Compositions Containing Chemerin Analogs and Methods of Use Thereof

Abstract

The present disclosure relates to peptide compositions and methods for treating inflammatory conditions such as ocular inflammation, retinal inflammation, dry eye, uveitis, allergic conjunctivitis, and inflammation due to nerve injury or neurodegeneration. The peptide compositions disclosed herein can also be used to treat pain such as neuropathic pain, ocular pain, chronic pain, pain due to chemotherapy or radiation, pain due to nerve injury or neurodegeneration, and pain due to inflammatory conditions, paresthesia, or allodynia.

Description

[Technical field]

[0001] Related Applications This application claims priority to and the benefit of U.S. Patent Application Publication No. 62 / 881,725, filed August 1, 2019, the contents of which are incorporated herein by reference in their entirety.

[0002] Incorporation by reference of sequence listing The contents of the text file named "OKYO-006_001WO_Sequence Listing_ST25.txt", created on June 28, 2020 and having a size of 16KB, are incorporated herein by reference in their entirety.

[0003] Technical Field The present disclosure relates generally to chemerin analogs and their use in the treatment of various diseases and conditions, including, but not limited to, inflammatory and pain conditions. [Background technology]

[0004] Background of the disclosure There are various inflammatory conditions, including, but not limited to, ocular inflammation, dry eye, uveitis (e.g., non-infectious uveitis), allergic conjunctivitis, retinal inflammation, and inflammation resulting from nerve injury or degeneration. Ocular inflammation can be caused by microbial infection of the eye. Such infections can be fungal, viral, or bacterial. Ocular inflammation can also be caused by trauma, autoimmune disease, chemical injury, contact lenses, or other external stimuli.

[0005] Dry eye, for example, is a multifactorial disease of the tears and ocular surface that involves inflammation in its pathogenesis. Dry eye is a common and chronic challenge, especially prevalent in older adults. In 2000, the prevalence in the United States was estimated to be about 17% in women and 12% in men, but has increased in recent years and is estimated to be over 50%. People with dry eye either do not produce enough tears or have poor tear quality. Tears are produced by several glands in and around the eyelids. Tear production tends to decrease with age, due to various medical conditions, or as a side effect of certain medications. Environmental conditions such as wind and dry weather can also decrease tear production by increasing tear evaporation. If normal tear production decreases or tears evaporate too quickly from the eye, dry eye symptoms may appear. In terms of tear quality, tears are made up of three layers: oil, water, and mucus. Each component protects and nourishes the front of the eye. The lubricating oil layer prevents the aqueous layer from evaporating, while the mucin layer spreads the tears evenly over the surface of the eye. When tears evaporate too quickly or are not spread evenly across the cornea due to a deficiency in any of the three tear films, dry eye symptoms can appear.

[0006] A common form of dry eye occurs when the aqueous layer of tears is insufficient, a condition also known as keratoconjunctivitis sicca (KCS).

[0007] Pain also includes a variety of pain conditions, including neuropathic pain, ocular pain, chronic pain, pain due to chemotherapy or radiation, pain due to nerve injury or neurodegeneration, and pain due to inflammatory conditions, paresthesia, or allodynia.

[0008] For example, neuropathic pain is a complex chronic pain condition that usually involves tissue damage. In neuropathic pain, the nerve fibers themselves may be injured, dysfunctional, or damaged. These injured nerve fibers send erroneous signals to other pain centers. The effects of nerve fiber damage include altered nerve function both at the site of injury and around the injury. Neuropathic pain is a serious health problem that affects millions of people worldwide, occurring in as many as 7% of the total population. Managing neuropathic pain in patients is complex, with an estimated 40-60% of people being resistant to existing pain relief therapies. An aging population, the diabetes epidemic, and patients with cancer and AIDS all contribute to the epidemic of intractable neuropathic pain, highlighting the urgent need to develop new treatments for this condition.

[0009] The eye is highly innervated by sensory nerve fibers, and inflammatory, ischemic, and even neoplastic lesions of the eye and orbit can cause pain. Ophthalmic causes of eye pain include dry eye and other forms of keratitis, acute angle-closure glaucoma, and intraocular inflammation. Sicca keratitis, or dry eye, is a very common cause of eye discomfort. These conditions are most commonly diagnosed by slit-lamp examination of the cornea, anterior segment, and anterior vitreous. Dry eye is exacerbated by visual tasks, especially computer work, that reduce blink frequency, which has various causes and consequences and results from conditions that reduce tear production or increase tear evaporation. Dry eye is one of the hallmarks of autoimmune Sjogren's syndrome. Evidence of fluorescein or rose bengal staining, abnormal tear breakup time, and / or reduced Schirmer's test may help confirm dry eye syndrome. Posterior segment examination by indirect ophthalmoscopy or slit-lamp biomicroscopy may show evidence of choroidal or retinal inflammation or posterior scleritis.

[0010] Chemotherapy or radiation therapy can cause peripheral neuropathy (painful numbness in the limbs) or paresthesias (numbness and tingling in the hands, feet, or any extremity of the body). Summary of the Invention [Problem to be solved by the invention]

[0011] The present disclosure addresses the needs of patients suffering from a variety of pain and / or inflammatory conditions such as those described above. [Means for solving the problem]

[0012] Disclosure Summary The present disclosure provides compositions and methods for treating or ameliorating at least one symptom of neuropathic pain, ocular pain, chronic pain, pain due to chemotherapy or radiation, pain due to nerve injury or neurodegeneration, pain due to an inflammatory condition paresthesia or allodynia, inflammation due to nerve injury or neurodegeneration, ocular inflammation, retinal inflammation, uveitis, allergic conjunctivitis, and / or dry eye.

[0013] In one embodiment, the present disclosure provides a composition comprising: 1 X 2 X 3 X 4 -Nle-PX 5 X 6 X 7 X 8 -Tic-X 9 A peptide comprising an amino acid sequence having the sequence of (SEQ ID NO:1), and a lipid entity linked to said peptide, wherein X 1 is selected from A, dA, and NorV; 2 is A or G; X 3 is F or dY; X 4 is F or Y; X 5 is S or dS; X 6 is Q or α-aminoadipic acid (Aad); 7 is selected from Y, F, and fF; X 8 is A or dA; Tic is (S)-1,2,3,4-tetrahydroisoquinoline-3-carboxylic acid; X 9 is dA or dS.

[0014] In some embodiments, the peptide comprises X 1 X 2 X3 X 4 -Nle-PX 5 X 6 X 7 X 8 -Tic-X 9 It consists of amino acids having the sequence of (SEQ ID NO: 1).

[0015] In some embodiments, the peptide is selected from AAFY-Nle-PSQYA-Tic-dA (SEQ ID NO: 2), AG-dY-F-Nle-P-dS-Aad-FA-Tic-dA (SEQ ID NO: 3), dA-G-dY-F-Nle-P-dS-Q-fF-dA-Tic-dA (SEQ ID NO: 4), NorV-G-dY-F-Nle-P-dS-QF-dA-Tic-dA (SEQ ID NO: 5), AG-dY-F-Nle-P-dS-QFA-Tic-dS (SEQ ID NO: 6), and dA-G-dY-F-Nle-P-dS-QF-dA-Tic-dA (SEQ ID NO: 7).

[0016] In some embodiments, the lipid entity is linked to the peptide via a linker entity.

[0017] In some embodiments, the lipid entity is linked at or near the N-terminus of the peptide.

[0018] In some embodiments, the lipid entity is linked at or near the C-terminus of the peptide.

