WO2024241070A1 - Cosmetic composition comprising an endolysin and a polyvinyl alcohol - Google Patents

Abstract

The present invention relates to a composition, notably a cosmetic composition, comprising, in a physiologically acceptable medium, at least one endolysin, in particular an endolysin derived from a Staphylococcus aureus phage, and at least one polyvinyl alcohol. The invention also relates to the use of such a composition for preventing and/or treating acne and/or eczema in an individual in need thereof, and to a non-therapeutic cosmetic process for caring for keratin materials, in particular the skin, notably dry skin, comprising at least one step of topical application of such a composition to said keratin materials.

Description

: Cosmetic composition comprising an endolysin and a polyvinyl alcohol

Technical field

The present invention relates to a composition, notably a cosmetic composition, comprising, in a physiologically acceptable medium, at least one endolysin, in particular an endolysin derived from a Staphylococcus aureus phage, and at least one polyvinyl alcohol.

The invention also relates in particular to the use of a composition of the invention for preventing and/or treating acne and/or eczema in an individual in need thereof.

Finally, the invention relates to a non-therapeutic cosmetic process for caring for keratin materials, in particular the skin, comprising the topical application to these keratin materials of a composition according to the invention.

Prior art

Human skin is permanently populated by a multitude of different microorganisms (bacteria, yeasts and fungi). The resident microbial flora, which is essential to the good health of the skin, consists mainly of propionibacteria (Cutibacterium aeries)' , staphylococci (Staphylococcus epidermidis and Staphylococcus hominis), corynebacteria and streptococci, and also of a fungal flora composed mainly of Malassezia.

Certain dermatological disorders are usually due to the disruption of the ecological balance of the resident flora following a preponderant colonization of opportunistic microorganisms that are not beneficial to the skin, such as Staphylococcus aureus, which is known to be associated with atopic dermatitis (eczema), greasy or hyperseborrhoeic skin, and acne.

To counteract this excess colonization by these opportunistic microorganisms that are not beneficial to the skin, it is common practice to use broad-spectrum antimicrobials or bacteriostats. However, the use of these compounds poses the problem of non- specificity of action targeting both the undesirable opportunistic flora and the resident beneficial flora, and the problem of the risk of the appearance of bacterial resistance or imbalances by selecting resistant bacteria, and also of problems of skin tolerance (irritation, allergies, etc.).

There is thus a need to find novel compounds with good antimicrobial efficacy which do not have the abovementioned drawbacks, and which have good skin tolerance. It has thus been demonstrated that endolysins from Staphylococcus aureus phages (i.e. phages infecting Staphylococcus aureus) can specifically target S. aureus and lyse, and thus specifically destroy, this bacterium while preserving the resident skin flora (WO 2012/150858). However, it is well known that one of the main obstacles to the application of endolysins targeting Staphylococcus species is a problem of the stability of these proteins and/or of their enzymatic activity, notably the maintenance of this activity over time.

Indeed, it has notably been observed that the lytic action of these enzymes is significantly impaired by interactions with raw materials, in particular when the endolysin is introduced into formulations, notably cosmetic formulations.

There is thus a need for a film-forming and/or viscosity-controlling agent that can be used in compositions, notably cosmetic compositions, without degrading the antimicrobial activity of the endolysins also present in these compositions.

Moreover, there is a need for a film-forming and/or viscosity-controlling agent which can be used in compositions, notably cosmetic compositions, without degrading (i) the antimicrobial activity of the endolysins used and (ii) the specificity of these endolysins for the bacteria targeted, in particular in the present invention Staphylococcus aureus.

There is also a need for a film-forming and/or viscosity-controlling agent which can be used in compositions, notably cosmetic compositions:

- without degrading the antimicrobial activity of the endolysins used;

- while retaining the specificity of these endolysins for the targeted bacteria, in particular Staphylococcus aureus in the present invention; and

- while also affording good sensory properties for the composition.

Disclosure of the invention

The aim of the present invention is to solve at least one abovementioned technical problem. Indeed, the inventors have now discovered that a polyvinyl alcohol (PVA) polymer, compared with other film-forming and/or viscosity-controlling agents known in the field of formulation, in particular cosmetic formulation, proves to be capable of maintaining good destruction performance of an endolysin, in particular a Staphylococcus aureus endolysin, many months after preparation of the composition.

Summary of the invention As mentioned above, the present invention thus relates to a composition, notably a cosmetic composition, comprising, in a physiologically acceptable medium:

(i) at least one endolysin, in particular an endolysin derived from a Staphylococcus aureus phage; and

(ii) at least one polyvinyl alcohol (PVA).

As illustrated in the examples hereinbelow, the Applicant has discovered, surprisingly, that a composition according to the invention, comprising an endolysin derived from a Staphylococcus aureus phage and at least one polyvinyl alcohol polymer, advantageously makes it possible to maintain the S. aureus destruction performance of the endolysin, even after a long period of time (for example 6 months).

Thus, the present invention also relates to the use of a composition of the invention for preventing and/or treating acne and/or eczema in an individual in need thereof.

The invention moreover relates to a non-therapeutic cosmetic process for caring for keratin materials, in particular the skin, notably acne-prone skin, comprising the topical application to these keratin materials of a composition according to the invention.

Detailed description

The term “cosmetic” means a composition that is compatible with keratin materials, in particular the skin, mucous membranes and the integuments. The composition according to the invention is non-therapeutic.

The term “keratin materials” is intended to denote in particular the skin, mucous membranes, fibres, eyelashes and the integuments.

The term “the skin” means all the skin of the body, and preferably the skin of the face, the scalp, the neckline, the neck, the arms and forearms, the eyelids, around the mouth or behind the ears, the hollow of the elbow, the back of the knees, the hands, the wrists and the ankles, or even more preferably the skin of the face (in particular the forehead, the nose, the cheeks and the chin), the neckline and the neck.

A composition according to the invention comprises a physiologically acceptable medium, i.e. one which has a pleasant colour, odour and feel and which does not give rise to any unacceptable discomfort, i.e. tingling, tightness or redness, that is liable to discourage the user from applying this composition. Needless to say, a person skilled in the art will take care to choose a physiologically acceptable medium such that the advantageous properties of the endolysin(s) of the invention are not, or are not substantially, adversely affected. Thus, by way of illustration, a physiologically acceptable medium may consist mainly of water and/or of one or more water-miscible organic solvent(s). A physiologically acceptable medium according to the invention preferentially has a pH between 4 and 8, more particularly between 4.5 and 7.5. Thus, a composition according to the invention may comprise one or more pH adjusters, for instance arginine. According to a preferred embodiment, a composition according to the invention comprises at least one pH adjuster, in particular one pH adjuster, and in particular arginine.

As used herein, the terms “treat” and “treatment” mean the alleviation of the symptoms associated with a specific disorder or condition and/or the elimination of said symptoms and also the complete disappearance of the disorder or condition under consideration.

In the context of the present invention, the terms “prevent” and “prevention” denote the reduction to a lesser degree of the risk or probability of occurrence of a given phenomenon.

Endolysin

A composition, notably a cosmetic composition, according to the invention is first characterized in that it comprises at least one endolysin.

In the embodiments herein, the endolysin may be a native bacteriophage endolysin or a recombinant endolysin and may be any endolysin known to the persons skilled in the art.

Herein, the terms bacteriophage lysin, bacteriophage endolysin and endolysin are used interchangeably. An endolysin may be selected from the group of endolysins defined in

WO20 11/023702, WO2012/146738, W02003/082184, WO2010/011960,

WO2010/149795, WO2010/149792, WO20 12/094004, WO2011/023702,

WO201 1/065854, WO2011/076432, WO201 1/134998, WO2012/059545,

WO2012/085259, WO2012146738, WO2018/091707, Exebacase™ (Lysin CF-301); SAL200™ or Tonabacase ; Auresine™ (Sigma-Aldrich SAE0083), and Ectolysin™ P128, which are herein incorporated by reference in their entirety.

In the embodiments herein, the endolysin may be a Staphylococcus-specific endolysin, meaning that it will lyse Staphylococcus, such as Staphylococcus aureus, efficiently but does not substantially lyse other bacteria than Staphylococcus or Staphylococcus aureus. In some embodiments, the endolysin will lyse Staphylococcus aureus, but not Staphylococcus epidermidis.

Most native Staphylococcus bacteriophage endolysins exhibiting peptidoglycan hydrolase activity, such as the Ply2638 endolysin, consist of a C-terminal cell wall-binding domain (CBD), a central N-acetylmuramoyl-L-Alanine amidase domain, and an N-terminal Alanylglycyl endopeptidase domain with cysteine, histidine-dependent amidohydrolases/peptidase (CHAP) homology, or in case of Ply2638, of an N-terminal alanyl-glycine endopeptidase domain with peptidase_M23 homology, the latter three domains exhibiting peptidoglycan hydrolase activity each with distinct target bond specificity and generally named as enzymatically active domains.

