Collagen hybridization peptides having a modified c-terminal and use thereof

Abstract

Disclosed are a series of novel collagen hybridizing peptides having a lower melting temperature due to modification at the C-terminus, and uses thereof.

Description

[0001] MLB Docket No. 127033-5008

[0002] COLLAGEN HYBRIDIZATION PEPTIDES HAVING A MODIFIED C-TERMINAL

[0003] AND USE THEREOF

[0004] BACKGROUND

[0005] The present disclosure relates to a series of novel collagen hybridizing peptides that have a modified C-terminus and thus have a lower melting temperature, and uses thereof.

[0006] Previous strategies include the addition of nitrobenzyl to the central glycine of the peptide sequences to prevent self-trimerization, the addition of large and bulky fluorescein tags to the side chain of (unnatural) aminoproline in the middle of the sequence, the incorporation of (unnatural) fluoroproline at all Xaa positions, or the formation of charged (positively or negatively charged) strands. Most of these strategies involve the addition of bulky functionalities (e g ., nitrophenyl groups or fluorescein tags) through modification of the unnatural amino acid or residue, ultimately increasing the complexity and cost for mass production.

[0007] Alternatively, ABC heterotrimers having all natural amino acid residues were prepared by forming positively charged chains using lysine residues, negatively charged chains using glutamate or aspartate, and neutrally charged chains containing Gly-Pro-Hyp residues, but the individual charged chains did not form triple helices. In addition, these sequences have a reduced affinity for folding with damaged and denatured collagen compared to the canonical Gly-Pro-Hyp motif.

[0008] Therefore, there is a need to develop a CHP design that is capable of interfering with self- trimerization to keep the CHP active while maintaining a high affinity for binding to denatured collagen.

[0009] The disclosure of U.S. Patent Application Publication No. 2023 / 0375558 is hereby incorporated by reference for all purposes. Tire disclosure of U.S. Patent Application Publication No. 2013 / 0164220 is also hereby incorporated by reference for all purposes herein.

[0010] PROBLEM TO BE SOLVED

[0011] Therefore, the present disclosure has been made in view of the above problems, and it is one object of the present disclosure to provide a collagen hybridizing peptide (CHP) that has a modification at the C-terminus and thus has a lower melting temperature.

[0012] It is another object of the present disclosure to provide a composition containing the CHP.

[0013] It is another object of the present disclosure to provide a method of treating denatured collagen including administering the CHP or the composition to a subject in need thereof.

[0014] It is another object of the present disclosure to provide a composition for treating denatured collagen containing the collagen hybridization peptide (CHP) as an active ingredient.

[0015] DB1 / 161378040.3 MLB Docket No. 127033-5008

[0016] It is another object of the present disclosure to provide a method of inhibiting or reducing an inflammatory response including administering the CHP or the composition to a subject in need thereof.

[0017] It is another object of the present disclosure to provide a composition for inhibiting or reducing an inflammatory response containing the collagen hybridization peptide (CHP) as an active ingredient.

[0018] It is another object of the present disclosure to provide a method of downregulating one or more immune-related cytokines selected from the group consisting of TLR ligands, interleukins, IFN- a, TLR-9, TNF-a, IL-1 , IL-6, CCL3, IL- 16, IL- 18, and TNF-a including administering the CHP or the composition to a subject in need thereof.

[0019] It is another object of the present disclosure to provide a composition for downregulating one or more immune-related cytokines selected from the group consisting of TLR ligands, interleukins, IFN-a, TLR-9, TNF-a, IL-10, IL-6, CCL3, IL-16, IL-18, and TNF-a containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0020] It is another object of the present disclosure to provide a method of preventing immune cell differentiation including administering the CHP or the composition to a subject in need thereof.

[0021] It is another object of the present disclosure to provide a composition for preventing immune cell differentiation containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0022] It is another object of the present disclosure to provide a method of downregulating T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils including administering the CHP or the composition to a subject in need thereof.

[0023] It is another object of the present disclosure to provide a composition for downregulating T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0024] It is another object of the present disclosure to provide a method of preventing differentiation of T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils including administering the CHP or the composition to a subject in need thereof.

[0025] It is another object of the present disclosure to provide a composition for preventing differentiation of T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0026] It is another object of the present disclosure to provide a method of preventing or treating a dry eye disease including administering the CHP or the composition to a subject in need thereof.

[0027] It is another object of the present disclosure to provide a pharmaceutical composition for preventing or treating a dry eye disease containing the collagen hybridizing peptide (CHP) as an active

[0028] DB1 / 161378040.3 MLB Docket No. 127033-5008 ingredient.

[0029] It is another object of the present disclosure to provide a method of preventing or treating atopic dermatitis including administering the CHP or the composition to a subject in need thereof.

[0030] It is another object of the present disclosure to provide a pharmaceutical composition for preventing or treating atopic dermatitis containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0031] It is another object of the present disclosure to provide a method of preventing or treating osteoarthritis including administering the CHP or the composition to a subject in need thereof.

[0032] It is another object of the present disclosure to provide a pharmaceutical composition for preventing or treating osteoarthritis containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0033] It is another object of the present disclosure to provide a method of preventing or treating rheumatoid arthritis including administering the CHP or the composition to a subject in need thereof.

[0034] It is another object of the present disclosure to provide a pharmaceutical composition for preventing or treating rheumatoid arthritis containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0035] It is another object of the present disclosure to provide a method of preventing or treating diabetic lesions including administering the CHP or the composition to a subject in need thereof.

[0036] It is another object of the present disclosure to provide a pharmaceutical composition for preventing or treating diabetic lesions containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0037] It is another object of the present disclosure to provide a method of preventing or treating psoriasis including administering the CHP or the composition to a subject in need thereof.

[0038] It is another object of the present disclosure to provide a pharmaceutical composition for preventing or treating psoriasis containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0039] It is another object of the present disclosure to provide a method of inhibiting checkpoints including administering the CHP or the composition to a subject in need thereof.

[0040] It is another object of the present disclosure to provide a composition for inhibiting checkpoints containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0041] It is another object of the present disclosure to provide a method of healing wounds including diabetic lesions including administering the CHP or the composition to a subject in need thereof.

[0042] It is another object of the present disclosure to provide a pharmaceutical composition for

[0043] DB1 / 161378040.3 MLB Docket No. 127033-5008 healing wounds including diabetic lesions containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0044] It is another object of the present disclosure to provide a method of preventing or treating GI inflammatory conditions including Crohn’s disease and ulcerative colitis including administering the CHP or the composition to a subject in need thereof.

[0045] It is another object of the present disclosure to provide a pharmaceutical composition for preventing or treating GI inflammatory conditions including Crohn’s disease and ulcerative colitis containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0046] It is another object of the present disclosure to provide a cosmetic composition for treating denatured collagen containing the collagen hybridization peptide (CHP) as an active ingredient.

[0047] It is another object of the present disclosure to provide a cosmetic composition for inhibiting or reducing an inflammatory response containing the collagen hybridization peptide (CHP) as an active ingredient.

[0048] It is another object of the present disclosure to provide a cosmetic composition for downregulating one or more immune-related cytokines selected from the group consisting of TLR ligands, interleukins, IFN-a, TLR-9, TNF-a, IL-ip, IL-6, CCL3, IL-16, IL-18, and TNF-a containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0049] It is another object of the present disclosure to provide a cosmetic composition for preventing immune cell differentiation containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0050] It is another object of the present disclosure to provide a cosmetic composition for downregulating T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0051] It is another object of the present disclosure to provide a cosmetic composition for preventing differentiation of T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0052] It is another object of the present disclosure to provide a cosmetic composition for preventing or treating a dry eye disease containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0053] It is another object of the present disclosure to provide a cosmetic composition for preventing or treating atopic dermatitis containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0054] It is another object of the present disclosure to provide a cosmetic composition for preventing or treating psoriasis containing the collagen hybridizing peptide (CHP) as an active

[0055] DB1 / 161378040.3 MLB Docket No. 127033-5008 ingredient.

[0056] It is another object of the present disclosure to provide a cosmetic composition for inhibiting checkpoints containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0057] It is another object of the present disclosure to provide a quasi-drug composition for treating denatured collagen containing the collagen hybridization peptide (CHP) as an active ingredient.

[0058] It is another object of the present disclosure to provide a quasi-drug composition for inhibiting or reducing an inflammatory response containing the collagen hybridization peptide (CHP) as an active ingredient.

[0059] It is another object of the present disclosure to provide a quasi-drug composition for downregulating one or more immune-related cytokines selected from the group consisting of TLR ligands, interleukins, IFN-a, TLR-9, TNF-a, IL-ip, IL-6, CCL3, IL-16, IL-18, and TNF-a containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0060] It is another object of the present disclosure to provide a quasi-drug composition for preventing immune cell differentiation containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0061] It is another object of the present disclosure to provide a quasi-drug composition for downregulating T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0062] It is another object of the present disclosure to provide a quasi-drug composition for preventing differentiation of T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0063] It is another object of the present disclosure to provide a quasi-drug composition for preventing or treating a dry eye disease containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0064] It is another object of the present disclosure to provide a quasi-drug composition for preventing or treating atopic dermatitis containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0065] It is another object of the present disclosure to provide a quasi-drug composition for preventing or treating osteoarthritis containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0066] It is another object of the present disclosure to provide a quasi-drug composition for preventing or treating rheumatoid arthritis containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0067] It is another object of the present disclosure to provide a quasi-drug composition for

[0068] DB1 / 161378040.3 MLB Docket No. 127033-5008 preventing or treating diabetic lesions containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0069] It is another object of the present disclosure to provide a quasi-drug composition for preventing or treating psoriasis containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0070] It is another object of the present disclosure to provide a quasi-drug composition for inhibiting checkpoints containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0071] It is another object of the present disclosure to provide a quasi-drug composition for healing wounds including diabetic lesions containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0072] It is another object of the present disclosure to provide a quasi-drug composition for preventing or treating GI inflammatory conditions including Crohn’s disease and ulcerative colitis containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0073] It is another object of the present disclosure to provide a method of reducing or preventing downregulation of immune cells including administering the composition containing the collagen hybridization peptide as an active ingredient to a subject in need thereof.

[0074] It is another object of the present disclosure to provide a method of preventing or treating cancer including administering the composition containing the collagen hybridization peptide as an active ingredient to a subject in need thereof.

[0075] It is another object of the present disclosure to provide a method of reducing, preventing or treating oxidative modification of natural collagen including administering the composition containing the collagen hybridization peptide as an active ingredient to a subject in need thereof.

[0076] It is another object of the present disclosure to provide a method of reducing, preventing or treating oxidative modification of natural collagen in a subject with diabetes including administering the composition containing the collagen hybridization peptide as an active ingredient to the subject in need thereof.

[0077] It is another object of the present disclosure to provide a method of reducing, preventing or treating oxidative modification of natural collagen in a subject with diabetic ketoacidosis including administering the composition containing the collagen hybridization peptide as an active ingredient to the subject in need thereof.

[0078] It is another object of the present disclosure to provide a method of reducing, preventing or treating oxidative modification of natural collagen in a subject with hyperglycemic hyperosmolar syndrome including administering the composition containing the collagen hybridization peptide as an active ingredient to the subject in need thereof.

[0079] DB1 / 161378040.3 MLB Docket No. 127033-5008

[0080] It is another object of the present disclosure to provide a method of reducing, preventing or treating oxidative modification of natural collagen in a subject with acidosis including administering the composition containing the collagen hybridization peptide as an active ingredient to the subject in need thereof.

[0081] It is another object of the present disclosure to provide a method of reducing, preventing or treating oxidative modification of natural collagen in a subject with homocystinuria including administering the composition containing the collagen hybridization peptide as an active ingredient to the subject in need thereof.

[0082] It is another object of the present disclosure to provide a method of reducing, preventing or treating oxidative modification of natural collagen in a subject with methylmalonic acidemia including administering the composition containing the collagen hybridization peptide as an active ingredient to the subject in need thereof.

[0083] It is another object of the present disclosure to provide a method of reducing, preventing or treating diabetic lesions in a subject including administering the composition containing the collagen hybridization peptide as an active ingredient to the subject in need thereof.

[0084] It is another object of the present disclosure to provide a method of reducing, preventing or treating diabetic retinopathy in a subject including administering the composition containing the collagen hybridization peptide as an active ingredient to the subject in need thereof.

[0085] It is another object of the present disclosure to provide a method of providing information for diagnosing fibrosis in a subject including administering the composition containing the collagen hybridization peptide as an active ingredient to the subject, and imaging the collagen hybridization peptide containing the detection moiety in vivo to determine the presence or progression of fibrosis in the subject.

[0086] MEANS TO SOLVE PROBLEM

[0087] Hereinafter, exemplary embodiments of the present disclosure will be described in detail. However, the present disclosure is not limited to the embodiments disclosed below and may be implemented in various forms. The embodiments described below are provided to enable those skilled in the art to implement and practice the embodiments of the present disclosure.

[0088] Materials, compositions, and components that may be used for the disclosed methods and compositions, may be used in conjunction therewith, may be used in the preparation of the compositions, or are products thereof, are disclosed. These and other materials are disclosed herein, and it is understood that, when combinations, subsets, interactions, groups, and the like of these materials are disclosed, each of the various individual and collective combinations and permutations

[0089] DB1 / 161378040.3 MLB Docket No. 127033-5008 of these compounds may not be explicitly disclosed, but each is specifically considered and described herein. For example, when peptide conjugates are disclosed and discussed, and a number of modifications that may be made on a number of molecules comprising the peptide conjugates are discussed, each and every combination and permutation and possible modification of the peptide conjugate is specifically considered unless specifically stated otherwise. Thus, when not only the classes of molecules A, B, C are disclosed, but also the classes of molecules D, E, F and the combination molecules, for example A-D, are disclosed, each is considered individually and collectively, although not individually mentioned. Thus, in this example, the combinations A-E, A- F, B-D, B-E, B-F, C-D, C-E, and C-F are each specifically considered and should be considered disclosed from the disclosure of the exemplary combinations of A, B and C; D, E, and F; and A-D. Similarly, any subset or combination thereof is also specifically considered and disclosed. Thus, for example, the subsets of A-E, B-F and C-E are specifically considered and should be considered disclosed from the disclosure of the exemplar} / combinations of A, B and C; D, E, and F; and A-D. This applies to all aspects of the present disclosure, including, but not limited to, the steps of preparing and using the disclosed compositions. Accordingly, it is understood that, when there are various additional steps that may be performed, each of these additional steps may be performed by any particular embodiment or combination of embodiments of the disclosed method, and it is understood that each such combination should be considered specifically and disclosed.

[0090] In accordance with one aspect of the present invention, there is provided a collagen hybridizing peptide (CHP), wherein the CHP comprises a sequence represented by Formula I: (Gly-X-Y)a-(Z) (Fonnula I) in which the CHP of the Fonnula I comprises a modification at the C-terminus, thereby lowering a melting temperature of the CHP; X and Y are any amino acid, provided at least one of X and Y is proline, hydroxyproline, or modified proline in each instance of (Gly-X-Y): Gly is glycine; Z is an optional spacer, which may be between (Gly-X-Y)a and the C-terminus; and a is a number from 1 to 40.

[0091] In some embodiments, the CHP comprises a modification at the C-terminus.

[0092] In some embodiments, the Formula I comprises a modification at the C-terminus.

[0093] In one embodiment, the modification at the C-terminus may comprise one or more molecules that yield a lower homotrimerization melting temperature for the CHP than that of the CHP bound with gelatin.

[0094] In another embodiment, the modification at the C-terminus may comprise one or more molecules that yield a lower homotrimerization melting temperature for the CHP than that of the CHP bound with damaged collagen.

[0095] In the CHP of any one of the previous embodiments, the modification at the C-terminus may

[0096] DB1 / 161378040.3 MLB Docket No. 127033-5008 comprise ataching one or more amino acids to the C-terminus of the CHP or the Formula I. In some embodiments, the modification at the C-terminus may comprise ataching one or more amino acids to the C-terminus of the CHP. In some embodiments, the modification at the C-terminus may comprise ataching one or more amino acids to the C-terminus of the Formula I.

[0097] In the CHP of any one of the previous embodiments, the modification at the C-terminus may comprise ataching one or more amino acids including one or more sequences represented by Formula II to the C-terminus of Formula I:

[0098] (Gly-B-B) (Formula II) in which B is independently in each instance phenylalanine, tryp tophan, tyrosine, aspartic acid, lysine, arginine, histidine, or any charged amino acid.

[0099] In the CHP of any one of the previous embodiments, (Z) may comprise at least one bioactive, the at least one bioactive comprising an amino acid sequence from 3 to 27 amino acid residues having bioactivity.

[0100] In the CHP of any one of the previous embodiments, the at least one bioactive may comprise at least one sequence selected from the group consisting of a leukocyte-associated immunoglobulin-like receptor binding sequence (LAIR-1 and / or LAIR-2), an integrin binding sequence, a discoidin domain receptor (DDR) binding sequence, a von Willebrand factor (VWF) sequence, a matrix metalloproteinase (MMP) binding sequence, a G6b-B recognition site sequence, an OSCAR domain sequence, a GPVI sequence, and a FcR-y domain sequence.

[0101] In the CHP of any one of the previous embodiments, the at least one bioactive may comprise a LAIR-1 receptor binding sequence.

[0102] In the CHP of any one of the previous embodiments, the at least one bioactive may comprise a LAIR-2 receptor binding sequence.

[0103] In the CHP of any one of the previous embodiments, the at least one bioactive may comprise a G6b-B recognition site sequence.

[0104] In the CHP of any one of the previous embodiments, the at least one bioactive may comprise a FcR-y domain sequence.

[0105] In the CHP of any one of the previous embodiments, the at least one bioactive may comprise an OSCAR domain sequence.

[0106] In the CHP of any one of the previous embodiments, the at least one bioactive may comprise a GPVI sequence.

[0107] In the CHP of any one of the previous embodiments, the at least one bioactive may comprise an integrin binding sequence.

[0108] In the CHP of any one of the previous embodiments, the at least one bioactive may comprise a

[0109] DB1 / 161378040.3 MLB Docket No. 127033-5008

[0110] DDR binding sequence.

[0111] In the CHP of any one of the previous embodiments, the at least one bioactive may comprise a VWF sequence.

[0112] In the CHP of any one of the previous embodiments, the at least one bioactive may comprise an MMP binding sequence.