[0019] In some embodiments, the lipid entity is selected from the group consisting of α-linolenic acid, γ-linolenic acid, stearidonic acid, eicosapentaenoic acid, docosahexaenoic acid, linoleic acid, dihomo-γ-linolenic acid, arachidonic acid, docosatetraenoic acid, palmitoleic acid, vaccenic acid, paulic acid, oleic acid, elaidic acid, gondoic acid, erucic acid, nervonic acid, mead acid, myristic acid, palmitic acid, stearic acid, 1,2-dipalmitoyl-sn-glycero-3-phosphoric acid, glyceryl-sulfate ... DPPE, GM1 ganglioside, GM2 ganglioside, GM3 ganglioside, 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC), 1,2-dioleoyl-sn-glycero-3-phospho-L-serine (DOPS), 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), glycosphingolipids, sphingolipids, phosphatidylinositol 4,5-bisphosphate (PIP 2 ), ceramide, cholesterol, ergosterol, phytosterol, hopanoids, steroids, and 17-carboxy-1-oxo-heptadecyl.

[0020] In some embodiments, the lipid entity is selected from the group consisting of α-linolenic acid, γ-linolenic acid, palmitic acid, myristic acid, vaccenic acid, oleic acid, and elaidic acid.

[0021] In some embodiments, the linker entity comprises ethylene glycol, polyethylene glycol, a peptide, aminoethylethanolamine (AEEA), inulin, a trisaccharide, or a combination thereof.

[0022] In some embodiments, the linker entity comprises polyethylene glycol (PEG).

[0023] In some embodiments, the linker entity is selected from the group consisting of: [ka] [ka] [ka]

[0024] In some embodiments, the combination of lipid entity and linker entity comprises palmitate-PEG8.

[0025] In some embodiments, the lipid entity is covalently attached to the peptide.

[0026] In some embodiments, the composition is water-soluble.

[0027] In one aspect, the present disclosure relates to a pharmaceutical composition comprising a peptide composition described herein and a pharma- ceutically acceptable carrier.

[0028] In one embodiment, the present disclosure relates to a method of treating pain in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a peptide composition or pharmaceutical composition described herein.

[0029] In some embodiments, the pain is ocular pain, chronic pain, neuropathic pain, pain due to chemotherapy or radiation, pain due to nerve injury or neurodegeneration, or pain due to an inflammatory condition, dysesthesias, or allodynia.

[0030] In some embodiments, the peptide composition or pharmaceutical composition is administered locally.

[0031] In some embodiments, the peptide composition or pharmaceutical composition is administered once per day, twice per day, or three times per day.

[0032] In some embodiments, the method further comprises administering an analgesic (e.g., paracetamol, a nonsteroidal anti-inflammatory drug, a COX-2 inhibitor, an opioid, or medical cannabis).

[0033] In some embodiments, the subject is a human.

[0034] In another aspect, the present disclosure relates to a method of treating an inflammatory condition in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a peptide composition or pharmaceutical composition described herein.

[0035] In some embodiments, the inflammatory condition is ocular inflammation.

[0036] In some embodiments, the inflammatory condition is dry eye disease, uveitis (eg, non-infectious uveitis), allergic conjunctivitis, or a retinal inflammatory disease.

[0037] In some embodiments, the inflammatory condition results from nerve injury or nerve degeneration.

[0038] In some embodiments, the peptide composition or pharmaceutical composition is administered locally.

[0039] In some embodiments, the peptide composition or pharmaceutical composition is administered once per day, twice per day, or three times per day.

[0040] In some embodiments, the subject is a human.

[0041] Any aspect or embodiment described herein can be combined with any other aspect or embodiment disclosed herein. While the present disclosure has been described in conjunction with its detailed description, the foregoing description is intended to illustrate, but not to limit, the scope of the disclosure, which is defined by the appended claims. Other aspects, advantages, and modifications are within the scope of the following claims.

[0042] The patent and scientific literature referenced herein establishes knowledge available to those skilled in the art. All U.S. patents and published or unpublished U.S. patent applications cited herein are incorporated by reference. All published foreign patents and patent applications cited herein are incorporated by reference. All other published references, publications, manuscripts, and scientific literature cited herein are incorporated by reference. [Brief description of the drawings]

[0043] [Figure 1] FIG. 1 is a graph showing that OK-101 significantly reduced the increase in corneal permeability caused by dry eye. The efficacy of OK-101 was comparable to that of cyclosporine (CS), the active ingredient in Restasis® (Allergan). Animal studies were performed with a single dose. OK-101 has the following structure: [ka]

[0044] [Diagram 2] FIG. 1 is a graph showing that OK-113 significantly reduced the increase in corneal permeability caused by dry eye. The efficacy of OK-113 was comparable to that of cyclosporine (CS), the active ingredient in Restasis® (Allergan). Animal studies were performed with a single dose. OK-113 has the following structure: [ka]

[0045] [Diagram 3] FIG. 1 is a graph showing that OK-101 significantly reduced dry eye-induced CD4+ T cell infiltration, a known biomarker of inflammation.

[0046] [Figure 4] 1 is a graph showing that OK-101 normalized dry eye-induced loss of goblet cell density.

[0047] [Diagram 5] 1 is a graph showing that OK-113 significantly reduced dry eye-induced CD4+ T cell infiltration.

[0048] [Figure 6] 1 is a graph showing that OK-113 normalized dry eye-induced loss of goblet cell density. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0049] Detailed Description of the Disclosure As described in more detail herein, there is an unmet need for novel strategies to treat pain, especially neuropathic pain, a disease that affects millions of people worldwide and is present in as many as 7% of the population.The management of patients with neuropathic pain is complex, and many patients do not respond to treatment or experience only partial relief.At the extreme, there is a substantial subpopulation with moderate to severe chronic intractable pain, and more effective and long-acting therapies are urgently needed.

[0050] CMKLR1 is a G protein-coupled receptor that has been shown to regulate nociception. The receptor is expressed in glial cells, dorsal root ganglion neurons, and immune cells. The endogenous ligand (agonist) of CMKLR1 is the 163 amino acid protein chemerin. Chemerin, also known as retinoic acid receptor response protein 2 (RARRES2), tazarotene-induced gene 2 protein (TIG2), or RAR response protein TIG2, is a protein that is encoded by the RARRES2 gene in humans. The homo sapiens amino acid sequence of chemerin is shown below in SEQ ID NO:8.

[0051] NCBI reference sequence: NP_002880.1 MRRLLIPLAL WLGAVGVGVA ELTEAQRRGL QVALEEFHKH PPVQWAFQET SVESAVDTPF PAGIFVRLEF KLQQTSCRKR DWKKPECKVR PNGRKRKCLA CIKLGSEDKV LGRLVHCPIE TQVLREAEEH QETQCLRVQR AGEDPHSFYF PGQFAFSKAL PRS (SEQ ID NO: 8).

[0052] Chemerin is inactive as preprochemerin (having SEQ ID NO: 8) and is activated via C- and N-terminal cleavage to form a chemerin fragment having the amino acid sequence of positions 21 to 157 of SEQ ID NO: 8, which can function as an agonist of CMKLR1. This chemerin fragment has the following amino acid sequence: ELTEAQRRGL QVALEEFHKH PPVQWAFQET SVESAVDTPF PAGIFVRLEF KLQQTSCRKR DWKKPECKVR PNGRKRKCLA CIKLGSEDKV LGRLVHCPIE TQVLREAEEH QETQCLRVQR AGEDPHSFYF PGQFAFS (SEQ ID NO: 9).

