WO2013131856A1

WO2013131856A1 - Antioxidant peptides, compositions comprising them and their uses

Info

Publication number: WO2013131856A1
Application number: PCT/EP2013/054291
Authority: WO
Inventors: Jean-Marc Sabatier; Nicolas ANDREOTTI; Bonabes-Olivier De Rouge
Original assignee: Tournoux Biotech
Priority date: 2012-03-05
Filing date: 2013-03-04
Publication date: 2013-09-12

Abstract

The present invention relates to peptides or derivatives thereof, cosmetic or pharmaceutical compositions comprising the same and uses thereof as antioxidant or antiallergenic agents.

Description

ANTIOXIDANT PEPTIDES, COMPOSITIONS COMPRISING THEM AND THEIR USES

FIELD OF THE INVENTION

BACKGROUND OF THE INVENTION

Antioxidants/reductants

Basics on oxidation-reduction reactions:

An antioxidant/reductant compound is a compound able to slow down or prevent the oxidation of other molecules, via oxidation-reduction reaction(s). During such reactions, there is a transfer of electrons from a suitable electron donor (antioxidant or reductant) to a suitable electron acceptor (oxidant). In some oxidation-reduction reactions, only electrons are transferred from the antioxidant/reductant to the oxidant (e.g. electron transfer between cytochromes), whereas in other reactions, both electrons and protons (H⁺) are transferred

(e.g. electrons and protons transfers between NADH and FAD). The oxidized and reduced forms of the compounds (e.g. peptides) operating in oxidation-reduction reactions are referred to as redox couples or pairs. The facility with which a given antioxidant/reductant (electron donor) gives up its electrons to an oxidant (electron acceptor) is quantitatively expressed as the oxidation-reduction potential (ORP) of the system. The ORP is measured

(with an ORP meter) in Volts as an electromotive force of a half-cell made up of both partners of an oxidation-reduction couple when compared to a reference half-cell (hydrogen electrode reaction). The antioxidant/reductant of an oxidation-reduction pair with large negative ORP will give up its electrons more readily than pairs with smaller negative or positive ORPs. This implies that a strong oxidant (e.g. characterized by a large positive ORP) possesses a very high affinity for electrons. The relationship between the standard ORP of a specific redox pair (E Ό), the observed potential (E), and the ratio of the concentrations of oxidant and antioxidant/reductant in the system is described by the Nernst equation:

E = E'o + 2.3 RT/nF log ([oxidant]/[reductant])

Where E is the observed potential with all concentrations at 1M;

E'o is the standard potential at pH 7;

R is the gas constant (8.3 J/deg/mol); T is the absolute temperature (Kelvin degrees);

n is the number of electrons being transferred;

F is the Faraday (96,500 J/V). When the observed potential is equal to the standard potential, the potential is referred to as the midpoint potential, in which the concentrations of oxidant and antioxidant/reductant are the same. The knowledge of ORPs for various redox reactions therefore allows one to predict the direction of electron flow or transfer when more than one redox pair is concerned. For example, the NAD⁺-NADH pair has a standard potential of -0.32 V, whereas the pyruvate/lactate pair possesses a potential of -0.19 V. The electrons will flow from the NAD⁺-NADH system to the pyruvate/lactate system (pyruvate + NADH + H⁺ (gives) lactate + NAD⁺). Of note, ORP values are dependent on buffer used for ORP measurements with an ORP meter; high pH buffer (basic buffer) showed negative ORP (reducing medium), whereas low pH buffer (acidic buffer) showed positive ORP (oxidant medium). Water ORP generally ranges between +90 to +110 mV. In water, a negative

ORP value of -40 mV for 1 mM reduced glutathione (GSH), a natural antioxidant, was obtained.

Benefits of antioxidant agents:

The oxidation is a chemical reaction that transfers electrons from a molecule (e.g. antioxidant/reductant compound) to another one (oxidant). Oxidation reactions produce free radicals which start chain reactions and oxidative stress (large rise in the cellular reduction potential or a marked decrease in the reducing capacity of the cellular redox couples, such a glutathione) that damage or kill cells by action on a variety of cellular components such as proteins, lipids and DNA. By interfering with oxidants (e.g. superoxide anion .0₂ ^~, hydroxyl radical .OH, hydrogen peroxide H₂0₂, alkoxy radical RO., peroxy radical ROO., peroxynitrite ONOO-, hypochlorous acid HOC1, organic hydroperoxide ROOH), antioxidant/reductant (or also called herein 'antioxidant') compounds inhibit these chain reactions through neutralization of reactive oxygen species (ROS) that are free radicals, oxygen ions and peroxides, both organic and inorganic. The reported harmful effects of ROS on cells are the (i) damage of nucleic acids (DNA and RNA), (ii) oxidations of specific amino acid residues in proteins, (iii) oxidations of fatty acids (polydesaturated) in lipids, referred to as lipid peroxidation, (iv) inactivation of some enzymes by oxidation of their related co-factors, and (v) induction of apoptosis (programmed cell death). Antioxidant compounds inhibit oxidation reactions by being oxidized themselves. Some antioxidants are consequently known to reduce oxidative stress; in that case they are also called antiradicals or radical scavengers. Reducing compounds such as polyphenols (epicatechin derivatives), ascorbic acid (vitamin C), tocopherols and tocotrienols (vitamin E), vitamin A, vitamin B2, β-carotene, lycopene, CoQIO, glutathione, thiols, tanins, uric acid, lipoic acid are known as potent antioxidants/reductants. Several natural antioxidant enzymes are also well characterized, such as superoxide dismutases (SOD), catalases, peroxiredoxins, and a number of peroxidases. The natural sources of antioxidant compounds are mainly fruits, cereals, vegetables and herbs (e.g. aloe vera, bilberry, grape seed or pine bark extracts, turmeric (curcumin), cascara sagrada, milk thistle, ginkgo biloba, to cite a few). Benefits of antioxidant/reductant compounds were reported for maintaining the integrity of blood vessel membranes (brain, heart) and treating or preventing cancer, cardiovascular, inflammatory and neurodegenerative diseases (e.g. Alzheimer's and Parkinson's diseases, dementia). Some of these compounds are also currently used industrially as preservatives in food and cosmetics. In the case of topically applied cosmetic compositions, such antioxidant compounds were also found to delay/slow skin damages due to oxidative stress, in particular skin ageing.

Allergens

Basics on allergy and allergens:

Allergy is a sort of disorder (over-reaction) of the immune system, also called atopy. Allergic reactions are caused by allergens, which are normally harmless environmental compounds. Allergy (type I hypersensitivity) can be characterized by an excessive activation of white blood cells (i.e. basophils and mast cells) by a type of immunoglobulin called IgE. This excessive activation of cells results in a severe inflammatory response. Among allergic reactions are asthma, eczema (atopic dermatitis), hay fever (allergic rhinitis), hives (urticaria), food allergies, etc. Anaphylaxis is the most severe, whole body and life-threatening type of allergic reaction. Among allergens (substances causing allergic reactions) are found chemicals, perfumes, pollen, mold spores, plants, dander, dust, foods, drugs (e.g. antibiotics, aspirin), insect venoms (e.g. wasps), parasites, viruses and bacteria. Most environmental allergens contact the skin or eyes, or are inhaled, explaining why most symptoms are affecting the skin, eyes, or the airways. The antagonist drugs currently used in pharmacotherapy block the action of allergic mediators or prevent activation of cells and degranulation processes. The most common drugs are cortisone, anti-histamines, dexamethasone, epinephrine/adrenaline, theophylline and cromolyn sodium. Anti- cholinergics, anti-leukotrienes, decongestants and mast cell stabilizers are also used. These drugs help to alleviate the symptoms of allergy and are required for the treatment of acute anaphylaxis, although they play a little role in the chronic treatment of allergic disorders.

The inventors have surprisingly found that specific short peptides can have high antioxidant properties and can consequently be useful in cosmetic and/or pharmaceutical compositions, in particular as antioxidant, antiallergen, antiradical and/or antiinflammatory agents. Peptides of the invention are more particularly useful to prevent oxidation of compounds in the cosmetic or pharmaceutical compositions and also to prevent or treat damages due or related to oxidative stress, in particular skin damages due or related to oxidative stress, and enhancing thereby the protection against ultraviolet (UV) exposure (photo-protection) or environmental aggressions, skin inflammation, irritation or reddening (erythema) and UV-induced DNA damage. They can also potently recognize and interact (in particular interact covalently) with allergenic substances (herein referred to as 'allergens'), being consequently useful in the detection of allergens (e.g. diagnostic kit), as well as in the treatment of disorders originating from allergen-induced reactions. Such anti-allergen peptides, and derivatives thereof, may be useful in dermatologic and/or pharmaceutical preparations to treat allergy, especially skin allergy and associated inflammation, such as allergic contact dermatitis. SUMMARY OF THE INVENTION

The present invention deals with specific peptides (and derivatives thereof) with antioxidant and/or antiallergenic properties, which can be particularly useful in cosmetic or pharmaceutical compositions. The invention also deals with a nucleic acid comprising a sequence, which encodes a peptide of the invention. Such compounds can be used in cosmetic or pharmaceutical (preferably dermatological) preparations either alone or in combination with additional compounds usually implemented in cosmetics or pharmaceutics, such as antioxidant(s), for instance vitamins C and E. The present invention also deals with peptides of the invention (and derivatives thereof) particularly useful in the (i) detection and/or classification of allergens, or (ii) pharmaceutical compositions to treat allergy.

