptidomimetic agent.

An analogue of an amino acid sequence of the invention may result from modifications resulting from mutation or variation in the sequences of the peptides according to the invention either from the deletion or from the insertion of one or more amino acids either the substitution of one or more amino acids or alternatively splicing. Many of these changes can be combined.

Advantageously, an analogue of an amino acid sequence of the invention may comprise conservative substitutions with respect to this amino acid sequence. By way of example of mutations which may be considered in the present invention, it may be mentioned, in a non-exhaustive manner, the replacement of one or more amino acid residues by amino acid residues having a subscript. similar hydropathic without substantially affecting the biological properties of the polypeptide. The hydropathic index is an index attributed to amino acids as a function of their hydrophobicity and charge (Kyte et al (1982) J. Mol Biol., 157: 105).

An amino acid sequence or an analogue thereof targeted by the present invention may be an amino acid sequence having undergone one or more post-translational processing (s).

By "post-translational processing (s)" is intended to encompass all the modifications that an amino acid sequence is likely to undergo at the end of its synthesis in a cell, such as, for example, a or phosphorylation (s), thiolation (s), acetylation (s), glycosylation (s), lipidation (s), such as farnesylation or palmitoylation, rearrangement Disulfide-type and / or cleavage-type structural bonding within the peptide sequence. An analogue of an amino acid sequence has, moreover, substantially the same biological activity as this amino acid sequence.

It is moreover known that a primary amino acid sequence may comprise sites specifically recognized by protease-type enzymes, such as trypsin, which, once the recognition of these effective sites will induce the cleavage of the sequence by proteolysis. This proteolysis results in the generation of various peptides, or fragments of amino acid sequences of the invention.

Consequently, the invention also extends to fragments of Dermicidin, possibly resulting from its proteolysis. Within the meaning of the invention, the term "fragment of an amino acid sequence" any portion of the amino acid sequence according to the invention comprising from 3 to 48 consecutive amino acids of said sequence.

According to one embodiment, an amino acid sequence that is suitable for the invention may be an amino acid sequence represented by a sequence chosen from SEQ ID NO: 11 to 20, in particular from SEQ ID NO: 16, SEQ ID NO : 20, an analogue, or a fragment thereof. According to a preferred embodiment, an amino acid sequence that is suitable for the invention may be an amino acid sequence represented by a sequence chosen from SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18, SEQ ID NO: 19, SEQ ID NO: 20, an analogue, or a fragment thereof. More preferably, an amino acid sequence suitable for the invention may be an amino acid sequence represented by the sequence SEQ ID NO: 20, an analogue or a fragment thereof.

According to another embodiment, a polypeptide that is suitable for the invention may also be a natural or synthetic amino acid sequence, which may optionally be obtained after enzymatic or chemical hydrolysis of Dermicidine or by chemical or biological synthesis, or by extraction from a biological tissue, such as the skin, naturally expressing this amino acid sequence or after transfection thereof, as well as the various post-translational forms thereof, or any natural or synthetic amino acids whose sequence completely or partially comprises a aforementioned amino acid sequence, for example variants and analogs.

Those skilled in the art can obtain an amino acid sequence according to the invention by means of recombinant DNA methods, for example those described in the manual "Molecular Cloning - A Laboratory Manual" (2nd edition). edition), Sambrook et al., 1989, Vol. I-III, Coldspring Harbor Laboratory, Coldspring Harbor Press, NY, (Sambrook).

According to one embodiment, the present invention also relates to nucleic acid sequences SEQ ID NO: 1 to SEQ ID NO: 10 coding respectively for the amino acid sequences SEQ ID NO: 11 to SEQ ID NO: 20 of the invention. invention and their implementation in the various uses and methods according to the invention.

For the purposes of the present invention, the term "nucleic acid sequence fragment" means a nucleic acid sequence encoding an amino acid sequence having a biological activity of the same nature as the amino acid sequence encoded by said sequence. For the purposes of the present invention, the term "nucleic acid sequence analogue" means a nucleic acid sequence having a sequence identity of at least 85%, preferably at least 90%, and more. preferably at least 95%, and even more preferably at least 99% with said sequence, and coding for an amino acid sequence having a biological activity of the same nature as the amino acid sequence encoded by said sequence.