[0053] In particular, the disclosure provides compositions comprising chemerin analogs that can retain some or all of the biological functions of the amino acid sequence of SEQ ID NO: 9, e.g., functioning as an agonist of CMKLR1. Chemerin analogs can be either full-length or fragment analogs of chemerin.

[0054] In one embodiment, the present disclosure provides a composition comprising: 1 X 2 X 3 X 4 -Nle-PX 5 X 6 X 7 X 8 -Tic-X 9A peptide comprising an amino acid having the sequence of (SEQ ID NO: 1), and a lipid entity linked to the peptide. In some embodiments, any one or more amino acids in the peptide can be either L-amino acids or D-amino acids. In some embodiments, any one or more amino acids in the peptide can be fluorinated. As used herein, a fluorinated amino acid is an amino acid in which one or more hydrogens are replaced with fluorine. As used herein, Nle refers to norleucine, and Tic refers to (S)-1,2,3,4-tetrahydroisoquinoline-3-carboxylic acid.

[0055] In some embodiments, X 1 is selected from A, dA, and NorV; 2 is A or G; X 3 is F or dY; X 4 is F or Y; X 5 is S or dS; X 6 is Q or α-aminoadipic acid (Aad); 7 is selected from Y, F, and fF; X 8 is A or dA; X 9 is dA or dS.

[0056] As used herein, NorV refers to norvaline.

[0057] As used herein, a lower case d before an amino acid indicates that the amino acid is a D-amino acid, for example, dA indicates D-alanine, dY indicates D-tyrosine, and dS indicates D-serine.

[0058] As used herein, a lower case f before an amino acid indicates that the amino acid is fluorinated, for example, fF indicates fluorinated phenylalanine.

[0059] In some embodiments, the peptide has 20 or fewer amino acids, 19 or fewer amino acids, 18 or fewer amino acids, 17 or fewer amino acids, 16 or fewer amino acids, 15 or fewer amino acids, 14 or fewer amino acids, 13 or fewer amino acids, or 12 or fewer amino acids.

[0060] In some embodiments, the peptide has at least 9 amino acids, at least 10 amino acids, at least 11 amino acids, or at least 12 amino acids.

[0061] Combinations of the above ranges for the number of amino acids are also possible, for example, a peptide can have about 9-20 amino acids, about 10-20 amino acids, or about 10-18 amino acids.

[0062] In some embodiments, the peptide comprises X 1 X 2 X 3 X 4 -Nle-PX 5 X 6 X 7 X 8 -Tic-X 9 It consists of amino acids having the sequence of (SEQ ID NO: 1).

[0063] In some embodiments, the peptide comprises AAFY-Nle-PSQYA-Tic-dA (SEQ ID NO:2), AG-dY-F-Nle-P-dS-Aad-FA-Tic-dA (SEQ ID NO:3), dA-G-dY-F-Nle-P-dS-Q-fF-dA-Tic-dA (SEQ ID NO:4), NorV-G-dY-F-Nle-P-dS-QF-dA-Tic-dA (SEQ ID NO:5), AG-dY-F-Nle-P-dS-QFA-Tic-dS (SEQ ID NO:6), or dA-G-dY-F-Nle-P-dS-QF-dA-Tic-dA (SEQ ID NO:7).

[0064] In some embodiments, the peptide comprises AAFY-Nle-PSQYA-Tic-dA (SEQ ID NO:2), AG-dY-F-Nle-P-dS-Aad-FA-Tic-dA (SEQ ID NO:3), dA-G-dY-F-Nle-P-dS-Q-fF-dA-Tic-dA (SEQ ID NO:4), NorV-G-dY-F-Nle-P-dS-QF-dA-Tic-dA (SEQ ID NO:5), AG-dY-F-Nle-P-dS-QFA-Tic-dS (SEQ ID NO:6), or dA-G-dY-F-Nle-P-dS-QF-dA-Tic-dA (SEQ ID NO:7).

[0065] According to the present disclosure, any of a variety of lipid entities can be utilized. According to various embodiments, the lipid entity can include an entity that can be inserted into a lipid bilayer (e.g., a cell membrane). In some embodiments, the lipid entity can be incorporated into lipid rafts of a lipid bilayer (e.g., a cell membrane).

[0066] In some embodiments, the lipid entity may contain saturated or unsaturated fatty acids. The numbers in the lipid name are used to describe the fatty acid chain of the lipid. The numbers are usually expressed in the format of (carbon number of fatty acid chain):(number of double bonds of fatty acid chain), for example, 16:0 indicates that there are 16 carbons in the fatty acid chain and zero double bonds. The saturated or unsaturated fatty acid may contain at least 4 carbons, at least 5 carbons, at least 6 carbons, at least 7 carbons, at least 8 carbons, at least 9 carbons, at least 10 carbons, or at least 15 carbons in the fatty acid chain. In some embodiments, the saturated or unsaturated fatty acid may contain about 4 to 24 carbons in the fatty acid chain. The number of double bonds in the fatty acid chain may range from 0 to 10, for example, 0 to 8, 0 to 6, 1 to 8, 1 to 6. For example, the lipid entity can be C22:0, C22:1, C22:2, C22:3, C22:4, C22:5, C22:6, C20:0, C20:1, C20:2, C20:3, C20:4, C20:5, C20:6, C18:0, C18:1, C18:2, C18:3, C18:4, C18:5, C18:6, C10:0, C10:1, C10:2, C10:3, C10:4, etc.

[0067] For example, lipid entities include α-linolenic acid, γ-linolenic acid, stearidonic acid, eicosapentaenoic acid, docosahexaenoic acid, linoleic acid, dihomo-γ-linolenic acid, arachidonic acid, docosatetraenoic acid, palmitoleic acid, vaccenic acid, paulic acid, oleic acid, elaidic acid, gondoic acid, erucic acid, nervonic acid, mead acid, myristic acid, palmitic acid, stearic acid, 1,2-dipalmitoyl-sn-glycero-3-phosphoethanoic acid, and 1,2-dipalmitoyl-sn-glycero-3-phosphoethanoic acid. dipalmitoylamine (DPPE), GM1 ganglioside, GM2 ganglioside, GM3 ganglioside, 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC), 1,2-dioleoyl-sn-glycero-3-phospho-L-serine (DOPS), 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), glycosphingolipids, sphingolipids, phosphatidylinositol 4,5-bisphosphate (PIP 2 ), ceramide, cholesterol, ergosterol, phytosterol, hopanoid, steroid, fluorinated-GM1, fluorinated-GM2, fluorinated-GM3, 17-carboxy-1-oxo-heptadecyl, and isoprenoid lipids (e.g., farnesyl (C-15) or geranylgeranyl (C-20)). In some embodiments, the lipid entity can be alpha-linolenic acid. In some embodiments, the lipid entity can be gamma-linolenic acid. In some embodiments, the lipid entity can be palmitic acid. In some embodiments, the lipid entity can be vaccenic acid. In some embodiments, the lipid entity can be oleic acid. In some embodiments, the lipid entity can be elaidic acid. In some embodiments, the lipid entity can be myristic acid. In some embodiments, the lipid entity can be 17-carboxy-1-oxo-heptadecyl.

[0068] The attachment of a lipid entity to a peptide is referred to herein as lipidation. In some embodiments, the lipid entity is covalently attached to the peptide. In some embodiments, lipidation refers to the attachment of a peptide to any compound that is soluble in a cell membrane (e.g., by reducing the equilibrium constant K assoc ≧10, including 10:1).

