WO2018085565A2 - Short bioactive peptides blocking activity of advanced glycation end products, compositions, and methods of use - Google Patents

Abstract

Advanced glycation end products (AGEs) represent a non-enzymatic post-translational protein modification. AGEs are very stable and the accumulation over time can be used as markers of carbonyl stress which are responsible for the malfunction of bioactive macromolecules in the body. Disclosed herein are a series of short peptides for blocking activity of advanced glycation end products (AGEs), the peptide being derived from a V-region of an sRAGE protein, wherein the V-region is given by SEQ ID NO:61. These peptides effectively inhibit the progress of a Maillard reaction, blocking the AGEs and inhibiting dicarbonyl induced cell apoptosis and death. The peptides can be used as, for example, an anti-glycation agent, a skin anti-aging agent, or an anti-diabetic complication agent.

Description

SHORT BIOACTIVE PEPTIDES BLOCKING ACTIVITY OF ADVANCED GLYCATION END PRODUCTS, COMPOSITIONS, AND METHODS OF USE

CROSS REFERENCE TO RELATED APPLICATION [0001] This application claims the benefit under 35 U.S.C. § 119(e) to U.S. Provisional Application No. 62/417,137, filed November 3, 2016, which application is hereby incorporated by reference in its entirety.

STATEMENT REGARDING SEQUENCE LISTING

[0002] The Sequence Listing associated with this application is provided in text format in lieu of a paper copy, and is hereby incorporated by reference into the specification. The name of the text file containing the Sequence Listing is 150145_404WO_SEQUENCE_LISTING. The text file is 14.9 KB, was created on October 23, 2017 and is being submitted electronically via EFS-Web.

FIELD OF THE INVENTION

[0003] The present disclosure relates to peptides for blocking activity of advanced glycation end products.

INTRODUCTION

[0004] Advanced glycation end-products (AGEs) represent a post-translational modification process between free reducing sugars and free amino groups in proteins, lipids, or nucleic acids that adversely affect the functional properties of the molecules. The initial reaction is a non-enzymatic glycosylation also known as Maillard reaction, at the ε-amino group of lysine or at its free amino group, although side chains of the amino acids cysteine, arginine or tryptophan can also be potential sites for glycation. The intermediate products are known, variously, as Amadori, Schiff base, and Maillard products, named after the researchers who first described them. After further oxidation, dehydration, and condensation, the Amadori products undergo further structural changes to finally yield highly stable AGE compounds. Once AGEs are formed, they are nearly irreversible. The reaction causes the formation of protein adducts

subsequently resulting in denaturing, browning, and cross-linking of the targeted proteins.

[0005] AGEs exert their deleterious actions not only by deactivating proteins but also through their interaction with specific receptors. The binding of ligands to certain AGE receptors stimulates various signaling pathways resulting in activation of the transcription factor nuclear factor kappa-B (NFkB) and subsequent transcription of many pro-inflammatory genes. AGE receptors and their ligands may be involved in the pathobiology of a wide range of diseases that share common features, such as enhanced oxidative stress, immune/inflammatory responses, and altered cell functions.

[0006] The accumulation of AGEs over time can be used as markers of carbonyl stress which are responsible for the malfunction of bioactive macromolecules in the body. AGEs are formed both endogenously via glucose and fructose metabolism and exogenously via diet and tobacco. The receptor of AGEs (RAGE) is a multi-ligand member of the immunoglobulin super-family of cell surface receptors and is highly expressed in many cell types including fibroblasts, keratinocytes, endothelial cells, immune cells, neurons, blood vessel walls, bone, and tumor cells, etc. RAGE activation induces multiple intracellular signaling pathways that have been implicated in the pathogenesis of serious systemic complications and aging. SUMMARY OF THE INVENTION

[0007] Provided herein is a series of short peptides that effectively inhibit the progress of a Maillard reaction, block the AGEs and/or inhibit dicarbonyl induced cell damage, inflammation, apoptosis and death. These manufactured peptides are promising therapeutics as antiglycation agents, skin anti-aging agents, and anti- diabetic complication agents.

[0008] In some embodiments, provided herein is a peptide for blocking the activity of advanced glycation end products (AGEs), the peptide being a 2-9 amino acid subsequence of SEQ ID NO:61.

[0009] In some embodiments, provided herein a composition including at least one peptide according to the disclosure and a pharmaceutically acceptable carrier.

[0010] In some embodiments, provided herein is a method for treating a skin condition in a subject comprising administering to the subject a medicament comprising a pharmaceutically acceptable carrier and at least one peptide according to the disclosure.

[0011] In some embodiments, the present disclosure provides a method for treating a condition, disease, or disorder in a subject comprising administering to the subject an effective amount of at least one peptide disclosed herein. The at least one peptide is comprised in a medicament comprising a pharmaceutically acceptable carrier. The condition, disease or disorder is diabetes, metabolic syndrome, diabetic skin chronic wounds, Alzheimer's disease, atherosclerosis, osteoporosis, osteoarthritis, cancer, cancer therapy associated condition, pollution associated assaults, body and skin detoxification, renal failure, diabetic retinopathy and glaucoma, periodontal disease, cystic fibrosis related diabetes, polycystic ovary syndrome, psoriasis associated condition, oxidative stress, complications associated with maternal chorioamnionitis or funisitis, or sarcopenia.

[0012] In some embodiments, provided herein is a method of inhibiting formation of AGEs in a food or beverage comprising contacting the food or beverage with at least one peptide according to the disclosure.

[0013] In some embodiments, provided herein is a method of manufacturing a skin care product for blocking activity of AGEs including: synthesizing a first peptide, wherein the first peptide is between 2 and 9 amino acids in length and is a

subsequence of SEQ ID NO:61 , and wherein the first peptide blocks activity of AGEs; and blending the first peptide with a carrier.

[0014] Additional embodiments will become apparent by consideration of the detailed description and accompanying drawings.

BRIEF DESCRIPTION OF THE DRAWINGS

[0015] Figure 1 shows the amino acid sequence of the AGE receptor isoform 1 of human (NCBI Reference Sequence: NP_001 127.1), where the underlined region indicates the active binding region of RAGE to AGEs from which the peptides disclosed herein have been generated.

[0016] Figure 2 shows the amino acid sequence of the V-region of the AGE receptor sequence of Figure 1. DETAILED DESCRIPTION OF THE INVENTION

[0017] Disclosed herein are a series of short peptides that provide the advantage of effectively inhibiting the progress of a Maillard reaction, blocking the AGEs and/or inhibiting dicarbonyl induced cell apoptosis and death. The peptides disclosed herein are advantageous as therapeutics, anti-glycation agents, skin anti-aging agents, and anti-diabetic complication agents. Accordingly, the peptides of the disclosure may be formulated as a composition or medicament (e.g., lotion, cream, etc.) that can be administered to a subject for the treatment or amelioration of a condition, disease or disorder.

[0018] In the present description, any concentration range, percentage range, ratio range, or integer range is to be understood to include the value of any integer within the recited range and, when appropriate, fractions thereof (such as one tenth and one hundredth of an integer), unless otherwise indicated. Also, any number range recited herein relating to any physical feature, such as polymer subunits, size or thickness, are to be understood to include any integer within the recited range, unless otherwise indicated. As used herein, the term "about" means ± 20% of the indicated range, value, or structure, unless otherwise indicated.

[0019] It should also be noted that the term "or" is generally employed in its sense including "and/or" (i.e., to mean either one, both, or any combination thereof of the alternatives) unless the content dictates otherwise.

[0020] Also, as used in this specification and the appended claims, the singular forms "a," "an," and "the" include plural referents unless the content dictates otherwise.

[0021] The terms "include," "have," "comprise" and their variants are used synonymously and to be construed as non-limiting.

[0022] The term "a combination thereof as used herein refers to one of the all possible combinations of the listed items preceding the term. For example, "A, B, C, or a combination thereof" is intended to refer to any one of: A, B, C, AB, AC, BC, or ABC. Similarly, the term "combinations thereof" as used herein refers to all possible combinations of the listed items preceding the term. For instance, "A, B, C, and combinations thereof is intended to refer to all of: A, B, C, AB, AC, BC, and ABC.

[0023] Advanced glycation end-products (AGEs) are a heterogeneous group of molecules but share some common characteristics including the formation of covalent cross-links between proteins, the effect of turning some foodstuffs a yellow-brown color, and the ability to generate fluorescence. Based on their properties AGEs can be categorized as: 1) fluorescent cross-linking AGEs such as pentosidine, 2-(2-furoyl)- 4(5)-2(2-fluranyl)-1 H-imidazole, glyxal-lysine dimer, and methylglyoxal-lysine dimer (MOLD); and 2) non-fluorescent and non-cross-linking agents such as N- carboxymethyllysine (CML), l\l£-(carboxyethyl)-lysine (CEL), and pyrraline. CML was the first described and represents the most prevalent AGE in vivo. Because CML can form in vitro from low density lipids (LDL) incubated with copper ions and glucose, it has been postulated that it can form both lipid and protein adducts. [0024] AGEs are formed and accumulated both endogenously and exogenously. Endogenous AGE formation occurs slowly in normal aging. Tissue levels of AGEs increase with age. In diabetes and metabolic syndrome, hyperglycemia can accelerate the formation of AGEs. Endogenously, glucose auto-oxidation or glycolysis can generate highly reactive dicarbonyl compounds such as glyoxal, methylglyoxal (MG), and 3-deoxyglucosone as metabolic by-products, all of which can interact with proteins to form AGEs.

[0025] Methylglyoxal (MG), a potent glycating agent, is produced mainly during glucose and fructose metabolism. In comparison to the parent sugar, MG is more likely to bind to arginine, lysine, and cysteine residues of proteins, causing the formation of a number of AGEs and subsequently more cross-linking to various proteins. In addition to formation of AGEs, MG reacts rapidly with RNA and DNA and has both mutagenic and clastogenic activities. It can induce numerous adverse reactions if not efficiently detoxified. MG is not only found at high levels in the blood of diabetic patients but also in hypertension, where it covalently depletes glutathione, leading to serious

toxicological effects. MG also contributes to dysfunction of adipose tissue during type 2 diabetes progressions.

[0026] Similar to MG, glyoxal is also a toxic dicarbonyl compound capable of damaging cells via AGE formation. One of the AGEs formed in the presence of glyoxal is CML, which has been used extensively as a biomarker for aging. CML can be formed by different pathways: glucose can be oxidized to glyoxal, which can react with protein to form CML; glucose can also react with protein to form fructose-lysine (an Amadori product), which then undergoes oxidative cleavage to form CML.

[0027] AGEs may also be introduced into organisms by means of exogenous sources such as tobacco and diet. The western style diet contains a substantial proportion of industrially processed foods that have been shown to contain high levels of AGEs, which is the main exogenous source and is associated with the development of several pathological conditions such as diabetes. Food preparation methods using high temperatures (frying, roasting and grilling) enhance the production of AGEs. In addition, MG can be generated by degradation of carbohydrates in food and beverages during food processing.

[0028] AGEs exert their deleterious actions not only by deactivating proteins but also through their interaction with specific receptors. Receptor for AGEs (RAGE) is a multi-ligand member of the immunoglobulin superfamily of cell surface receptors. It is a pattern recognition receptor binding to various other molecules in addition to AGEs. Six receptors that recognize and bind AGEs have been identified, among which RAGE is the best characterized and most extensively studied. RAGE is a type I transmembrane protein receptor, expressed mainly on the surface in many cell types. The binding of ligands to RAGE stimulates various signaling pathways resulting in activation of the transcription factor nuclear factor kappa-B (NFkB) and subsequent transcription of many pro-inflammatory genes. RAGE and its ligands are intimately involved in the pathobiology of a wide range of diseases that share common features, such as enhanced oxidative stress, immune/inflammatory responses, and altered cell functions. RAGE is highly expressed in fibroblasts, keratinocytes, immune cells, neurons, blood vessel walls, bone, and tumor cells. RAGE activation by high serum and tissue levels of AGEs induces multiple intracellular signaling pathways that have been implicated in the pathogenesis of serious complications including diabetes and diabetes associated atherosclerosis, cardiovascular disease, nephropathy and chronic inflammatory conditions.