[0113] In the CHP of any one of the previous embodiments, the at least one bioactive may comprise a LAIR-1 and / or LAIR-2 receptor binding sequence comprising a sequence having at least 85% sequence identity to any one of SEQ ID NO: 1-27, 56-64, 93, 94, and 98-100.

[0114] In the CHP of any one of the previous embodiments, the at least one bioactive may comprise a G6b-B recognition site sequence comprising a sequence having at least 85% sequence identity to anyone of SEQ ID NO: 28-46, 95, and 96.

[0115] In the CHP of any one of the previous embodiments, the at least one bioactive may comprise a FcR-y domain sequence comprising a sequence having at least 85% sequence identity to any one of SEQ ID NO: 47-55 and 97.

[0116] In the CHP of any one of the previous embodiments, the at least one bioactive may comprise a GPVI domain sequence comprising a sequence having at least 85% sequence identity to any one of SEQ ID NO: 65-83 and 101-103.

[0117] In the CHP of any one of the previous embodiments, the at least one bioactive may comprise an OSCAR domain sequence comprising a sequence having at least 85% sequence identity to any one of SEQ ID NO: 84-92 and 104.

[0118] In the CHP of any one of the previous embodiments, the at least one bioactive may comprise an integrin binding sequence comprising a sequence having at least 85% sequence identity to any one of SEQ ID NO: 107-114.

[0119] In the CHP of any one of the previous embodiments, the at least one bioactive may comprise a sequence selected from the group consisting of SEQ ID NO: 1-104 and 107-114.

[0120] In the CHP of any one of the previous embodiments, (Z) may comprise at least one linker.

[0121] In the CHP of any one of the previous embodiments, the at least one linker may comprise one or more selected from the group consisting of: Ahx (6-aminohexanoic acid), 12-aminododecanoic acid, N-Fmoc-N"-succinyl-4,7,10-trioxa-l,13-tridecanediamine (TEDS), PEG, and a glycine linker (GGG)3-20.

[0122] In the CHP of any one of the previous embodiments, the length of the linker may be 11-45 A.

[0123] In the CHP of any one of the previous embodiments, (Z) may comprise absence or at least one amino acid sequence.

[0124] In the CHP of any one of the previous embodiments, (Z) may comprise at least one repeat of DB1 / 161378040.3 MLB Docket No. 127033-5008

[0125] (Gly-X-Y)a.

[0126] In the CHP of any one of the previous embodiments, the CHP may further comprise one or more of an acetyl capping group, a linker, a cut site, and a bound drug molecule attached at the N- terminus of the CHP. In some embodiments, the CHP may further comprise one or more of an acetyl capping group, a linker, a cut site, and a bound drug molecule attached at the N-terminus of the Formula I.

[0127] In the CHP of any one of the previous embodiments, the cut site is one or more selected from a group consisting of MMP cut-sites, cathepsin cut sites, pH-sensitive cut sites / linker, reducible di-sulfide cut-site / linkers.

[0128] In the CHP of any one of the previous embodiments, the drug molecule is one or more selected from a group consisting of immunomodulators, anti-cancer agents, anti-fibrotic agents, and antiinflammatory agents.

[0129] In the CHP of any one of the previous embodiments, the CHP may further comprise one or more sequence represented by Formula III attached at the N-terminus of Formula I:

[0130] (dmg)-(linker) (Formula III)

[0131] In the CHP of any one of the previous embodiments, the CHP may further comprise one or more sequence represented by Formula IV attached at the N-terminus of Formula I:

[0132] (drug)-(cut site)-(linker) (Formula IV)

[0133] In any one of the previous embodiments, (Z) is bioactive, and the bioactive includes an amino acid sequence from 3 to 27 amino acid residues having bioactivity, wherein the bioactive comprises at least one leukocyte-associated immunoglobulin-like receptor (LAIR-1 and / or LAIR-2) binding sequence, and wherein the amino acid sequence of the bioactive includes at least one citrulline residue.

[0134] In any one of the previous embodiments, (Z) is bioactive, and the bioactive includes any amino acid capable of post-translational modification by a sugar in a glycation reaction.

[0135] In any one of the preceding embodiments, the CHP further includes at least one detection moiety.

[0136] In any of the preceding embodiments, the at least one detection moiety is a dye selected from the group consisting of Alexa Fluor dyes, cyanine dyes, sulfo-cyanine dyes, indocyanine dyes, Tide Flour dyes, TAMRA, FITC, 5-FAM, carboxyfluorescein, coumarin dyes and rhodamine dyes; a gold particle; or a label selected from the group consisting of prednisolone acetate, triamcinolone acetonide and lipid-based artificial tears.

[0137] In accordance with another aspect of the present invention, there is provided a composition comprising the CHP of any one of the preceding embodiments.

[0138] In an embodiment, each individual CHP does not form a triple helix with other CHPs.

[0139] DB1 / 161378040.3 MLB Docket No. 127033-5008

[0140] In another embodiment, the composition may further comprise a carrier.

[0141] In the composition of any one of the previous embodiments, the composition may be a cosmetic composition, and the carrier may comprise at least one selected from the group consisting of micelles, dendrites, lipids, microemulsions, nanoemulsions, solid lipid nanoparticles, polymers, gels, lens, surfactants, cyclodextrins, inserts, nanostructured lipid carriers, liposomes, transfersomes, ethosomes, niosomes, collagen matrix, extracellular matrix, and artificial extracellular matrix.

[0142] In the composition of any one of the previous embodiments, the composition may be a topical cream, a saline solution, gel, polymer, or a solution for systemic injection within a subject.

[0143] In the composition of any one of the previous embodiments, a CHP may be bound directly to a collagen matrix.

[0144] In accordance with another aspect of the present invention, there is provided a method of treating denatured collagen in a subject, comprising administering the CHP or the composition according to any one of the previous embodiments to the subject.

[0145] In an embodiment, the administering may be performed by injection.

[0146] In another embodiment, the administering may be performed by micro-dermal injection.

[0147] In the method of any one of the previous embodiments, the administering may be performed by topical application.

[0148] In accordance with another aspect of the present invention, there is provided a method of inhibiting or reducing an inflammatory response in a subject, comprising administering the CHP or the composition according to any one of the previous embodiments to the subject.

[0149] In an embodiment, the administering may be performed by injection.

[0150] In another embodiment, the administering may be performed by micro-dermal injection.

[0151] In the method of any one of the previous embodiments, the administering may be performed by topical application.

[0152] In accordance with another aspect of the present invention, there is provided a method of dow nregulating immune-related cytokines, including, but not limited to, TLR ligands, interleukins, IFN-a, TLR-9, TNF-a, IL- 1 , IL-6, CCL3, IL-16, IL-18, and / or TNF-a activity in a subject, comprising administering the CHP or the composition according to any one of the previous embodiments to the subject.

[0153] In accordance with another aspect of the present invention, there is provided a method of preventing the differentiation of immune cells comprising administering the CHP or the composition according to any one of the previous embodiments to the subject.

[0154] In accordance with another aspect of the present invention, there is provided a method of downregulating T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils

[0155] DB1 / 161378040.3 MLB Docket No. 127033-5008 comprising administering the CHP or the composition according to any one of the previous embodiments to the subject.

[0156] In accordance with another aspect of the present invention, there is provided a method of preventing the differentiation of T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils comprising administering the CHP or the composition according to any one of the previous embodiments to the subject.

[0157] In an embodiment, the administering may be performed by injection.

[0158] In another embodiment, the administering may be performed by micro-dermal injection.

[0159] In the method of any one of the previous embodiments, the administering may be performed by topical application.

[0160] In accordance with another aspect of the present invention, there is provided a method of preventing or treating dry eye disease in a subject, comprising administering the CHP or the composition according to any one of the previous embodiments to the subject.

[0161] In an embodiment, the administering may be performed by local injection, intravenous injection, topical application, or physical application.

[0162] In another embodiment, the CHP or composition may be administered as a putty, mesh, patch, adhesive, cream, sutures, gel or eyedrops.

[0163] In accordance with another aspect of the present invention, there is provided a method of preventing or treating atopic dermatitis in a subject, comprising administering the CHP or the composition according to any one of the previous embodiments to the subject.

[0164] In an embodiment, the administering may be performed by local injection, intravenous injection, topical application, or physical application.

[0165] In another embodiment, the CHP or composition may be administered as a putty, mesh, patch, adhesive, cream, sutures, or eyedrops.

[0166] In accordance with another aspect of the present invention, there is provided a method of preventing or treating osteoarthritis in a subject, comprising administering the CHP, or the composition according to any one of the previous embodiments to the subject.

[0167] In an embodiment, the administering may be performed by local injection, intravenous injection, topical application, or physical application.

[0168] In another embodiment, the CHP or composition may be administered as a putty, mesh, patch, adhesive, cream, sutures, or eyedrops.

[0169] In accordance with another aspect of the present invention, there is provided a method of preventing or treating rheumatoid arthritis in a subject, comprising administering the CHP, or the composition according to any one of the previous embodiments to the subject.

[0170] DB1 / 161378040.3 MLB Docket No. 127033-5008

[0171] In an embodiment, the administering may be performed by local injection, intravenous injection, topical application, or physical application.

[0172] In another embodiment, the CHP or composition may be administered as a putty, mesh, patch, adhesive, cream, sutures, or eyedrops.

[0173] In accordance with another aspect of the present invention, there is provided a method of preventing or treating diabetic lesions in a subject, comprising administering the CHP or the composition according to any one of the previous embodiments to the subject.

[0174] In an embodiment, the administering may be performed by local injection, intravenous injection, topical application, or physical application.

[0175] In another embodiment, the CHP or composition may be administered as a putty, mesh, patch, adhesive, cream, sutures, or eyedrops.

[0176] In accordance with another aspect of the present invention, there is provided a method of preventing or treating psoriasis in a subject, comprising administering the CHP or the composition according to any one of the previous embodiments to the subject.

[0177] In an embodiment, the administering may be performed by local injection, intravenous injection, topical application, or physical application.

[0178] In another embodiment, the CHP or composition may be administered as a putty, mesh, patch, adhesive, cream, sutures, or eyedrops.

[0179] In accordance with another aspect of the present invention, there is provided a method of checkpoint inhibition in a subject, comprising administering the CHP or the composition according to any one of the previous embodiments to the subject.

[0180] In an embodiment, the administering may be performed by local injection, intravenous injection, topical application, or physical application.

[0181] In another embodiment, the CHP or composition may be administered as a putty, mesh, patch, adhesive, cream, sutures, or eyedrops.

[0182] In accordance with another aspect of the present invention, there is provided a method of wound healing including diabetic lesions in a subject, comprising administering the CHP or the composition according to any one of the previous embodiments to the subject.

[0183] In an embodiment, the administering may be performed by local injection, intravenous injection, topical application, or physical application.

[0184] In another embodiment, the CHP or composition may be administered as a putty, mesh, patch, adhesive, cream, sutures, or eyedrops.

[0185] In the method of any one of the previous embodiments, the at least one bioactivc may comprise an integrin binding sequence comprising a sequence having at least 85% sequence identity to

[0186] DB1 / 161378040.3 MLB Docket No. 127033-5008 any one of SEQ ID NO: 107-109, and wherein the integrin binding sequence may be specific for alpha5-betal integrin.

[0187] In the method of any one of the previous embodiments, the at least one bioactive comprising an integrin binding sequence may enhance cell adhesion and / or collagen production.

[0188] In the method of any one of the previous embodiments, the at least one bioactive comprising an integrin binding sequence specifically may not bind to integrins other than alphao-betal integrin.

[0189] In accordance with another aspect of the present invention, there is provided a method of treating GI inflammatory conditions such as Crohn’s and Ulcerative Colitis in a subject, comprising administering the CHP or the composition according to any one of the previous embodiments to the subject.

[0190] In an embodiment, the administering may be performed by local injection, intravenous injection, topical application, or physical application.

[0191] In another embodiment, the CHP or composition may be administered as a putty, mesh, patch, adhesive, cream, sutures, or eyedrops.

[0192] In accordance with another aspect of the present disclosure, there is provided a composition for treating denatured collagen containing the collagen hybridization peptide (CHP) as an active ingredient.

[0193] In accordance with another aspect of the present disclosure, there is provided a composition for inhibiting or reducing an inflammatory response containing the collagen hybridization peptide (CHP) as an active ingredient.

[0194] In accordance with another aspect of the present disclosure, there is provided a composition for downregulating one or more immune-related cytokines selected from the group consisting of TLR ligands, interleukins, IFN-a, TLR-9, TNF-a, IL-1 , IL-6, CCL3, IL-16, IL-18, and TNF-a containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0195] In accordance with another aspect of the present disclosure, there is provided a composition for preventing immune cell differentiation containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0196] In accordance with another aspect of the present disclosure, there is provided a composition for downregulating T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0197] In accordance with another aspect of the present disclosure, there is provided a composition for preventing differentiation of T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0198] In accordance with another aspect of the present disclosure, there is provided a

[0199] DB1 / 161378040.3 MLB Docket No. 127033-5008 pharmaceutical composition for preventing or treating a dry eye disease containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0200] In accordance with another aspect of the present disclosure, there is provided a pharmaceutical composition for preventing or treating atopic dermatitis containing the collagen hybridizing peptide (CHP) of any one of claims 1-35 as an active ingredient.

[0201] In accordance with another aspect of the present disclosure, there is provided a pharmaceutical composition for preventing or treating osteoarthritis containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0202] In accordance with another aspect of the present disclosure, there is provided a pharmaceutical composition for preventing or treating rheumatoid arthritis containing the collagen hybridizing peptide (CHP) of any one of claims 1-35 as an active ingredient.

[0203] In accordance with another aspect of the present disclosure, there is provided a pharmaceutical composition for preventing or treating diabetic lesions containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0204] In accordance with another aspect of the present disclosure, there is provided a pharmaceutical composition for preventing or treating psoriasis containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0205] In accordance with another aspect of the present disclosure, there is provided a composition for inhibiting checkpoints containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0206] In accordance with another aspect of the present disclosure, there is provided a pharmaceutical composition for healing wounds including diabetic lesions containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0207] In accordance with another aspect of the present disclosure, there is provided a pharmaceutical composition for preventing or treating GI inflammatory conditions including Crohn’s disease and ulcerative colitis containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0208] In accordance with another aspect of the present disclosure, there is provided a cosmetic composition for treating denatured collagen containing the collagen hybridization peptide (CHP) as an active ingredient.

[0209] In accordance with another aspect of the present disclosure, there is provided a cosmetic composition for inhibiting or reducing an inflammatory response containing the collagen hybridization peptide (CHP) as an active ingredient.

[0210] In accordance with another aspect of the present disclosure, there is provided a cosmetic composition for downregulating one or more immune-related cytokines selected from the group

[0211] DB1 / 161378040.3 MLB Docket No. 127033-5008 consisting of TLR ligands, interleukins, IFN-a, TLR-9, TNF-a, IL-ip, IL-6, CCL3, IL-16, IL-18, and TNF-a containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0212] In accordance with another aspect of the present disclosure, there is provided a cosmetic composition for preventing immune cell differentiation containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0213] In accordance with another aspect of the present disclosure, there is provided a cosmetic composition for downregulating T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0214] In accordance with another aspect of the present disclosure, there is provided a cosmetic composition for preventing differentiation of T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0215] In accordance with another aspect of the present disclosure, there is provided a cosmetic composition for preventing or treating a dry eye disease containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0216] In accordance with another aspect of the present disclosure, there is provided a cosmetic composition for preventing or treating atopic dennatitis containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0217] In accordance with another aspect of the present disclosure, there is provided a cosmetic composition for preventing or treating psoriasis containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0218] In accordance with another aspect of the present disclosure, there is provided a cosmetic composition for inhibiting checkpoints containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0219] In accordance with another aspect of the present disclosure, there is provided a quasi-drug composition for treating denatured collagen containing the collagen hybridization peptide (CHP) as an active ingredient.

[0220] In accordance with another aspect of the present disclosure, there is provided a quasi-drug composition for inhibiting or reducing an inflammatory response containing the collagen hybridization peptide (CHP) as an active ingredient.

[0221] In accordance with another aspect of the present disclosure, there is provided a quasi-drug composition for downregulating one or more immune-related cytokines selected from the group consisting of TLR ligands, interleukins, IFN-a, TLR-9, TNF-a, IL-ip, IL-6, CCL3, IL-16, IL-18, and TNF-a containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0222] In accordance with another aspect of the present disclosure, there is provided a quasi-drug

[0223] DB1 / 161378040.3 MLB Docket No. 127033-5008 composition for preventing immune cell differentiation containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0224] In accordance with another aspect of the present disclosure, there is provided a quasi-drug composition for downregulating T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0225] In accordance with another aspect of the present disclosure, there is provided a quasi-drug composition for preventing differentiation of T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0226] In accordance with another aspect of the present disclosure, there is provided a quasi-drug composition for preventing or treating a dry eye disease containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0227] In accordance with another aspect of the present disclosure, there is provided a quasi-drug composition for preventing or treating atopic dermatitis containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0228] In accordance with another aspect of the present disclosure, there is provided a quasi-drug composition for preventing or treating osteoarthritis containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0229] In accordance with another aspect of the present disclosure, there is provided a quasi-drug composition for preventing or treating rheumatoid arthritis containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0230] In accordance with another aspect of the present disclosure, there is provided a quasi-drug composition for preventing or treating diabetic lesions containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0231] In accordance with another aspect of the present disclosure, there is provided a quasi-drug composition for preventing or treating psoriasis containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0232] In accordance with another aspect of the present disclosure, there is provided a quasi-drug composition for inhibiting checkpoints containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0233] In accordance with another aspect of the present disclosure, there is provided a quasi-drug composition for healing wounds including diabetic lesions containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0234] In accordance with another aspect of the present disclosure, there is provided a quasi-drug composition for preventing or treating GI inflammatory conditions including Crohn’s disease and

[0235] DB1 / 161378040.3 MLB Docket No. 127033-5008 ulcerative colitis containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0236] In accordance with another aspect of the present disclosure, there is provided a method of reducing or preventing downregulation of immune cells including administering a composition containing the collagen hybridization peptide as an active ingredient to a subject in need thereof.

[0237] In accordance with another aspect of the present disclosure, there is provided a method of preventing or treating cancer including administering a composition containing the collagen hybridization peptide as an active ingredient to a subject in need thereof.

[0238] In accordance with another aspect of the present disclosure, there is provided a method of reducing, preventing or treating oxidative modification of natural collagen including administering a composition containing the collagen hybridization peptide as an active ingredient to a subject in need thereof.