[0069] In some embodiments, lipidation can include one or more of the following: attachment of diacylglycerol to the side chain of the N-terminal cysteine ​​of the peptide via a sulfur atom; O-octanoyl attachment to a serine or threonine of the peptide; and attachment of S-archaeoyl to a cysteine ​​of the peptide. In some embodiments, lipidation can occur, for example, at any lysine, glutamic acid, aspartic acid, serine, threonine, cysteine, and / or tyrosine.

[0070] In some embodiments, lipidation may include fluorination. Fluorination may include one or more C 6 F 13 Without wishing to be bound by theory, it is believed that one or more C 6 F 13 It is believed that the presence of the chains may enable the lipid entities to separate from the hydrocarbon lipid membrane components (see J. Am. Chem. Soc. 2007, 129, 9037-9043; J. Phsy. Chem. B, 2008, 112, 8250-8256; J. Am. Chem. Soc., 2009, 131, 12091-12093).

[0071] In some embodiments, the lipid entity may be attached to an amino acid residue that may be at or near the N-terminus of the peptide. In some embodiments, the lipid entity may be attached to an amino acid residue that may be at or near the C-terminus of the peptide.

[0072] In some embodiments, the presence of at least one alkene in the structure of lipid entity provides increased fluidity in membrane (i.e., greater ability to move in membrane) compared to a similar lipid entity that lacks at least one alkene.In some embodiments, lipid entities with higher fluidity can provide enhanced activity against targets with low density in membrane (e.g., receptors, ion channels, or enzymes).Without wishing to be bound by theory, lipid entities with higher ability to move in membrane may encounter targets with low density faster than lipid entities with lower mobility in membrane.

[0073] The lipid entity can be linked to the peptide via a linker entity. Thus, the composition can have the formula: [ka]

[0074] In some embodiments, the linker entity can have a length of about 2 Å to 300 Å, inclusive. In some embodiments, the linker entity is 30 Å to 150 Å, inclusive.

[0075] In some embodiments, the linker entity is attached to an amino acid residue that may be at or near the N-terminus of the peptide. In some embodiments, the linker entity may be attached to an amino acid residue that may be at or near the C-terminus of the peptide.

[0076] In some embodiments, the linker entity can include one or more amino acids, which can be either natural or synthetic. For example, the linker entity can include glycine, alanine, leucine, methionine, phenylalanine, tryptophan, lysine, glutamine, glutamic acid, serine, proline, valine, isoleucine, cysteine, tyrosine, histidine, arginine, asparagine, aspartic acid, threonine, or combinations thereof.

[0077] In some embodiments, the linker entity can comprise a peptide ("peptide linker"), e.g., about 2-20 amino acid residues in length, or about 5-10 amino acid residues in length. According to various embodiments, the peptide linker can be designed such that one or more alpha helices are formed between the peptides described herein and the lipid entity. In some embodiments, the peptide linker can comprise multiple alpha helices. In some embodiments, the multiple alpha helices are contiguous. In some embodiments, the multiple alpha helices are 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, or more alpha helices.

[0078] In some embodiments, the peptide linker can include a repeating unit, such as a plurality of repeating glycine-asparagine (GN) units. In some embodiments, the peptide linker can include an epitope tag (e.g., a c-Myc tag) or other marker to allow for identification and / or characterization of the provided agents and their fates in vitro and / or in vivo.

[0079] In some embodiments, the peptide linker can include the amino acid sequence of GGK or GG.

[0080] In some embodiments, the linker entity can comprise a non-peptide entity ("non-peptide linker"). In some embodiments, the linker entity can comprise ethylene glycol and / or aminoethylethanolamine (AEEA). In some embodiments, the non-peptide linker can be a synthetic polymer. According to various embodiments, the synthetic polymer can be of any of a variety of lengths. In some embodiments, the linker entity comprising a synthetic polymer comprises a monomeric unit of a polymer. In some embodiments, the linker entity comprising a synthetic polymer comprises two or more monomeric units of a synthetic polymer (e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 100 or more monomeric units).

[0081] In some embodiments, the linker entity can comprise polyethylene glycol (PEG). In some embodiments, the average number of ethylene glycol units in the PEG is 2-20, such as 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20. In some embodiments, the linker entity can comprise PEG8, which means that the average number of ethylene glycol units is 8.

[0082] Non-limiting examples of suitable polymer linkers include linkers having one or more monomer units according to one of the following formulas: [ka] In the formula, n represents an integer equal to or greater than 1. In some embodiments, n is an integer from 2 to 50, 4 to 24, and / or 8 to 24 (inclusive).

[0083] In some embodiments, the linker entity can have any one of the following structures: [ka] [ka] [ka]

[0084] In some embodiments, the linker entity may comprise a monosaccharide, oligosaccharide, or polysaccharide, such as glucose, fructose, galactose, inulin, or a trisaccharide.

[0085] In some embodiments, the linker entity may include 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDAC), benzophenone-4-isothiocyanate, bis-((N-iodoacetyl)piperazinyl)sulfonerhodamine, succinimidyl 2-(2-pyridyldithio)propionate (SPDP), 4-azido-2,3,5,6-tetrafluorobenzoic acid (ATFB), (N-((2-pyridylthio)ethyl)-4-azidosalicylamide), succinimidyl trans-4-(maleimidylmethyl)cyclohexane-1-carboxylate (SMCC), and / or N-(t-BOC)-aminooxyacetic acid. Those skilled in the art will be able to identify additional potential linker entities according to known methods.

[0086] In some embodiments, the linker entity may include both peptide and non-peptide entities, for example, the linker entity may include both PEG and a peptide (e.g., a peptide having the amino acid sequence GGK or GG).

[0087] In some embodiments, the linker entity is formed, at least in part, as a result of a click reaction, such as an azide-alkyne Huisgen cycloaddition reaction.

[0088] Additional examples of lipid entities, linker entities, and methods of lipidation can be found in US20160052982 and US20190022168, the contents of each of which are incorporated herein by reference.

[0089] Any embodiment of the lipid entity described herein can be combined with any embodiment of the linker entity described herein. For example, combinations of lipid entities and linker entities include palmitic acid-γ-glutamic acid, palmitic acid-lysine, palmitic acid-γ-glutamic acid-lysine, myristic acid-lysine, 17-carboxy-1-oxo-heptadecyl-γ-glutamic acid-(AEEA) 2 , AEEA-PEG, or 17-carboxy-1-oxo-heptadecyl-γ-glutamic acid-(AEEA)2 - May contain lysine.

[0090] In some embodiments, the combination of lipid entity and linker entity is [ka] where the wavy line indicates the point of attachment to the peptide.

[0091] In some embodiments, when lipidation occurs at the N-terminus of the peptide, the peptide may be selected from the group consisting of palmitic acid, myristic acid, palmitic acid-γ-glutamic acid, palmitic acid-γ-glutamic acid-2xOEG, 17-carboxy-1-oxo-heptadecyl-γ-glutamic acid-(AEEA). 2 Alternatively, cholesterol may be attached to the N-terminus of the peptide.

[0092] In some embodiments, when lipidation occurs at the C-terminus of the peptide, it is selected from palmitic acid-lysine, palmitic acid-gamma-glutamic acid-lysine, myristic acid-lysine, or 17-carboxy-1-oxo-heptadecyl-gamma-glutamic acid-(AEEA). 2 The -lysine may be attached to the C-terminus of the peptide.

[0093] In some embodiments, the compositions described herein are water soluble.

