Phospholipase c inhibitor peptides and uses thereof

Abstract

The present invention provides a peptide or a pharmaceutically, veterinary, or cosmetically acceptable salt thereof, wherein the peptide comprises at least 5 consecutive amino acid residues of a sequence at least 80% identical to the sequence TTDEGTASSEVNATEEMSTLVN (SEQ ID NO: 1) or the sequence EQAWMASYKYVGATTNIHPYLS (SEQ ID NO: 2); the peptide has a length equal to or lower than 22 amino acid residues. The invention also provides a pharmaceutical, veterinary, and cosmetic compositions comprising the peptides, and therapeutic and cosmetic uses thereof.

Description

[0001] Phospholipase C inhibitor peptides and uses thereof

[0002] This application claims the benefit of European Patent Application EP23383327.6 filed on December 20th, 2023.

[0003] Technical Field

[0004] The present invention belongs to the field of pharmacy, veterinary and cosmetics. In particular, it relates to therapeutic and cosmetic peptides that are capable of inhibiting phospholipase C activity, and to their pharmaceutical, veterinary and cosmetic uses.

[0005] Background Art

[0006] Phospholipase C (PLC) is a family of enzymes that includes 6 subfamilies, PLC-p, 5, y, E, ( and q, and 13 isoforms in humans. PLCs are activated through G protein-coupled receptors and its activation generates downstream pathways being PIP2 hydrolyzed in DAG and IP3 and resulting in elevated Ca+2 and PKG activation.

[0007] PLC activation has been widely characterized as a mediator in several diseases or disorders, including pain, cancer, inflammation, skin disease and cardiovascular disease (Kadamur G and Ross EM, "Mammalian phospholipase C", Annual review of physiology, 2013, vol. 15, pp 127-154; Cocco L et al., "Phosphoinositidespecific phospholipase C in health and disease", Journal of Lipid Research, 2015, vol. 56 (10), pp.1853- 1860). Among other effects, PLC signaling cascade induces direct neuronal potentiation and consequent peripheral inflammatory sensitization.

[0008] Although the PLC family is a promising therapeutic and cosmetic target, non-specific PLC inhibitors have revealed undesired side effects. Therefore, specific inhibition of PLC subtypes and isoforms has been proposed to allow for safer therapies for pathological conditions.

[0009] PLC-beta3 has been suggested as a potential target for treating skin diseases and conditions, as it is widely expressed in peripheral sensory neurons, and has been described as a key isoform involved in pain and itch signaling (Shi T-JS, et al., "Phospholipase Cp3 in mouse and human dorsal root ganglia and spinal cord is a possible target for treatment of neuropathic pain" PNAS, 2008, vol. 105(50), pp. 20004-20008). PLC-beta3 has also been disclosed to play a main role in pruritogenic-elicited itch through histaminergic and non- histaminergic fibers, due to several endogenous and exogenous stimuli (Luo J, et al., "Molecular and cellular mechanisms that initiate pain and itch", Cellular and molecular life sciences, 2015, vol. 72(17), pp. 3201 - 3223).

[0010] An oncogenic role of PLC-beta4 has been previously described, and it has been postulated as an interesting target for antitumoral therapies, in particular for therapies against uveal melanoma (Phan HTN et al., " Uveal melanoma-associated mutations in PLC|34 are constitutively activating and promote melanocyte proliferation and tumorigenesis", Sci Signal, 2021, Dec 14; 14(713)). PLC-beta4 has also been disclosed to be involved in inflammatory pain signaling (Miyata M. et al., " Role of Thalamic Phospholipase Cp4 Mediated by Metabotropic Glutamate Receptor Type 1 in Inflammatory Pain" J Neurosci, 2003 Sep 3; vol. 23(22); pp. 8098-8108).

[0011] U73122 is probably the most widely used agent to inhibit PLC-beta activity. However, several recent reports have shown that U73122 may not be selective for PLC-beta, in fact, effects on numerous cellular proteins have been reported, including on telomerase, 5-lipoxygenase, histamine H1 receptor, and calcium channels, which may cause undesired side-effects. Similarly, small peptides previously described to inhibit PLC enzymes also suffer from indirect effects, as well as from limited efficacy or bioavailability.

[0012] Therefore, there is still a need for improved PLC inhibitors, in particular PLC-beta inhibitors, that are efficient, specific, and safe for therapeutic and cosmetic purposes, in particular for treating diseases or disorders associated with PLC-beta activation.

[0013] Summary of Invention

[0014] The present inventors have surprisingly found that very small synthetic peptides derived from the C-terminal end sequence of the X-Y loop of the PLC-beta enzymes can be used for the specific and efficient inhibition of PLC-beta enzyme activity.

[0015] The X-Y loop of all PLC-beta proteins has an unconserved N-terminal region and two conserved elements —a highly acidic stretch and a conserved C-terminal region that forms the lid helix. The X-Y loop has been described to have autoin hibitory activity on the PLC-beta proteins. However, the sequence of whole X-Y loop was thought to be required for a proper inhibition of the catalytic center. More particularly, it was described in the prior art that the lid helix is insufficient to inhibit PLC-beta activity on its own as it also requires the presence of the adjacent acidic stretch, which has probably hindered the development of small inhibitory peptides based on this sequence (Lyon AM et al., "Molecular mechanisms of Phospholipase C |33 autoinhibition”, Structure. 2014 Dec 2;22(12): 1844-1854).

[0016] Notwithstanding the above, the present inventors have unexpectedly found that small peptides containing as little as 6 amino acids derived exclusively from the C-terminal region of the X-Y loop — i.e., without any amino acid from the acidic stretch— are by themselves capable of strongly inhibiting PLC-beta activity when administered exogenously into cells. Advantageously, the lack of any acidic stretch in their sequence provides the peptides of the invention with a very high bioavailability and makes them particularly well suited for both in vitro and in vivo administration. Moreover, from the extensive data provided below in several cellular models, it is apparent that the peptides of the invention can be used in therapeutic and cosmetic indications that benefit from PLC-beta activity inhibition.

[0017] Thus, in a first aspect, the invention provides a peptide or a pharmaceutically, veterinary, or cosmetically acceptable salt thereof, wherein the peptide comprises at least 5 consecutive amino acid residues of a sequence at least 80% identical to the sequence TTDEGTASSEVNATEEMSTLVN (SEQ ID NO: 1) or the sequence EQAWMASYKYVGATTNIHPYLS (SEQ ID NO: 2); the peptide has a length equal to or lower than 22 amino acid residues, particularly wherein the peptide is capable of inhibiting PLC-beta activity.

[0018] In a second aspect, the invention provides a pharmaceutical, veterinary, or cosmetic composition comprising: (i) a therapeutically or cosmetically effective amount of a peptide or a pharmaceutically, veterinary, or cosmetically acceptable salt thereof as defined in the first aspect, and (ii) at least one pharmaceutically, veterinary, or cosmetically acceptable excipient, diluent, or carrier.

[0019] In a third aspect, the invention provides a peptide or a pharmaceutically, veterinary, or cosmetically acceptable salt thereof as defined in the first aspect, or, alternatively, the pharmaceutical or veterinary composition as defined in the second aspect, for use as a medicament.

[0020] In a fourth aspect, the invention provides a peptide or a pharmaceutically, veterinary, or cosmetically acceptable salt thereof as defined in the first aspect, or, alternatively, the pharmaceutical or veterinary composition as defined in the second aspect, for use in the treatment or prevention of a disease or disorder mediated by PLC activation; particularly by PLC-beta activation.

[0021] In a fifth aspect, the invention provides the use of (i) a peptide or a pharmaceutically, veterinary, or cosmetically acceptable salt thereof as defined in the first aspect, or, alternatively, (ii) the cosmetic composition as defined in the second aspect, for the cosmetic, non-therapeutic treatment or care of the skin, hair, nails or mucous membranes.

[0022] In a sixth aspect, the invention provides a method of cosmetic, non-therapeutic treatment or care of the skin, hair, nails or mucous membranes of a subject comprising administering a cosmetically effective amount of (i) a peptide or a pharmaceutically, veterinary, or cosmetically acceptable salt thereof as defined in the first aspect, or (ii) a cosmetic composition as defined in the second aspect, to the subject.

[0023] In a seventh aspect, the invention provides a polynucleotide that encodes the peptide as defined in the first aspect.

[0024] In an eighth aspect, the invention provides an expression vector comprising the polynucleotide as defined in the seventh aspect.

[0025] In a nineth aspect, the invention provides a host cell comprising the peptide as defined in the first aspect, the polynucleotide as defined in the seventh aspect, or the expression vector as defined in the eighth aspect.

[0026] In a tenth aspect, the invention provides a method for producing the peptide as defined in the first aspect, the method comprising culturing the host cell as defined in the nineth aspect.

[0027] In an eleventh aspect, the invention provides a kit of parts comprising the peptide as defined in the first aspect, the pharmaceutical, veterinary, or cosmetic composition as defined in the second aspect, the polynucleotide as defined in the seventh aspect, or the expression vector as defined in the eighth aspect, and instructions for its use.

[0028] In a twelfth aspect, the invention provides an in vitro use of a peptide or a pharmaceutically, veterinary, or cosmetically acceptable salt thereof as defined in the first aspect, or, alternatively, the pharmaceutical or veterinary composition as defined in the second aspect, for the inhibition of PLC activity, particularly PLC-beta activity.

[0029] In a thirteenth aspect, the invention provides an in vitro method for inhibiting PLC activity, particularly PLC- beta activity, in a sample, the method comprising contacting the sample with a peptide or a pharmaceutically, veterinary, or cosmetically acceptable salt thereof as defined in the first aspect, or, alternatively, the pharmaceutical or veterinary composition as defined in the second aspect.

[0030] Detailed description of the invention

[0031] All terms as used herein in this application, unless otherwise stated, shall be understood in their ordinary meaning as known in the art. Other more specific definitions for certain terms as used in the present application are as set forth below and are intended to apply uniformly throughout the specification and claims unless an otherwise expressly set out definition provides a broader definition.

[0032] As used herein, the indefinite articles "a” and "an” are synonymous with "at least one” or "one or more”. Unless indicated otherwise, definite articles used herein, such as "the” also include the plural of the noun.

[0033] As above explained, the invention provides in a first aspect a peptide or a pharmaceutically, veterinary, or cosmetically acceptable salt thereof, that comprises at least 5 consecutive amino acid residues of a sequence at least 80% identical to SEQ ID NO: 1 or SEQ ID NO: 2. SEQ ID NO: 1 corresponds to a fragment of the C- terminal end of the X-Y loop of the human protein PLC-beta-3, and SEQ ID NO: 2 corresponds to the equivalent fragment of the human protein PLC-beta-4, As used herein, the terms "peptide", "polypeptide,” and "protein" are used interchangeably, and refer to a compound comprised of two or more amino acid residues covalently linked by peptide bonds. In the present invention, the term "amino acid” refers to a molecule containing both an amino group and a carboxyl group. Suitable amino acids include, without limitation, alpha amino acids, such as the L-isomers of alpha-amino acids of the 20 common naturally occurring alpha-amino acids: alanine, arginine, asparagine, aspartic acid, cysteine, glutamine, glutamic acid, glycine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine, and valine; natural beta-amino acids (e.g., beta-alanine); and unnatural amino acids. Each one of the amino acids forming the peptide of the invention can have, independently from the others, L- or D-configuration. The peptides of the present invention may exist as stereoisomers or mixtures of stereoisomers; for example, the amino acids that make them up can have L- configuration, D-configuration or be racemic independently from each other. Therefore, it is possible to obtain isomeric mixtures as well as racemates or diastereomeric mixtures or pure diastereomers or enantiomers, depending on the number of asymmetric carbons and what isomers or isomeric mixtures are present. In one embodiment, the structures of the peptides of the present invention are pure isomers, i.e., enantiomers or diastereomers. In a more particular embodiment, the structures of the peptides of the present invention consist of amino acids having the L- configuration (L-amino acids).

