Peptide, composition comprising said peptide and uses thereof, in particular cosmetic uses

Abstract

The peptide has from 3 to 10 amino acids comprising at least the sequence K*(Ac)GH or K*(Ac)HG and may further comprise an N-terminus modification, preferably an acylation, and/or a C-terminus modification; K* is selected from the group consisting of lysine, ornithine, diaminobutyric acid, diaminopropionic acid and a hydroxylated derivative thereof; K*(Ac) corresponds to a lysine, ornithine, diaminobutyric acid, diaminopropionic acid or a hydroxylated derivative thereof, acetylated on the amine of their lateral hydrocarbon chain. The two preferred peptides are Pal-K(Ac)GH and Pal-K(Ac)HG. This peptide can be used for a cosmetic treatment, in particular anti-aging, anti-wrinkle and fine lines, to improve the mechanical properties of the skin, firmness/tonicity/elasticity/flexibility, to increase the density and volume of the skin, for a restructuring, healing effect, and/or to fight stretch marks.

Claims

1. A peptide having a length of from 3 to 6 amino acids and which comprises at least one peptide sequence K*(Ac)GH or K*(Ac)HG, wherein K* is selected from the group consisting of lysine, ornithine, diaminobutyric acid, diaminopropionic acid, and a hydroxylated derivative thereof; K*(Ac) represents a lysine, ornithine, diaminobutyric acid, diaminopropionic acid or a hydroxylated derivative thereof, which is acetylated on the amine of the lateral hydrocarbon chain thereof; the amino acids of the peptide other than K*(Ac)GH or K*(Ac)HG, when present, are each independently selected from the group consisting of lysine, histidine, glycine, alanine, and K*; the peptide is modified at the N-terminal with a modification selected from the group consisting of —CO—R.sub.1 and —SO.sub.2—R.sub.1; and/or the peptide is modified at the C-terminal with a modification selected from the group consisting of —OR.sub.1, —NH.sub.1, —NH.sub.1R.sub.2, and —NR.sub.1R.sub.2; and R.sub.1 and R.sub.2, when present, independently of one another, are chosen from an alkyl, aryl, aralkyl, alkylaryl, alkoxy and aryloxy group, which is linear, branched, cyclic, polycyclic, unsaturated, hydroxylated, carbonylated, phosphorylated and/or sulfurated, said group having from 1 to 24 carbon atoms and may optionally have in its carbon backbone an O, S and/or N heteroatom.

2. The peptide of claim 1, wherein K* is lysine and K*(Ac) represents an s-acetylated lysine.

3. The peptide of claim 1, wherein the R.sub.1 and/or R.sub.2 group comprises from 3 to 24 carbon atoms.

4. The peptide of claim 1, which is modified at the N-terminal with —CO-R.sub.1.

5. The peptide of claim 4, wherein —CO—R.sub.1 is an octanoyl, decanoyl, lauroyl, myristoyl, palmitoyl, stearoyl, biotinoyl, elaidoyl, oleoyl or lipoyl group.

6. The peptide of claim 1, which is modified at the N-terminal and not modified at the C terminal.

7. The peptide of claim 1, represented by the formula (1) or (2):
X-K*(Ac)GH-Z   (1)
or X-K*(Ac)HG-Z   (2) wherein X represents, when present, the modification at the N-terminal, and Z represents, when present, the modification at the C-terminal.

8. The peptide of claim 7, which is Pal-K(Ac)GH or Pal-K(Ac)HG.

9. A cosmetic composition comprising as an active ingredient an effective amount of at least one peptide of claim 1 in a physiologically acceptable medium.

10. The composition of claim 9, further comprising at least one additional active ingredient selected from the group consisting of vitamin B3 compounds, niacinamide, tocopherol, retinoid compounds, hexamidine, a-lipoic acid, resveratrol, DHEA, hyaluronic acid, and peptides.

11. A method for a non-therapeutic cosmetic treatment to improve the general condition of the skin and/or its appendages and to treat their imperfections, comprising applying to the skin and/or its appendages at least one peptide of claim 1.

12. The method of claim 11, wherein the at least one peptide is applied topically to the skin and/or its appendages.

13. The method of claim 11, wherein the non-therapeutic cosmetic treatment is an anti-aging treatment of the skin and/or its appendages.

14. The method of claim 13, wherein the anti-aging treatment is for a treatment: of wrinkles and fine lines; and/or for improving the mechanical properties of the skin; and/or for increasing the density and volume of the skin; and/or for fighting stretch marks; and/or for improving the homogeneity and/or radiance of complexion.