The invention also relates to cosmetic or pharmaceutical compositions comprising in a physiologically acceptable support at least one peptide of the invention, said peptide being particularly useful as an antioxidant agent or as antiallergenic agent.

The invention also relates to a process for the treatment of disease related or due to an oxidative stress and/or to allergen-induced reactions, especially of the skin.

The invention also deals with a process to retain or improve skin or hair in or to healthy and fresh conditions by protecting them from damage by oxidation reaction. It aims also at providing a process to prevent or treat skin damage due to oxidative stress, in particular enhancing thereby the protection against ultraviolet (UV) exposure (photo -protection), or treating skin inflammation, irritation or reddening (erythema) and UV-induced DNA damage.

DETAILED DESCRIPTION OF THE INVENTION

PEPTIDES

The peptides of the invention comprise the following sequence (seq. ID n°l):

VYMNR YYKL, or variants thereof defined as comprising seq. ID n°l with deletion, substitution, and/or addition of one, two, three or more amino acids therein.

Peptides of the invention can be more particularly defined as comprising the following sequence seq. ID n°2:

XI -X2-YM-X3-NR-X4-KY-X5-K-X6-X7

wherein:

XI is vacant, or is an amino acid sequence consisting of RCR,

X2 is vacant, or V;

X3 is vacant, or is an amino acid sequence consisting of RG;

X4 is vacant, or C;

X5 is vacant, or Y;

X6 is vacant or L; and X7 is vacant, or an amino acid sequence selected in the group consisting of RCR, CCK and CLK.

According to a particular embodiment, the peptides of the invention consist (essentially) of the sequence seq ID n° 2.

According to a preferred embodiment, the peptides of the invention comprise the sequence seq. ID n°2 wherein at least two, or preferably at least three, of XI, X2, X3, X4, X5, X6, and X7 are not vacant.

According to specific embodiments, peptides of the invention can be more particularly defined as comprising sequence seq. ID n°2 with one of the following specific features: where X6 is L, X7 is vacant or an amino acid sequence CLK,

where X6 is vacant, X7 is CCK or RCR,

where X4 is C, XI, X3 and X7 are vacant and X2, X5, and X6 are not vacant,

where X4 is vacant, XI, X2, X3, X5, X6 and X7 are as identified above,

where XI is RCR, X7 is not RCR,

where X7 is RCR, XI is not RCR.

Whenever it is possible (i.e., not contradictory), some (in particular two or three) of the above specific features can be cumulative.

According to a specific embodiment, the peptides of the invention comprise (or preferably consist of) the sequence seq ID n° 2 with one, two or three of (or optionally all) the following features:

- X2 is V, and/or

- XI is vacant, and/or

- X3 is vacant, and/or

- X5 is Y. The peptides of the invention are more specifically from nine amino acid residues to twenty amino acid residues in length, preferably from ten to thirteen amino acid residues in length. The present invention discloses more specifically peptides or their derivatives comprising all, or part, of the amino acid sequences from (1), (3) to (8).

Some antioxidant/reductant peptides of the present invention were selected from domains of redox enzymes, especially NADH/FMN oxidoreductase (EmoB).

In a specific embodiment, the peptides of the invention comprise (or advantageously consist of) one of the following peptide sequences:

VYMNR YYKCCK (SEQ ID n° 3)

VYMNR YYKL (SEQ ID n° 1 )

RCRVYMNRKYYKL (SEQ ID n° 4)

VYMNRKYYKLCLK (SEQ ID n° 5)

VYMNRCKYYKL (SEQ ID n° 6)

YMNRKYKRCR (SEQ ID n° 7)

VYMRGNRKYYKL (SEQ ID n° 8)

According to the invention, the term 'comprise(s)' or 'comprising' can be generally interpreted such that all of the specifically mentioned features and any optional, additional and unspecified features are included; it can also be interpreted as the expression "consisting of where only the specified features are included, unless otherwise specified.

The peptides, which primary structures are detailed in this invention, are potent antioxidant and antiallergenic agents. In view of those properties, the peptides of the invention are particularly useful as active ingredients (preferably as antioxidant, antiallergen, antiradical and/or antiinflammatory agents) in cosmetic or pharmaceutical compositions. They are more particularly useful for treating diseases or disorders related to oxidative stress, such as cancers, and more particularly in topically applied cosmetic or dermato logical compositions, treating thereby environmental stress, UV-induced cellular damage on the skin and resulting skin ageing. They are also more particularly useful as interacting molecules with allergens in diagnostic kit(s), and also as active ingredients

(preferably as anti-allergy and/or anti-inflammatory agents) in dermatologic or pharmaceutical compositions, treating thereby diseases or disorders related to allergy or related or due to allergen-induced reactions, and more particularly in topically applied dermato logical compositions, treating thereby skin disorders originating from allergen- induced reactions.