By "analog of a nucleic acid sequence" is meant a nucleic acid sequence, possibly resulting from the degeneracy of the nucleic acid code, and coding for an amino acid sequence according to the invention, in particular as defined above. According to a preferred embodiment, a nucleic acid sequence of the invention may be represented by at least one sequence selected from SEQ ID NO: 6 or SEQ ID NO: 10 and more preferably SEQ ID NO: 10. A nucleic acid sequence of the invention may be of any possible origin, namely animal, in particular mammalian and even more particularly human, either plant or micro-organisms, such as, for example, viruses, phages, or bacteria among others, or fungi, without prejudging the fact that they are present naturally or not in said original organism. According to one embodiment, the invention also relates to the isolated and purified nucleic acid sequences coding for an amino acid sequence considered according to the invention, as well as to their analogs and fragments. A nucleic acid sequence according to the invention may comprise a sense, antisense or interferential sequence corresponding to a sequence coding for a polypeptide according to the invention. Compositions Cosmetic Composition The present invention also relates to cometic compositions comprising in an cosmetically acceptable medium an effective amount of at least one amino acid sequence of the invention. A composition of the invention may contain adjuvants customary in the field under consideration, such as hydrophilic or lipophilic gelling agents, preservatives, antioxidants, solvents, perfumes, fillers, filters, odor absorbers and materials. dyes, oils, water waxes and polyols. The amounts of the various constituents of the compositions according to the invention are those conventionally used in the fields under consideration. The amount of amino acid sequence, or nucleic acid sequence of the invention, or active agent according to the invention contained in a composition of the invention, also called "effective amount" is, of course depending on the nature of the asset and the effect sought and may therefore vary to a large extent. To give an order of magnitude, a composition may contain an amino acid sequence, or a nucleic acid sequence, or an active agent according to the invention in an amount representing from 0.00001% to 2.5% of the total weight of the composition, in particular in an amount representing from 0.001% to 2% of the total weight of the composition, and more particularly in an amount representing from 0.1% to 1% of the total weight of the composition. Within the meaning of the present invention, the term "effective amount" of a compound of the invention, a sufficient and necessary amount of this compound to obtain a desired effect, and more particularly a cosmetic effect or care with regard to acne skin. Thus, and without being in any way limiting within the meaning of the present invention, those skilled in the art will be able to refer to the examples below, in which the said at least one amino acid sequence represents 0.01% of the composition.

Pharmaceutical Composition The present invention also relates to pharmaceutical compositions, especially dermatological compositions comprising in a physiologically acceptable medium an effective amount of at least one amino acid sequence in accordance with the invention, or at least one nucleic acid sequence in accordance with the invention. to the invention, or at least one active agent capable of modulating the activity, the expression or the maturation of said amino acid sequence or of said nucleic acid sequence.

For the purposes of the present invention, the term "modulating agent" or "active agent capable of modulating the expression, the processing or the activity of an amino acid sequence or nucleic acid sequence conforming to the invention "any compound capable of acting, directly or indirectly, on at least one amino acid sequence or a nucleic acid sequence in accordance with the invention, or on an element of an intracellular signaling pathway or extracellular, or a metabolic pathway, or transcriptional and / or translational regulation involving said amino acid sequence or said nucleic acid sequence.

For the purposes of the invention, the term "modulate", with respect to a given effect, the action of stimulating or inhibiting this effect. For the purposes of the present invention, the term "physiologically acceptable medium" is intended to mean a medium that is suitable for the administration of a composition topically to the skin, the scalp, or the lips, or orally or by the route. parenteral such as the intradermal or subcutaneous route. The amounts of the various constituents of the compositions according to the invention are those conventionally used in the fields under consideration.

The amount of amino acid sequence, or nucleic acid sequence of the invention, or active agent according to the invention contained in a composition of the invention, also called "effective amount" is, of course depending on the nature of the asset and the effect sought and may therefore vary to a large extent. To give an order of magnitude, a composition may contain an amino acid sequence, or a nucleic acid sequence, or an active agent according to the invention in an amount representing from 0.00001% to 5% of the total weight of the composition, in particular in an amount representing from 0.001% to 5% of the total weight of the composition, preferentially from 0.1 to 5 and more particularly in an amount representing from 0.5% to 5% of the total weight of the composition and even more preferably 2.5% to 5%. According to another embodiment, a composition at least one additional active therapeutic agent.

