Compounds and methods for treating neuropathic ocular pain

Abstract

The present disclosure relates to compositions and methods for treating ocular inflammation and / or ocular pain, e.g., neuropathic ocular pain. One aspect of the present disclosure relates to methods of administering soluble lipidated ligand agents comprising human chemerin fragments or chemerin analogs for the treatment of neuropathic ocular pain.

Description

COMPOUNDS AND METHODS FOR TREATING NEUROPATHIC OCULAR PAINCROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the benefit of and priority to U.S. Provisional Patent Application No. 63 / 662,990, filed June 21, 2024, the entire contents of which are incorporated by reference herein for all purposes.INCORPORATION BY REFERENCE OF SEQUENCE LISTING

[0002] The instant application contains a Sequence Listing which has been submitted electronically in XML format and is hereby incorporated by reference in its entirety. Said XML copy, created on June 15, 2025, is named OKYO_009_001WO_ST26.xml and is 6,276 bytes in size.BACKGROUND OF THE DISCLOSURE

[0003] Neuropathic ocular pain, e.g., neuropathic corneal pain (NCP), is a complex disease affecting the somatosensory nervous system characterized by different underlying etiologies and pathogenesis. After injury to peripheral axons, various inflammatory mediators are released, resulting in sensitization of peripheral nerves and lowering of their thresholds. These inflammatory mediators can affect adjacent nociceptors, activating silent receptors and intensifying the peripheral pain signaling process, leading to peripheral and central sensitization over time. NCP can be caused by among other factors, dry eye disease (DED), refractive surgery, herpes simplex keratitis, herpes zoster ophthalmicus, Sjogren’s syndrome, and chemical bums, but it can also be systemic in origin, as in small fiber polyneuropathy or fibromyalgia.

[0004] One cause of NCP is dry eye disease (DED), which is a multifactorial disease of the tears and the ocular surface, with inflammation playing a part in its pathogenesis. Dry eye disease is a common and often chronic problem, particularly in older adults. Recent global estimates show that about 5-50% of people suffer from dry eye disease, depending on the region. People with dry eyes either do not produce enough tears or their tears are of a poor quality. Tears are produced by several glands in and around the eyelids. Tear production may diminish with age, with various medical conditions or as a side effect of certain medicines. Environmental conditions, such as wind and dry climates, can also decrease tear volume dueto increased tear evaporation. When the normal amount of tear production decreases or tears evaporate too quickly from the eyes, symptoms of dry eye can develop.

[0005] The present disclosure addresses the need of patients suffering from neuropathic ocular pain, e.g., NCP, caused by factors including but not limited to, dry eye disease.SUMMARY OF THE DISCLOSURE

[0006] The present disclosure provides compositions and methods for treating or ameliorating at least one symptom of neuropathic ocular pain or ocular inflammation, e.g., neuropathic corneal pain, in a subject in need thereof.

[0007] In some aspects, provided herein is a method of treating neuropathic corneal pain in a subject in need thereof, the method comprising: administering to the subject a therapeutically effective amount of a soluble lipidated ligand agent comprising: a ligand entity comprising: (a) a human chemerin fragment having the sequence of YFPGQFAFS (SEQ ID NO: 2) or (b) a chemerin analog having the sequence of Y*FLPS*QFA*-Tic-S (SEQ ID NO: 3), wherein the * denotes D-amino acids, and wherein Tic represents (S)-l,2,3,4-tetrahydroisoquinoline- 3 -carboxylic acid; at least one lipid entity covalently linked to the ligand entity at or near the N-terminus, at or near the C-terminus, or at or near both the N-terminus and C-terminus; and an optional linker entity connecting the ligand entity to the at least one lipid entity; wherein the soluble lipidated ligand agent is formulated in a concentration of about 0.05% or about 0.1% w / v; and wherein the soluble lipidated agent is topically administered about 4 times daily to the eye of the subject.