[0029] Structurally, RAGE is composed of an N-terminal extracellular domain with a ligand-engaging V-region and two cytosolic-regions; a single transmembrane domain and a C-terminal highly charged, short cytoplasmic domain essential for signal transduction. A soluble form, in contrast to the membrane bound form, has been characterized and is called sRAGE.

[0030] Figure 1 shows the amino acid sequence of isoform 1 of human RAGE. The inventors focused on the V-region located in the N-terminal portion of the human isoform 1 of RAGE, which is underlined in Figure 1 and shown separately in Figure 2. Prior to this study, it was unknown whether the entire N-terminal 77-amino acid region shown in Figure 2 was required to bind AGE. For example, it was unknown 1) whether a linear sequence, meaning a continuous sequence from the amino to the carboxy terminus (N to C), is required for the binding; 2) whether a secondary structure is required, meaning does the binding site require a 3-dimensional configuration; 3) whether more than one active binding sites would be present in the 77-amino acid long sequence shown in Figure 2; and 4) the minimal number of amino acids required for binding.

[0031] The inventors first utilized a "shotgun" approach to randomly generate overlapping sequences from 5 to 9 amino acids in length along the 77 amino acid region. These sequences are represented by SEQ ID NOs 1-17 (Table 1).

Significantly, these sequences are solely in s/7/co-designed sequences which do not exist or occur naturally, e.g. in microbes, animals to plants. The binding activity was found encouraging as 10 out of the 17 sequences showed positive AGE binding (Table 2) suggesting there are multiple binding sites present in the 77-amino acid region shown in Figure 2, and the binding sites are linear, meaning a continuous linear sequence (from N-C) is required for binding. The next step was to find out the minimal number of amino acids required for the binding, therefore shorter fragments of 2-4 amino acids in length were generated, also using in silico design. These sequences are represented by SEQ ID NOs 18 to 59, shown in Table 1 , along with an indication of whether the peptides are in an amidated (NH2) or free acid (OH) form.

Table 1. Amino acid sequence of peptides

28 HB2306 LEWK-NH2

29 HB2288 KGAP-OH

30 HB2309 KGAP-NH2

31 HB2291 GGGP-OH

32 HB2295 TGRT-OH

33 HB2311 TGRT-NH2

34 HB2313 EWKL-OH

35 HB2321 GPWD-OH

36 HB2322 GPWD-NH2

37 HB2276 FLP-OH

38 HB2304 FLP-NH2

39 HB2285 LP-OH

40 HB2307 LP-NH2

41 HB2290 GGP-OH

42 HB2286 FL-OH

43 HB2308 FL-NH2

44 HB2293 GRT-OH

45 HB2310 GRT-NH2

46 HB2296 SPQ-OH

47 HB2297 SPQ-NH2

48 HB2292 SPG-OH

49 HB2294 TGR-OH

50 HB2287 GP-OH

51 HB2314 GP-NH2

52 HB2299 SP-OH

53 HB2300 SP-NH2

54 HB2315 TG-NH2

55 HB2316 TG-OH

56 HB2317 PQ-NH2

57 HB2319 PQ-OH

58 HB2320 RT-NH2

59 HB2318 RT-OH

[0032] The peptides were tested for blocking AGEs as described in Example 1 , and the results are shown in Table 2. Carnosine was used as antiglycation positive control in all of the experiments conducted in this study. As shown in Table 2, the peptides which effectively bind to a mixture of AGEs include SEQ ID NOs 1 , 4, 5, 8, 9, 12, 13, 14, 15, 16, 18, 19, 20, 21 , 22, 23, 24, 25, 27, 28, 29, 32, 33, 34, 35, 36, 38, 39, 41 , 42, 44, 45, 46, 47, 48, 49, 52, 54, 55, 56, 57, 58, and 59. The specificity of each peptide was also tested for binding to CEL and CML. As shown in Table 3, most peptides also significantly bind to one or both of CEL and CML. [0033] Several peptides did not show binding using a polyclonal antibody against AGEs (Table 2) but showed specific binding to either CEL or CML using monoclonal antibodies; these peptides are SEQ ID NOs 30, 31 , 37, 40, 43, and 51 (Table 3). The disclosed peptides not only effectively bind various AGEs but at the cellular level also reduce MG-induced apoptosis to keratinocytes (Table 4). As the data shown in Table 4 indicate, the peptides that significantly reduced keratinocyte apoptosis include SEQ ID NOs 1 , 5, 8, 12, 13, 18, 20, 21 , 22, 23, 24, 25, 26, 27, 29, 30, 31 , 32, 33, 35, 36, 37, 38, 39, 40, 41 , 43, 44, 45, 47, 48, 49, 52, 54, 55, and 58.

[0034] Although several potent AGE binding peptides did not show significant protection for methylgloxal induced apoptosis to keratinocytes, they did show significant protection for human fibroblasts against methylglyoxal induced cell death (Table 5); these peptides are SEQ ID NOs 42, 56, 57, 59. All the sequences binding to AGEs, and/or suppressing/reducing keratinocyte apoptosis, and/or protecting fibroblast survival are useful for reducing or blocking the toxic effects due to AGEs. Such activity, with the help of a proper formulation and delivery system, may be used in treating conditions associated with AGEs from aging to other chronic inflammatory conditions including, but not limited to, diabetes, metabolic syndrome, diabetic skin chronic wounds, Alzheimer's disease, atherosclerosis, osteoporosis, osteoarthritis, cancer therapy, pollution associated assaults, body and skin detoxification, renal failure, diabetic retinopathy and glaucoma, periodontal disease, cystic fibrosis related diabetes, polycystic ovary syndrome, psoriasis associated condition, and oxidative stress, among others.

[0035] Many of the peptides also effectively block AGE formation between sugar and collagen. When incubated with calf collagen and glucose, many of the peptides significantly inhibit the Maillard reaction between glucose and collagen as shown in

Table 6. These peptides include SEQ ID NOs 3, 8, 9, 12, 13, 14, 23, 24, 25, 29, 34, 35, 38, 39, 40, 41 , 42, 46, 48, 49, 52, 53, 55 and 58. These peptides can be useful to inhibit the Maillard reaction in a subject's body or elsewhere (e.g. added to a food or beverage product) so as to prevent this and similar reactions in food or beverages containing sugar and collagen.

[0036] The peptides disclosed herein were synthesized using standard Fmoc (9- fluorenylmethoxycarbonyl) solid-phase chemistry, although those skilled in the art will recognize that other synthesis technologies may be used. As the peptides are prepared synthetically, the disclosed peptides are essentially free of post-translational modifications, although various modifications may be added in a controlled manner to confer certain desired properties on the peptides. For example, the peptides may be prepared as either amidated (NH2) or free acid (OH) sequences using standard amino acids; Table 1 indicates the form of each peptide disclosed herein. Amidation of the carboxy-terminus may render the inventive peptides less susceptible to protease degradation and increase their solubility compared to the free acid form, therefore providing heightened therapeutic potency. The peptides may include L- or D-amino acid enantiomers, either containing residues of one enantiomeric form or a

combination of both forms.

[0037] Furthermore, the peptides may be modified on one or both of the N- terminus and the C-terminus. For example, N-terminus lipidation or acylation may improve peptide penetration across skin without altering the bioactive function of the peptide, which may provide for enhanced skin penetration. Examples of saturated or unsaturated fatty acids that can be used to provide a C12-18 lipid-component to the compounds disclosed herein include lauric acid, myristic acid, palmitic acid, stearic acid, myristoleic acid, palmitoleic acid, oleic acid, and linoleic acid, although other lipid sources may also be used. The carboxy-terminus of the peptides can be modified to be acidic (-COOH) or amidated (e.g., -CONH2, -CONHR, or -CONR2). Amidation of the carboxy-terminus may confer advantageous properties on the peptides, for example rendering the inventive peptides less susceptible to protease degradation and/or increasing their polarity compared to the free acid forms, therefore providing heightened therapeutic potency. Also the peptide functional groups that may typically be modified include hydroxyl, amino, guanidinium, carboxyl, amide, phenol, imidazol rings or sulfhydryl.

[0038] Peptides may also be conjugated to soluble or insoluble carrier molecules to modify their solubility properties as needed and to control (e.g. increase) the local concentrations of peptides in targeted tissues. Examples of soluble carrier molecules include, but are not limited to, polymers of polyethyleneglycol (PEG) and

polyvinylpyrrolidone; examples of insoluble polymers include, but not limited to, silicates, polystyrene, and cellulose. To enhance their stability and provide controlled release, peptides may be micro-encapsulated using liposome technology or via nano- technology in the form of nano-emulsions, nanoliposomes, or other types of nanoparticles or nanomaterials including those disclosed in Sharma et al. (2012 Int J Pharm Pharm Sci, Vol 4, Issue 3, 57-66, incorporated herein by reference in its entirety), using methods known to those skilled in the art. In general, the peptides may be produced using any method known to those skilled in the art such as those disclosed in Merrifield (J Am Chem Soc. 85:2149, 1963); Carpino et al. (J Org Chem. 51 :3732, 1986); Merrifield et al. (Anal Chem. 38: 1905, 1966); or Kent et al. [High Yield Chemical Synthesis Of Biologically Active Peptides On An Automated Peptide

Synthesizer Of Novel Design, IN: PEPTIDES 1984 (Ragnarsson, ed.) Almqvist and Wiksell Int., Stockholm (Sweden), pp. 185-188], all of which are herein incorporated by reference in their entirety.

[0039] In various embodiments, included herein are compositions for using the above-described peptides, such as in formulations or as therapeutic agents. Also included are methods of using the above described peptides, such as in formulations or as therapeutic agents. These methods may involve the use of a single peptide, or multiple peptides in combination or combined with other peptides having collagen stimulating activity or anti-inflammatory activity, or combined with materials such as antioxidants. In certain instances, the inventive composition can be disposed within devices placed upon, in, or under the skin. Such devices include transdermal patches, implants, and injections which release the substances in such a manner as to contact the skin or hair follicle either by passive and/or active release mechanisms. The compositions used to deliver the peptides in the methods described herein can be in the form of an aerosol, emulsion, liquid, lotion, solution, gel, micro-encapsulation, cream, paste, ointment, powder, foam, or other pharmaceutically-acceptable formulation. The delivery methods may also include nanotechnology or other advanced technologies to facilitate the penetration and delivery of peptides to desired areas or organs. Furthermore, the peptides can be delivered using less involved formulations such as deionized/distilled water, PBS, or standard medical saline solutions.

[0040] The formulation may optionally have cosmetic appeal, and/or contain other agents such as retinoids, vitamin C, vitamin E or other peptides that can act as adjuvants for the therapeutic action of the inventive peptides. Antibiotics can also be added to the formulation in order to ward off infection, thereby permitting maximal healing processes to occur.

[0041] The formulation may contain protease inhibitors. A protease inhibitor can be selected to specifically target proteases that would be expected to degrade the selected bioactive peptide; such a selection would be determined based on the length and/or sequence of the bioactive peptide. However, protease inhibitors need not necessarily be selected in any specific manner; for example, in some embodiments a protease inhibitor cocktail, which contains two or more inhibitors, can be employed in the formulation. The following types of protease inhibitors can be incorporated in the formulation: serine protease inhibitors, cysteine protease inhibitors, aspartate protease inhibitors, metalloproteinase inhibitors, thiol protease inhibitors, and/or threonine protease inhibitors. The protease inhibitor used in formulation may be a peptide, protein, or chemical. Non-limiting examples of such inhibitors are the serpins, which include alpha-1-antitrypsin, complement 1 -inhibitor, antithrombin, alpha-1 - antichymotrypsin, plasminogen activator inhibitor 1 , and neuroserpin, or chemicals including, but not limited to, ursolic acid and tranexamic acid that can act as adjuvant for the therapeutic action of the inventive peptides.