[0239] In accordance with another aspect of the present disclosure, there is provided a method of reducing, preventing or treating oxidative modification of natural collagen in a subject with diabetes including administering a composition containing the collagen hybridization peptide as an active ingredient to the subject in need thereof.

[0240] In accordance with another aspect of the present disclosure, there is provided a method of reducing, preventing or treating oxidative modification of natural collagen in a subject with diabetic ketoacidosis including administering a composition containing the collagen hybridization peptide as an active ingredient to the subject in need thereof.

[0241] In accordance with another aspect of the present disclosure, there is provided a method of reducing, preventing or treating oxidative modification of natural collagen in a subject with hyperglycemic hyperosmolar syndrome including administering a composition containing the collagen hybridization peptide as an active ingredient to the subject in need thereof.

[0242] In accordance with another aspect of the present disclosure, there is provided a method of reducing, preventing or treating oxidative modification of natural collagen in a subject with acidosis including administering a composition containing the collagen hybridization peptide as an active ingredient to the subject in need thereof.

[0243] In accordance with another aspect of the present disclosure, there is provided a method of reducing, preventing or treating oxidative modification of natural collagen in a subject with homocystinuria including administering a composition containing the collagen hybridization peptide as an active ingredient to the subject in need thereof.

[0244] In accordance with another aspect of the present disclosure, there is provided a method of reducing, preventing or treating oxidative modification of natural collagen in a subject with methylmalonic acidemia including administering a composition containing the collagen hybridization

[0245] DB1 / 161378040.3 MLB Docket No. 127033-5008 peptide as an active ingredient to the subject in need thereof.

[0246] In accordance with another aspect of the present disclosure, there is provided a method of reducing, preventing or treating diabetic lesions in a subject including administering a composition containing the collagen hybridization peptide as an active ingredient to the subject in need thereof.

[0247] In accordance with another aspect of the present disclosure, there is provided a method of reducing, preventing or treating diabetic retinopathy in a subject including administering the composition containing the collagen hybridization peptide as an active ingredient to the subject in need thereof.

[0248] In accordance with another aspect of the present disclosure, there is provided a method of providing information for diagnosing fibrosis in a subject including administering a composition containing the collagen hybridization peptide as an active ingredient to the subject, and imaging the collagen hybridization peptide containing the detection moiety in vivo to determine presence or progression of fibrosis in the subject.

[0249] In another embodiment, the fibrosis may be selected from the group consisting of neovascular age-related macular degeneration (nAMD), diabetic retinopathy, glaucoma, particularly fibrosis in the trabecular meshwork, neovascular glaucoma, corneal scarring, conjunctiva, postcataract surgery, retinopathy of prematurity, and proliferative vitreoretinopathy.

[0250] DETAILED DESCRIPTION

[0251] It will be understood that, although the terms “first,” “second,” etc., may be used herein to describe various elements, these terms should not be construed as limiting these elements. These terms are used only to distinguish one element from another. For example, within the scope of the exemplary embodiment, a “first” element may be referred to as a “second” element, and similarly, a “second” element may be referred to as a “first” element. In addition, the term “and / or” includes any combination of one or more of the related mentioned items.

[0252] It must be noted that, as used herein and the appended claims, the singular forms “a,” “an,” and “the” include plural forms unless the context clearly indicates otherwise. Thus, for example, the expression “a peptide” includes a plurality of peptides including such peptide, and the expression “the peptide” includes one or more peptides and equivalents thereof and the like known to those skilled in the art. Similarly, the term “or” is intended to include “and” unless the context clearly indicates otherwise.

[0253] As used herein, the term “at least one” in relation to a list of one or more elements means at least one element selected from any one or more of the elements in the list of elements, but docs not necessarily include at least one of all elements specifically listed in the list of elements, and does not

[0254] DB1 / 161378040.3 MLB Docket No. 127033-5008 exclude any combination of elements in the list of elements. This definition also allows that other elements may optionally be present in addition to the elements specifically identified in the list of elements to which the term “at least one” relates, whether or not it is related to those elements specifically identified. Thus, as a non-limiting example, “at least one of A and B” (or, equivalently , “at least one of A or B,” or, equivalently, “at least one of A and / or B”) may refer to, in one embodiment, at least one A, optionally including more than one B, where B is not present (and, optionally, including an element other than A); in another embodiment, at least one B, optionally including more than one B, where A is not present (and, optionally, including an element other than A); in yet another embodiment, at least one A, optionally including more than 1, and at least one B, optionally including more than 1 (and optionally including other elements).

[0255] It should be understood that the ranges mentioned herein are shorthand expressions including all values within that range. For example, the range from 1 to 50 includes any number of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, or 50, a combination thereof, or a subrange thereof.

[0256] The terms “comprising,” “including,” “having,” and the like are used interchangeably and have the same meaning. Similarly, the terms “comprises,” “includes,” “has” and the like are used interchangeably and have the same meaning. Specifically, each term is defined consistently with the term “comprising” as generally defined in U.S. patent law and thus is construed as an open-ended term meaning “at least the following” while not excluding additional features, limitations, aspects, etc. Thus, for example, “a device having components a, b, and c” means that the device includes at least components a, b, and c. Similarly, the phrase “a method comprising steps a, b, and c” means that the method includes at least steps a, b, and c. Furthermore, steps and processes may be described schematically herein in a particular order, but those skilled in the art will recognize that the order of steps and processes may vary, unless a particular order is clearly mentioned by context.

[0257] As used herein, the term “about” refers to a numerical value, including, for example, integers, fractions, and percentages, whether or not explicitly mentioned. The term “about” generally refers to a range of numerical values (e.g., + / - 5, 6, 7, 8, 9, or 10% of the stated value) that those skilled in the art would consider equivalent to the stated value (e.g., having the same function or result). In some cases, the term “about” may include a numerical value rounded to the nearest significant figure. In general, the term “about” includes a value for a given amount that falls at least within a corresponding margin of error for preparation, formulation, and / or measurement.

[0258] As used herein, the term “collagen” may refer to collagen derived from any type of tissue (e.g., bone, dermis, tendon, ligament, or the like). Collagen may refer to a molecule in which three

[0259] DB1 / 161378040.3 MLB Docket No. 127033-5008 alpha chains of a polyproline Il-like structures fold together to form a triple helix. In addition, this may be applied to any protein that contains a triple helical region, including collagen types I -XXVIII and bacterial collagen. As used herein, the term “collagen” may refer to any form of collagen, including artificial collagen and processed, or otherwise modified collagen. In some embodiments, the collagen is selected from type I collagen, type II collagen, type III collagen, type IV collagen, type V collagen, type VI collagen, type VII collagen, type VIII collagen, type IX collagen, type X collagen. type XI collagen, type XII collagen, type XIII collagen, type XIV collagen, type XV collagen, type XVI collagen, type XVII collagen, type XVIII collagen, type XIX collagen, type XX collagen, type XXI collagen, type XXII collagen, type XXIII collagen, type XXIV collagen, type XXV collagen, type XXVI collagen, type XXVII collagen, type XXVIII collagen, and combinations thereof.

[0260] As used herein, the term “proline or modified proline” refers to the amino acid proline, natural and unnatural isomers thereof, various isomers including both natural and unnatural isomers thereof, and analogues and variants thereof. In an embodiment, the modified proline includes an electron withdrawing group. Examples of the modified proline include, but are not limited to, hydroxyproline, methylated proline, 4-fluoroproline, and 4-chloroproline.

[0261] As used herein, the tenn “hydroxyproline” refers to hydroxy group-substituted proline, which is a type of the modified proline.

[0262] As used herein, the term “citrullinated arginine” refers to deaminated and citrullinated arginine in which arginine is substituted with citrulline. Thus, “citrullinated arginine” may also be interchangeably used with “citrulline.”

[0263] In some embodiments, the method excludes collecting a sample from a subject. In some embodiments, the term “subject” used herein may refer to a human, an animal, or a microorganism, or a cell culture from any of the groups mentioned above. Examples of the animal include, but are not limited to. vertebrates such as primates, rodents, domestic animals, or game animals. The primates include chimpanzees, cynomolgus monkeys, spider monkeys, and macaques (e.g., Rhesus). The rodents include mice, rats, woodchucks, ferrets, rabbits, and hamsters. Domestic animals and game animals include cattle, horses, pigs, deer, bison, buffalo, moose, felines (e.g., domestic cats), and canines (e g., dogs, foxes, wolves). The fish include Chondrichthyes (cartilaginous fish) and Osteichthyes (bony fish). The subject may be a mammal. Hie mammal may be a human, a nonhuman primate, a mouse, a rat, a dog. a cat. a horse, or a cow, but is not limited thereto. In addition, the methods described herein may be used to diagnose and / or treat livestock or pets. This term does not denote a particular age or gender. Thus, this tenn is intended to encompass all gender including male and female, adults, newborns, and fetuses.

[0264] The term “prevention” refers to any action that inhibits or delays the occurrence, propagation, DB1 / 161378040.3 MLB Docket No. 127033-5008 and recurrence of a specific disease, and the term '‘treatment” refers to partial or complete relief, alleviation, palliation, delayed onset, inhibition of progression, reduction in severity, and / or reduction in incidence of one or more symptoms or characteristics of a specific disease, disorder, and / or condition. For example, the prevention or treatment may be performed by administration to a subject who does not have signs of the disease, disorder, and / or condition and / or to a subject who has only early signs of the disease, disorder, and / or condition, to reduce the risk of developing pathology associated with the disease, disorder, and / or condition.

[0265] The present applicant has designed collagen hybridizing peptides (CHPs) which have reduced self-trimerization dependent on the generation of interstrand steric hindrance between hydroxyproline (Hyp, O) residues of one chain and fluoroproline (flp, f) residues of an adjacent chain. This method successfully reduced the stability of CHP homotrimers from 65°C for (GPOjp CHP to 27°C for (GfOf CHP, as measured by circular dichroism (CD) (Gly is glycine, P is proline. 0 is hydroxyproline. and f is 2S,4S-4-fluoroproline, in the cis conformation). The method worked well but required the use of the unnatural amino acid 2S,4S-4-fluoroproline, which is much more expensive than standard proline residues and has different properties from CHPs containing all natural amino acid residues. The method has some beneficial properties, such as high stability due to low enzymatic recognition, but also some undesirable properties, such as reduced cellular recognition potential.

[0266] Therefore, the present inventors provide a new method for designing CHPs that are resistant to self-trimerization while still retaining the ability to target and bind to damaged collagen strands.

[0267] The new design is based on the theory in which the CHP first nucleates at the C-terminus of the peptide and then folds along the entire sequence toward the N-terminus, thereby hybridizing with denatured collagen strands. Furthermore, the new design contains only naturally occurring amino acids and thus is expected to provide higher biomimicry and substantially lower preparation costs compared to when using unnatural residues.

[0268] Specifically, the present applicants decided to incorporate two Phe residues into the X-Y positions of the last triplet on the C-terminus thereof and found that this was unable to form a homotrimer. This was achieved due to steric hindrance on the C-terminus since the Phe residues include a large phenyl ring in the side chain thereof. This severely limited tire ability of the adjacent CHP strand to nucleate and fold on the C-terminus thereof. In addition, the present applicant obtained similar results when incorporating charged residues into the C-terminal Gly-X-Y motif. By introducing two glutamic acid (Glu, E) residues, the ability of CHP to self-trimerize was hindered using charge repulsion and some steric hindrance. Several initial sequences were synthesized to perform proof-of-conccpt studies based on self-trimerization and binding to damaged collagen, and it was found that the melting point and binding affinity could be fine-tuned to meet the needs of the present applicant.

[0269] DB1 / 161378040.3 MLB Docket No. 127033-5008

[0270] The present applicant has developed a rational design that integrates data having the rules for triple-helix folding (i.e., each third glycine residue) and the disruption of triple-helix stability to create novel peptides that are resistant to self-trimerization, but it is important not to completely inhibit the ability thereof to fold on denatured collagen strands. This subtle approach of interfering with self- trimerization, but not full trimerization, of CHP means that the stability of the homotrimer may be precisely controlled to maintain single strands below main temperatures, such as body temperature (37°C), room temperature (~20-22°C), or skin temperature (~30-33°C). The stability of the triple helix may be increased or decreased by intelligent selection and placement of amino acids within the CHP sequence. This may be achieved using naturally occurring amino acids, which causes simplified synthesis, reduced cost, maintenance of the ability to biosynthesize CHP by yeast or bacteria, and thus usefulness for cosmetics. In addition, unnatural amino acids may be incorporated to further modulate the ability of CHP to bind to denatured collagen.

[0271] Since the bioACTIVE CHP, which is truncated at the C-terminus, addresses the upstream problem of cell migration into damaged areas, a bioACTIVE CHP that binds to damaged collagen and helps modulate cellular responses has broad applicability. Two causes of cell migration into damaged areas are (1) loss of mechanotransduction between cells and the microenvironment due to mechanically poor ECM, and (2) loss of signaling sites (integrins, LAIRs, DDRs, and the like) found on collagen by enzymatic cleavage, both of which are important for cell-to-cell communication in tissues.

[0272] For example, by increasing the local concentration of integrin sites at the site of cell adhesion, cell migration to the site of damage or retention and attachment to the ECM at that site can be promoted. Once attached, the cells can perform other functions, such as releasing MMPs to help break down damaged collagen or increasing collagen production to replace and repair the damaged microenvironment. Thus, by increasing the integrin areas at the damaged site, the number of adherent cells can be increased to increase the rate of wound healing, and collagen production can be increased to restore the tissue to the original state thereof. The RGD-CHP design utilizes the ability of CHP to target and bind to damaged collagen and the ability of RGD to be recognized without being present in the triple helix. Alternatively, a combination of a triple helix or two CHPs is effective, and thus GFOGER bioACTIVE may be used to achieve increased cell attachment by providing attachment sites.

[0273] In accordance with one aspect of the present disclosure, there is provided a collagen hybridizing peptide (CHP) including a sequence represented by Formula I: [Formula I]

[0274] (Gly-X-Y)a-(Z) wherein the CHP includes a modification at the C -terminus and thus has a lower melting temperature;

[0275] DB1 / 161378040.3 MLB Docket No. 127033-5008

[0276] X and Y are any amino acids, provided at least one of X or Y is proline, hydroxyproline, or modified proline in each unit of (Gly-X-Y);

[0277] Gly is glycine;

[0278] Z is an optional spacer; and a is a natural number from 1 to 40.

[0279] For example, the modification at the C-terminus may include one or more molecules that impart a lower homotrimerization melting temperature to the CHP than that of the CHP bound with gelatin.

[0280] For example, the modification at the C-terminus may include one or more molecules that impart a lower homotrimerization melting temperature to the CHP than that of the CHP bound with damaged collagen.

[0281] For example, in some embodiments, the modification at the C-terminus may include attaching one or more amino acids to the C-terminus of Formula I, but is not limited thereto.

[0282] For example, in some embodiments, the modification at the C-terminus may include attaching one or more amino acids including one or more sequences represented by Formula II to the C-terminus of Formula I:

[0283] [Fonnula II]

[0284] (Gly-B-B) wherein B is each independently phenylalanine, tryptophan, tyrosine, aspartic acid, lysine, arginine, histidine, or any charged amino acid.

[0285] For example, (Z) may include at least one bioactive and the at least one bioactive may include an amino acid sequence having 3 to 27 amino acid residues having bioactivity.

[0286] For example, the at least one bioactive may include at least one sequence selected from the group consisting of a leukocyte-associated immunoglobulin-like receptor binding sequence (LAIR-1 and / or LAIR-2), an integrin binding sequence, a discoidin domain receptor (DDR) binding sequence, a von Willebrand factor (VWF) sequence, a matrix metalloproteinase (MMP) binding sequence, a G6b-B recognition site sequence, an OSCAR domain sequence, a GPVI sequence, and a FcR-y domain sequence, but is not limited thereto.

[0287] For example, the at least one bioactive may include a LAIR-1 receptor binding sequence.

[0288] For example, the at least one bioactive may include a LAIR-2 receptor binding sequence.

[0289] For example, the at least one bioactive may include a G6b-B recognition site sequence.

[0290] For example, the at least one bioactive may include a FcR-y recognition site sequence.

[0291] For example, the at least one bioactivc may include an OSCAR recognition site sequence.

[0292] For example, the at least one bioactive may include a GPVI recognition site sequence.

[0293] DB1 / 161378040.3 MLB Docket No. 127033-5008

[0294] For example, the at least one bioactive may include an integrin recognition site sequence.

[0295] For example, the at least one bioactive may include a DDR recognition site sequence.

[0296] For example, the at least one bioactive may include a VWF recognition site sequence.

[0297] For example, the at least one bioactive may include an MMP recognition site sequence.

[0298] For example, the at least one bioactive may include a LAIR-1 and / or LAIR-2 receptor binding sequence including a sequence having at least 85% sequence identity to any one of SEQ ID NOs: 1-27, 56-64, 93, 94, and 98-100, but is not limited thereto.

[0299] For example, the at least one bioactive may include a G6b-B recognition site sequence including a sequence having at least 85% sequence identity to any one of SEQ ID NOs: 28-46, 95, and 96, but is not limited thereto.

[0300] For example, the at least one bioactive may include a FcR-y domain sequence including a sequence having at least 85% sequence identity to any one of SEQ ID NOs: 47-55 and 97, but is not limited thereto.

[0301] For example, the at least one bioactive may include a GPVI domain sequence including a sequence having at least 85% sequence identity to any one of SEQ ID NOs: 65-83 and 101-103, but is not limited thereto.

[0302] For example, the at least one bioactive may include an OSCAR domain sequence including a sequence having at least 85% sequence identity to any one of SEQ ID NOs: 84-92 and 104, but is not limited thereto.

[0303] For example, the at least one bioactive may include an integrin binding sequence including a sequence having at least 85% sequence identity to any one of SEQ ID NOs: 107-114, but is not limited thereto.

[0304] For example, the at least one bioactive may include a sequence selected from the group consisting of SEQ ID NOs: 1-104 and 107-114. but is not limited thereto.

[0305] CHP sequences used in some embodiments are disclosed in Table 1.

[0306] Table 1

[0307] DB1 / 161378040.3 MLB Docket No. 127033-5008

[0308] DB1 / 161378040.3 MLB Docket No. 127033-5008

[0309] DB1 / 161378040.3 MLB Docket No. 127033-5008

[0310] DB1 / 161378040.3 MLB Docket No. 127033-5008

[0311] DB1 / 161378040.3 MLB Docket No. 127033-5008

[0312] The Ac- is an acetyl (capping) group at the N-terminus, the G- is glycine, the P- is proline, the 0- is hydroxyproline, the f- is 2S.4S-4-fluoroproline. x is any integer from 1-7, y is any integer from 1- 7, and z is any integer from 1-5.

