Peptide containing hydrogel compositions

Abstract

There is provided a hydrogel composition comprising a hydrogel system and a peptide; wherein the hydrogel system comprises; one or more thickener, and a solvent system comprising one or more organic solvent and one or more surfactant. Also provided are methods of making said hydrogel composition, methods and uses of said hydrogel composition, and the use of said hydrogel composition in a method of treating wounds to the skin of a subject.

Description

[0001] Peptide Containing Hydrogel Compositions

[0002] Field of the Invention

[0003] The present invention relates to hydrogel compositions comprising hydrogel systems and peptides, methods of making said hydrogel compositions, and methods of using said hydrogel compositions to stimulate the production of extracellular proteins in humans, in a non- therapeutic cosmetic treatment to improve the condition of the skin and / or lines and / or wrinkles and / or imperfections and in methods of treating a wound in the skin of a subject.

[0004] Background of the Invention

[0005] The largest component of normal skin is the dermal extracellular matrix (ECM) which is a gellike matrix produced by the cells that it surrounds. The ECM comprises two major elements, structural proteins and proteoglycans. Changes to skin come about naturally over time (ageing) as a result of changes in the composition and crosslinked state of the elements of the ECM which cause changes at the structural and behavioural level. Structural changes affect the integrity of the skin. Ageing skin exhibits both an increased degradation of dermal ECM proteins (such as collagen, laminin, elastin, fibronectin) and reduced production of the same proteins.

[0006] Ageing changes in the skin can occur intrinsically as a consequence of time and extrinsically as a consequence of external mechanisms, including UV-related damage and pollution. Extrinsic mechanisms can result in the increased availability of reactive oxygen species (UV- ROS). These reactive species are known to cause ECM fragmentation and the upregulation of Matrix Metallopeptidases (MMPs) and other ECM proteases. ECM fragmentation may bring about damage to and / or remodelling of the structural ECM proteins and reduce the integrity of the skin.

[0007] W02022106055A1 discloses peptides for use in cosmetic beauty applications, and compositions comprising said peptide. The document teaches the benefits of the peptide in relation to enhancing production of dermal ECM proteins (such as fibrillin, fibronectin, decorin and collagen) in aging skin. Cosmetic patches comprising active components are known. It is thought that they enhance the efficacy of any cosmetic or therapeutic agent when compared to the corresponding liquid composition by increasing the contact time between the skin and the composition comprising active agents when compared to an equivalent liquid or cream application without a patch. Cosmetic patches comprising hydrogel compositions are used as they allow a high loading of active material into the patch and have the ability to donate and absorb fluid, thus maintaining a moist but not overly saturated, wet environment.

[0008] Summary of the Invention

[0009] The present inventors have developed a hydrogel composition for the provision of peptides to the skin.

[0010] The present invention therefore provides a hydrogel composition comprising a hydrogel system and a peptide. The hydrogel system comprises one or more thickener, and a solvent system comprising one or more organic solvent and one or more surfactant.

[0011] In some embodiments, the peptide comprises a first tetrapeptide, or a second peptide, or a third tetrapeptide, or a fourth tetrapeptide or a combination of the first tetrapeptide and the second peptide.

[0012] The first tetrapeptide (a) is selected from the group consisting of tetrapeptides having the amino acid sequence U-LSXX-Z wherein L is used to denote amino acid Leucine and S is used to denote Serine, as per the internationally recognised single letter code for amino acids, X denotes an amino acid independently selected from the group consisting of Valine (V), Aspartic acid (D), Proline (P), Glycine (G), at the N-terminal end, U is selected from the group consisting of H, -CO-R1, -SO2-R1or a biotinyl group, at the C-terminal end, Z is selected from the group consisting of OH, OR1, NHR1or NR'R2; R1and R2are independently selected from the group consisting of alkyl, aryl, aralkyl, alkylaryl, alkoxy, saccharide and aryloxy group, which may be linear, branched, cyclical, polycyclic, unsaturated, hydroxylates, carbonylated, phosphorylated and / or sulphurous, said groups comprising from 1 to 24 carbon atoms and being capable of including one or more heteroatoms O, S and / or N.

[0013] The second peptide (b) is selected from the group consisting of tetrapeptides having the amino acid sequence U-GPXG-Z wherein G is used to denote amino acid glycine and P denotes the amino acid proline, as per the internationally recognised single letter code for amino acids, X denotes an amino acid independently selected from the group consisting of Lysin(K), Glutamic acid (E), Proline (P) and Serine (S) and mixtures thereof, or a pentapeptide having the amino acid sequence according to SEQ ID No: 17, at the N-terminal end, U is selected from the group consisting of H, -CO-R1, -SO2-R1or a biotinyl group, at the C-terminal end, Z is selected from the group consisting of OH, O R1, NHR1or NR’R2; R1and R2are independently selected from the group consisting of alkyl, aryl, aralkyl, alkylaryl, alkoxy, saccharide and aryloxy group, which may be linear, branched, cyclical, polycyclic, unsaturated, hydroxylates, carbonylated, phosphorylated and / or sulphurous, said groups comprising from 1 to 24 carbon atoms and being capable of including one or more heteroatoms O, S and / or N.

[0014] The third tetrapeptide (c) is selected from the group consisting of tetrapeptides having the amino acid sequence U-XXGD-Z wherein G is used to denote amino acid Glycine and D is used to denote amino acid Aspartic acid, as per the internationally recognised single letter code for amino acids; X denotes an amino acid selected from the group consisting of Glutamic acid (E), Lysine (K), Leucine (L), Alanine (A), Isoleucine (I), Arginine (R) and mixtures thereof; at the N-terminal end, U is selected from the group consisting of H, -CO-R1, -SO2-R1or a biotinyl group; at the C-terminal end, Z is selected from the group consisting of OH, OR1, NHR1or NR'R2; R1and R2are independently selected from the group consisting of alkyl, aryl, aralkyl, alkylaryl, alkoxy, saccharide and aryloxy group, which may be linear, branched, cyclical, polycyclic, unsaturated, hydroxylates, carbonylated, phosphorylated and / or sulphurous, said groups comprising from 1 to 24 carbon atoms and being capable of including one or more heteroatoms O, S and / or N.

[0015] The fourth tetrapeptide (d) is a tetrapeptide having the amino acid sequence U-QTAV-Z wherein Q is used to denote amino acid Glutamine, T is used to denote amino acid Threonine, A is used to denote amino acid Alanine and V is used to denote amino acid Valine; at the N- terminal end, U is independently selected from the group consisting of octanoyl (C8), decanoyl (CIO), lauroyl (C 12), myristoyl (C 14), palmitoyl (C16), stearoyl (C 18), biotinoyl, elaidoyl, oleoyle, lipoyle; at the C-terminal end, Z is selected from the group consisting of OH, OR1, NHR1or NR'R2; R1and R2are independently selected from the group consisting of alkyl, aryl, aralkyl, alkylaryl, alkoxy, saccharide and aryloxy group, which may be linear, branched, cyclical, polycyclic, unsaturated, hydroxylates, carbonylated, phosphorylated and / or sulphurous, said groups comprising from 1 to 24 carbon atoms and being capable of including one or more heteroatoms O, S and / or N. In some embodiments, the peptide comprises (e) palmitoyl tetrapeptide-7 (Pal-GQPR), palmitoyl tripeptide-1 (Pal-GHK), acetyl dipeptide-1 cetyl ester (N-acetyl-YR hexadecyl ester), a combination of Pal-GQPR and Pal-GHK or a combination of all three.

[0016] The present inventors have identified that specific hydrogels and solvent system combinations are particularly beneficial for the delivery of the peptides at least to the lower epidermis, where they provide benefits in relation to enhancing production of dermal ECM proteins. Without wishing to be bound by theory, it is thought that the combination of hydrophilic hydrogel structure and solvents selected for their affinity to the skin drives the solvents and dissolved peptides from the hydrogel composition into the skin along a diffusion gradient. Enhanced delivery of the peptides to the skin is the result of this process. When the peptide comprises an R1or R2group as defined above, such as when U is selected from -CO-R1, -SO2-R1, the affinity of the peptide for the skin is increased, further driving the peptide from the hydrogel composition into the skin.

[0017] In some embodiments of the first aspect, the organic solvent is a diol, an isosorbide or derivative thereof, or a combination thereof.

[0018] In some embodiments of the first aspect, the one or more surfactant is selected from Glyceryl Oleate Citrate, Sorbitan Laurate, Polyglyceryl-4 Laurate, Polyglyceryl-4 Laurate / Succinate, Lauryl Glucoside, Myristyl Glucoside, Polyglyceryl-6 Laurate, Polyglyceryl- 10 Oleate, Laureth-23, Oleth-20, Ceteareth-25, a polysorbate, or combinations therein. In some embodiments of the first aspect, the one or more surfactant is selected from Glyceryl Oleate Citrate, Sorbitan Laurate, Polyglyceryl-4 Laurate, Polyglyceryl-4 Laurate / Succinate, Lauryl Glucoside, Myristyl Glucoside, Polyglyceryl-6 Laurate, Polyglyceryl- 10 Oleate, Laureth-23, Oleth-20, Ceteareth-25, polysorbate 20, or combinations therein. Preferably the surfactant is a polysorbate, and most preferably the surfactant is polysorbate 20.

[0019] In some embodiments of the first aspect, the surfactant is present in an amount of from 0.1 wt.% to 5 wt.%.

[0020] In some embodiments of the first aspect, the diol is propanediol. In some embodiments the diol is present in an amount of from 1 to 40 wt.%. In some embodiments of the first aspect, the isosorbide or derivative thereof is an isosorbide ether, optionally dimethylisosorbide. In some embodiments, the isosorbide or derivative thereof is present in an amount of from 2 to 30 wt.%.

[0021] In some embodiments of the first aspect, the solvent system comprises both a diol and an isosorbide or derivative thereof, wherein the diol is present in an amount of from 1 to 10 wt.% and the isosorbide or derivative thereof is present in an amount of from 2 to 40 wt.%.

[0022] In some embodiments of the first aspect where the solvent system comprises both a diol and an isosorbide ether, the diol is present in an amount of from 1 to 10 wt.% and the isosorbide ether is present in an amount of from 2 to 40 wt.%.

[0023] In some embodiments of the first aspect, the solvent system further comprises glycerin in addition to the organic solvent.

[0024] The use of a diol and / or glycerin in the hydrogel composition provides the additional benefit of making the hydrogel composition substantive to the skin. This means that users can affix patches of the hydrogel composition to their skin in the location that they are to work, with no additional attachment means being necessary.

[0025] In some embodiments of the first aspect, the one or more thickener is selected from carrageenan, amorphophallus konjac root extract, caesalpinia spinosa gum, sclerotium gum, agar, locust bean gum, cellulose, starch, chitin, hyaluronic acid, guar gum, gellan gum, xanthan gum, chitosan, phosphoryl choline, acrylic acid, sodium polyacrylate, polyvinyl acetate (PVAc) or combinations thereof.

[0026] In some embodiments of the first aspect, the one or more thickener is present in an amount of from 0. 1 to 5 wt.%.

[0027] In a second aspect, the present invention provides a method of preparing the hydrogel composition of the first aspect, the method comprising providing the hydrogel system and adding the peptide. In some embodiments, the method further comprises the steps of preparing the hydrogel system, the steps comprising providing the one or more thickener; providing the solvent system; and combining the one or more thickener and solvent system to provide the hydrogel system.

[0028] In a third aspect, the present invention provides a method for stimulating the production of extracellular proteins in humans, the method comprising administering to the skin of said human the hydrogel composition of the first aspect.

[0029] In a further aspect, the present invention provides a use of the hydrogel composition of the first aspect in a non-therapeutic cosmetic treatment to improve the condition of the skin and / or lines and / or wrinkles and / or imperfections such as the appearance of scars.

[0030] In a sixth aspect, the present invention provides the hydrogel composition of the first aspect for use in a method for treating a wound in the skin of a subject, the method comprising applying the hydrogel composition of the present invention to the wound in the skin of the subject.

[0031] Detailed Description of the Invention

[0032] The present invention is directed to hydrogel composition comprising a hydrogel system and a peptide. The hydrogel system comprises one or more thickener, and a solvent system comprising one or more organic solvent and one or more surfactant.