[0034] Amino acids used in the preparation of the peptides of the present invention may be prepared by organic synthesis, or obtained by other routes, such as, for example, degradation or isolation from a natural source.

[0035] As used herein, the term "pharmaceutical, veterinary, or cosmetical acceptable salt” refers to those salts which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of humans and lower animals without undue toxicity, irritation, allergic response and the like, and are commensurate with a reasonable benefit / risk ratio. Pharmaceutical, veterinary, and cosmetical acceptable salts are well known in the art. Examples of salts include salts of an amino group formed with inorganic acids such as hydrochloric acid, hydrobromic acid, phosphoric acid, sulfuric acid and perchloric acid or with organic acids such as acetic acid, trifluoroacetic acid, oxalic acid, maleic acid, tartaric acid, citric acid, succinic acid or malonic acid or by using other methods used in the art such as ion exchange. Other pharmaceutical, veterinary and cosmetical acceptable salts include adipate, alginate, ascorbate, aspartate, benzenesulfonate, benzoate, bisulfate, borate, butyrate, camphorate, camphorsulfonate, citrate, cyclopentanepropionate, digluconate, dodecylsulfate, ethanesulfonate, formate, fumarate, glucoheptonate, glycerophosphate, gluconate, hemisulfate, heptanoate, hexanoate, hydroiodide, 2-hydroxy-ethanesulfonate, lactobionate, lactate, laurate, lauryl sulfate, malate, maleate, malonate, methanesulfonate, 2-naphthalenesulfonate, nicotinate, nitrate, oleate, oxalate, palmitate, pamoate, pectinate, persulfate, 3-phenylpropionate, phosphate, picrate, pivalate, propionate, stearate, succinate, sulfate, tartrate, thiocyanate, p-toluenesulfonate, undecanoate, valerate salts, and the like. Salts derived from appropriate bases include alkali metal, alkaline earth metal, and ammonium. Representative alkali or alkaline earth metal salts include sodium, lithium, potassium, calcium, magnesium, and the like. Further pharmaceutical and cosmetical acceptable salts include, when appropriate, nontoxic ammonium, quaternary ammonium, and amine cations formed using counterions such as halide, hydroxide, carboxylate, sulfate, phosphate, nitrate, lower alkyl sulfonate and aryl sulfonate.

[0036] The preparation of pharmaceutically, veterinary, or cosmetically acceptable salts of the peptides of the invention can be carried out by methods known in the art. For instance, such salts may be prepared by reacting the peptides with a stoichiometric amount of the appropriate pharmaceutically, cosmetically or veterinary acceptable base or acid in water or in an organic solvent or in a mixture of them. The peptides and their salts may differ in some physical properties but they are equivalent for the purposes of the present invention.

[0037] In all embodiments of the invention referring to the peptides, their pharmaceutically, veterinary, or cosmetically acceptable salts are always contemplated even if they are not specifically mentioned.

[0038] In one embodiment of the first aspect, the peptide has a length equal to or lower than 22, 21, 20, 19, 18, 17, 16, 15, 14, 13, 12, 11 , 10, 9, 8, 7, 6, or 5 amino acid residues. In a more particular embodiment, the peptide has a length of 22, 21, 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, or 5 amino acid residues. In a more particular embodiment, the peptide has a length of 6 amino acid residues.

[0039] In one embodiment of the invention, the peptide is with the proviso that the peptide does not consist of the amino acid sequence IEVNATEEM (SEQ ID NO: 23).

[0040] In the present invention the term "identical" or "identity" refers to the percentage of positions that are identical in the two sequences when the sequences are optimally aligned. If, in the optimal alignment, a position in a first sequence is occupied by the same amino acid as the corresponding position in the second sequence, the sequences exhibit identity with respect to that position. The percentage of identity determines the number of identical amino acids over a defined length in a given alignment. Thus, the level of identity between two sequences or ("percent sequence identity") is measured as a ratio of the number of identical positions shared by the sequences with respect to the number of positions compared (i.e., percent sequence identity = (number of identical positions / total number of positions compared) x 100). A gap, i.e., a position in an alignment where an amino acid is present in one sequence but not in the other, is regarded as a position with non-identical amino acid and is counted as a compared position.

[0041] A number of mathematical algorithms for rapidly obtaining the optimal alignment and calculating identity between two or more sequences are known and incorporated into a number of available software programs. For purposes of the present invention, the sequence identity between amino acid sequences is preferably determined using algorithms based on global alignment, such as the Needleman-Wunsch algorithm (Needleman and Wunsch, 1970, J. Mol. Biol. 48: 443-453), preferably implemented in the Needle program of the EMBOSS package (EMBOSS: The European Molecular Biology Open Software Suite, Rice et al., 2000, Trends Genet. 16: 276-277); or the BLAST Global Alignment tool (Altschul et al., "Basic local alignment search tool”, 1990, J. Mol. Biol, v. 215, pages 403-410), using default settings. Local alignment also can be used when the sequences being compared are substantially the same length.

[0042] In a particular embodiment, the peptide comprises or consists of at least 5 consecutive amino acid residues of a sequence at least 80 %, at least 81 %, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100 % identical to SEQ ID NO: 1 or SEQ ID NO: 2. In another embodiment, the peptide comprises or consists of at least 6 consecutive amino acid residues of a sequence at least 80 %, at least 81 %, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100 % identical to SEQ ID NO: 1 or SEQ ID NO: 2. In another embodiment, the peptide comprises or consists of 6 consecutive amino acid residues of a sequence at least 80 %, at least 81 %, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100 % identical to SEQ ID NO: 1 or SEQ ID NO: 2.

[0043] In one embodiment, the peptide comprises or consists of at least 5 or at least 6 consecutive amino acid residues of a sequence at least 75 %, at least 76 %, at least 77 %, at least 78 %, at least 79 %, at least 80 %, at least 81 %, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100 % identical to SEQ ID NO: 4 or SEQ ID NO: 16. In one embodiment, the peptide comprises or consists of 6 consecutive amino acid residues of a sequence at least 75 %, at least 76 %, at least 77 %, at least 78 %, at least 79 %, at least 80 %, at least 81 %, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100 % identical to SEQ ID NO: 4 or SEQ ID NO: 16. In a particular embodiment, the peptide consists of 6 consecutive amino acid residues of a sequence at least 75 % identical to SEQ ID NO: 4 or SEQ ID NO: 16.

[0044] In one embodiment, the peptide comprises or consists of a sequence selected from SEQ ID NO: 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 , 12, 13, 14, 15, 16, 17, 18, 19, 20, 21 , 22, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49, 50 and 51 (i.e., SEQ ID NO: 1-22 and 24-51). In a particular embodiment, the peptide comprises or consists of a sequence selected from SEQ ID NO: 5-14, 1 -22, and 24-51 . In a more particular embodiment, the peptide comprises or consists of SEQ ID NO: 28. In a more particular embodiment, the peptide comprises or consists of SEQ ID NO: 37. In another particular embodiment, the peptide comprises or consists of SEQ ID NO: 51.

[0045] In some embodiments, the peptide comprises or consists of a sequence selected from SEQ ID NO: 1-22 and 24-51 , or a functional variant that comprises up to 1, up to 2, up to 3, up to 4, or up to 5 amino acid changes. The term "functional variant" refers to variants that maintain the biological activity of the peptide, more particularly that maintain the capacity to inhibit PLC-beta activity (e.g., PLC-beta-3 and / or PLC-beta-4 activity).

[0046] In one embodiment, the peptide comprises at least 5 consecutive amino acid residues of the sequence SEQ ID NO: 1 or the sequence SEQ ID NO: 2, or a variant of SEQ ID NO: 1 or SEQ ID NO: 2 that comprises up to 1 , up to 2, up to 3, up to 4, or up to 5 amino acid changes, the peptide has a length equal to or lower than 22 amino acid residues; and the peptide is capable of inhibiting PLC-beta activity; particularly PLC-beta-3 activity and / or PLC-beta-4 activity.

[0047] In one embodiment, the peptide comprises at least 6 consecutive amino acid residues of the sequence SEQ ID NO: 4 or the sequence SEQ ID NO: 16, or a variant of SEQ ID NO: 4 or SEQ ID NO: 16 that comprises up to 1 , up to 2, up to 3, or up to 4 amino acid changes, the peptide has a length equal to or lower than 22 amino acid residues; and the peptide is capable of inhibiting PLC-beta activity; particularly PLC-beta-3 activity and / or PLC-beta-4 activity.

[0048] In one embodiment, the peptide is capable of inhibiting PLC-beta activity. In a more particular embodiment, the peptide is capable of inhibiting PLC-beta-3 and / or PLC-beta-4 activity. In a more particular embodiment, the peptide is capable of specifically inhibiting PLC-beta-3 or PLC-beta-4 activity. The skilled person can readily test the PLC-beta-inhibiting activity of a peptide following routine methods known in the art, for example those disclosed in the examples below, without the need of inventive skill. For example, a way to test if a peptide is capable of inhibiting PLC-beta activity is by an acetylcholine response test shown in example 4 below. Briefly, HEK-293 cells are with Fluo-4 AM for 1 h, then contacted with the peptide at 100 uM for 1 h, and then treated with 1 uM acetylcholine twice for 30 s. Finally intracellular calcium levels revealed by Fluo-4 are used as indicator of acetylcholine response. If the peptide inhibits the acetylcholine response to levels similar to a positive control (e.g. U73122), the peptide has PLC-beta-3 or PLC-beta-4 inhibiting activity.

[0049] In one embodiment, the peptide further comprises an N-terminal and / or C-terminal modification. It is well- known in the state of the art how to modify (i.e., derivatize) the terminal ends of a peptide, for example, to improve its stability or efficacy. It should be appreciated that any of the peptide sequences provided herein are provided with or without any N-terminal and / or C-terminal modification.

[0050] As used herein, "N-terminal modification" refers to the modification of the NH2 of the N-terminal residue of the peptide, resulting in a compound being linked to the NH moiety. Examples of N-terminal modifications include, but are not limited to acetylation. As used herein, "C-terminal modification" refers to the modification of the carboxyl moiety of the C-terminal residue of the peptide. Examples of C-terminal modifications include, but are not limited to amidation, esterification and reduction. Thus, in a particular embodiment, the N-terminal and / or C-terminal modification comprises or consists of a compound selected from the group consisting of phosphoryl group, glycosyl group, acyl group, alkyl group, carboxyl group, hydroxyl group, biotinyl group, ubiquitinyl group, and amido group; particularly, wherein the acyl group is selected from the group consisting of acetyl group, lauroyl group, myristoyl group, and palmitoyl group.

[0051] In another embodiment, the N-terminal modification comprises or consists of a compound selected from the group consisting of acetyl group, urea group, formyl group, carbamate group, pyroglutamyl group, and sulfonamide group, and / or the C-terminal modification comprises or consists of a compound selected from the group consisting of amido group, N-alkyl amido group, or aldehyde group.

[0052] In a more particular embodiment, the N-terminal modification comprises or consists of an acyl group, optionally selected from the group consisting of acetyl group, myristoyl group, and palmitoyl group, and / or the C-terminal modification comprises or consists of an amido group. In a more particular embodiment, the N- terminal modification comprises or consists of an acetyl group and / or the C-terminal modification comprises or consists of an amido group. That is, the N-terminal end of the peptide is acetylated and the C-terminal end is amidated. N-terminal acetylation, in which the charge of the N-terminal is removed to mimic the state of the native protein, results in a peptide that is more stable and its resistance to enzymatic degradation of exopeptidases is highly enhanced. C-terminal amidation removes the negative charge of the carboxylic acid and, when this charge is neutralized, the hydrophobicity of the molecule is increased. Peptides with C-terminal amidation have enhanced stability and ability to disrupt cell membranes.