The endolysin herein may be a recombinant endolysin, such as a recombinant

endolysin, in particular a recombinant Staphylococcus-s^QciUc chimeric endolysin comprising one or more heterologous domains.

In general, endolysins are comprised of different subunits (domains); e.g. a cell wall-binding domain (CBD) and one or more enzymatic domains having peptidoglycan activity, such as an amidase domain, a M23 domain and a CHAP (cysteine, histidine-dependent amidohydrolases/peptidases) domain. An example of a Staphylococcus-s^QciUc chimeric endolysin comprising one or more heterologous domains is an endolysin comprising an amidase domain of bacteriophage Ply2638, a M23 domain of lysostaphin (S. siniulans) and a cell wall-binding domain of bacteriophage Ply2638.

Such Staphylococcus-specific chimeric endolysin is a preferred endolysin and is extensively described in WO2012/150858, which is herein incorporated by reference in its entirety. Other preferred endolysins are extensively described in W02013/169104, which is herein incorporated by reference in its entirety. Other preferred endolysins are extensively described in WO2016/ 142445, which is herein incorporated by reference in its entirety. Other preferred endolysins according to the invention are extensively described in WO20 17/046021, which is herein incorporated by reference in its entirety.

In the embodiments herein, the endolysin may comprise a domain having at least 80% sequence identity with a domain depicted in WO2012/150858, W02013/169104, WO2016/142445, or WO2017/046021.

In the embodiments herein, the endolysin may have at least 80% sequence identity with an endolysin depicted in WO2012/150858, W02013/169104, WO2016/142445, W02017/046021, such as the endolysin with the amino acid as set forward in SEQ ID NO: 29 in WO2012/150858.

According to one particular embodiment, the endolysin is an endolysin derived from a Staphylococcus aureus phage.

For the purposes of the present invention, “endolysin derived from a Staphylococcus aureus phage” means a native or recombinant protein such as an enzyme or nucleic acid molecule encoding for same derived from one or more bacteriophage(s) that are capable of lysing the wall of bacteria of the species Staphylococcus aureus.

The endolysin notably comprises one or more domains for binding to the bacterial wall of Staphylococcus aureus and/or one or more domains for lysing the bacterial wall of Staphylococcus aureus, said binding domain(s) and lysis domain(s) for the Staphylococcus aureus bacterial wall being derived from one or more identical or different bacteriophage(s) that are capable of lysing the wall of bacteria of the species Staphylococcus aureus.

The endolysin used in the context of the present invention may be in native or recombinant form, in particular in recombinant form.

According to a particular embodiment, the endolysin comprises a first protein sequence comprising a domain for binding to the cell wall of species of the genus Staphylococcus.

In particular, the first protein sequence is derived from the endolysin of the 02638a bacteriophage of S. aureus.

For each of the amino acid or nucleic acid sequences of interest, reference sequences are described herein. The present description also encompasses amino acid or nucleic acid sequences (for example enzyme amino acid sequences), having specific percentages of amino acid or nucleotide identity with a reference sequence.

For obvious reasons, throughout the present description, a specific nucleic acid sequence or a specific amino acid sequence which respects, respectively, the nucleotide or amino acid identity under consideration, must also lead to the production of a protein (or enzyme) which displays the desired biological activity. As used herein, the “percentage identity” between two nucleic acid sequences or between two amino acid sequences is determined by comparing the two optimally aligned sequences through a comparison window.

The portion of the nucleotide or amino acid sequence in the comparison window may thus comprise additions or deletions (e.g. “gaps”) relative to the reference sequence (which does not comprise these additions or deletions) so as to achieve an optimal alignment between the two sequences.

The terms “sequence homology” or “sequence identity” or “homology” or “identity” are used interchangeably herein. For the purposes of the invention, this means that in order to determine the percentage of sequence homology or sequence identity of two amino acid sequences or two nucleic acid sequences, the sequences are aligned for optimal comparison. In order to optimize the alignment between the two sequences, gaps may be introduced in either of the two sequences being compared. This alignment may be performed over the entire length of the sequences being compared. The alignment may also be performed over a shorter length, for example over about twenty, fifty, one hundred or more nucleic acids/bases or amino acids. Sequence identity is the percentage of identical matches between the two sequences over the reported aligned region.

The comparison of sequences and the determination of the percentage of sequence identity between two sequences may be performed using a mathematical algorithm. A person skilled in the art knows that several different computer programs are available for aligning two sequences and determining the identity between two sequences (Kruskal, J.B. (1983) An overview of sequence comparison In D. Sankoff and J.B. Kruskal, (ed.), Time warps, string edits and macromolecules: the theory and practice of sequence comparison, pages 1-44 Addison Wesley).

The percentage of sequence identity between two amino acid sequences or between two nucleotide sequences may be determined using the Needleman-Wunsch algorithm for the alignment of two sequences. (Needleman, S.B. and Wunsch, C.D. (1970) J. Mol. Biol. 48, 443-453). The algorithm allows the alignment of both amino acid sequences and nucleotide sequences. The Needleman-Wunsch algorithm was implemented in the NEEDLE computer program.

For the purposes of the invention, the NEEDLE program of the EMBOSS software package was used (version 2.8.0 or higher, EMBOSS: The European Molecular Biology Open Software Suite (2000) Rice, P. Longden, J. and Bleasby, A. Trends in Genetics 16, (6) pages 276- 277, http://emboss.bioinformatics.nl/). For protein sequences, EBLOSUM62 is used for the substitution matrix. For nucleotide sequences, EDNAFULL is used. The optional parameters used are a space opening penalty of 10 and a space extension penalty of 0.5. No end gap penalty is added. In the Output section, Yes has been indicated in response to the question “Brief identity and similarity” and “SRS pairwise” has been indicated as the output alignment format.

After alignment by the NEEDLE program described above, the percentage of sequence identity between a query sequence and a sequence of the invention is calculated as follows: Number of matching positions in the alignment showing an identical amino acid or nucleotide in the two sequences divided by the total length of the alignment after subtracting the total number of gaps in the alignment. The identity defined here may be obtained from NEEDLE using the NOBRIEF option and is labelled in the output of the program as “longest identity”.

The similarity of nucleotide and amino acid sequences, i.e. the percentage identity of the sequences, may be determined by sequence alignments using several other known algorithms, preferably the mathematical algorithm of Karlin and Altschul (Karlin & Altschul (1993) Proc. Natl. Acad. Sci. USA 90: 5873-5877), with hmmalign (HMMER package, http://hmmer.wustl.edu/) or with the CLUSTAL algorithm (Thompson, J.D., Higgins, D.G. & Gibson, T.J. (1994) Nucleic Acids Res. 22, 4673-80) available, for example, at https://www.ebi.ac.uk/Tools/msa/clustalo/ or the GAP program (mathematical algorithm of the University of Iowa) or the mathematical algorithm of Myers and Miller (1989 - Cabios 4: 11-17) or Clone Manager 9. The preferred parameters used are the default parameters as defined at https://www.ebi.ac.uk/Tools/msa/clustalo/.

The degree of sequence identity (sequence matching) may be calculated using, for example, BLAST, BLAT or BlastZ (or BlastX). A similar algorithm is incorporated in the BLASTN and BLASTP programs of Altschul et al. (1990) J. Mol. Biol., 215, 403-410. BLAST polynucleotide searches are performed with the BLASTN program, score = 100, word length = 12, so as to obtain polynucleotide sequences homologous to the nucleic acids which encode the protein of interest.

BLAST protein searches are performed with the BLASTP program, score = 50, word length = 3, to obtain amino acid sequences homologous to the SHC polypeptide. To obtain gapped alignments for comparison, Gapped BLAST is used as described in Altschul et al. (1997) Nucleic Acids Res. 25, 3389-3402. When using the BLAST and Gapped BLAST programs, the default settings of the respective programs are used. Sequence matching analysis may be complemented by established homology mapping techniques such as Shuffle-LAGAN (Brudno M., Bioinformatics 2003b, 19 Suppl. 1 : 154-162) or Markov random fields. Where reference is made to percentages of sequence identity in the present patent application, these percentages are calculated relative to the total length of the longest sequence, unless otherwise indicated.

In particular embodiments, the percentage identity between two sequences is determined using CLUSTAL O (version 1.2.4).

Thus, according to a particular embodiment, the first protein sequence comprises a protein sequence comprising at least 80%, in particular at least 90%, more particularly at least 95% sequence identity with the reference amino acid sequence SEQ ID NO: 1.

The term “at least 80% sequence identity between two sequences” means that the first sequence may comprise 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the second sequence, whether they are amino acid sequences or nucleic acid sequences.

In particular, the first protein sequence consists of an amino acid sequence of reference SEQ ID NO: 1.