[0008] In some embodiments, the subject in need thereof has a Visual Analog Scale (VAS) pain intensity of at least 30 units, and the method decreases VAS pain intensity by at least 10 units. In some embodiments, the method decreases VAS pain intensity by at least 20 units. In some embodiments, the method decreases VAS pain intensity by at least 30 units.

[0009] In some embodiments, the subject in need thereof has positive In Vivo Confocal Microscopy (IVCM) findings, wherein positive IVCM findings are decreased nerve density and evidence of microneuromas.

[0010] In some embodiments, the subject does not have dry eye disease.

[0011] In some aspects, the present disclosure relates to a method of treating neuropathic ocular pain in a subject in need thereof, the method comprising: administering to the subject a therapeutically effective amount of a soluble lipidated ligand agent comprising: a ligand entity comprising: (a) a human chemerin fragment having thesequence of YFPGQFAFS (SEQ ID NO: 2) or (b) a chemerin analog having the sequence of Y*FLPS*QFA*-Tic-S (SEQ ID NO: 3), wherein the * denotes D-amino acids, and wherein Tic represents (S)-l,2,3,4-tetrahydroisoquinoline-3-carboxylic acid; at least one lipid entity covalently linked to the ligand entity at or near the N-terminus, at or near the C-terminus, or at or near both the N-terminus and C-terminus; and an optional linker entity connecting the ligand entity to the at least one lipid entity; wherein the soluble lipidated ligand agent is formulated in a concentration of about 0.05% or about 0.1% w / v; and wherein the soluble lipidated agent is administered about 4 times daily.

[0012] In some embodiments, the method comprises administering the soluble lipidated ligand agent to the eye of a subject, optionally wherein the soluble lipidated agent is administered topically to the eye of the subject.

[0013] In some embodiments, the method comprises administering a soluble lipidated agent in a formulation comprising about 0.05% w / v of the soluble lipidated agent. In some embodiments, the soluble lipidated agent is administered in a formulation comprising about 0.1% w / v of the soluble lipidated agent.

[0014] In some embodiments, the method comprises administering a soluble lipidated agent, wherein the lipid entity of the soluble lipidated agent is linked to the chemerin fragment or chemerin analog through a linker. In some embodiments, the linker comprises polyethylene glycol, a peptide, or a combination thereof. In some embodiments, the linker is selected from the group consisting of:

[0015] In some embodiments, the method comprises administering the soluble lipidated agent, wherein the chemerin fragment or chemerin analog comprises at least 10 amino acids, at least 15 amino acids, at least 20 amino acids, at least 30 amino acids, at least 40 amino acids, at least 50 amino acids, at least 60 amino acids, at least 70 amino acids, at least 80 amino acids, at least 90 amino acids, at least 100 amino acids, at least 110 amino acids, at least 120 amino acids, at least 130 amino acids, at least 140 amino acids, or at least 150 amino acids.

[0016] In some embodiments, the method comprises administering a soluble lipidated agent wherein the chemerin fragment consists of the sequence YFPGQFAFS (SEQ ID NO: 2).

[0017] In some embodiments, the method comprises administering a soluble lipidated agent wherein the chemerin analog consists of the sequence Y*FLPS*QFA*-Tic-S (SEQ ID NO: 3), wherein the * denotes D-amino acids, and wherein Tic represents (S)-l, 2,3,4- tetrahydroisoquinoline-3-carboxylic acid.

[0018] In some embodiments, the method comprises administering the soluble lipidated agent wherein the chemerin fragment or chemerin analog is resistant to proteolysis.