The present invention relates to the cosmetic use of an effective amount of at least one amino acid sequence as an active agent to prevent the appearance of imperfection and / or treat acne-prone skin.

In another aspect, the present invention relates to a cosmetic or non-therapeutic method for preventing the appearance of imperfections and / or treating acne-prone skin in an individual in need thereof, the method comprising at least one step of to administer to said individual, at least one composition comprising as active agent at least one amino acid sequence of the invention, in particular as defined above. A method or a use of the invention makes it possible to prevent the appearance of imperfection and / or to treat acne-prone skin. A method or a use of the invention makes it possible to maintain a healthy skin. For the purposes of the invention, the term "healthy skin" means a skin that does not have imperfections.

Preferably, a method of the invention may comprise the application topically to at least a portion of the skin of an individual in need thereof, in particular on the skin of the face, back and / or décolleté, at least one layer of a topical composition of the invention.

A cosmetic process according to the invention may be carried out daily, for example for a single application per day, or a divided administration in two or three times a day, for example once in the morning and once. the evening. A cosmetic process according to the invention may be carried out over a period of time varying from one week to several weeks, or even several months, this period being able to be repeated after periods of non-treatment for several months or even several years. . By way of example, a cosmetic process according to the invention can be provided for applying a cosmetic composition of the invention, for example, at a rate of 1, 2 or 3 times per day, or more, and generally on a extended duration of at least 4 weeks, or even 4 to 15 weeks, with one or more periods of interruption where applicable. The present invention relates to the therapeutic use of an effective amount of at least one amino acid or nucleic acid sequence of the invention or at least one agent for modulating the activity, the expression or maturation of said amino acid sequence or said nucleic acid sequence, and in particular a modulating agent as defined above, as active agent to prevent the occurrence of imperfections and / or prevent and / or treat acne skin. In another aspect, the present invention relates to a therapeutic method, for preventing the appearance of imperfections and / or preventing and / or treating acne skin in an individual in need thereof, the method comprising at least one step of administering to said at least one composition comprising as active agent (i) at least one amino acid sequence of the invention or (ii) at least one nucleic acid sequence of the invention, or (iii) at least one less a modulating agent for the activity, expression or maturation of said sequences of the invention, in particular as defined above. A method or a use of the invention allows to find a healthy skin, manifested by a smooth skin without imperfections.

A method or a use of the invention makes it possible to prevent and / or reduce the presence of a microrelief of the skin. Preferably, a method of the invention may comprise the application topically to at least a portion of the skin of an individual in need thereof, in particular on the skin of the face, back and / or décolleté, at least one layer of a topical composition of the invention. A topically therapeutic method according to the invention may advantageously comprise the application of a composition of the invention, in combination simultaneously, sequentially or separately in time with an additional cosmetic or dermatological composition distinct from the composition of the invention. invention and intended for the care of the skin. A pharmaceutical process according to the invention may be carried out on a daily basis, for example on the basis of a single application per day, or of a divided administration in two or three times a day, for example once in the morning and in the morning. times in the evening. A pharmaceutical process according to the invention can be carried out over a period of time varying from one week to several weeks, or even several months, this period being able to be repeated after periods of non-treatment for several months, or even several years. . By way of example, a pharmaceutical method according to the invention may provide for an application of a composition of the invention, for example, at a rate of 1, 2 or 3 times per day, or more, and generally over a period of time. extended by at least 4 weeks, or even 4 to 15 weeks, with one or more periods of interruption if necessary.

In a second embodiment, the present invention relates to the field of biomarkers of the skin, and more particularly of the acne skin, and to their use as targets or cosmetic active agents. These factors can be used as bio-markers of the skin, as screening targets or even as cosmetic actives.

There is also a need for new biomarkers for characterizing a state of the skin, and in particular acne-prone or acne-prone skin, or imperfections Biomarker To the inventors' knowledge, this antimicrobial protein of the cutaneous surface has never been classified as variant acne skin and even less as one of the most relevant biomarkers emerging from this type of study.