The protein sequences described herein may be encoded by one or more allelic variants.

An allelic variant denotes any one of two or more alternative forms of a gene occupying the same chromosomal locus. A preferred nucleic acid variant is a nucleotide sequence that contains one or more silent mutations. Alternatively or in combination, a nucleic acid variant may also be obtained by introducing nucleotide substitutions, which do not result in another amino acid sequence of the polypeptide encoded by the nucleotide sequence, but which correspond to the use of codons of the host organism intended for the production of the polypeptide of the invention. According to a preferred embodiment, a nucleic acid variant encodes a polypeptide still having its biological function. More preferably, a nucleotide sequence variant encodes a polypeptide displaying binding to the cell wall of species of the genus Staphylococcus and/or lytic activity. Even more preferably, a nucleic acid variant encodes a polypeptide displaying increased binding to the cell wall of species of the genus Staphylococcus and/or lytic activity, as defined hereinbelow. Nucleic acids encoding a polypeptide displaying binding to the cell wall of species of the genus Staphylococcus and/or lytic activity may be isolated from any microorganism.

All these variants may be obtained using techniques known to those skilled in the art, such as library screening by hybridization (Southern blot procedures) under low to medium to high hybridization conditions. Low to medium to high stringency conditions means prehybridization and hybridization at 42°C in 5X SSPE, 0.3% SDS, 200 pg/ml sheared and denatured salmon sperm DNA, and either 25%, 35% or 50% formamide for low to medium to high stringencies, respectively. Next, the hybridization reaction is washed three times for 30 minutes using for each wash 2XSSC, 0.2% SDS and at 55°C, 65°C or 75°C for low to medium to high stringencies.

According to a particular embodiment, the first protein sequence is encoded by a nucleic acid sequence comprising at least 80%, in particular at least 90%, more particularly at least 95% sequence identity with the reference nucleic acid sequence SEQ ID NO: 2.

In particular, the first protein sequence is encoded by a nucleic acid sequence consisting of a reference nucleic acid sequence SEQ ID NO: 2.

The binding of a domain to the peptidoglycan cell wall of Staphylococcus genera may be evaluated using assays that are well known to those skilled in the art. In a preferred embodiment, an immunohistochemical technique and/or a gene fusion technique resulting in labelled constructs of either domain are used to evaluate the specific binding of peptides, polypeptides or proteins to the peptidoglycan cell wall of Staphylococcus genera. The signal quantification processes used in the abovementioned immunohistochemical or fusion techniques are well known in the art.

In one embodiment, the binding to the peptidoglycan cell wall of Staphylococcus may be quantified using a fluorescent fusion construct comprising a polypeptide comprising a domain included in a first protein sequence as described previously. Such a cell wall binding assay is described in detail by Loessner et al. (Molecular Microbiology 2002, 44(2): 335- 349). In said assay, a solution comprising said fluorescent fusion construct or a negative control, preferably green fluorescent protein (GFP), is subjected to Staphylococcus cells, preferably S. aureus cells, more preferably S. aureus BB255, for a specified period of time, after which the cells are sedimented by centrifugation together with the bound fluorescent fusion constructs. The fluorescent signal of Staphylococcus cells exposed to a fluorescent fusion construct, subtracted from the fluorescent signal of Staphylococcus cells exposed to a negative control, preferably GPF, is a measure of cell binding for the purposes of the present invention.

Examples of evaluation of the binding of an endolysin to the cell wall of species of the genus Staphylococcus suitable according to the invention are notably illustrated in WO 2012/150858 Al.

Preferably, in the context of the present text, a protein sequence will be said to comprise a domain for binding to the peptidoglycan cell wall of Staphylococcus genera when, using this assay, an increase in the fluorescent signal of the sedimented cells is detected. The binding is preferably said to be specific. Preferably, an endolysin is described comprising a domain which displays a binding capacity, as defined herein, of at least 50, 60, 70, 80, 90 or 100, 150 or 200% of the peptidoglycan cell wall binding of the O2638a bacteriophage endolysin of S. aureus (Ply2638) encoded by the reference nucleic acid sequence SEQ ID NO: 5.

According to a particular embodiment, the binding activity to the cell wall of species of the genus Staphylococcus is measured by an immunohistochemical technique and/or a gene fusion technique, in particular a fluorescent fusion technique, more particularly fusion with a green fluorescent protein.

According to a particular embodiment, the endolysin comprises a protein sequence comprising at least 80%, in particular at least 90%, more particularly at least 95% sequence identity with an amino acid sequence chosen from the group consisting of the amino acid sequences of references SEQ ID NO: 3 and SEQ ID NO: 4.

According to a particular embodiment, the endolysin comprises a protein sequence encoded by a nucleic acid sequence comprising at least 80%, in particular at least 90%, more particularly at least 95% sequence identity with the reference nucleic acid sequence SEQ ID NO: 5.

In particular, the protein sequence may be encoded by a nucleic acid sequence consisting of a reference nucleic acid sequence SEQ ID NO: 5. According to a particular embodiment, the endolysin may also comprise a heterologous protein sequence.

The term “heterologous protein sequence” is intended to denote a protein sequence, i.e. an amino acid sequence or a nucleic acid sequence encoding the protein sequence, which is not naturally functionally linked as a neighbouring sequence to said first protein sequence. As used herein, the term “heterologous” may mean “recombinant”. The term “recombinant” refers to a genetic entity different from that generally found in nature. When applied to a nucleotide sequence or nucleic acid molecule, this means that said nucleotide sequence or nucleic acid molecule is the product of various combinations of cloning, restriction and/or ligation steps, and other procedures that result in the production of a construct that is different from a sequence or molecule found in nature.

Such protein or nucleic acid recombination methods are well known to those skilled in the art.

According to a particular embodiment, the endolysin comprises a heterologous protein sequence comprising a lytic domain. Preferably, said lytic domain shows a peptidoglycan hydrolase activity.

“Peptidoglycan hydrolase activity”, also defined herein as “lytic activity”, may be evaluated via processes that are well known to those skilled in the art. In one embodiment, the lytic activity may be evaluated spectrophotometrically by measuring the decrease in turbidity of substrate cell suspensions. Preferably, the lytic activity may be evaluated spectrophotometrically by measuring the decrease in turbidity of a suspension of S. aureus, the turbidity being quantified by measuring the OD595 spectrophotometrically (Libra S22, Biochrom). More preferably, 200 nM of a polypeptide encoded by a nucleic acid molecule as identified herein are incubated with a suspension of S. aureus having an initial ODeoo of 1 ± 0.05, as evaluated spectrophotometrically (Libra S22, Biochrom), in PBS buffer pH 7.4, 120 mM sodium chloride for 30 min at 37°C. The decrease in turbidity is calculated by subtracting the OD595 after 30 min of incubation from the OD595 before 30 min of incubation. In the context of the present text, a protein sequence will be said to comprise a lytic domain when, using this assay, a decrease in turbidity of at least 10, 20, 30, 40, 50 or 60% is detected. Preferably, a decrease of at least 70% is detected. Preferably, an endolysin is described comprising a domain which shows lytic activity of at least 50, 60, 70, 80, 90, 100, 150 or 200% or more of a lytic activity of the 02638a bacteriophage endolysin of S. aureus (Ply2638) encoded by the reference nucleic acid sequence SEQ ID NO: 5.

According to a particular embodiment, the lytic activity of the endolysin is measured spectrophotometrically by measuring the decrease in turbidity of a suspension of S. aureus.

In one embodiment, the endolysin is possibly not encoded by an amino acid sequence comprising or consisting of an amino acid sequence chosen from the group consisting of the amino acid sequences of references SEQ ID NO: 3 and SEQ ID NO: 4.

In particular, the endolysin is possibly not encoded by a nucleic acid sequence comprising or consisting of the reference nucleic acid sequence SEQ ID NO: 5, encoding the 02638 bacteriophage endolysin of S. aureus.

According to a particular embodiment, the heterologous protein sequence comprises a lytic domain, said lytic domain comprising a second and a third protein sequence, said second protein sequence comprising an endopeptidase M23 domain and said third protein sequence comprising an amidase domain.

An endopeptidase domain as used herein preferably cleaves pentaglycine cross-bridges (Trayer, H.R. and Buckley, C.E. (1970) Molecular properties of lysostaphin, a specific bacteriolytic agent for Staphylococcus aureus. J. Biol. Chem. 245, 4842-4846) which are found in the cell wall of Staphylococcus genera, preferably in the cell wall of S. aureus, S. simulans and S. carnosus.

An amidase domain as used herein preferably hydrolyses substrates containing gammaglutamyl.

The functionality and activity of such domains in a polypeptide may be confirmed by characterizing the cleavage products upon incubation of said polypeptides containing any of these domains with a purified peptidoglycan. According to a particular embodiment, the endopeptidase and/or amidase activity of the endolysin is measured by characterizing the cleavage products.