[0053] In one embodiment, the N-terminal group of the peptide is a monoradical of formula — N HRi ; the C-terminal group of the peptide is a monoradical of formula -C(0)-R2; Ri is a monoradical selected from -H and -C(O)- (Ci-C2o)alkyl; R2 is a monoradical selected from -OH and -NR3R4 radical; and R3 and R4 are independently selected from -H and -(C1-C 10) alkyl. In a more particular embodiment, the N-terminal group of the peptide is a monoradical of formula -NH-C(O)-CH3 and / or the C-terminal group of the peptide is a monoradical of formula -C(O)-NH2.

[0054] In one embodiment, the peptide is a linear peptide or a circularized peptide (i.e., cyclic peptide). There are several methods in the art to circularize peptides known to the skilled person.

[0055] In one embodiment, the peptide is conjugated to a cell penetrating peptide. In present invention the term "cell penetrating peptide” or "CPP” refers to short peptides that facilitate cellular uptake of various molecular cargo, including other peptides. The "cargo" is associated to peptides via the C(t) or N(t)-end, either through chemical linkage via covalent bonds or through non-covalent interactions. The function of the CPPs are to deliver the cargo into cells, a process that commonly occurs through endocytosis. The conjugation of the CPP to the peptide provided in the present invention can be performed following well-known routine protocols, such as solid phase synthesis or solution selective capping. (Copolovici D. M. et al., "Cell-Penetrating Peptides: Design, Synthesis, and Applications”, 2014, ACS Nano, 2014, 8 (3), pp 1972-1994). In another embodiment of the first aspect of the invention, the cell penetrating peptide is a polycationic CPP, polyArg or penetratine.

[0056] The peptides of the invention may also comprise localization sequences, such as cytoplasmic localization sequences, export sequences, such as nuclear export sequences, or other localization sequences, as well as sequence tags that are useful for solubilization, purification, or detection of the peptides. Suitable protein tags provided herein include, but are not limited to, biotin carboxylase carrier protein (BCCP) tags, myc-tags, calmodulin-tags, FLAG-tags, hemagglutinin (HA)-tags, polyhistidine tags, also referred to as histidine tags or His-tags, maltose binding protein (MBP)-tags, nus-tags, glutathione- S- transferase (GST)-tags, green fluorescent protein (GFP)-tags, thioredoxin-tags, S-tags, Softags ( e.g ., Softag 1, Softag 3), strep-tags , biotin ligase tags, FIAsH tags, V5 tags, and SBP-tags. Additional suitable sequences will be apparent to those of skill in the art.

[0057] In a particular embodiment of the first aspect, the peptide is an isolated, synthetic, or recombinant peptide. "Isolated” means altered or removed from its natural state.

[0058] Synthesis of the peptides of invention and their therapeutically or cosmetically acceptable salts can be carried out according to the conventional methods, known in the prior art, such as solid phase peptide synthesis methods, enzymatic synthesis or any combination (Palomo JM, "Solid-phase peptide synthesis: an overview focused on the preparation of biologically relevant peptides" RSC Advances, 2014, issue 62; Coin I. et al., " Solid-phase peptide synthesis: from standard procedures to the synthesis of difficult sequences", Nature Protocols, 2007, vol. 2, pp, 3247-3256). The peptides of the invention can also be obtained by means of recombinant DNA technology, which belongs to the common general knowledge of the skilled person. Peptides can be synthetized with an amidated C-terminus and / or modified in their N-terminus (e.g. acetylation, palmitoylation or myristoylation) as described, for example, in WO2013064583 or WO2019238686.

[0059] As above indicated, the present invention also relates to a pharmaceutical, veterinary or cosmetic composition comprising an effective amount of a peptide of the invention, together with pharmaceutically, veterinary, or cosmetically acceptable excipients, diluents, or carriers.

[0060] The expression "therapeutically effective amount" as used herein, refers to the amount of peptide that, when administered is sufficient to prevent development of, or alleviate to some extent, one or more of the symptoms of the disease which is addressed. The particular dose of the peptide administered according to this invention will of course be determined by the particular circumstances surrounding the case, including the peptide administered, the route of administration, the particular condition being treated, and the similar considerations.

[0061] The expression "cosmetically effective amount", in the context of the present invention, refers to the amount of a peptide that when it is applied (for example topically) is sufficient for cosmetically treating the skin, hair, nails or mucous membranes. The particular dose of the peptide administered according to this invention for cosmetic purposes will of course be determined by the particular circumstances surrounding the case. In one embodiment of the invention, the "cosmetically effective amount” of the peptide of the invention is higher, equal or lower than the amount needed to achieve a therapeutic effect.

[0062] The expression "pharmaceutically or veterinary acceptable excipient, diluent or carrier" means that the excipients, diluents, or carriers are suitable for the preparation of compositions for pharmaceutical or medical uses in humans and animals. Each component must be pharmaceutically acceptable in the sense of being compatible with the other ingredients of the pharmaceutical or veterinary composition. It must also be suitable for use in contact with tissues or organs of humans and animals without excessive toxicity, irritation, allergic response, immunogenicity or other problems or complications consistent with a reasonable risk / benefit relationship.

[0063] The expression "cosmetically acceptable excipient, diluent, or carrier" means that the excipients, diluents, or carriers are suitable for the preparation of compositions for cosmetic use. Each component must be cosmetically acceptable in the sense of being compatible with the other ingredients of the cosmetic composition. It must also be suitable for use in contact with tissues or organs of humans and animals, in particular the skin, without excessive toxicity, irritation, allergic response, immunogenicity or other problems or complications consistent with a reasonable risk / benefit relationship.

[0064] Examples of suitable acceptable excipients, diluents or carriers are solvents, dispersion media, liquid vehicles, dispersion or suspension aids, surface active agents, isotonic agents, thickening or emulsifying agents, preservatives, solid binders, lubricants and the like. Except insofar as any conventional excipient medium is incompatible with a substance or its derivatives, such as by producing any undesirable biological effect or otherwise interacting in a deleterious manner with any other component(s) of the pharmaceutical, veterinary or cosmetical composition, its use is contemplated to be within the scope of this invention.

[0065] The pharmaceutical, veterinary, or cosmetic composition may be formulated for buccal, oral, topical, or transdermal administration, including adhesive patches, non-adhesive patches, occlusive patches, micropatches. Systemic administration such as oral or parenteral administration are also contemplated. For the purposes of the invention, the term "parenteral" includes nasal, atrial, ophthalmic, rectal, urethral, vaginal, subcutaneous, intradermal, intravascular, intravenous, intramuscular, intraocular, intravitreal, intracorneal, intraspinal, intramedullary, Intracranial, intracervical, intracerebral, intrameningeal, intraarticular, intrahepatic, intrathoracic, intratracheal, intrathecal and intraperitoneal, as well as any other similar injection or infusion technique.

[0066] The election of the type of formulation will depend upon the nature of the active compound, the site of administration, the kinetics and duration of release of the compound of the invention, and the condition to be treated. In a particular embodiment, the pharmaceutical, veterinary, or cosmetic composition of the invention is a topical composition. In another particular embodiment, the composition of the invention is a transdermal composition. In another particular embodiment, the composition of the invention is a buccal or oral composition.

[0067] Bucal and oral formulations include solid and liquid formulations such as tablets, capsules, pills, solutions, emulsions, syrups, elixirs. The skilled in the art will know the appropriate excipients to be used in these formulations. Non-limiting examples of excipients include disintegrants such as microcrystalline cellulose, corn starch, sodium starch, glycolate, and alginic acid; binders such as starch, pregelatinized starch, polyvinyl pyrrolidone (PVP), copovidone, gum acacia, xanthan gum, gum tragacanth, cellulose derivatives, such as hydroxypropylmethyl cellulose (HPMC), hydroxypropyl cellulose (HPC), ethylcellulose (EC) and carboxymethyl cellulose (CMC); lubricants such as magnesium stearate or sodium laurilsulfate; glidants such as colloidal silicon dioxide, starch and talc, sweeteners such as sucrose, glucose, fructose, maltose, lactose, sorbitol, xylitol, mannitol, maltulose, isomaltulose, maltitol, isomaltitol, lactulose, and lactitol; coloring and flavoring agents.

[0068] Generally, topical or transdermal formulations include creams, multiple emulsions such as, for example, emulsions of oil and / or silicone in water, emulsions of water / oil / water or water / silicone / water and emulsions and oil / water / oil or silicone / water / silicone, anhydrous compositions, aqueous dispersions, oils, milks, balms, foams, gels, pomades, powders, lotions, cream gels, hydroalcoholic solutions, hydro glycol solutions, hydrogels, liniments, soaps, shampoos, conditioners, serums, polysaccharide films, ointments, mousses, ointments powders, bars, dry, pastes, pencils and vaporizers, sprays, including "leave on" formulations and "rinse-off" formulations, wherein the compound is dispersed or dissolved in suitable excipients.

[0069] Topical or transdermal application can be incorporated by techniques known to those skilled in the art to various types of solid accessories such as, for example, and without limitation, sense bandages, gauzes, t- shirts, socks, stockings, underwear, girdles, gloves, diapers, napkins, dressings, bedspreads, wipes, adhesive patches, non-adhesive patches, occlusive patches, microelectronic patches or face masks, or may be incorporated into various make-up line products such as makeup bottoms, such as fluid makeup bottoms and make-up lotions, cleansing lotions, make-up removers, concealers, eye shadows, lipsticks, lip protectors, lip gloss and powders, among others.

[0070] The topical compositions defined above comprise pharmaceutical, veterinary or cosmetic excipients or carriers appropriate for topical administration, including, binders, emollients, skin permeation enhancers, emulsifiers, surfactants, thickening agents, viscosity increasing agents, pH regulators, antioxidants, preservative agents, solvents, dyestuffs, pigments, perfumes or mixtures thereof. The excipients or carriers used have affinity for the skin, are well tolerated, are stable, and are used in an amount suitable to provide the desired consistency and ease of application. The appropriate excipients and / or carriers, and their amounts, can readily be determined by those skilled in the art according to the type of formulation being prepared. The carriers of the compositions of the invention may be liquid carriers such as water, oils, including those of petroleum, animal, vegetable or synthetic origin, such as for example peanut oil, soybean oil, mineral oil, sesame oil, castor oil, polysorbates, sorbitan esters, ether sulfates, sulfates, betaines, glycosides, maltosides, fatty alcohols, nonoxynols, poloxamers, polyoxyethylenes, polyethylene glycols, dextrose, glycerol, digitonine.

[0071] Examples of binders include, without limitation, polyvinyl pyrrolidone, alginates, traganth. Examples of emollients or skin permeation enhancers include, without limitation, lauryl alcohol, oleyl alcohol, eucalyptol, sodium lauryl sulfate, glyceryl monooleate, sorbitan monooleate, isopropyl myristate, glyceryl myristate, propylene glycol, dimethyl isosorbide, Isopropyl Palmitate, oleic acid, dimethylsulfoxide, dimethylacetamide, dimethylformamide, surfactants, azone (1-dodecylazacycloheptan-2-one), alcohol, urea, ethoxydiglycol, acetone, polyethylene glycol among others. If desired, the compositions of the invention may be applied in the local areas to be treated by iontophoresis, sonophoresis, electroporation, microelectrical patches, mechanical pressure, osmotic pressure gradient, occlusive cure, microinjections or injections without needles by pressure, as for example oxygen-injections, or any combination thereof, to achieve enhanced penetration of the compound of the invention. The area of application will be determined by the nature of the condition to be treated or prevented.