[0019] In some embodiments, the method comprises administering a soluble lipidated agent wherein the lipid entity is selected from the group consisting of a-linolenic acid, y-linolenic acid, stearidonic acid, eicosapentaenoic acid, docosahexaenoic acid, linoleic acid, dihomo-y- linolenic acid, arachidonic acid, docosatetraenoic acid, palmitoleic acid, vaccenic acid, paullinic acid, oleic acid, elaidic acid, gondoic acid, erucic acid, nervonic acid, mead acid, myristic acid, palmitic acid, stearic acid, l,2-dipalmitoyl-sn-glycero-3 -phosphoethanolamine (DPPE), GM1 ganglioside, GM2 ganglioside, GM3 ganglioside, 1,2-dipalmitoyl-sn-glycero- 3 -phosphocholine (DPPC), l,2-dioleoyl-sn-glycero-3-phospho-L-serine (DOPS), 1,2- dioleoyl-sn-glycero-3 -phosphocholine (DOPC), a glycosphingolipid, a sphingolipid, phosphatidylinositol 4, 5 -bisphosphate (PIP2), a ceramide, cholesterol, ergosterol, phytosterol, a hopanoid, and a steroid. In some embodiments, the lipid entity is selected from the group consisting of a-linolenic acid, y-linolenic acid, palmitic acid, vaccenic acid, oleic acid, and elaidic acid. In some embodiments, the lipid entity is linked at or near the N-terminus of the chemerin fragment or chemerin analog. In some embodiments, the lipid entity is linked at or near the C-terminus of the chemerin fragment or chemerin analog.

[0020] In some embodiments, the method comprises treating a human subject.

[0021] In some embodiments, the chemerin fragment has the structure

[0022] In some embodiments, the method of treating comprises administering the soluble lipidated ligand agent, wherein the method of treating regenerates nerves in the subject, optionally wherein the method reduces microneuromas in the cornea of the subject, increases the sensitivity of the cornea of the subject, and / or increases dendritiform cells in the cornea of the subject.

[0023] In some embodiments, the neuropathic ocular pain of the subject is associated with dry eye disease (DED), refractive surgery, herpes simplex keratitis, herpes zoster ophthalmicus, Sjogren’s syndrome, or chemical bums.

[0024] In some embodiments, the neuropathic ocular pain of the subject is systemic in origin, e.g., the neuropathic ocular pain is associated with small fiber polyneuropathy or fibromyalgia.

[0025] In some embodiments, the method comprises administering the soluble lipidated ligand agent formulated in an ophthalmic formulation comprising (a) about 0.1% to about 0.5% w / v of NaCl; (b) about 25 mM to about 100 mM of phosphate buffered saline; and (c) about 0.05% or about 0.1% w / v of the soluble lipidated ligand agent; wherein the formulation has a pH of about 6.5 to about 8.5, and an osmolality of about 200 to about 450 mOsm / Kg.

[0026] Any aspect or embodiment described herein can be combined with any other aspect or embodiment as disclosed herein. While the disclosure has been described in conjunction with the detailed description thereof, the foregoing description is intended to illustrate and not limit the scope of the disclosure, which is defined by the scope of the appended claims. Other aspects, advantages, and modifications are within the scope of the following claims.

[0027] The patent and scientific literature referred to herein establishes the knowledge that is available to those with skill in the art. All United States patents and published or unpublished United States patent applications cited herein are incorporated by reference. All published foreign patents and patent applications cited herein are hereby incorporated by reference. All other published references, documents, manuscripts and scientific literature cited herein are hereby incorporated by reference.BRIEF DESCRIPTION OF THE DRAWINGS

[0028] FIG. 1 depicts line graphs demonstrating improvement of conjunctival Sum Lissamine Green staining. Data are the change from baseline using 0-8 scale in above median population. P-values are determined against placebo, based on ANCOVA model.

[0029] FIGs. 2A-2B depict bar graphs demonstrating improvement of conjunctival Sum Lissamine Green Staining, in intent-to-treat (FIG.2A) and above median populations.

[0030] FIG. 3 depicts line graphs demonstrating improvement in conjunctival sum staining in intent-to-treat above median population. P values are vs placebo based on ANCOVA Model

[0031] FIG. 4 depicts bar graphs demonstrating conjunctival sum staining at least 25% change from baseline.