[0094] Administration of the Composition The therapeutically effective amount of the composition according to the present disclosure can vary within a wide range and can be determined by methods known in the art. For example, the composition can be administered according to body weight. Such dosage is adjusted to the individual requirements in each particular case, including the particular composition administered, the route of administration, the condition being treated, as well as the patient being treated. In another embodiment, the composition can be administered in a fixed dose, e.g., a dose that is not adjusted according to body weight. In general, for oral or parenteral administration to adults, a daily dosage of about 0.5 mg to about 1000 mg is appropriate, although the upper limit may be exceeded if indicated. The dosage can be about 5 mg to about 500 mg, e.g., about 5 mg to about 400 mg, about 5 mg to about 300 mg, about 5 mg to about 200 mg per day. The daily dosage can be administered as a single dose or divided doses, or, for parenteral administration, as a continuous infusion.

[0095] A therapeutically effective amount of a composition is that amount that provides an objectively discernible improvement as recognized by the clinician or other qualified observer.

[0096] The present disclosure also provides a pharmaceutical composition comprising a peptide composition as described herein and a pharma- ceutically acceptable carrier. The pharmaceutical composition can be included in a container, pack, or dispenser with instructions for administration. The compositions described herein can be administered topically, orally, intranasally, transdermally, pulmonary, inhalation, buccal, sublingually, intraperitoneally, subcutaneously, intramuscularly, intravenously, rectally, intrapleurally, intrathecally, or parenterally. In one embodiment, the composition is administered topically. For example, the composition is administered in the form of eye drops. In one embodiment, the composition is administered orally. In some embodiments, the composition is administered intravenously. In some embodiments, the composition is administered intrathecally. Those skilled in the art will recognize the advantages of certain routes of administration.

[0097] Dosage regimens utilizing the compositions described herein are selected according to a variety of factors, including the species, ethnicity, age, weight, sex, and medical condition of the patient; the severity of the condition being treated; the route of administration; the renal and hepatic function of the patient; and the particular composition being used. An ordinarily skilled physician or veterinarian can readily determine and prescribe the effective amount of the drug required to prevent, counter, or arrest the progress of the condition.

[0098] The compositions described herein can be administered once a day, twice a day, three times a day, four or more times a day, or once every few days.

[0099] Techniques for formulating and administering the compositions disclosed herein can be found in Remington: the Science and Practice of Pharmacy, 19th edition, Mack Publishing Co., Easton, Pa. (1995). In some embodiments, the lipidated peptides described herein, and their pharma- ceutically acceptable salts, are used in pharmaceutical preparations in combination with a pharma- ceutically acceptable carrier or diluent. Suitable pharma- ceutically acceptable carriers include inert solid fillers or diluents, as well as sterile aqueous or organic solutions. The lipidated peptides will be present in such pharmaceutical compositions in an amount sufficient to provide the desired dosage within the range described herein.

[0100] In some embodiments, the pharmaceutical composition may be formulated for topical administration. Formulations suitable for topical administration include, but are not limited to, liquid and / or semi-liquid preparations, such as liniments, lotions, oil-in-water and / or water-in-oil emulsions, such as creams, ointments, and / or pastes, and / or solutions, and / or suspensions. Topically administrable formulations may contain, for example, about 1% to about 10% (w / w) of active ingredient, although the concentration of active ingredient may be as high as the solubility limit of the active ingredient in the solvent. Formulations for topical administration may further include one or more additional ingredients.

[0101] In some embodiments, pharmaceutical composition can be formulated for oral administration.Oral formulations including pharmaceutical composition described herein can be formulated into any commonly used oral form, such as tablet, capsule, pill, troche, lozenge, pastille, cachet, pellet, medicated chewing gum, granule, bulk powder, effervescent or non-effervescent powder or granule, liquid, emulsion, suspension, liquid, wafer, sprinkle, elixir, syrup, buccal form, and oral liquid.Capsule can contain active compound and inert filler and / or diluent, such as pharmaceutically acceptable starch (e.g. corn, potato, or tapioca starch), sugar, artificial sweetener, powdered cellulose, such as crystalline and microcrystalline cellulose, fine flour, gelatin, gum, etc. Useful tablets can be prepared by conventional compression, wet granulation or dry granulation methods and utilize pharma- ceutically acceptable diluents, binders, lubricants, disintegrants, surface modifiers (including surfactants), suspending agents, or stabilizers (including, but not limited to, magnesium stearate, stearic acid, talc, sodium lauryl sulfate, microcrystalline cellulose, calcium carboxymethylcellulose, polyvinylpyrrolidone, gelatin, alginic acid, gum acacia, xanthan gum, sodium citrate, complex silicates, calcium carbonate, glycine, dextrin, sucrose, sorbitol, dicalcium phosphate, calcium sulfate, lactose, kaolin, mannitol, sodium chloride, talc, dry starch, powdered sugar). In some embodiments, surface modifiers include nonionic and anionic surface modifiers. For example, surface modifiers include, but are not limited to, poloxamer 188, benzalkonium chloride, calcium stearate, cetostearyl alcohol, cetomacrogol emulsifying wax, sorbitan esters, colloidal silicon dioxide, phosphate salts, sodium dodecyl sulfate, magnesium aluminum silicate, and triethanolamine.Oral formulations herein can utilize standard delayed or time-release formulations to alter the absorption of active compounds.Oral formulations can also consist of administering the active ingredient in water or fruit juice, with appropriate solubilizers or emulsifiers as needed.

[0102] treatment method The compositions described herein can be used to treat a variety of conditions including pain, such as neuropathic pain, ocular pain, chronic pain, pain due to chemotherapy or radiation, pain due to nerve injury or neurodegeneration, and pain due to inflammatory conditions, paresthesia, or allodynia. The compositions described herein can also be used to treat inflammatory conditions, such as ocular inflammation, retinal inflammation, dry eye, uveitis, allergic conjunctivitis, and inflammation due to nerve injury or neurodegeneration.

[0103] In one embodiment, the present disclosure provides a method for treating neuropathic pain using the compositions described herein. Neuropathic pain according to the present disclosure is pain initiated or caused by a primary lesion or dysfunction in the nervous system. Neuropathic pain can be divided into "peripheral" (originating from the peripheral nervous system) and "central" (originating from the brain or spinal cord). For example, neuropathic pain syndromes include post-herpetic neuralgia (caused by varicella zoster virus), nerve root avulsion injury, painful traumatic mononeuropathy, painful polyneuropathy (especially due to diabetes), central pain syndromes (which can be caused by virtually any lesion at any level of the nervous system), post-operative pain syndromes (e.g., post-mastectomy syndrome, post-thoracotomy syndrome, or phantom pain), and complex regional pain syndromes (e.g., reflex sympathetic dystrophy or caustic pain).

[0104] Neuropathic pain has typical symptoms such as paresthesia (spontaneous or evoked burning pain, often with an overlapping tingling component), but the pain can also be deep and aching. Other sensations such as hyperesthesia, hyperalgesia, allodynia (pain with normal oxygen stimuli), and hypersensitivity (particularly unpleasant and exaggerated pain response) can also occur. The compositions of the present disclosure can be administered to improve at least one of these symptoms.

[0105] Current treatments for neuropathic pain generally only aim to reduce symptoms by suppressing nerve activity. Thus, treatment options such as nonsteroidal anti-inflammatory drugs (NSAIDS), antidepressants, anticonvulsants, baclofen, neuromodulation, or opiates, do not target specific etiology, but mainly relieve symptoms through nonspecific reduction of neuronal hyperexcitability. The compositions of the present disclosure can be administered in combination with current treatments for treating neuropathic pain. For example, the compositions of the present disclosure can be administered in combination with NSAIDs, antidepressants, anticonvulsants, baclofen, neuromodulation, or opiates for treating neuropathic pain.