The term 'peptide' refers to an isolated polymer of nine to twenty amino acid residues; polypeptides, oligopeptides, and proteins can then be included here within the definition of peptide. This term also does not specify or exclude post-expression modifications of peptides; for example, peptides which include the covalent attachment of glycosyl groups, acetyl groups, phosphate groups, lipid groups, and the like, are expressly encompassed by the term peptide.

The peptides are from nine amino acid residues to twenty amino acid residues in length, preferably from ten amino acid residues to thirteen amino acid residues in length, advantageously 10, 11, 12, or 13 amino acid residues.

The peptides according to the invention include also fragments, analogs and derivatives thereof. Said fragments, analogs and derivatives of the peptides described hereinabove have substantially the same protective functions and antioxidant/reductant and/or antiallergenic properties as the above defined peptides. More particularly, they can be used as active ingredients in cosmetic or pharmaceutical compositions to prevent or treat disorders or diseases related to an oxidative stress, in particular skin ageing due to oxidative stress, and to enhance thereby the protection against ultraviolet (UV) exposure

(photo-protection) or environmental aggressions, skin reddening (erythema) and UV- induced DNA damage. They can also be used as active ingredients (preferably as anti- allergy and/or anti-inflammatory agents) in dermatologic or pharmaceutical compositions, treating thereby diseases or disorders related to allergy or related or due to allergen- induced reactions, and more particularly in topically applied dermatological compositions, treating thereby skin disorders originating from allergen-induced reactions.

A peptide fragment is a polypeptide having a sequence that is entirely the same as part but not all of the given peptide sequence.

A specific embodiment of a peptide of interest according to the present invention, includes, but is not limited to, a peptide molecule which is resistant to proteolysis, a peptide in which a or the -CONH- amide peptide bond is modified and replaced by a (CH₂-NH) reduced bond, a (NH-CO) retro-inverso bond, a (CH₂-0) methylene-oxy bond, a (CH₂-S) thiomethylene bond, a (CH₂-CH₂) carba bond, a (CO-CH₂) ceto -methylene bond, a (CHOH-CH₂) hydroxyethylene bond), a (N-N) bond, a E-alcene bond or also a -CH=CH- bond. The invention also encompasses a peptide or a fragment thereof, in which at least one peptide bond has been modified as described above. Such fragments may be "freestanding", i.e. not part of or fused to other peptides, or they may be comprised within a single larger peptide of which they form a part or region.

However, several fragments may be comprised within a single larger peptide. Peptides according to the invention can present either carboxylic (i.e. -COO^") or carboxyl- amidated (i.e. -CONH₂) C-terminal extremity. The peptides of the invention can be 'free' or 'blocked' at N-termini (for instance, acetyl moiety), with either -COO^" (carboxylate) or - CONH₂ (carboxylamide) at C-termini. In a particular embodiment, the peptides according to the invention can present a (or several) lipid moiety (moieties), covalently or not covalently attached. The lipid moiety (moieties) can be any lipid presenting from C3 to C20 carbon atoms, preferably linked at N- or C- terminal extremity. According to another embodiment, peptides of the invention can comprise at least two or more consecutive amino acid sequences of the peptides (1) to (9), wherein said sequences are linear- or branched- attached. The number of repetitions of the peptide sequences can vary from 2 to 16, preferably from 2-4. Said repeated sequences can be interrupted (or linked) by one or several, identical or different, linker(s). Said linkers can be of different natures, including amino acid derivatives, in particular bifunctional amino acid derivatives, including K, ornithine, or any other suitable linker, such as bifunctional hydrocarbonated derivatives, including diaminobutyrate and diaminopropionate, providing said linker does not change substantially the desired effect (antioxidant/reductant or antyallergy) of the peptides according to the present invention.

The amino acid residues (or derivatives) within the peptides according to the invention can be L- and/or D-amino acid residues. Peptides containing all D-amino acid residues are also included, such as antisense peptides. Peptide sequences defined herein are represented by one-letter symbols for L- and D- amino acid residues as follows:

A (alanine) L (leucine)

R (arginine) K (lysine)

N (asparagine) M (methionine)

D (aspartic acid) F (phenylalanine)

C (cysteine) P (proline)

Q (glutamine) S (serine)

E (glutamic acid) T (threonine)

G (glycine) W (tryptophan)

H (histidine) Y (tyrosine)

I (iso leucine) V (valine)

The peptides of the invention can be produced by any chemical or genetic technique commonly known in the art. They can be produced using a peptide synthesizer with standard solid-phase synthesis approaches (Fmoc/t-butyl or Boc/benzyl chemistry, as described in Merrifield R.B., Science 232, 341-347, 1986). COMPOSITIONS, USES AND TREATMENTS