[0032] FIG. 5 depicts bar graphs showing significant improvement in tear film break up time, as change from baseline using 0-10 scale in intent-to-Treat population. P values are vs placebo based on Wilcoxon rank sum test

[0033] FIG. 6 depicts bar graphs showing significant improvement in pain, shown as change from baseline using 0-100 Visual Analog Scale (VAS) in Intent-To-Treat Population. P values are vs placebo based on Wilcoxon rank sum test.

[0034] FIGs. 7A-7B depicts bar graphs showing improvement in burning / stinging for responders improving at least 30 units in VAS (FIG. 7 A) and responders improving at least 10 units in VAS (FIG. 7B). Data are the change from baseline to intent-to-treat above median population. P-values are determined against placebo, based on Wilcoxon rank sum test.

[0035] FIG. 8 depicts bar graphs demonstrating responders with improvement in both at day 85, in intent to treat population.

[0036] FIG. 9 depicts line graphs demonstrating conjunctival sum Lissamine Green staining post-controlled adverse environment. Data shown is change from baseline in intent-to-treat above median population. P values are vs placebo based on ANCOVA Model.

[0037] FIG. 10 depicts line graphs demonstrating improvement in burning / stinging (VAS 1- 100) as change from baseline in intent-to-treat above median population. P values are vs placebo based on Wilcoxon rank sum test.

[0038] FIG. 11 depicts the clinical trial design for OK-101.

[0039] FIGs. 12A - 12B depict line graphs showing improvement in conjunctival sum staining shown as change from baseline in intent-to-treat population with Lissamine Green staining (FIG. 12A) and Fluorescein staining (FIG. 12B). P values are vs placebo based on ANCOVA Model.

[0040] FIG. 13 depicts line graphs showing significant improvement in tear film break up time shown as change from baseline in intent-to-treat population. P values are vs placebo based on Wilcoxon rank sum test.

[0041] FIG. 14 depicts line graphs showing significant improvement in pain, shown as change from baseline in intent-to-treat population. P values are vs placebo based on Wilcoxon rank sum test.

[0042] FIGs. 15A - 15B depict line graphs showing significant improvement in burning / stinging (FIG. 15 A) and blurred vision (FIG. 15B), shown as change from baseline in Intent-To-Treat Population. P values are vs placebo based on Wilcoxon rank sum test.

[0043] FIG. 16 depict bar graphs showing significant improvement in symptoms by patient reported daily diary within first two weeks. P values are vs placebo based on Wilcoxon rank sum test.

[0044] FIGs. 17A-17D depict line graphs demonstrating significant improvement in symptoms by patient reported daily diary within the first two weeks, for pain (FIG. 17 A), burning / stinging (FIG. 17B), itching (FIG. 17C), and eye dryness (FIG. 17D). P values are vs placebo based on Wilcoxon rank sum test.

[0045] FIG. 18 depicts bar graphs showing percentage treatment emergent adverse events between treated and placebo groups.DETAILED DESCRIPTION OF THE DISCLOSURE

[0046] In some aspects, provided herein is a method of treating neuropathic corneal pain in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of a soluble lipidated ligand agent comprising a ligand entity comprising: (a) a human chemerin fragment having the sequence of YFPGQFAFS (SEQ ID NO: 2) or (b) a chemerin analog having the sequence of Y*FLPS*QFA*-Tic-S (SEQ ID NO: 3), wherein the * denotes D-amino acids, and wherein Tic represents (S)-l,2,3,4-tetrahydroisoquinoline- 3 -carboxylic acid; at least one lipid entity covalently linked to the ligand entity at or near the N-terminus, at or near the C-terminus, or at or near both the N-terminus and C-terminus; and an optional linker entity connecting the ligand entity to the at least one lipid entity; wherein the soluble lipidated ligand agent is formulated in a concentration of about 0.05% or about 0.1% w / v; and wherein the soluble lipidated agent is topically administered about 4 times daily to the eye of the subject.