[0313] For example, (Z) may include at least one linker, but is not limited thereto.

[0314] Specifically, the at least one linker may include at least one of Ahx (6-aminohexanoic acid); other linkers such as 12-aminododecanoic acid and N-Fmoc-N"-succinyl-4,7,10-trioxa-l,13- tridecanediamine (TTDS); a polymer such as PEG; or a glycine linker (GGG)3 2o, but is not limited thereto.

[0315] In this case, the length of the linker used may affect the ability of the RGD moiety to promote cell attachment. The RGD should enter the globular head on the cell receptor and a linker with a length of 11 to 45A may affect cell attachment.

[0316] In addition, (Z) may be absent or may include at least one amino acid sequence, but is not limited thereto.

[0317] Alternatively, (Z) may include at least one (Gly-X-Y)arepeat. In this case, Gly. X, Y, and a are as defined above.

[0318] For example, (Z) may include, but is not limited to, at least one bioactive, a linker, at least one (Gly-X-Y)arepeat unit, or a combination thereof, and the bioactive, linker, Gly, X, Y and a are as defined above.

[0319] For example, (Z) may further include at least one of an acetyl capping group, a linker, a cut site, or a bound drug molecule attached at the N-terminus of Formula 1.

[0320] Specifically, the cut site may include at least one selected from group consisting of MMP cut sites, cathepsin cut sites, pH-sensitive cut sites / linker, and reducible di-sulfide cut site / linkers, but is not limited thereto.

[0321] Specifically, the drug molecule may include one or more selected from group consisting of immunomodulators, anti-cancer agents, anti-fibrotic agents, and anti-inflammatory agents, but is not limited thereto.

[0322] DB1 / 161378040.3 MLB Docket No. 127033-5008

[0323] Specifically, the CHP may further include, but is not limited to, at least one sequence represented by Formula III attached at the N-terminus of Formula I:

[0324] [Formula III]

[0325] (dmg)-(linker)

[0326] More specifically, the CHP may further include, but is not limited to, at least one sequence represented by Formula IV attached at the N-terminus of Formula I:

[0327] [Formula IV]

[0328] (drug)-(cut site)-(linker)

[0329] In some embodiments, in the CHP, (Z) is bioactive and the bioactive includes an amino acid sequence having 3 to 27 amino acid residues having bioactivity, wherein the bioactive includes at least one leukocyte-associated immunoglobulin-like receptor (LAIR-1 and / or LAIR-2) binding sequence, and wherein the amino acid sequence of the bioactive includes at least one citrulline residue.

[0330] In some embodiments, in the CHP, (Z) is bioactive and the bioactive includes any amino acid capable of post -translational modification by a sugar in a glycation reaction. Since (Z) includes the amino acid, it binds to AGEs preferentially rather than natural collagen, is preferentially glycated, and may be freely separated and discharged. Therefore, the CHP helps maintain a normal, healthy vascular system. Furthermore, since the CHP prevents oxidative modification of natural collagen and suppresses plaque formation by AGEs, it is useful for the prevention or treatment of various diseases mediated thereby.

[0331] In accordance with another aspect of the present disclosure, there is provided a composition containing the CHP.

[0332] In an embodiment, one CHP contained in the CHP does not form a triple helix with other CHPs.

[0333] In addition, the composition may further contain a pharmaceutically acceptable carrier. In this case, the carrier may include, but is not limited to, at least one selected from the group consisting of micelles, dendrites, lipids, microemulsions, nanoemulsions, solid lipid nanoparticles, polymers, gels, lenses, surfactants, cyclodextrins, inserts, nanostructured lipid carriers, liposomes, transfersomes, ethosomes, niosomes, collagen matrices, extracellular matrices, and artificial extracellular matrices.

[0334] For example, the composition may be provided in the form of a topical cream, saline solution, gel, polymer, or solution for systemic injection into a subject, but is not limited thereto.

[0335] For example, in the composition, the CHP in the composition directly binds to the collagen matrix.

[0336] In accordance with another aspect of the present disclosure, there is provided a method of treating denatured collagen including administering the CHP or the composition to a subject in need

[0337] DB1 / 161378040.3 MLB Docket No. 127033-5008 thereof.

[0338] In accordance with another aspect of the present disclosure, there is provided a composition for treating denatured collagen containing the collagen hybridization peptide (CHP) as an active ingredient.

[0339] In accordance with another aspect of the present disclosure, there is provided a method of inhibiting or reducing an inflammatory response including administering the CHP or the composition to a subject in need thereof.

[0340] In accordance with another aspect of the present disclosure, there is provided a composition for inhibiting or reducing an inflammatory response containing the collagen hybridization peptide (CHP) as an active ingredient.

[0341] In accordance with another aspect of the present disclosure, there is provided a method of downregulating one or more immune-related cytokines selected from the group consisting of TLR ligands, interleukins, IFN-a, TLR-9, TNF-a, IL-1 , IL-6, CCL3, IL-16, IL-18, and TNF-a including administering the CHP or the composition to a subject in need thereof.

[0342] In accordance with another aspect of the present disclosure, there is provided a composition for downregulating one or more immune-related cytokines selected from the group consisting of TLR ligands, interleukins, IFN-a, TLR-9, TNF-a, IL-10, IL-6, CCL3, IL-16, IL-18, and TNF-a containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0343] In accordance with another aspect of the present disclosure, there is provided a method of preventing immune cell differentiation including administering the CHP or the composition to a subject in need thereof.

[0344] In accordance with another aspect of the present disclosure, there is provided a composition for preventing immune cell differentiation containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0345] In accordance with another aspect of the present disclosure, there is provided a method of downregulating T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils including administering the CHP or the composition to a subject in need thereof.

[0346] In accordance with another aspect of the present disclosure, there is provided a composition for downregulating T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0347] In accordance with another aspect of the present disclosure, there is provided a method of preventing differentiation of T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils including administering the CHP or the composition to a subject in need thereof.

[0348] In accordance with another aspect of the present disclosure, there is provided a composition

[0349] DB1 / 161378040.3 MLB Docket No. 127033-5008 for preventing differentiation of T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0350] In accordance with another aspect of the present disclosure, there is provided a method of preventing or treating a dry eye disease including administering the CHP or the composition to a subject in need thereof.

[0351] In accordance with another aspect of the present disclosure, there is provided a pharmaceutical composition for preventing or treating a dry eye disease containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0352] In accordance with another aspect of the present disclosure, there is provided a method of preventing or treating atopic dennatitis including administering the CHP or the composition to a subject in need thereof.

[0353] In accordance with another aspect of the present disclosure, there is provided a composition for preventing or treating atopic dermatitis containing the collagen hybridizing peptide (CHP) as an active ingredient. In accordance with another aspect of the present disclosure, there is provided a pharmaceutical composition for preventing or treating atopic dermatitis containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0354] In accordance with another aspect of the present disclosure, there is provided a method of preventing or treating osteoarthritis including administering the CHP or the composition to a subject in need thereof.

[0355] In accordance with another aspect of the present disclosure, there is provided a pharmaceutical composition for preventing or treating osteoarthritis containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0356] In accordance with another aspect of the present disclosure, there is provided a method of preventing or treating rheumatoid arthritis including administering the CHP or the composition to a subject in need thereof.

[0357] In accordance with another aspect of the present disclosure, there is provided a pharmaceutical composition for preventing or treating rheumatoid arthritis containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0358] In accordance with another aspect of the present disclosure, there is provided a method of preventing or treating diabetic lesions including administering the CHP or the composition to a subject in need thereof.

[0359] In accordance with another aspect of the present disclosure, there is provided a pharmaceutical composition for preventing or treating diabetic lesions containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0360] DB1 / 161378040.3 MLB Docket No. 127033-5008

[0361] In accordance with another aspect of the present disclosure, there is provided a method of preventing or treating psoriasis including administering the CHP or the composition to a subject in need thereof.

[0362] In accordance with another aspect of the present disclosure, there is provided a pharmaceutical composition for preventing or treating psoriasis containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0363] In accordance with another aspect of the present disclosure, there is provided a method of inhibiting checkpoints including administering the CHP or the composition to a subject in need thereof.

[0364] In accordance with another aspect of the present disclosure, there is provided a composition for inhibiting checkpoints containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0365] In accordance with another aspect of the present disclosure, there is provided a method of healing wounds including diabetic lesions including administering the CHP or the composition to a subject in need thereof.

[0366] In accordance with another aspect of the present disclosure, there is provided a pharmaceutical composition for healing wounds including diabetic lesions containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0367] In accordance with another aspect of the present disclosure, there is provided a method of preventing or treating GI inflammatory conditions including Crohn’s disease and ulcerative colitis including administering the CHP or the composition to a subject in need thereof.

[0368] In accordance with another aspect of the present disclosure, there is provided a pharmaceutical composition for preventing or treating GI inflammatory conditions including Crohn’s disease and ulcerative colitis containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0369] As used herein, the term “administration” means introducing a predetermined substance into a patient in an appropriate manner and the composition may be administered through any general route as long as it can reach the target tissue. Specifically, the composition may be administered intraperitoneally, intravenously, intramuscularly, subcutaneously, intradermally, orally, topically, intranasally. intrapulmonary, or rectally, but is not limited thereto.

[0370] The composition may be prepared into the formulation described above and may be administered by injection, micro-dermal injection, intravenous injection, intratumoral injection, subcutaneous injection, topical application, or oral administration, but is not limited thereto.

[0371] Meanwhile, the composition may be administered in a pharmaceutically effective amount, and the term “pharmacally effective amount” refers to an amount sufficient to treat a disease at a

[0372] DB1 / 161378040.3 MLB Docket No. 127033-5008 reasonable benefit / risk ratio applicable to all medical treatments, without causing side effects. The effective dose may be easily determined by those skilled in the art based on factors including the gender, age, weight, health condition, and type and severity of the disease of the patient, activity of the drug, sensitivity of the patient to the drug, administration method, administration time, administration route, and excretion rate, treatment period, drugs used in combination or simultaneously, and other factors well known in the pharmaceutical field. In general, the active ingredient may be administered at a dose of about 0.001 mg / kg / day to 1,000 mg / kg / day. For example, the active ingredient may be administered at a dose of 0.01 mg / kg / day to 100 mg / kg / day, or 0.01 mg / kg / day to 35 mg / kg / day, but is not limited thereto. For oral administration, the active ingredient is preferably the range of 50 to 500 mg / kg and may be administered once or more per day. Specifically, the active ingredient is generally administered to an adult patient weighing 70 kg at a dose of 0.07 to 7,000 mg / day, preferably 0.7 to 2,500 mg / day and may be administered once or several times per day in divided doses at regular intervals depending on the judgment of physicians or pharmacists.

[0373] In some embodiments, the administration of the composition for treating denatured collagen may be performed by injection, but is not limited thereto.

[0374] In other embodiments, the administration of the composition for treating denatured collagen may be performed by micro-dennal injection, but is not limited thereto.

[0375] In yet other embodiments, the administration of the composition for treating denatured collagen may be performed by topical application, but is not limited thereto.

[0376] In some embodiments, the administration of the composition for inhibiting or reducing an inflammatory response may be performed by injection, but is not limited thereto.

[0377] In other embodiments, the administration of the composition for inhibiting or reducing an inflammatory response may be performed by micro-dermal injection, but is not limited thereto.

[0378] In yet other embodiments, the administration of the composition for inhibiting or reducing an inflammatory response may be performed by topical application, but is not limited thereto.

[0379] In some embodiments, the administration of the compositions for downregulating immune- related cytokines, for preventing immune cell differentiation, for downregulating T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils, and / or for preventing differentiation of T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils may be performed by injection, but is not limited thereto.

[0380] In other embodiments, the administration of the compositions for downregulating immune- related cytokines, for preventing immune cell differentiation, for downregulating T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils, and / or for preventing differentiation of T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils may be performed by micro-dermal

[0381] DB1 / 161378040.3 MLB Docket No. 127033-5008 injection, but is not limited thereto.

[0382] In other embodiments, the administration of the compositions for downregulating immune- related cytokines, for preventing immune cell differentiation, for downregulating T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils, and / or for preventing differentiation of T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils may be performed by local application, but is not limited thereto.

[0383] In some embodiments, the administration of the composition for preventing or treating GI inflammatory conditions including dry eye, atopic dermatitis, osteoarthritis, rheumatoid arthritis, diabetic lesions, psoriasis, and / or Crohn’s disease and ulcerative colitis; and the composition for inhibiting checkpoints; and / or the composition for healing wounds including diabetic lesions may be performed by topical injection, intravenous injection, topical application, or physical application, but is not limited thereto.

[0384] For this purpose, the composition may be administered in the form of, but is not limited to, a putty, mesh, patch, plaster, cream, suture, gel, and / or eye drops.

[0385] In some embodiments, the at least one bioactive of CHP contained in the composition for healing wounds including diabetic lesions include an integrin binding sequence including a sequence having at least 85% sequence identity to any one of SEQ ID NOs: 107 to 109, and the integrin binding sequence may be specific for alpha5-betal integrin.

[0386] In addition, the at least one bioactive including the integrin binding sequence may enhance cell adhesion and / or collagen production.

[0387] Furthermore, the at least one bioactive including the integrin binding sequence may not specifically bind to any integrin other than alpha5-betal integrin.

[0388] In an embodiment, the composition may be provided in the form of a food composition for preventing or ameliorating the aforementioned diseases.

[0389] For example, the composition may be used before or after the onset of the disease, simultaneously with or separately from a drug for treating the disease, to prevent or ameliorate the aforementioned diseases.

[0390] As used herein, the tenn “amelioration” means any action that at least reduces a parameter related to the treatment condition, such as the degree of symptoms.

[0391] In accordance with another aspect of the present disclosure, there is provided a cosmetic composition for treating denatured collagen containing the collagen hybridization peptide (CHP) as an active ingredient.

[0392] In accordance with another aspect of the present disclosure, there is provided a cosmetic composition for inhibiting or reducing an inflammatory response containing the collagen hybridization

[0393] DB1 / 161378040.3 MLB Docket No. 127033-5008 peptide (CHP) as an active ingredient.

[0394] In accordance with another aspect of the present disclosure, there is provided a cosmetic composition for downregulating one or more immune-related cytokines selected from the group consisting of TLR ligands, interleukins, IFN-a, TLR-9, TNF-a, IL- 1 p, IL-6, CCL3, IL-16, IL- 18, and TNF-a containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0395] In accordance with another aspect of the present disclosure, there is provided a cosmetic composition for preventing immune cell differentiation containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0396] In accordance with another aspect of the present disclosure, there is provided a cosmetic composition for downregulating T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0397] In accordance with another aspect of the present disclosure, there is provided a cosmetic composition for preventing differentiation of T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0398] In accordance with another aspect of the present disclosure, there is provided a cosmetic composition for preventing or treating a dry eye disease containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0399] In accordance with another aspect of the present disclosure, there is provided a cosmetic composition for preventing or treating atopic dermatitis containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0400] In accordance with another aspect of the present disclosure, there is provided a cosmetic composition for preventing or treating psoriasis containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0401] In accordance with another aspect of the present disclosure, there is provided a cosmetic composition for inhibiting checkpoints containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0402] As used herein, the term “cosmetic composition” may be prepared in the form of a general emulsified or solubilized formulation. Examples of the emulsified formulation include nourishing toners, creams, essences, and the like, and examples of the solubilized formulation include toners, and the like. The cosmetic composition may be prepared into a formulation selected from the group consisting of solutions, suspensions, emulsions, pastes, gels, creams, lotions, powders, soaps, cleansing agents containing surfactants, oils, ampoules, powder foundations, emulsified foundations, wax foundations, and sprays, but is not limited thereto. Specifically, the cosmetic composition may be prepared as a formulation of a non-irritating cosmetic skin protectant, skin toner, nourishing toner,

[0403] DB1 / 161378040.3 MLB Docket No. 127033-5008 nourishing cream, massage cream, essence, eye cream, serum, cleansing cream, cleansing foam, cleansing water, pack, spray, or powder.

[0404] In addition, the cosmetic composition may further contain at least one cosmetically acceptable carrier that is mixed with general skin cosmetic materials and may be appropriately prepared by mixing common ingredients such as oils, water, surfactants, moisturizers, lower alcohols, thickeners, chelating agents, pigments, preservatives, fragrances, and the like, but is not limited thereto.

[0405] The cosmetically acceptable carrier contained in the cosmetic composition varies depending on the formulation.

[0406] When the formulation of the cosmetic composition is an ointment, paste, cream, or gel, the carrier ingredients may include an animal oil, vegetable oil. wax, paraffin, starch, tragacanth, cellulose derivatives, polyethylene glycol, silicone, bentonite, silica, talc, zinc oxide, or a mixture thereof.

[0407] When the formulation of the cosmetic composition is a powder or spray, the carrier ingredients may include lactose, talc, silica, aluminum hydroxide, calcium silicate, polyamide powder or a mixture thereof, and particularly, when the formulation of the cosmetic composition is a spray, a propellant such as chlorofluorohydrocarbon, propane / butane or dimethyl ether may be further contained.

[0408] When the formulation of the cosmetic composition is a solution or an emulsion, the carrier ingredients may include a solvent, a solubilizer or an emulsifier, for example, water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, or 1,3- butyl glycol oil, and particularly, cottonseed oil, peanut oil, com germ oil, olive oil, castor oil and sesame oil, glycerol aliphatic ester, polyethylene glycol or fatty acid ester of sorbitan.

[0409] When the formulation of the cosmetic composition is a suspension, the carrier ingredients may include liquid diluents such as water, ethanol or propylene glycol, suspending agents such as ethoxylated isostearyl alcohol, polyoxyethylene sorbitol ester and polyoxyethylene sorbitan ester, microcrystalline cellulose, aluminum metahydroxide, bentonite, agar or tragacanth, or the like.

[0410] When the formulation of the cosmetic composition is soap, the carrier ingredients may include alkali metal salts of fatty acids, fatty acid hemiester salts, fatty acid protein hydrolysates, isethionates, lanolin derivatives, aliphatic alcohols, vegetable oils, glycerol, sugars, and the like.