[0106] In another aspect, the present disclosure provides a method for treating eye pain using the compositions described herein.Eye pain can occur simultaneously with many conditions, including but not limited to trauma from accidental or surgical injury, uveitis, dry eye, and diabetic neuropathy.Standard treatment for eye pain is usually either topical administration of NSAIDs or oral administration of painkillers such as NSAIDS or opioids such as hydrocodone.In some embodiments, the compositions of the present disclosure can be administered in combination with NSAIDs or opioids to treat eye pain.

[0107] In another aspect, the present disclosure provides a method for treating chronic pain using the compositions described herein. The compositions described herein can be used as non-opioid analgesics for treating chronic pain.

[0108] In another aspect, the present disclosure provides a method of treating pain resulting from chemotherapy or radiation. Commonly used chemotherapy drugs such as vincristine sulfate, paclitaxel, and cisplatin can cause what is known as chemotherapy-induced peripheral neuropathy or paresthesias.

[0109] In another aspect, the disclosure provides methods of treating pain resulting from nerve injury or nerve degeneration using the compositions described herein.

[0110] In another aspect, the disclosure provides methods of treating pain resulting from an inflammatory condition, dysesthesias, or allodynia using the compositions described herein.

[0111] The compositions described herein can be administered in combination with analgesics to treat pain. Examples of analgesics include, but are not limited to, paracetamol, NSAIDs, COX-2 inhibitors, opioids, and medical cannabis.

[0112] In another aspect, the present disclosure provides a method for treating ocular inflammation or retinal inflammation using the compositions described herein. Ocular inflammation can be caused by ocular microbial infection. Such infection can be fungal, viral, or bacterial. Current therapies for treating ocular inflammation include topical administration of anti-cytokine or anti-inflammatory agents. In some embodiments, the compositions of the present disclosure can be administered in combination with anti-cytokine or anti-inflammatory agents for treating ocular inflammation.

[0113] Anti-cytokine or anti-inflammatory agents include, but are not limited to, NF-kappa B inhibitors, glucocorticoids such as corticosteroids, e.g., flucinolonone; NSAIDs such as sulindac and tepoxalin; antioxidants such as dithiocarbamates; and other compounds such as autologous blood-derived products, e.g., sulfasalazine [2-hydroxy-5-[-4-[C2-pyridinylamino)sulfonyl]azo]benzoic acid], clonidine, and orthocaine.

[0114] The compositions described herein can also be used to treat dry eye. Dry eye is primarily caused by the breakdown of the preocular tear film, leading to dehydration of the exposed external surface. Dry eye can be diagnosed by a comprehensive eye examination. The examination, which focuses on evaluating the quantity and quality of tears produced by the eye, can include: (a) a patient's medical history to determine the patient's symptoms and note any general health problems, medications, or environmental factors that may be contributing to the dry eye problem; (b) an external examination of the eye (including eyelid structure and blinking); (c) evaluation of the eyelids and cornea using bright light and magnification; and (d) measurement of tear quantity and quality for abnormalities. Special dyes can be placed in the eye to better observe tear flow and highlight changes in the external surface of the eye caused by insufficient tear production.

[0115] Without wishing to be bound by theory, there is a theoretical justification that ocular inflammation as a result of pro-inflammatory cytokines and growth factors plays a major role in the underlying cause of dry eye.Therefore, topically administered anti-cytokine or anti-inflammatory agents are often used to treat dry eye.In some embodiments, the composition of the present disclosure can be administered in combination with anti-cytokine or anti-inflammatory agents to treat dry eye.

[0116] In some embodiments, a therapeutically effective amount for treating ocular inflammation is an amount that reduces the level of inflammation in a subject by at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 95% compared to a placebo.

[0117] In some embodiments, a therapeutically effective amount for treating dry eye is an amount that increases tear production in a subject by at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 100%, or at least 150% compared to placebo.

[0118] The compositions described herein can be used to treat uveitis, including non-infectious uveitis. Uveitis refers to inflammation in one or both eyes of a patient. Uveitis is a widespread inflammatory disease of the eye, specifically the uvea. There are three basic layers of the eye: the sclera and cornea on the outside, the retina on the inside, and the uvea in between. The uvea is primarily made up of blood vessels and connective tissue containing pigment cells. The various parts of the uvea are the iris in front, the ciliary body in the middle, and behind these, the choroid, which surrounds most of the eye. Sometimes, uveitis affects parts of the eye other than the uvea, such as the retina, vitreous, and optic nerve. The type of uveitis can be based on which part of the eye is affected. For example, anterior uveitis is inflammation of the front of the eye, called iritis or iridocyclitis; intermediate uveitis is inflammation of the middle part of the eye, or squamous cellulitis or vitritis; posterior uveitis is inflammation of the back of the eye, such as choroiditis, retinal vasculitis, retinitis, neuroretinitis, retina-choroiditis, or chorioretinitis.

[0119] Symptoms of uveitis generally include redness, blurred vision, pain, light sensitivity, and floaters or flashes of light.

[0120] Non-infectious uveitis may be due to injury to the eye or disease elsewhere in the patient's body. Typically, steroids or immunosuppressants are used to treat non-infectious uveitis. In some embodiments, the compositions described herein can be used in combination with steroids or immunosuppressants to treat non-infectious uveitis.

[0121] The compositions described herein can be used to treat allergic conjunctivitis. Allergic conjunctivitis is an inflammation of the eye caused by an allergic reaction to substances such as pollen or mold spores. Antihistamines, decongestants, or steroids are usually used to treat allergic conjunctivitis. In some embodiments, the compositions described herein can be used in combination with antihistamines, decongestants, or steroids to treat allergic conjunctivitis.

[0122] The compositions described herein can be used to treat inflammation resulting from nerve injury or nerve degeneration.

[0123] The compositions described herein can be used to prevent any one of the diseases, disorders, or conditions described herein.For example, the compositions described herein can be used to prevent neuropathic pain, eye pain, eye inflammation, dry eye, uveitis, or allergic conjunctivitis.

[0124] For combination therapies including a first therapeutic agent (e.g., a composition of the present disclosure) and a second therapeutic agent (e.g., an anti-inflammatory agent, an opioid, an NSAID, or an antidepressant), the first therapeutic agent can be administered simultaneously with the second therapeutic agent; the first therapeutic agent can be administered before the second therapeutic agent; or the first therapeutic agent can be administered after the second therapeutic agent. Administration of the first and second therapeutic agents can be separated by minutes or hours, e.g., 1 hour, 2 hours, 3 hours, 4 hours, 5 hours, or 6 hours.

[0125] definition Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the art to which the present invention pertains. Although other methods and materials similar or equivalent to those described herein can be used in the practice of the present invention, the preferred materials and methods are described herein. It is to be understood that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting.

[0126] The term "at least one" as used herein in connection with a list of one or more elements should be understood to mean at least one element selected from any one or more of the elements in the list of elements, but not necessarily including at least one of every element specifically listed in the list of elements and excluding combinations of elements in the list of elements. This definition also allows for elements other than those specifically identified in the list of elements to which the term "at least one" refers, whether or not related to the specifically identified elements, may optionally be present. 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") can refer, in one embodiment, to at least one A, optionally including more than one A, and no B (and optionally including elements other than B); in another embodiment, to at least one B, optionally including more than one B, and no A (and optionally including elements other than A); in yet another embodiment, to at least one A, optionally including more than one A, and including at least one B, optionally including more than one B (and optionally including other elements).