[0033] All percentages used herein are by weight of the hydrogel composition unless otherwise specified.

[0034] Hydrogel Composition

[0035] Hydrogels are macromolecular networks swollen partially or to equilibrium with a suitable fluid, normally an aqueous fluid.

[0036] The hydrogel composition of the present invention is a solid composition at room temperature. That is, when heated above room temperature it will undergo a phase change to become a liquid composition. The hydrogel composition of the present invention may be formed into a patch for application to the skin. The patch may have any suitable dimensions for application to the skin.

[0037] Hydrogel composition of the present invention comprises a hydrogel system. The hydrogel system comprises one or more thickener in combination with a solvent system comprising one or more organic solvent and one or more surfactant.

[0038] In some embodiments the thickener is selected from carrageenan, amorphophallus konjac root extract, caesalpinia spinosa gum, sclerotium gum, agar, locust bean gum, cellulose, starch, chitin, hyaluronic acid, guar gum, gellan gum, xanthan gum, chitosan, phosphoryl choline, acrylic acid, sodium polyacrylate, polyvinyl acetate (PVAc) or combinations thereof.

[0039] In preferred embodiments, the thickener is selected from guar gum, gellan gum, xanthan gum, and combinations thereof. For example, the composition may comprise guar gum and gellan gum, gellan gum and xanthan gum, guar gum and xanthan gum, or guar gum, gellan gum and xanthan gum. Guar gum, gellan gum, xanthan gum, in particular the combination of guar gum, gellan gum and xanthan gum provides a solid structure network with suitable mechanical properties such as flexibility for use in a cosmetic patch.

[0040] The one or more thickener of the hydrogel system provides a hydrophilic, three-dimensional structure network that holds the solvent system and the peptides dissolved therein. When applied to the skin, this hydrophilic network is less favourable to the solvent system than the skin, and so the solvents move from the composition to the skin. The peptides dissolved in the solvent system are therefore transported into the skin.

[0041] The composition may comprise at least 0.1 wt.% of the one or more thickener, such as at least 0.2 wt.%, at least 0.3 wt.% at least 0.4 wt.% at least 0.5 wt.%, at least 0.6 wt.%, at least 0.7 wt.%, at least 0.8 wt.% at least 0.9 wt.% or at least 1 wt.% of the one or more thickener. In some embodiments, the composition comprises 5 wt.% or less of the one or more thickener, such as 4.5 wt.% or less, 4 wt.% or less, 3.5 wt.% or less, 3 wt.% or less 2.5 wt.% or less or 2 wt.% or less of the one or more thickener.

[0042] In some embodiments, the composition may comprise from 0.1 wt.% to 5 wt.% of the one or more thickener, for example, the composition may comprise from 0.3 to 4 wt.%, from 0.5 to 3 wt.%, from 0.7 to 2.5 wt.% or from 1 to 2 wt.% of one or more thickener. The hydrogel system of the present invention further comprises a solvent system having one or more organic solvent and one or more surfactant. An organic solvent is one that comprises at least one carbon atom. Examples of organic solvents include ethers and alcohols.

[0043] As would be apparent to the skilled person, the composition further comprises water. Water may be considered part of the solvent system.

[0044] The solvent system comprises an organic solvent. The solvent system may comprise at least one solvent selected from a diol, an isosorbide or derivative thereof, or a combination thereof.

[0045] The diol is a compound with a melting point below room temperature having two alcohol moieties. In some embodiments, the diol is selected from propanediol, butanediol, pentanediol, hexanediol or combinations thereof. In some preferred embodiments, the alcohol is propanediol. The skilled person would understand that room temperature is 20 °C.

[0046] The diol may be present in an amount of at least 1 wt.%, such as at least 5 wt.%, at least 10 wt.% or at least 15 wt.%. The diol may be present in an amount of 40 wt.% or less, such as 35 wt.% or less or 30 wt.% or less. The diol may be present in an amount of from 1 to 40 wt.%, such as from 10 to 35 wt.%, from 15 to 30 wt.%, or from 15 to 25 wt.% of the composition. Where the solvent system does not comprise an isosorbide or derivative thereof, the diol may be present in an amount of at least 10 wt.%, such as from 10 to 35 wt.% from 15 to 30 wt.%, or from 15 to 25 wt.%.

[0047] The isosorbide or derivative thereof is a compound with a melting point below room temperature optionally having the structure of Formula (I).

[0048] Formula (I) Wherein each R3is independently selected from H, methyl, ethyl, branched or linear propyl, branched or linear butyl, or -CO-R4, wherein R4is independently selected from H, methyl, ethyl, branched or linear propyl, branched or linear butyl.

[0049] In some embodiments, isosorbide or derivative thereof is an isosorbide ether. In some embodiments, R3of Formula (I) is independently selected from methyl, ethyl, branched or linear propyl, branched or linear butyl. In some embodiments, the isosorbide ether may be selected from dimethyl isosorbide, diethyl isosorbide, dipropyl isosorbide, dibutyl isosorbide and combinations thereof. In some preferred embodiments, the isosorbide ether is dimethyl isosorbide.

[0050] In some embodiments, isosorbide or derivative thereof is an isosorbide ester. In some embodiments, R3of Formula (I) R3is equal to -CO-R4, wherein R4is independently selected from H, methyl, ethyl, branched or linear propyl, branched or linear butyl, optionally wherein R4is selected from methyl, ethyl, branched or linear propyl, branched or linear butyl.

[0051] The isosorbide or derivative thereof may be present in an amount of at least 2 wt.%, such as at least 4 wt.% or at least 6 wt.%. The isosorbide or derivative thereof may be present in an amount of 30 wt.% or less, such as 25 wt.% or less or 20 wt.% or less. The isosorbide or derivative thereof may be present in an amount of from 2 to 30 wt.%, such as from 4 to 25 wt.%, from 6 to 20 wt.%, or from 6 to 15 wt.% of the composition.

[0052] In some embodiments where the solvent system comprises both a diol and an isosorbide or derivative thereof, the diol is present in an amount of from 1 to 10 wt.% and the isosorbide or derivative thereof is present in an amount of from 2 to 40 wt.%, such as from 2 to 30 wt.%, such as from 4 to 25 wt.%, from 6 to 20 wt.%, or from 6 to 15 wt.%.

[0053] The solvent system further comprises a surfactant. In some embodiments the surfactant is a non-ionic surfactant. In some embodiments, the surfactant is selected from polysorbate 20, polysorbate 40, polysorbate 60, polysorbate 80, laureth-23, sodium oleate, potassium oleate, or combinations thereof. In some embodiments, the surfactant is a polysorbate-type non-ionic surfactant. In some embodiments, the surfactant is selected from polysorbate 20, polysorbate 40, polysorbate 60, polysorbate 80 or combinations thereof. In some preferred embodiments the surfactant is polysorbate 20. The surfactant may be present in an amount of at least 0.1 wt.%, such as at least 0.2 wt.%, at least 0.3 wt.%, such as at least 0.4 wt.% or at least 0.5 wt.%. The surfactant may be present in an amount of 5 wt.% or less, such as 4 wt.%, less or 3 wt.% or less, or 2 wt.% or less. The surfactant may be present in an amount of from 0.1 wt.% to 5 wt.%, such as from 0.2 to 4 wt.%, from 0.3 to 3 wt.%, from 0.4 to 2 wt.%, from 0.5 to 2 wt.%, or from 0.7 to 1.3 wt.%.

[0054] In some embodiments, the solvent system comprises a diol, an isosorbide ether or a combination thereof, and a polysorbate-type non-ionic surfactant.

[0055] In some embodiments the solvent system comprises a diol selected from propanediol, butanediol, pentanediol, hexanediol and combinations thereof, an isosorbide ether selected from dimethyl isosorbide, diethyl isosorbide, dipropyl isosorbide, dibutyl isosorbide and combinations thereof, and a polysorbate selected from polysorbate 20, polysorbate 40, polysorbate 60, polysorbate 80 or combinations thereof.

[0056] In some embodiments the solvent system comprises a diol selected from propanediol, butanediol, pentanediol, hexanediol and combinations thereof and a polysorbate selected from polysorbate 20, polysorbate 40, polysorbate 60, polysorbate 80 or combinations thereof.

[0057] In some embodiments the solvent system comprises an isosorbide ether selected from dimethyl isosorbide, diethyl isosorbide, dipropyl isosorbide, dibutyl isosorbide and combinations thereof, and a polysorbate selected from polysorbate 20, polysorbate 40, polysorbate 60, polysorbate 80 or combinations thereof.

[0058] The present inventors have found that a solvent system comprising propanediol and polysorbate 20, or one comprising dimethyl isosorbide and polysorbate 20, or one comprising propanediol, dimethyl isosorbide and polysorbate 20 is particularly effective in delivering the peptides used in the present invention to the skin of a user. This has been determined using Hansen Solubility Parameters, as discussed in the examples herein.

[0059] Therefore, in some preferred embodiments, the solvent system comprises a solvent selected from propanediol, dimethyl isosorbide or a combination thereof, and a surfactant selected from polysorbate 20. In other preferred embodiments the solvent system comprises propanediol and polysorbate 20. For example, the solvent system may comprise from 10 to 20 wt.% of propanediol and from 0.1 to 2 wt.% of polysorbate 20.

[0060] In some preferred embodiments the solvent system comprises dimethyl isosorbide and polysorbate 20. For example, the solvent system may comprise from 6 to 20 wt.% of dimethyl isosorbide and from 0.1 to 2 wt.% of polysorbate 20.

[0061] In some preferred embodiments, the solvent system comprises propanediol, dimethyl isosorbide and polysorbate 20. For example, the solvent system comprises from 1 to 10 wt.% propanediol, from 6 to 20 wt.% of dimethyl isosorbide and from 0.1 to 2 wt.% of polysorbate 20.

[0062] The solvent system may further comprise glycerin in addition to the organic solvent. Glycerin may be present in an amount of at least 5 wt.%, such as at least 10 wt.% or at least 15 wt.%. glycerine may be present in an amount of 40 wt.% or less, such as 35 wt.% or less or 30 wt.% or less. Glycerin may be present in an amount of from 5 to 40 wt.%, such as from 10 to 35 wt.%, from 15 to 30 wt.%, or from 15 to 25 wt.% of the composition.

[0063] Therefore, in some embodiments, the solvent system comprises a diol, an isosorbide ether or a combination thereof, glycerin, and a polysorbate-type non-ionic surfactant. For example, the solvent system may comprise a solvent selected from propanediol, dimethyl isosorbide or a combination thereof, glycerin, and a surfactant selected from polysorbate 20.

[0064] As will be understood by the skilled person, the solvent system of a hydrogel comprises water. Water may be included in the solvent system to the saturation point of the one or more thickener following the addition of other parts of the solvent system, peptide and components such as preservatives and pH adjusters to the one or more thickener. That is, water may be included in the composition to equilibrium. In some embodiments, water is present in the composition in an amount of at least 35 wt.%, such as 40 wt.% or 45 wt.%. In some embodiments, water is present in the composition in an amount of 85 wt.% or less, such as 80 wt.% or less, 75 wt.% or less, 70 wt.% or less or, 65 wt.% or less. In some embodiments, is present in the composition in an amount of from 35 wt.% to 85 wt.%, such as from 40 to 85 wt.%. In some embodiments the hydrogel system of the composition comprises two or more thickeners selected from guar gum, gellan gum and xanthan gum, water, and propanediol and / or dimethyl isosorbide. In some embodiments this hydrogel system further comprises glycerin.

[0065] In some embodiments, the hydrogel system comprises from 0.1 wt.% to 5 wt.% of the two or more thickeners, optionally selected from guar gum, gellan gum and xanthan gum, from 0.1 wt.% to 5 wt.% of a polysorbate-type non-ionic surfactant, from 5 to 40 wt.% of glycerin, at least 40 wt.% of water, and either i) 10 to 35 wt.% of a diol, ii) from 6 to 20 wt.% of an isosorbide ether or iii) from 1 to 10 wt.% of a diol and from 6 to 20 wt.% of an isosorbide ether.

[0066] The hydrogel composition of the present invention is a solid composition at room temperature. That is, when heated above room temperature it will undergo a phase change to become a liquid composition.