[0411] In accordance with another aspect of the present disclosure, there is provided a quasi-drug composition for treating denatured collagen containing the collagen hybridization peptide (CHP) as an active ingredient.

[0412] In accordance with another aspect of the present disclosure, there is provided a quasi-drug composition for inhibiting or reducing an inflammatory response containing the collagen hybridization peptide (CHP) as an active ingredient.

[0413] In accordance with another aspect of the present disclosure, there is provided a quasi-drug

[0414] DB1 / 161378040.3 MLB Docket No. 127033-5008 composition for downregulating one or more immune-related cytokines selected from the group consisting of TLR ligands, interleukins, IFN-a, TLR-9, TNF-a, IL-ip, IL-6, CCL3, IL-16, IL-18, and TNF-a containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0415] In accordance with another aspect of the present disclosure, there is provided a quasi-drug composition for preventing immune cell differentiation containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0416] In accordance with another aspect of the present disclosure, there is provided a quasi-drug composition for downregulating T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0417] In accordance with another aspect of the present disclosure, there is provided a quasi-drug composition for preventing differentiation of T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0418] In accordance with another aspect of the present disclosure, there is provided a quasi-drug composition for preventing or treating a dry eye disease containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0419] In accordance with another aspect of the present disclosure, there is provided a quasi-drug composition for preventing or treating atopic dermatitis containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0420] In accordance with another aspect of the present disclosure, there is provided a quasi-drug composition for preventing or treating osteoarthritis containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0421] In accordance with another aspect of the present disclosure, there is provided a quasi-drug composition for preventing or treating rheumatoid arthritis containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0422] In accordance with another aspect of the present disclosure, there is provided a quasi-drug composition for preventing or treating diabetic lesions containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0423] In accordance with another aspect of the present disclosure, there is provided a quasi-drug composition for preventing or treating psoriasis containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0424] In accordance with another aspect of the present disclosure, there is provided a quasi-drug composition for inhibiting checkpoints containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0425] In accordance with another aspect of the present disclosure, there is provided a quasi-drug

[0426] DB1 / 161378040.3 MLB Docket No. 127033-5008 composition for healing wounds including diabetic lesions containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0427] In accordance with another aspect of the present disclosure, there is provided a quasi-drug composition for preventing or treating GI inflammatory conditions including Crohn’s disease and ulcerative colitis containing the collagen hybridizing peptide (CHP) as an active ingredient.

[0428] As used herein, the term ‘"quasi-drug” refers to a product that is used for the purpose of diagnosing, treating, ameliorating, alleviating, curing, or preventing diseases of humans or animals, and has a milder effect than drugs. For example, according to the Pharmaceutical Affairs Act of the Republic of Korea, quasi-drugs are products other than those used for the purpose of drugs and include products that are used for the treatment or prevention of diseases of humans or animals and have a milder effect on the human body or do not directly affect the human body, and the like.

[0429] The quasi-drug composition according to the present disclosure may be prepared in a fonn selected from the group consisting of body cleansers, foams, soaps, masks, ointments, creams, lotions, essences, and sprays, but is not limited thereto.

[0430] For example, when the composition containing CHP according to the present disclosure is used as a quasi-drug additive, the composition containing CHP according to the present disclosure maybe added as is or used in combination with other quasi-drugs or quasi-drug ingredients, and may be used appropriately according to a conventional method.

[0431] In accordance with another aspect of the present disclosure, there is provided a method of reducing or preventing downregulation of immune cells including administering the composition containing the collagen hybridization peptide as an active ingredient to a subject in need thereof.

[0432] In accordance with another aspect of the present disclosure, there is provided a method of preventing or treating cancer including administering the composition containing the collagen hybridization peptide as an active ingredient to a subject in need thereof.

[0433] For example, these aspects may be achieved using the CHP wherein (Z) is bioactive and the bioactive includes an amino acid sequence from 3 to 27 amino acid residues having bioactivity, wherein the bioactive includes at least one leukocyte-associated immunoglobulin-like receptor (LAIR- 1 and / or LAIR-2) binding sequence, and wherein the amino acid sequence of the bioactive includes at least one citrulline residue.

[0434] The CHP containing a LAIR- 1 / 2 binding sequence including citrullinated arginine may bind to LAIR- 1 / 2 expressed on immune cells such as T cells, macrophages, natural killer cells, B cells, and dendritic cells to maintain the active immune cells, and may further effectively identify and destroy cancer cells in a tumor microenvironment.

[0435] In accordance with another aspect of the present disclosure, there is provided a method of

[0436] DB1 / 161378040.3 MLB Docket No. 127033-5008 reducing, preventing or treating oxidative modification of natural collagen including administering the composition containing the collagen hybridization peptide as an active ingredient to a subject in need thereof.

[0437] In accordance with another aspect of the present disclosure, there is provided a method of reducing, preventing or treating oxidative modification of natural collagen in a subject with diabetes including administering the composition containing the collagen hybridization peptide as an active ingredient to the subject in need thereof.

[0438] In accordance with another aspect of the present disclosure, there is provided a method of reducing, preventing or treating oxidative modification of natural collagen in a subject with diabetic ketoacidosis including administering the composition containing the collagen hybridization peptide as an active ingredient to the subject in need thereof.

[0439] In accordance with another aspect of the present disclosure, there is provided a method of reducing, preventing or treating oxidative modification of natural collagen in a subject with hyperglycemic hyperosmolar syndrome including administering the composition containing the collagen hybridization peptide as an active ingredient to the subject in need thereof.

[0440] In accordance with another aspect of the present disclosure, there is provided a method of reducing, preventing or treating oxidative modification of natural collagen in a subject with acidosis including administering the composition containing the collagen hybridization peptide as an active ingredient to the subject in need thereof.

[0441] In accordance with another aspect of the present disclosure, there is provided a method of reducing, preventing or treating oxidative modification of natural collagen in a subject with homocystinuria including administering the composition containing the collagen hybridization peptide as an active ingredient to the subject in need thereof.

[0442] In accordance with another aspect of the present disclosure, there is provided a method of reducing, preventing or treating oxidative modification of natural collagen in a subject with methylmalonic acidemia including administering the composition containing the collagen hybridization peptide as an active ingredient to the subject in need thereof.

[0443] In accordance with another aspect of the present disclosure, there is provided a method of reducing, preventing or treating diabetic lesions in a subject including administering the composition containing the collagen hybridization peptide as an active ingredient to the subject in need thereof.

[0444] In accordance with another aspect of the present disclosure, there is provided a method of reducing, preventing or treating diabetic retinopathy in a subject including administering the composition containing the collagen hybridization peptide as an active ingredient to the subject in need thereof.

[0445] DB1 / 161378040.3 MLB Docket No. 127033-5008

[0446] For example, the aspects may be achieved using CHP that includes any amino acid capable of post-translational modification by a sugar in a glycation reaction as a bioactive in (Z).

[0447] Specifically, the CHP preferentially takes up AGEs compared to natural collagen, glycolates, separates and discharges the same, thereby maintaining a healthy vascular system.

[0448] In accordance with another aspect of the present disclosure, there is provided a method of providing information for diagnosing fibrosis in a subject including administering the composition containing the collagen hybridization peptide as an active ingredient to the subject, and imaging the collagen hybridization peptide containing the detection moiety in vivo to determine the presence or progression of fibrosis in the subject.

[0449] In another embodiment, the fibrosis is selected from the group consisting of neovascular age- related macular degeneration (nAMD), diabetic retinopathy, glaucoma, particularly fibrosis in the trabecular meshwork, neovascular glaucoma, corneal scarring, conjunctiva, post-cataract surgery, retinopathy of prematurity, and proliferative vitreoretinopathy.

[0450] As used herein, the term “food” includes meat, sausage, bread, chocolate, candy, snacks, confectionery, pizza, ramen, other noodles, gum, dairy products including ice cream, various soups, beverages, tea, drinks, alcoholic beverages, vitamin complexes, health functional foods, and health foods, and includes all foods in the conventional sense.

[0451] For example, since the food composition is consumed on a daily basis, it is expected to be highly effective in amelioration of diseases and is very useful to improve health. In addition, the food composition may further contain a bromatologically acceptable food additive.

[0452] As used herein, the term “functional food” has the same meaning as the term “food for special health use (FoSHU)” and refers to a food having strong medical and pharmaceutical effects that has been processed to efficiently provide bioregulatory functions as well as nutrition supply functions. As used herein, the term “functional” means obtaining beneficial effects for health purposes, such as controlling nutrients or exhibiting physiological effects with regard to the structures and functions of the human body. The food may be prepared by a method commonly used in the art, and the preparation may be performed using raw materials and ingredients commonly added in the art. The food composition may be prepared into various fonnulations and, unlike general drugs, it is advantageously free of side effects that may occur upon long-term take of drugs since it uses food as a raw material, and is highly portable, and thus can be consumed as a supplement to enhance the effect of ameliorating diseases.

[0453] The term “health food” refers to a food having an effect of actively maintaining or improving health beyond that of a general food, and the temi “health supplement food” refers to a food ingested for the purpose of health supplement. In some cases, the terms “health functional food,” “health food,” DB1 / 161378040.3 MLB Docket No. 127033-5008 and “health supplement food” are used interchangeably.

[0454] Specifically, the health food means a food product that is prepared by adding the compound to food materials such as beverages, teas, spices, gums, and confectionery, or by preparing the compound in the form of a capsule, powder, suspension, or the like, and has a specific health effect when consumed, and is advantageously free of side effects that may occur when taking drugs for a long period of time because it is prepared from food as a raw material unlike general drugs.

[0455] The food composition may further contain a physiologically acceptable carrier. There is no particular limitation as to the type of carrier, and any carrier may be used as long as it is commonly used in the relevant art.

[0456] In addition, the food composition may contain additional ingredients commonly used in food compositions to improve fragrance, flavor, aesthetics and the like. For example, the food composition may contain vitamins A, C, D, E, Bl. B2. B6, B12. niacin, biotin, folate, pantothenic acid, or the like. Further, the food composition may contain minerals such as zinc (Zn), iron (Fe), calcium (Ca), chromium (Cr), magnesium (Mg), manganese (Mn), copper (Cu), and chromium (Cr); and amino acids such as lysine, tryptophan, cysteine, and valine.

[0457] In addition, tire food composition may contain food additives such as preservatives (potassium sorbate, sodium benzoate, salicylic acid, sodium dehydroacetate, and the like), bactericides (bleaching powder and highly bleaching powder, sodium hypochlorite, and the like), antioxidants (butylated hydroxyanisole, BHA), butylated hydroxytoluene (BHT and the like), colorants (coal tar dyes, and the like), color developers (sodium nitrite, sodium nitrite, and the like), bleaching agents (sodium sulfite), seasonings (MSG, monosodium glutamate, and the like), sweeteners (dulcin, cyclamate, saccharin, sodium, and the like), flavorings (vanillin, lactones, and the like), leavening agents (alum, D-potassium hydrogen tartrate, and the like), reinforcing agents, emulsifiers, thickeners (glutinating agents), coating agents, gum bases, de-foaming agents, solvents, and enhancers. The additives may be selected depending on the type of food and used in an appropriate amount.

[0458] When the food composition is used as a food additive, the composition may be added as it is or used in combination with other foods or food ingredients, and may be used appropriately according to a conventional method. Tire amount of the active ingredient may be appropriately determined depending on the purpose of use. such as prevention, health, or therapeutic treatment. In general, the composition is added in an amount of 15 wt% or less, preferably 35 wt% or less, based on the weight of raw material in the process of preparing a food or beverage. However, in case of long-term intake for health and hygiene or health control, the amount of active ingredients may be below the range defined above.

[0459] In an embodiment, the food composition may be used as a health beverage composition. In this case, the food composition may contain various flavoring agents or natural carbohydrates as

[0460] DB1 / 161378040.3 MLB Docket No. 127033-5008 additional ingredients, like a conventional beverage. The natural carbohydrates include monosaccharides such as glucose and fructose; disaccharides such as maltose and sucrose; polysaccharides such as dextrin and cyclodextrin; and sugar alcohols such as xylitol, sorbitol, or erythritol. Sweeteners may include natural sweeteners such as thaumatin and stevia extracts, synthetic sweeteners such as saccharin or aspartame, and the like. The content of the natural carbohydrate may be generally about 0.01 to 0.04 g. specifically about 0.02 to 0.03 g. with respect to 100 mL of the health beverage composition.

[0461] Furthermore, the health beverage composition may contain various nutrients, vitamins, electrolytes, flavors, coloring agents, pectic acid, salts of pectic acid, alginic acid, salts of alginic acid, organic acids, protective colloid thickeners, pH regulators, stabilizers, preservatives, glycerin, alcohol, or carbonating agents. In addition, the health beverage composition may contain fruit pulp for the preparation of natural fruit juice, fruit juice beverage, or vegetable beverage. These ingredients may be used alone or in combination. The content of these additives is not particularly important, but is typically selected within the range of 0.01 to 0.1 parts by weight with respect to 100 parts by weight of the health beverage composition.

[0462] The novel CHP design of the present disclosure is intended for use in research and diagnosis of cosmetics and therapeutics that keep the monomeric form thereof active at room temperature (20-22°C), remain stable during storage, and still retain high binding affinity for damaged / denatured / remodeled collagen. It was found that, when the CHP is stored at room temperature, it can keep the monomeric form active and can still bind to denatured collagen when applied to the skin (~32-36°C). Therefore, the CHP is essential for topically applied cosmetics. For cosmetic applications, only natural amino acids are used due to the low production margins and the ability thereof to claim to be “totally natural" since all amino acids are naturally synthesized in vivo. Cosmetic applications include anti -aging, antiwrinkle. age prevention, or moisturization. These cosmetics may be applied topically.

[0463] Two examples of cosmetic bioACTIVE CHPs disclosed herein include integrin binding motifs having different mechanisms of action. One cosmetic application of the C-terminally truncated CHP involves disposing the bioACTIVE sequence in the middle of the CHP, as shown in FIG. 1. Since collagen exists as a stable triple-helix structure when intact, most cellular interactions with cell signaling sites found in collagen require a triple-helix structure. A specific bioactive moiety is disposed in the center of the CHP binding domain, which ensures that, when CHP binds to a damaged single chain collagen strand to form a triple helix, the bioACTIVE integrin moiety will have a correct structure and will be recognized by cells via a2pi and aipi integrins. This design is disclosed in the previous PCT application (PCT / US2022 / 075819) of the present applicant. Other bioACTIVE CHP designs that incorporate the RGD sequence are implemented by delivering the active site to the monomer through

[0464] DB1 / 161378040.3 MLB Docket No. 127033-5008 interaction with av 1 and av 3 integrins.

[0465] When the ECM is damaged due to disease, injury, or natural aging, collagen strands, which are a major component of the ECM, are enzymatically cleaved by a family of matrix metalloproteinases (MMPs) or damaged by mechanical forces (e g., muscle rupture, skin abrasion / cut, bone fracture). Once collagen is damaged and the triple-helix structure thereof is lost, it is continuously denatured at body temperature, leaving a mechanically harsh microenvironment on cells. The ECM exhibits mechanical stability, is biologically active, and interacts and communicates with cells through a number of receptors, including integrins, OSCAR, GPIV, DDR, LAIR, and uPARAP / Endol80 . Many integrin binding sites, such as GFOGER, require a triple-helix structure to be recognized by and interact with cells. However, the RGD (Arg-Gly-Asp) cell attachment site does not have these structural limitations and does not need to be a triple helix to be recognized by cells. Cells may be attached to denatured collagen via av03 and avpi integrins that recognize the RGD site. According to the literature, when the level of collagen damage is high, more RGD sites are exposed and may interact with cells. Physiologically, this provides a method for cells to enter the affected area, attach, and initiate damage repair by first removing the damaged portion and then generating fresh collagen to replace the damaged portion.

[0466] By providing more RGD sites in the damaged area, more cells may be introduced into the corresponding area to facilitate wound healing. Interestingly, adding the RGD sequence alone to the solution (without binding ability) inhibits the ability of cells to attach to the ECM due to the superior soluble form thereof compared to the RGD sequence still attached to the damaged collagen strands. Therefore, when the cellular receptor for RGD binds to the soluble RGD, it can no longer interact with the RGD sequence attached to collagen, and thus the level of cell adhesion decreases. For wound healing, RGD-CHP may be preloaded onto 3D tissue scaffolds to form a “super collagen” mesh for cell migration into the wound, may be added directly to the wound as a solution to increase the local concentration of cell attachment sites at the edge surrounding the wound, or a combination of both methods. The RGD-CHP may be more readily attached to collagen scaffolds, which have better biomimetic properties compared to artificial polymer-based scaffolds, due to the different active sites contained in natural collagen. Uris approach depends on the ability of CHP to attach to damaged collagen in a harsh cellular microenvironment and the signaling of RGD to aid cell attachment to the ECM.

[0467] The C-terminally truncated CHP of the present disclosure may be combined with many bioACTIVE moieties with a variety of applications, such as increased collagen production for anti-aging or anti -wrinkle effects.

[0468] The inclusion of unnatural amino acids provides beneficial properties that are more suitable for DB1 / 161378040.3 MLB Docket No. 127033-5008 therapeutic or diagnostic purposes, such as avoiding recognition by enzymatic cleavage (greater enzyme stability), extending retention time at the target site, and increasing minimum effective concentration (MEC) or maximum tolerated dose (MTD). These properties are still being investigated. Some identified applications may include atopic dermatitis, psoriasis, dry eye disease, age-related macular degeneration, wound healing, and cancer therapeutics. While these applications are integrated into different bioACTIVE moieties, the C-terminal truncations allow for fine-tuning of the properties of the bioACTIVE CHP for targeting, binding, and stability at or about body temperature.

[0469] Finally, the novel C-terminal design may be used for further development of current RUO production lines to optimize binding time or concentration to achieve optimal signal intensity for imaging. This may be useful for automated tissue staining platforms at a part of the cost of previous designs involving unnatural amino acids.

[0470] The C -terminally truncated CHP according to some embodiments has several advantages over conventional CHP designs.

[0471] 1. The new design affects only the nucleation site of the CHP fold, but requires only one or two unfavorable amino acid residues, so the region where negative influence is present is much shorter than that of conventional (GfO)g CHPs, which have steric hindrance throughout the entire sequence to achieve similar results in self-trimerization resistance. Thus, the new design can maintain high affinity in most regions within damaged collagen, excluding regions with potentially similar charges or regions containing amino acids with large side chains. By affecting only one or two amino acid residues on the C -terminus, residues with high folding ability into the middle or N-terminus of the CHP sequence maybe integrated, and folding and binding ability to naturally degraded collagen may be increased. For example, the use of proline at the position X or hydroxyproline at the position Y of the Gly-X-Y motif provides high binding affinity.