[0127] The term "and / or" as used herein should be understood to mean "either or both" of the elements so conjoined, i.e., elements that are conjunctively present in some cases and disjunctively present in other cases. Multiple elements listed with "and / or" should be interpreted in the same manner, i.e., "one or more" of the elements so conjoined. Other elements than the elements specifically identified by the term "and / or" can optionally be present, whether or not related to the specifically identified elements. Thus, as a non-limiting example, a reference to "A and / or B", when used in conjunction with open-ended language such as "comprises", can refer in one embodiment to only A (optionally including elements other than B); in another embodiment to only B (optionally including elements other than A); in yet another embodiment to both A and B (optionally including other elements); and so forth.

[0128] The terms "peptide", "polypeptide" and "protein" are used interchangeably herein and typically refer to a molecule that includes a chain of two or more amino acids (e.g., most typically L-amino acids, but also including, e.g., D-amino acids, modified amino acids, amino acid analogs, and amino acid mimetics). Peptides can be naturally occurring, synthetically produced, or recombinantly expressed. Peptides can also include additional groups that modify the amino acid chain, such as functional groups added via post-translational modifications. Examples of post-translational modifications include, but are not limited to, acetylation, alkylation (including methylation), biotinylation, glutamylation, glycylation, glycosylation, isoprenylation, lipoylation, phosphopantetheinylation, phosphorylation, selenylation, and C-terminal amidation. The term peptide also includes peptides that include modifications of the amino and / or carboxyl termini. Modifications of the terminal amino group include, but are not limited to, desamino, N-lower alkyl, N-di-lower alkyl, and N-acyl modifications. Modifications of the terminal carboxy group include, but are not limited to, amide, lower alkyl amide, dialkyl amide, and lower alkyl ester modifications (e.g., where the lower alkyl is C 1~C 4 The term peptide also includes modifications of amino acids between the amino and carboxy termini, such as, but not limited to, those listed above. The term peptide also includes peptides that have been modified to include one or more detectable labels.

[0129] As used herein, the term "amino acid residue" refers to an amino acid that is incorporated into a peptide by an amide bond or an amide bond mimetic. The amino acid can be either natural or synthetic.

[0130] The terminal amino acid at one end of a peptide chain typically has a free amino group (i.e., the amino terminus). The terminal amino acid at the other end of the chain usually has a free carboxyl group (i.e., the carboxy terminus). The amino acids that make up a peptide are usually numbered sequentially, starting from the amino terminus and proceeding toward the carboxy terminus of the peptide.

[0131] The term "analog" as used herein refers to a variant or mutant polypeptide having one or more amino acid modifications compared to the wild type.

[0132] As used herein, the terms "treat," "treating," "treatment," and the like refer to reducing or ameliorating a disorder and / or its associated symptoms. Although not exclusive, it will be understood that treating a disorder or condition does not require that the disorder or its associated symptoms be completely eliminated. The terms "treat," "treating," or "treatment" do not include prevention.

[0133] As used herein, the term "preventing" refers to reducing or eliminating the onset of symptoms or complications of a disease, condition, or disorder.

[0134] As used herein, a "subject" can be any mammal, such as a human, non-human primate, mouse, rat, dog, cat, cow, horse, pig, sheep, goat, camel. In some embodiments, the subject is a human.

[0135] The term "pharmaceutical composition" refers to a mixture of the peptide composition disclosed herein with other chemical components, such as diluents or carriers. The pharmaceutical composition facilitates administration of the peptide composition to an organism, such as a human. The pharmaceutical composition can also be obtained by reacting the compound with an inorganic or organic acid, such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, salicylic acid, and the like.

[0136] "Pharmaceutically acceptable carrier" means a carrier useful for preparing a pharmaceutical composition that is generally safe, non-toxic, and not biologically or otherwise harmful, and includes excipients acceptable for veterinary and human pharmaceutical use. Pharmaceutical carriers useful for preparing the compositions herein can be solid, liquid, or gaseous; that is, the compositions can take the form of tablets, pills, capsules, suppositories, powders, enteric coated or other protected preparations (e.g., binding to ion exchange resins or packaging in lipid-protein vesicles), sustained release preparations, solutions, suspensions, elixirs, aerosols, and the like. Carriers can be selected from a variety of oils, including those of petroleum, animal, vegetable, or synthetic origin, such as peanut oil, soybean oil, mineral oil, sesame oil, and the like. Water, saline, aqueous dextrose, and glycols are preferred liquid carriers, particularly for injectable solutions (when isotonic with blood). For example, formulations for intravenous administration include sterile aqueous solutions of the active ingredient, prepared by dissolving the solid active ingredient in water to produce an aqueous solution, and sterilizing the solution. Suitable pharmaceutical excipients include starch, cellulose, talc, glucose, lactose, talc, gelatin, malt, rice, flour, chalk, silica, magnesium stearate, sodium stearate, glycerol monostearate, sodium chloride, dried skim milk, glycerol, propylene glycol, water, ethanol, etc. The composition may be subjected to conventional pharmaceutical additives such as preservatives, stabilizers, wetting agents, or emulsifiers, salts for adjusting osmotic pressure, buffers, etc. Suitable pharmaceutical carriers and their formulations are described in Remington's Pharmaceutical Sciences by EW Martin. Such compositions will in any event contain an effective amount of active compound together with a suitable carrier to prepare the appropriate dosage form for proper administration to the recipient.

[0137] The term "therapeutically effective amount" as used herein refers to an amount of a formulation to treat, ameliorate, or prevent a specified disease or condition, or to exhibit a detectable therapeutic or inhibitory effect. This effect can be detected by any assay method known in the art. The precise effective amount for a subject will vary depending on the subject's weight, size, and health; the nature and extent of the condition; and the therapeutic agent or combination of therapeutic agents selected for administration. The therapeutically effective amount for a given situation can be determined by routine experimentation that is within the skill and judgment of the clinician.

[0138] As used herein, the singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise. Thus, for example, reference to a "solvent" includes a combination of two or more such solvents, reference to a "peptide" includes one or more peptides or mixtures of peptides, reference to a "drug" includes one or more drugs, reference to a "device" includes one or more devices, etc. As used herein, unless specifically stated or clear from the context, the term "or" is understood to be inclusive and encompasses both "or" and "and."

[0139] Throughout this specification the word "comprising" or variations such as "comprises" or "comprising" will be understood to mean the inclusion of a stated element, integer, or step, or group of elements, integers, or steps, but not the exclusion of other elements, integers, steps, or groups of elements, integers, steps.

[0140] As used herein, unless specifically stated or clear from the context, the term "about" is understood to mean within 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, 0.5%, 0.1%, 0.05%, or 0.01% of the stated value. Unless clear from the context, all numerical values ​​provided herein are modified by the term "about." EXAMPLES

[0141] Example 1 Synthesis of lipidated peptides

[0142] All peptides were prepared using the Fmoc-tBu method. This utilizes an Fmoc protecting group at the N-terminus with a side chain protecting group that is either tBu or Trt depending on the specific functional group present. Activation utilized diisopropoylcarbodiimide in the presence of hydroxybenzotriazole (HOBT) to minimize racemization in DMF. Removal of Fmoc is facilitated using 20% ​​piperidine in DMF at a ratio of 10 ml / g resin. The peptide resin is washed with DMF and IpOH (10 ml of solvent per g of resin) after the deblocking and coupling steps. Once the linear peptide sequence is complete, the peptide is cleaved from the solid support and simultaneously deprotected at the side chain using acidolytic cleavage with TFA and cationic scavengers (water, triisopropylsilane, 3,6-dioxa-1,8-octanedithiol (DODT), thioanisole, phenol) that are added depending on the specific side chain protecting group present. Cleavage times are typically 60 min to 120 min. The crude peptide is isolated by filtration to remove spent resin beads, followed by precipitation in ice-cold methyl t-butyl ether. The precipitated peptide is collected using a sintered glass funnel and dried in a drying oven until a constant weight is obtained.