[0072] Examples of emulsifiers or surfactants include, without limitation, polysorbate 80, polysorbate 20, sorbitan groups (spans), lecithin and Potassium Hexadecyl Hydrogen Phosphate.

[0073] Examples of thickening or viscosity increasing agents include, without limitation, synthetic polymers such as carbomers (cross linked polymers of acrylic acid), cellulosic polymers such as hydroxypropyl methylcellulose, methylcellulose, sodium carboxy methylcellulose, and hydroxypropyl cellulose and block copolymers based on ethylene oxide and propylene oxide (pluronic compounds).

[0074] Examples of pH regulators or buffering agents include, without limitation, monobasic sodium phosphate, dibasic sodium phosphate, sodium benzoate, potassium benzoate, sodium citrate, sodium acetate, sodium tartrate, diethanolamine, triethanolamine, sodium hydroxide, citric acid.

[0075] Examples of antioxidant agents include, without limitation, ascorbic acid, sodium ascorbate, sodium bisulfite, sodium sulfite, sodium metabisulfate, curcumin, tetrahydrocurcumin, diacetyl tetrahydrocurcumin, resveratrol, quercetin, hesperidin, myricetin, naringin, alpha-lipoic acid, monothioglycerol, butylated hydroxy anisole (BHA), butylated hydroxy toluene (BHT), propyl gallate.

[0076] Examples of preservatives include, without limitation, methylparaben, methylparaben sodium, propylparaben, propylparaben sodium, benzalkonium chloride, diazolidinyl urea, benzethonium chloride, chlorocresol, thiomarsal, sorbic acid, potassium sorbate and benzyl alcohol. Examples of solvents include, without limitation, ethanol, isopropyl alcohol, water, propylene glycol, polyethylene glycol, substituted glycols such as cremophor, or mixtures thereof.

[0077] The peptides of the present invention may also be adsorbed on solid organic polymers or solid mineral supports such as, for example, and without limiting sense tale, bentonite, silica, starch or maltodextrin among others.

[0078] The pharmaceutical, veterinary or cosmetic composition of the invention may be immediate or sustained release systems. The term "sustained release" is used herein to refer to a system for the delivery of a compound which provides for the gradual release of said compound over a period of time and preferably, but not necessarily, with relatively constant release levels of the compound at over a period of time. Examples of sustained release or carrier systems include, without limitation, liposomes, mixed liposomes, oleosomes, niosomes, etosomes, milliparticles, microparticles, nanoparticles and solid lipid nanoparticles, nanostructured lipid carriers, sponges, cyclodextrins, vesicles, micelles, mixed micelles of surfactants, Phospholipidsurfactant mixed micelles, millispheres, microspheres and nanospheres, lipospheres, milicapsules, microcapsules and nanocapsules, as well as in microemulsions and nanoemulsions, which may be added to achieve greater penetration of the active ingredient and / or improve the pharmacokinetic and pharmacodynamic properties thereof. Particular examples of sustained release or carrier systems are liposomes, mixed micellar phospholipid and microemulslons, more particularly water-in-oil microemulsions with internal reverse micelle structure.

[0079] The formulations of the pharmaceutical, veterinary, or cosmetical compositions described herein may be prepared by any method known or hereafter developed in the art of pharmacology and cosmetics. In general, such preparatory methods include the step of bringing the active ingredient (the peptide) into association with an excipient and / or one or more other accessory ingredients, and then, if necessary and / or desirable, shaping and / or packaging the product into a desired single- or multi-dose unit.

[0080] A pharmaceutical, veterinary, or cosmetic composition of the invention may be prepared, packaged, and / or sold in bulk, as a single unit dose, and / or as a plurality of single unit doses. As used herein, a "unit dose” is discrete amount of the pharmaceutical, veterinary, or cosmetic composition comprising a predetermined amount of the active ingredient.

[0081] The pharmaceutical, veterinary, and cosmetic composition of the present invention may be administered either alone or in combination with one or more other therapeutic or cosmetic agents. By "in combination with,” it is not intended to imply that the agents must be administered at the same time and / or formulated for delivery together, although these methods of delivery are within the scope of the invention. The compositions can be administered concurrently with, prior to, or subsequent to, one or more other desired therapeutics or cosmetic procedures. In general, each agent will be administered at a dose and / or on a time schedule determined for that agent. Additionally, the invention encompasses the delivery of the peptide or pharmaceutical, veterinary, or cosmetic compositions in combination with agents that may improve their bioavailability, reduce and / or modify their metabolism, inhibit their excretion, and / or modify their distribution within the body.

[0082] As above indicated, in a third aspect the present invention provides the peptide or compositions of the invention for use as a medicament. And in a fourth aspect, the invention provides the peptide or compositions of the invention for use in the treatment or prevention of a disease or disorder mediated by PLC activation; particularly by PLC-beta activation.

[0083] A "disease” is a state of health of an animal wherein the animal cannot maintain homeostasis, and wherein if the disease is not ameliorated then the animal's health continues to deteriorate. In contrast, a "disorder” in an animal is a state of health in which the animal is able to maintain homeostasis, but in which the animal's state of health is less favorable than it would be in the absence of the disorder. Left untreated, a disorder does not necessarily cause a further decrease in the animal's state of health.

[0084] The term "treatment”, as used herein, unless otherwise indicated, refers to any type of therapy which is aimed at terminating, preventing, ameliorating or reducing the susceptibility to a clinical condition or existing disease as described herein, including complete curing of a disease as well as amelioration or alleviation of said disease. Thus, "treatment,” "treating,” and their equivalent terms refer to obtaining a desired pharmacologic or physiologic effect, covering any treatment of a pathological condition or disorder in a subject. The term "prevention” or "preventing", as used herein, means, but is not limited to, a process of prophylaxis in which a subject is exposed to the polypeptide of the invention prior to the induction or onset of the disease process.

[0085] In a particular embodiment of the fourth aspect, the disease or disorder mediated by PLC activation is selected from the group consisting of cancer, inflammation, skin disease, pain, diabetes, atopia, and cardiovascular disease. In a more particular embodiment, the cancer is melanoma, particularly uveal melanoma. In another particular embodiment, the pain is inflammatory pain.

[0086] In a particular embodiment of the fourth aspect, the peptide or the pharmaceutical or veterinary composition is for use in the treatment of inflammation, optionally wherein inflammation is selected from the group consisting of conjunctivitis, allergic conjunctivitis, itchy eyes, chronic urticaria, eczema, prurigo, itchy skin, and erythema exudative. The involvement of PLC-beta activity in inflammation has been disclosed, for example, in Han SK et al., "Phospholipase Cbeta 3 mediates the scratching response activated by the histamine H1 receptor on C-fiber nociceptive neurons", Neuron., 2006, vol. 52(4), pp. 691-703; Xiao W et al., "Phospholipase C-|33 regulates FcsRI-mediated mast cell activation by recruiting the protein phosphatase SHP-1", Immunity, 2011, vol. 34(6), pp. 893-904; Descorbeth M et al., "Role of oxidative stress in high-glucose- and diabetes-induced increased expression of Gq / 11a proteins and associated signaling in vascular smooth muscle cells", Free Radio Biol Med, 2010, vol. 49(9), pp. 1395-405. In a particular embodiment of the fourth aspect, the peptide or the pharmaceutical or veterinary composition is for use in the treatment of skin disease, optionally wherein skin disease is selected from the group consisting of pruritus (itch), sensitive skin, atopic dermatitis, contact dermatitis, allergic dermatitis, psoriasis, rosacea, acne vulgaris, and psoriatic arthritis. The inhibition of PLC-beta activity has been associated to the relief of skin disease, for example, in atopic dermatitis (Ando T. et al., "Critical role for mast cell Stat5 activity in skin inflammation", Cell Rep., 2014, vol. 6(2), pp. 366-76).

[0087] In a particular embodiment of the fourth aspect, the peptide or the pharmaceutical or veterinary composition is for use in the treatment of pain, optionally wherein the pain is selected form the group consisting of postherpetic neuralgia, shingles (herpes zoster), diabetic neuropathy, postmastectomy pain syndrome, oral neuropathic pain, trigeminal neuralgia, temperomandibular joint disorders, cluster headache, osteoarthritis, arthritis pain, rhinopathy, oral mucositis, cutaneous allergy, detrusor hyperreflexia, loin pain / hematuria syndrome, neck pain, back pain, amputation stump pain, reflex sympathetic dystrophy and pain due to skin tumor. In another embodiment, the pain is neuropathic pain. The inhibition of PLC-beta activity for relieving pain has been disclosed, for example, in Shi TJ et al., "Phospholipase C{beta}3 in mouse and human dorsal root ganglia and spinal cord is a possible target for treatment of neuropathic pain", Proc Natl Acad Sci U S A, 2008, vol. 105(50), pp. 20004-8.

[0088] In a particular embodiment of the fourth aspect, the peptide or the pharmaceutical or veterinary composition is for use in the treatment of cardiovascular disease, wherein the cardiovascular disease is vascular hyperreactivity or atherosclerosis. The involvement of PLC-beta activity in cardiovascular disease has been disclosed, for example, in Tappia P S et al., "Upregulation of Phospholipase C Gene Expression Due to Norepinephrine-Induced Hypertrophic Response", Cells., 2022, vol. 11(16), pp. 2488.

[0089] In a further embodiment, the peptide or the pharmaceutical or veterinary composition of the invention is for use in the treatment of neurodegenerative disease, optionally wherein the neurodegenerative disease is selected from the group consisting of a Alzheimer's type senile dementia, a non-Alzheimer's type senile dementia, Parkinson's disease, facial nerve (Bell's) palsy, glaucoma, Huntington's chorea, amyotrophic lateral sclerosis (ALS), Alper's disease, Batten disease, Cockayne syndrome, Guillain-Barre syndrome, Lewy body disease, and Creutzfeldt-Jakob disease.

[0090] As explained above, the invention provides in a fifth aspect the use of the peptide or the composition of the invention for the cosmetic, non-therapeutic treatment or care of the skin, hair, nails or mucous membranes.

[0091] In a particular embodiment of the fifth or sixth aspect, the cosmetic, non-therapeutic treatment or care of the skin, hair, nails or mucous membranes comprises inhibiting melanogenesis. The involvement of PLC-beta activity in melanogenesis has been described, for example, in Bellono NW et al., " UV light activates a Gaq / 11 -coupled phototransduction pathway in human melanocytes", J Gen Physiol., 2014, vol. 143(2), pp. 203-14. In another particular embodiment of the fifth or sixth aspect, the cosmetic, non-therapeutic treatment or care of the skin, hair, nails or mucous membranes is selected from the group consisting of depigmentation or whitening or lightening in colour of the skin (e.g. depigmentation or whitening or lightening in colour of age spots; depigmentation or whitening or lightening in colour of the skin of dark eye circles); maintenance or improvement of skin luminosity, skin elasticity, skin resistance, skin firmness, skin tensile strength, skin smoothness, skin hydration, skin barrier function, skin microcirculation, or skin trone; treatment or prevention of the symptoms of skin aging (e.g. skin photoaging), skin fatigue, skin wrinkles (e.g. periorbital wrinkles), skin redness, skin itchiness, eye bags, skin sensory discomfort, unregulated pigmentation, excessive perspiration, unregulated sebum production, and unregulated hair growth.