The present invention relates to the use of at least one amino acid sequence of the invention, or at least one nucleic acid sequence of the invention, as a biomarker of a skin condition. Preferably, a use according to the invention makes it possible to characterize a state of the skin such as acne skin. The present invention relates to an in vitro or ex vivo method for characterizing an acne state of the skin of a subject comprising at least the steps of: offetuer. :: :: in an .isole sample of a skin of a tnesult.7 qualitative quarrying experience. :: Maturation I Activity of the biornarqueu r. b) compaiet..this measurement carried out in step a) at a reference point, c.) Conclude: ipu a decrease in e.ctivity, es.pmSSion or inaturation of said biornarquer by - Such a measure of nonthetic or acne-like behavior of the subject is defined by the meaning of the invention or by the meaning of the invention. pre-sence: the content or the degree of aelivity characteristic of a biological, pathological or pathological process, or the effect of the administration of the disease. The preferred active agent is a dermicidin or an acid sequence derived therefrom which is derived from the protein, or an analog thereof or a fragment thereof; a number of different acids coding for a serum of hydrolyzed acid, in particular as a biomarker for the evaluation of a state of the epidermis, which may have one of the eagles of the trolifera "anomalous", such as: P. eiém..v. According to one embodiment, a decrease in ace`.ivrte, expression or L.>: maturalior_ (said biumarqueur may be indicative of a vascular acneio-c 'z% trend According to: a. ::: - Node of realization, A: aueinentutWi - ..: 4 ..: r .... eibity: .... dez I, kijr,: zsrioh .. or of the maturation of said biôntarq -eur to be: iindieetNe.: enti .:.: treatment: cosi-netique effieace..reour e ... erc.er a beeiioue effect: on the -;.:.: eru .., :: 15-i-eérendeilen-Wii a. effect to skin acneelut or tendanoc, for use in accordance with the invention. a decrease or an increase in: activity, expressioti or delainiaturation said Bioniarqueur can be determined by comparison with a reference measurement obtained by any method known to those skilled in the art. A "reference measurement" for a given parameter, is a qualitative or quantitative measure of this parameter carried out under so-called "control" or "normal" conditions, for example determined in a reference sample, or determined in a sample in the absence of a treatment alleged to have an effect on the parameter. For example, a reference measurement for an amino acid sequence or a nucleic acid sequence according to the invention may be a quantitative or qualitative value relating to the expression, the maturation or the activity of said sequences determined in a physiologically healthy skin sample, or determined in a skin sample, especially acne skin, before a cosmetic or therapeutic treatment. Preferably, a reference measurement is a statistical measurement, that is to say having been repeated on different samples so as to obtain an average.

The reference measurement can be performed in parallel or sequentially with the test measurement. It can also be a "historical" measure, that is to say carried out prior to the test measurement, and stored, for example in a database, for later use.

A comparison of the test measurement with a reference measurement, and an observation of a deviation or absence of deviation between the two measurements, makes it possible to derive information as to the measured parameter, for example the decrease or the increase. expression, processing or activity of an amino acid sequence or a nucleic acid sequence according to the invention.

Such information may subsequently be used to determine the healthy or acne-prone and / or acne-like nature of a skin. According to one embodiment, the invention relates to a method, in particular in vitro or ex vivo, for characterizing a state of acne-prone or acne-prone skin. The qualitative or quantitative measurement of the expression, maturation or the activity of a nucleic acid sequence of the invention may be determined by any method known to those skilled in the art. By way of example of methods that are suitable for the invention, mention may be made of the polymerase chain reaction (PCR), quantitative (Q-PCR) or not, in the presence or absence of reverse transcriptase (RT-PCR or Q-RT-PCR), Northern-blot, ribonuclease protection assay, microarray methods, transcriptomic chip methods, oligonucleotide microarray methods, hybridization methods if you.

By way of example of agents suitable for the detection of a nucleic acid sequence of the invention, and in particular of an mRNA sequence, mention may be made of labeled nucleic acid probes which can be hybridizing to a nucleic acid sequence of the invention. Such a nucleic acid probe can be easily obtained by any method known to those skilled in the art. Thus, the nucleic acid sequences according to the invention can be used to produce sense and / or antisense oligonucleotide primers, which hybridize under conditions of high stringency to at least one of the sequences SEQ ID NO: 1 , SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, an analogue or fragment thereof. The expression of a nucleic acid sequence may also be determined, indirectly, by determining the expression of the amino acid sequence encoded by said sequence, by means of any technique known in the art, such as Western blot, ELISA, BRADFORD or LOWRY method, or as indicated below. The qualitative or quantitative measurement, expression, maturation, or activity of an amino acid sequence of the invention may be performed by any method known to those skilled in the art.