[0067] The solvent system of the present invention may comprise a single phase, which is an aqueous phase. That is, in some embodiment the solvent system of the present invention does not comprise an oil phase, or is not an emulsion comprising an oil phase.

[0068] In some embodiments, the hydrogel composition of the present invention is suitable for topical application. The composition of the present invention may be formed into a patch for application to the skin. The patch may have any suitable dimensions for application to the skin.

[0069] Peptides

[0070] The hydrogel composition of the present invention comprises a peptide.

[0071] The peptide of the hydrogel composition may comprise a first tetrapeptide, a second peptide, a third tetrapeptide, a fourth tetrapeptide or a combination of the first tetrapeptide and the second peptide.

[0072] The first tetrapeptide (a) has generic formulation U-LSXX-Z wherein L is used to denote amino acid Leucine and S is used to denote Serine, as per the internationally recognised single letter code for amino acids. X denotes an amino acid selected from the group consisting of Valine (V), Aspartic acid (D), Proline (P), Glycine (G) and mixtures thereof. At the N-terminal end, U is selected from the group consisting of H, -CO-R1, -SO2-R1or a biotinyl group. At the C- terminal end, Z is selected from the group consisting of OH, O R1, NHR1or NR’R2. R1and R2are independently selected from the group consisting of alkyl, aryl, aralkyl, alkylaryl, alkoxy, saccharide and aryloxy group, which may be linear, branched, cyclical, polycyclic, unsaturated, hydroxylates, carbonylated, phosphorylated and / or sulphurous, said groups comprising from 1 to 24 carbon atoms and being capable of including one or more heteroatoms O, S and / or N.

[0073] The tetrapeptide of the present invention is preferably selected from the group consisting of SEQ ID No: 1, SEQ ID No: 2, SEQ ID No: 3, SEQ ID No: 4, SEQ ID No: 5, SEQ ID No: 6, SEQ ID No: 7, SEQ ID No: 8, SEQ ID No: 9, SEQ ID No: 10, SEQ ID No: 11 and SEQ ID No: 12.

[0074] In another embodiment of the present invention the tetrapeptide is preferably selected from the group consisting of SEQ ID No: 1, SEQ ID No: 9 and SEQ ID No: 8.

[0075] In another embodiment of the present invention the tetrapeptide is preferably selected from the group consisting of SEQ ID No: 1.

[0076] In another embodiment of the present invention the tetrapeptide is preferably selected from the group consisting of SEQ ID No: 9.

[0077] In another embodiment of the present invention the tetrapeptide is preferably selected from the group consisting of SEQ ID No: 8.

[0078] In a preferred embodiment the tetrapeptide combination of the present invention, tetrapeptide a) is selected from the group consisting of U-LSVD-Z, U-LSVP-Z, U-LSVG-Z, U-LSDV-Z, U- LSDP-Z, U-LSDG-Z, U-LSPV-Z, U-LSPD-Z, U-LSPG-Z, U-LSGV-Z, U-LSGD-Z and U- LSGP-Z. In another embodiment, the tetrapeptides is selected from the group consisting of U- LSVD-Z, U-LSPG-Z and U-LSPD-Z. In another embodiment the tetrapeptide a) is Pal-LSVD- OH. In another embodiment the tetrapeptide a) is Pal-LSPG-OH. In another embodiment the tetrapeptide a) is Pal-LSPD-OH.

[0079] The second peptide (b) has generic amino acid sequence U-GPXG-Z wherein G is used to denote amino acid glycine and P denotes the amino acid proline, as per the internationally recognised single letter code for amino acids, X denotes an amino acid selected from the group consisting of Lysine (K), Glutamic acid (E) and Serine (S) and mixtures thereof, or a pentapeptide having the amino acid sequence according to SEQ ID No: 17, at the N-terminal end, U is selected from the group consisting of H, -CO-R1, -SO2-R1or a biotinyl group, at the C-terminal end, Z is selected from the group consisting of OH, OR1, NHR1or NR'R2. R1and R2are independently selected from the group consisting of alkyl, aryl, aralkyl, alkylaryl, alkoxy, saccharide and aryloxy group, which may be linear, branched, cyclical, polycyclic, unsaturated, hydroxylates, carbonylated, phosphorylated and / or sulphurous, said groups comprising from 1 to 24 carbon atoms and being capable of including one or more heteroatoms O, S and / or N.

[0080] In a preferred embodiment, peptide b) is selected from the group consisting of SEQ ID No: 13, SEQ ID No: 14, SEQ ID No: 15, SEQ ID No: 16 and SEQ ID No: 17.

[0081] In a further preferred embodiment of the present invention peptide b) is U-GPKG-Z.

[0082] In a further preferred embodiment of the present invention peptide b) is U-GPEG-Z.

[0083] In a further preferred embodiment of the present invention peptide b) is U-GPSG-Z.

[0084] The third tetrapeptide (c) has the generic formulation U-XXGD-Z wherein G is used to denote amino acid Glycine and D is used to denote amino acid Aspartic acid, as per the internationally recognised single letter code for amino acids. X denotes an amino acid selected from the group consisting of Glutamic acid (E), Lysine (K), Leucine (L), Alanine (A), Isoleucine (I), Arginine (R) and mixtures thereof. At the N-terminal end, U is selected from the group consisting of H, -CO-R1, -SO2-R1or a biotinyl group. At the C-terminal end, Z is selected from the group consisting of OH, OR1, NHR1or NR'R2. R1and R2are independently selected from the group consisting of alkyl, aryl, aralkyl, alkylaryl, alkoxy, saccharide and aryloxy group, which may be linear, branched, cyclical, polycyclic, unsaturated, hydroxylates, carbonylated, phosphorylated and / or sulphurous, said groups comprising from 1 to 24 carbon atoms and being capable of including one or more heteroatoms O, S and / or N.

[0085] The third tetrapeptide is preferably selected from the group consisting of SEQ ID No: 25, SEQ ID No: 26, SEQ ID No: 27, SEQ ID No: 28, SEQ ID No: 29, SEQ ID No: 30, SEQ ID No: 31, SEQ ID No: 32, SEQ ID No: 33, SEQ ID No: 34, SEQ ID No: 35, SEQ ID No: 36, SEQ ID No: 37, SEQ ID No: 38, SEQ ID No: 39, SEQ ID No: 40, SEQ ID No: 41, SEQ ID No: 42, SEQ ID No: 43, SEQ ID No: 44, SEQ ID No: 45, SEQ ID No: 46, SEQ ID No: 47, SEQ ID No: 48, SEQ ID No: 49, SEQ ID No: 50, SEQ ID No: 51, SEQ ID No: 52, SEQ ID No: 53 and SEQ ID No: 54.

[0086] In another embodiment the third tetrapeptide is preferably selected from the group consisting of SEQ ID No: 25, SEQ ID No: 36, SEQ ID No: 44 and SEQ ID No: 51 .

[0087] In another embodiment the third tetrapeptide is preferably selected from the group consisting of SEQ ID No: 25.

[0088] In another embodiment the third tetrapeptide is preferably selected from the group consisting of SEQ ID No: 36.

[0089] In another embodiment the third tetrapeptide is preferably selected from the group consisting of SEQ ID No: 44.

[0090] In another embodiment the third tetrapeptide is preferably selected from the group consisting of SEQ ID No: 51.

[0091] In a preferred embodiment the third tetrapeptide is selected from the group consisting of U- EKGD-Z, U-ELGD-Z, U-EAGD-Z, U-EIGD-Z, U-ERGD-Z, U-KEGD-Z, U-KLGD-Z, U- KAGD-Z, U-KIGD-Z, U-KRGD-Z, U-LEGD-Z, U-LKGD-Z, U-LAGD-Z, U-LIGD-Z, U- LRGD-Z, U-IEGD-Z, U-IKGD-Z, U-ILGD-Z, U-IAGD-Z, U-IRGD-Z, U-REGD-Z, U-RKGD- Z, U-RLGD-Z, U-RAGD-Z, U-RIGD-Z, U-AEGD-Z, U-AKGD-Z, U-ALGD-Z, U-AIGD-Z and U-ARGD-Z. In another embodiment, the third tetrapeptides is selected from the group consisting of U-EKGD-Z, U-LKGD-Z, U-IRGD-Z and U-AKGD-Z. In another embodiment the third tetrapeptide is U-EKGD-Z. In another embodiment the third tetrapeptide is U-LKGD-Z. In another embodiment the third tetrapeptide is U-IRGD-Z. In another embodiment the third tetrapeptide is U-AKGD-Z.

[0092] The fourth tetrapeptide (d) has the amino acid sequence U-QTAV-Z wherein Q is used to denote amino acid Glutamine, T is used to denote amino acid Threonine, A is used to denote amino acid Alanine and V is used to denote amino acid Valine. At the N-terminal end, U is selected from the group consisting of H, -CO-R1, -SO2-R1or a biotinyl group. At the C-terminal end, Z is selected from the group consisting of OH, OR1, NHR1or NR'R2. R1and R2are independently selected from the group consisting of alkyl, aryl, aralkyl, alkylaryl, alkoxy, saccharide and aryloxy group, which may be linear, branched, cyclical, polycyclic, unsaturated, hydroxylates, carbonylated, phosphorylated and / or sulphurous, said groups comprising from 1 to 24 carbon atoms and being capable of including one or more heteroatoms O, S and / or N.

[0093] Where, at the N-terminal, U is H then the amino acid is not modified. When, at the C-terminal, Z is OH then the amino acid is not modified. The tetrapeptide is thus not in derivatised form. When other than U is H and Z is OH, then the tetrapeptide is derivatised. Derivation of the tetrapeptide is intended to increase the bioavailability of the peptide by improving the ability of the tetrapeptide to pass through the skin. An increase in bioavailability can also be achieved through vectoring, for example by encapsulation of the peptide.

[0094] In a preferred embodiment of the present invention the tetrapeptide is modified at the N- terminal and / or the C-terminal end.

[0095] In a preferred embodiment of the present invention, R1and / or R2is an alkyl chain of from 1 to 24 carbon atoms, preferably a lipophilic alkyl chain of 3 to 24 carbon atoms.

[0096] In a further preferred embodiment of the present invention U is an acyl group -CO-R1and Z is selected from the group consisting of OH, methoxy, ethoxy and NH2, preferably OH. In a further embodiment, U is preferably independently selected from the group consisting of octanoyl (C8), decanoyl (CIO), lauroyl (C12), myristoyl (C 14), palmitoyl (C16), stearoyl (Cl 8), biotinoyl, elaidoyl, oleoyle and lipoyle. In a preferred embodiment of the present U is independently selected from lauroyl (C 12), myristoyl (C14) and palmitoyl (C16).

[0097] Peptides of embodiments of the preferred invention where U is an acyl group and Z is selected from the group consisting of OH, methoxy, ethoxy and NH2 have a particular affinity for the skin. It is thought that when the hydrophilic environment of the hydrogel composition is brought into contact with the skin, the peptide is driven out of the patch and into the skin along a diffusion gradient. The solvent system of the present invention further enhances this process, encouraging delivery of the peptide to the skin.

[0098] In a further preferred embodiment Z is OH and U is independently selected from the group consisting of palmitoyl (C l 6), myristoyl (C l 4) and lauroyl (C l 2). Most preferably U is palmitoyl (Cl 6) and Z is OH. The tetrapeptides may comprise amino acids in the D- or L- configuration. The tetrapeptides may comprise an acid C-terminus such as -CO2H.

[0099] The amino acids making up the tetrapeptides according to the invention may be optically pure, be made up of L or D isomers or a mixture thereof. L isomers are those present in the natural state and may be preferred.

[0100] The present invention also envisages and includes further derivatives of the tetrapeptide, including for example modification and / or addition of a chemically functional group to one or more of the amino acids but without a change in the carbon skeleton. The present invention also envisages and includes further analogues of the tetrapeptide, including modification and / or addition of a chemically functional group to one or more of the amino acids with a change in the carbon skeletal and complexes of the tetrapeptide with other species such as a metal ion (e.g. copper, zinc, manganese, magnesium, and others).