[0092] It would be appreciated that all embodiments of the peptides of the first aspect are also meant to apply to the compositions of the second aspect, the peptides for use in the third and fourth aspect, the uses of the fifth and twelfth aspects, and the methods of the sixth and thirteenth aspects of the invention.

[0093] As above indicated, in a seventh aspect the invention provides a polynucleotide that encodes the peptide of the invention.

[0094] In one embodiment of the seventh aspect, the polynucleotide is DNA (single or double stranded) or RNA. In another embodiment, the polynucleotide is an isolated, synthetic, or recombinant polynucleotide. In a more particular embodiment, the polynucleotide comprises a sequence optimized for expression in one or more cell types. For example, the polynucleotide sequence may be optimized for expression in a mammalian cell (e.g., a HEK 293T cell). The polynucleotide sequence may be codon optimized for expressing in a mammalian cell using Integrated DNA Technologies (IDT), GeneArt, CoIler, and GenScript.

[0095] The invention also provides in an eighth aspect an expression vector comprising the polynucleotide of the seventh aspect. Examples of suitable expression vectors include those conventionally used in biomedicine and known to the skilled person, which includes vectors designed for expression in prokaryotic or eukaryotic cells. For example, vectors can be expressed in bacterial cells such as Escherichia coll, insect cells (using baculovirus expression vectors), yeast cells, or mammalian cells. Alternatively, expression vectors can be transcribed and translated in vitro, for example using T7 promoter regulatory sequences and T7 polymerase.

[0096] As above explained, in a twelfth and thirteenth embodiment, the invention provides the in vitro use of the peptides of the invention for inhibiting PLC activity.

[0097] In a particular embodiment of the twelfth and thirteenth aspects, the inhibition of PLC activity consists of the inhibition of PLC-beta3 and / or inhibition of PLC-beta4 activity.

[0098] The sequences provided in the present application are disclosed in Table 1 below: Table 1:

[0099] "Ac" stands for acetyl; "palm" stands for palmitoyl, "myr" stands for myristoyl, and "-NH2" refers to C-terminal amidation.

[0100] Throughout the description and claims the word "comprise" and variations of the word, are not intended to exclude other technical features, additives, components, or steps. Furthermore, the word "comprise” encompasses the case of "consisting of'. Additional objects, advantages and features of the invention will become apparent to those skilled in the art upon examination of the description or may be learned by practice of the invention. The following examples are provided by way of illustration, and they are not intended to be limiting of the present invention. Furthermore, the present invention covers all possible combinations of particular and preferred embodiments described herein.

[0101] For reasons of completeness, various aspects of the invention are set out in the following numbered clauses:

[0102] 1. A peptide or a pharmaceutically, veterinary, or cosmetically acceptable salt thereof, wherein: the peptide comprises at least 5 consecutive amino acid residues of a sequence at least 80% identical to the sequence TTDEGTASSEVNATEEMSTLVN (SEQ ID NO: 1) or the sequence EQAWMASYKYVGATTNIHPYLS (SEQ ID NO: 2); the peptide has a length equal to or lower than 22 amino acid residues; and with the proviso that the peptide does not consist of the amino acid sequence IEVNATEEM (SEQ ID NO: 23).

[0103] 2. The peptide according to clause 1, wherein the peptide is capable of inhibiting PLC-beta activity; particularly PLC-beta-3 activity and / or PLC-beta-4 activity. 3. The peptide according to any one of clauses 1-2, wherein the peptide comprises or consists of at least 6 consecutive amino acid residues of a sequence at least 75 % identical to the sequence TASSEVNATEEM (SEQ ID NO: 4) or the sequence ASYKYVGATTNIH (SEQ ID NO: 16).

[0104] 4. The peptide according to any one of clauses 1-3, wherein the peptide consists of 6 consecutive amino acid residues of a sequence at least 75 % identical to the sequence TASSEVNATEEM (SEQ ID NO: 4) or the sequence ASYKYVGATTNIH (SEQ ID NO: 16).

[0105] 5. The peptide according to any one of clauses 1-4, wherein the peptide consists of a sequence selected from SEQ ID NO: 5-14, 17-22, and 24-51.

[0106] 6. The peptide according to any one of clauses 1-5, which further comprises an N-terminal and / or C-terminal modification.

[0107] 7. The peptide according to clause 6, wherein the N-terminal and / or C-terminal modification comprises a compound selected from the group consisting of phosphoryl group, glycosyl group, acyl group, alkyl group, carboxyl group, hydroxyl group, biotinyl group, ubiquitinyl group, and amido group; particularly, wherein the acyl group is selected from the group consisting of acetyl group, lauroyl group, myristoyl group, and palmitoyl group.

[0108] 8. The peptide according to clause 6 or 7, wherein the N-terminal modification consists of an acetyl group and / or the C-terminal modification consists of an amido group.

[0109] 9. A pharmaceutical, veterinary, or cosmetic composition comprising:

[0110] (I) a therapeutically or cosmetically effective amount of a peptide or a pharmaceutically, veterinary, or cosmetically acceptable salt thereof, wherein the peptide comprises at least 5 consecutive amino acid residues of a sequence at least 80% identical to the sequence TTDEGTASSEVNATEEMSTLVN (SEQ ID NO: 1) or the sequence EQAWMASYKYVGATTNIHPYLS (SEQ ID NO: 2); the peptide has a length equal to or lower than 22 amino acid residues; optionally, wherein the peptide further comprises an N-terminal and / or C-terminal modification; and

[0111] (II) at least one pharmaceutically, veterinary, or cosmetically acceptable excipient, diluent, or carrier.

[0112] 10. A peptide or a pharmaceutically, veterinary, or cosmetically acceptable salt thereof, wherein the peptide comprises at least 5 consecutive amino acid residues of a sequence at least 80% identical to the sequence TTDEGTASSEVNATEEMSTLVN (SEQ ID NO: 1) or the sequence EQAWMASYKYVGATTNIHPYLS (SEQ ID NO: 2); the peptide has a length equal to or lower than 22 amino acid residues; optionally, wherein the peptide further comprises an N-terminal and / or C-terminal modification; or, alternatively, the pharmaceutical or veterinary composition as defined in clause 9, for use as a medicament.

[0113] 11 . A peptide or a pharmaceutically, veterinary, or cosmetically acceptable salt thereof, wherein the peptide comprises at least 5 consecutive amino acid residues of a sequence at least 80% identical to the sequence TTDEGTASSEVNATEEMSTLVN (SEQ ID NO: 1) or the sequence EQAWMASYKYVGATTNIHPYLS (SEQ ID NO: 2); the peptide has a length equal to or lower than 22 amino acid residues; optionally, wherein the peptide further comprises an N-terminal and / or C-terminal modification; or, alternatively, the pharmaceutical or veterinary composition as defined in clause 9, for use in the treatment or prevention of a disease or disorder mediated by PLC activation; particularly by PLC-beta activation.

[0114] 12. The peptide or the composition for use according to clause 11, wherein the disease or disorder mediated by PLC activation is selected from the group consisting of cancer, inflammation, skin disease, pain, diabetes, atopia, and cardiovascular disease.

[0115] 13. Use of:

[0116] (i) a peptide or a pharmaceutically, veterinary, or cosmetically acceptable salt thereof, wherein the peptide comprises at least 5 consecutive amino acid residues of a sequence at least 80% identical to the sequence TTDEGTASSEVNATEEMSTLVN (SEQ ID NO: 1) or the sequence EQAWMASYKYVGATTNIHPYLS (SEQ ID NO: 2); the peptide has a length equal to or lower than 22 amino acid residues; optionally, wherein the peptide further comprises an N-terminal and / or C-terminal modification; or, alternatively,

[0117] (ii) the cosmetic composition as defined in clause 9, for the cosmetic, non-therapeutic treatment or care of the skin, hair, nails, or mucous membranes.

[0118] 14. The use according to clause 13, wherein the cosmetic, non-therapeutic treatment or care is selected from the group consisting of depigmentation or whitening or lightening in colour of the skin; maintenance or improvement of skin luminosity, skin elasticity, skin resistance, skin firmness, skin tensile strength, skin smoothness, skin hydration, skin barrier function, skin microcirculation, or skin tone; treatment or prevention of the symptoms of skin aging, skin fatigue, skin wrinkles, skin redness, skin itchiness, eye bags, skin sensory discomfort, unregulated pigmentation, excessive perspiration, unregulated sebum production, and unregulated hair growth. 15. The pharmaceutical, veterinary, or cosmetic composition according to clause 9, the peptide or the composition for use according to any one of clauses 10-12, or the use according to any one of clauses 13-14, wherein the peptide is as defined in any one of clauses 1-8.

[0119] Examples

[0120] Example 1 - Peptide synthesis

[0121] All peptides used in the Examples were synthetized using the standard Fmoc solid-phase method as described in WO2013064583. The identity of the peptides obtained was confirmed by ESI-MS.

[0122] Example 2 - M3-acetylcholine activation

[0123] Recent studies have showed a direct interaction between M3 receptor and PLCp3, where the expression and activation of M3 in CHO cells drives PLCp3 localization at the plasma membrane and enhances the efficiency of GPCRs signaling (Kan W, et al., "M3 muscarinic receptor interaction with phospholipase C p3 determines its signaling efficiency", J Biol Chem., 2014, vol. 289(16), pp. 11206-18).

[0124] The muscarinic acetylcholine M3 receptor (M3) is expressed by several skin cells and appendix, such as melanocytes, keratinocytes, fibroblast or sweat glands, suggesting a role in cutaneous pigmentation, wound healing, or sweating. After ACh interaction with M3 receptor, phospholipase C (PLC) is activated increasing calcium release from the intracellular stores. Therefore, PLC inhibition is postulated as a promising target to modulate this cell signaling pathway.

[0125] Thus, the aim of this study was to evaluate the effect of various peptides of the invention in the activity of M3- acetylcholine activation. For this purpose, an epithelial stable cell line constitutively expressing human muscarinic receptor M3 (CHO-k1-hM3; Perkinelmer, ValiScreen Muscarinic M3) was used to assess peptides activity.

[0126] Cells were seeded in 96-well plates at 70% cell confluence in complete Ham's F-12 medium. After 24 hours, medium was removed and cells were incubated with a solution of the peptide at the indicated concentrations or DMSO or PBS-1% NH4OH as control (vehicles).

[0127] After 24 hours, the medium of CHO-k1-hM3 cells was replaced by Fluo 4-NW dye solution in assay buffer containing probenecid. Then, Peptides and vehicles were added. All treatments were incubated 60 minutes at 37 °C in CO2 incubator following manufacturer's instructions (Fluo-4 NW Calcium Assay kit, Molecular Probes).. Afterwards, M3 activity was determined by reading the fluorescence in a fluorescence plate reader (Fluostar Galaxy, BMG LabTechnologies) using 485 nm filter for excitation and 520 nm for emission, recording signal before and after M3 activation with acetylcholine in a range of 10-20 nM. Atropine at 100 nM was used as positive control for inhibition.

[0128] The human M3 receptor mediated response was quantified from raw data in Arbitrary Fluorescence Units (AFU) as the ratio between signals before and after M3 activation with acetylcholine (Fmax / Fmin). Inhibition values of M3 activity were calculated as percentages by considering maximum fluorescence corresponding to acetylcholine M3 activation (100%response) and minimum fluorescence to atropine treated cells (0% response), as follows:

[0129] % hM3R response=([Xi-mean (0%)] x 100) / (mean (100%)-mean (0%) where XI are the values of Fmax / Fmin. Each peptide was normalized with respect to its corresponding vehicle.

[0130] Table 2 shows the %inhibition values of M3 activity obtained for the indicated peptides of invention.