According to a particular embodiment, the endopeptidase and/or amidase activity of the endolysin may be measured by measuring the optical density of bacteria in the presence of the endolysin. Such methods are notably described in Park et al. (Characterization of an endolysin, LysBPS13, from a Bacillus cereus bacteriophage, FEMS Microbiol. Lett. 2012 Jul.; 332(1): 76-83) and in Grishin et al. (A Simple Protocol for the Determination of Lysostaphin Enzymatic Activity, Antibiotics (Basle). 2020 Dec. 17; 9(12): 917).

Preferably, each of the protein sequences and nucleotide sequences encoding the second or third domain is of bacterial or bacteriophage origin.

According to a particular embodiment, said second and third protein sequences are derived, independently of each other, from an enzyme chosen from the group consisting of the endolysin from O2638a bacteriophage from S. aureus and the lysostaphin from S. simulans. In particular, one of the second and third protein sequences is derived from the endolysin of 02638a bacteriophage of S. aureus and the other sequence of the second and third protein sequences is derived from the lysostaphin of S. simulans.

According to a particular embodiment, said second protein sequence comprises at least 80%, in particular at least 90%, more particularly at least 95% sequence identity with the reference amino acid sequence SEQ ID NO: 6 and said third protein sequence comprises at least 80%, in particular 90%, more particularly 95% sequence identity with the reference amino acid sequence SEQ ID NO: 8.

According to a particular embodiment, said second protein sequence is encoded by a nucleic acid sequence comprising at least 80%, in particular at least 90%, more particularly at least 95% sequence identity with the reference nucleic acid sequence SEQ ID NO: 7 and said third protein sequence is encoded by a nucleic acid sequence comprising at least 80%, in particular at least 90%, more particularly at least 95% sequence identity with the reference nucleic acid sequence SEQ ID NO: 9.

According to a particular embodiment, said second protein sequence is encoded by a nucleic acid sequence consisting of the reference nucleic acid sequence SEQ ID NO: 7 and said third protein sequence is encoded by a nucleic acid sequence consisting of the reference nucleic acid sequence SEQ ID NO: 9.

According to a particular embodiment, the endolysin comprises a protein sequence comprising at least 80%, in particular at least 90%, more particularly at least 95% sequence identity with an amino acid sequence chosen from the group consisting of the amino acid sequences of references SEQ ID NO: 10, and SEQ ID NO: 11.

In particular the endolysin may comprise a protein sequence consisting of the reference amino acid sequence SEQ ID NO: 10.

According to a particular embodiment, the endolysin comprises a protein sequence encoded by a nucleic acid sequence comprising at least 80%, in particular at least 90%, more particularly at least 95% sequence identity with a reference nucleic acid sequence SEQ ID NO: 12. In particular, the endolysin may comprise a protein sequence encoded by a nucleic acid sequence consisting of a reference nucleic acid sequence SEQ ID NO: 12.

An endolysin comprising a protein sequence encoded by a reference nucleic acid sequence SEQ ID NO: 12 differs from the 02638a bacteriophage endolysin of S. aureus in that the N-terminal M23 endopeptidase domain is substituted with an M23 endopeptidase domain from the lysostaphin of S. simulans.

Endolysins that are suitable for use in the invention may be obtained via any method known to those skilled in the art for producing recombinant proteins. In particular, endolysins according to the invention may be obtained by introducing one or more gene(s) of interest, such as the nucleic acid sequences described previously, into the genome of a host organism via a vector.

In another aspect, a nucleic acid construct comprising at least one of the nucleic acid sequences as defined previously is described. This nucleic acid construct may comprise a first nucleic acid sequence encoding a polypeptide comprising a cell wall binding domain, also possibly comprising a second and third nucleic acid sequence as defined previously.

Also described is an expression vector comprising such a nucleic acid construct. Preferably, an expression vector comprises a nucleotide sequence as mentioned previously, which is operatively linked to one or more control sequences, which direct the production or expression of the encoded polypeptide in a cell, a subject or a cell-free expression system. An expression vector may be considered a recombinant expression vector. This vector may consist of a plasmid, cosmid, bacteriophage or virus which is transformed by the introduction of a nucleic acid molecule according to the invention. Such transformation vectors according to the host organism to be transformed are well known to those skilled in the art and widely described in the literature.

Another subject described herein is a process for transforming host organisms by integration of at least one nucleic acid sequence as described, which transformation may be performed via any suitable means known and widely described in the specialized literature, more particularly via the vector described above.

In another aspect, a cell is described, which comprises a nucleic acid construct or an expression vector as defined previously. A cell may be any microbial, prokaryotic or eukaryotic cell, which is suitable for expressing an endolysin that is suitable for use in the invention. In a preferred embodiment, said cell is an E. coli cell. In an even more preferred embodiment, said cell is E. coli CLlblue MRF.

The endolysin obtained may then be purified according to purification methods known in the art such as column chromatography, high performance liquid chromatography, etc.

In a particular embodiment, one or more of the protein sequences as defined in the text may comprise a sequence encoding a tag to facilitate the purification of the resulting endolysin. Preferably, said tag is chosen from, but is not limited to, a group consisting of a FLAG tag, a poly(His) tag, an HA tag and an Myc tag. More preferably, said tag is a 6xHis tag. Even more preferably, said tag is an N-terminal 6xHis tag identical to SEQ ID NO: 13.

Endolysins that are suitable for use according to the invention and also methods for producing them are notably described in patent application WO 2012/150858 Al.

An endolysin derived from a Staphylococcus aureus phage that is suitable for use according to the invention may be present in the composition in freshly prepared form or in lyophilized form. In the present text, the term “freshly prepared” is preferably defined as storage for no more than 2 days after production at 1.63 mg/mL in lyophilization buffer (50 mM Tris, 500 mM sucrose, 200 mM mannitol, 0.05% polysorbate 20 + 50% glycerol) at -20°C followed by thawing immediately prior to evaluating the lytic activity in an assay as identified herein. The term “lyophilized” refers to an endolysin that has been dehydrated by lyophilization, which consists in freezing the protein and then dehydrating it to remove the water.

After lyophilization, an endolysin in freeze-dried form may undergo a subsequent reconstitution step by addition of water. In one embodiment, the lyophilization and reconstitution may be performed by dialysis against three changes of 300 ml lyophilization buffer (50 mM phosphate or Tris, 500 mM sucrose, 200 mM mannitol, pH 7.4) aliquot and freezing in the gas phase of liquid nitrogen. The lyophilization may be performed under standard conditions, preferably at -40°C and under vacuum at 75 mTorr for 60 minutes, before increasing the temperature over 5 hours to -10°C and further increasing over 60 minutes to -10°C at the same vacuum levels. As a final step, the temperature is preferably increased to 25 °C over 10 hours. The samples are reconstituted by adding water.

A composition according to the invention may comprise a content of endolysin(s), in particular endolysin(s) derived from a Staphylococcus aureus phage, ranging from 0.0001% to 0.1 % by weight relative to the total weight of the composition, in particular from 0.0005% to 0.01% by weight relative to the total weight of the composition, more particularly from 0.001% to 0.005% by weight relative to the total weight of the composition.

Polymer of polyvinyl alcohol type

A composition according to the invention comprises at least one polyvinyl alcohol (PVA). In particular, a PVA of a composition according to the invention includes at least units of formula (I):

[Formula I]

Said polyvinyl alcohol may be formed by polymerization of one or more vinyl ester monomers by mass polymerization, solution polymerization, emulsion polymerization, and suspension polymerization. The vinyl ester monomers may comprise various aliphatic acids, such as vinyl formate, vinyl acetate, vinyl butyrate, vinyl pivalate, and vinyl versatate. The vinyl ester polymer thus obtained may be saponified to form a polyvinyl alcohol. The saponification (or hydrolysis) reaction may be total or partial.

Preferably, the polyvinyl alcohol used in the context of the present invention has a degree of hydrolysis ranging from 65% to 99%, more preferentially ranging from 75% to 95%, better still ranging from 87% to 89%.

Preferably, the polyvinyl alcohol used in the context of the present invention has a degree of polymerization ranging from 100 to 2500, more preferentially ranging from 200 to 1000, better still ranging from 300 to 600.

According to a particular embodiment, the polyvinyl alcohol has a mass-average molecular weight from 1000 to 1 000 000 g/mol, preferably from 10 000 to 500 000 g/mol and even more preferentially from 50 000 to 400 000 g/mol.

The PVA may advantageously be present in a composition according to the invention in a content which is usual for a cosmetic composition, in particular in a content which is usual for it to play its role as a film-forming and viscosity-controlling agent in a composition according to the invention, in particular in a cosmetic composition according to the invention, more particularly in a content which is usual for it to play its role as a film- forming and viscosity-controlling agent when it is the sole film-forming and/or viscosity-controlling agent of a composition according to the invention.