[0042] In some embodiments, the peptides disclosed herein are incorporated into an aqueous solution. In some embodiments, the peptides are incorporated into a cosmetically or pharmaceutically acceptable solvent. The solubility of a peptide can be readily determined based on the charged or hydrophobic nature of the amino acid sequence and any modification at the N-terminal or C-terminal ends. Generally, a pharmaceutically acceptable formulation according to embodiments includes any carrier suitable for use in or on humans. Such pharmaceutically or cosmetically acceptable carriers include, but are not limited to, ethanol, propanol, isopropanol, propylene glycol, butylene glycol, polyethylene glycol, dimethyl sulfoxide, glycerol, silica, alumina, starch, or any combination thereof. In some embodiments, the pharmaceutically acceptable carrier or cosmetically acceptable carrier does not consist of water or does not consist essentially of water.

[0043] In some embodiments, disclosed herein are methods of treating a condition, disease or disorder in a subject comprising administering to the subject an effective amount of at least one of the peptides of this disclosure. "Treatment," "treating" or "ameliorating" refers to medical management of a condition, disease, or disorder of a subject (e.g., patient), which may be therapeutic,

prophylactic/preventative, or a combination treatment thereof. An "effective amount" or "therapeutically effective amount" of a peptide or composition described herein refers to that amount of peptide or composition sufficient to result in amelioration of one or more symptoms of the condition, disease, or disorder being treated in a statistically significant manner. In some embodiments, the condition, disease, or disorder is associated with skin aging. For example, the method comprises administering at least one peptide disclosed herein to reduce or reverse the effects or appearance of aging with regard to skin (e.g. , anti-aging). In some embodiments, the condition, disease, or disorder is a chronic inflammatory conditions including, but not limited to, diabetes, metabolic syndrome, diabetic skin chronic wounds, Alzheimer's disease, atherosclerosis, osteoporosis, osteoarthritis, cancer therapy, pollution associated assaults, body and skin detoxification, renal failure, diabetic retinopathy and glaucoma, periodontal disease, cystic fibrosis related diabetes, polycystic ovary syndrome, psoriasis associated condition, and oxidative stress, complications associated with maternal chorioamnionitis or funisitis, sarcopenia, among others.

[0044] In some embodiments, the peptides disclosed herein are useful for the treatment or amelioration of aging and symptoms associated with aging. Examples of symptoms of aging include muscle weakness (e.g., sarcopenia), atherosclerosis, walking disability, Alzheimer's disease, and skin aging, such as drying, thinning, spots, decreased elasticity, increased stiffening, and wrinkles.

[0045] In some embodiments, the peptides disclosed herein are useful for body and skin detoxification. A skin detoxification means to allow a sufficient amount of time after direct contact of the peptides with skin to inhibit the interaction of sugar with skin cell components e.g., proteins, DNA, RNA and lipids to form AGEs, or inhibit the interaction of AGEs with its receptors to further damaging skin cells. Therefore, skin detoxification can result in a statistically significant reduction in the appearance of dry skin, rash, dull looking skin, sagging skin, wrinkles, fine lines, dark spots, uneven tone, or other inflammatory effects due to the toxic and inflammatory effects of AGEs compared to a control subject that did not receive treatment with the peptide disclosed herein. In some embodiments, a sufficient amount of time of direct contact with the skin includes, for example, 1-24 hours, 1-2 days, 3-4 days, 5-6 days, 1 week, 2 weeks, 3 weeks, 4 weeks, 2 months, 3 months, 4 months, 5 months, 6 months, 7 months, 8 months, 9 months, 10 months, 1 1 months, 12 months, or more. Similarly, taking peptides orally or systemically inhibits the formation of AGEs or the interaction of AGEs with its receptors, therefore reducing toxic and inflammatory effects of AGEs and one or more associated symptoms, e.g., dry skin, rash, dull looking skin, sagging skin, wrinkles, fine lines, dark spots, uneven tone, or other inflammatory effects.

[0046] In some embodiments, the peptides disclosed herein are useful for the treatment or amelioration of oxidative stress and symptoms associated with oxidative stress. Examples of oxidative stress and symptoms of oxidative stress include inflammation (e.g., osteoporosis, osteoarthritis, arthritis, psoriasis), cancer (e.g., prostate cancer, breast cancer, gastric cancer, lung cancer), cancer therapy associated condition, renal failure. [0047] In some embodiments, the peptides disclosed herein are useful for the treatment or amelioration of complications associated with or caused by diabetes. Examples of diabetic complications include metabolic syndrome, hyper-coagulation of the blood, diabetic skin chronic wounds, skin ulcer, osteoporosis, periodontitis, cystic fibrosis related diabetes, diabetic retinopathy, diabetic nephropathy and glaucoma.

[0048] In some embodiments, the peptides disclosed herein are useful for the treatment or amelioration of symptoms associated with exposure to pollution, such as indoor and outdoor air pollution. Examples of pollution include carbonyl compounds (e.g., formaldehyde, acetaldehyde, hexanal, glyoxal, and methylglyoxal), oxygenated organic compounds (e.g, glyoxal, methylglyoxal, glycolaldehyde, and diacetyl). Exemplary symptoms of exposure to pollution include headache; eye, nose, or throat irritation; dry cough; dry, irritated, or itchy skin; dizziness and nausea; difficulty in concentrating; fatigue; and sensitivity to odors.

[0049] The peptides disclosed herein may be used in a cosmetic or

pharmaceutical composition in effective concentrations to achieve the desired effect. In some embodiments, the concentration of the peptide in the composition can be between 0.1 parts per million (ppm) and 500 ppm or between 0.1 ppm and

100,000ppm. However, the ultimate concentration employed may vary outside these ranges, depending on the nature of the wound/tissue condition, the bio-activity of the inventive peptide, and/or the use of any adjuvant or technique to obtain enhanced composition absorption. In various embodiments, the concentration of the peptide in the composition is between 0.1 ppm and 100,000ppm; between 1 ppm and 50,000ppm; between 1 ppm and 10,000ppm; between 10 ppm and 5000 ppm; between 10ppm and 100,000 ppm. In various embodiments, the concentration of the peptide in the composition is at least about 0.1 ppm, at least about 0.5 ppm, at least about 1 ppm, at least about 5 ppm, at least about 10 ppm, at least about 50 ppm, at least about 100 ppm, at least about 500 ppm, at least about 1 ,000 ppm, at least about 5,000 ppm, at least about 10,000 ppm, or at least about 50,000 ppm. In various embodiments, the concentration of the peptide in the composition is no more than about 0.5 ppm, no more than about 1 ppm, no more than about 5 ppm, no more than about 10 ppm, no more than about 50 ppm, no more than about 100 ppm, no more than about 500 ppm, no more than about 1 ,000 ppm, no more than about 5,000 ppm, no more than about 10,000 ppm, no more than about 50,000 ppm, or no more than about 100,000ppm.

[0050] In some embodiments, the peptides disclosed herein may be prepared as a premix peptide formulation. A premix peptide formulation refers to a mixture containing at least one active peptide of the current disclosure plus at least one other ingredient disclosed herein, such as water, glycerin, a stabilizer, an oil, an emollient, an emulsifier, a humectant, a thickener, a neutralizer, a surfactant, a preservative, or any combination thereof. A premix may exist in the form of solid (powder), semi-solid (emulsion), or solution. It may be made by agitation, stirring, or mixing under high shear force. A premix is not a final formulation or product for use with a subject.

Instead, it can be mixed with different base formulations to generate various final products.

[0051] The peptides disclosed herein may be stabilized or are stable in a premix. As used herein "stabilized" refers to the absence of or reduction of loss of activity over a period of time in a statistically significant manner compared to non-stabilized control peptide. A peptide is "stable" in a premix (or another formulation) if no more than 50% of the peptide is degraded for at least 1 month. In some embodiments, any of the peptides disclosed herein in a premix may be stabilized or stable for at least 6 months, at least 12 months, at least 18 months, at least 24 months, at least 30 months, at least 36 months, at least 42 months, at least 48 months, at least 54 months, or at least 60 months.

[0052] The concentration of a peptide in a premix formulation is higher than the final formulation for use by a subject. In some embodiments, a premix formulation may comprise any of the peptide disclose herein at a concentration between 1 ppm and 100,000 ppm, such as between 1 ppm to 100 ppm, 100 ppm to 500 ppm, 500 ppm to 1 ,000 ppm, 1 ,000 ppm to 5,000 ppm, 5,000 ppm to 10,000 ppm, 10,000 ppm to 50,000 ppm, and 50,000 ppm to 100,000 ppm.

[0053] The CTFA Cosmetic Ingredient Handbook, Second Edition (1992) or the CTFA International Cosmetic Ingredient Dictionary & Handbook (13th Ed. 2010)

(published by the Cosmetic, Toiletry, and Fragrance Combination, Inc., Washington, D.C.) describes a wide variety of non-limiting cosmetic and pharmaceutical ingredients commonly used in the skin care industry, which are suitable for use in the compositions disclosed herein. Examples of these ingredient classes include: abrasives, absorbents, aesthetic components such as fragrances, pigments, colorings/colorants, essential oils, skin sensates, astringents, etc. (e.g. clove oil, menthol, camphor, eucalyptus oil, eugenol, menthyl lactate, witch hazel distillate), anti-acne agents, anti- caking agents, antifoaming agents, antimicrobial agents (e.g., iodopropyl

butylcarbamate), antioxidants, binders, biological additives, buffering agents, bulking agents, chelating agents, chemical additives, cosmetic biocides, denaturants, drug astringents, external analgesics, film formers or materials, opacifying agents, pH adjusters, propellants, reducing agents, sequestrants, skin bleaching and lightening agents (e.g. hydroquinone, kojic acid, ascorbic acid, magnesium ascorbyl phosphate, ascorbyl glucosamine), skin-conditioning agents (e.g. humectants), skin soothing and/or healing agents (e.g. panthenol and its derivatives, aloe vera, pantothenic acid and its derivatives, allantoin, bisabolol, and dipotassium glycyrrhizinate), skin treating agents, thickeners, and vitamins, and derivatives thereof.

[0054] In certain embodiments, the peptides are formulated in a lotion or cream that includes both water soluble ingredients and oil soluble ingredients. These water soluble and oil soluble ingredients may comprise (but not be limited to) one or more moisturizing components, occlusive agents, humectants, emollients, emulsifiers, thickeners, diluents, fragrances, preservatives, and neutralizers.

[0055] More particularly, the peptides may be formulated in a lotion or cream that may include one or more occlusive agents which creates a barrier that blocks water from escaping the skin. Such occlusive agents may be present at a percentage of final volume of about 0.1 %-10%, about 0.5%-5%, about 0.001 %-1 %, about 1 %-10%, about 10%-50%, or about 50%-99%and may include but are not limited to petrolatum, mineral oil, dimethicone, isopropyl myristate, or any combination thereof. One or more humectants may also be included in order to attract water to the skin. Such humectants may be present at a percentage of final volume of 0.1 %-10%, about 0.5%-5%, about 1 %-5%, about 0.001 about 1 %-10%, about 10%-50%, or about 50%-99% and include but are not limited to glycerin, propylene glycol, PEG, silicones, urea, pyrrolidone carboxylic acid and salts thereof, aloe, honey, hyaluronic acid, or any combination thereof. One or more emollients may also be added to improve the feel of the lotion on the skin and/or reduce the tackiness and greasiness caused by the other moisturizing ingredients. Such emollients may be present at a percentage of final volume of about 0.1 %-10%, about 0.5%-5%, about 1 %-5%, about 0.001 %-1 %, about 1 %-10%, about 10%-50%, or about 50%-99%, and may include, but are not limited to, plant oils such as coconut oil, natural butters, cetyl esters, Argan oil, certain silicones, mineral oil, petrolatum, fatty acids, squalane , polymers, lenitin or any combination thereof. In certain embodiments, the lotion also contains emulsifiers to help combine the oil soluble ingredient with the water soluble ingredients. Such emulsifiers may be present at a percentage of final volume of about 0.1 %-10%, about 0.5%-5%, or about 1 %-5%, about 0.001-1 %, about 1 %-10%, about 10%-50%, or about 50%-99% and include but are not limited to lecithin, polyglyceryl oleate, glyceryl stearate, glyceryl oleate, sorbitan stearate, sorbitan oleate, laureth-3, PEG-8 beeswax, glycol distearate, stearic acid, polysorbates, creammaker^®ceteareth-20, gelmaker^®, gum Arabic, prehydrated, PEG, polysorbate stearic acid, cetearyl alcohol, cetyl phosphate, ceteareth, glyceryl stearate citrate; shea butter glycerides, creammaker®-wax, myristic acid, brassica glycerides and brassica alcohol, or any combination thereof. Other ingredients such as thickeners may also be provided to help keep the formula stable and make it more appealing to use. Such thickeners may be present at a percentage of final volume of about 0.1 %-20%, about 1 %-10%, or about 3%-7%, about 0.001-1 %, about 1 %-10%, about 10%-50%, or about 50%-99% and include but are not limited to carbomer, natural wax, stearyl palmitate, or any combination thereof. Other components such as one or more diluents at a percentage of final volume of about 50%-95%, about 60%-90%, about 70%-85%, 0.001 %-1 %, 1 %-10%, 10%-50%, 50%- 99% and include but not be limited to water, botanical extracts, or any combination thereof. Fragrances or preservatives may be present at a percentage of final volume of about 0.01 %-3%, about 0.05%-2%, or about 0.1 %-1.5%, 0.001 %-1 %, 1 %-10%, 10%- 50%, 50-99% and include but not limited to methylparaben, propylparaben, benzylalcohol-DHA, phenoxyethanol disodium EDTA, or any combination thereof. Neutralizers may be present at a percentage of final volume of about 0.05%-5%, about 0.1 %-3%, or about 0.2%-1.5%, 0.001 %-1 %, 1 %-10%, 10%-50%, 50%-99% and include but not limited to triethanolamine. Colorants may also be included to make a well-rounded formulation.