[0131] Table 2

[0132] As shown in the results above, all the peptides of the invention tested, which are derived from the C-terminal fragment of the X-Y loop of PLC-beta3 (SEQ ID NO: 24, 28 41, and 43) or the PLC-beta4 (SEQ ID NO: 44-46) and bearing different terminal modifications, provided an efficient inhibition of human M3 activity induced by acetylcholine. These results clearly demonstrate the utility of the peptides of the invention for treating diseases or disorder associated with PLC-beta activity.

[0133] Example 3 - Inhibition of UVA-induced melanogenesis

[0134] The aim of this study was to evaluate the effect of various peptide of the invention in UVA-induced melanogenesis in human primary melanocytes. For this purpose, HEMn-MP primary cells (moderately pigmented normal human melanocytes) (Cascade Biologies™, Thermo Fisher Scientific) isolated from human skin were cultured in supplemented medium (Medium 254, 1% HMGS and 1% penicillin-streptomycin). Above 80% confluence, cells were trypsinized with Trypsin-EDTA solution (3 min, room temperature) and neutralized with Trypsin Neutralizer Solution. Then cells were then seeded in 12-well-plate in supplemented medium. Melanin accumulation assays were performed 5 days after seeding (37°C, 5% CO2).

[0135] After 3 days cell seeding, medium was replaced by treatments in supplemented medium, the indicated peptides were incubated for 24h and vehicle. Two identical plates were prepared with all testing conditions. On the irradiation day, treatments were refreshed. One plate was left in the dark at room temperature while the other was irradiated with UVA at 5J / cm2for 30 min, except for one capped non-treated control for using it as basal condition. Then, treatments were replaced by freshly medium, and cells were incubated 24h (37°C, 5% CO2).

[0136] After 24 hours of recovery, cells were lysed with PBS-1% tritonXI 00 and lysates were centrifuged at 13.000xg for 15 min. Pellet was dissolved with 1 N NaOH to extract melanin for 1 .5 h at 90 °C in a water bath. A direct relationship between optic density (OD) at 405 nm and amount of melanin accumulated by the cells was established by using a synthetic melanin standard curve to interpolate sample values. Supernatants were used to determine total protein concentration through BOA Absorbance assay (Thermoscientific) following commercial instructions.

[0137] Raw data were processed to calculate the corresponding melanin accumulation as follow:

[0138] The melanin quantity of each sample was determined from linear fit in the standard curve (Mi). Then, melanin concentration was normalized by its respective total protein amount (MPi=Mi / Pi).

[0139] The percentage of melanin accumulation after treatments without UVA induction was determined: % Mel accum = 100

[0140] Where MPmin= (melanin / protein) mean of non-stimulated vehicle-treated cells or irradiated-capped vehicle- treated cells; MPi= (melanin / protein) of UVA-stimulated treated-cells.

[0141] Table 3

[0142] The percentage of UVA-induced melanin after treatments was determined:

[0143] % UVA-induced melanin

[0144] Where MPmin= (melanin / protein) mean of non-stimulated or irradiated-capped, vehicle-treated cells (0% UVA- induced melanin); MPmax= (melanin / protein) mean of UVA-stimulated, vehicle-treated cells (100% UVA- induced melanin); MPj= (melanin / protein) of stimulated treated-cells.

[0145] Table 4

[0146] As shown in Table 4 above, all the peptides of the invention tested provided a strong inhibition of melanogenesis in human primary melanocytes, in particular the Ac-SSEVNA-NH2 (SEQ ID NO. 28) that reduced the production of melanin in more than 90 %. These results clearly demonstrate the utility of the peptides of the invention for treating diseases or disorder associated with unregulated pigmentation, and also for the cosmetic treatment of the skin.

[0147] Example 4 - Acetylcholine response

[0148] The HEK293 immortalized cell line has been widely used for transfection experiments, although endogenously expresses some G protein coupled receptors such as the acetylcholine muscarinic M3 receptor. According to previous studies of protein expression, HEK293 cells express the isoform PLCp3, which is known to be associated with the M3 receptor as above described. In contrast, PLCp4 levels are low.

[0149] Thus, the aim of this study was to evaluate the effect of various peptides of the invention on the acetylcholine response. For this purpose, intracellular calcium fluxes triggered by acetylcholine were analyzed in HEK-293 cell line (ATCC) using the calcium imaging technique. HEK293 cells were maintained in supplemented medium (DMEM, high glucose, pyruvate, no glutamine containing 10% Heat Inactivated Foetal Bovine Serum, 1% Penicillin-Streptomycin and 1% L-glutamine). Cells were seeded on pre-treated Poly-L-Lysin 12 mm-glass coverslip and cultured at 37°C and 5% CO2.

[0150] Calcium imaging recordings were performed 48 h after seeding. Measurements were carried out in an inverted fluorescent microscope (ZEISS Axiovert 200b; Software HCImage v. DIA. HAMAMATSU) coupled to a Hamamatsu FLASH 4.0 LT camera (C11440-42U30, Hamamatsu, Sunayama-cho Japan).

[0151] Non-treated cells on coverslips were incubated with 6 pig / mL Fluo-4 AM prepared in prewarmed external solution with 0.05% pluronic acid for 1 h at 37°C (Fluo-4 NW Calcium Assay kit, Molecular Probes). In cells treated with the peptides at 100 piM or U-73122 at 2 pi M as the reference PLC inhibitor, the compounds were preincubated with the dye for 1 h at 37°C. Afterwards, cells were washed with external solution for at least 20 min at 37°C. Then, coverslips were mounted onto the microscope set up. Stimuli were applied in perfusion following the protocol described below. Two pulses of acetylcholine 1 piM were applied during 30 s separated by a 3 min wash with external solution to recover a stable basal line. Finally, ionomycin 10 piM was applied briefly to check the integrity of cells.

[0152] Changes in the intracellular calcium levels revealed by Fluo-4 dye (exciting the fluorophore at 483 nm during 200-300 ms and collecting the emission at 512 nm) were registered as indicator of acetylcholine response. Fluorescence images were taken every 3 s during stimuli applications and responses.

[0153] Fluorescence intensity of individual cells was adjusted to baseline zero subtracting the background signal. The response to each stimulus was analyzed quantifying the fluorescence peak and subtracting the fluorescence average 30 s before applying it. Only cells with responses equal or greater than 20 were considered significant.

[0154] Acetylcholine responses (Facetyichoiine) were normalized against the ionomycin response (Fionomycin) per cell, obtaining the maxim response called % Fmax:

[0155] % FmaX — 100 X F Acetylcholine / F Ionomycin

[0156] Only acetylcholine and ionomycin responding cells were considered. Finally, the acetylcholine response in cells treated was normalized against non-treated cells considered the 100%.

[0157] Percentage inhibition was calculated as % Inhibition = 100 - % Fmax. Table 5

[0158] As shown in the results above, treatment with various peptides of the invention provided a strong inhibition of calcium fluxes triggered by acetylcholine in HEK293 cells, which was in most cases equivalent to the inhibition achieved by the PLC inhibitor U73122.

[0159] Example 5 - Neural cell integrity in diabetic model

[0160] Methylglyoxal (MGO), an endogenous reactive carbonyl compound that is a physiological metabolite of glucose, mediates rapid non-enzymatic glycation of proteins, lipids, and DNA to promote formation of advanced glycation end products (AGEs), which renders irreversible damage to these macromolecules. The accumulation of MGO and advanced glycation end products (AGEs) has been detected in diabetes, aging and age-related neural disorders. Elevated levels of MGO and formation of AGEs in diabetic patients contribute to itch and loss of pain sensation (hypoalgesia) associated with peripheral diabetic neuropathy and neuronal damage.

[0161] The aim of this study was to evaluate the protective effect of various peptides of the invention on neural cell integrity (neurite count and total sum of length) in a neural degeneration model, in particular in methylglyoxal (MGO)-induced neural degeneration in neuron-like SH-SY5Y cells (Dwane S, et al., "Optimising parameters for the differentiation of SH-SY5Y cells to study cell adhesion and cell migration", BMC Res Notes. 2013 Sep 11 ;6:366). For this purpose, differentiated neuron-like SH-SY5Y cells were exposed to MGO and the impact on the neural cell integrity (neurite count and length ) was analyzed.

[0162] SH-SY5Y cells were seeded in 96-well plates at 20% cell confluence in 100 piL of supplemented DMEM / F12. After 24 hours, cell synchronization was carried out for 72 hours with 0.4 piM aphidicolin in supplemented medium during 72 hours at 37 °C and 5% CO2. Then, medium was removed, and cells were incubated for 4 days with 200 ng / mL hNGF (R&D Systems) and the indicated peptide at 100 pi M or DMSO as vehicle in presence or absence of 400 piM methylglyoxal (MGO) (Sigma) at 37 °C in CO2 incubator. Afterwards, neurite outgrowth and cell number were measured in the same sample using membrane and nucleus fluorescents probes respectively (Neurite OutGrowth Staining Kit from Fisher Scientific) after microscopic image capture with automated microscope (InCell Analyser 6000) using 555 nm filter for excitation and 565 nm for emission with the cell membrane indicator and exciting at 358 nm and collecting emission at 461 nm for nucleus stain.

[0163] Finally, captured images were analysed with Imaged software using different plugins that are freely available, Neuron J plugin for measurement of number and sum of length (pm) of neurites, and Cell counter plugin to quantify nucleus number.

[0164] The neurite outgrowth percentage was calculated in two ways from ratio between the sum of length (pm) or number of neurites and cell number of each image and subsequent normalization of ratio values respect to vehicle-treated cells (100 %), NN (neurite number) % and NTO (neurite total outgrowth) % respectively. The neurite regeneration percentage was calculated in two ways by double normalization of ratio values respect to vehicle-treated cells (0%) and vehicle NGF-treated cells (100 %), NNR (neurite number regeneration) % and NTOR (neurite total outgrowth regeneration) % respectively.

[0165] Table 6 below details neurite regeneration percentage values obtained for two peptides of invention expressed as neurite number regeneration (NNR %) and neurite total outgrowth regeneration (NTOR %)

[0166] Table 6

[0167] % neurite regeneration

[0168] As shown in the results above, the treatment with the peptides of the invention provided a strong neurite regeneration effect, clearly indicating that the peptides of the invention are useful for the treatment or prevention of diabetes, aging, and age-related neural disorders. Also, these results suggest that the peptides of the invention are useful for treating itch and loss of pain sensation (hypoalgesia) associated with peripheral diabetic neuropathy and neuronal damage. Example 6 - Neuronal excitability

[0169] Chloroquine-induced pruritus is the most characteristic side-effect of this common treatment and prophylactic of malaria. Chloroquine binding mouse MrgprA3 (MrgprXI in human) and NK1R provoke a histamine- independent itch response mediated by PLC.

[0170] Next, the effects of the peptides of the invention on PLC mediated neuronal excitability induced by chloroquine or bradikinin in differentiated neuroblastoma F11 cell line culture were evaluated (Pastor! V et al., "M. Serum-deprived differentiated neuroblastoma F-11 cells express functional dorsal root ganglion neuron properties". PeerJ. 2019;7:e7951).

[0171] For this purpose, hybridoma F11 cell line was maintained in culture in supplemented DMEM, High Glucose, Glutamax Supplement (1% penicillin-streptomycin, 10% FBS). Cells were seeded on pre-treated Poly-L-Lysin and Laminin 12 mm-glass coverslip. To get differentiated state, cells were maintained during 14 days in a deprived serum media (1% FBS) supplemented with 25 ng / mL of NGF. Non-differentiated cells were kept for 7 days in maintenance media (10% FBS). Cells were cultured at 37°C in 5% CO2.