In another embodiment, the present invention relates to compositions, preferably cosmetic or pharmaceutical compositions, comprising the peptides of the invention, in particular as antioxidant or antiallergenic agents, preferably in a physiologically acceptable support. The pharmaceutical composition is preferably a dermatological composition or a composition useful for the treatment for ear, nose or throat, such as an aerosol composition. The composition of the invention is particularly useful for the treatment of a disorder or disease due or related to an oxidative stress, including cancer, and/or allergen-induced reactions, including related inflammation. According to a specific embodiment, the composition can be used for the treatment of skin damage due to an allergic reaction, in particular allergy- related inflammation, irritation, or reddening of skin. According to another embodiment, the composition can be used for the treatment of respiratory diseases due to allergic reactions, in particular hay-fever and asthma. In another embodiment, the present invention discloses methods for preparing cosmetic or pharmaceutical compositions comprising in a physiologically acceptable support at least one peptide of the invention, in particular as an active ingredient, more specifically as an antioxidant, anti-allergy and/or anti-inflammatory agent. The pharmaceutical composition is preferably a dermatological or aerosol composition.

The term physiologically, or preferably cosmetically, dermato logically, or ear, nose or throat, acceptable carrier or support, as used herein, means that the compositions are suitable for use in contact with human, in particular human skin and mucosae, without risk of toxicity, incompatibility, instability, allergic response, and the like.

The compositions of the invention can be administered according to various routes, via the oral route, the nasal route, with a buccal or enteric absorption, via the mucous membrane, subcutaneous, intramuscular route, the transdermal route (as a patch or gel) or topically onto the skin, or through inhalation.

Preferably, the peptides of the invention are administered topically on the skin using cosmetic or dermatologic compositions, or are inhaled through aerosol or similar compositions.

The peptides of the invention can be topically applied in admixture with suitable cosmetic or dermatological compounds (excipients, stabilizers, etc.). Such compounds are selected in accordance with the intended form, i.e. creams, moisturizers, lotions, pomades, ointments, and the like, and consistent with conventional cosmetic or dermatological practices. The peptides may thus be formulated in various forms, including solid, semisolid and liquid forms, such as tablets, capsules, gel, emulsions, etc.

The peptides agents of the present invention can also be delivered via oral inhalation or intranasal administration. Appropriate dosage forms for such administration, such as an aerosol formulation or a metered dose inhaler, may be prepared by conventional techniques.

For administration by inhalation, the peptides may be delivered in the form of an aerosol spray presentation from pressurized packs or a nebulizer, with the use of a suitable propellant, e.g. dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, a hydrofluoroalkane such as tetrafluoroethane or heptafluoropropane, carbon dioxide or other suitable gas. In the case of a pressurized aerosol, the dosage unit may be determined by providing a valve to deliver a metered amount. Capsules and cartridges of e.g. gelatin for use in an inhaler or insufflator may be formulated containing a powder mix of a peptide of the invention and a suitable powder base such as lactose or starch. Additionally, the compositions of the present invention may be formulated in a sustained, long acting release form to provide the rate controlled release of any one or more of the components or active ingredients to optimize the antioxidant and/or anti-inflammatory effects of peptides. Suitable dosage forms for sustained release include liposomes or controlled release polymeric matrices impregnated with the active components.

The peptides of the invention can be used in a substantially similar manner to other known antioxidant or anti-allergy agents. For the peptides of the invention, the dose to be administered can vary depending on the particular peptide employed because of the varying antioxidant and/or antiallergen potency of the peptides. The dosage to be administered is not subject to definite bounds, but it will usually be an effective amount, or the equivalent on a molar basis of the redox active free form (e.g. mM concentration). Someone skilled in the art of cosmetics and pharmaceutical compositions will be able to ascertain, without undue experimentation, appropriate protocols for the effective delivery of the peptides or derivatives of this present invention, such as by referring to the earlier published studies on antioxidants. Without limiting the invention, the composition preferably comprises from 0.001 to 40%, more preferably from 0.01 to 10%, by weight of the peptide of the invention with respect to the total weight of the composition.

The peptides of the invention can be used alone or in combination with other active compounds, including natural antioxidant agents (e.g. coenzyme Q10, vitamin(s) C and/or

E) and/or other anti-allergy agents.