[0056] An example of a cream or lotion comprises the following phase ingredient quantity (%), A: glyceryl stearate 6, cetearyl alcohol 2.0, stearic acid 2.0, ethylhexyl stearate 3.0, caprylic/capric triglyceride 3.0, cetyl dimethicone 2.5, dimethicone 1.5, B: glycerin 5.5, Deionized Water 57.70, C: active peptide premix 2.5, urea 2.0, sodium bisulfate 39% 0.3, sodium ascorbyl phosphate 1.5, Z: preservative/perfume 0.5. Heat phase A and B separately to approx 70-75°C. Add phase A to phase B with stirring. Homogenize, then cool with gentle stirring below 40°C and add phase C, then Z.

[0057] In some embodiments, the peptides disclosed herein are combined or formulated with, for example, Palmitoyl hexapeptide-14, Heptapeptide-7, Hexapeptide- 21 , Oligopeptide-10, SD alcohol 40-B, dimethicone, cyclopentasiloxane, glycerin, neopentyl glycol diheptanoate, hydroxyethyl acrylate/sodium acryloyldimethyltaurate copolymer, polysilicone-1 1 , isohexadecane, pheyl trimethicone, sodium hydroxide, caprylyl glycol, hexylene glycol, polysorbate-20, polysorbate-60, ethylhexyglycerine, phenoxyehtanol, polyethylene, cocamidopropyl betaine, sorbitol, Peg-100 sterate, glyceryle sterate, glycol sterate, calendula officinalis extract, lecithin, tetrasodium EDTA, hydroxyethlcellulose, carbomer, magnesium aluminum silicate, triethanolamine, phenoxyethanol, sodium hyaluronate, algae extract, Artemisia vulgaris (Mugwort) extract, Aleo Barbadensis Leaf Juice Powder, cyclopentasiloxane, ubiquinone, Disodium EDTA, ammonium lauryl sulfate, lauramidoproply betaine, cetyl betaine, lauramide DEA, Melaleuca alternifolia (tea tree) leaf oil, Cucumis sativus (cucumber) fruit extract, Salvia officinalis (sage) leaf extract, parfum, tocopherol, ascorbic acid, citric acid, malic acid, salicylic acid, methylchloroisothiaxolinone, methylisothiazolinone, methylparaben, proplyparaben, caramel, butylene glycol, sodium PCA, ascorbyl glucoside, Chamomilla recutita (matricaria) flower extract, Camellia oleifera (green tea) leaf extract, Hamamelis virginiana (Witch Hazel) extract, propylene glycol, phenoxyethanol, caprylyl glycol, Carthamus tinctorius (Safflower) seed oil, papain powder, Ananas sativus (pineapple) fruit powder, Saponaria officinalis (Soapwort) leaf extract, Carica papaya fruit extract, tocopheryl acetate, Helainthus annuus (sunflower) seed oil, Glycyrrhiza glabra (licorice) root extract, Butyrospermum parkii (shea butter), Olea europaea (olive) fruit oil, Theobroma cacao (cocoa) seed butter, Limnanthes alba (meadowfoam) seed oil, Calendula Officinalis (Marigold) Extract, Jojoba esters, simmondsia chinensis (Jojoba) seed oil, trehalose, Palmaria palmate, Thermus ferment, squalane, Vitamin A, Vitamin, C, Vitamin E, evening primrose oil, carrot, avocado oil, seabuckthorn, soybean oil, green tea, copper aminoacetylamino imidazolyl propanoate (copper peptide), retinyl palmitate, Echinacea purpurea (coneflower) extract, ceramide-2, hydrolyzed soybean fiber, Centella asiatica (gotu kola) extract, Vitis vinifera (grape) seed extract, pichia/resveratrol ferment extract, bisbolol, Malic acid, mandelic acid, Peg-8/SMDI copolymer, carosine, Swiss apple stem cell extract, Swiss alpine rose stem cell extract, grape stem cell extract, Cannabis Sativa Seed Oil, Hexanoyl Dipeptide-3 Norleucine Acetate, Pentapeptide-48, Pentapeptide-59, sunflower sprout extract, Tara (Caesalpinia spinose) extract, PerfectionPeptide P7, carnosine, silymarin, or any combination thereof.

[0058] In some embodiments, the peptides disclosed herein are incorporated into cosmetic delivery systems, pharmaceutical delivery systems, or sustained release systems. Such systems are related to a delivery system of a compound that provides either a targeted release of this compound e.g., delivery to intestine avoiding stomach enzymes or gradual release of this compound during a period of time and preferably, although not necessarily, with relatively constant compound release levels over a period of time. Examples include, but not limited to, liposomes, mixed liposomes, oleosomes, niosomes, ethosomes, milliparticles, microparticules, nanoparticles and solid nanoparticules, nanostructured lipid carriers, sponges, cyclodextrins, vesicles, micelles, mixed micelles of surfactants, surfactant-phospholipiod mixed micelles, millispheres, microspheres and nanosperes, lipospheres, millicapsules, microcapsules and nanocapsules, or with the help of cell penetrating peptides to achieve a greater penetration of the active ingredient and/or improve its pharcokinetic and

pharmacodynamic properties. The inventive peptides and associated compositions may be administered to subjects such as humans and animals, including mammals. Application may also be made in combination with conventional and/or experimental materials such as tissue grafts, skin substitutes, tissue culture products and dressings. The peptides disclosed herein can also be adsorbed on solid organic polymers or solid mineral supports. The compositions which contain the peptides can also be

incorporated into fabrics, non-woven fabrics and medical devices which are in direct contact with the skin. Examples include, but are not limited to, gauzes (woven and non- woven, impregnated, nonadherent, packing, debriding); compression bandages and system; wound fillers and cleansers; contact layers; collagens; amniotic membranes; acellular human dermis; acellular matrices and combination products; and various commonly used dressings, including those listed below. Furthermore, the cosmetic or pharmaceutical compositions can be applied to local areas to be treated by means of iontophoresis, sonophoresis, electroporation, microelectric pateches, mechanical pressure, osmotic gradient, occlusive cure, microinjections or needle-free injections by means of pressure, such as injection by oxygen pressure, or any combination thereof.

[0059] Furthermore, the cosmetic or pharmaceutical compositions containing the peptides disclosed herein, their stereoisomers or their cosmetically or pharmaceutically acceptable salts can be used in different types of formulations for oral administration, preferably in the form of oral cosmetics and pharmaceutical drugs, such as, but not limited to, capsules, tablets, powders, granules, chewing gum, solutions, suspensions, syrups, polysaccharide films, jellies or gelatins, and any other form known by the person skilled in the art. In addition, the peptides disclosed herein can be incorporated into any forms of functional food, fortified food or nutritional supplements e.g., dietary bars or powders. The powders can be dissolved in water, juices, soda, dairy products, or soya derivatives. Thus, in various embodiments, are methods of treatment which include application of one or more of the disclosed peptides in a pharmaceutically- acceptable carrier composition and may be applied to treat one or more condition, including those disclosed herein. [0060] List of commonly used dressings

[0061] In general, the composition can be administered topically, orally, or parenterally including nasal, rectal, vaginal, ocular or subcutaneous implantation or injection directly into a specific body part, transdermally, systemically, or by any other method known to those of skill in the art to be useful to deliver the inventive peptides to the target tissue. Compositions may also be applied in an in vitro or ex vivo manner, either to cells or patient grafts growing in culture, for example.

[0062] The compositions disclosed herein may contain one or more additional agents that exert skin care activity. Besides the bioactive peptide component, the compositions disclosed herein can contain other active agents such as hyaluronic acid, niacinamide, phytantriol, farnesol, bisabolol, salicylic acid, retinol, retinoic acid, alphahydroxy acids, ascorbic acid and alguronic acid. It is expected that certain additional active agents will act synergistically with the bioactive peptide component, and/or will enhance the shelf-life of the formulation.

[0063] Further, the abbreviations for the amino acids follow conventional usage:

[0064] Details on techniques for formulation and administration of

pharmaceuticals may be found in the latest edition of Remington's Pharmaceutical Sciences (Mack Publishing Co, Easton Pa.). Although local topical delivery is desirable, there are other means of delivery, for example: oral, parenteral, aerosol, intramuscular, intraocular, intravenous via dialysis fluid, subcutaneous,

transcutaneous, intramedullary, intrathecal, intraventricular, intravenous,

intraperitoneal, or intranasal administration. The peptides disclosed herein can be formulated in a number of carrier vehicles, for example, in a spray; an aerosol; a water and an oil-type emulsion; an oil and water-type emulsion; a face cream or body cream; a sun lotion or after-sun lotion; or other topical administration vehicle. Additionally, the peptides, and compositions containing them, may provide useful features for inclusion in general skin care and cosmetic formulations, such as various skin cosmetics, skin creams, lotions, sunscreens, and therapeutic lotions or creams. [0065] Areas for application

[0066] AGEs are toxic by-products of metabolism and are also acquired from diet, especially from high-temperature processed food. They promote oxidative damage to proteins, lipids and nucleotides. AGEs are strongly associated with chronic illnesses which account for about two-thirds of all premature deaths and 75% of all medical costs in the United States. AGE modified proteins are signaling molecules associated with several vascular and neurological complications. AGEs proved to be a marker of negative outcome that contribute to the pathophysiology of aging and long term complications of diabetes, atherosclerosis, osteoporosis, Alzheimer's disease, renal failure, and other chronic inflammatory conditions.

[0067] Diabetes is characterized by a high blood glucose level. The chronic hyperglycemia increases basal rates of nonenzymatic glycation and higher serum AGE levels were detected in type 2 diabetic patients which are about 50% greater than that of healthy age-matched controls. The subsequent loss of function of plasma proteins due to AGEs directly contributes to endothelial injury through irreversible glycation of collagen and other subendothelial structural proteins of the blood vessel. AGE formation in ECM also interferes with matrix-cell interactions, with alterations in signaling and adhesion, which may be an important initial event in diabetic

microangiopathy.

[0068] AGE deposits have been found in atherosclerotic plaques and within myocardium fibers and associated with aortic stiffness and correlated with disease severity. The cardiovascular complications are the leading cause of mortality in patients with diabetes mellitus. Plasma AGEs such as CML and pentosidine levels has been shown to be an independent predictor of both re-hospitalization and mortality in heart failure patients. Hyperglycemia induces the formation of AGE-modified low- density lipoprotein (LDL), resulting in a reduction in its plasma clearance therefore contributing to atherosclerosis.