As methods for detecting the expression, the maturation, or the activity of an amino acid sequence, mention may be made of Western blot, blot-blot, dot-blot, Single or multiplexed Enzyme Linked Immuno-Sorbent Assay (ELISA) methods, proteomics or glycomic methods, staining polypeptides in a silver-based dye polyacrylamide gel, Coomassie blue or SYPRO, immunofluorescence, UV absorption, immunohistochemical methods in classical electron or confocal microscopy, FRET (fluorescence resonance energy transfer), TR-FRET methods (time resolved FRET / FRET in time resolution), fluorescence lifetime imaging microscopy (FLIM) methods, fluorescence spectral imaging microscopy (FSPIM) methods, and spectroscopic imaging. uorescence), methods of FRAP (fluorescence recovery after photobleaching), reporter gene methods, AFM (atomic force microscopy) methods, surface plasmon resonance methods, microcalorimetry methods, flow cytometry methods, biosensor methods, radioimmunoassay (RIA) methods, isoelectric focusing methods, and enzymatic assays, methods using peptide chips, chips sugar, antibody chips, mass spectrometry methods, SELDI-TOF spectrometry methods (Ciphergen). More generally, immunoenzymatic assay methods from more quantitative and sensitive protein solutions can in particular be used. These ELISA methods combine couples of capture antibodies and specific detection of the targeted antigen. Specifically developed commercial or polyclonal, monoclonal or recombinant antibodies can be used. High capacity multiplexed ELISA techniques can also be implemented. It is thus possible to mention the Luminex antibody-type multiplexed approach (for example Bioplex from Bio-Rad), of the antibody-on-flat surface type ("antibodies-arrays") (for example, an approach proposed by MesoScale Discovery). One can also mention the microfluidic cartridge technology developed by Korean company NanoEntek. In a recent approach a new technique based on antibodies called "Quenchbodies" proposed by the Japanese company Ushio allows to use only one antibody. In particular, it may be advantageous to detect the expression of an amino acid sequence of the invention by means of an antibody, where appropriate in a labeled form. Such an antibody can be labeled with a directly detectable or detectable substance by reaction with another reagent. By "antibodies" is generally meant monoclonal or polyclonal antibodies, as well as immunoglobulin fragments capable of binding an antigen and which may be produced by any genetic engineering technique known to those skilled in the art. by enzymatic or chemical cleavage of intact antibody. An antibody that can be used as a skin condition assessment tool can be obtained by any method known to those skilled in the art, as described in "Antibodies: A Laboratory Manual". , Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY (1990). According to a preferred embodiment, it may be advantageous to detect the expression of an amino acid sequence of the invention by means of a differential proteomic method "iTRAQ". Such a method is known to those skilled in the art and can advantageously be implemented as described in the examples below. In particular, the term "activity" with respect to an amino acid sequence of the invention means an antimicrobial activity, an activity for stimulating cell survival, an activity for stimulating cell proliferation, or a activity of reducing or preventing aged skin and / or skin signs of aging, as indicated above. Such activity can be determined by any method known to those skilled in the art, such as, for example, by evaluating the proliferation or survival of epidermal cells in cultures, such as keratinocytes, or by evaluating the activity. antimicrobial on cultured bacteria, such as Staphylococcus aureus, Escherichia colt or P. acnes. Preferably, the determination of a state of the skin or the characterization of the effectiveness of a cosmetic treatment of the skin can be carried out by measuring the variation of the expression of an amino acid sequence of the skin. invention, and preferably represented by a sequence selected from SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18, SEQ ID NO: 19 or SEQ ID NO: 20, an analogue or a fragment thereof. this.