[0172] Calcium imaging recordings were carried out 14 days for differentiated or 7 days for non-differentiated conditions after seeding cells, respectively. Measurements were carried out in an inverted fluorescent microscope (ZEISS Axiovert 200b; Software HCImage v. DIA. HAMAMATSU).

[0173] Differentiated cells were incubated with FLUO 4-AM dye (Invitrogen) at 6 pig / mL for 1h at 37°C, in presence or absence of the indicated peptide at 100 piM following of a rinsing with external solution at least for 20 min at 37°C. Then, coverslips were mounted onto the microscope set up and stimuli were applied by perfusion. Control non-differentiated and differentiated cells were exposed to 1 mM Chloroquine for 30 s or 1 piM bradykinin for 15 s, followed by 40 mM of KCI and then lonomycin 10 piM for 15 s, washing with external solution in between them. Differentiated cells were incubated with the peptide at 100 piM for 1 h. Changes in the intracellular calcium levels revealed by Fluo-4 dye (Ex / Em=492 / 506 nm) were registered as indicator of neuronal induced activity. Fluorescence images (fields) were taken every 3 s during stimuli applications and responses and every 10 s during external solution cleaning between stimuli.

[0174] Fluorescence recording of each individual cell was adjusted to baseline zero by subtraction of blank signal. Response against each stimulus was determined by subtraction of florescence peak signal and previous baseline average. Stimulus Response (F) per cell was normalized by lonomycin response quantitation (F for each stimulus obtaining the percentage of maxim response, called % Fmax. For chloroquine and bradykinin response, only cells with also KCI response were considered:

[0175] % Cloroquine Fmax= 100x FCQ <KCI (+» / F % Bradykinin Fmax= 100x FBK <KCI (+» / F ionomycinw

[0176] Table 7

[0177] As shown in the results above, treatment with a peptide of the invention provided a reduced neuronal response induced by chloroquine, bradykinin and KCI on the sensory neuron model differentiated F11 cell line. These results highlight the utility of the peptides of the invention in alleviating pain and itch.

[0178] Example 7 - Neuronal excitability PLC-beta-4

[0179] To confirm the effects of another peptide of the invention on PLC mediated neuronal excitability induced by bradykinin in a neuronal model, the neuroblastoma F11 cell line culture was used as above explained.

[0180] For this purpose, intracellular calcium fluxes triggered by bradykinin were analyzed in undifferentiated F11 cells using the calcium imaging technique. The hybridoma F11 cell line was maintained in culture in supplemented DMEM, High Glucose, Glutamax Supplement (1% penicillin-streptomycin, 10% FBS). Cells were seeded on pre-treated Poly-L-Lysin and Laminin 12 mm-glass coverslip. Cells were cultured at 37°C and 5% CO2.

[0181] Calcium imaging recordings were performed 48 h after seeding. Measurements were carried out in an inverted fluorescent microscope (ZEISS Axiovert 200b; Software HCImage v. DIA. HAMAMATSU) coupled to a Hamamatsu FLASH 4.0 LT camera (C11440-42U30, Hamamatsu, Sunayama-cho Japan).

[0182] Non-treated cells on coverslips were incubated with 6 pig / mL Fluo-4 AM prepared in prewarmed external solution with 0.05% pluronic acid for 1 h at 37°C (Fluo-4 NW Calcium Assay kit, Molecular Probes). In cells treated with AG2104 at 100 piM or U73122 at 2 piM as the reference PLC inhibitor, the compound was preincubated with the dye for 1 h at 37°C. Afterwards, cells were washed with external solution for at least 20 min at 37°C. Then, coverslips were mounted onto the microscope set up. Stimuli were applied in perfusion following the protocol described below. Two pulses of bradykinin 10 pi M were applied during 30 s separated by a 3 min wash with external solution to recover a stable basal line. Finally, ionomycin 1 pi M was applied briefly to check the integrity of cells. Changes in the intracellular calcium levels revealed by Fluo-4 dye (exciting the fluorophore at 483 nm during 200 ms and collecting the emission at 512 nm) were registered as indicator of neuronal induced activity. Fluorescence images were taken every 3 s during stimuli applications and responses.

[0183] Fluorescence intensity of individual cells was adjusted to baseline zero subtracting the background signal. The response to each stimulus was analyzed quantifying the fluorescence peak and subtracting the fluorescence average 30 s before applying it. Only cells with responses equal or greater than 20 were considered significant.

[0184] Bradykinin responses (Fsradykinin) were normalized against the ionomycin response (Fionomycin) per cell, obtaining the maxim response called % Fmax

[0185] % FmaX — 100

[0186] Only bradykinin and ionomycin responding cells were considered. Finally, the bradykinin response in cells treated was normalized against non-treated cells considered the 100%.

[0187] Percentage inhibition was calculated as % Inhibition = 100 - % Fmax.

[0188] Table 8

[0189] As shown in the results above, treatment with another peptide of the invention provided a reduced neuronal response induced by bradykinin on the sensory neuron model F11 cell line. Notably, the inhibitory effect of the peptide was even stronger than that obtained with the PLC inhibitor U73122.

[0190] Example 8 - Neuronal excitability in diabetic model

[0191] Next, it was evaluated the effects of a peptide on the invention on PLC mediated neuronal excitability induced by bradikinin or KCI in a diabetic-induced neuronal model induced by chronic exposure to 800 nM of methylglyoxal (MGO) on differentiated F11 neuronal cells (Pastori V. et al. supra).

[0192] For this purpose, hybridoma F11 cell line was maintained in culture in supplemented DMEM, High Glucose, Glutamax Supplement (1 % penicillin-streptomycin, 10% FBS). Cells were seeded on pre-treated Poly-L-Lysin and Laminin 12 mm-glass coverslip. To get differentiated state, cells were maintained during 14 days in a deprived serum media (1% FBS) supplemented with 25 ng / mL of NGF and 800 nM MGO. Cells were cultured at 37°C in 5% CO2.

[0193] Calcium imaging recordings were carried out 14 or 7 days after seeding cells, respectively. Measurements were carried out in an inverted fluorescent microscope (ZEISS Axiovert 200b; Software HCImage v. DIA. HAMAMATSU).

[0194] MGO treated were incubated with FLUO 4-AM dye (Invitrogen) at 6 pig / mL for 1h at 37°C, in presence or absence of the peptide at 100 piM following of a rinsing with external solution at least for 20 min at 37°C. Then, coverslips were mounted onto the microscope set up and stimuli were applied by perfusion. Bradykinin 1 piM, 40 mM KCI and lonomycin 10 piM were applied for 15 s, with 5 min wash with external solution between stimuli to recover stable basal line. Changes in the intracellular calcium levels revealed by Fluo-4 dye (Ex / Em=492 / 506 nm) were registered as indicator of neuronal induced activity. Fluorescence images (fields) were taken every 3 s during stimuli applications and responses and every 10 s during external solution cleaning between stimuli.

[0195] Fluorescence recording of each individual cell was adjusted to baseline zero by subtraction of blank signal. Response against each stimulus was determined by subtraction of florescence peak signal and previous baseline average. Stimulus Response (F) per cell was normalized by lonomycin response quantitation (F for each stimulus u obtaining the percentage of maxim response, called % Fmax. For brady kinin response, only cells with also KCI response were considered:

[0196] %

[0197] Table 9

[0198] As shown in the results above, treatment with the peptide of the invention provided a reduced neuronal response induced by bradykinin and KCI in a diabetic-induced neuronal model on differentiated F11 neuronal cells. These results clearly demonstrate the utility of the peptides of the invention for treating pain or sensory disfunction, such as itchiness, in diabetic conditions. Example 9 - TRPV1 sensitization

[0199] The activation of TRPV1 generates a cationic flow that triggers the firing of action potentials transducing the information from the peripheral nervous system to the central nervous system. Under inflammatory conditions, activation of PLC signaling cascade promotes TRPV1 sensitization through direct phosphorylation reducing its threshold gating (Mohapatra DP. et al., "Regulation of Ca2-i~dependent desensitization in the Vanilloid Receptor TRPV1 by Calcineurin and cAMP-dependent Protein Kinase", J Biol Che., 2005, vol. 280(14), pp. 13424-32).

[0200] The aim of this study was to evaluate the effect of various peptides of the invention on TRPV1 sensitization induced via PLC activation by bradykinin.

[0201] For this purpose, peptides were assessed using planar multielectrode arrays technology on primary cultures of neonatal rat dorsal root ganglia using the excess / remains of already isolated sensory neurons.

[0202] Neonatal dorsal root ganglia (DRGs) from Wistar rats were isolated, dissociated and cells were seeded in a drop on microelectrode array chambers previously coated with poly-l-lysine and laminin, using as culture medium DMEM glutamax, 10% FBS and 1% P / S, supplemented with mouse 2.5S NGF 50 ng / mL and 1.25 pg / mL cytosine arabinoside as previously described (Butron L, et al., "Design and validation of neuronal exocytosis blocking peptides as potential novel antiperspirants". Exp Dermatol., 2023 Apr 3). All experiments were performed after 48 h cell seeding.

[0203] Extracellular recordings were performed using multiple electrode planar arrays of 60-electrode thin MEA chips, with 30 pm diameter electrodes and 200 pm inter-electrode spacing with an integrated reference electrode (Multichannel Systems GmbH). The electrical activity of primary sensory neurons was recorded by the MEA1060 System (Multi Channel Systems GmbH1), and MC_Rack software version 4.3.0 at a sampling rate of 25 kHz. To study the activity of the indicated peptides under sensitization condition, the peptides were evaluated on bradykinin mediated PLC neuronal TRPV1 sensitization protocol. U73122 at 5 piM was used as a control of inhibition. All compounds were pre-incubated for 1 h before starting electrical activity measurements at 100 piM and 10 piM (SEQ ID NO: 28, 24), at 1 piM (SEQ ID 42, 40, 47, 48, 49, 46) or at 100 piM (SEQ ID NO: 34). For that, three shorts 15s-applications (defined as P1 , P2 and P3, respectively) of capsaicin at 500 nM. Between P1 and P2, cells were washed with external solution for 2 min and 30 s, using continuous perfusion system. Between P2 and P3, after washing for 1 min and 30 s with external solution, cells were treated with the PLC activator bradykinin at 1 pi M for 7 min. At the end of each protocol 40 mM KCI pulse was applied to confirm appropriate cell culture excitability and viability. All measurements were performed at ~34.5°C.

[0204] Data were analyzed using MC_RACK spike sorter and Neuroexplorer Software (Nex Technologies). An evoked spike was defined when the amplitude of the neuronal electrical activity overcame a threshold set at - 15 pV. The recorded signals were processed, and mean spike frequency was extracted. TRPV1 potentiation induced by bradykinin via PLC was analyzed as ratio P3 / P2 of mean spike frequency. Then, TRPV1 -mediated responses were normalized to control conditions as percentage.

[0205] Table 10

[0206] As shown in the results above, all the peptides of the invention assayed, bearing different terminal modifications, provided a strong inhibition of TRPV1 mediated neuronal excitability potentiation induced by bradykinin. Finally, the reduction of peptide length from the C-terminus seemed not to affect efficacy on PLC mediated sensitization.

[0207] These results clearly demonstrate the utility of the peptides of the invention for treating diseases or disorder mediated by TRPV1, such as pain, pruritus (itch), sensitive skin, atopic dermatitis, contact dermatitis, allergic dermatitis, psoriasis, rosacea, psoriatic arthritis or inflammation, radiotherapy induced dermatitis or chemotherapy -mediated skin sensitization.

[0208] Example 10 - Acetylcholine response

[0209] The aim of this study was to evaluate the effect of various peptides of the invention on the acetylcholine response, intracellular calcium fluxes triggered by acetylcholine were analyzed in HEK-293 cell line (ATCC).