[0069] In addition, diabetics are known to have a hypercoagulable phenotype also called hypercoagulable state. The blood coagulation system is a tightly regulated balance of procoagulant and anticoagulant factors, disruption of which can cause clinical complications. Studies in both humans and mice have revealed that hyperglycemia leads to reduced activity of the anticoagulant plasma protein antithrombin III, and methylglyoxal has been directly linked to such activity inhibition. Other sources of dicarbonyls are in sugar-containing foods and beverages such as bread, coffee, honey, wine, and beer etc. These dicarbonyls including methylglyoxal are major precursors in the formation of intracellular AGEs in endothelial cells, triggers carbonyl stress and activates a series of inflammatory responses leading to

accelerated vascular damage.

[0070] The accumulation of increased levels of AGEs and free methylglyoxal has been demonstrated in diabetic skin, which is responsible for the formation of crosslinks of matrix proteins. Diabetic skin ulcer is a major and increasing public health concern worldwide. It usually causes substantial morbidity, impairs quality of life, and results in high treatment costs for patients. In diabetic skin ulcer, AGEs and dicarbonyl agents e.g., methylglyoxal, can cause resident cells to undergo phenotypic changes and impair the capacity of cell proliferation and movement. Both AGEs and free methylglyoxal contribute to delayed wound healing, which is associated with oxidative stress, keratinocyte injury, dysfunction and apoptosis, and defects in cell adhesion and endothelial dysfunction.

[0071] Hyperglycemia causes nephropathy through the formation of AGEs.

Several studies have demonstrated that AGEs damage the kidneys through AGE- RAGE interaction, deposition of AGEs, or by in situ glycation. AGEs directly affect the structural integrity of the renal tissue through the cross-linking of matrix proteins (collagen). An inverse correlation has been evident between renal function and serum levels of AGEs. Chronic kidney disease and AGEs participate in what has been termed a "vicious cycle", as this condition is associated with increased oxidative stress and the production of reactive carbonyl compounds and AGEs. The ability of AGE to cross-link with ECM exacerbates glomerulosclerosis. Activation of RAGE by circulating AGEs on mesangial cells, tubules, and podocytes all increase the production of intracellular reactive oxygen species and upregulation of transcription factor of nuclear factor kappa B, which further increase the production of some growth factors and inflammatory cytokines that contribute to the further decline of renal function.

[0072] Alzheimer's disease (AD) is the most common dementia disorder of later life. AGEs have long been considered as potent molecules promoting neuronal cell death and contributing to neurodegenerative disorders. In fact, one study suggests that AGEs co-localize with Αβ in microglial cells in the brains of Alzheimer's individuals and a marked increase in AGE accumulation is observed at later stages of the disease, which is associated with the formation of insoluble deposits such as amyloid plaques and neurofibrillary tangles. AGEs are also known to activate glia, resulting in inflammation and neuronal dysfunction. The interaction between AGEs and RAGE elicits intracellular oxidative stress, proinflammatory response and apoptosis all of which contribute to nerve cell damage. The brain has high energy requirements and glucose is the main energy substrate for the brain. Glucose utilization and glycolysis generate dicarbonyl by-products e.g., glyoxal, methyloglyxoal etc. As previously mentioned, methyloglyoxal is one of the most potent glycating agents present in cells, making its accumulation highly deleterious. In humans the cerebrospinal fluid of AD patients shows high levels of methylogyoxal. Direct comparison of toxicity in primary cultures of mouse cortical neurons and astrocytes demonstrated that neurons are 6- fold more susceptible toward methylglyoxal toxicity than astrocytes.

[0073] Both type I and type II diabetes are known to compromise bone microstructure. Hyperglycemia and insulin resistance in type II diabetes induce osteoblast apoptosis and uncoupling of bone turnover. Advanced alterations of bone proteins by glycation are clearly detectable in osteoporotic bone. In bone, AGE adducts form predominantly on the long-living and abundant matrix protein collagen type I (COL I). The intermolecular cross-linking and side-chain modifications in COL I reduce the solubility and flexibility and increase the stiffness of the protein, thereby contributing to skeletal fragility. Elevated serum AGEs have been connected to poor grip strength in older community-dwelling women. AGEs also impair collagen's ability to dissipate energy. Mechanical testing of in vitro glycated human bone specimens revealed that AGE accumulation dramatically reduces the capacity of organic and mineralized matrix to creep and causes bone to fracture under impact at low levels of strain typically associated with fall. The study suggests that patients with osteoporosis had significantly higher serum levels of pentosidine and CML than the non-osteoporotic age matched controls, and AGE modified proteins may be a cause of disturbed bone remodeling in osteoporosis. The adhesion of osteoblasts to bone extracellular matrix can modulate diverse aspects of their physiology such as growth, differentiation and mineralization, but study has shown that AGE modification of collagen impairs the integrin-mediated adhesion of osteoblastic cells to the matrix therefore contributing to the pathogenesis of diabetic osteopenia.

[0074] AGEs have been implicated in vision loss associated with macular degeneration, cataract formation, diabetic retinopathy, and glaucoma. The

accumulation of AGEs has been detected at the site of the corneal epithelium and epithelial basement membrane in diabetic rats, monkeys, and humans. It was shown that AGEs were elevated in tears of diabetic patients. It was also evident that AGEs in ocular tissues mediate aberrant crosslinking of extracellular matrix proteins and disruption of endothelial junctional complexes that affect cell permeability and mediate angiogenesis and breakdown of the inner blood-retinal barrier. AGEs also severely affect cellular metabolism by disrupting ATP production, enhancing oxidative stress, and modulating gene expression of anti-angiogenic and anti-inflammatory genes. Treatment with aminoguanidine, an inhibitor of AGEs, prevented corneal structural abnormalities in diabetic rats.

[0075] As mentioned previously, apoptosis is a potential mechanism by which AGEs may exert effects. AGEs induce apoptosis in retinal pericytes and corneal epithelium, and increases in corneal epithelial cell apoptosis contributes to delayed epithelial wound healing in diabetic cornea. Senile cataracts are associated with progressive oxidation, fragmentation, cross-linking, insolubilization, and yellow pigmentation of lens crystallins. Both glucose and ascorbic acid metabolism generate high levels of dicarbonyl compounds that participate in the glycation process of lens proteins. A large spectrum of AGEs and precursors, e.g. methyloglyoxal, glyoxal, the Amadori products of glucose and lysine, and ascorbic acid, has been quantitated in the human lens. Therefore, modifications of AGEs predispose lens crystallins toward aggregation and cataractogenesis.

[0076] The lack of metabolic control has been implicated in the high susceptibility of diabetic patients to periodontal disease. Periodontitis is a globally prevalent inflammatory disease which causes the destruction of the periodontal structures (i.e. alveolar bone, periodontal ligament and root cementum) and potentially leads to tooth loss. Type II diabetes patients have a 2- to 5-fold higher risk for periodontal diseases. The chronic accumulation of AGEs in diabetes provides a potent and sustained stimulus to activation of the receptor RAGE and of tissue-destructive inflammatory cascades in diabetic periodontium. AGEs/RAGE interaction and activation are the primary concern in the development and progression of periodontitis. Enhanced RAGE expression in the periodontium of an individual with diabetes mellitus leads to exaggerated inflammation and impaired repair, which then results in accelerated and severe periodontal destruction. Blockade of RAGE decreases the generation of the proinflammatory cytokines and metalloproteinases in gingival tissue leading to suppression in alveolar bone loss. Recently, oxidative stress and increased level of AGEs have been proposed to be responsible for the malfunctioning of dental implants.

[0077] Diabetes also causes worsening lung function in cystic fibrosis (CF). CF related diabetes (CFRD) prevalence increases with age; with more than 50% of CF patients having diabetes by age 40. CFRD patients have a higher mortality than CF alone and there is a strong association between CFRD and deterioration in lung function and clinical status. The peptides disclosed herein can be formulated and delivered topically to the lung with or without the combination of other existing CF medications and provide an alternative to reduce the inflammatory status of the CFRD lung.

[0078] AGEs and receptor activation have been implicated to be responsible for the ovulation failure that characterizes polycystic ovary syndrome (PCOS). The ovary is the main regulator of female fertility. Changes in maternal health and physiology can disrupt intraovarian homoeostasis thereby compromising oocyte competence and fertility. Both clinical and experimental research support the role of dicarbonyl overload and AGEs as key contributors to perturbations of the ovarian microenvironment leading to lower fertility. In ovarian tissue samples, women with PCOS have increased AGEs and RAGE expression in theca and granulosa cell layers compared with normal women. The accumulation of AGE, pentosidine, and CML in the follicular fluid and AGE in serum negatively correlated with follicular growth, fertilization and embryonic development.

[0079] Maternal infection and inflammation are known risk factors for premature birth. For example, many premature infants are exposed to inflammatory stimuli even prior to birth, with additional inflammatory insults often occurring as a result of the subsequent resuscitation. Accordingly, the peptides disclosed herein can be administered to pregnant women to reduce inflammation associated with AGEs.

[0080] Psoriasis is a chronic inflammatory skin disease with a genetic

predisposition and a very complex aetiopathogenesis. It was found that the

concentrations of AGEs and anti-CML and anti-CEL antibodies were significantly higher in the active phase of psoriasis patients than those in healthy individuals. In addition, patients with psoriasis vulgaris (PV) had significantly increased blood levels of total peroxide concentration (TPX) and methylglyoxal compared to healthy controls. Methylgloxal serum level reflects increased oxidative stress as well as carbonyl stress and is positively correlated with psoriasis area and severity index. Psoriasin was originally identified in psoriasis and its production is induced by reactive oxygen species. Psoriasin increases the expression of ROS and VEGF and acts through RAGE to promote endothelial cell proliferation. Treatments with the peptides disclosed herein are thought to block the interaction of AGEs and RAGE thereby inhibiting psoriasin-induced ROS generation, endothelial cell proliferation, and angiogenesis. [0081] In some embodiments, the peptides disclose herein may be used to treat or ameliorate osteoarthritis. Aging and diabetes are known to be major causes which affect the progression of osteoarthritis (OA). OA is a progressive degenerative joint disease with signs and symptoms of inflammation, including joint pain, swelling, and stiffness leading to significant functional impairment and disability in older adults. Accumulation of AGEs in cartilage chondrocytes shows the decreased proteoglycan and collagen synthesis, which leads to stiffness and brittleness of the articular cartilage. AGEs can also up-regulate the production of MMPs and IL-6 and IL-8 which mediate cartilage degradation, leading to joint destruction. Rheumatoid arthritis is a chronic inflammatory disease of the synovium, a membrane lining the non-weight- bearing surfaces of the joint.

[0082] In some embodiments, the peptides disclose herein may be used to treat or ameliorate loss of muscle mass and strength (sarcopenia). Sarcopenia is a serious problem among older populations. One third of women and half of men older than 60 suffer from sarcopenia in the U.S. The pathogenesis of sarcopenia is multifactorial and may involve hormonal changes, oxidative stress and inflammation, changes in vasculature, and activity levels. It was shown that pentosidine concentrations were 200% higher in a group of older individuals with a mean age of 78 compared with their younger counterparts with a mean age of 25, suggesting therefore that AGEs may contribute to the decline of muscular function observed in aging. It has been confirmed that women with higher concentrations of CML had less grip strength than women with lower CML concentrations in the blood, and that therefore women with higher AGE levels have more severe muscle weakness. Women with elevated serum CML levels are at an increased risk of developing severe walking disability. Older adults have increased cross-linking of collagen, and deposition of AGEs in skeletal muscles and AGEs may also contribute to this condition by increasing oxidative stress and inflammation.

[0083] In some embodiments, the peptides disclose herein may be used to treat or ameliorate the risk of cancer. A molecular link between glycotoxins and cancer biology involves the activation of an inflammatory cycle of persistent oxidative stress that favors cancer development. In vitro studies of breast, prostate, and lung cancer cell lines imply a possible contribution of AGEs to cancer proliferation, migration, invasion, and survival. Studies support a direct link between RAGE activation and proliferation, survival, migration, and invasion of tumor cells. Detection of AGEs e.g., CML and argpyrimidine, has been reported in various human tumors. In addition, a higher accumulation of AGEs has been observed in malignant tissues (such as prostate cancer) compared to benign ones.