[0472] 2. The present applicant generally focuses only on influencing self-trimerization rather than binding. Most approaches affect binding to self and damaged collagen.

[0473] 3. The present applicant focuses on the C -terminus, which is the area where the peptide and collagen are folded. Others focus on the entire peptide.

[0474] 4. Compared to (GPP)? CHP of Stuart Therapeutics, the design of the present application allows for greater customization of the amino acid sequence that may be integrated. Meanwhile, the design thereof alters the sequence with amino acids that further interfere with the triple helix fold, resulting in a peptide that already has a low melting point (Tm) and therefore has little or no binding affinity for naturally denatured collagen.

[0475] 5. Tire bioACTIVE CHP requires high binding affinity for native collagen (required to have a triple-helix structure) so that cells can recognize the active site, but must resist self-trimerization at a

[0476] DB1 / 161378040.3 MLB Docket No. 127033-5008 temperature of 20 to 22°C to maintain the active state at room temperature.

[0477] 6. The cyclized peptide developed by Waseda University resists self-trimerization, but the (GfO)gCHP of the present applicant exhibited worse binding performance to denatured collagen in tissue sections when tested. This may be due to the larger size of the collagen strands and potentially some steric hindrance upon access to the binding site of the peptide. In addition, since this is a cyclized peptide, the synthesis thereof is more complicated, which increases the cost for mass production.

[0478] 7. RGD-CHP may provide an alternative mechanism for integrin activation and may be easily used to decorate collagen-based scaffolds or meshes without strong cross-linkers.

[0479] An important feature of the novel peptide of the present disclosure is the provision of CHP that blocks self-trimerization while maintaining binding affinity to the target by introducing bulky or charged amino acid residues at the C-terminus of the collagen hybridization peptide sequence, so that the CHP is useful for prevention or treatment of related diseases, as well as diagnosis by combination with various bioactive sequences.

[0480] BRIEF DESCRIPTION OF THE DRAWINGS

[0481] The above and other objects, features and other advantages of the present disclosure will be more clearly understood from the following detailed description taken in conjunction with the accompanying drawings, in which:

[0482] Figure 1. Schematic diagram illustrating the design of a C-terminally truncated CHP according to one embodiment of the present disclosure.

[0483] Figure 2. Circular Dichroism Evaluation. (A) The thermal melting curve evaluating the melting temperatures of the C-terminally modified CHPs compared to a sequence without a bulky or charged residue on the C-terminal. The two CHPs without GPO triplets and a disrupted or modified C- terminus (red and green traces) showed no thermal transition from a higher order to more disorder, as is evidenced by a lack of a sigmoidal curve, and instead showed a linear decrease over the temperature range. (B) This graph depicts the first derivative of the thermal melting curves of the three CHPs that showed a thermal transition in (A). The minimum point of the first derivative curve corresponds to the melting temperature (Tm) of the homotrimer, which is the inflection point where 50% of the CHPs are in a triple helix and 50% of the CHPs are in a monomeric fonn. All CHPs are 33 amino acids in length and were run at a concentration of 150 pM. They were allowed to fold at 4°C for 72 hours prior to being run. CD measurements were taken from 4-90°C at a rate of 0.5°C / min and evaluated at 225 nm.

[0484] Figure 3. CHP staining 4 hours @ 37°C.

[0485] Figure 4. Control staining 4 hours @ 37°C. Human skin tissue stained with positive and

[0486] DB1 / 161378040.3 MLB Docket No. 127033-5008 negative controls. The Biotin-GFOGER hexapeptide (left) shows no signal, indicating no binding to denatured collagen. This result is expected since that sequence alone is not long enough to form a triple helix with denatured collagen. Alternatively, the CHP does bind with denatured collagen, represented by the green color. This CHP does not have a disrupted or modified C-terminus, and we expected this to bind. The signal intensity seen is similar to the signal intensities seen for the C- terminally disrupted or modified CHPs. which resist self-trimerization (shown in figure 4).

[0487] Figure 5. 45°C binding. Human tissue staining with biotin-labeled CHPs overnight at 45°C. These histology images show that even though they have weak affinity to form a homotrimer with themselves, they are still able to bind to denatured collagen and remain bound even at temperatures above their melting temperature. Although the CHP without a disrupted or modified C-terminus had the most binding, each C-terminally disrupted or modified CHP tested here still had strong signal intensity at 45°C, and the CHPs that included two extra GPO repeats showed higher signal intensity than CHPs without the triplet to enhance binding.

[0488] Figure 6. Schematic representation of the RGD-Ahx-(GPP)8 CHP. The design incorporates a triple helical CHP region (right), a linker (middle), and an active site sequence (left). Each component has several alternatives that can be used.

[0489] Figure 7. CD Spectra and melting curve. Circular dichroism analysis of RGD-Ahx-(GPP)8. (a) The CD spectra evaluating the secondary structure of the peptide, the peak at 225nm is the characteristic peak for the collagen triple helix, (b) The thermal melting curve evaluated at 225 nm. The sigmoidal transition indicated the CHP is going from a higher order structure to more disorder as the temperature increases. The inflection point of the transition is the melting temperature (Tm) where 50% of the CHP is folded in a triple helix and 50% of the CHPs are in a monomeric form.

[0490] Temperature readouts were taken from 4-90°C at a heating rate of 0.5°C / min.

[0491] Figure 8. First Derivative of Melting Curve. First derivative of thermal melting curve. This confirms the absolute melting temperature of the CHP. The local minima show the melting temperature of the CHP sequence, which was found to be 35.01°C.

[0492] Figure 9. FLAIR2 Binding Concentration Curve. FLAIR2 shows a dose-dependent response for recombinant LAIR binding where the scramble does not, and CHP2HmT begins to self-trimerize at higher concentrations.

[0493] Figure 10. FLA1R2 reduces T cell activation in non-linear dose-dependent manner. FLA1R2 shows the same shape of dose response shown by Linde Meyyard (discover of LAIR) with collagen — inverse due to readout type.

[0494] Figure 11 . Alamar Blue - FLAIR2. Alamar Blue is a cell viability assay commonly used in biological research to assess cell proliferation, cytotoxicity, and metabolic activity. No change in the DB1 / 161378040.3 MLB Docket No. 127033-5008 cell viability over various concentrations of FLAIR2. Suggesting high levels of safety.

[0495] Figure 12. CHP2L Scramble Control.

[0496] Figure 13. Biotin-CHP Positive Control.

[0497] Figure 14. CHP2L.

[0498] Figure 15. HT2 / CHP 2.2 / FLAIR2.

[0499] Hereinafter, the present disclosure will be described in more detail with reference to the following examples. However, it will be obvious to those skilled in the art that the following examples are provided only for illustration of the present disclosure and should not be construed as limiting the scope of the present disclosure.

[0500] EXAMPLES

[0501] Example 1: Design of Sequence

[0502] The prior LAIR bioACTIVE CHP developed by the present applicant includes the following sequence:

[0503] Ac-S-(Gly-X-Y)n-bioactive-(Gly-X-Y)mwherein Ac is an acetyl capping group; S is 0 to 10 spacer molecules: X is proline or modified proline; Gly is glycine; Y is hydroxyproline or any other amino acid: n is a natural number of 1 to 20; m is a natural number of 1 to 20; and bioactive is an amino acid sequence including 3 to 27 amino acid residues having bioactivity.

[0504] In a specific embodiment of the present disclosure, a novel C-tenninally truncated CHP was constructed by incorporating amino acid residues that have the same basic design as the conventional design, but interfere with nucleation of the triple helix fold at the C-tenninus thereof (FIG. 1).

[0505] Example 2: Examples of Specific Sequence Designs and Melting Point Testing

[0506] In some embodiments, Biotin-G3-(GPP)4-GFOGER-(GPP)3-GFF (SEQ ID NO: 122) containing the bulky group phenylalanine (F) or Biotin-G3-(GPP)4-GFOGER-(GPP)3-GEE (SEQ ID NO: 123) containing the charged group glutamate (E) were tested as novel C -terminally truncated CHPs. The last two residues of the sequence used to demonstrate how interrupting the C-tenninal nucleation affects the ability to self-trimerize were Phe or Glu. These CHPs were folded at 4°C for 72 hours prior to CD evaluations to allow sufficient time for folding. The CHP (charged or bulky) has no Tm when measured by CD over the range of 4 to 90°C using a step size of 0.5°C / min (FIG. 2, red and green solid lines). This shows the great impact of alteration of the C-tcrminus on the overall stability, since the "normal" CHP sequence Biotin-G3-(GPP)4-GFOGER-(GPP)4 (SEQ ID No: 127) has a Tm of 32.5°C.

[0507] DB1 / 161378040.3 MLB Docket No. 127033-5008

[0508] The method for engineering the CHP sequence having a truncated C-terminus was further evaluated to achieve higher binding affinity to collagen. CHP samples including two GPP repeats replaced with GPO repeats, Biotin-G3-(GPP)3-GPO-GFOGER-GPO-(GPP)2-GEE (SEQ ID NO: 124) and Biotin-G3- (GPP)3-GPO-GFOGER-GPO-(GPP)2-GFF (SEQ ID NO: 125), were tested. Trn was measured using CD spectroscopy, and the result showed that the CHP had a Tm of 29°C and 28°C. respectively (FIG. 2, blue and pink solid lines). In order to determine whether or not these CHPs retained the ability thereof to bind to denatured collagen strands, human skin tissue was stained and imaged by fluorescence microscopy. The result of staining showed that the CHPs still retained the ability thereof to be folded into triple helices with denatured collagen strands, despite not self-trimerizing (FIG. 3). Compared to the positive control (Biotin-G3-(GPP)4-GFOGER-(GPP)4 CHP; SEQ ID NO: 126) and the negative control (Biotin-GFOGER hexapeptide) (FIG. 4), the C-terminally modified CHPs were found to exhibit qualitatively higher signal intensities. Overall, this initial data set demonstrated that simply introducing a charged or bulky group at the C-terminus thereof drastically affects the self-trimerization ability. The binding ability to denatured collagen was analyzed in human skin tissue. The result showed that they had similar or slightly higher signal intensities compared to “normal” Biotin-G ,- (GPP)4-GFOGER-(GPP)4, which has a 4.5°C higher Tm. Furthermore, the binding ability of these novel CHP designs was tested while increasing the temperature at which the binding is possible in tissue. It was found that the CHPs were capable of maintaining a high level of binding even at 45°C (FIG. 5).

[0509] Example 3: CHP having bioactive moiety at N-terminus and characterization thereof

[0510] The RGD bio ACTIVE moiety is attached to the N-terminus of CHP (FIG 7). Tire mechanism of action to recognize the RGD integrin moiety is different from that of other sequences and thus is an important distinction. Here, the active moiety does not need to have a triple-helix structure, and thus is attached to the N-terminus of the CHP sequence, rather than being located in the middle of the CHP sequence, thus providing a substantial advantage. The CHP sequence according to the present disclosure is RGD-AHX-(GPP)S-GXY, wherein AHX is 6-aminohexanoic acid (used to provide flexibility so that the RGD moiety can be accessed and recognized by cells), the repeating glycine (G)- proline (P)-proline (P) sequence enables the RGD moiety to be localized and anchored in the extracellular matrix (ECM), and X and Y are any charged or bulky amino acid residues such as phenylalanine, glutamic acid, tryptophan, and arginine that prevent self-trimerization. The (GPP)s CHP base targets and binds to each alpha helix of damaged collagen molecules within the ECM, thus localizing the RGD active site to promote cell homing and adhesion. Tire (GPP)s amino acid sequence was selected not only for the ability thereof to bind to damaged collagen, but also for the lack of post-

[0511] DB1 / 161378040.3 MLB Docket No. 127033-5008 translationally modified residues (meaning that they could be produced by yeast or bacteria) such as hydroxyproline, and has a low melting point (35.01°C). This indicates that the CHP is further modified by introducing truncation into the C-terminus thereof and thus has the ability to resist self- trimerization at body temperature (FIGS. 7 to 8).

[0512] However, a more comprehensive version of RGD-CHP may be represented by the following formula: L-S-(Gly-X-Y)n -Gly-Z-Z; wherein L is a bioactive moiety including an amino acid sequence including 3 to 10 residues in length; S is one or more spacer molecules, X is proline or a modified proline, Y is any amino acid, or proline, hydroxyproline or any modified proline, n means the number of repetitions of the CHP binding domain and is a natural number of 4 to 15, and Z is a charged amino acid residue or an amino acid residue having a bulky side chain.

[0513] Example 4: Additional modification possibilities

[0514] There are several elements of the CHP design that may be modified or adjusted and still maintain the desired effect. In fact, the present applicant further studied the following modifications or combinations of modifications that may be used to optimize the difference between low self-binding and high denatured collagen binding:

[0515] 1. It is known that the overall CHP length may be varied and, as the number of Gly-X-Y repeats increases, the binding ability (of CHP to all denatured collagen) increases.

[0516] 2. More specifically, the triple helix CHP region that binds to damaged collagen may be modified as long as it contains a Gly-X-Y motif in its sequence. For hybridization with damaged collagen, when X is proline and Y is hydroxyproline, the sequence has been shown to have a ven high binding affinity. However, the amino acid residues at positions X and Y may be manipulated to adjust the overall Tm of the CHP to suit a specific purpose. The number (n) of Gly-X-Y repeats may range from 4 to 15 depending on the desired properties of the CHP.

[0517] 3. The number of bulky or charged amino acids (e.g., Phe or Glu) at the C-terminus may be increased or decreased.

[0518] 4. Tire position of the bulky or charged amino acid may be shifted from the very end of the C- tenninus to three or four residues inward of the sequence. For example, instead of having a sequence terminus of . . . GPP-GPP-GZZ (wherein Z is any charged or bulky amino acid residue), the sequence may be terminated with GPP-GPZ-GPZ, GPP-GZZ-GPP. GPP-GZP-GZP, or the like.

[0519] 5. Aspartic acid, lysine, arginine or histidine may be replaced as a charged residue, instead of glutamic acid. In addition, phenylalanine may be replaced with tryptophan or tyrosine, which also contain a bulky phenyl ring in the side chain.

[0520] 6. A mixture of the aforementioned amino acids may be used at three or four different

[0521] DB1 / 161378040.3 MLB Docket No. 127033-5008 positions on the C-terminus of CHP.

[0522] 7. Unnatural amino acids may be introduced into the center or N-terminus of CHP in order to increase the binding affinity of the sequence, but still terminate the C-terminus in the manner described above. Some residues of interest are 2S.4R-4-fluoroproline as a trans form. When the amino acid is disposed at the Y position, it actually supports proper pre-alignment at the phi-psi angle of poly-proline, such as a poly -proline type II helix, which allows for better trimerization. This is due to the chlorine atom having a stronger electron-withdrawing group compared to the hydroxyproline residues that are usually placed at the position Y. Another unnatural amino acid that may be tested is the aza-glycine residue, which increases the triple helix stability of the formed helix by increasing the formation of cross-strand hydrogen bonds. In addition to inclusion of these residues in the CHP sequence, the number of these residues included in the sequence, the position thereof in the CHP sequence, and the mixing thereof in a single CHP sequence may be varied.

[0523] 8. There are several RGD derivatives that include different amino acids at both ends of the sequence that affect cellular responses. The RGD moiety is recognized by cells on collagen, fibronectin, and vitronectin. Therefore, research has been conducted to investigate the affinity of the RGD moiety based on the amino acids surrounding the RGD triplet. Therefore, cell attachment may still be achieved using other RGD-containing sequences. Some sequences include RGDS (fibronectin), GRGD, GRGDSP, GRGDG, or RGDV (vitronectin). In addition, cyclized versions of RGD have also been shown to be effective.

[0524] 9. The length of the linker used may affect the ability of the RGD moiety to promote cell attachment. According to the literature, RGD must be able to fit inside the globular head on the cell receptor, and linkers of 11-45 A in length may affect cell attachment.

[0525] 10. The type of linker may also be changed. Although the use of Ahx (6-aminohexanoic acid) has been described above, other linkers such as 12-aminododecanoic acid and N-Fmoc-N"- succinyl-4,7,10-trioxa-l,13-tridecanediamine (TTDS) or even polymeric linkers such as PEG may be used.

[0526] Unless otherwise specified, it should be understood that all numbers expressing quantities of ingredients, properties, such as molecular weights and reaction conditions, used in this disclosure and claims are modified in all cases by the term “about / ’ Accordingly, unless otherwise specified, the numerical parameters set forth in this disclosure and appended claims are approximations that may vary depending upon the desired properties sought to be obtained by the present disclosure. At the very least, and not as an attempt to limit the application of the doctrine of equivalents to the scope of the claims, each numerical parameter should at least be construed by applying ordinary rounding techniques

[0527] DB1 / 161378040.3 MLB Docket No. 127033-5008 in consideration of the number of significant digits reported.

[0528] Notwithstanding that the numerical ranges and parameters setting forth the broad scope of the present disclosure are approximations, the numerical values set forth in the specific examples are reported as precisely as possible. Any numerical value, however, inherently contains certain errors necessarily resulting from the standard deviation found in their respective testing measurements.

[0529] At the very least, and not as an attempt to limit the application of the doctrine of equivalents to the scope of the claims, each numerical parameter should at least be construed in light of the number of reported significant digits and by applying ordinary rounding techniques.

[0530] The grouping of alternative elements or embodiments of the present disclosure should not be construed as limiting. Each group member may be referred to and claimed individually or in combination with the other members of the group or other elements herein. It is anticipated that one or more members of a group may be included in, or deleted from, a group for reasons of convenience and / or patentability. When any such inclusion or deletion occurs, the disclosure is deemed to satisfy all Markush group descriptions used in the appended claims, including the modified group.

[0531] Specific embodiments of the present disclosure including the best mode known to the present inventors for carrying out the present disclosure are described herein. Of course, variations of the described embodiments will be apparent to those skilled in the art upon who read the foregoing descriptions. The present inventors expect that those skilled in the art employ such variations as appropriate and intend that the present disclosure is implemented otherwise than as specifically described herein. Accordingly, the present disclosure encompasses all modifications and equivalents of the subject matter recited in the claims appended hereto as permitted by the corresponding law. Furthermore, any combination of all possible variations of the elements described above is encompassed by the present disclosure unless otherwise indicated herein or otherwise clearly contradicted by context.