[0047] In some embodiments, the subject has a Visual Analog Scale (VAS) pain intensity of at least 30 units, and the method decreases VAS pain intensity by at least 10 units. In some embodiments, the method decreases VAS pain intensity by at least 20 units. In some embodiments, the method decreases VAS pain intensity by at least 30 units.

[0048] VAS pain intensity may be determined after at least 14 days of treatment, at least 21 days of treatment, at least one month of treatment, at least two months of treatment, or at least three months of treatment.

[0049] In some embodiments, the subject has positive / / ? Vivo Confocal Microscopy (IVCM) findings, wherein positive IVCM findings are decreased nerve density and evidence of microneuromas.

[0050] some aspects, provided herein is a method of treating neuropathic ocular pain in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of a soluble lipidated ligand agent comprising: a ligand entity comprising: (a) a human chemerin fragment having the sequence of YFPGQFAFS (SEQ ID NO.: 2) or (b) a chemerin analog having the sequence of Y*FLPS*QFA*-Tic-S (SEQ ID NO.: 3), wherein the * denotes D-amino acids, and wherein Tic represents (S)-l,2,3,4-tetrahydroisoquinoline- 3 -carboxylic acid; at least one lipid entity covalently linked to the ligand entity at or near the N-terminus, at or near the C-terminus, or at or near both the N-terminus and C-terminus; and an optional linker entity connecting the ligand entity to the at least one lipid entity; wherein the soluble lipidated ligand agent is formulated in a concentration of about 0.05% or about 0.1% w / v; and wherein the soluble lipidated agent is administered about 4 times daily.

[0051] Without wishing to be bound by theory, it is believed that dosing of the soluble lipidated ligand agent 4 times daily (QID), e.g., about every 4-6 hours, facilitates pharmacodynamics of the soluble lipidated ligand agent. QID may facilitate near-constant occupancy, and activation, of CMKLR1 in the eye. This constant activation of CMKLR1 may reduce off-target immune effects while maintaining the therapeutic range, facilitating significant benefits in efficacy. High efficacy pain relief is particularly desirable in neuropathic ocular pain, where patients often experience chronic, debilitating discomfort that is poorly managed by existing therapies.

[0052] CMKLR1 is a G protein-coupled receptor which has been shown to modulate nociception. This receptor is expressed in glia, dorsal root ganglion neurons, and immune cells. The endogenous ligand (agonist) for CMKLR1 is chemerin, a 163 amino acid protein. Chemerin, also known as retinoic acid receptor responder protein 2 (RARRES2), tazarotene- induced gene 2 protein (TIG2), or RAR-responsive protein TIG2 is a protein that in humans is encoded by the RARRES2 gene. The amino acid sequence of chemerin, in the inactive pre- prochemerin (homo sapiens) form is shown below in SEQ ID NO: 1.

[0053] NCBI Reference Sequence: NP_002880.1MRRLLIPLAL WLGAVGVGVA ELTEAQRRGL QVALEEFHKH PPVQWAFQET SVESAVDTPF PAGIFVRLEF KLQQTSCRKR DWKKPECKVR PNGRKRKCLA CIKLGSEDKV LGRLVHCPIE TQVLREAEEH QETQCLRVQR AGEDPHSFYF PGQFAFSKAL PRS (SEQ ID NO: 1).

[0054] Pre-prochemerin (having SEQ ID NO: 1) is activated through cleavage of the C- terminus and N-terminus to form a chemerin fragment having an amino acid sequence fromposition 21 to 157 of SEQ ID NO: 1, which can function as an agonist for 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: 4).

[0055] The human chemerin fragments or chemerin analogs of the disclosure may retain some or all of the biological functions of the amino acid sequence of SEQ ID NO: 4, e.g., functioning as an agonist of CMKLR1.