[0084] In some embodiments, the peptides disclose herein may be used to treat or ameliorate skin aging. One of the hallmarks of aging skin is the accumulation of AGEs resulted from the overlapping of an intrinsic chronological aging (individual, genetic) and of an extrinsic aging (external factors like UV, pollution, and lifestyle). As such the human skin becomes drier, thinner, spots appear, elasticity decreases and stiffening increases, together with the appearance of wrinkles over time. In skin, glycation is involved in a very complex aging process and simultaneously affects, directly and indirectly, certain cells, their synthesis, and the organization of the matrix. At the cutaneous level, glycation affects cells including endothelial cells, fibroblasts, keratinocytes, and structural proteins such as collagen, elastin, glycoproteins, and glycosaminoglycans. Glycation of collagens has been linked to the development of skin dullness and the decrease in skin elasticity and evidence supports that methylglyoxal induced AGEs deform the elasticity of the collagen molecules and reduce inherent collagen fibril viscoelasticity. AGEs were reported to accumulate in dermal elastin and collagens and such glycation modified dermal ECM further affects growth,

differentiation and motility of fibroblasts, cytokine response, metaloproteinase and vascular hemostasis. Collagen cross-linking by AGEs has been increasingly implicated as a central factor in the onset and progression of connective tissue disease. RAGE is highly expressed in the skin and upregulated by AGEs and TNF-a. In elderly subjects, the accumulation of AGEs causes decreased elasticity and increased stiffening.

Histological data indicate that changes in dermis including structural, biochemical, molecular, architectural, and functional changes start before the age of 35 and increase rapidly due to intrinsic processes and are amplified by UV exposure.

Therefore 35 is the age threshold that represents the best time for the initiation of anti- aging therapy, which may include a combination of products that both stimulate protein synthesis and inhibit the glycation process.

[0085] Over the past two decades, there has been increasing awareness regarding the potential impact of both indoor and outdoor air pollution on health.

Carbonyl compounds not only participate in glycation endogenously but are also present in high levels in pollution. It has been reported that formaldehyde,

acetaldehyde, hexanal, glyoxal, and methylglyoxal were the dominant compounds in rainwater of an urban atmosphere. These carbonyl compounds are toxic and also formed in mainstream cigarette smoke. Both glyoxal and methylglyoxal are important elements in the cooking emissions. A pilot hazardous airborne carbonyls study was carried out in Hong Kong and Mainland China. Workplace air samples in 14 factories of various types of manufacturing and industrial operations were collected and analyzed for a panel of 21 carbonyl compounds. The factories can be classified into five general categories, including food processing, electroplating, textile dyeing, chemical manufacturer, and petroleum refinery. The study found that glyoxal and methylglyoxal existed at variable levels in the selected workplaces, ranging from 0.2% to 5.5%.

[0086] It has been assumed that the majority of pollutants enter the body through breathing, however, results of one study suggest that inhalation and skin passage delivered roughly equal amounts of certain pollutants into the blood and urine. Even more interesting, wearing clothing left in a closet or on the back of a chair in a room with polluted air also enhances the uptake of airborne pollutants

(www.skininc.com/skinscience/physiology/Skin-Matches-Lung-Absorption-of- Phthalates-358850721.html). Modern society spends a considerable amount of time indoors, and research findings have demonstrated that some air pollutants occur as frequently in indoor air as in outdoor air and may be associated with adverse health effects. On average, U.S. citizens spend 80% or more of their daily lives indoors whether at home, work, or in other commercial buildings, and over the last two decades there has been an increasing awareness regarding the potential impact of indoor air pollution on health. The term "sick building syndrome" (SBS) has been used to describe situations in which no specific illness or cause, aside from time spent indoors, explains adverse health effects experienced by building occupants. Indoor air quality-related health issues cost businesses billions of dollars annually due to factors including decreases in worker productivity and time off from work. Several oxygenated organic compounds, such as glyoxal, methylglyoxal, glycolaldehyde, and diacetyl, have been identified as possible reaction products of indoor environment chemistry. These compounds are sensitizers. Affected individuals often complain of associated symptoms such as headache; eye, nose, or throat irritation; dry cough; dry or itchy skin; dizziness and nausea; difficulty in concentrating; fatigue; and sensitivity to odors.

[0087] An in silico approach was used to target the region of sRAGE that has been experimentally confirmed to bind AGEs for the design of a series of short peptides from 2 to 9 amino acids in length. Before the current disclosure, it was unknown as to how many binding sites are located in sRAGE; the minimal length of sequence required to bind to AGE; and whether the binding site involves linear sequences or whether a secondary structure is involved in the binding. Furthermore, as shown in the present disclosure, these peptides not only effectively bind to AGEs and prevent AGE-RAGE interactions, but also effectively block dicarbonyl stress such as methylglyoxal-induced cell apoptosis and cell death, thereby blocking the signal transduction pathways that ultimately lead to cellular damage and cell death. Such activity has not previously been reported or disclosed. These peptides are novel as they are generated using an in silico design method which do not exist or occur in nature and the peptides produced by these methods cannot be isolated from microbes, animals, or plants.

[0088] Among other properties, the peptides disclosed herein significantly protect human skin keratinocytes against AGEs and precursors such as methyloglyoxal- induced apoptosis and improve fibroblast survival. These peptides are promising therapeutic agents for neutralizing AGEs, alleviating AGE deposition, and blocking AGE-RAGE interactions and thus can be used for improving medical conditions associated with AGEs. With proper formulation these peptides can be delivered systemically, topically, or via other routes, for the prevention of AGE formation as well as for the treatment, with and/or without the help of current available medical regimes, of AGE-associated complications and diseases. In addition to therapeutic agents, the disclosed peptides effectively inhibit the Maillard reaction, as shown in Table 6, to block sugar and protein from forming AGEs. These peptides can be used as Maillard reaction inhibitors, for example if mixed into foods and beverages containing proteins such as collagen, and therefore it may be possible to suppress the Maillard reaction to prevent the deterioration of the foods and beverages.

[0089] EXAMPLES

[0090] The following non-limiting Examples are intended to be purely illustrative, and show specific experiments that were carried out in accordance with embodiments disclosed herein:

[0091] Example 1. Screening for AGE binding peptides

[0092] For initial screening for AGE-binding peptides, binding to a mixture of various AGEs was conducted using an OxiSelect Advanced Glycation End Product (AGE) ELISA kit (Cat# STA-317) purchased from Cell BioLabs Inc (Atlanta GA). This kit uses a standard AGE-BSA that was prepared by reacting BSA with glycolaldehyde followed by extensive dialysis and column purification. According to the product manual the AGE-BSA contains CML, pentosidine, and other AGE structures. The AGEs are probed with an anti-AGE polyclonal antibody, followed by an HRP- conjugated secondary antibody. The current study design was that first AGE-BSA and a proper amount of peptide were mixed and the mixture was incubated at room temperature from 4-20 hours. Next the mixture was transferred to a 96-well protein binding plate supplied in the kit and incubated at +4°C for overnight. The ELISA assay was processed according to the manufacturer's instructions.

[0093] The principle of the experimental design is such that if a peptide binds to AGE it will compete against anti-AGE antibody for one or more binding sites.

Therefore, competitive binding by peptides is expected to block access of the anti-AGE antibody, which subsequently results in a reduced amount of AGE available for antibody binding. AGE-BSA in the absence of a peptide competing for the AGE binding site, on the other hand, is expected to be fully accessible by the anti-AGE antibody, resulting in maximal reading which is taken as 100% binding/0% reduction. The binding activity of a peptide to AGEs is calculated as percentage reduction, as indicated in the formula below. The cutoff value was chosen to be equal or more than approximately 70-80% of the standard control of carnosine-induced activity and a value of≥30% is considered significant. The results are shown in Table 2. In most cases the peptides outperform the reference ("gold") standard carnosine.

Percentage

reduction 1 x 100%

Table 2. Binding activity of peptide to AGEs.

HB2289 SLFLP-OH 77

HB2222 NSLFL-NH2 54

HB2223 FLPAVG-NH2 47

HB2224 VGIQNEG-NH2 85

HB2225 QNEGIFR-NH2 0

HB2275 SLFL-OH 64

HB2303 SLFL-NH2 75

HB2277 LFLP-OH 35

HB2278 VGIQ-OH 71

HB2302 VGIQ-NH2 80

HB2281 GAPK-OH 55

HB2305 GAPK-NH2 60

HB2282 SPQG-OH 80

HB2298 SPQG-NH2 0

HB2283 LEWK-OH 47

HB2306 LEWK-NH2 83

HB2288 KGAP-OH 58

HB2309 KGAP-NH2 0

HB2291 GGGP-OH 0

HB2295 TGRT-OH 70

HB231 1 TGRT-NH2 87

HB2313 EWKL-OH 77

HB2321 GPWD-OH 53

HB2322 GPWD-NH2 56

HB2276 FLP-OH 0

HB2304 FLP-NH2 63

HB2285 LP-OH 56

HB2307 LP-NH2 0

HB2290 GGP-OH 44

HB2286 FL-OH 76

HB2308 FL-NH2 0

HB2293 GRT-OH 63

HB2310 GRT-NH2 50

HB2296 SPQ-OH 71

HB2297 SPQ-NH2 68

HB2292 SPG-OH 58

HB2294 TGR-OH 63

HB2287 GP-OH 0

HB2314 GP-NH2 0

HB2299 SP-OH 69

HB2300 SP-NH2 0

HB2315 TG-NH2 39

HB2316 TG-OH 70

HB2317 PQ-NH2 72

HB2319 PQ-OH 63 58 HB2320 RT-NH2 81

59 HB2318 RT-OH 62

[0094] The binding of each peptide to individual types of AGEs was tested using specific monoclonal antibodies in an ELISA assay. OxiSelect™ N^£-(carboxylmethyl) lysine ELISA kit (Cell Biolab, Atlanta, GA) was purchased for evaluating binding activity to N^£-(carboxylmethyl) lysine (CML). Standard CML-BSA was diluted to 10ug/ml and mixed with an equal volume of reduced BSA to make a stock solution. The peptide (10C^g/ml) was mixed in the stock solution containing CML-BSA at a final volume of 100 μΙ and incubated at room temperature for 5hr. 100 μΙ of the mixture was transferred to a 96-well protein binding plate together with the standard and controls. The plate was then incubated at 4°C overnight. The next day the plate was subjected to an ELISA assay according to the manufacturer's instructions.

[0095] The specificity of peptide binding to N^£-(carboxyethyl) lysine (CEL) was evaluated using OxiSelect™ N^£-(carboxyethyl) lysine (CEL) ELISA kit (Cell BioLabs Inc., Atlanta, GA). Briefly, standard N^£-(carboxyethyl) lysine-BSA was diluted to 10 μg/ml and mixed with equal volume of 10 μg/ml reduced BSA to make a stock solution. Next, peptide (100 μg/ml) was added to an equal volume of the stock solution containing methylglyoxal-BSA and incubated at room temperature for 5hr. At the end of incubation, 100 μΙ of the reaction mixture was transferred to the 96-well protein binding plate and incubated overnight at +4°C. The next day the plate was subjected to an ELISA assay according to the manufacturer's instructions. Percentage block as reported in Table 3 was calculated as indicated above for Table 2 and values of≥15% and≥24% are considered significant for blocking CEL and CML, respectively. The cutoff value was chosen to be equal to or more than the standard control carnosine- induced activity. ND refers to "not done" and applies to all Tables.