The peptides set forth herein may also be used as laboratory reagents. The peptide of the present invention may be employed as a redox couple to favour in vitro oxidative folding of reduced peptides/proteins, similar to the use of redox glutathione (GSH/GSSG pair). The peptide of the present invention may also be employed in in vitro assays to identify or assess a variety of compounds as potential allergens. Thus, the invention relates to a use of at least one peptide as defined in the invention as a tool (a molecule) to detect allergens in an in vitro assay. According to this assay, where one of the used peptide of the invention interacts with the test compounds, said compounds are identified or determined as potential allergens. The test compounds can be of any type, such as a natural or synthetic compound or a mixture of compounds, including plant or animal extracts and parts. Such compounds may come from chemicals, perfumes, pollen, mold spores, plants, dander, dust, foods, drugs (e.g. antibiotics, aspirin), insect venoms (e.g. wasps), parasites, viruses, bacteria, animals, humans, or the like.

The invention also deals with a method, preferably a cosmetic or dermatologic process, to prevent or treat skin damage due to oxidative stress, in particular by enhancing the protection against ultraviolet (UV) exposure (photo -protection), environmental damage, skin inflammation, irritation or reddening and UV-induced DNA damage, by applying topically onto the skin or hair an efficient amount of the peptide of the invention or a composition comprising the same. It also deals with a process, more specifically a cosmetic process, of treatment of ageing skins and/or fighting in a curative and/or preventive manner the phenomena of cutaneous ageing comprising applying to the surface of the skin or hair an effective amount of the peptide such as defined in the present invention or a composition comprising the same. Moreover, it concerns a process, more specifically a cosmetic process, of treatment of skin and/or hair against any type of external aggressions, especially UV and/or other environmental damages, comprising applying to the skin surface an effective amount of the peptide such as defined above or a composition comprising said peptide.

Another embodiment of the invention discloses the use of a peptide defined above for the preparation of composition, in particular a dermatological composition, for the treatment of a disorder or disease due or related to an oxidative stress, in particular cancer or inflammatory disorder or disease, and more specifically skin cancer or psoriasis. Peptides of the invention, more specifically peptide of Seq ID n° 4, were tested for cell viability on several colonic cancer cell lines that produced either high or low amounts of ROS. The peptides of the invention were found to strongly and specifically inhibit cell lines that highly produced ROS. It appears that such induced inhibition of cell proliferation is dose-dependent, with an activity obtained in the low micromolar concentration range. In this system, peptides of the invention, and more specifically peptide of seq id n° 4, were found to be more potent than other bioactive compounds, including some of the reference antioxidants (e.g. Trolox or also called 6-hydroxy-2,5,7,8-tetramethylchroman-2- carboxylic acid).

The peptides of the invention are particularly suited for the treatment of cancers, such as solid tumors or lymphoid tumors. Specific examples include prostate cancer, ovarian cancer, pancreas cancer, lung cancer, breast cancer, liver cancer, head and neck cancer, colon cancer, bladder cancer, non-Hodgkin's lymphoma cancer, leukemia (acute lymphoblastic leukemia, chronic myeloid leukemia, acute myeloid leukemia) and melanoma.

Another embodiment of the invention discloses the use of a peptide defined above for the preparation of a pharmaceutical composition, in particular a dermatological composition, for the treatment of a disorder or disease due or related to an allergy (or allergen-induced reactions), including allergy-related inflammatory disorder or disease.

The present invention relates to a method for the treatment of a disorder or disease, and in particular a disorder or disease related or due to an oxidative stress, in particular cancer or inflammatory disorder or disease, and more specifically skin cancer or psoriasis, comprising administering to a mammal (in particular a patient) in need of such treatment an effective amount of at least one peptide of the present invention as defined herein.

The invention also deals with a method for the treatment of a disorder or disease due or related to an allergy (or allergen-induced reactions), including allergy-related inflammatory disorder or disease, allergy, allergy-related irritation and/or inflammation, comprising the administration to a mammal (preferably an animal or human being) in need thereof of an efficient amount of the peptide of the invention or a composition comprising the same, in particular in enhancing protection by trapping or neutralizing allergen(s). Preferably, the method is a dermatological method to treat skin allergy, allergy-related irritation, inflammation and/or reddening, by applying topically onto the skin of a subject in need thereof an efficient amount of the peptide of the invention or a composition comprising the same, in particular in enhancing skin protection by trapping or neutralizing allergen(s). It also deals with a method, more specifically a dermatologic method, of treatment of allergy and/or fighting the phenomena of allergy comprising applying to the surface of the skin an effective amount of the peptide such as defined in the present invention or a composition comprising the same. Moreover, it concerns a dermatological method for treatment of skin against any type of external aggressions, especially the contact with one or more allergenic substance(s), comprising applying to the skin surface an effective amount of the peptide such as defined above or a composition comprising said peptide.

As mentioned before, allergic reactions are caused by allergens, which are normally harmless environmental compounds. Allergy (type I hypersensitivity) can be characterized by an excessive activation of white blood cells (i.e. basophils and mast cells) by a type of immunoglobulin called IgE. This excessive activation of cells results in a severe inflammatory response. Peptides of the invention by trapping allergen(s) allow allergic reactions to be diminished, abolished or prevented.