The methods of the invention are particularly advantageous in that their implementation does not require the use of an invasive technique. A small epidermis ilium: thus be obtained by stripping techniques and directly analyzed by a conventicumeik analysis technique entlue man of art These strlappings are tacky surfaces applied to the epidermis surface as 3M Blendere, the commercial squam of the squam. CuDERM). eyanoarrylate glue or varnished stripping method, Thanks to these "strippings", eerneocytes: adherents and the content: - 1s spaces: intercellular can be pre-treated 33llinis thereafter has an extfacticihnermettant to have accee in the case of protein. The sampling of a sample which is a result of a process of the invention can also be carried out directly on the skin surface, for example by means of a suspension. of the type described above: spiral as described in patent FR 2 6 7 778 associated with a fluidic circuit, or simply by the addition of a drop, of. buffer on the surface of the skin. For information. other methods of harvesting may be mentioned, such as methods of scraping by means of a secondary system by means of a bi-series system. The technique allows for the collection of squares, which can be analyzed directly by different techniques for determining the levels of minerals: aix, amino acids or xylidene. Advantageously, one of the markers of the invention can be put into print for pre-selection, more efficient and more rigorous selection of individuals; - in view: - to evaluate the efficacy of treatment or aedfcosmetics for the care of the skin. Also, a momarqueur of the invention can at once be set to indecedernate to improve the effectiveness of an age, tif, al ej; Viro or Eizaleinent, a biomarker of the invention can be used to establish a personalized advice of a cosmetic treatment to induce an action of sound. - protil expre.ssl) n of shared subscribers. 3 () According to yet another of its objects, the present invention relates to the use (i) of at least one amino acid sequence of the invention, or (ii) a nucleic acid sequence of invention, for screening for active agents capable of modulating the activity, expression or maturation of said amino acid sequence or said nucleic acid sequence. For the purposes of the invention, the term "expression" with respect to an amino acid sequence, for example a protein or a peptide, or a nucleic acid sequence, for example an mRNA, its content or the variation of its content with respect to a reference. For the purposes of the invention, the term "maturation" with respect to an amino acid sequence, for example a protein or a peptide, or a nucleic acid sequence, for example an mRNA, the modifications who follow their synthesis in a cellular environment. For example, in the case of an amino acid sequence, "processing" is understood to mean post-translational modifications, such as the glycosylation or farnesylation of certain amino acids, or the proteolytic steps leading to the elimination of so-called "signal" or "secretory" sequences or the release of sequences with particular biological properties. In the case of a nucleic acid sequence, the term "maturation" is understood to mean, for example, the alternative splicing of an mRNA. Within the meaning of the invention, the term "activity", with respect to an amino acid sequence, for example a protein or a peptide, the biological activity of the amino acid sequence, if appropriate after processing, such as an enzymatic activity, an agonist or antagonist activity with respect to a receptor, or an activator or an inhibitor of an enzyme, a so-called "structure" activity, or antimicrobial activity. Within the meaning of the invention, the term "activity" refers to a nucleic acid sequence, for example an mRNA, its translation. According to another of its objects, the present invention relates to the use (i) of at least one amino acid sequence of the invention, or (ii) at least one nucleic acid sequence of the invention, to characterize the effectiveness of a treatment of acne-prone or acne-prone skin.

According to another advantage, the present invention makes it possible to propose novel active agents that are suitable for the prevention and / or treatment of acne-prone or acne-prone skin.

Screening There is still a need for new tools for screening active agents that are suitable for skin care, and more particularly useful for preventing and / or treating acne-prone or acne-prone skin. The object of the present invention is to satisfy these needs.

According to one of its aspects, the present invention relates to the use of an amino acid sequence, of nucleic acids of the invention, for screening or in a screening method, in particular in vitro or ex vivo, of active agents, particularly suitable for skin care.