Table 3. Binding activity of peptides towards CEL and/or CML

HB2213 LEWKLNTGR-NH2 0 35

HB2214 TGRTEA-NH2 15 28

HB2215 TEAWKVK-NH2 ND ND

HB2216 AWKVKSPQ-NH2 ND ND

HB2217 SPQGGGP-NH2 8 37

HB2218 GPWDSVR-NH2 11 34

HB2219 PWDSV-NH2 ND ND

HB2220 RVLPNG-NH2 ND ND

HB2221 SLFLP-NH2 44 41

HB2289 SLFLP-OH 24 22

HB2222 NSLFL-NH2 23 35

HB2223 FLPAVG-NH2 ND ND

HB2224 VGIQNEG-NH2 73 32

HB2225 QNEGIFR-NH2 ND 20

HB2275 SLFL-OH 8 18

HB2303 SLFL-NH2 22 38

HB2277 LFLP-OH 0 25

HB2278 VGIQ-OH 14 12

HB2302 VGIQ-NH2 90 0

HB2281 GAPK-OH 24 33

HB2305 GAPK-NH2 20 20

HB2282 SPQG-OH 32 22

HB2298 SPQG-NH2 0 18

HB2283 LEWK-OH 23 23

HB2306 LEWK-NH2 66 33

HB2288 KGAP-OH 0 17

HB2309 KGAP-NH2 0 24

HB2291 GGGP-OH 19 0

HB2295 TGRT-OH 10 30

HB2311 TGRT-NH2 87 0

HB2313 EWKL-OH 36 22

HB2321 GPWD-OH 33 31

HB2322 GPWD-NH2 22 21

HB2276 FLP-OH 25 0

HB2304 FLP-NH2 28 12

HB2285 LP-OH 32 10

HB2307 LP-NH2 29 19

HB2290 GGP-OH 37 2

HB2286 FL-OH 28 11

HB2308 FL-NH2 34 19

HB2293 GRT-OH 29 9

HB2310 GRT-NH2 28 3

HB2296 SPQ-OH 27 5

HB2297 SPQ-NH2 0 24

HB2292 SPG-OH 1 1 13 49 HB2294 TGR-OH 24 19

50 HB2287 GP-OH 0 ND

51 HB2314 GP-NH2 15 2

52 HB2299 SP-OH 31 0

53 HB2300 SP-NH2 9 9

54 HB2315 TG-NH2 17 0

55 HB2316 TG-OH 25 11

56 HB2317 PQ-NH2 47 2

57 HB2319 PQ-OH 0 0

58 HB2320 RT-NH2 28 4

59 HB2318 RT-OH 13 12

[0096] Example 2. Activity of peptide to block methylglyoxal induced keratinocyte apoptosis

[0097] Human skin keratinocytes (ATCC CRL-2404, American Type Culture Collection, Manassas, VA, USA) were grown in serum-free keratinocyte growth media supplemented with 5ng/ml human recombinant epithelial growth factor (EGF) (Life Technologies™, Grand Island, N.Y.). The cells were then trypsinized and seeded into a 96-well plate. The plate was incubated in 37°C, 5% C02 for 8hr to allow attachment of cells to the plate. Methylgloxal was purchased from Sigma-Aldrich (St. Louis, MO). Each peptide (100 μg) was pre-incubated with methylglyoxal (50 μg) for 6hr then transferred to a 96-well plate containing human keratinocytes. Carnosine was used as protective positive control. The cells were further incubated overnight prior to the apoptosis assay. Apoptotic cells release histone-associated DNA-fragments that were quantified using the Cell Death Detection ELISA^P|US kit purchased from Roche (Roche Diagnosis GmbH, Mannheim, Germany). Briefly, at the end of treatment the supernatant was removed and cells were lysed using lysis buffer. The cell lysate was centrifuged then 20μΙ of the supernatant (cytoplasmic fraction) was carefully transferred into the streptavidin-coated plate; ELISA analysis then proceeded according to the manufacturer's instructions. A 20% protection/reduction in apoptosis was considered significant. The cutoff value was chosen to be equal to or more than approximately 70-80% of the standard control carnosine-induced activity. Table 4 shows some peptides outperform the reference ("gold") standard carnosine.

Table 4. Percentage reduction of methylglyoxal induced apoptosis on human skin keratinocytes Percentage (%)

SEQ ID HB No. Sequence (N-C) reduction of methylgloxal NO. induced apoptosis

- methylglyoxal - 0

- Carnosine - 47

1 HB2210 KGAPK-NH2 36

2 HB221 1 PKKPPQ-NH2 ND

3 HB2212 PQRLEWK-NH2 ND

4 HB2213 LEWKLNTGR-NH2 ND

5 HB2214 TGRTEA-NH2 40

6 HB2215 TEAWKVK-NH2 ND

7 HB2216 AWKVKSPQ-NH2 ND

8 HB2217 SPQGGGP-NH2 45

9 HB2218 GPWDSVR-NH2 ND

10 HB2219 PWDSV-NH2 ND

1 1 HB2220 RVLPNG-NH2 ND

12 HB2221 SLFLP-NH2 48

13 HB2289 SLFLP-OH 52

14 HB2222 NSLFL-NH2 ND

15 HB2223 FLPAVG-NH2 ND

16 HB2224 VGIQNEG-NH2 ND

17 HB2225 QNEGIFR-NH2 ND

18 HB2275 SLFL-OH 48

19 HB2303 SLFL-NH2 15

20 HB2277 LFLP-OH 23

21 HB2278 VGIQ-OH 39

22 HB2302 VGIQ-NH2 30

23 HB2281 GAPK-OH 37

24 HB2305 GAPK-NH2 41

25 HB2282 SPQG-OH 50

26 HB2298 SPQG-NH2 51

27 HB2283 LEWK-OH 42

28 HB2306 LEWK-NH2 6

29 HB2288 KGAP-OH 47

30 HB2309 KGAP-NH2 51

31 HB2291 GGGP-OH 25

32 HB2295 TGRT-OH 64

33 HB231 1 TGRT-NH2 56

34 HB2284 EWKL-OH 18

35 HB2321 GPWD-OH 47

36 HB2322 GPWD-NH2 31

37 HB2276 FLP-OH 21

38 HB2304 FLP-NH2 38

39 HB2285 LP-OH 41 40 HB2307 LP-NH2 58

41 HB2290 GGP-OH 40

42 HB2286 FL-OH 15

43 HB2308 FL-NH2 38

44 HB2293 GRT-OH 63

45 HB2310 GRT-NH2 68

46 HB2296 SPQ-OH 0

47 HB2297 SPQ-NH2 38

48 HB2292 SPG-OH 40

49 HB2294 TGR-OH 65

50 HB2287 GP-OH 0

51 HB2314 GP-NH2 0

52 HB2299 SP-OH 39

53 HB2300 SP-NH2 17

54 HB2315 TG-NH2 26

55 HB2317 TG-OH 42

56 HB2316 PQ-NH2 11

57 HB2319 PQ-OH 8

58 HB2320 RT-NH2 61

59 HB2318 RT-OH 14

[0098] Example 3. Protection of human skin fibroblast from methylglyoxal induced cell death

[0099] Human skin fibroblasts (ATCC CRL-7481 , American Type Culture Collection, Manassas, VA, USA) were grown in 96-well plates in Dulbecco's modified Eagle's medium (DMEM; 4 mM L-glutamine, 4.5 g/L glucose) adjusted to contain 1.5 g/L sodium bicarbonate and supplemented with 10% fetal bovine serum (FBS). After the cells reached >95% confluence the culture medium was replaced with fresh medium containing methylglyoxal (40ug/ml) and each peptide at 100ug/ml. The cells were incubated in a 37°C, 5% C02 incubator overnight and cell survival was measured using the XTT assay (from ATCC) according to the manufacturer's instructions.

Results are shown in Table 5. A protection of 10% and above is considered significant. The cut off value was chosen to be equal to or more than approximately 70-80% of the standard control carnosine-induced activity and, as Table 5 shows, some peptides outperform carnosine.

Table 5. Protection of human skin fibroblasts against methylglyoxal induced cell death SEQ ID HB No. Sequence Percentage of protection of

NO. fibroblast survival

- Methylglyoxal - 0

- Carnosine - 13

1 HB2210 KGAPK-NH2 ND

2 HB2211 PKKPPQ-NH2 ND

3 HB2212 PQRLEWK-NH2 ND

4 HB2213 LEWKLNTGR-NH2 ND

5 HB2214 TGRTEA-NH2 ND

6 HB2215 TEAWKVK-NH2 ND

7 HB2216 AWKVKSPQ-NH2 ND

8 HB2217 SPQGGGP-NH2 ND

9 HB2218 GPWDSVR-NH2 ND

10 HB2219 PWDSV-NH2 ND

11 HB2220 RVLPNG-NH2 ND

12 HB2221 SLFLP-NH2 19

13 HB2289 SLFLP-OH 23

14 HB2222 NSLFL-NH2 ND

15 HB2223 FLPAVG-NH2 ND

16 HB2224 VGIQNEG-NH2 ND

17 HB2225 QNEGIFR-NH2 ND

18 HB2275 SLFL-OH 20

19 HB2303 SLFL-NH2 6

20 HB2277 LFLP-OH ND

21 HB2278 VGIQ-OH 41

22 HB2302 VGIQ-NH2 5

23 HB2281 GAPK-OH 33

24 HB2305 GAPK-NH2 5

25 HB2282 SPQG-OH 21

26 HB2298 SPQG-NH2 ND

27 HB2283 LEWK-OH ND

28 HB2306 LEWK-NH2 14

29 HB2288 KGAP-OH 13

30 HB2309 KGAP-NH2 ND

31 HB2291 GGGP-OH ND

32 HB2295 TGRT-OH 36

33 HB231 1 TGRT-NH2 8

34 HB2313 EWKL-OH 19

35 HB2321 GPWD-OH 34

36 HB2322 GPWD-NH2 35

37 HB2276 FLP-OH 4

38 HB2304 FLP-NH2 ND

39 HB2285 LP-OH ND 40 HB2307 LP-NH2 ND

41 HB2290 GGP-OH ND

42 HB2286 FL-OH 35

43 HB2308 FL-NH2 ND

44 HB2293 GRT-OH ND

45 HB2310 GRT-NH2 ND

46 HB2296 SPQ-OH ND

47 HB2297 SPQ-NH2 ND

48 HB2292 SPG-OH 39

49 HB2294 TGR-OH 50

50 HB2287 GP-OH ND

51 HB2314 GP-NH2 ND

52 HB2299 SP-OH ND

53 HB2300 SP-NH2 ND

54 HB2315 TG-NH2 ND

55 HB2316 TG-OH 29

56 HB2317 PQ-NH2 24

57 HB2319 PQ-OH 24

58 HB2320 RT-NH2 38

59 HB2318 RT-OH 27

[00100] Example 4. Activity of peptides to block AGE formation between sugar and collagen

[00101] Calf skin collagen (0.1 % solution in 0.1 M acetic acid) was purchased from Sigma-Aldrich. A 2xglycation stock solution was prepared which contained 3% glucose and 0.22% NaN3. Briefly, 300 μΙ of collagen was mixed in a sterile 1.5ml Eppendorf tube with an equal volume of 2xglycation stock solution. Peptide was then added at final concentration of 2mg/ml. The mixture was incubated at 37°C for 3 month. Next, the mixture was diluted in PBS and the formation of AGE was quantified using the OxiSelect Advanced Glycation End Product (AGE) ELISA kit (Cat# STA-317) purchased from Cell BioLabs (Atlanta GA), and the results are shown in Table 6. Carnosine was used as a positive control which blocked AGE formation between collagen and sugar. Collagen alone in PBS was used as negative control. A value≥25 is considered significant. The cut off value was chosen to be equal to or more than approximately 70-80% of the standard control carnosine-induced activity. Some of the peptides outperform carnosine, as shown in Table 6.