[0532] The specific embodiments disclosed herein may be further limited in the claims by the use of the phrases ‘'consisting of’ or “consisting essentially of.” When used in the claims, whether as filed or added by amendment, the transitional phrase “consisting of’ excludes any element, step, or ingredient not specified in the claim. The transitional phrase “consisting essentially of’ limits the scope of the claim to those materials or steps specified and those that do not substantially affect the basic and novel characteristic(s). The embodiments of the present disclosure so claimed are essentially or explicitly described and enabled herein.

[0533] It should be understood that the embodiments of the present disclosure disclosed herein are illustrative of the principles of the present disclosure. Other modifications that may be employed fall within the scope of the present disclosure. Thus, by way of example, and not limitation, alternative configurations of the present disclosure may be employed in accordance with the teachings herein.

[0534] DB1 / 161378040.3 MLB Docket No. 127033-5008

[0535] Accordingly, the present disclosure is not limited to what has been precisely set forth and described.

[0536] While the present disclosure has been described and illustrated herein with reference to various specific materials, procedures, and examples, it should be understood that the present disclosure is not limited to the specific combinations of materials and procedures selected for the purposes for which they are intended. Many variations of these details are contemplated, as will be appreciated by those skilled in the art. It is intended that the disclosure and examples be considered illustrative only, with the true scope and spirit of the present disclosure being indicated by the claims that follow. All references, patents, and patent applications cited herein are hereby incorporated by reference in their entirety.

[0537] From the foregoing description, it will be apparent to those skilled in the art that the present disclosure may be implemented in other specific forms without departing from the spirit or essential characteristics thereof. In this regard, it should be understood that the embodiments described above are illustrative in all respects and not restrictive. The scope of the present disclosure should be construed as encompassing within the scope of the present disclosure all changes or modifications derived from the meaning and scope of the following claims and equivalents thereof, rather than the foregoing detailed description.

[0538] EMBODIMENTS

[0539] Embodiment 1. A collagen hybridizing peptide (CHP), wherein the CHP comprises a sequence represented by Formula I:

[0540] (Gly-X-Y)a-(Z) (Formula I) in which X and Y are independently any amino acid, provided at least one of X and Y is proline, hydroxyproline, or modified proline in each instance of (Gly-X-Y); Gly is glycine; Z is an optional spacer; and a is a number from 1 to 40, and the CHP or the Formula I comprises a modification at the C-terminus, thereby lowering a melting temperature of the CHP.

[0541] Embodiment 2. The CHP according to embodiment 1, wherein the modification at the C-terminus comprises one or more molecules that yield a lower homotrimerization melting temperature for the CHP than that of the CHP bound with gelatin.

[0542] Embodiment 3. Tire CHP according to embodiment 1 or embodiment 2, wherein the modification at the C-terminus comprises one or more molecules that yield a lower homotrimerization melting temperature for the CHP than that of the CHP bound with damaged collagen.

[0543] Embodiment 4. The CHP according to any one of the preceding embodiments, wherein the modification at the C-terminus comprises attaching one or more amino acids to the C-terminus of the CHP or the Formula I.

[0544] Embodiment 5. The CHP according to any one of the preceding embodiments, wherein the

[0545] DB1 / 161378040.3 MLB Docket No. 127033-5008 modification at the C-terminus comprises attaching one or more amino acids including one or more sequences represented by Formula II to the C-terminus of the Formula I:

[0546] (Gly-B-B) (Formula II) in which B is independently in each instance phenylalanine, tryptophan, tyrosine, aspartic acid, lysine, arginine, histidine, or any charged amino acid.

[0547] Embodiment 6. The CHP according to any one of the preceding embodiments, wherein (Z) comprises at least one bioactive, the at least one bioactive comprising an amino acid sequence from 3 to 27 amino acid residues having bioactivity.

[0548] Embodiment 7. The CHP according to any one of the preceding embodiments, wherein the at least one bioactive comprises at least one sequence selected from the group consisting of a leukocyte- associated immunoglobulin-like receptor binding sequence (LAIR-1 and / or LAIR-2), an integrin binding sequence, a discoidin domain receptor (DDR) binding sequence, a von Willebrand factor (VWF) sequence, a matrix metalloproteinase (MMP) binding sequence, a G6b-B recognition site sequence, an OSCAR domain sequence, a GPVI sequence, and a FcR-y domain sequence.

[0549] Embodiment 8. The CHP according to any one of the preceding embodiments, wherein the at least one bioactive comprises a LAIR-1 receptor binding sequence.

[0550] Embodiment 9. The CHP according to any one of the preceding embodiments, wherein the at least one bioactive comprises a LAIR-2 receptor binding sequence.

[0551] Embodiment 10. The CHP according to any one of the preceding embodiments, wherein the at least one bioactive comprises a G6b-B recognition site sequence.

[0552] Embodiment 11 . The CHP according to any one of the preceding embodiments, wherein the at least one bioactive comprises a FcR-y domain sequence.

[0553] Embodiment 12. The CHP according to any one of the preceding embodiments, wherein the at least one bioactive comprises an OSCAR domain sequence.

[0554] Embodiment 13. The CHP according to any one of the preceding embodiments, wherein the at least one bioactive comprises a GPVI sequence.

[0555] Embodiment 14. The CHP according to any one of the preceding embodiments, wherein the at least one bioactive comprises an integrin binding sequence.

[0556] Embodiment 15. The CHP according to any one of the preceding embodiments, wherein the at least one bioactive comprises a DDR binding sequence.

[0557] Embodiment 16. The CHP according to any one of the preceding embodiments, wherein the at least one bioactive comprises a VWF sequence.

[0558] Embodiment 17. The CHP according to any one of the preceding embodiments, wherein the at least one bioactive comprises an MMP binding sequence.

[0559] DB1 / 161378040.3 MLB Docket No. 127033-5008

[0560] Embodiment 18. The CHP according to any one of the preceding embodiments, wherein the CHP comprises a sequence having at least 85% sequence identity to any one of SEQ ID NO: 1-27, 56-64, 93, 94, and 98-100.

[0561] Embodiment 19. The CHP according to any one of the preceding embodiments, wherein the CHP comprises a sequence having at least 85% sequence identity to any one of SEQ ID NO: 28-46, 95, and 96.

[0562] Embodiment 20. The CHP according to any one of the preceding embodiments, wherein the CHP comprises a sequence having at least 85% sequence identity to any one of SEQ ID NO: 47-55 and 97. Embodiment 21 . The CHP according to any one of the preceding embodiments, wherein the CHP comprises a sequence having at least 85% sequence identity to any one of SEQ ID NO: 65-83 and 101-103.

[0563] Embodiment 22. The CHP according to any one of the preceding embodiments, wherein the CHP comprises a sequence having at least 85% sequence identity to any one of SEQ ID NO: 84-92 and 104.

[0564] Embodiment 23. The CHP according to any one of the preceding embodiments, wherein the CHP comprises a sequence having at least 85% sequence identity to any one of SEQ ID NO: 107-114. Embodiment 24. The CHP according to any one of the preceding embodiments, wherein the CHP comprises a sequence selected from the group consisting of SEQ ID NO: 1-104 and 107-114. Embodiment 25. The CHP according to any one of the preceding embodiments, wherein (Z) comprises at least one linker.

[0565] Embodiment 26. The CHP according to any one of the preceding embodiments, wherein the at least one linker comprises one or more selected from the group consisting of Ahx (6-aminohexanoic acid), 12-aminododecanoic acid, N-Fmoc-N"-succinyl-4,7,10-trioxa-l,13-tridecanediamine (TTDS), PEG, and a glycine linker (GGG)3-20.

[0566] Embodiment 27. The CHP according to embodiment 26, wherein the length of the linker is 1 1-45 A.

[0567] Embodiment 28. The CHP according to any one of the preceding embodiments, wherein (Z) is absent.

[0568] Embodiment 29. Tire CHP according to any one of the preceding embodiments, wherein (Z) comprises at least one amino acid sequence. Embodiment 30. The CHP according to any one of the preceding embodiments, wherein (Z) comprises at least one repeat of (Gly-X-Y)a. Embodiment 31 . The CHP according to any one of the preceding embodiments, wherein the CHP further comprises one or more of an acetyl capping group, a linker, a cut site, and a bound drug molecule attached at the N-terminus of Formula I.

[0569] DBl / 161378040.3 MLB Docket No. 127033-5008

[0570] Embodiment 32. The CHP according to embodiment 31, wherein the CHP further comprises the cut site, the cut site comprising one or more selected from group consisting of MMP cut sites, cathepsin cut sites, pH-sensitive cut sites / linker, reducible di-sulfide cut site / linkers.

[0571] Embodiment 33. The CHP according to embodiment 31, wherein the CHP further comprises the drug molecule, the drug molecule comprising one or more selected from group consisting of immunomodulators, anti-cancer agents, anti-fibrotic agents, and anti-inflammatory agents.

[0572] Embodiment 34. The CHP according to any one of the preceding embodiments, wherein the CHP further comprises one or more sequence represented by Formula III attached at the N -terminus of Formula I:

[0573] (drug)-(linker) (Formula III)

[0574] Embodiment 35. The CHP according to any one of the preceding embodiments, wherein the CHP further comprises one or more sequence represented by Formula IV attached at the N-tenninus of Formula I:

[0575] (drug)-(cut site)-(linker) (Formula IV)

[0576] Embodiment 36. A composition comprising the CHP of any one of the preceding embodiments.

[0577] Embodiment 37. Tire composition according to embodiment 36, wherein each individual CHP does not form a triple helix with other CHPs.

[0578] Embodiment 38. The composition according to embodiment 36 or 37, further comprising a carrier. Embodiment 39. The composition according to any one of embodiments 36-38, wherein the composition is a cosmetic composition, and the carrier comprises at least one selected from the group consisting of micelles, dendrites, lipids, microemulsions, nanoemulsions, solid lipid nanoparticles, polymers, gels, lens, surfactants, cyclodextrins, inserts, nano structured lipid carriers, liposomes, transfersomes, ethosomes, niosomes, collagen matrix, extracellular matrix, and artificial extracellular matrix.

[0579] Embodiment 40. The composition according to any one of embodiments 36-39, wherein the composition is atopical cream, a saline solution, gel, polymer, or a solution for systemic injection within a subject.

[0580] Embodiment 41. Tire composition according to any one of embodiments 36-40, in which a CHP is bound directly to collagen matrix.

[0581] Embodiment 42. A method of treating denatured collagen in at subject, comprising administering the CHP of any one of embodiments 1-35, or the composition of any one of embodiments 36-41.

[0582] Embodiment 43. The method according to embodiment 42, wherein the administering is performed by injection.

[0583] Embodiment 44. The method according to embodiment 43, wherein the administering is perforated

[0584] DB1 / 161378040.3 MLB Docket No. 127033-5008 by micro-dermal injection.

[0585] Embodiment 45. The method according to embodiment 42, wherein the administering is performed by topical application.

[0586] Embodiment 46. A method of inhibiting or reducing an inflammatory response in a subject, comprising administering the CHP of any one of embodiments 1-35, or the composition of any one of embodiments 36-41.

[0587] Embodiment 47. The method according to embodiment 46, wherein the administering is performed by injection.

[0588] Embodiment 48. The method according to embodiment 46, wherein the administering is performed by micro-dennal injection.

[0589] Embodiment 49. The method according to embodiment 46, wherein the administering is perfonned by topical application.

[0590] Embodiment 50. A method of downregulating immune-related cytokines including, but not limited to, TLR ligands, interleukins, IFN-a, TLR-9, TNF-a, IL- Ip, IL-6, CCL3, IL- 16, IL- 18, and / or TNF-a activity in a subject, comprising administering the CHP of any one of embodiments 1-35, or the composition of any one of embodiments 36-41.

[0591] Embodiment 51. A method of preventing the differentiation of immune cells comprising administering the CHP of any one of embodiments 1-35, or the composition of any one of embodiments 36-41.

[0592] Embodiment 52. A method of downregulating T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils comprising administering the CHP of any one of embodiments 1-35, or the composition of any one of embodiments 36-41.

[0593] Embodiment 53. A method of preventing the differentiation of T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils comprising administering the CHP of any one of embodiments 1-35, or the composition of any one of embodiments 36-41.

[0594] Embodiment 54. The method according to any one of embodiments 50-53, wherein the administering is performed by injection.

[0595] Embodiment 55. Tire method according to any one of embodiments 50-53, wherein the administering is performed by micro-dermal injection.

[0596] Embodiment 56. The method according to any one of embodiments 50-53, wherein the administering is performed by topical application.

[0597] Embodiment 57. A method of preventing or treating dry eye disease in a subject, comprising administering the CHP of any one of embodiments 1-35, or the composition of any one of embodiments 36-41.

[0598] DB1 / 161378040.3 MLB Docket No. 127033-5008

[0599] Embodiment 58. The method according to embodiment 57, wherein the administering is performed by local injection, intravenous injection, topical application, or physical application.

[0600] Embodiment 59. The method according to embodiment 57, wherein the CHP or composition is administered as a putty, mesh, patch, adhesive, cream, sutures, gel or eyedrops.

[0601] Embodiment 60. A method of preventing or treating atopic dennatitis in a subject, comprising administering the CHP of any one of embodiments 1-35, or the composition of any one of embodiments 36-41.

[0602] Embodiment 61. The method according to embodiment 60, wherein the administering is performed by local injection, intravenous injection, topical application, or physical application.

[0603] Embodiment 62. The method according to embodiment 60, wherein the CHP or composition is administered as a putty, mesh, patch, adhesive, cream, sutures, eyedrop or gel.

[0604] Embodiment 63. A method of preventing or treating osteoarthritis in a subject, comprising administering the CHP of any one of embodiments 1-35, or the composition of any one of embodiments 36-41.

[0605] Embodiment 64. The method according to embodiment 63, wherein the administering is performed by local injection, intravenous injection, topical application, or physical application.

[0606] Embodiment 65. The method according to embodiment 63, wherein the CHP or composition is administered as a putty, mesh, patch, adhesive, cream, sutures, or eyedrops.

[0607] Embodiment 66. A method of preventing or treating rheumatoid arthritis in a subject, comprising administering the CHP of any one of embodiments 1-35, or the composition of any one of embodiments 36-41.

[0608] Embodiment 67. Tire method according to embodiment 66, wherein the administering is perforated by local injection, intravenous injection, topical application, or physical application.

[0609] Embodiment 68. The method according to embodiment 66, wherein the CHP or composition is administered as a putty, mesh, patch, adhesive, cream, sutures, or eyedrops.

[0610] Embodiment 69. A method of preventing or treating diabetic lesions in a subject, comprising administering the CHP of any one of embodiments 1-35, or the composition of any one of embodiments 36-41.

[0611] Embodiment 70. The method according to embodiment 69, wherein the administering is perfonned by local injection, intravenous injection, topical application, or physical application.

[0612] Embodiment 71. The method according to embodiment 69, wherein the CHP or composition is administered as a putty, mesh, patch, adhesive, cream, sutures, or eyedrops.

[0613] Embodiment 72. A method of preventing or treating psoriasis in a subject, comprising administering the CHP of any one of embodiments 1-35, or the composition of any one of embodiments 36-41.

[0614] DB1 / 161378040.3 MLB Docket No. 127033-5008

[0615] Embodiment 73. The method according to embodiment 72, wherein the administering is performed by local injection, intravenous injection, topical application, or physical application.

[0616] Embodiment 74. The method according to embodiment 72, wherein the CHP or composition is administered as a putty, mesh, patch, adhesive, cream, sutures, or eyedrops.

[0617] Embodiment 75. A method of checkpoint inhibition in a subject, comprising administering the CHP of any one of embodiments 1-35, or the composition of any one of embodiments 36-41.

[0618] Embodiment 76. The method according to embodiment 75, wherein the administering is performed by local injection, intravenous injection, topical application, or physical application.

[0619] Embodiment 77. The method according to embodiment 75, wherein the CHP or composition is administered as a putty, mesh, patch, adhesive, cream, sutures, or eyedrops.

[0620] Embodiment 78. A method of wound healing including diabetic lesions in a subject, comprising administering the CHP of any one of embodiments 1-35, or the composition of any one of embodiments 36-41.

[0621] Embodiment 79. The method according to embodiment 78, wherein the administering is performed by local injection, intravenous injection, topical application, or physical application.

[0622] Embodiment 80. Tire method according to embodiment 78, wherein the CHP or composition is administered as a putty, mesh, patch, adhesive, cream, sutures, or eyedrops.

[0623] Embodiment 81. The method according to any one of embodiments 78-80, wherein the at least one bioactive comprises an integrin binding sequence comprising a sequence having at least 85% sequence identity to any one of SEQ ID NO: 107-109, and wherein the integrin binding sequence is specific for alpha5-betal integrin.

[0624] Embodiment 82. Tire method according to any one of embodiments 78-81, wherein the at least one bioactive comprising an integrin binding sequence enhances cell adhesion and / or collagen production. Embodiment 83. The method according to any one of embodiments 78-82, wherein the at least one bioactive comprising an integrin binding sequence specifically does not bind to integrins other than alpha5-betal integrin.

[0625] Embodiment 84. A method of treating GI inflammatory conditions such as Crohn’s and Ulcerative Colitis in a subject, comprising administering the CHP of any one of embodiments 1-35, or the composition of any one of embodiments 36-41.

[0626] Embodiment 85. The method according to embodiment 84, wherein the administering is perfonned by local injection, intravenous injection, topical application, or physical application.

[0627] Embodiment 86. The method according to embodiment 84, wherein the CHP or composition is administered as a putty, mesh, patch, adhesive, cream, sutures, eyedrops, or gel.

[0628] Embodiment 87. A composition for treating denatured collagen comprising the collagen

[0629] DB1 / 161378040.3 MLB Docket No. 127033-5008 hybridization peptide (CHP) of any one of embodiments 1-35 as an active ingredient.

[0630] Embodiment 88. A composition for inhibiting or reducing an inflammatory response comprising the collagen hybridization peptide (CHP) of any one of embodiments 1-35 as an active ingredient. Embodiment 89. A composition for downregulating one or more immune-related cytokines selected from the group consisting of TLR ligands, interleukins, IFN-ot, TLR-9, IL- ip, IL-6, CCL3, IL-16, IL- 18, and TNF-a comprising the collagen hybridizing peptide (CHP) of any one of embodiments 1-35 as an active ingredient.

[0631] Embodiment 90. A composition for preventing immune cell differentiation comprising the collagen hybridizing peptide (CHP) of any one of embodiments 1-35 as an active ingredient.