15. The method of claim 11, further comprising applying to the skin and/or its appendages at least one additional active ingredient selected from the group consisting of vitamin B3 compounds, niacinamide, tocopherol, retinoid compounds, hexamidine, a-lipoic acid, resveratrol, DHEA, hyaluronic acid, and peptides.

Description

DETAILED DESCRIPTION

(1) The following examples describe and illustrate certain aspects of the invention.

(2) A) Example of Manufacturing of a Peptide According to the Invention: the Pal-K(Ac)HG

(3) The peptide Pal-K(Ac)HG is prepared by peptide synthesis. The glycine is coupled with a resin via its terminal acid function (with a coupling agent for example DCC (diclyclohexylcarbodiimide)/NHS (N-hydroxysuccinimide) or HBTU (2-(1H-benzotriazol-1-yl) 3,3-tetramethyluronium hexafluorophosphate)/HOBT (1-hydroxy-benzotriazole)). The glycine thus protected is then reacted with a derivative of histidine in the presence of a coupling agent, and then the same operation is carried out to add the previously acetylated lysine. The latter is then acylated on its amine function with an activated palmitic acid derivative (palmitoyl chloride for example) in the presence of a base. After cleavage of the peptide resin, precipitation of the peptide, washing and drying, the acetylated palmitoyl-lysyl-glycyl-histidine product is obtained in solid form.

(4) This procedure is applicable to obtain Pal-K(Ac)HG by initially inverting glycine and histidine.

(5) B) Preparation of a Composition According to the Invention Comprising the Pal-K(Ac)HG Peptide of l'example A) or the Pal-K(Ac)GH peptide

(6) Starting Materials: The pure peptide, synthesized according to the synthesis method explained above; Excipient: mixture of fatty esters, chosen in order to form an oily matrix, for example intended to form a anhydrous composition water for the subsequent formulation of anhydrous cosmetic compositions.

(7) Procedure: The peptide is mixed with the excipient and stirred gently and heated until solubilization and total clarity.

(8) C) In Vitro Tests

(9) The peptides according to the invention have a number of remarkable effects presented below. Peptides prepared according to A) above and dissolved in an excipient were in vitro tested and showed activities which are presented hereinafter.

(10) 1) ELISA Assays

(11) Protocol

(12) Cultured normal human fibroblasts (NHF) are brought into contact with the test products or their excipient (negative control) for 72 hours. At the end of the contact, the culture supernatants are removed and the syntheses of the dermal macromolecules are estimated by ELISA assays. An estimation of the cell viability is carried out by Hoechst assay and makes it possible to weigh the data obtained.

(13) Results for the Pal-K(Ac)HG

(14) TABLE-US-00002 TABLE 1 Collagen I % change/control (significance - Student test) Concentration Pal-KHG Pal-K(Ac)HG 7 ppm /  +40 (p < 0.05) 10 ppm  +41 (p < 0.05) +138 (p < 0.01) 12.5 ppm +123 (p < 0.01) / 15 ppm / +241 (p < 0.01)

(15) TABLE-US-00003 TABLE 2 % change/control Collagen IV (significance-Student test) Concentration Pal-KHG Pal-K(Ac)HG 10 ppm  +45 (p < 0.01)  +67 (p < 0.01) 12.5 ppm   +141 (p < 0.01) / 15 ppm / +155 (p < 0.01)

(16) TABLE-US-00004 TABLE 3 % change/control Fibronectin (significance-Student test) Concentration Pal-KHG Pal-K(Ac)HG 10 ppm +128 (p < 0.01) +128 (p < 0.01)

(17) TABLE-US-00005 TABLE 4 % change/control Hyaluronic acid (significance-Student test) Concentration Pal-KHG Pal-K(Ac)HG 3 ppm −14 (nsd) +259 (p < 0.01) 7 ppm  +4 (nsd) +142 (p < 0.01) 10 ppm  +11 (nsd) +123 (p < 0.01) 12.5 ppm   +33 (p < 0.05) +119 (p < 0.01) nsd: non significant data

(18) The results show that the Pal-K(Ac)HG peptide according to the invention stimulates the synthesis of collagens I and IV, fibronectin and hyaluronic acid on normal human fibroblasts at concentrations of a few ppm and in significant proportions. The results also show that the Pal-K(Ac)HG peptide according to the invention is advantageously more active than its non-acetylated version on lysine (Pal-KHG) for collagens I and IV, and hyaluronic acid very largely from 3 ppm, whereas there is no decreased activity on fibronectin.