A composition according to the invention may comprise a polyvinyl alcohol content ranging from 0.1% to 20% by weight relative to the total weight of the composition, in particular from 0.5% to 10%, even more preferentially from 1% to 5% by weight relative to the total weight of the composition.

A composition according to the invention may comprise one or more endolysins according to the invention and one or more polyvinyl alcohols in a mass ratio of endolysin(s)/PVA of between 0.000 land 0.1 and in particular between 0.001 and 0.01.

A composition according to the invention may comprise a single type of polyvinyl alcohol or at least two or more polyvinyl alcohols.

Examples of polyvinyl alcohols that are suitable for use according to the invention are those sold under the names “Selvol® Ultalux FP” (a polyvinyl alcohol having a degree of polymerization of about 300-600 and a degree of hydrolysis of about 87-89), “Selvol® Ultalux FA” (a polyvinyl alcohol having a degree of polymerization of about 900-1500 and a degree of hydrolysis of about 87-89), “Selvol® Ultalux FF” (a polyvinyl alcohol having a degree of polymerization of about 1500-2500 and a degree of hydrolysis of about 87-89) and “SELVOL® 107” (a polyvinyl alcohol having a degree of polymerization of about 350-700 and a degree of hydrolysis of about 98-99) by the company Sekisui Specialty Chemicals®.

Additional ingredients

In addition to the abovementioned compounds, a composition according to the invention may of course comprise one or more additional ingredient(s), other than a polyvinyl alcohol. It goes without saying that a person skilled in the art will take care to choose this or these additional ingredient(s) such that the advantageous properties of the endolysin(s) of the invention are not, or are not substantially, adversely affected.

The additional ingredients are present in the compositions in a content which is usual for each of them in a cosmetic composition, in particular in a content which is usual for each of them and which enables them to retain its cosmetic properties, more particularly in a content which is usual for each of them and which enables them, when they are each the only ingredient of a composition according to the invention having this property, to retain its cosmetic property.

Thus, a composition according to the invention may also comprise one or more of the ingredients, other than a PVA, chosen from surfactants ; fatty substances; colorants; preserving agents; fragrances; pH adjusters such as organic acids such as citric acid; antioxidants; hydrophilic gelling agents such as hydroxypropylmethylcellulose; amino acids such as arginine; carbohydrates; chelating agents; sugar alcohols; cosmetic active agents; and mixtures thereof.

According to a particular embodiment, a composition according to the invention comprises less than 2% by weight, in particular less than 1% by weight, of hydroxypropylmethylcellulose relative to the total weight of the composition; more particularly, a composition according to the invention is free of hydroxypropylmethylcellulose. A composition according to the invention may be in any presentation form normally used in the cosmetics field.

It may notably be in the form of an aqueous or aqueous-alcoholic solution, which may be gelled, a dispersion of the lotion type, which may be a two-phase dispersion, an oil-in-water or water-in-oil emulsion or a multiple emulsion, a gel, notably an aqueous gel, or even a dispersion of oils in an aqueous phase, notably using spherules, it being possible for these spherules to be polymeric particles or, better still, lipid vesicles of ionic and/or nonionic type. In particular, a composition according to the invention may be in the form of a gel, notably an aqueous gel. It may also be an anhydrous composition. The term “anhydrous composition” means a composition containing less than 10% by weight of water, in particular less than 5% by weight of water, more particularly less than 2% by weight of water or even less than 0.5% by weight of water, and notably free of water, the water not being added during the preparation of the composition but corresponding to the residual water provided by the mixed ingredients. The composition may be of more or less fluid liquid consistency.

A composition according to the invention is preferentially suitable for topical administration. Thus, a composition according to the invention may comprise all the constituents usually employed in the envisaged topical application and administration.

A composition according to the invention may advantageously be in the form of an emulsion, notably obtained by dispersion of an aqueous phase in a fatty phase (W/O) or of a fatty phase in an aqueous phase (O/W), of liquid or semi-liquid consistency of the milk type, or of soft consistency, or even of multiple emulsion (W/O/W or O/W/O). These compositions are prepared according to the usual known methods.

More particularly, a composition according to the invention may be intended for topical application and may preferably be in the form of an emulsion, preferably an oil-in-water emulsion. Preferably, such an emulsion is not intended to be rinsed off after application.

A composition according to the invention is preferentially intended to be applied to a skin. Preferably, the skin is the skin of the face, scalp, neckline, neck, arms or forearms, or even more preferably the skin of the face (in particular of the forehead, nose, cheeks, chin), neckline and neck. The composition may alternatively be in the form of a face and/or body care or makeup product, and may be packaged, for example, in the form of a cream in a jar or a fluid in a tube or a pump bottle or a dropper bottle.

The composition according to the invention may be manufactured via any known process generally used in the cosmetics field.

The ingredients are mixed before forming, in the order and under conditions readily determined by a person skilled in the art.

According to a particular mode of the invention, other agents intended to embellish the appearance and/or texture of the skin may also be added to the composition according to the invention.

Uses and processes

According to one of its aspects, the present invention relates to the cosmetic use, notably the topical use, of a composition according to the invention for preventing and/or treating acne and/or eczema in an individual in need thereof.

According to yet another one of its aspects, the present invention relates to a non-therapeutic cosmetic process for caring for keratin materials, in particular the skin, notably acne-prone skin, comprising the topical application to these keratin materials of a composition according to the invention.

A skin may in particular be a skin presenting acne or at risk of presenting acne and/or a skin presenting eczema or at risk of presenting eczema.

The cosmetic uses and processes considered according to the invention are non-therapeutic. The cosmetic uses and processes of the invention are preferentially performed by topically administering a composition according to the invention. As such, the composition of the cosmetic use is in particular suitable for topical administration.

Topical administration consists of the external application to the skin of cosmetic compositions according to the usual techniques for the use of these compositions.

By way of illustration, the cosmetic use or process according to the invention may be performed by topical, for example daily, application of at least one composition according to the invention, which may be formulated, for example, in the form of a cream, gel, serum, lotion, emulsion or cleansing milk, in particular in gel form. The application may be repeated for example 1 to 2 times a day over a day or more and generally over a prolonged period of at least 3 days, at least 1 week, at least 4 weeks, or even 4 to 15 weeks, with, if appropriate, one or more interruption periods.

According to one embodiment, the application is daily (once a day) and generally over an extended period of at least 3 days, at least 1 week, at least 4 weeks or even 4 to 15 weeks, with one or more interruption periods if necessary.

According to one embodiment, the cosmetic treatment process according to the invention may comprise a single application.

Throughout the description, including the claims, the terms “between ... and ...”, and “ranging from ... to ...” should be understood as meaning limits included, unless otherwise specified.

The examples that follow illustrate the present invention without limiting the scope thereof. In the examples, unless otherwise specified, the temperature is room temperature (20°C) and is expressed in degrees Celsius, and the pressure is atmospheric pressure.

Examples

Protocol (experimental conditions)

Preparation of the working culture of Staphylococcus aureus

Preparation of the working suspension of Staphylococcus aureus (S. aureus from a lyophilizate ATCC 6538 (according to the recommendations of the standard NF EN 12353). The lyophilizate is rehydrated in Trypticase soy broth, plated on Trypticase soy agar (TSA plates) and incubated for 24 hours at 32.5°C. The cells are then recovered and resuspended in a commercial cryoprotectant solution with cryobeads for storage at -80°C for a maximum of 14 months (long-term storage stock).

From a cryobead of this -80°C stock, subculturing is performed on TSA slant agar which is then incubated for 24 hours at 32.5°C to obtain the stock culture. This stock culture is stored at 4°C for a maximum of 9 weeks. The working culture is obtained by subculturing the stock culture onto Trypticase soy agar and incubating it for 24 hours at 35°C. The working suspension is prepared by suspending the cells of this working culture in a trypton salt diluent. This suspension is calibrated between 1 and 3 x 10⁸ CFU (colony forming units)/mE by measuring the absorbance at 620 nm. Example 1

Evaluation of the antimicrobial activity of the samples against S. aureus

Six formulae were prepared according to the information given in Table 1 below. All the formulae are aqueous mixtures, comprising a qsp amount of water and also a preserving agent.

Hydroxypropylmethylcellulose (HPMC) at 1%, used here as a comparative, is indeed also a film-forming and/or viscosity-controlling polymer commonly used in the cosmetics industry.