The screened active agents may in particular be suitable for the care of acne-prone skin, and / or for the prevention and / or treatment of acne and imperfections. A use or a method of the invention may comprise comparing a measurement of the activity, expression or processing of an amino acid sequence or nucleic acid sequence according to the invention. invention to a reference measurement. A reference measurement may be as previously defined. In particular, a reference measurement may be a quantitative or qualitative value relating to the expression, the maturation or the activity of said sequences determined in a sample in the absence of tested active agent. Thus, a reference measurement can be obtained by repeating the steps of a process of the invention, and in particular the steps a), b) and c) of a method of the invention as defined above, in the following way: absence of biological or chemical compounds to be tested. A comparison of the test measurement with a reference measurement, and an observation of a deviation or absence of deviation between the two measurements, makes it possible to derive information as to the effect of the tested active agent. The quantitative or qualitative determination of the expression, the maturation or the activity of an amino acid sequence or of a nucleic acid sequence of the invention may be carried out by any method known to man of art, and especially as described above. According to one embodiment, the screening of an active agent capable of modulating the activity of an amino acid sequence of the invention can be done by measuring the activity or the expression of a target molecule. belonging to the signaling or metabolism pathways in which said amino acid sequence may be involved, such as, for example, a reporter gene system. According to one embodiment, a method of the invention can be implemented in a cell-free system, i.e. in a system not comprising cells but reproducing cellular functions, or in an isolated cell sample. A method according to the invention can be carried out on an isolated cell sample, an acellular sample, an isolated amino acid sequence or an isolated nucleic acid sequence of the invention, obtained by cutaneous biopsy, from cells in culture, in particular from an epidermal model, or a non-invasive skin surface sample, in particular by stratum corneum tape-stripping or by simple washing, as previously described. Advantageously, as a cellular sample that is suitable for the invention, there may be mentioned a sample of keratinocytes or any other cell type of the skin expressing an amino acid sequence of the invention. In a preferred manner, the screening of an active agent can be carried out by measuring the variation of the expression, in the presence and in the absence of the screened active agent, of an amino acid sequence of the invention, and preferably represented by a sequence selected from SEQ ID NO: 16, SEQ ID NO: 17, or SEQ ID NO: 18, an analogue or a fragment thereof. The active agents resulting from a screening according to the invention can be advantageously used for cosmetic or therapeutic purposes, in particular for the care of the acne-prone skin or for the treatment of the acne epau. According to one embodiment, a The active agent of the invention may be a stimulating agent for the activity, expression or maturation of an amino acid sequence of the invention. As a modulating agent that can be screened according to a use or a process in accordance with the invention, mention may also be made of antibodies or interfering RNAs. For the purposes of the present invention, "a" must be understood, unless otherwise indicated, in the sense of "at least one".

The examples and figures below are presented for illustrative and not limiting of the invention. EXAMPLES Example 1 Determination of the inhibitory activity of Dermicidin on the growth of P. acnes. A micromethode was developed in microplate.