Among disorders or diseases due or related to allergy are contact dermatitis, asthma, eczema (atopic dermatitis), hay fever (allergic rhinitis), hives (urticaria), food allergies, etc. Anaphylaxis is the most severe, whole body and life-threatening type of allergic reaction. Among allergens (substances causing allergic reactions) are found chemicals, perfumes, pollen, mold spores, plants, dander, dust, foods, drugs (e.g. antibiotics, aspirin), insect venoms (e.g. wasps), parasites, viruses and bacteria. Most environmental allergens contact the skin or eyes, or are inhaled, explaining why most symptoms are affecting the skin, eyes, or the airways. According to the invention, 'treatment' or 'treating' includes both curative and preventive treatments. Accordingly, and as curative treatment, the compounds may be used at very early stages of damages due or related to an oxidative stress and/or allergen-induced reactions, in particular at very early stages of a damaged skin due to an oxidative stress and/or allergen-induced reactions, or before early onset, or after significant progression. Peptides of the invention can be used in humans with existing damaged state, including at early or late stages of damaged state. The peptides of the invention will not necessarily treat the subject who has already damaged condition, in particular damaged skin (for instance aged skin with wrinkles) but will delay or slow the progression or prevent further skin damages due to UV, environment, or to a contact with allergenic substance(s), ameliorating thereby the subject's conditions (more specifically skin conditions). By delaying the onset of allergy (more specifically skin allergy) or ageing skin, peptides of the invention have prevented the individual from getting damaged condition (more specifically damaged skin) during the period in which the individual would normally have gotten a damaged condition (more specifically damaged skin) or reduce the rate of development of the damaged condition (more specifically damaged skin) or some of its effects but for the administration (more specifically topical administration) of compounds of the invention up to the time the individual ultimately gets damaged condition (more specifically damaged skin). In treating the above conditions, the peptides of the invention are administered (more specifically topically) in an effective amount.

As an alternative, the peptides of the invention can also be applied topically on the skin or hair as a preventive treatment to those who do not have a damaged skin or hair yet.

The following are examples provided solely to illustrate the present invention and are not intended to limit the scope of the invention, as described herein. EXAMPLES

Chemical Synthesis of peptides of the invention

Na-fluoren-9-ylmethyloxycarbonyl (Fmoc)-L-amino acids and reagents used for peptide synthesis were obtained from Perkin-Elmer (Shelton, CT, U.S.A.). The Wang resin was purchased from Advanced ChemTech (Cambridge, U.K.). Solvents were analytical grade products from SDS (Peypin, France). The peptides were produced by chemical solid-phase synthesis using an automated peptide synthesizer (Model 433A, Applied Biosystems Inc.). In all cases, peptide chains were assembled stepwise on 0.50 mmol of Fmoc-amide resin (0.68 mmol equivalent of amino group/g) using 1 mmol of N-(9-fluorenyl)methyloxycarbonyl(Fmoc) amino acid derivatives. The side chain-protecting groups used for trifunctional residues were: trityl

(Trt) for Cys; tert-butyl (t-Bu) for Asp and Glu; 2,2,4,6,7-pentamethyldihydrobenzofuran- 5-sulfonyl (Pbf) for Arg; and tert-butyloxycarbonyl (Boc) for Lys. N-amino groups were deprotected by successively treating with 18 and 20% (v/v) piperidine/N- methylpyrrolidone for 3 and 8 min, respectively. After three washes with N- methylpyrrolidone, the Fmoc-amino acid derivatives were coupled (20 min) as their hydroxybenzotriazole active esters in N-methylpyrrolidone (2.5-fold excess). After complete assembly of the peptides, and removal of their N-terminal Fmoc groups, the peptide resins (from 1.2 to 1.6 g) were treated under stirring for 2h at 25°C with mixtures of TFA (trifluoroacetic acid)/water/thioanisole/ethanedithiol (88:5:5:2, by vol.) in the presence of crystalline phenol (0.5 g) in final volumes of 30 ml per gram of peptide resins.

The peptide mixtures were filtered, precipitated and washed twice with cold diethyloxide. The crude peptides were pelleted by centrifugation (3,000g ; 10 min). The crude peptides were then dissolved in water and freeze dried. Peptides were purified to homogeneity by reversed-phase HPLC (Phenomenex, CI 8 jupiter 5 μιη, 250 mm x 10 mm) by means of a 90-min linear gradient of 0.08% (v/v) TF A/0-30% acetonitrile in 0.1% (v/v) TFA/water at a flow rate of 3 ml/min (λ= 230 nm). The purity and identity of the peptides were assessed by analytical CI 8 reversed-phase HPLC (Phenomenex, CI 8 Onyx monolithic 5 μιη, 100 mm x 4.6 mm) using a 40 min linear gradient of 0.08% (v/v) TF A/0-60% acetonitrile in 0.1% (v/v) TFA/water at a flow rate of 1 ml/min, and molecular mass determination by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry. Peptide reactivity with potential allergenic substances

Reactivity of compounds (potential allergens) with peptides of the invention is gathered the following table. The results are expressed as a percentage of peptide depletion.