At TO months (i.e. between 1 and 7 days of the manufacture of the formulae) and T6 months of the manufacture of the formulae, 20-gram aliquots of each formula are inoculated with 0.2 ml of a calibrated suspension of S. aureus. After homogenization, the microorganism content in the product represents a concentration of S. aureus of 10⁶ CFU per gram of product, i.e. a 1% inoculation of a suspension of 10⁸ CFU per mF (the inoculum content is determined by spreading the suspension on Trypticase soy agar plates and incubating for 24 hours at 35°C). After t = 30 minutes and t = 60 minutes of contact at room temperature (20°C ± 3°C), 1 gram of the mixture is weighed and 9.0 mF of Eugon ET100 supp broth are then added, followed by mixing until fully homogenized. This mixture is then serially diluted in Eugon FT 100 supp broth to 1/100 dilution.

The dilutions are spread on Trypticase soy agar plates and incubated at 35 °C for 48 hours until the surviving colonies of S. aureus are counted.

The antimicrobial activity on S. aureus is expressed in logarithmic abatement relative to the initial content and associated with the activity of the active compound, which is the endolysin of sequence SEQ ID NO: 10 by comparing the counts of surviving S. aureus in formulae with and without the endolysin of sequence SEQ ID NO: 10, in the presence of each of the polymers tested.

This method is an adaptation of the challenge test method described in the standard “ISO 11930 Cosmetics - Microbiology - Evaluation of the antimicrobial protection of a cosmetic product.

PVA and HPMC are present in the compositions of the present example in a content usual for each of them in a cosmetic composition, in particular in a content usual for each of them enabling it to play its role as a film-forming agent and/or viscosity control agent in a composition of the invention. Results and conclusions

The results are expressed as logarithmic Staphylococcus aureus abatement relative to the inoculated content. The abatements are thus between 0 log (no antimicrobial activity observed in the tested condition i.e. complete loss of activity of the endolysin of sequence SEQ ID NO: 10) and -5.4 log (maximum reduction observable under the test conditions i.e. maintenance of endolysin activity in the tested condition). The formula tested is thus particularly active when the values are close to -5.4.

A first evaluation of the antimicrobial activity of the endolysin is taken at TO (i.e. between 1 day and 7 days after the manufacture of the formulae), and a second at T6 months from the date of manufacture of the formulae. This evaluation is performed at these two times according to the protocol described above, i.e. measurement of the activity of the endolysin in the formula after 30 minutes of contact and 60 minutes of S’.t/z/rez/.s/formula contact.

The results are given in Table 1 hereinbelow.

[Table 1]

Table 1

The results show that the combination of PVA with the endolysin of sequence SEQ ID NO: 10 makes it possible to maintain better antibacterial performance against S. aureus of the endolysin - as shown by the reduction Log results in Table 1 - compared to the performance obtained by replacing PVA with HPMC.

Indeed, at T6M, the reduction Logs are much higher with PVA than with the two different HPMC concentrations.

Example 2

Three compositions A to C were prepared according to the information and contents given in Table 2 below. Compositions A and B, outside the invention, comprise a gel base (water + preserving agent) and 1% hydroxypropylmethylcellulose (HPMC) (see Example 1). Composition C, according to the invention, comprises a gel base (water + preserving agent) and 2% PVA (see also Example 1). The contents are given as mass percentages. 0.0025652% of the endolysin of sequence SEQ ID NO: 10 was added to the compositions under a laminar flow hood to avoid any microbial contamination.

The preparation of the compositions depended on the desired viscosity of each.

Compositions A to C were prepared by adding 1.08 mL of an aqueous solution containing 0.095% by weight of endolysin of sequence SEQ ID NO: 10 to 38.9 g of a gel base (water + preserving agent) in a 50 mL Falcon tube. The tube was placed in a shaker so as to homogenize the formulation at room temperature.

The compositions were prepared by mixing 1.08 mL of an aqueous solution containing 0.095% by weight of endolysin of sequence SEQ ID NO: 10 with 38.9 g of a gel base using a mortar and pestle for 5 minutes. Samples of approximately 5 mL were aliquoted into 15 mL tubes and placed under storage conditions at 25°C, 40°C and 45°C.

A control formulation was prepared in a similar manner to compositions A to C, using sterile storage buffer instead of endolysin (SB). Samples for viscosity measurements were also prepared in the same manner for a total volume of 50 mL using 1.35 mL of endolysin and 48.65 g of gel base in a Falcon tube. The various polymers (PVA or HPMC) are present in the compositions of the present example in a content usual for each of them in a cosmetic composition, in particular in a content usual for each of them to play its role as a film-forming and viscosity-controlling agent in a composition of the invention. [Table 2]

Table 2

A destruction assay study was conducted on all of the above compositions and also on the control composition with 2.7% by weight of an aqueous solution containing 0.095% by weight of endolysin of sequence SEQ ID NO: 10 at TO so as to measure the capacity of the endolysin of sequence SEQ ID NO: 10 to digest S. aureus. Only one concentration of S. aureus was used in this study, and the ODeoo was set at 0.15.

Results and conclusions

In this study, a destruction capacity of greater than 90%, i.e. a reduction log of 1, was considered acceptable for formulations comprising the endolysin of sequence SEQ ID NO: 10. The results of this study are given in Table 3 below. This study compares the change over time at TO (i.e. between 1 day and 7 days after manufacture of the formulae), T7 (7 days after TO), T28 (28 days after TO), and T84 (84 days after TO) and of temperature (at 25°C, 40°C and 45°C) of the various compositions, observing the change in the endolysin destruction values as a function of the values at TO for all formulae. It also made it possible to highlight the robust formulae (little or no change in endolysin stability over time) and the less robust formulas (loss of endolysin destruction capacity over time) among the formulae that remained.

[Table 2]

The results of the study show that all compositions comprising the endolysin of sequence SEQ ID NO: 10 are stable at 25°C after three months of storage with a destruction assay of greater than 90%. After 84 days of storage at 40°C and 45°C, the comparative formulae A and B show a reduction in the antimicrobial potency of the endolysin of sequence SEQ ID NO: 10, as evidenced by the reduced degree of destruction.

Thus, it is clear that the systems according to the invention comprising an endolysin in combination with at least one PVA are robust systems for maintaining the antimicrobial activity against S. aureus of the endolysin of sequence SEQ ID NO: 10 over time, in a cosmetic composition environment.

In contrast, a system comprising an endolysin in combination with a film-forming and viscosity-controlling agent other than PVA, such as hydroxypropylmethylcellulose at a concentration equal to 1% or 2% by weight, shows a reduction in the antimicrobial activity of the endolysin of sequence SEQ ID NO: 10 after 84 days at 40°C and 45 °C. This system is thus less resistant, in particular over time, than a system according to the invention.

Sequence listing

SEQ ID NO: 1 CBD-2638 (PROTEIN) WKQNKDGIWYKAEHASFTVTAPEGIITRYKGPWTGHPQAGVLQKGQTIKYDEQK FDGHVWVSWETFEGETVYMPVRTWDAKTGKVGKLWGEIK

SEQ ID NO: 2 CBD-2638 (NUCLEIC ACID)

TGGAAACAGAATAAAGATGGCATTTGGTATAAAGCTGAACATGCTTCGTTCAC

AGTGACAGCACCAGAGGGAATTATCACAAGATACAAAGGTCCTTGGACTGGTC

ACCCACAAGCTGGTGTATTACAAAAAGGTCAAACGATTAAATATGATGAGGTT CAAAAATTTGACGGTCATGTTTGGGTATCGTGGGAAACGTTTGAGGGCGAAAC

TGTATACATGCCGGTACGCACATGGGACGCTAAAACTGGTAAAGTTGGTAAGT TGTGGGGCGAAATTAAATAA

SEQ ID NO: 3 PLY2638 (PROTEIN)

MLTAIDYLTKKGWKISSDPRTYDGYPKNYGYRNYHENGINYDEFCGGYHRAFDV

YSNETNDVPAVTSGTVIEANDYGNFGGTFVIRDANDNDWIYGHLQRGSMRFVVG DKVNQGDIIGLQGNSNYYDNPMSVHLHLQLRPKDAKKDEKSQVCSGLAMEKYDI TNLNAKQDKSKNGSVKELKHIYSNHIKGNKITAPKPSIQGVVIHNDYGSMTPSQYL PWLYARENNGTHVNGWASVYANRNEVLWYHPTDYVEWHCGNQWANANLIGFE VCESYPGRISDKLFLENEEATLKVAADVMKSYGLPVNRNTVRLHNEFFGTSCPHR

SWDLHVGKGEPYTTTNINKMKDYFIKRIKHYYDGGKLEVSKAATIKQSDVKQEV KKQEAKQIVKATDWKQNKDGIWYKAEHASFTVTAPEGIITRYKGPWTGHPQAGV

LQKGQTIKYDEVQKFDGHVWVSWETFEGETVYMPVRTWDAKTGKVGKLWGEIK

SEQ ID NO: 4 PLY2638 (PROTEIN)