[0632] Embodiment 91. A composition for downregulating T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils comprising the collagen hybridizing peptide (CHP) of any one of embodiments 1-35 as an active ingredient.

[0633] Embodiment 92. A composition for preventing differentiation of T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils comprising the collagen hybridizing peptide (CHP) of any one of embodiments 1-35 as an active ingredient.

[0634] Embodiment 93. A pharmaceutical composition for preventing or treating a dry eye disease comprising the collagen hybridizing peptide (CHP) of any one of embodiments 1-35 as an active ingredient.

[0635] Embodiment 94. A composition for preventing or treating atopic dermatitis comprising the collagen hybridizing peptide (CHP) of any one of embodiments 1-35 as an active ingredient.

[0636] Embodiment 95. A pharmaceutical composition for preventing or treating osteoarthritis comprising the collagen hybridizing peptide (CHP) of any one of embodiments 1-35 as an active ingredient. Embodiment 96. A pharmacal composition for preventing or treating rheumatoid arthritis comprising the collagen hybridizing peptide (CHP) of any one of embodiments 1-35 as an active ingredient.

[0637] Embodiment 97. A pharmaceutical composition for preventing or treating diabetic lesions comprising the collagen hybridizing peptide (CHP) of any one of embodiments 1-35 as an active ingredient.

[0638] Embodiment 98. A pharmacal composition for preventing or treating psoriasis comprising the collagen hybridizing peptide (CHP) of any one of embodiments 1-35 as an active ingredient. Embodiment 99. A composition for inhibiting checkpoints comprising the collagen hybridizing peptide (CHP) of any one of embodiments 1-35 as an active ingredient.

[0639] Embodiment 100. A pharmaceutical composition for healing wounds including diabetic lesions comprising the collagen hybridizing peptide (CHP) of any one of embodiments 1-35 as an active

[0640] DB1 / 161378040.3 MLB Docket No. 127033-5008 ingredient.

[0641] Embodiment 101. A pharmaceutical composition for preventing or treating GI inflammatory conditions including Crohn’s disease and ulcerative colitis comprising the collagen hybridizing peptide (CHP) of any one of embodiments 1-35 as an active ingredient.

[0642] Embodiment 102. A cosmetic composition for treating denatured collagen comprising the collagen hybridization peptide (CHP) of any one of embodiments 1-35 as an active ingredient.

[0643] Embodiment 103. A cosmetic composition for inhibiting or reducing an inflammatory response comprising the collagen hybridization peptide (CHP) of any one of embodiments 1-35 as an active ingredient.

[0644] Embodiment 104. A cosmetic composition for downregulating one or more immune-related cytokines selected from the group consisting of TLR ligands, interleukins, IFN-a, TLR-9, IL-ip, IL-6, CCL3, IL- 16, IL- 18, and TNF-a comprising the collagen hybridizing peptide (CHP) of any one of embodiments 1-35 as an active ingredient.

[0645] Embodiment 105. A cosmetic composition for preventing immune cell differentiation comprising the collagen hybridizing peptide (CHP) of any one of embodiments 1-35 as an active ingredient.

[0646] Embodiment 106. A cosmetic composition for downregulating T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils comprising the collagen hybridizing peptide (CHP) of any one of embodiments 1-35 as an active ingredient.

[0647] Embodiment 107. A cosmetic composition for preventing differentiation of T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils comprising the collagen hybridizing peptide (CHP) of any one of embodiments 1-35 as an active ingredient.

[0648] Embodiment 108. A cosmetic composition for preventing or treating a dry eye disease comprising the collagen hybridizing peptide (CHP) of any one of embodiments 1-35 as an active ingredient.

[0649] Embodiment 109. A cosmetic composition for preventing or treating atopic dermatitis comprising the collagen hybridizing peptide (CHP) of any one of embodiments 1-35 as an active ingredient.

[0650] Embodiment 110. A cosmetic composition for preventing or treating psoriasis comprising the collagen hybridizing peptide (CHP) of any one of embodiments 1-35 as an active ingredient.

[0651] Embodiment 111. A cosmetic composition for inhibiting checkpoints comprising the collagen hybridizing peptide (CHP) of any one of embodiments 1-35 as an active ingredient.

[0652] Embodiment 112. A quasi-drug composition for treating denatured collagen comprising the collagen hybridization peptide (CHP) of any one of embodiments 1-35 as an active ingredient.

[0653] Embodiment 113. A quasi-drug composition for inhibiting or reducing an inflammatory response comprising the collagen hybridization peptide (CHP) of any one of embodiments 1-35 as an active ingredient.

[0654] DB1 / 161378040.3 MLB Docket No. 127033-5008

[0655] Embodiment 114. A quasi-drug composition for downregulating one or more immune-related cytokines selected from the group consisting of TLR ligands, interleukins, IFN-a, TLR-9, IL- 1 p, IL-6, CCL3, IL- 16, IL- 18, and TNF-a comprising the collagen hybridizing peptide (CHP) of any one of embodiments 1-35 as an active ingredient.

[0656] Embodiment 115. A quasi-drug composition for preventing immune cell differentiation comprising the collagen hybridizing peptide (CHP) of any one of embodiments 1-35 as an active ingredient. Embodiment 116. A quasi-drug composition for downregulating T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils comprising the collagen hybridizing peptide (CHP) of any one of embodiments 1-35 as an active ingredient.

[0657] Embodiment 117. A quasi-drug composition for preventing differentiation of T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils comprising the collagen hybridizing peptide (CHP) of any one of embodiments 1-35 as an active ingredient.

[0658] Embodiment 118. A quasi-drug composition for preventing or treating a dry eye disease comprising the collagen hybridizing peptide (CHP) of any one of embodiments 1-35 as an active ingredient. Embodiment 119. A quasi-drug composition for preventing or treating atopic dermatitis comprising the collagen hybridizing peptide (CHP) of any one of embodiments 1-35 as an active ingredient. Embodiment 120. A quasi-drug composition for preventing or treating osteoarthritis comprising the collagen hybridizing peptide (CHP) of any one of embodiments 1-35 as an active ingredient. Embodiment 121. A quasi-drug composition for preventing or treating rheumatoid arthritis comprising the collagen hybridizing peptide (CHP) of any one of embodiments 1-35 as an active ingredient.

[0659] Embodiment 122. A quasi-drug composition for preventing or treating diabetic lesions comprising the collagen hybridizing peptide (CHP) of any one of embodiments 1-35 as an active ingredient. Embodiment 123. A quasi-drug composition for preventing or treating psoriasis comprising the collagen hybridizing peptide (CHP) of any one of embodiments 1-35 as an active ingredient. Embodiment 124. A quasi-drug composition for inhibiting checkpoints comprising the collagen hybridizing peptide (CHP) of any one of embodiments 1-35 as an active ingredient.

[0660] Embodiment 125. A quasi-drug composition for healing wounds including diabetic lesions comprising the collagen hybridizing peptide (CHP) of any one of embodiments 1-35 as an active ingredient. Embodiment 126. A quasi-drug composition for preventing or treating GI inflammatory conditions including Crohn’s disease and ulcerative colitis comprising the collagen hybridizing peptide (CHP) of any one of embodiments 1-35 as an active ingredient.

[0661] Embodiment 127. The CHP according to any one of embodiments 1-35, wherein (Z) is bioactivc and comprises an amino acid sequence including 3 to 27 amino acid residues having bioactivity, wherein

[0662] DB1 / 161378040.3 MLB Docket No. 127033-5008 the bioactive comprises at least one leukocyte-associated immunoglobulin-like receptor (LAIR-1 and / or LAIR-2) binding sequence, and wherein the amino acid sequence of the bioactive includes at least one citrulline residue.

[0663] Embodiment 128. A method of reducing or preventing downregulation of immune cells comprising administering a composition comprising the collagen hybridization peptide of embodiment 127 as an active ingredient to a subject in need thereof.

[0664] Embodiment 129. A method of preventing or treating cancer comprising administering a composition comprising the collagen hybridization peptide of embodiment 127 as an active ingredient to a subject in need thereof.

[0665] Embodiment 130. The CHP according to any one of embodiments 1-35, wherein (Z) is bioactive and comprises any amino acid capable of post-translational modification by a sugar in a glycation reaction. Embodiment 131. A method of reducing, preventing or treating oxidative modification of natural collagen comprising administering a composition comprising the collagen hybridization peptide of embodiment 130 as an active ingredient to a subject in need thereof.

[0666] Embodiment 132. A method of reducing, preventing or treating oxidative modification of natural collagen in a subject with diabetes comprising administering a composition comprising the collagen hybridization peptide of embodiment 130 as an active ingredient to the subject in need thereof. Embodiment 133. A method of reducing, preventing or treating oxidative modification of natural collagen in a subject with diabetic ketoacidosis comprising administering a composition comprising the collagen hybridization peptide of embodiment 130 as an active ingredient to the subject in need thereof.

[0667] Embodiment 134. A method of reducing, preventing or treating oxidative modification of natural collagen in a subject with hyperglycemic hyperosmolar syndrome comprising administering a composition comprising the collagen hybridization peptide of embodiment 130 as an active ingredient to the subject in need thereof.

[0668] Embodiment 135. A method of reducing, preventing or treating oxidative modification of natural collagen in a subject with acidosis comprising administering a composition comprising the collagen hybridization peptide of embodiment 130 as an active ingredient to the subject in need thereof. Embodiment 136. A method of reducing, preventing or treating oxidative modification of natural collagen in a subject with homocystinuria comprising administering a composition comprising the collagen hybridization peptide of embodiment 130 as an active ingredient to the subject in need thereof.

[0669] Embodiment 137. A method of reducing, preventing or treating oxidative modification of natural collagen in a subject with methylmalonic acidemia comprising administering a composition

[0670] DB1 / 161378040.3 MLB Docket No. 127033-5008 comprising the collagen hybridization peptide of embodiment 130 as an active ingredient to the subject in need thereof.

[0671] Embodiment 138. A method of reducing, preventing or treating diabetic lesions in a subject comprising administering a composition comprising the collagen hybridization peptide of embodiment 130 as an active ingredient to the subject in need thereof.

[0672] Embodiment 139. A method of reducing, preventing or treating diabetic retinopathy in a subject comprising administering a composition comprising the collagen hybridization peptide of embodiment 130 as an active ingredient to the subject in need thereof.

[0673] Embodiment 140. The CHP according to any one of embodiments 1 to 35, further comprising at least one detection moiety.

[0674] Embodiment 141. The CHP according to embodiment 140, wherein the at least one detection moiety is a dye selected from the group consisting of Alexa Fluor dyes, cyanine dyes, sulfo-cyanine dyes, indocyanine dyes, Tide Flour dyes, TAMRA, FITC, 5 -FAM, carboxyfluorescein, coumarin dyes and rhodamine dyes; a gold particle; or a label selected from the group consisting of prednisolone acetate, triamcinolone acetonide and lipid-based artificial tears.

[0675] Embodiment 142. A method of providing information for diagnosing fibrosis in a subject comprising: administering a composition comprising the collagen hybridization peptide of embodiment 140 or 141 as an active ingredient to the subject; and imaging the collagen hybridization peptide comprising the detection moiety in vivo to determine presence or progression of fibrosis in the subject.

[0676] Embodiment 143. The method according to embodiment 142, wherein the fibrosis is selected from the group consisting of neovascular age-related macular degeneration (nAMD), diabetic retinopathy, glaucoma, particularly fibrosis in the trabecular meshwork, neovascular glaucoma, corneal scarring, conjunctiva, post-cataract surgery, retinopathy of prematurity, and proliferative vitreoretinopathy. Embodiment 144. The CHP according to any one of embodiments 1 to 35, wherein (Z) comprises at least one bioactive, a linker, at least one (Gly-X-Y)arepeat unit, or a combination thereof. Embodiment 145. The CHP according to any one of embodiments 1 to 35, wherein the CHP comprises a sequence having at least 85% sequence identity to any one of SEQ ID NOs: 115-127.

[0677] DB1 / 161378040.3

Claims

MLB Docket No. 127033-5008CLAIMS1 . A collagen hybridizing peptide (CHP), wherein the CHP comprises a sequence represented by Formula I:(Gly-X-Y)a-(Z) (Formula I) in which X and Y are independently any amino acid, provided at least one of X and Y is proline, hydroxyproline, or modified proline in each instance of (Gly-X-Y); Gly is glycine; Z is an optional spacer; and a is a number from 1 to 40, and the CHP or the Formula I comprises a modification at the C-terminus, thereby lowering a melting temperature of the CHP.

2. The CHP according to claim 1, wherein the modification at the C-terminus comprises one or more molecules that yield a lower homotrimerization melting temperature for the CHP than that of the CHP bound with gelatin or damaged collagen.

3. The CHP according to claim 1 or 2, wherein the modification at the C-terminus comprises attaching one or more amino acids including one or more sequences represented by Formula II to the C-terminus of the Formula I:(Gly-B-B) (Formula II) in which B is independently in each instance phenylalanine, tryptophan, tyrosine, aspartic acid, lysine, arginine, histidine, or any charged amino acid.

4. The CHP according to any one of the preceding claims, wherein (Z) comprises at least one bioactive, the at least one bioactive comprising an amino acid sequence from 3 to 27 amino acid residues having bioactivity, wherein the bioactive comprises at least one leukocyte-associated immunoglobulin-like receptor (LAIR-1 and / or LAIR-2) binding sequence, and wherein the amino acid sequence of the bioactive includes at least one citrulline residue.

5. The CHP according to any one of the preceding claims, wherein the at least one bioactive comprises at least one sequence selected from the group consisting of a leukocyte-associated immunoglobulin-like receptor binding sequence (LAIR-1 and / or LAIR-2), an integrin binding sequence, a discoidin domain receptor (DDR) binding sequence, a von Willebrand factor (VWF) sequence, a matrix metalloproteinase (MMP) binding sequence, a G6b-B recognition site sequence, an OSCAR domain sequence, a GPVI sequence, and a FcR-y domain sequence.DB1 / 161378040.3MLB Docket No. 127033-50086. The CHP according to any one of the preceding claims, wherein the CHP comprises a sequence having at least 85% sequence identity to any one of SEQ ID NOs: 1-104 and 107-114.

7. The CHP according to any one of the preceding claims, wherein the CHP comprises a sequence having at least 85% sequence identity to any one of SEQ ID NOs: 115-127.

7. The CHP according to any one of the preceding claims, wherein (Z) comprises at least one linker, wherein the at least one linker comprises one or more selected from the group consisting of Ahx (6-aminohexanoic acid), 12-aminododecanoic acid, N-Fmoc-N"-succinyl-4,7, 10-trioxa-l,13- tridecanediamine (TTDS), PEG, and a glycine linker (GGG)3-20; at least one amino acid sequence; at least one repeat of (Gly-X-Y)a; or at least one bioactive, a linker, at least one (Gly-X-Y)arepeat unit, or a combination thereof.

8. The CHP according to any one of the preceding claims, wherein the CHP further comprises one or more of an acetyl capping group, a linker, a cut site, and a bound drug molecule attached at the N-terminus of Fonnula I.

9. The CHP according to any one of the preceding claims, wherein the CHP further comprises the cut site, the cut site comprising one or more selected from group consisting of MMP cut sites, cathepsin cut sites, pH-sensitive cut sites / linker, and reducible di-sulfide cut site / linkers.

10. The CHP according to any one of the preceding claims, wherein the CHP further comprises the drug molecule, the drug molecule comprising one or more selected from group consisting of immunomodulators, anti-cancer agents, anti-fibrotic agents, and anti-inflammatory agents.

11. The CHP according to any one of the preceding claims, wherein the CHP further comprises one or more sequence represented by Formula III attached at the N-terminus of Formula I:(drug)-(linker) (Formula 111).

12. The CHP according to any one of the preceding claims, wherein the CHP further comprises one or more sequence represented by Formula IV attached at the N-tcrminus of Formula I:(drug)-(cut site)-(linker) (Formula IV).DB1 / 161378040.3MLB Docket No. 127033-500813. The CHP according to any one of the preceding claims, further comprising at least one detection moiety, wherein the at least one detection moiety is a dye selected from the group consisting of Alexa Fluor dyes, cyanine dyes, sulfo-cyanine dyes, indocyanine dyes, Tide Flour dyes, TAMRA, FITC, 5-FAM, carboxyfluorescein, coumarin dyes and rhodamine dyes; a gold particle; or a label selected from the group consisting of prednisolone acetate, triamcinolone acetonide and lipid-based artificial tears.

14. A composition comprising the CHP of any one of the preceding claims.

15. The composition according to claim 14, further comprising a carrier.

16. A method comprising administering the CHP of any one of claims 1 to 13, or the composition of claim 14 or 15, wherein the method is used for treating denatured collagen in a subject; inhibiting or reducing an inflammatory response in a subject; downregulating immune-related cytokines including, but not limited to, TLR ligands, interleukins, IFN-a, TLR-9, TNF-a, IL- 10, IL-6, CCL3, IL-16, IL-18, and / or TNF-a activity in a subject; preventing the differentiation of immune cells; downregulating T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils; preventing the differentiation of T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils; preventing or treating dry eye disease in a subject; preventing or treating atopic dermatitis in a subject; preventing or treating osteoarthritis in a subject; preventing or treating rheumatoid arthritis in a subject; preventing or treating diabetic lesions in a subject; preventing or treating psoriasis in a subject; checkpoint inhibition in a subject; wound healing including diabetic lesions in a subject; and treating GI inflammatory conditions such as Crohn's and Ulcerative Colitis in a subject.

17. The method according to claim 16, wherein the at least one bioactive comprises an integrin binding sequence comprising a sequence having at least 85% sequence identity to any one of SEQ ID NO: 107-109, and wherein the integrin binding sequence is specific for alpha5-betal integrin.

18. A composition comprising the collagen hybridization peptide (CHP) of any one of claims 1-13 as an active ingredient, wherein the composition is used for treating denatured collagen; inhibiting or reducing an inflammatory response; downrcgulating one or more immune-related cytokines selected from the group consisting of TLR ligands, interleukins, IFN-a, TLR-9, IL-10, IL-6, CCL3, IL-16, IL-18, and DB1 / 161378040.3MLB Docket No. 127033-5008TNF-a; preventing immune cell differentiation; downregulating T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils; preventing differentiation of T cells, B cells, natural killer cells, macrophages, monocytes, and neutrophils; or inhibiting checkpoints, wherein the composition comprises at least one selected from the group consisting of a pharmacal composition, a cosmetic composition, a quasi-drug composition.DBl / 161378040.3

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