[0101] Tetrapeptides are also envisaged in the form of salts, including hydrochloric salt, or acetate. The combination of tetrapeptides of the present invention show synergistic benefit in that they offer markedly improved levels of fibrillin- 1 production in human dermal fibroblast cells in comparison to the peptides singularly.

[0102] The tetrapeptide may be present in the hydrogel composition in an amount of from of from O. lOppm to 10,000ppm, preferably from 0.50ppm to 5,000ppm, more preferably from Ippm to lOOOppm, and most preferably from Ippm to 500ppm. These are based on a wt.% basis. Thus 100,000ppm is 10% by weight of the hydrogel composition.

[0103] Where the hydrogel composition comprises a combination of a first tetrapeptide and a second peptide, the first tetrapeptide and second peptide may be present in the hydrogel composition in a weight ratio of from 20:80 to 80:20, such as 30:70 to 80:20, 40:60 to 80:20, 50:50 to 80:20, 60:40 to 80:20, or 70:30 to 80:20, based on the total weight of the first tetrapeptide and a second peptide. The first tetrapeptide and second peptide may be present in the hydrogel composition in a weight ratio of from 20:80 to 80:20, such as 20:80 to 70:30, 20:80 to 60:40, 20:80 to 50:50, 20:80 to 40:60, or 20:80 to 30:70, based on the total weight of the first tetrapeptide and a second peptide. The first tetrapeptide and second peptide may be present in the hydrogel composition in a weight ratio of from 20:80 to 80:20, such as 30:70 to 70:30, or 40:60 to 60:40, based on the total weight of the first tetrapeptide and a second peptide. In some embodiments, the peptide of the hydrogel composition may comprise (e) palmitoyl tetrapeptide-7 (Pal-GQPR, SEQ ID No: 19), palmitoyl tripeptide- 1 (Pal-GHK), acetyl dipeptide- 1 cetyl ester (N-acetyl-YR hexadecyl ester), a combination of Pal-GQPR and Pal- GHK or a combination of all three.

[0104] Peptide (e) may be present in the hydrogel composition in an amount of from of from O. lOppm to 10,000ppm, preferably from 0.50ppm to 5,000ppm, more preferably from Ippm to lOOOppm. These are based on a wt.% basis. Thus 100,000ppm is 10% by weight of the hydrogel composition.

[0105] Additional Actives

[0106] The hydrogel compositions of the invention may comprise additional active components. These active components may have a cosmetic benefit to the appearance of the skin.

[0107] Vitamin B and Derivatives:

[0108] In a preferred embodiment, the hydrogel composition of the present invention comprises vitamin B, vitamin B derivatives, vitamin Bl to vitamin B12 and their derivatives; which are preferably chosen from niacinamide, panthenol, pantothenic acid, pyridoxine, pyridoxine hydrochloride, biotin, and combinations thereof. Preferably the vitamin B, vitamin B derivatives, vitamin Bl to vitamin B 12 and their derivatives are present in an amount of from 0.1 to 5 wt.% or more preferably from 1 to 4% wt.%. In particular niacinamide is the chosen Vitamin B. Niacinamide is most preferably present in an amount from 1 to 4% wt.%.

[0109] Vitamins:

[0110] The hydrogel compositions may comprise one or more vitamins. The hydrogel compositions may comprise ascorbates, for example vitamin C, vitamin C derivatives, ascorbic acid, ascorbyl glucoside, ascorbyl palmitate, magnesium ascorbyl phosphate, sodium ascorbyl phosphate and ethyl ascorbic acid. The hydrogel composition may comprise vitamin B, vitamin B derivatives, vitamin Bl to vitamin B 12 and their derivatives. In a further embodiment the hydrogel composition comprising the Vitamin B3 derivative niacinamide. In an alternative embodiment the hydrogel composition comprises vitamin K, vitamin K derivatives, vitamin H, vitamin D, vitamin D derivatives and mixtures thereof. In an alternative embodiment of the hydrogel composition comprises vitamin E, vitamin E derivatives such as tocopherol and tocopheryl acetate, and provitamins thereof, such as panthenol and mixtures thereof.

[0111] In a further embodiment the hydrogel composition comprises retinoid compounds, including retinoic acid, retinaldehyde, retinol and derivatives thereof. In one embodiment the hydrogel composition comprises retinyl palmitate, retinyl acetate, retinyl retinoate, retinyl proprionate, retinyl ascorbate, retinyl linoleate, retinyl retinoate, retinyl sunflowerseedate and mixtures thereof.

[0112] The vitamin compounds may be included as the substantially pure material, or as an extract obtained by suitable physical and / or chemical isolation from natural (e. g. plant) sources. In some embodiments, when vitamin compounds are present in the hydrogel compositions of the instant invention, the hydrogel compositions comprise from about 0.0001% to 10%, more preferably from 0.001% to 8%, still more preferably from 0.01% to 5%, and still more preferably from 0. 1% to 5%, by weight of the hydrogel composition, of the vitamin compound.

[0113] Additional Peptides:

[0114] The hydrogel compositions of the invention may comprise further peptides. Preferably said additional peptides are selected from the group consisting of dipeptides, tripeptides, additional tetrapeptides, pentapeptides and mixtures thereof. By tripeptides, it is meant compound comprising an uninterrupted sequence of three amino acids. By tetrapeptides, it is meant a compound comprising an uninterrupted sequence of four amino acids and when using further tetrapeptides, the tetrapeptides are referred to as ‘additional tetrapeptides’ . By pentapeptide it is meant a compound comprising an uninterrupted sequence of five amino acids.

[0115] Dipeptides

[0116] The hydrogel compositions may comprise a dipeptide selected from the group consisting of acetyl dipeptide 1 cetyl ester, acetyl dipeptide 3 aminohexanoate, azelaoyl bisdipeptide 10, coumaroyl dipeptide 3, dicetyl dipeptide 9, dipeptide diamino butyroyl benzylamide diacetate, dipeptide 1, dipeptide 10, dipeptide 11, dipeptide 12, dipeptide 15, dipeptide 16, dipeptide 17, dipeptide 18, dipeptide 19, dipeptide 2, dipeptide 20, dipeptide 3, dipeptide 4, dipeptide 5, dipeptide 6, dipeptide 7, dipeptide 8, dipeptide 8 HCL, dipeptide 9, hexanoyl dipeptide 3 norleucine acetate, methyl undecylenoyl dipeptide 16, nicotinoyl dipeptide 22, nicotinoyl dipeptide 23, nicotinoyl dipeptide 24, nicotinoyl dipeptide 26, oleoyl dipeptide 15, palmitoyl dipeptide 10, palmitoyl dipeptide 13, palmitoyl dipeptide 17, palmitoyl dipeptide 5 diaminobutyroyl hydroxythreonine, palmitoyl dipeptide 5 diaminohydroxybutyrate, palmitoyl dipeptide 7 and mixtures thereof.

[0117] Dipeptides are preferably incorporated into the hydrogel composition at a level of from 0.1 to 50000ppm, more preferably from 1 to 5000 ppm, most preferably from 10 to 500ppm.

[0118] Tripeptides

[0119] The hydrogel compositions may comprise a tripeptide. Said tripeptide may be naturally occurring or of synthetic origin. Suitable tripeptides include tripeptide 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, derivatives thereof and mixtures thereof.

[0120] Particularly preferred tripeptides comprise one or more His-based tripeptides. However, another suitable tripeptide may be Arg-Lys-Arg. Particularly preferred tripeptides are based on the structure Gly-His-Lys and its analogs and derivatives thereof. These are collectively known herein as GHK-tripeptides. Indeed, the preferred tripeptide in accordance with this aspect of the invention has this exact sequence of amino acids. Analogs of the preferred tripeptide useful herein include those in which one or more of the three amino acids are reorganized or rearranged within the sequence (e.g., Gly-Lys-His) and / or where no more than two amino acids are substituted (e.g., His-Ala-Orn). However, most preferably, amino acids substituted for Gly include an aliphatic side chain such as, without limitation, beta-Ala, Ala, Vai, Leu, Pro, Sarcosine (Sar) and lie. Most preferred are Ala, Leu and He. The most preferable amino acid substituted for Lys or His include those having a side chain that includes, predominantly, a charged nitrogen at a pH of 6, such as, without limitation, Pro, Lys, Arg, His, Desmosine and Isodesmosine. Most preferably, Lys is replaced with Orn, Arg, or Citrulline.

[0121] Derivatives are also considered to be encompassed by the term GHK-tripeptides in accordance with the present invention, (and therefore also the more generic term tripeptides). Derivatives of GHK-tripeptides in accordance with the present invention include derivatives of the substituted and rearranged tripeptides described herein. These derivatives include, inter alia, acyl-derivatives, which are tripeptides substituted with one or more straight-chain or branched- chain, long or short chain, saturated or unsaturated, substituted with a hydroxy, amino, acyl amino, sulfate or sulfide group, or unsubstituted, which can be derived from acetic acid, capric acid, lauric acid, myristic acid, octanoic acid, palmitic acid, stearic acid, behenic acid, linoleic acid, linolenic acid, lipoic acid, oleic acid, isostearic acid, elaidoic acid, 2-ethylhexaneic acid, coconut oil fatty acid, tallow fatty acid, hardened tallow fatty acid, palm kernel oil fatty acid, lanolin fatty acid and the like. Preferable examples of the acyl group include an acetyl group, a palmitoyl group, an elaidoyl group, a myristyl group, a biotinyl group and an octanoyl group. These may be substituted or unsubstituted. When substituted, they are preferably substituted with hydroxyl or sulphur comprising groups such as, without limitation SO3H, SH or S-S.

[0122] His-based tripeptides include at least one histidine amino acid. The other two amino acids in the sequence may be the same or different. Thus, contemplated are, without limitation, His- Xaa-Xaa, His-Xaa-Xbb, His-Xbb-Xaa, Xbb-His-Xbb, Xbb-His-Xaa, Xaa-His-Xbb, Xaa-Xaa- His, Xaa-Xbb-His, Xbb-Xaa-His and Xbb-Xbb-His, where Xaa and Xbb are two different amino acids, although either can be His. Preferably, at least one of the other amino acids is Gly, betaAla, Ala, Vai, Leu, Pro, Sarcosine (Sar) or lie. Preferably, at least one of the other amino acids is Pro, Lys, Arg, His, Desmosine and Isodesmosine. Most preferably, Lys is replaced with Orn, Arg, or Citrulline.

[0123] Derivatives are also considered to be encompassed by the term His-based tripeptides in accordance with the present invention, (and therefore also the more generic term tripeptides). These derivatives include, inter alia, acyl-derivatives, which are tripeptides substituted with one or more straight-chain or branched-chain, long or short chain, saturated or unsaturated substituted or unsubstituted acyl group(s) having from 1 to 29 carbon atoms. The acyl groups which can be used are the same as those described for the GHK-tripeptides.

[0124] Particularly preferred embodiments of tripeptides in accordance with the present invention include N-Acyl-Gly-His-Lys and most preferably, N-Palmitoyl-Gly-His-Lys. Preferred commercially available tripeptide and tripeptide derivative comprising hydrogel compositions include Biopeptide-CL from SEDERMA, Maxilip(R) from SEDERMA, Biobustyl(R) from SEDERMA. The tripeptides where included are preferably incorporated into the hydrogel composition in amounts of from O. lOppm to 10,000ppm, preferably from 0.50ppm to 5,000ppm, more preferably from Ippm to lOOOppm, and most preferably from Ippm to 500ppm. These are again based on a % w / w basis. Thus 100,000ppm is 10% by weight of the emulsion.

[0125] Additional tetrapeptides

[0126] The hydrogel composition may comprise an additional tetrapeptide. These may be one or more rigin-based tetrapeptides, one or more ALAMCAT-tetrapeptides or mixtures thereof. These tetrapeptides may be naturally occurring or of synthetic origin. Suitable tetrapeptides for use in the present hydrogel composition include those selected from the group consisting of well- known tetrapeptide 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 ,20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 34, 35, derivatives thereof and mixtures thereof.