MRGSHHHHHHGSMLTAIDYLTKKGWKISSDPRTYDGYPKNYGYRNYHENGINYD

EFCGGYHRAFDVYSNETNDVPAVTSGTVIEANDYGNFGGTFVIRDANDNDWIYG

HLQRGSMRFVVGDKVNQGDIIGLQGNSNYYDNPMSVHLHLQLRPKDAKKDEKSQ VCSGLAMEKYDITNLNAKQDKSKNGSVKELKHIYSNHIKGNKITAPKPSIQGVVIH NDYGSMTPSQYLPWLYARENNGTHVNGWASVYANRNEVLWYHPTDYVEWHCG NQWANANLIGFEVCESYPGRISDKLFLENEEATLKVAADVMKSYGLPVNRNTVRL HNEFFGTSCPHRSWDLHVGKGEPYTTTNINKMKDYFIKRIKHYYDGGKLEVSKAA TIKQSDVKQEVKKQEAKQIVKATDWKQNKDGIWYKAEHASFTVTAPEGIITRYKG PWTGHPQAGVLQKGQTIKYDEVQKFDGHVWVSWETFEGETVYMPVRTWDAKT

GKVGKLWGEIK

SEQ ID NO: 5 PLY2638 (NUCLEIC ACID)

ATGCTAACTGCTATTGACTATCTTACGAAAAAAGGTTGGAAAATATCATCTGA

CCCTCGCACTTACGATGGTTACCCTAAAAACTACGGCTACAGAAATTACCATG

AAAACGGCATTAATTATGATGAGTTTTGTGGTGGTTATCATAGAGCTTTTGATG

TTTACAGTAACGAAACTAACGACGTGCCTGCTGTTACTAGCGGAACAGTTATT

GAAGCAAACGATTACGGTAATTTTGGTGGTACATTCGTTATTAGAGACGCTAA

CGATAACGATTGGATATATGGGCATCTACAACGTGGCTCAATGCGATTTGTTGT

AGGCGACAAAGTCAATCAAGGTGACATTATTGGTTTACAAGGTAATAGCAACT

ATTACGACAATCCTATGAGTGTACATTTACATTTACAATTACGCCCTAAAGACG

CAAAGAAAGATGAAAAATCACAAGTATGTAGTGGTTTGGCTATGGAAAAATAT

GACATTACAAATTTAAATGCTAAACAAGATAAATCAAAGAATGGGAGCGTGA

AAGAGTTGAAACATATCTATTCAAACCATATTAAAGGTAACAAGATTACAGCA

CCAAAACCTAGTATTCAAGGTGTGGTCATCCACAATGATTATGGTAGTATGAC

ACCTAGTCAATACTTACCATGGTTATATGCACGTGAGAATAACGGTACACACG

TTAACGGTTGGGCTAGTGTTTATGCAAATAGAAACGAAGTGCTTTGGTATCATC

CGACAGACTACGTAGAGTGGCATTGTGGTAATCAATGGGCAAATGCTAACTTA

ATCGGATTTGAAGTGTGTGAGTCGTATCCTGGTAGAATCTCGGACAAATTATTC

TTAGAAAATGAAGAAGCGACATTGAAAGTAGCTGCGGATGTGATGAAGTCGT

ACGGATTACCAGTTAATCGCAACACTGTACGTCTGCATAACGAATTCTTCGGA

ACTTCTTGTCCACATCGTTCGTGGGACTTGCATGTTGGCAAAGGTGAGCCTTAC

ACAACTACTAATATTAATAAAATGAAAGACTACTTCATCAAACGCATCAAACA

TTATTATGACGGTGGAAAGCTAGAAGTAAGCAAAGCAGCAACTATCAAACAAT

CTGACGTTAAGCAAGAAGTTAAAAAGCAAGAAGCAAAACAAATTGTGAAAGC

AACAGATTGGAAACAGAATAAAGATGGCATTTGGTATAAAGCTGAACATGCTT

CGTTCACAGTGACAGCACCAGAGGGAATTATCACAAGATACAAAGGTCCTTGG

ACTGGTCACCCACAAGCTGGTGTATTACAAAAAGGTCAAACGATTAAATATGA

TGAGGTTCAAAAATTTGACGGTCATGTTTGGGTATCGTGGGAAACGTTTGAGG

GCGAAACTGTATACATGCCGGTACGCACATGGGACGCTAAAACTGGTAAAGTT

GGTAAGTTGTGGGGCGAAATTAAATAA SEQ ID NO: 6 M23-LST (PROTEIN)

AATHEHSAQWLNNYKKGYGYGPYPLGINGGMHYGVDFFMNIGTPVKAISSGKIV

EAGWSNYGGGNQIGLIENDGVHRQWYMHLSKYNVKVGDYVKAGQIIGWSGSTG YSTAPHLHFQRMVNSFSNSTAQDPMPFLKSAGY

SEQ ID NO: 7 M23-LST (NUCLEIC ACID)

GCTGCAACACATGAACATTCAGCACAATGGTTGAATAATTACAAAAAAGGATA

TGGTTACGGTCCTTATCCATTAGGTATAAATGGCGGTATGCACTACGGAGTTGA

TTTTTTTATGAATATTGGAACACCAGTAAAAGCTATTTCAAGCGGAAAAATAG

TTGAAGCTGGTTGGAGTAATTACGGAGGAGGTAATCAAATAGGTCTTATTGAA

AATGATGGAGTGCATAGACAATGGTATATGCATCTAAGTAAATATAATGTTAA

AGTAGGAGATTATGTCAAAGCTGGTCAAATAATCGGTTGGTCTGGAAGCACTG

GTTATTCTACAGCACCACATTTACACTTCCAAAGAATGGTTAATTCATTTTCAA

ATTCAACTGCCCAAGATCCAATGCCTTTCTTAAA GAGCGCAGGATAT

SEQ ID NO: 8 AMI-2638 (PROTEIN)

NKITAPKPSIQGVVIHNDYGSMTPSQYLPWLYARENNGTHVNGWASVYANRNEV

LWYHPTDYVEWHCGNQWANANLIGFEVCESYPGRISDKLFLENEEATLKVAADV

MKSYGLPVNRNTVRLHNEFFGTSCPHRSWDLHVGKGEPYTTTNINKMKDYFIKRI KHYYDG

SEQ ID NO: 9 AMI-2638 (NUCLEIC ACID)

GGTAACAAGATTACAGCACCAAAACCTAGTATTCAAGGTGTGGTCATCCACAA

TGATTATGGTAGTATGACACCTAGTCAATACTTACCATGGTTATATGCACGTGA

GAATAACGGTACACACGTTAACGGTTGGGCTAGTGTTTATGCAAATAGAAACG

AAGTGCTTTGGTATCATCCGACAGACTACGTAGAGTGGCATTGTGGTAATCAA

TGGGCAAATGCTAACTTAATCGGATTTGAAGTGTGTGAGTCGTATCCTGGTAG

AATCTCGGACAAATTATTCTTAGAAAATGAAGAAGCGACATTGAAAGTAGCTG

CGGATGTGATGAAGTCGTACGGATTACCAGTTAATCGCAACACTGTACGTCTG

CATAACGAATTCTTCGGAACTTCTTGTCCACATCGTTCGTGGGACTTGCATGTT GGCAAAGGTGAGCCTTACACAACTACTAATATTAATAAAATGAAAGACTACTT

CATCAAACGCATCAAACATTATTATGACGGT

SEQ ID NO: 10 M23-LST AMI2638 CBD2638 (PROTEIN)

AATHEHSAQWLNNYKKGYGYGPYPLGINGGMHYGVDFFMNIGTPVKAISSGKIV

EAGWSNYGGGNQIGLIENDGVHRQWYMHLSKYNVKVGDYVKAGQIIGWSGSTG

YSTAPHLHFQRMVNSFSNSTAQDPMPFLKSAGYGKAGGTVTPTPNTGELLRPKDA

KKDEKSQVCSGLAMEKYDITNLNAKQDKSKNGSVKELKHIYSNHIKGNKITAPKP

SIQGVVIHNDYGSMTPSQYLPWLYARENNGTHVNGWASVYANRNEVLWYHPTD

YVEWHCGNQWANANLIGFEVCESYPGRISDKLFLENEEATLKVAADVMKSYGLP

VNRNTVRLHNEFFGTSCPHRSWDLHVGKGEPYTTTNINKMKDYFIKRIKHYYDGG

KLEVSKAATIKQSDVKQEVKKQEAKQIVKATDWKQNKDGIWYKAEHASFTVTAP

EGIITRYKGPWTGHPQAGVLQKGQTIKYDEVQKFDGHVWVSWETFEGETVYMPV RTWDAKTGKVGKLWGEIK

SEQ ID NO: 11 M23-LST AMI2638 CBD2638 (PROTEIN)