Table 6. Activity of peptides to block sugar and collagen to form AGEs (%) block of sugar and

SEQ ID HB No. Sequence collagen to form AGEs in NO. vitro

- Sugar+collagen - 0

- Collagen alone - -

- carnosine - 35

1 HB2210 KGAPK-NH2 19

2 HB2211 PKKPPQ-NH2 21

3 HB2212 PQRLEWK-NH2 28

4 HB2213 LEWKLNTGR-NH2 19

5 HB2214 TGRTEA-NH2 22

6 HB2215 TEAWKVK-NH2 ND

7 HB2216 AWKVKSPQ-NH2 ND

8 HB2217 SPQGGGP-NH2 25

9 HB2218 GPWDSVR-NH2 36

10 HB2219 PWDSV-NH2 15

1 1 HB2220 RVLPNG-NH2 19

12 HB2221 SLFLP-NH2 45

13 HB2289 SLFLP-OH 44

14 HB2222 NSLFL-NH2 57

15 HB2223 FLPAVG-NH2 15

16 HB2224 VGIQNEG-NH2 15

17 HB2225 QNEGIFR-NH2 ND

18 HB2275 SLFL-OH 17

19 HB2303 SLFL-NH2 20

20 HB2277 LFLP-OH ND

21 HB2278 VGIQ-OH 21

22 HB2302 VGIQ-NH2 1

23 HB2281 GAPK-OH 30

24 HB2305 GAPK-NH2 28

25 HB2282 SPQG-OH 33

26 HB2298 SPQG-NH2 24

27 HB2283 LEWK-OH 23

28 HB2306 LEWK-NH2 15

29 HB2288 KGAP-OH 26

30 HB2309 KGAP-NH2 18

31 HB2291 GGGP-OH ND

32 HB2295 TGRT-OH 15

33 HB231 1 TGRT-NH2 20

34 HB2313 EWKL-OH 36

35 HB2321 GPWD-OH 52

36 HB2322 GPWD-NH2 15

37 HB2276 FLP-OH 0 38 HB2304 FLP-NH2 30

39 HB2285 LP-OH 25

40 HB2307 LP-NH2 35

41 HB2290 GGP-OH 65

42 HB2286 FL-OH 44

43 HB2308 FL-NH2 10

44 HB2293 GRT-OH 16

45 HB2310 GRT-NH2 21

46 HB2296 SPQ-OH 40

47 HB2297 SPQ-NH2 10

48 HB2292 SPG-OH 39

49 HB2294 TGR-OH 29

50 HB2287 GP-OH ND

51 HB2314 GP-NH2 10

52 HB2299 SP-OH 45

53 HB2300 SP-NH2 26

54 HB2315 TG-NH2 10

55 HB2316 TG-OH 30

56 HB2317 PQ-NH2 10

57 HB2319 PQ-OH 10

58 HB2320 RT-NH2 42

59 HB2318 RT-OH 15

[00102] Example 5. Formulation of the peptides in a lotion

A skin lotion is prepared from the following components according to the methods below:

Procedure:

[00103] In a suitable container, the diluent (Item #1) is heated to 75°C. Items #2 through #5 (water soluble ingredients) are then added to the heated diluent. Items #6 through #1 1 (oil soluble ingredients) are separately mixed together. The mixture of oil soluble ingredients are slowly mixed with the mixture of water soluble ingredients and further mixed for thirty (30) minutes. The resulting mixture is then cooled to 25°C and fragrance (Item #12) is added after the temperature drops below 40°C. The pH and viscosity are checked and adjusted to a pH of 5.0-5.5 and a viscosity of 15,000-20,000. The peptides-containing suspension are then added to the resulting lotion.

[00104] Example 6. Formulation of the peptides in a cream

[00105] A skin cream is prepared according to the methods below:

[00106] All percentages are % of final volume unless otherwise indicated. 0.1-2% thickener (such as natural wax or stearyl palmitate) is dispersed in 55-75% diluent (such as water or water mixed with botanical extracts) at 75-80°C while stirring until a homogeneous gel is formed. An oil soluble phase is separately made by mixing 2-6% melted emulsifier (such as CreamMaker blend or a polysorbate) and 10-35% melted emollient (such as one or more plant oil or natural butter). The oil soluble mixture is then combined with the homogenous gel and stirred intensively until an emulsion is formed and then gently mixed while the emulsion is cooling. Sensitive components like 0.1 %-10% active ingredients (peptide, anti-aging ingredients, skin whitening ingredients, antioxidants, etc.), 0.1-1.0% fragrances, and 0.1-1.0% preservatives (such as benzylalcohol-DHA) are added after the mixture is cooled (40-30°C) to keep their properties intact.

[00107] The various embodiments described above can be combined to provide further embodiments. All of the U.S. patents, U.S. patent application publications, U.S. patent applications, foreign patents, foreign patent applications and non-patent publications referred to in this specification and/or listed in the Application Data Sheet are incorporated herein by reference in their entirety except where incorporation of a reference or a portion thereof contradicts with the present disclosure. Aspects of the embodiments can be modified, if necessary to employ concepts of the various patents, applications and publications to provide yet further embodiments.

[00108] These and other changes can be made to the embodiments in light of the above-detailed description. In general, in the following claims, the terms used should not be construed to limit the claims to the specific embodiments disclosed in the specification and the claims, but should be construed to include all possible embodiments along with the full scope of equivalents to which such claims are entitled. Accordingly, the claims are not limited by the disclosure.

Claims

What is claimed is: 1. A peptide for blocking activity of advanced glycation end products (AGEs), wherein the peptide is between 2 and 9 amino acids in length and is a subsequence of SEQ ID NO:61.

2. The peptide of claim 1 , wherein the peptide is selected from the group consisting of SEQ ID NOs: 1 , 4, 5, 8, 9, 12, 13, 14, 15, 16, 18, 19, 20, 21 , 22, 23, 24, 25, 27, 28, 29, 32, 33, 34, 35, 36, 38, 39, 41 , 42, 44, 45, 46, 47, 48, 49, 52, 54, 55, 56, 57, 58, and 59.

3. The peptide of claim 2, wherein the peptide blocks binding of AGEs to proteins and other cell components.

4. The peptide of claim 1 , wherein the peptide is selected from the group consisting of SEQ ID NOs: 1 , 4, 5, 8, 9, 12, 13, 14, 16, 19, 20, 21 , 22, 23, 24, 25, 27, 28, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 49, 51 , 52, 54, 55, 56, and 58.

5. The peptide of claim 4, wherein the peptide blocks binding of at least one of the AGEs selected from N^£-(carboxyethyl) lysine (CEL), and N^£-(carboxymethyl) lysine (CML), pentosidine and other AGE structures.

6. The peptide of claim 1 , wherein the peptide is selected from the group consisting of SEQ ID NOs: 1 , 5, 8, 12, 13, 18, 20, 21 , 22, 23, 24, 25, 26, 27, 29, 30, 31 , 32, 33, 35, 36, 37, 38, 39, 40, 41 , 43, 44, 45, 47, 48, 49, 52, 54, 55, and 58.

7. The peptide of claim 6, wherein the peptide reduces methylglyoxal-induced apoptosis of keratinocytes.

8. The peptide of claim 1 , wherein the peptide is selected from the group consisting of SEQ ID NOs: 12, 13, 18, 21 , 23, 25, 28, 29, 32, 34, 35, 36, 42, 48, 49, 55, 56, 57, 58, and 59.

9. The peptide of claim 8, wherein the peptide reduces methylglyoxal-induced cell death of human skin fibroblasts.

10. The peptide of claim 1 , wherein the peptide is selected from the group consisting of SEQ ID NOs: 3, 8, 9, 12, 13, 14, 23, 24, 25, 29, 34, 35, 38, 39, 40, 41 , 42, 46, 48, 49, 52, 53, 55, and 58.

11. The peptide of claim 10, wherein the peptide blocks AGE formation in a solution of collagen and sugar.

12. The peptide of any one of claims 1-11 , wherein the peptide is in an amidated form.

13. The peptide of any one of claims 1-12, wherein the peptide is in a free acid form.

14. The peptide of any one of claims 1-13, wherein the peptide comprises either or both L- and D-amino acid enantiomers.

15. The peptide of any one of claims 1-14, wherein the peptide is conjugated to a carrier molecule or is lipidated.

16. A composition comprising at least one peptide according to any one of claims 1-15 and a pharmaceutically acceptable carrier.

17. The composition of claim 16, wherein the peptide is present in a concentration ranging from about 0.1 ppm to about 500ppm.

18. The composition of claim 16, wherein the peptide is present in a concentration ranging from about 0.1 ppm to about 100,000 ppm.

19. The composition of claim 16, wherein the composition is in the form of an aerosol, emulsion, nanoemulsion, nanoliposome, liquid, lotion, solution, gel, micro- encapsulation, cream, paste, ointment, powder, or foam, or is incorporated in a device adapted for application to the surface of skin or under skin tissue.

20. The composition of any one of claims 16-19, comprising a plurality of peptides.

21. A method for treating a skin condition in a subject comprising administering to the subject a medicament comprising a pharmaceutically acceptable carrier and an effective amount of at least one peptide according to any one of claims 1-15.

22. The method of claim 21 , wherein administering to the subject a medicament comprises administering the medicament to skin or associated mucosal tissue of the subject.

23. The method of claims 21 or 22, wherein administering to the subject a medicament comprises administering the medicament to the subject topically, orally, transdermal^, or systemically.

24. The method of claim 21 , wherein administering to the subject at least one peptide comprises administering to the subject at least one peptide selected from the group consisting of SEQ ID NOs: 1 , 3, 4, 5, 8, 9, 12, 13, 14, 15, 16, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49, 51 , 52, 53, 54, 55, 56, 57, 58, 59, or any combination thereof.

25. A method for treating a condition, disease, or disorder in a subject comprising administering to the subject an effective amount of at least one peptide according to any one of claims 1-15.

26. The method of claim 25, wherein the at least one peptide is comprised in a medicament comprising a pharmaceutically acceptable carrier.

27. The method of claim 25 or 26, wherein the condition, disease, or disorder is a chronic inflammatory condition.

28. The method of any one of claims 25-27, wherein the condition, disease or disorder is aging, diabetic complications, exposure to pollution, oxidative stress, body or skin detoxification, diabetes, metabolic syndrome, diabetic skin chronic wounds, Alzheimer's disease, atherosclerosis, osteoporosis, osteoarthritis, cancer, cancer therapy associated condition, pollution associated assaults, body and skin detoxification, renal failure, diabetic retinopathy and glaucoma, periodontal disease, cystic fibrosis related diabetes, polycystic ovary syndrome, psoriasis associated condition, oxidative stress, complications associated with maternal chorioamnionitis or funisitis, or sarcopenia.

29. A method of inhibiting formation of AGEs in a food or beverage comprising contacting the food or beverage with at least one peptide according to any one of claims 1-15.

30. A method of manufacturing a skin care product for blocking activity of AGEs comprising synthesizing a first peptide, wherein the first peptide at least one peptide according to any one of claims 1-15, and wherein the first peptide blocks activity of AGEs; and

blending the first peptide with a carrier.

31. The method of claim 30, wherein the carrier is selected from the group consisting of polyethyleneglycol (PEG), polyvinylpyrrolidone, silicate, polystyrene, and cellulose.

32. The method of claim 30 or 31 , wherein the first peptide is encapsulated using liposomes, nanoliposomes, nanoparticles, or nanomaterials.

33. The method of any one of claims 30-32, wherein the first peptide is lipidated.

34. The method of any one of claims 30-33, further comprising

synthesizing a second peptide, wherein the second peptide is at least one peptide according to any one of claims 1-15, and wherein the second peptide blocks activity of AGEs; and

blending the second peptide and the first peptide with a carrier.

35. A method of manufacturing a skin care product for blocking activity of AGEs comprising synthesizing a peptide, wherein the peptide at least one peptide according to any one of claims 1-15; and

preparing a lotion comprising one or more of a diluent, thickener, humectant, occlusive, protease inhibitors or emulsifier, and

blending the peptide with the lotion.

36. A peptide of any one of claims 1-15, for use in a method of treating a condition, disease, or disorder in a subject.

37. The peptide for use according to claim 36, wherein the peptide comprises a pharmaceutically acceptable carrier.

38. The peptide for use according to claim 36 or 37, wherein the condition, disease, or disorder is a chronic inflammatory condition.

39. The peptide for use according to any one of claims 36-38, wherein the condition, disease or disorder is aging, diabetic complications, exposure to pollution, oxidative stress, body or skin detoxification, diabetes, metabolic syndrome, diabetic skin chronic wounds, Alzheimer's disease, atherosclerosis, osteoporosis, osteoarthritis, cancer, cancer therapy associated condition, pollution associated assaults, body and skin detoxification, renal failure, diabetic retinopathy and glaucoma, periodontal disease, cystic fibrosis related diabetes, polycystic ovary syndrome, psoriasis associated condition, oxidative stress, complications associated with maternal chorioamnionitis or funisitis, or sarcopenia.