(19) Results for the Pal-K(Ac)GH

(20) TABLE-US-00006 TABLE 5 % change/control Collagen I (significance-Student test) Concentration Pal-KGH Pal-K(Ac)GH 3 ppm +81 (p < 0.01) +71 (p < 0.01) 5 ppm / +75 (p < 0.01) 7 ppm +69 (p < 0.05) +119 (p < 0.01)  10 ppm  +123 (p < 0.01)  /

(21) TABLE-US-00007 TABLE 6 % change/control Collagen IV (significance-Student test) Concentration Pal-KGH Pal-K(Ac)GH 10 ppm +59 (p < 0.01) +80 (p < 0.01)

(22) TABLE-US-00008 TABLE 7 % change/control Fibronectin (significance-Student test) Concentration Pal-KGH Pal-K(Ac) GH   10 ppm +45(p < 0.01) / 12.5 ppm / +35 (p < 0.01)

(23) TABLE-US-00009 TABLE 8 % change/control Elastin (significance-Student test) Concentration Pal-KGH Pal-K(Ac) GH  7 ppm −62 (p < 0.05) / 10 ppm / +207 (p < 0.01)

(24) TABLE-US-00010 TABLE 9 % change/control Hyaluronic acid (significance-Student test) Concentration Pal-KGH Pal-K(Ac)GH 3 ppm / +111 (p < 0.01) 7 ppm +113 (p < 0.05) +244 (p < 0.01) 10 ppm  +118 (p < 0.05) +260 (p < 0.01) 12.5 ppm   / +437 (p < 0.01)

(25) The results show that the peptide Pal-K(Ac)GH according to the invention stimulates the synthesis of collagens I and IV, fibronectin, elastin and hyaluronic acid on normal human fibroblasts at concentrations of a few ppm and in significant proportions. The results also show that the Pal-K(Ac)GH peptide according to the invention is advantageously more active than its non-acetylated version on lysine (the Pal-KGH) for collagens I (at 7 ppm) and IV (at 10 ppm), elastin above 10 ppm, and hyaluronic acid very largely from 3 ppm.

(26) 2) Immunofluorescence Assays

(27) Protocol

(28) Normal Human Fibroblasts (NHF) are grown for 24 h. The cells are placed in contact with the test products or their excipient at different concentrations for 6 days for collagen I or 14 days for elastin (DMEMc 5% FCS). The synthesis of collagen I and elastin produced by the cells in the form of extracellular matrix is then quantified by immuno-marking on the attached layers. A count of the Hoechst-labeled nuclei is performed in parallel in order to have an estimate of the viability and to weight the data.

(29) Results

(30) TABLE-US-00011 TABLE 10 % change/control Collagen I (significance-Student test) Concentration Pal-KHG-OH Pal-K(Ac)HG-OH  3 ppm +20 (p < 0.05) +42 (p < 0.01)  7 ppm +34 (p < 0.01) / 10 ppm / +45 (p < 0.01) 15 ppm / +120 (p < 0.01) 

(31) TABLE-US-00012 TABLE 11 % change/control Collagen IV (significance-Student test) Concentration Pal-KHG-OH Pal-K(Ac)HG-OH 10 ppm +217 (p < 0.01) +231 (p < 0.01)

(32) TABLE-US-00013 TABLE 12 % change/control Elastin (significance-Student test) Concentration Pal-KHG-OH Pal-K(Ac)HG-OH 3 ppm / +326 (p < 0.01) 7 ppm / +438 (p < 0.01) 10 ppm  / +221 (p < 0.01)

(33) The results confirm that the Pal-K(Ac)HG peptide according to the invention stimulates the synthesis of collagens I and IV and show that it strongly stimulate the synthesis of elastin. The results also confirm that the Pal-K(Ac)HG peptide according to the invention is advantageously more active than its non-acetylated version on lysine (Pal-KHG) for collagens I and IV and advantageously exhibits very high activity on the elastin whereas the non-acetylated version had no activity on this target.

(34) D) GALENIC

(35) Various formulations are described below. Additional cosmetic active ingredients, if appropriate in support and/or in addition to the activity of the active ingredient according to the invention, may be added in the appropriate phase according to their hydrophobic or hydrophilic nature. These ingredients can be of any category according to their function(s), the place of application (body, face, neck, bust, hands, hair, eyelashes, eyebrows, hair, etc.) and targeted consumer, for example anti-oxidant, moisturizing, nourishing, protective, smoothing, remodeling, volumizing (lipofiling), acting on the shine of the complexion, against the spots, anti dark circles, anti-glycation, myorelaxing, anti-redness, anti-stretch marks, etc. They are mentioned above in the description.