MRGSHHHHHHGSAATHEHSAQWLNNYKKGYGYGPYPLGINGGMHYGVDFFMNI

GTPVKAISSGKIVEAGWSNYGGGNQIGLIENDGVHRQWYMHLSKYNVKVGDYVK

AGQIIGWSGSTGYSTAPHLHFQRMVNSFSNSTAQDPMPFLKSAGYGKAGGTVTPT

PNTGELLRPKDAKKDEKSQVCSGLAMEKYDITNLNAKQDKSKNGSVKELKHIYSN

HIKGNKITAPKPSIQGVVIHNDYGSMTPSQYLPWLYARENNGTHVNGWASVYAN

RNEVLWYHPTDYVEWHCGNQWANANLIGFEVCESYPGRISDKLFLENEEATLKV

AADVMKSYGLPVNRNTVRLHNEFFGTSCPHRSWDLHVGKGEPYTTTNINKMKDY

FIKRIKHYYDGGKLEVSKAATIKQSDVKQEVKKQEAKQIVKATDWKQNKDGIWY

KAEHASFTVTAPEGIITRYKGPWTGHPQAGVLQKGQTIKYDEVQKFDGHVWVSW

ETFEGETVYMPVRTWDAKTGKVGKLWGEIK

SEQ ID NO: 12 M23-LST AMI2638 CBD2638 (NUCLEIC ACID)

GCTGCAACACATGAACATTCAGCACAATGGTTGAATAATTACAAAAAAGGATA

TGGTTACGGTCCTTATCCATTAGGTATAAATGGCGGTATGCACTACGGAGTTGA

TTTTTTTATGAATATTGGAACACCAGTAAAAGCTATTTCAAGCGGAAAAATAG

TTGAAGCTGGTTGGAGTAATTACGGAGGAGGTAATCAAATAGGTCTTATTGAA AATGATGGAGTGCATAGACAATGGTATATGCATCTAAGTAAATATAATGTTAA

AGTAGGAGATTATGTCAAAGCTGGTCAAATAATCGGTTGGTCTGGAAGCACTG

GTTATTCTACAGCACCACATTTACACTTCCAAAGAATGGTTAATTCATTTTCAA

ATTCAACTGCCCAAGATCCAATGCCTTTCTTAAAGAGCGCAGGATATGGAAAA

GCAGGTGGTACAGTAACTCCAACGCCGAATACAGGTGAGCTCTTACGCCCTAA

AGACGCAAAGAAAGATGAAAAATCACAAGTATGTAGTGGTTTGGCTATGGAA

AAATATGACATTACAAATTTAAATGCTAAACAAGATAAATCAAAGAATGGGA

GCGTGAAAGAGTTGAAACATATCTATTCAAACCATATTAAAGGTAACAAGATT

ACAGCACCAAAACCTAGTATTCAAGGTGTGGTCATCCACAATGATTATGGTAG

TATGACACCTAGTCAATACTTACCATGGTTATATGCACGTGAGAATAACGGTA

CACACGTTAACGGTTGGGCTAGTGTTTATGCAAATAGAAACGAAGTGCTTTGG

TATCATCCGACAGACTACGTAGAGTGGCATTGTGGTAATCAATGGGCAAATGC

TAACTTAATCGGATTTGAAGTGTGTGAGTCGTATCCTGGTAGAATCTCGGACA

AATTATTCTTAGAAAATGAAGAAGCGACATTGAAAGTAGCTGCGGATGTGATG

AAGTCGTACGGATTACCAGTTAATCGCAACACTGTACGTCTGCATAACGAATT

CTTCGGAACTTCTTGTCCACATCGTTCGTGGGACTTGCATGTTGGCAAAGGTGA

GCCTTACACAACTACTAATATTAATAAAATGAAAGACTACTTCATCAAACGCA

TCAAACATTATTATGACGGTGGAAAGCTAGAAGTAAGCAAAGCAGCAACTATC

AAACAATCTGACGTTAAGCAAGAAGTTAAAAAGCAAGAAGCAAAACAAATTG

TGAAAGCAACAGATTGGAAACAGAATAAAGATGGCATTTGGTATAAAGCTGA

ACATGCTTCGTTCACAGTGACAGCACCAGAGGGAATTATCACAAGATACAAAG

GTCCTTGGACTGGTCACCCACAAGCTGGTGTATTACAAAAAGGTCAAACGATT

AAATATGATGAGGTTCAAAAATTTGACGGTCATGTTTGGGTATCGTGGGAAAC

GTTTGAGGGCGAAACTGTATACATGCCGGTACGCACATGGGACGCTAAAACTG

GTAAAGTTGGTAAGTTGTGGGGCGAAATTAAATAA

SEQ ID NO: 13 6XHIS N-TERMINAL TAG

MRGSHHHHHHGS

Claims

Claims

1. Composition, notably a cosmetic composition, comprising, in a physiologically acceptable medium:

(i) at least one endolysin; and

(ii) at least one polyvinyl alcohol.

2. Composition according to Claim 1, in which the endolysin is an endolysin derived from a Staphylococcus aureus phage.

3. Composition according to Claim 1 or 2, in which the endolysin comprises a first protein sequence comprising a domain for binding to the cell wall of species of the genus Staphylococcus .

4. Composition according to Claim 3, in which the first protein sequence is derived from the endolysin of the O2638a bacteriophage of S. aureus.

5. Composition according to Claim 3 or 4, in which the first protein sequence comprises a protein sequence comprising at least 80%, in particular at least 90%, more particularly at least 95% sequence identity with the reference amino acid sequence SEQ ID NO: 1.

6. Composition according to any one of Claims 1 to 4, in which he endolysin comprises a protein sequence comprising at least 80%, in particular at least 90%, more particularly at least 95% sequence identity with an amino acid sequence chosen from the group consisting of the amino acid sequences of references SEQ ID NO: 3 and SEQ ID NO: 4.

7. Composition according to any one of Claims 3 to 5, in which said endolysin also comprises a heterologous protein sequence.

8. Composition according to Claim 7, in which the heterologous protein sequence comprises a lytic domain, said lytic domain comprising a second and a third protein sequence, said second protein sequence comprising an endopeptidase M23 domain and said third protein sequence comprising an amidase domain.

9. Composition according to Claim 8, in which said second and third protein sequences are derived, independently of each other, from an enzyme chosen from the group consisting of the endolysin from 02638a bacteriophage of S. aureus and the lysostaphin from S. simulans, in particular one of the second and third protein sequences is derived from the endolysin of 02638a bacteriophage of S. aureus and the other sequence of the second and third protein sequences is derived from the lysostaphin of S. simulans.

10. Composition according to Claim 8 or 9, in which said second protein sequence comprises at least 80%, in particular at least 90 %, more particularly at least 95 % sequence identity with the reference amino acid sequence SEQ ID NO: 6 and said third protein sequence comprises at least 80%, in particular 90%, more particularly 95% sequence identity with the reference amino acid sequence SEQ ID NO: 8.

11. Composition according to any one of Claims 1 to 4 and 6 to 9, in which the endolysin comprises a protein sequence comprising at least 80%, in particular at least 90%, more particularly at least 95% sequence identity with an amino acid sequence chosen from the group consisting of the amino acid sequences of references SEQ ID NO: 10, and SEQ ID NO: 11, in particular the endolysin comprises a protein sequence consisting of the reference amino acid sequence SEQ ID NO: 10.

12. Composition according to any one of Claims 1 to 11 , in which the endolysin, in particular the endolysin derived from a Staphylococcus aureus phage, is present in an amount ranging from 0.0001% to 0.1% by weight, in particular 0.0005% to 0.01% by weight, more particularly 0.001% to 0.005% by weight relative to the total weight of the composition.

13. Composition according to any one of Claims 1 to 12, the polyvinyl alcohol including at least one of units of formula (I):

14. Composition according to any one of Claims 1 to 13, in which the polyvinyl alcohol has a mass-average molecular weight from 1000 to 1 000 000 g/mol, preferably from 10 000 to 500 000 g/mol and even more preferentially from 50 000 to 400 000 g/mol.

15. Composition according to any one of Claims 1 to 14, in which the polyvinyl alcohol is present in a content ranging from 0.1% to 20% by weight relative to the total weight of the composition, in particular from 0.5% to 10%, even more preferentially from 1% to 5% by weight relative to the total weight of the composition.

16. Composition according to any one of Claims 1 to 15, the composition being suitable for topical administration.

17. Cosmetic use, notably the topical use, of a composition according to any one of Claims 1 to 16, for preventing and/or treating acne and/or eczema in an individual in need thereof.

18. Cosmetic use according to Claim 17, in which the composition is suitable for topical administration.

19. Non-therapeutic cosmetic process for caring for keratin materials, in particular the skin, notably acne-prone skin, comprising the topical application to said keratin materials of a composition according to any one of Claims 1 to 16.