Δ ORP (mV) of the peptide is measured as mentioned above.

Peptide reactivity with potential allergenic substances was assessed by measuring peptide depletion using Cis-reversed-phase high pressure liquid chromatography (HPLC) upon a 24h incubation period of the peptide with potential allergens (25°C in the dark). The reaction medium (final volume of 100 μί) consisted of 40 μΙ_, of peptide solution at 1.25mM concentration, 35 of 100 mM Tris-HCl buffer at pH 8.3, 20 acetronitrile, and 5 μί of potential allergen solution (solution at 100 mM potential allergen dissolved in acetronitrile (ACN) or acetronitrile :dimethylsulfoxide (DMSO) 50:50 (v/v) depending on product solubility). HPLC analyses of reaction media were carried out on C₁₈ Onyx monolith 100 column (4.6 mm x 150 mm) with a 20-min linear gradient of 0.08% (v/v) trifiuoroacetic acid, 0-60% acetronitrile in 0.1% (v/v) trifiuoroacetic acid, ¾0 at a flow rate of lml/min (λ = 230 nm). Analyses of eluted products (i.e. peptide, peptide-allergen complex (in particular covalent complex), and 'unreactive' product) were achieved by mass spectrometry.

The asterisk (*) indicates an amidated C-terminus of peptide; NS corresponds to a non- sensitizing compound; (-) stands for 'not determined'. In the Table, a value of ' 100' indicates a full reactivity of the peptide with the allergen, which corresponds to 100% peptide depletion under the experimental conditions used.

Claims

1. A peptide comprising the following sequence (seq. ID n°l): VYMNR YYKL, or variants thereof defined as comprising seq. ID n°l with deletion, substitution, and/or addition of one, two, three or more amino acids therein and having from nine amino acid residues to twenty amino acid residues in length.

2. The peptide of claim 1, wherein it comprises the following sequence seq. ID n°2:

XI -X2-YM-X3-NR-X4-KY-X5-K-X6-X7

wherein:

XI is vacant, or is an amino acid sequence consisting of RCR,

X2 is vacant, or V;

X3 is vacant, or is an amino acid sequence consisting of RG;

X4 is vacant, or C;

X5 is vacant, or Y;

X6 is vacant or L;

X7 is vacant, or an amino acid sequence selected in the group consisting of RCR, CCK or CLK.

3. The peptide of claim 1 or 2, wherein it is from nine amino acid residues to twenty amino acid residues in length, preferably from ten to thirteen amino acid residues in length.

4. The peptide of one of claims 1-3, wherein it comprises one of the following peptide sequences:

V YMNRKY YKC CK (SEQ ID n° 3)

VYMNRKYYKL (SEQ ID n° 1)

RCRVYMNRKYYKL (SEQ ID n° 4)

VYMNRKYYKLCLK (SEQ ID n° 5)

VYMNRCKYYKL (SEQ ID n° 6)

YMNRKYKRCR (SEQ ID n° 7)

VYMRGNRKYYKL (SEQ ID n° 8).

5. A cosmetic or pharmaceutical composition comprising at least one peptide as defined in any of the preceding claims in a physiologically acceptable support.

6. The composition according to claim 5, wherein the peptide is in combination with at least one other active compound, including a natural antioxidant agent and/or another anti- allergy agent.

7. The composition according to claim 5 or 6, useful for the treatment of a disorder or disease due or related to an oxidative stress and/or allergen-induced reactions.

8. The composition according to anyone of claims 5-7, useful for the treatment of cancer or inflammatory disorder or disease, and more specifically skin cancer or psoriasis.

9. The composition according to anyone of claims 5-7, useful for the treatment of skin damage due to an allergic reaction, in particular allergy-related inflammation, irritation or reddening of skin.

10. The composition according to anyone of claims 5-7, useful for the treatment of respiratory disorders due to allergic reactions, in particular hay-fever or asthma.

11. The composition according to anyone of claims 5-7, useful for the treatment of skin damage due to oxidative stress, in particular enhancing thereby the protection against ultraviolet exposure, or treating environmental damage, skin inflammation, irritation or reddening and/or UV-induced DNA damage.

12. A use of at least one peptide as defined in any of the preceding claims 1-4, as a redox couple to favour in vitro oxidative folding of reduced peptides/proteins.

13. A use of at least one peptide as defined in any of the preceding claims 1-4, as a tool to identify or assess allergens in an in vitro assay.