[0127] Rigin-based tetrapeptides in accordance with the present invention are based on the structure Gly-Gln-Pro-Arg (Rigin) and include its analogs and derivatives thereof. Rigin is an additional tetrapeptide. Analogs of the tetrapeptide rigin useful in accordance with the present invention include those in which one or more of the four amino acids are reorganized or rearranged within the sequence and / or where no more than two of the amino acids are substituted (e.g., Ala-Gln- Thr-Arg. More preferably, at least one of the amino acids within the sequence is Pro or Arg and most preferably the tetrapeptide includes both Pro and Arg although their order and position may vary. The amino acid substitutions can be from amongst any amino acid as defined herein. Particularly preferred rigin-based tetrapeptides include Xaa-Xbb-Arg-Xcc, Xaa-Xbb-Xcc-Pro, Xaa-Xbb-Pro-Arg, wherein Xaa-Xbb-Pro-Xcc, Xaa-Xbb-Xcc-Arg, Xaa, Xbb and Xcc may be the same or different and selected from the following Xaa is Glu or the amino acids that may be substituted therefore, Xbb is Gin or the amino acids that may be substituted therefore and Xcc may be Pro or Arg or the amino acids substituted therefore. The most preferable amino acids substituted for Gly include an aliphatic side chain such as, without limitation, beta-Ala, Ala, Vai, Leu, Pro, Sarcosine (Sar) and lie. The most preferable amino acids substituted for Gin include a side chain that includes an amine group that is predominantly uncharged at neutral pH (pH 6-7) such as, without limitation, Asn, Lys, Orn, 5-hydroxyproline, Citrulline and Canavanine. When Arg is substituted, it is preferably replaced with an amino acid having a side chain that includes, predominantly, a charged nitrogen at a pH of 6, such as, without limitation, Pro, Lys, His, Desmosine and Isodesmosine. Derivatives are also considered to be encompassed by the term rigin-base tetrapeptides, (and therefore also the more generic term tetrapeptides). Derivatives include derivatives of the substituted and rearranged rigin-based tetrapeptides described herein. These derivatives include, inter alia, acyl-derivatives, which are tetrapeptides substituted with one or more straight-chain or branched-chain, long or short chain, saturated or unsaturated, substituted with a hydroxy, amino, amino acyl, sulfate or sulfide group or unsubstituted having from 1 to 29 carbon atoms. N-acyl-derivatives include those acyl groups which can be derived from acetic acid, capric acid, lauric acid, myristic acid, octanoic acid, palmitic acid, stearic acid, behenic acid, linoleic acid, linolenic acid, lipoic acid, oleic acid, isostearic acid, elaidoic acid, 2- ethylhexaneic acid, coconut oil fatty acid, tallow fatty acid, hardened tallow fatty acid, palm kernel oil fatty acid, lanolin fatty acid and the like. Preferable examples of the acyl group include an acetyl group, a palmitoyl group, an elaidoyl group, a myristyl group, a biotinyl group and an octanoyl group. These may be substituted or unsubstituted. When substituted, they are preferably substituted with hydroxyl or sulphur comprising groups such as, without limitation SO3H, SH or S-S.

[0128] Derivatives are also considered to include peptide-divalent ion complexes. Cu2+-peptide derivatives are preferred as this may provide increased biological effect compared to the peptide alone.

[0129] ALAMCAT tetrapeptides are tetrapeptides which include at least one amino acid including an aliphatic group comprising side chain. These amino acids include, without limitation, Gly, betaAla, Ala, Vai, Leu, Sarcosine (Sar) and lie. These tetrapeptides also include at least one amino acid including at least one NH2 comprising side chain. These amino acids include a side chain that has an amine group that is predominantly uncharged at neutral pH (pH 6-7) such as, without limitation, Gin, Asn, Lys, Orn, 5 -hydroxyproline, Citrulline and Canavanine. The ALAMCAT- tetrapeptides also include at least one amino acid having at least one side chain including at least one cationic amine (predominant species is charged such as NH3+, NH2+, etc. -basic amino acids which are positively charged at pH 6.0). These amino acids include, without limitation, Pro, Arg, Lys, His, Desmosine and Isodesmosine. The remaining amino acid can be any amino acid, but is preferably one comprising an aliphatic group, pendant amino group or pendant cationic group. Derivatives are also considered to be encompassed by the term ALAMCAT- tetrapeptides in accordance with the present invention, (and therefore also the more generic term tetrapeptides). These derivatives include, inter alia, acyl-derivatives, which are tetrapeptides substituted with one or more straight-chain or branched-chain, substituted or unsubstituted long or short chain, saturated or unsaturated acyl group(s) having from 1 to 29 carbon atoms. The acyl groups which can be used are the same as those described for the riginbased tetrapeptides.

[0130] Preferred embodiments include Peptide E, arg-ser-arg-lys, N-acyl-Gly-Gln-Pro-Arg peptides, most preferably N-palmitoyl-Gly-Gln-Pro-Arg.

[0131] Preferred commercially available sources of tetrapeptides include RIGIN, EYELISS, Haloxyl, and MATRIXYL 3000, which comprise between 50 to 500 ppm of palmitoyl-Gly-Gln-Pro-Arg, and other ingredients, such as peptides, chaicones and an excipient, commercially available from SEDERMA, France. Tego Pep 417 available from Evonik. These may be used to produce hydrogel compositions of the present invention by adding thereto at least one tripeptide as described herein.

[0132] The additional tetrapeptides when used are preferably incorporated into the hydrogel composition in amounts from 0.1 ppm (0.00001% w / w also referred to herein as "weight percent", "weight %" or simply by weight) to 10,000 ppm (1.0% w / w), preferably from 0.5 ppm to 1000 ppm (0.1% w / w), and most preferably from 1 ppm to 500ppm (0.05% w / w) by weight of the hydrogel composition.

[0133] The combination of tripeptides and additional tetrapeptides, can be particularly preferred. When present, the preferred ratio of additional tetrapeptide to tripeptide, or indeed the ratio of molecules having four amino acids to those having three amino acids can range from 100: 1 to 1 : 100; more preferably from 50: 1 to 1 : 50, even more preferably from 30: 1 to 1 :30 and even more preferably between 10: 1 to 1 : 10. Most preferably, the ratio of additional tetrapeptide to tripeptide ranges from between 3: 1 to 1 :3. These ratios are on a weight basis (% w / w-e.g. mg of pure peptide per Kilogram in the final formulation). In a particularly preferred embodiment, the amount of tripeptide used is greater than the amount of additional tetrapeptide used when considered in terms of their amounts in parts per million, again based on overall weight of the hydrogel composition. In a particularly preferred embodiment, the hydrogel composition comprises an additional tetrapeptide of the sequence Gly-Gln-Pro-Arg, its analogs and derivatives in combination with one or more tripeptide of the sequences Gly-His-Lys, its analogs and derivatives. Pentapeptides

[0134] The hydrogel compositions may optionally comprise a pentapeptide, derivatives of pentapeptides, and mixtures thereof. As used herein, "pentapeptides" refers to both the naturally occurring pentapeptides and synthesized pentapeptides. Also, useful herein are naturally occurring and commercially available hydrogel compositions that comprise pentapeptides. Suitable pentapeptides are those selected from the group consisting of pentapeptide 1, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 21, 22, 23, 24, 25, 26, 28, 29, 30, 31, 33, 34, 35, 36, 38, 39, derivatives thereof and mixtures thereof.

[0135] Suitable pentapeptides for use herein are the pentapeptide, lys-thr-thr-lys-ser, Arg-asp-lys-tyr- val (pentapeptide-1) and derivatives thereof. A preferred commercially available pentapeptide derivative-comprising hydrogel composition is Matrixyl which comprises 100 ppm of palmitoyl-lys-thr-thr-lys-ser and is commercially available from Sederma, France.

[0136] The pentapeptides when used are preferably incorporated into the hydrogel composition in amounts from 0.1 ppm (0.00001% w / w also referred to herein as "weight percent", "weight %" or simply by weight) to 10,000 ppm (1.0% w / w), preferably from 0.5 ppm to 1000 ppm (0.1% w / w), and most preferably from 1 ppm to 500ppm (0.05% w / w) by weight of the hydrogel composition.

[0137] Hexapeptides

[0138] The hydrogel compositions may optionally comprise a hexapeptide, derivatives of hexapeptides, and mixtures thereof. As used herein, "hexapeptides" refers to both the naturally occurring hexapeptides and synthesized hexapeptides. Also, useful herein are naturally occurring and commercially available compositions that comprise hexapeptides.

[0139] The hexapeptides when used are preferably incorporated into the hydrogel composition in amounts from 0.1 ppm (0.00001% w / w also referred to herein as "weight percent", "weight %" or simply by weight) to 10,000 ppm (1.0% w / w), preferably from 0.5 ppm to 1000 ppm (0.1% w / w), and most preferably from 1 ppm to 500ppm (0.05% w / w) by weight of the composition. Matrix metalloproteinase inhibitors (MMPi):

[0140] The term "matrix metalloproteinase inhibitor" relates to all molecule and / or plant or bacterial extracts having an inhibitory activity on at least one of the matrix metalloproteinases expressed or synthetized by or in the skin. The family of the matrix metalloproteinases is formed of several well-defined groups on the basis of their resemblance regarding structure and substrate specificity (Woessner J. F. 1991, Faseb Journal, vol. 5, , 2145). Among these groups, there are collagenases able to degrade fibrillar collagens (MMP-1 or interstitial collagenase, MMP-8 or neutrophil collagenase, MMP- 13 or collagenase 3, MMP- 18 or collagenase 4), gelatinases degrading type IV collagen or other denatured collagen form (MMP-2 or A gelatinase (72 kDa), MMP-9 or B gelatinase (92 kDa)), stromelysins (MMP-3 or stromelysin 1, MMP- 10 or stromelysin 2, MMP-1 1 or stromelysin 3) whose broad spectrum of activity targets proteins of the dermal extracellular matrix such as glycoproteins (fibronectin, laminin), proteoglycanes etc., matrilysin (MMP-7), metalloelastase (MMP- 12) or metalloproteinases (MMP- 14, MMP- 15, MMP- 16 and MMP- 17). Metalloproteinases (MMPs) are proteases that use a metal, (mostly zinc) coordinated to 3 cysteine residues and to a methionine in their active site, that degrade macromolecular components of the dermal extracellular matrix and of basal layers at neutral pH (collagen, elastin, etc.). This group of enzymes is inactivated by metal chelators. The principal activity regulators of MMPs are the tissue inhibitors of metalloproteinases or TIMPs such TIMP-I, TIMP-2, TIMP-3 and TIMP-4 (Woessner J. F., Faseb Journal, 1991). Furthermore, MMP expression is also regulated by growth factors, cytokines, oncogene products (ras, jun), or also matrix constituents.

[0141] The term "matrix metalloproteinase inhibitors " according to the present invention means all molecules able to reduce the MMP’s activity regarding the gene expression (transcription and translation) or regarding the activation of the zymogen form of the MMP, or else regarding the local control of active forms. Furthermore, the metalloproteinase inhibitors according to the present invention can also be MMP-1 inhibitors of natural or synthetic origin. The terms "natural origin" or "synthetic origin" mean both a metalloproteinase inhibitor at a pure state or in solution at different concentrations, but natural origin termed inhibitors are obtained by different extraction methods from a natural element (for example lycopene from a tomato) whereas the inhibitors of synthetic origin are all obtained via chemical synthesis.

[0142] Preferred MMPi are selected from the group consisting of retinoid, N-acetyl cysteine, glutathione, 2-furildioxime, vitamin C, flavones, isoflavones, hydrolysed rice protein, alfalfa extract, white lupin, zizyphus jujube extract, dihydroxy methyl chromone, kudzu extract, vitis vinifera extract, Oenothera biennis extract Anogeissus leiocarpus extract and mixtures thereof.

[0143] Where present, MMPi are present at a level of from 0.01% to 10%, more preferably 0.1 % to 5% and most preferably from 0.5% to 2.5% by weight of the hydrogel composition.

[0144] Skin Conditioning Agent:

[0145] The hydrogel compositions may optionally comprise a skin conditioning agent. Said skin conditioning agents may preferably be selected from the group consisting of humectants, emollients, moisturisers, or mixtures thereof. Where present, they are preferably present at a level of from 0.01% to 20%, more preferably from 0. 1% to 10%, most preferably from 0.5% to 7% by weight of the hydrogel composition.