(36) 1) Cream Form, for Example an Antiaging Day Cream for the Face

(37) TABLE-US-00014 Ingredient (INCI name) Weight % Phase A Sorbitan Stearate 3.00 Cyclopentasiloxane (and) Cyclohexasiloxane 2.00 Ethylhexyl Palmitate 3.00 Glyceryl Stearate (and) PEG-100 Stearate 3.00 Ethylhexyl Methoxycinnamate 1.00 Ethylhexyl Dimethyl PABA 1.00 Phase B Demineralized water Qsp 100 Ultrez 10 (Carbomer) 0.40 Phase C Glycerin 5.00 Preservative qs Phase D Peptide according to the invention in a fatty excipient 3.00 Phase E Potassium Sorbate 0.10 Phase F Sodium Hydroxide 30% 0.60 Demineralized water 6.00 Phase G Perfume 0.10

(38) Protocol: Weigh phase A and heat it at 75° C. in a water bath. Weigh phase B and let it swell for 20 minutes. Melt phase C until dissolved and add it to phase B. Heat phase (B+C) at 75° C. in a water bath. Pour phase A into the phase (B+C) under Staro stirring. Extemporaneously add phase D to phase (A+B+C). At about 45° C. add phase E and neutralize with phase F. Homogenize well. At 35° C., add phase G. Homogenize well. PH: 6.20.

(39) Example of Ingredients that can be Added to this Formulation: CALMOSENSINE™: soothing active for sensitive skins comprising the Tyr-Arg lipo-dipeptide. It reduces discomfort feelings. SEBULESS™: purifying sebo-regulator ingredient comprising a Syringa vulgaris extract, which mattifies and refreshes complexion, fades the inflammatory blemishes. PRODIZIA™: active ingredient comprising an extract of Albizia julibrissin, fighting the signs cutaneous fatigue: dark circles, under eye bags, dull complexion and drawn features, by repairing and protection the skin against the caused by damages of glycation and glycoxydation. PACIFEEL™: active ingredient comprising a natural extract of the Mirabilis jalapa plant also known as the Marvel of Peru, which alleviates cutaneous discomfort, fades redness of sensitive and reactive skin and strengthens and hydrates the epidermis. MAJESTEM™: active agent based on plant cells obtained by in vitro cell culture titrated in leontopodic acid; tightens the sagging neck skin, lifts the cheeks smoothes out wrinkles around the eyes, especially crow's feet wrinkles.

(40) 2) Gel Form, for Example a Firming Gel for the Body

(41) TABLE-US-00015 Weight Ingredient (INCI name) % Phase A Demineralized water Qsp 100 Ultrez 10 (Carbomer) 0.20 Phase B PEG 400 5.00 Preservatives qs Phase C Dimethicone 4.00 Pemulen TR2 (Acrylates/C10-30 Alkyl Acrylate Cross Polymer) 0.20 Phase D Tween 20 (Polysorbate 20) 1.00 Peptide of the invention in a fatty excipient 2.00 Phase E Potassium Sorbate 0.10 Phase F Sodium hydroxide30% 0.60 Demineralized water 5.00 Phase G Perfume 0.10

(42) Protocol: Disperse Ultrez 10 in water and let it swell for 15 minutes. Heat phase B until dissolved and add it to phase A. Weigh and mix phase C. Mix phase D and add it to phase C; homogenize thoroughly. Add phase (C+D) to phase (A+B). Then add phase E. Leave to swell for 1 hour. Homogenize thoroughly. Neutralize with phase F. Finally, add phase G. pH: 6.10.

(43) Example of Ingredients that can be Added to this Formulation: AQUALANCE™: osmo-protector moisturising active ingredient comprising homarine and erythritol. LEGANCE™: anti-aging active marketed by Sederma, corresponding to a Zingiber zerumbet Smith extract obtained by CO.sub.2 supercritical in a water-soluble excipient and titrated in zerumbone ingredient. It is a global anti-aging ingredient for legs. It improves their appearance and comfort by reducing water retention, improving microcirculation and refining adipose tissue. BODYFIT™: slimming/firming active ingredient comprising glaucine marketed by Sederma. BODYFIT™ reduces the appearance of cellulite and helps to improve drainage and water distribution in the tissues. JUVINITY™: active marketed by Sederma reducing signs of aging on the face and neckline, smoothing wrinkles, densifying and restructuring the dermis.