[0143] The crude peptides are then redissolved in an appropriate buffer system (e.g., 20% AcOH in water) and purified by preparative reverse-phase HPLC. After purification, fractions meeting the minimum pooling criteria of >95% are pooled and subsequently lyophilized. The final lyophilized peptides are analyzed by analytical RP-HPLC, ESI-MS, and other analytical methods such as Karl Fischer for water analysis, elemental nitrogen analysis for peptide content, amino acid analysis, etc.

[0144] Cell culture assays

[0145] A PhatHunter® β-Arrestin GPCR cell line was used that co-expresses a ProLink™ (PK) tagged GPCR (human chemokine-like receptor 1, CMKLR1) and an enzyme acceptor (EA) tagged β-arrestin. Activation of the GPCR-PK induces recruitment of β-arrestin-EA, forcing complementation of two β-galactosidase enzyme fragments (EA and PK). The resulting functional enzyme hydrolyzes a substrate to generate a chemiluminescent signal. The PathHunter® β-Arrestin assay uses a technology developed by DiscoverX called Enzyme Fragment Complementation (EFC), which uses β-galactosidase (β-Gal) as a functional reporter, to monitor GPCR activation in a homogenous, non-imaging assay format. The enzyme is split into two inactive complementary parts (the enzyme acceptor EA and the ProLink PK) that are expressed as a fusion protein in cells. The EA is fused to β-arrestin and the PK is fused to the GPCR of interest. Upon GPCR activation and re-recruitment of β-arrestin to the receptor, complementation of the ED and EA occurs, restoring β-Gal activity as measured using chemiluminescent PathHunter® detection reagents.

[0146] Assay Design:

[0147] (a) Cell Treatment: 1. PathHunter cell lines co-expressing ProLink™ (PK) tagged GPCR (human chemokine-like receptor 1, CMKLR1) and enzyme acceptor (EA) tagged β-arrestin were grown from freezer stocks following standard procedures. 2. Cells were seeded in a total volume of 20 μL into white-walled 384-well microplates and incubated at 37°C for the appropriate time before testing.

[0148] (b) Agonist format: 1. For agonist measurements, cells were incubated with peptide to induce a response. 2. Intermediate dilutions of peptide stocks were made to generate 5X samples in assay buffer. 3. 5 μL of 5X peptide was added to cells and incubated at 37° C. or room temperature for 90-180 minutes. Vehicle concentration was 1%.

[0149] (c) Signal detection: 1. Assay signal was generated by a single addition of 12.5 or 15 μL (50% v / v) of PathHunter detection reagent cocktail, followed by incubation for 1 hour at room temperature. 2. After signal generation, the microplate was read using a PerkinElmer Envision® instrument for chemiluminescent signal detection.

[0150] (d) Data Analysis: 1. Compound activity was analyzed using the Chemical and Biological Information System (CBIS) data analysis suite (ChemInnovation, CA). 2. For agonist mode assays, percent activity was calculated using the following formula: % Activity = 100% x (mean RFU of test samples - mean RFU of vehicle control) / (mean MAX control ligand - mean RFU of vehicle control). RFU means relative fluorescence units.

[0151] [Table 1]

[0152] Example 2 In Figures 1 and 2, the assessment of corneal barrier function is performed as follows. Corneal staining was measured by the permeation of Oregon Green Dextran (OGD). For baseline assessment, 0.5 μL of OGD was injected into the cornea of ​​both eyes, and the mice were housed in the dark for 1 min, then washed with balanced salt solution (BSS) and imaged. For corneal permeability assessment on day 5, 0.5 μL of OGD was injected into the cornea of ​​both eyes, and the mice were immediately housed in the dark for 1 min, followed by euthanasia and immediate imaging. The eyes were washed with 2 mL of BSS. Digital images were captured and the mean fluorescence intensity was measured using NIS Elements imaging software (Nikon).

[0153] In Figures 3 and 5, immunohistochemistry is used to detect CD4+ T cells. Enucleated mouse eyes with intact conjunctiva were suspended in optimal cutting temperature (OCT) compound and flash frozen in liquid nitrogen. Immunohistochemistry was performed on 6 micrometer frozen sections to detect and count the number of cells in the conjunctival epithelium that stained positively for CD4, biotinylated secondary antibody, and NovaRED peroxidase. Nikon imaging software was used to count positively stained cells in the conjunctiva.

[0154] In Figures 4 and 6, conjunctival goblet cell (GC) measurements are performed as follows: Enucleated mouse eyes with intact conjunctiva were fixed in 10% formalin and embedded in paraffin. Six micrometer sections were stained with periodic acid-Schiff's reagent. GC density in the superior and inferior conjunctiva was measured using Nikon imaging software.

Claims

1. 1. A composition comprising a peptide selected from AAFY-Nle-PSQYA-Tic-dA (SEQ ID NO:2), AG-dY-F-Nle-P-dS-Aad-FA-Tic-dA (SEQ ID NO:3), dA-G-dY-F-Nle-P-dS-Q-fF-dA-Tic-dA (SEQ ID NO:4), NorV-G-dY-F-Nle-P-dS-QF-dA-Tic-dA (SEQ ID NO:5), AG-dY-F-Nle-P-dS-QFA-Tic-dS (SEQ ID NO:6), and dA-G-dY-F-Nle-P-dS-QF-dA-Tic-dA (SEQ ID NO:7), wherein palmitic acid-PEG8 is linked at or near the N-terminus of the peptide. Here, Nle refers to norleucine, Tic refers to (S)-1,2,3,4-tetrahydroisoquinoline-3-carboxylic acid, dA refers to D-alanine, dY refers to D-tyrosine, dS refers to D-serine, Aad refers to α-aminoadipic acid, fF refers to fluorinated phenylalanine, and NorV refers to norvaline.

2. A pharmaceutical composition comprising the composition of claim 1 and a pharma- ceutically acceptable carrier.

3. 10. The composition of claim 1 for use in a method for treating pain in a subject in need thereof.

4. 4. The composition of claim 3, wherein the pain is ocular pain; chronic pain; neuropathic pain; pain due to chemotherapy or radiation therapy; pain due to nerve injury or neurodegeneration; or pain due to an inflammatory condition, paresthesia, or allodynia.

5. The method further comprises administering an analgesic; 4. The composition of claim 3, optionally wherein the analgesic is paracetamol, a nonsteroidal anti-inflammatory drug, a COX-2 inhibitor, an opioid, or medical cannabis.

6. 10. The composition of claim 1 for use in a method of treating an inflammatory condition in a subject in need thereof.

7. 7. The composition of claim 6, wherein the inflammatory condition is due to ocular inflammation; dry eye disease; anterior uveitis; intermediate uveitis; posterior uveitis allergic conjunctivitis; retinal inflammatory disease; or nerve damage or degeneration.

Images