[0146] Preferred skin conditioning agents are selected from the group consisting of guanidine, urea, glycolic acid and glycolate salts, salicylic acid, lactic acid and lactate salts, aloe vera, shea butter, polyhydroxy alcohols, such as sorbitol, mannitol, xylitol, erythritol, glycerol, hexanetriol, butanitriol, (di) propylene glycol, butylene glycol, hexylene glycol, polyethylene glycol, sugars (e.g. fructose, glucose, xylose, honey, mannose, xylose), gluconodeltalactone, and starches and their derivatives, pyrrolidone, carboxylic acid, hyaluronic acid and salts thereof, lactamide monoethanolamine, acetamide monoethanolamine, panthenol, allantoin and mixtures thereof.

[0147] In some embodiments said skin conditioning agent is selected from the group consisting of glycerine, arabinogalactan, butylene glycol, hyaluronic acid, shea butter, propylene glycol, ethylhexyl glycerine, caprylyl glycol, hyaluronate and mixtures thereof.

[0148] In a preferred embodiment, the hydrogel composition comprises skin conditioning agents selected from ethylhexyl glycerine, caprylyl glycol, saccharide isomerate, butylene glycol, pentylene glycol and xylitylglcoside, anhydroxylitol, xylitol or combinations thereof, most preferably ethylhexyl glycerine, caprylyl glycol or a combination thereof. In some embodiments these skin conditioning agents are present in an amount of from 0.01 to 5 wt.%, such as from 0.1 to 5 wt.%. Antioxidant Agent:

[0149] The hydrogel compositions may optionally comprise an antioxidant agent. Suitable antioxidant agents may include: a) ascorbic acid its salts, esters, glucosides and glucosamines, particularly sodium ascorbyl phosphate, magnesium ascorbyl phosphate, ascorbyl palmitate and ethyl ascorbic acid b) vitamin E (tocopherol) and its esters, particularly tocopheryl acetate, as well as Dimethyl methoxy chromanol which is a synthetic analogue of gamma tocopherol, available from Lipotec S.A. polygon Industrial Camri Rai, under the tradename Lipochroman-6 c) herbal extracts, particularly gingko biloba, such as that available under the trade name "Gingko Biloba Leaf Powder" from Univar PLC, morus alba, such as that available under the trade name "Mulberry Concentrate" from Solabia, origanum vulgare, such as that available under the trade name "Pronalen Origanum HSC" from S Black Ltd, panax ginseng, such as that available under the trade name "Panax ginseng 1.1 extract 4294"from S Black Ltd or "Phytexcell Panax ginseng" available from Croda Chemicals Ltd, birch extract such as those available from Cosmetochem (U. K.) Ltd under the trade names "Super Herbasol Extract Birch" and "HP Herbasol Betula" and those available from Blagden Chemicals under the tradenames "Phytelene of Birch" and "Aqueous Spray Dried Birch", camellia sinensis, such as that available under the trade name "Herbal Extract Green Tea 75% Solids" from Nichimen Europe, rosmarinus officinalis, such as that available under the trade name "Pronalen Rosemary" from S. Black, Acerola cherry powder, such as that available as Acerola PE from Gee Lawson, Emblica extract sold under the tradename Emblica™ by Merck Speciality chemicals, and Grape Seed oil, such as that available from Chesham Chemicals Limited.

[0150] The amounts of antioxidant agents used in the cosmetic hydrogel composition are expressed as dry weights, as understood by a man skilled in the art. The total amount of antioxidant agents optionally present in the hydrogel composition may range from 0.005% to 10% by weight, preferably 0.5% to 5%, most preferably 0.2% to 1.5% by weight of the hydrogel composition.

[0151] Particularly preferred synergistic combinations of antioxidant agents suitable for inclusion in the hydrogel composition are selected from the group consisting of : i) panax ginseng, morus alba and magnesium ascorbyl phosphate; ii) panax ginseng, morus alba and sodium ascorbyl phosphate; iii) panax ginseng, morus alba and rosmarinus officinalis; iv) ginkgo biloba, phyllanthus emblica and Dimethylmethoxy chromanol; v) morus alba, camellia sinensis and dimethylmethoxy chromanol; vi) morus alba, camellia sinensis and tocopheryl acetate; vii) panax ginseng, morus alba and origanum vulgare, viii) camellia sinensis, tocopheryl acetate and dimethylmethoxychromanol, viv) morns alba, tocopheryl acatete and dimethylmethoxy chromanol .

[0152] In these preferred combinations (a) the panax ginseng is preferably present in an amount of 0.005% to 0.1%, more preferably 0.01% to 0.05% by weight of the hydrogel composition; (b) the morns alba is preferably present in an amount of 0.0005% to 0.01%, more preferably 0.001% to 0.005% by weight of the hydrogel composition; (c) the sodium, magnesium ascorbyl phosphate or ethyl ascorbic acid is preferably present in an amount of 0.05% to 2.5%, preferably 0.1% to 2%, most preferably 0.15% to 1.5% by weight of the hydrogel composition;(d) the rosmarinus officinalis or origanum vulgare or phyllanthus emblica is preferably present in an amount of 0.01% to 0.5%, more preferably 0.05% to 0.2% by weight of the hydrogel composition e) the dimethylmethoxy chromanol is preferably present in an amount of 0.0005% to 0.1%, more preferably from 0.005% to 0.05% by weight of the hydrogel composition; f) the camellia sinensis is preferably present in an amount of 0.005% to 0.2%, more preferably from 0.01% to 0.1% and the g) Tocopherol acetate is preferably present in an amount of 0.01 to 0.5%, more preferably from 0.05% to 0.25%

[0153] In some embodiments the antioxidant agent comprises a Camellia japonica extract in combination with a second antioxidant agent is selected from the group consisting of dimethylmethoxy chromanol, Myrtus communis leaf extract, green tea extract, Quercus petraea fruit extract, Phyllanthus emblica fruit extract, ginkgo extract, ferulic acid, Argania spinosa leaf extract, Pinus pinaster bark extract, and resveratrol and optionally a third antioxidant agent selected from the group consisting of dimethylmethoxy chromanol, Myrtus communis leaf extract, green tea extract, Quercus petraea fruit extract, Phyllanthus emblica fruit extract, ginkgo extract, ferulic acid, Argania spinosa leaf extract, Pinus pinaster bark extract, and resveratrol, wherein the third antioxidant is not the same as the second antioxidant agent.

[0154] In some embodiments the antioxidant agent comprises Argania spinosa leaf extract, Pinus pinaster bark extract and resveratrol. In some embodiments the antioxidant agent comprises Myrtus communis leaf extract, ginkgo extract and ferulic acid. In some embodiments the antioxidant agent comprises Myrtus communis leaf extract, Quercus petraea fruit extract and Pinus pinaster bark extract. In some embodiments the antioxidant agent comprises green tea extract, Quercus petraea fruit extract and ferulic acid. Salicylic Acid Compound:

[0155] The hydrogel compositions may comprise a salicylic acid compound, its esters, its salts, or combinations thereof. In one embodiment of the hydrogel compositions of the present invention, the salicylic acid compound preferably comprises from 0.0001% to 25%, more preferably from 0.001% to 15%, even more preferably from 0.01% to 10%, still more preferably from 0.1% to 5%, and even more preferably from 0.01% to 2%, more preferably 0.1% to 2% by weight of the hydrogel composition, of salicylic acid.

[0156] Other components:

[0157] The hydrogel composition may comprise preservatives, such as phenoxyethanol and / or potassium sorbate. The skilled person would understand that other preservatives and the concentrations that preservatives should be used in are known in the art. The hydrogel composition may also comprise sodium hydroxide, possibly added in an aqueous solution such as a 30 wt.% by weight of the solution in water solution.

[0158] Method of Preparation

[0159] The invention further provides a method of preparing the hydrogel composition of the present invention. The method comprises providing the hydrogel system and adding to the hydrogel system the peptide to provide the hydrogel composition of the present invention.

[0160] In some embodiments the hydrogel composition is a patch suitable for application to the skin. In these embodiments the method may further comprise the additional step of moulding the hydrogel composition to form a patch, optionally by pouring the hydrogel composition heated above room temperature, such as to at least 70°C in a mould for forming the hydrogel composition into a patch and allowing the hydrogel composition to cool.

[0161] In some embodiments, the method further comprises the steps of forming the hydrogel system. These steps may comprise providing the one or more thickener as defined herein, providing the solvent system as defined herein; and combining the one or more thickener and the solvent system to provide the hydrogel system. In some embodiments, the method therefore comprises:

[0162] (i) providing the one or more thickener as defined herein;

[0163] (ii) providing the solvent system as defined herein;

[0164] (iii) combining the thickener and the solvent system, to provide the hydrogel system;

[0165] (iv) adding to the hydrogel system the peptide as defined herein; and

[0166] (v) moulding the loaded hydrogel composition to form a patch.

[0167] In some embodiments the method comprises the steps of adding one of the ether, the organic solvent of the solvent system to the one or more thickener, mixing the resulting combination, and then adding the surfactant of the solvent system.

[0168] In some embodiments, the method further comprises heating the combination of the one or more thickener and solvent system, optionally with mixing, before the addition of the peptide. For example, the method may comprise heating the combination of the thickener and solvent system to from 50°C to 99°C, such as from 60°C to 95°C or from 80°C to 95°C. Where the method comprises heating the combination, the method may further comprise cooling the hydrogel system formed to a temperature of 80°C or less, such as 75°C or less or from 5°C to 80°C, before adding the peptide.

[0169] Where glycerin is present, this may be added along with the organic solvent. The method may comprise an additional step of first heating the water of the solvent system to from 70 to 99 °C, performing steps (i) to (iii) above less the water of the solvent system, and adding to the heated water the resulting combination of step (iii).

[0170] In some embodiments, the method further comprises adding a base to the hydrogel system before the addition of the peptide. For example, sodium hydroxide may be added at this point, for example a sodium hydroxide solution having a concentration of from 10 to 50 wt.% sodium hydroxide in water may be added in an amount of from 0.01 to 0.1 wt.%.

[0171] Use and Method of Use

[0172] The present invention also provides a method for stimulating the production of extracellular proteins in humans, the method comprising administering to the skin of said human a hydrogel composition described herein, optionally wherein the hydrogel composition is moulded into a patch. The present invention therefore also provides the use of the hydrogel composition described herein, optionally formed into a patch, in the method for stimulating the production of extracellular proteins in humans.

[0173] The present invention also provides a non-therapeutic cosmetic method to improve the condition of the skin and / or lines and / or wrinkles and / or imperfections, the method comprising application of the hydrogel composition of the present invention to the skin of a human.

[0174] The present invention also provides a use of the hydrogel composition of the present invention for the non-therapeutic cosmetic treatment to improve the condition of the skin and / or lines and / or wrinkles and / or imperfections, such as the appearance of scars.

[0175] The present invention provides the hydrogel composition of the present invention for use in a method for treating a wound in the skin of a subject, the method comprising applying the hydrogel composition of the present invention to the wound in the skin of the subject. That is, the present invention further provides a method for treating a wound in the skin of a subject, the method comprising applying the hydrogel composition of the present invention to the wound in the skin of the subject.

[0176] The subject may be a human or animal. In a preferred embodiment the subject is a human.

[0177] To heal a wound effectively, the skin barrier needs to be restored quickly to prevent further damage and infection (healing speed). After closing, wound remodelling occurs to reduce the severity of or prevent scarring (healing maturity). The present inventors have identified that specific peptide combinations are effective in accelerating wound closure (healing speed) and enhance the quality of the repair (healing maturity), thus providing an improved outcome for patients and consumers.

[0178] Brief Description of the Drawings

[0179] Figure 1 is a schematic diagram showing a HSP sphere within a 3D HSP space.

[0180] Figure 2 is a HSP space showing a HSP sphere of a solute, which surrounded by and containing smaller HSP spheres of solvents. Figure 3 is a depth profile analysis following 2 hours exposure, using a Franz cell system, of porcine skin to a hydrogel composition in accordance with the present invention

[0181] Figure 4a and 4b provide depth profile analysis for upper layers of the porcine skin, area under the curve (AUC) analysis that was performed after best fit polynomial functions were applied to the curves for both the formulation (Figure 4B) and for the hydrogel compositions (Figure 4A) containing Pal-GHK.