(44) 3) Compact Powder Form

(45) TABLE-US-00016 Ingredient (INCI name) Weight % Phase A Talc Qsp 100 Kaolin 2.00 Calcium Stearate 1.00 Mica 4.00 Silica 1.00 Bismuth Oxychloride 2.00 Potassium Sorbate qs Phenoxyethanol qs Phase B Unipure Black LC 989 HLC [CI 77499 (and) Hydrogenated 0.20 Lecithin] Unipure Red LC 381 HLC [CI 77491 (and) Hydrogenated 0.60 Lecithin] Unipure Yellow LC 182 HLC [CI 77492 (and) Hydrogenated 1.00 Lecithin] Covapearl Star Gold 2302 AS [CI 77891 (and) CI 77491 (and) 0.50 Synthetic Fluorphlogopite (and) Triethoxycaprylylsilane] Covapearl Brown 838 HLC [CI 77491 (and) Mica (and) 1.00 Hydrogenated Lecithin) Covapearl Dark Blue 637 [CI 77510 (&) CI 77891 (&) Mica] 0.10 Phase C Crodamol PTIS-LQ-(MV) [Pentaerythrityl Tetraisostearate] 4.00 Peptide of the invention in a fatty matrix 3.00 Phase D Perfume 0.30

(46) Protocol: Weigh phase A and mix. Weigh phase B and pour it into phase B. Pour A+B into the mixer and mix. Add phase C to A+B in several times and mix each time. Add phase D. Check homogeneity at each step.

(47) Example of Ingredients that can be Added to this Formulation: VEGESOME MOIST 24™: ingredient marketed by SEDERMA designed for the formulation of moisturizing powder makeup ; it is a powder consisting of hollow particles 25 microns (Lycopodium clavatum exins) loaded with an Imperata cylindrica extract having moisturizing properties.

(48) 4) Other Cream Form (Face or Body)

(49) TABLE-US-00017 Ingredient (INCI name) Weight % Phase A Arlacel 170 (Glyceryl Stearate (and) PEG-100 Stearate) 5.50 Abil Wax 2434 (Stearoxy Dimethicone) 3.00 Acetulan (Cetyl Acetate (and) Acetylated Lanolin Alcohol) 1.50 Crodacol C 90 (Cetyl Alcohol) 1.50 Mineral Oil 3.00 Shea Butter 5.00 Unsaponifiable Shea 1.00 Parsol MCX (Ethylhexyl Methoxicinnamate) 3.50 Phase B Demineralized water Qs 100 Phase C Carbopol 940 (Carbomer) 0.20 Phase D Demineralized water 2.00 Triethanolamine 99% 0.20 Phase E Propylene Glycol 0.10 Mixed Parabens Phase F Sodium Hydroxide 30% 5.00 Demineralized water qs Phase G Peptide according to the invention in an hydrophilic matric 2.00

(50) Protocol: Weigh phase A and heat it at 75° C. in a water bath. Weigh phase B and let it swell for 20 minutes. Melt phase C until dissolved and add it to phase B. Heat phase (B+C) at 75° C. using a water bath. Pour phase A into the (B+C) phase under Staro stirring. Extemporaneously add phase D to phase (A+B+C). At about 45° C. add phase E and neutralize with phase F. Homogenize well. At 35° C., add phase G. Homogenize well. PH: 6.20.

(51) Example of Ingredients that can be Added to this Formulation: SUBLISKIN™: active ingredient that moisturizes and smooths the skin while allowing it to resist to external aggressions. VENUCEANE™: active marketed by Sederma comprising a Thermus thermophiles biotechnological extract, that prevents visible signs of photo-aging (spots, wrinkles, dryness . . . ), protects cell structures from damages caused by UV and strengthens skin integrity. KOMBUCHKA™: active ingredient acting on complexion marketed by Sederma. INTENSLIM™: slimming active ingredient marketed by Sederma corresponding to a synergistic combination of extracts obtained by Globularia cordifolia plant cell culture, Zingiber zerumbet Smith titrated in zerumbone and vegetable caffeine obtained by supercritical CO.sub.2 extraction. CITYSTEM™: active ingredient based on plant cells obtained in vitro from Marrubium vulgare with a high concentration of Forsythoside B; used against the attacks of pollution: makes the skin soft and smooth, refines the skin texture, reduces the visibility of comedones, leaving the skin radiant and purified.