[0210] HEK293 cells using the calcium imaging technique. HEK293 cells were maintained in supplemented medium (DMEM, high glucose, pyruvate, no glutamine containing 10% Heat Inactivated Foetal Bovine Serum, 1% Penicillin-Streptomycin and 1% L-glutamine). Cells were seeded on pre-treated Poly-L-Lysin 12 mm-glass coverslip and cultured at 37°C and 5% CO2. Calcium imaging recordings were performed 48 h after seeding. Measurements were carried out in an inverted fluorescent microscope (ZEISS Axiovert 200b; Software HCImage v. DIA. HAMAMATSU) coupled to a Hamamatsu FLASH 4.0 LT camera (C11440-42U30, Hamamatsu, Sunayama-cho Japan).

[0211] Non-treated cells on coverslips were incubated with 6 pig / mL Fluo-4 AM prepared in prewarmed external solution with 0.05% pluronic acid for 1 h at 37°C (Fluo-4 NW Calcium Assay kit, Molecular Probes). In cells treated with the indicated peptides at 100 piM or U73122 at 2 piM as the reference PLC inhibitor, the compound was preincubated with the dye for 1 h at 37°C. Afterwards, cells were washed with external solution for at least 20 min at 37°C. Then, coverslips were mounted onto the microscope set up. Stimuli were applied in perfusion following the protocol described below. Two pulses of acetylcholine 1 piM were applied during 30 s separated by a 3 min wash with external solution to recover a stable basal line. Finally, ionomycin 10 pi M was applied briefly to check the integrity of cells. Changes in the intracellular calcium levels revealed by Fluo-4 dye (exciting the fluorophore at 483 nm during 200-300 ms and collecting the emission at 512 nm) were registered as indicator of acetylcholine response. Fluorescence images were taken every 3 s during stimuli applications and responses.

[0212] Fluorescence intensity of individual cells was adjusted to baseline zero subtracting the background signal. The response to each stimulus was analyzed quantifying the fluorescence peak and subtracting the fluorescence average 30 s before applying it. Only cells with responses equal or greater than 20 were considered significant.

[0213] Acetylcholine responses were normalized against the ionomycin response per cell, obtaining the maxim response called % Fmax % FmaX — 100

[0214] Only acetylcholine and ionomycin responding cells were considered. Finally, the acetylcholine response in cells treated was normalized against non-treated cells considered the 100%.

[0215] Percentage inhibition was calculated as % Inhibition = 100 - % Fmax.

[0216] Table 11

[0217] As shown in the results above, treatment with peptides of the invention provided a strong inhibition of calcium fluxes triggered by acetylcholine in HEK293 cells.

[0218] Citation List

[0219] Kadamur G and Ross EM, "Mammalian phospholipase C", Annual review of physiology, 2013, vol. 15, pp 127-154

[0220] Cocco L et al., "Phosphoinositide-specific phospholipase C in health and disease", Journal of Lipid Research, 2015, vol. 56 (10), pp.1853-1860

[0221] Shi T-JS, et al., "Phospholipase Cp3 in mouse and human dorsal root ganglia and spinal cord is a possible target for treatment of neuropathic pain" PNAS, 2008, vol. 105(50), pp. 20004-20008

[0222] Luo J, et al., "Molecular and cellular mechanisms that initiate pain and itch", Cellular and molecular life sciences, 2015, vol. 72(17), pp. 3201-3223

[0223] Phan HTN et al., " Uveal melanoma-associated mutations in PLCp4 are constitutively activating and promote melanocyte proliferation and tumorigenesis", Sci Signal, 2021, Dec 14; 14(713)

[0224] Miyata M. et al., " Role of Thalamic Phospholipase Cp4 Mediated by Metabotropic Glutamate Receptor Type 1 in Inflammatory Pain" J Neurosci, 2003 Sep 3; vol. 23(22); pp. 8098-8108

[0225] Lyon AM et al., "Molecular mechanisms of Phospholypase C p3 autoinhibition”, Structure. 2014 Dec 2;22(12): 1844-1854

[0226] Copolovici D. M. et al., "Cell-Penetrating Peptides: Design, Synthesis, and Applications”, 2014, ACS Nano, 2014, 8 (3), pp 1972-1994

[0227] Dwane S, et al., "Optimising parameters for the differentiation of SH-SY5Y cells to study cell adhesion and cell migration", BMC Res Notes. 2013 Sep 11 ;6:366

[0228] Pastor! V et al., "M. Serum-deprived differentiated neuroblastoma F-11 cells express functional dorsal root ganglion neuron properties". PeerJ. 2019;7:e7951

[0229] Han SK et al., "Phospholipase Cbeta 3 mediates the scratching response activated by the histamine H1 receptor on C-fiber nociceptive neurons", Neuron., 2006, vol. 52(4), pp. 691-703

[0230] Xiao W et al., "Phospholipase C-p3 regulates FceRI-mediated mast cell activation by recruiting the protein phosphatase SHP-1", Immunity, 2011, vol. 34(6), pp. 893-904

[0231] Descorbeth M et al., "Role of oxidative stress in high-glucose- and diabetes-induced increased expression of Gq / 11a proteins and associated signaling in vascular smooth muscle cells", Free Radio Biol Med, 2010, vol. 49(9), pp. 1395-405

[0232] Shi TJ et al., "Phospholipase C{beta}3 in mouse and human dorsal root ganglia and spinal cord is a possible target for treatment of neuropathic pain", Proc Natl Acad Sci U S A, 2008, vol. 105(50), pp. 20004-8

[0233] Tappia P S et al., "Upregulation of Phospholipase C Gene Expression Due to Norepinephrine-Induced Hypertrophic Response", Cells., 2022, vol. 11(16), pp. 2488

[0234] Bellono NW et al., " UV light activates a Gaq / 11 -coupled phototransduction pathway in human melanocytes", J Gen Physiol., 2014, vol. 143(2), pp. 203-14

[0235] Palomo JM "Solid-phase peptide synthesis: an overview focused on the preparation of biologically relevant peptides" RSC advances, 2014, issue 62

[0236] Coin I. et al., " Solid-phase peptide synthesis: from standard procedures to the synthesis of difficult sequences", Nature Protocols, 2007, vol. 2, pp, 3247-3256

[0237] Kan W, et al., "M3 muscarinic receptor interaction with phospholipase C p3 determines its signaling efficiency", J Biol Chem., 2014, vol. 289(16), pp. 11206-18

[0238] Mohapatra DP. et al., "Regulation of Ca2-<~dependent desensitization in the Vanilloid Receptor TRPV1 by Calcineurin and cAMP-dependent Protein Kinase", J Biol Che., 2005, vol. 280(14), pp. 13424-32

[0239] Butron L, et al., "Design and validation of neuronal exocytosis blocking peptides as potential novel antiperspirants". Exp Dermatol., 2023 Apr 3

[0240] Ando T. et al., "Critical role for mast cell Stat5 activity in skin inflammation", Cell Rep., 2014, vol. 6(2), pp. 366-76

[0241] WC2013064583

[0242] WO2019238686

Claims

Claims1. A peptide or a pharmaceutically, veterinary, or cosmetically acceptable salt thereof, wherein: the peptide comprises at least 5 consecutive amino acid residues of a sequence at least 80% identical to the sequence TTDEGTASSEVNATEEMSTLVN (SEQ ID NO: 1) or the sequence EQAWMASYKYVGATTNIHPYLS (SEQ ID NO: 2); the peptide has a length equal to or lower than 22 amino acid residues; and the peptide is capable of inhibiting PLC-beta activity.

2. The peptide according to claim 1, wherein the peptide is capable of inhibiting PLC-beta-3 activity and / or PLC-beta-4 activity.

3. The peptide according to claim 1 or 2, with the proviso that the peptide does not consist of the amino acid sequence IEVNATEEM (SEQ ID NO: 23).

4. The peptide according to any one of claims 1-3, wherein the peptide comprises or consists of at least 6 consecutive amino acid residues of a sequence at least 75 % identical to the sequence TASSEVNATEEM (SEQ ID NO: 4) or the sequence ASYKYVGATTNIH (SEQ ID NO: 16).

5. The peptide according to any one of claims 1-4, wherein the peptide consists of 6 consecutive amino acid residues of a sequence at least 75 % identical to the sequence TASSEVNATEEM (SEQ ID NO: 4) or the sequence ASYKYVGATTNIH (SEQ ID NO: 16).

6. The peptide according to any one of claims 1-5, wherein the peptide consists of a sequence selected from SEQ ID NO: 5-14, 17-22, and 24-51.

7. The peptide according to any one of claims 1-6, which further comprises an N-terminal and / or C-terminal modification.

8. The peptide according to claim 7, wherein the N-terminal and / or C-terminal modification comprises a compound selected from the group consisting of phosphoryl group, glycosyl group, acyl group, alkyl group, carboxyl group, hydroxyl group, biotinyl group, ubiquitinyl group, and amido group; particularly, wherein the acyl group is selected from the group consisting of acetyl group, lauroyl group, myristoyl group, and palmitoyl group.

9. The peptide according to claim 7 or 8, wherein the N-terminal modification consists of an acetyl group and / or the C-terminal modification consists of an amido group.

10. A pharmaceutical, veterinary, or cosmetic composition comprising:(i) a therapeutically or cosmetically effective amount of a peptide or a pharmaceutically, veterinary, or cosmetically acceptable salt thereof as defined in any one of claims 1-9; and(II) at least one pharmaceutically, veterinary, or cosmetically acceptable excipient, diluent, or carrier.11 . A peptide or a pharmaceutically, veterinary, or cosmetically acceptable salt thereof as defined in any one of claims 1-9 or a pharmaceutical or veterinary composition as defined in claim 10, for use as a medicament.

12. A peptide or a pharmaceutically, veterinary, or cosmetically acceptable salt thereof as defined in any one of claims 1-9 or a pharmaceutical or veterinary composition as defined in claim 10, for use in the treatment or prevention of a disease or disorder mediated by PLC activation; particularly by PLC-beta activation.

13. The peptide or the composition for use according to claim 12, wherein the disease or disorder mediated by PLC activation is selected from the group consisting of cancer, inflammation, skin disease, pain, diabetes, atopia, and cardiovascular disease.

14. Use of a peptide or a pharmaceutically, veterinary, or cosmetically acceptable salt thereof as defined in any one of claims 1-9 or a cosmetic composition as defined in claim 10, for the cosmetic, non-therapeutic treatment or care of the skin, hair, nails, or mucous membranes.

15. The use according to claim 14, wherein the cosmetic, non-therapeutic treatment or care is selected from the group consisting of depigmentation or whitening or lightening in colour of the skin; maintenance or improvement of skin luminosity, skin elasticity, skin resistance, skin firmness, skin tensile strength, skin smoothness, skin hydration, skin barrier function, skin microcirculation, or skin tone; treatment or prevention of the symptoms of skin aging, skin fatigue, skin wrinkles, skin redness, eye bags, unregulated pigmentation, excessive perspiration, unregulated sebum production, and unregulated hair growth.

16. A kit of parts comprising the peptide as defined in any one of claims 1-9 or the composition as defined in claim 10, and instructions for its use.

17. An in vitro use of the peptide or a pharmaceutically, veterinary, or cosmetically acceptable salt thereof as defined in any one of claims 1-9 or the composition as defined in claim 10, for the inhibition of PLC activity, particularly PLC-beta activity.

18. An in vitro method for inhibiting PLC activity, particularly PLC-beta activity, in a sample, the method comprising contacting the sample with a peptide or a pharmaceutically, veterinary, or cosmetically acceptable salt thereof as defined in any one of claims 1-9 or the composition as defined in claim 10.