[0182] Figure 5 provides analysis data for individual pieces of hydrogel compositions after they had been used in 2-hour Franz cell runs.

[0183] Examples

[0184] Tetrapeptide synthesis

[0185] The tetrapeptides of the present disclosure with generic formulation pal-X’X2X3X4-OH are prepared by peptidic synthesis. In a first step the N-terminal of X4is coupled with a resin via the terminal acid functionality in the presence of a coupling agent. The N terminal amine is then reacted with the next amino acid in the sequence X3in the presence of a coupling agent. The same process is repeated until the required sequence is obtained and a suitable C terminal functionality added. Suitable coupling agents include DCC (dicyclohexylcarbodiimide)ZNHS (N-hydroxysuccinimide) or HBTU (2-(lH-benzotriazole-l-yl)-l,l,3,3-tetramethyluronium hexafluorophosphate) / HOBT (1-hydroxy-benzotriazole)). The resulting peptide is then cleaved from the resin in an acidic medium and after precipitation, washing and drying, the peptide is obtained in solid form.

[0186] Solvent Identification

[0187] Hansen Solubility Parameter (HSP) information was used to screen solvents, allowing the identification of solvent system candidates selected for suitability in cosmetic applications. These solvent systems were then further investigated to identify suitability in peptide delivery applications. Hansen Solubility Parameter (HSP):

[0188] HSP refers to three different parameters of a molecule or compound, i.e., 5D = Dispersion forces (Van der Waals), 5P = Polarity, and 5H = Hydrogen bonding (see Figure 1). Many molecules and compounds have HSP values, which (once determined) can be plotted in a HSP space (see Figure 2). Knowledge of these parameters for a given solute (e.g., a peptide or skin) and solvent enables one to determine if the solute will dissolve in the solvent. A solvent, with similar HSP values to a solute, is likely to dissolve the solute. In other words, the more similar the HSP values of a solute are to those of a solvent, the more likely that solute will dissolve in the solvent.

[0189] HSP Sphere:

[0190] The HSP values of a solute and a solvent do not need to be identical for complete solubility to occur. The solvents for a given solute tend to have HSP values that are close in 3D proximity to the HSP values of the solute. Likewise, molecules that are incapable of dissolving a given solute have HSP values that are more distal from those of the solute. Thus, by testing the ability of several different solvents, each with known HSP values, to dissolve a given solute, a boundary (i.e., a HSP sphere) can be generated for the solute with the HSP values of the solute being in the centre of sphere (see Figure 2). Thus, solvents with HSP values within the sphere can be used to dissolve the solute, whereas potential solvents with HSP values outside the sphere cannot be used to dissolve the solute. The diameter / radius of a sphere varies from solute to solute.

[0191] Identifying the HSP Sphere:

[0192] Identifying the HSP sphere according to the invention may comprise referring to reference HSP values, e.g., for skin and / or the two or more test solvents. Reference HSP values can be obtained by determining the chemical structure of the relevant chemical using Simplified molecular- input line-entry system (SMILES) notification, and then using the software, HSPiP (https: / / www.hanseii-solubility.com / dowiiloads.php; and / or the software, Formulating For Efficacy™ (https: / / www.jwsolutionssoftware.com / ). Reference HSP values can also be obtained from various websites online, including, for example, .specialchem.com.

[0193] Identifying the HSP sphere (for one or more of members selected from the group consisting of skin, the peptide and the two or more test solvents) may comprise using one or more different reference solvents, i.e., known solvents with known HSP values. Identifying the HSP sphere may comprise determining the HSP values using one or more, two or more, three or more, four or more, five or more, six or more, seven or more, eight or more, nine or more, 10 or more, 11 or more, 12 or more 13 or more, 14 or more, 15 or more, 16 or more, 17 or more, 18 or more, 19 or more, 20 or more, 25 or more, 30 or more, 35 or more, 40 or more, 45 or more, or 50 or more different reference solvents.

[0194] Identifying the HSP sphere may comprise ranking each reference solvents based on their ability to dissolve skin, the peptide and / or a test solvent by using the qualitative criteria of a solubility rank.

[0195] The solubility rank may use the following qualitative criteria:

[0196] Reference solvents that are ranked 1 completely dissolve skin, the peptide or a test solvent and thus fall within the relevant HSP sphere. Reference solvents that are ranked 2, 3, 4 or 5 may, or may not fall within the HSP sphere of skin, the HSP sphere of the peptide or the HSP sphere of a test solvent. Reference solvents that are ranked 6 do not dissolve skin, the peptide or a test solvent and thus fall outside the relevant HSP sphere.

[0197] Reference solvents with a solubility rank of 4, 3, 2 or 1 may be used to identify the HSP sphere of skin, the HSP sphere of the peptide, or the HSP sphere of a test solvent. Preferably, reference solvents with a solubility rank of 3, 2 or 1 are used to identify the HSP sphere of skin, the HSP sphere of the peptide, or the HSP sphere of a test solvent. More preferably, reference solvents with a solubility rank of 2 or 1 are used to identify the HSP sphere of skin, the HSP sphere of the peptide, or the HSP sphere of a test solvent. Most preferably, reference solvents with a solubility rank of 1 are used to identify the HSP sphere of skin, the HSP sphere of the peptide, or the HSP sphere of a test solvent. Once the HSP sphere of the skin and the peptide have been identified, solvents having an HSP that bridges this gap can be identified. Around 150 solvents having a suitable HSP were identified. Solvents from this group were selected based on their known tolerance on the skin, compatibility in desired formulation types. The present inventors then identified solvent systems comprising various levels of selected solvents from this filtered group for further investigation.

[0198] The amount of peptide dissolved in a given solvent system that would enter the skin from this solvent system can be calculated using Formulating For Efficacy™ (https: / / www.iwsolutionssoftware.com / ) software.

[0199] Table 1 provides the calculated peptide IN (that is, amount of peptide that would transition into the skin) for seven solvent systems. Wt.% is of the solvent system and the calculations only consider selected solvents in the system, ignoring other components that might make the solvent system up to 100 wt.%.

[0200] Solvent systems comprising polysorbate 20 are calculated to provide better penetration of peptide into the skin than systems without polysorbate 20. It can also be seen that of systems comprising 1 wt.% polysorbate 20, those comprising 20 or 30 wt.% of propanediol provide better penetration of peptide into the skin than those solvent systems with a lower amount of propanediol.

[0201] Table 1

[0202] The invention will now be further described through reference to specific example formulations and the associated methods of manufacture. Example 1 - Hydrogel composition 1

[0203] Method of manufacture

[0204] 1. In the main vessel add water and potassium sorbate. Heat to 85-90°C and stir for 2 minutes.

[0205] 2. In a separate vessel weigh out glycerin, propanediol, phenoxyethanol, caprylyl glycol & ethylhexylglycerin, gellan gum, cyamopsis tetragonoloba (Guar) gum and xanthan gum. Heat to 85-90°C and stir for 2 minutes.

[0206] 3. Add step 2 to step 1 slowly with stirring maintaining the temperature at 85-90°C.

[0207] 4. Continue to stir at 85-90°C for 45 minutes.

[0208] 5. Add polysorbate 20 and stir for 5 minutes.

[0209] 6. Add sodium hydroxide and stir for 5 minutes. 7. Cool to 75°C.

[0210] 8. Add Niacinamide dissolved in a small amount of water.

[0211] 9. Add palmitoyl tetrapeptide-94 (Pal-GPKG) & palmitoyl tetrapeptide-95 (Pal-LSVD) and stir for 5 minutes.

[0212] 10. Pour into moulds and allow to cool to room temperature. Example 2 - Hydrogel composition 2 Method of manufacture

[0213] 1. In the main vessel add water and potassium sorbate. Heat to 85-90°C and stir for 2 minutes.

[0214] 2. In a separate vessel weigh out glycerin, propanediol, phenoxyethanol, caprylyl glycol & ethylhexylglycerin, gellan gum, cyamopsis tetragonoloba (Guar) gum and xanthan gum. Heat to 85-90°C and stir for 2 minutes.

[0215] 3. Add step 2 to step 1 slowly with stirring maintaining the temperature at 85-90°C.

[0216] 4. Continue to stir at 85-90°C for 45 minutes.

[0217] 5. Add polysorbate 20 and stir for 5 minutes. 6. Add sodium hydroxide and stir for 5 minutes.

[0218] 7. Cool to 75°C.

[0219] 8. Add Niacinamide dissolved in a small amount of water.

[0220] 9. Add palmitoyl tetrapeptide-94 (Pal-GPKG) & palmitoyl tetrapeptide-95 (Pal-LSVD) and stir for 5 minutes. 10. Pour into moulds and allow to cool to room temperature.

[0221] Example 3 - Hydrogel composition 3

[0222] Method of manufacture

[0223] 1. In the main vessel add water and potassium sorbate. Heat to 85-90°C and stir for 2 minutes. 2. In a separate vessel weigh out glycerin, dimethylisosorbide, phenoxyethanol, caprylyl glycol & ethylhexylglycerin, gellan gum, cyamopsis tetragonoloba (Guar) gum and xanthan gum. Heat to 85-90°C and stir for 2 minutes.

[0224] 3. Add step 2 to step 1 slowly with stirring maintaining the temperature at 85-90°C.

[0225] 4. Continue to stir at 85-90°C for 45 minutes. 5. Add polysorbate 20 and stir for 5 minutes.

[0226] 6. Add sodium hydroxide and stir for 5 minutes.

[0227] 7. Cool to 75°C.

[0228] 8. Add Niacinamide dissolved in a small amount of water. 9. Add palmitoyl tetrapeptide-94 (Pal-GPKG) & palmitoyl tetrapeptide-95 (Pal-LSVD) and stir for 5 minutes.

[0229] 10. Pour into moulds and allow to cool to room temperature. Example 4 - Hydrogel composition 4

[0230] Method of manufacture

[0231] 1. In the main vessel add water and potassium sorbate. Heat to 85-90°C and stir for 2 minutes.

[0232] 2. In a separate vessel weigh out glycerin, propanediol, phenoxyethanol, caprylyl glycol & ethylhexylglycerin, gellan gum, cyamopsis tetragonoloba (Guar) gum and xanthan gum. Heat to 85-90°C and stir for 2 minutes.

[0233] 3. Add step 2 to step 1 slowly with stirring maintaining the temperature at 85-90°C.

[0234] 4. Continue to stir at 85-90°C for 45 minutes.

[0235] 5. Add polysorbate 20 and stir for 5 minutes. 6. Add sodium hydroxide and stir for 5 minutes.

[0236] 7. Cool to 75°C.

[0237] 8. Add Niacinamide dissolved in a small amount of water.

[0238] 9. Add palmitoyl tetrapeptide -7 & palmitoyl tripeptide-1 and acetyl dipeptide-1 cetyl ester and stir for 5 minutes.

[0239] 10. Pour into moulds and allow to cool to room temperature.

[0240] Example 5 - Hydrogel composition 5

[0241] Method of manufacture

[0242] 1. In the main vessel add water and potassium sorbate. Heat to 85-90°C and stir for 2 minutes. 2. In a separate vessel weigh out glycerin, propanediol, phenoxyethanol, caprylyl glycol

[0243] & ethylhexylglycerin, gellan gum, cyamopsis tetragonoloba (Guar) gum and xanthan gum. Heat to 85-90°C and stir for 2 minutes.

[0244] 3. Add step 2 to step 1 slowly with stirring maintaining the temperature at 85-90°C. 4. Continue to stir at 85-90°C for 45 minutes.

[0245] 5. Add polysorbate 20 and stir for 5 minutes.

[0246] 6. Add sodium hydroxide and stir for 5 minutes.

[0247] 7. Cool to 75°C.

[0248] 8. Add Niacinamide dissolved in a small amount of water.

[0249] 9. Add water tetrapeptide-7 & palmitoyl tripeptide-1 and acetyl dipeptide-1 cetyl ester and stir for 5 minutes.

[0250] 10. Pour into moulds and allow to cool to room temperature.