The principle of the assay is based on the contact of decreasing concentrations of antimicrobial peptide (DCD-1L) with an identical P.acnes inoculum in culture medium suitable for P. acnes growth. After incubation of the 48h microplate with the P. acnes strain, the Optical Density of the sample (620 nm) is measured and the results are expressed as a percentage of calculated growth relative to a growth control without antimicrobial peptide (negative control). . In addition, one also puts in contact with trichlosan in order to have a positive control. The minimum inhibitory concentration or MIC is the smallest concentration of antibiotic sufficient to inhibit, in the laboratory that is to say in vitro, the growth of a strain of bacteria (bacterial colony). The range of concentrations chosen ranged from 5 to 100 μg / mL. Each concentration was tested in triplicate. The inhibitory activity of the peptides was evaluated on the following organ: - P. acnes ATCC 6919 (Gram + bacterium) Name of the antimicrobial peptide Provider information CMI batch reference in p, g / mL P. recombinant human HBD2 cells Interchim (Serotec ) product code PHP161X, batch 221210 75 RNASE 7 Extraction Human Plv (foot horn)> 100 recombinant human S100A7 Interchim (Protera) folding verified by NMR batch EN00510- 23/24/25> 100 Lacritin Synthetized by 100 hybrogenics Ref: Hgx 2238v1 Dermcidine (DCD1L) SEQ ID NO: 20 Synthetized by ppt It is observed that the growth of P. acnes is strongly inhibited in the presence of a low concentration of the DCD1L peptide. EXAMPLE 2 Cosmetic Composition A cosmetic composition is prepared comprising the following ingredients (% by weight): Dermcidin antioxidant 1 0.05 40.0 0.30 isopropanol water preservative qs 100% I Example 3: pharmaceutical composition A therapeutic composition comprising the following ingredients is prepared: following ingredients (% by weight): Dermicidine 5.00 Isopropanol 40.00 Preservative 0.30 Water qs 100% Example 4: Cream for the care of the face A cosmetic composition is prepared comprising the following ingredients (% by weight): Arachidyl behenyl alcohol / arachidylglusoside 2.0 Isohexadecane 7.0 Dermicidine 5.00 Glycerin 2.0 Water qs 100% 25 BIBLIOGRAPHY Anderson, RK and WL Kenney (1987). "Effect of age on heat-activated sweat glans." Physiol. 63 (3): 1089-109. Baechle, D., Flad, T., et al. (2006). "Cathepsin D is present in human eccrine sweat and involved in the postsecretory processing of the antimicrobial peptide DCD-1L" J. Biol. Chem. 281 (9): 5406-5415. Cunningham, T.J., L. Hodge et al. (1998) J. Neurosci 18 (18): 7047-7060. Flad, T., Bogumil R. et al. (2002) "Detection of dermcidin-derived peptides in sweat by ProteinChip Technology. J. Immunol. Methods 270 (1): 53-62. Kenney, W.L. and S.R. Fowler (1988) "Methylcholine-activated eccrine sweat gland density and output as a function of age" J. Appl. Physiol. 65 (3): 1082-1086. Kyte et al., (1982) J. Mol. Biol., 157: 105. Mehul B. et al., (2000) "Identification and Cloning of a New Calmodulin-like Protein from Human Epidermis" J. Biol. Chem. 275 (17): 12841-12847 Moreira, D.F., B.E. Strauss et al. (2008) Genes up-and-down-regulated by dermcidin in breast cancer: a microarray analysis Genet. Mol. Res. 7 (3): 925-932. Niyonsaba, F. A. Suzuki et al. (2009) "The human antimicrobial peptide dermcidin activates normal human keratinocytes" Br. J. Dermatol. 160 (2): 243-249. Rieg, S.C., Garbe et al. (2004) "Dermcidin is constitutively produced by eccrine sweat gland and is not induced in epidermal cells under inflammatory skin conditions" Br. J. Dermatol. 151 (3): 534-539. Rieg S., Steffen II et al. (2005) "Deficiency of dermcidin-derived antimicrobial peptides in patients with atopic dermatitis correlates with an impaired innate defense of human skin in vivo" J. Immunol. 174 (12): 8003-8010. Sakurada K., T. Akutsu, et al., (2009) "Detection of dermcidin for sweat identification by real-time RT-PCR and ELISA" Forensic. Sci. Int. 2010 Jan. 30, 194 (1-3): 80-4. Epub 2009 Nov. 13.

Sambrook et al., 1989, Vol. I-III, Coldspring Harbor Laboratory, Coldspring Harbor Press, NY. Schittek, B, R. Hipfel et al. (2001): "Dermcidin: a novel human antibiotic secret peptide by sweat tassels" Nat. Immunol. 2 (12): 1133-11137.

Watchorn, T. M. N. Dowidar et al. (2005) "The chachectic mediator proteolysis inducing factor activates NF-kappaB and STAT3 in human kupffer cells and monocytes" Int. J. Oncol. 27 (4): 1105-1111. Wiese et al., (2007) "Protein labeling by iTRAQ: A new tool for quantitative mass spectrometry in proteome research" Proteomics 7 (3): 340-350 Zieske et al. (2006) "A perspective on the use of iTRAQ reagent technology for protein complex and profiling studies" J. Exp. Bot. 57 (7): 1051-1058.

Claims (1)

CLAIMS 1, frequency of amino acids or a composition containing it selected from F, Q ID No. 11 SEQ. Or a fragment thereof for the treatment of 1-1 skin with a tendency to stenic and / or the prevention of parathyroidism: d irupt-nreetions. pence according to claim 1, wherein said amino acid sequence selected from. DRY? ID NO: it to SEQ ID NO: 20 where a fragment thereof is selected. SEQ: D NO: 16, SEQ ID NO: 17, SEC) ID NO: 18, SEQ ID NO. SEQ ID NO: 20 Or a fragment thereof, Sequence according to any one of the preceding claims, characterized in that said amino acid sequence selected from SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID 15D. NO. 18, SEQ ID NO: 19, SEQ ID NO: 20 or a thwirtent thereof, is SEQ ID NO 20, an ANALOGUE QU a fragment thereof, 4. Sequence Scion Pune queiconee of the claims Pre.current . cairàctérisèe. in that said amine acid equivalence is from 0.00001% to 5% of the total weight of the composition by% of the total amount of the composition, and more particularly in an amount of 0.001% by weight. s 1, to the total weight of the ompostin.