[0251] Examples of Enhanced Delivery of Active Ingredients by Hydrogel

[0252] Ex vivo analysis was carried out using Franz Cell Methodology1and 3D OrbiSIMS Analysis2to demonstrate the enhanced delivery of active ingredients provided by the use of hydrogels in accordance with the present invention.

[0253] FRANZ CELL METHODOLOGY

[0254] For ex vivo analysis, porcine skin samples were obtained post-slaughter from the ears of pigs reared specifically for food. Underlying cartilage and excess subcutaneous fat were removed from the skin, creating full thickness skin sections of approximately 2 mm. The skin was stored at -20°C until required. The porcine skin was mounted dermal side down in a Franz- type static diffusion cell set-up1, with an exposed surface area of approx. 1.1 cm2. An infinite dose of the cosmetic formulation, containing the Pal-GHK peptide (Palmitoyl Tripeptide-1), was applied to the donor chamber, with the porcine skin exposed to the formulations for 2 hours in a water bath set to 36.5 °C. The hydrogel composition, containing the hydrogel system and Pal-GHK peptide, were placed on top of the porcine skin and the Franz cell assembled ensuring contact was maintained between the hydrogel composition and the skin. Hydrogel systems without Pal-GHK were also tested. Sink conditions were maintained throughout the experiments. After 2 hours, the Franz cells were dismantled, the hydrogels and any excess formulation was removed with a dry sponge and the porcine skin was wiped with a sponge soaked in Teepol Solution (3 % v / v). Next, the skin was dehydrated under vacuum at room temperature for 24 hours before 3D OrbiSIMS analysis. In addition, samples of the hydrogel compositions containing Pal-GHK were also dehydrated after the 2-hour Franz cell run and 3D OrbiSIMS analysis used to compared to samples that were not run in the Franz cell system. 3D ORBISIMS ANALYSIS

[0255] The 3D OrbiSIMS combines secondary ion mass spectrometry with the high mass-resolving power of an OrbitrapTM mass analyser, facilitating in situ label free molecular analysis and the identification of organic species in complex solid samples, including biological tissues2. 3D OrbiSIMS analysis of porcine skin samples was performed on a Hybrid SIMS instrument (IONTOF GmbH) under the following conditions; A 20 KeV Ar3000+ analysis beam with a diameter of 20pm was used as the primary ion source. Samples were analysed at ambient temperature across a 400 x 400pm area in positive polarity with sawtooth raster mode and a total crater size of 486 x 486pm. Duty cycle was set to 4.4% and cycle time to 200ps. Mass spectra were recorded at a resolution of 240,000 at m / z 200 in the mass range of 75 to 1, 125 m / z. Both data acquisition and the subsequent data processing were performed using SurfaceLab 7 software (IONTOF GmbH). Ionisation of Pal-GHK was used as a marker to track penetration of active ingredients through the porcine skin, with ion intensity plotted against sputter time, which represents depth. Area under the curve analysis was performed to compare penetration of Pal-GHK in the different systems.

[0256] RESULTS

[0257] Depth profile analysis after 2-hour Franz cell experiments revealed there was considerably more Pal-GHK detected in the porcine skin with the hydrogel composition, compared to the formulation (Figure 3). There was no Pal-GHK detected in the porcine skin where hydrogel systems without Pal-GHK were applied. Not only did the data reveal considerably more Pal- GHK detected in the upper layers of the porcine skin after use of the hydrogel compositions, the data also suggested that Pal-GHK penetrated deeper into the skin after 2 hours, with the ion clearly detected beyond a sputter time of 1000 seconds. Sputter times in this region are known to represent depth to epidermal layers of the skin.

[0258] For the upper layers of the porcine skin area under the curve (AUC) analysis was performed after best fit polynomial functions were applied to the curves for both the formulation and hydrogel compositions containing Pal-GHK (Figures 4A & 4B). X-axis limits were set at 2.5 and 60. The results showed a greater than 6-fold increase in the ion intensity for the hydrogel composition (AUC = 12,114.5) compared to the formulation (AUC = 1928.4). Analysis of 9 separate pieces of the hydrogel compositions themselves after a 2-hour Franz cell run revealed that only approximately 10% of the Pal-GHK was detectable compared to samples of the hydrogel systems that had not been put on porcine skin (Figure 5). This suggests that over 90% of the Pal-GHK peptide was delivered into the skin within 2 hours.

[0259] Overall, these data suggest enhanced delivery of active ingredients into the skin using the hydrogel composition compared to a traditional cosmetic formulation.

[0260] REFERENCES 1. Franz TJ (1975) Percutaneous absorption; On the relevance of in-vitro data. J Invest

[0261] Dermatol 64: 190-195

[0262] 2. Passarelli MK, Pirkl A, Moellers R, et al (2017) The 3D OrbiSIMS - label-free metabolic imaging with subcellular lateral resolution and high mass-resolving power. Nature Methods 14: 1175-1186.

Claims

45CLAIMS1. A hydrogel composition comprising a hydrogel system and a peptide; wherein the hydrogel system comprises; one or more thickener, and a solvent system comprising one or more organic solvent and one or more surfactant.

2. The hydrogel composition of claim 1, wherein the peptide comprises a first tetrapeptide, or a second peptide, or a third tetrapeptide, or a fourth tetrapeptide or a combination of the first tetrapeptide and the second peptide, wherein:(a) the first tetrapeptide is selected from the group consisting of tetrapeptides having the amino acid sequence U-LSXX-Z wherein L is used to denote amino acid Leucine and S is used to denote Serine, as per the internationally recognised single letter code for amino acids, X denotes an amino acid independently selected from the group consisting of Valine (V), Aspartic acid (D), Proline (P), Glycine (G), at the N-terminal end, U is selected from the group consisting of H, -CO-R1, -SO2-R1or a biotinyl group, at the C-terminal end, Z is selected from the group consisting of OH, OR1, NHR1or NR'R2; R1and R2are independently selected from the group consisting of alkyl, aryl, aralkyl, alkylaryl, alkoxy, saccharide and aryloxy group, which may be linear, branched, cyclical, polycyclic, unsaturated, hydroxylates, carbonylated, phosphorylated and / or sulphurous, said groups comprising from 1 to 24 carbon atoms and being capable of including one or more heteroatoms O, S and / or N;(b) the second peptide is selected from the group consisting of tetrapeptides having the amino acid sequence U-GPXG-Z wherein G is used to denote amino acid glycine and P denotes the amino acid proline, as per the internationally recognised single letter code for amino acids, X denotes an amino acid independently selected from the group consisting of Lysin(K), Glutamic acid (E), Proline (P) and Serine (S) and mixtures thereof, or a pentapeptide having the amino acid sequence according to SEQ ID No: 17, at the N-terminal end, U is selected from the group consisting of H, -CO-R1, -SO2-R1or a biotinyl group, at the C-terminal end, Z is selected from the group consisting of OH, O R1, NHR1or NR'R2; R1and R2are independently selected from the group consisting of alkyl, aryl, aralkyl, alkylaryl, alkoxy, saccharide and aryloxy group, which may be linear, branched, cyclical, polycyclic, unsaturated, hydroxylates,46 carbonylated, phosphorylated and / or sulphurous, said groups comprising from 1 to 24 carbon atoms and being capable of including one or more heteroatoms O, S and / or N;(c) the third tetrapeptide is selected from the group consisting of tetrapeptides having the amino acid sequence U-XXGD-Z wherein G is used to denote amino acid Glycine and D is used to denote amino acid Aspartic acid, as per the internationally recognised single letter code for amino acids; X denotes an amino acid selected from the group consisting of Glutamic acid (E), Lysine (K), Leucine (L), Alanine (A), Isoleucine (I), Arginine (R) and mixtures thereof; at the N-terminal end, U is selected from the group consisting of H, -CO-R1, -SO2-R1or a biotinyl group; at the C-terminal end, Z is selected from the group consisting of OH, OR1, NHR1or NR'R2; R1and R2are independently selected from the group consisting of alkyl, aryl, aralkyl, alkylaryl, alkoxy, saccharide and aryloxy group, which may be linear, branched, cyclical, polycyclic, unsaturated, hydroxylates, carbonylated, phosphorylated and / or sulphurous, said groups comprising from 1 to 24 carbon atoms and being capable of including one or more heteroatoms O, S and / or N;(d) the fourth tetrapeptide is a tetrapeptide having the amino acid sequence U-QTAV-Z wherein Q is used to denote amino acid Glutamine, T is used to denote amino acid Threonine, A is used to denote amino acid Alanine and V is used to denote amino acid Valine; at the N- terminal end, U is independently selected from the group consisting of octanoyl (C8), decanoyl (CIO), lauroyl (C 12), myristoyl (C 14), palmitoyl (C16), stearoyl (C 18), biotinoyl, elaidoyl, oleoyle, lipoyle; at the C-terminal end, Z is selected from the group consisting of OH, OR1, NHR1or NR'R2; R1and R2are independently selected from the group consisting of alkyl, aryl, aralkyl, alkylaryl, alkoxy, saccharide and aryloxy group, which may be linear, branched, cyclical, polycyclic, unsaturated, hydroxylates, carbonylated, phosphorylated and / or sulphurous, said groups comprising from 1 to 24 carbon atoms and being capable of including one or more heteroatoms O, S and / or N.

3. The hydrogel composition of claim 1 , wherein the peptide comprises:(e) palmitoyl tetrapeptide-7 (Pal-GQPR), or palmitoyl tripeptide-1 (Pal-GHK), or acetyl dipeptide- 1 cetyl ester (N-acetyl-YR hexadecyl ester), or a combination of Pal-GQPR and Pal- GHK, or a combination of all three.

4. The hydrogel composition of any one of the preceding claims, wherein the organic solvent is a diol, an isosorbide or derivative thereof, or a combination thereof.

475. The hydrogel composition of claim 4, wherein the organic solvent is a diol present in an amount of from 1 to 40 wt.%, wherein, optionally, the diol is propanediol.

6. The hydrogel composition of claim 4, wherein the organic solvent is an isosorbide or derivative thereof present in an amount of from 2 to 30 wt.%, optionally wherein the isosorbide or derivative thereof is isosorbide ether, optionally wherein the isosorbide ether is dimethyl isosorbide.

7. The hydrogel composition of claim 4, wherein the organic solvent is a combination of diol and an isosorbide or derivative thereof, wherein the diol is present in an amount of from 1 to 10 wt.% and the isosorbide or derivative thereof is present in an amount of from 2 to 40 wt.%.

8. The hydrogel composition of any one of the preceding claims, wherein the one or more surfactant is: a) a polysorbate; b) selected from polysorbate 20, polysorbate 40, polysorbate 60, polysorbate 80, or combinations thereof; and / or c) present in an amount of from 0.1 wt.% to 5 wt.%.

9. The hydrogel composition of any one of the preceding claims, wherein the solvent system comprises glycerin in addition to the organic solvent.

10. The hydrogel composition of any one of the preceding claims, wherein the one or more thickener is:(a) present in an amount of from 0.1 wt.% to 5 wt.%, and / or(b) selected from carrageenan, amorphophallus konjac root extract, caesalpinia spinosa gum, sclerotium gum, agar, locust bean gum, cellulose, starch, chitin, hyaluronic acid, guar gum, gellan gum, xanthan gum, chitosan, phosphoryl choline, acrylic acid, sodium polyacrylate, polyvinyl acetate (PVAc) or combinations thereof.

11. A method of preparing the hydrogel composition of any one of the preceding claims, the method comprising providing the hydrogel system and adding the peptide to provide the hydrogel composition.

12. The method of claim 11 wherein the method further comprises the steps of preparing the hydrogel system, the steps comprising providing the one or more thickener; providing the solvent system; and combining the one or more thickener and solvent system to provide the hydrogel system.

13. A method for stimulating the production of extracellular proteins in humans, the method comprising administering to the skin of said human the hydrogel composition of any one of claims 1 to 10.

14. A use of the hydrogel composition of any one of claims 1 to 10 in a non-therapeutic cosmetic treatment to improve the condition of the skin and / or lines and / or wrinkles and / or imperfections.

15. The hydrogel composition of any one of claims 1 to 10 for use in a method for treating a wound in the skin of a subject, the method comprising applying the hydrogel composition of the present invention to the wound in the skin of the subject.

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