[0056] As provided herein, a human chemerin fragment of the disclosure comprises the amino acid sequence of YFPGQFAFS (SEQ ID NO: 2). In some embodiments, the chemerin fragment comprises at least 10 amino acids, at least 20 amino acids, at least 30 amino acids, at least 40 amino acids, at least 50 amino acids, at least 60 amino acids, at least 70 amino acids, at least 80 amino acids, at least 90 amino acids, at least 100 amino acids, at least 110 amino acids, at least 120 amino acids, at least 130 amino acids, at least 140 amino acids, or at least 150 amino acids. In some embodiments, the chemerin fragment comprises about 5-150 amino acids, about 5-120 amino acids, about 5-100 amino acids, about 5-80 amino acids, about 5-50 amino acids, or about 5-30 amino acids. In some embodiments, the chemerin fragment has an amino acid sequence from position 21 to 157 of SEQ ID NO: 1.

[0057] As provided herein, a chemerin analog comprises the sequence of Y*FLPS*QFA*- Tic-S (SEQ ID NO: 3), wherein the * denotes D-amino acids, and wherein Tic represents (S)- l,2,3,4-tetrahydroisoquinoline-3-carboxylic acid. The chemerin analog can be an analog of either the full length or fragment of chemerin. The chemerin analog can retain some or all of the biological functions of the amino acid sequence of SEQ ID NO: 4, e.g., functioning as an agonist of CMKLR1. The chemerin analog can comprise at least three amino acid modifications, at least five amino acid modifications, or at least ten amino acid modifications. In some embodiments, the amino acid modification is an amino acid substitution. The chemerin analog can be more resistant to proteolysis compared to the unmodified polypeptide. The chemerin analog of SEQ ID NO: 3 is found to be resistant to proteolysis. See Shimamura el al., “Identification of a stable chemerin analog with potent activity toward ChemR23,” Peptides 30, 2009, 1529-1538, the contents of which are incorporated by reference.

[0058] In some embodiments, the chemerin fragment or analog has the structure

[0059] Without wishing to be bound by theory, the pharmacological properties of chemerin or a fragment or analog thereof can be modulated by the choice of the lipid entity. Any of a variety of lipid entities may be utilized in accordance with the present disclosure. According to various embodiments, a lipid entity can comprise an entity capable of insertion into a lipid bilayer (e.g., a cell membrane). In some embodiments, a lipid entity is capable of incorporating into a lipid raft in a lipid bilayer (e.g., a cell membrane).

[0060] In some embodiments, the lipid entity can comprise a saturated or unsaturated fatty acid. The numbers in the lipid name are used to describe the fatty acid chains on the lipid. The numbers are generally presented in the format (number of carbons in fatty acid chain): (number of double bonds in fatty acid chain), e.g., 16:0 would be 16 carbons in the fatty acid chain with zero double bonds. The saturated or unsaturated fatty acid can include 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 can include about 4-24 carbons in the fatty acid chain. The number of double bonds in the fatty acid chain can be in the range of 0- 10, e.g., 0-8, 0-6, 1-8, 1-6. For example, the lipid entity can be C22:0, C22: l, C22:2, C22:3, C22:4, C22:5, C22:6, C20:0, C20: l, C20:2, C20:3, C20:4, C20:5, C20:6, C18:0, C18: l, C18:2, C18:3, C18:4, C18:5, C18:6, C10:0, C10: l, C10:2, C10:3, C10:4, etc.

[0061] For example, the lipid entity can be selected from the group consisting of a-linolenic acid, y-linolenic acid, stearidonic acid, eicosapentaenoic acid, docosahexaenoic acid, linoleic acid, dihomo-y-linolenic acid, arachidonic acid, docosatetraenoic acid, palmitoleic acid, vaccenic acid, paullinic acid, oleic acid, elaidic acid, gondoic acid, erucic acid, nervonic acid, mead acid, myristic acid, palmitic acid, stearic acid, l,2-dipalmitoyl-sn-glycero-3- phosphoethanolamine (DPPE), GM1 ganglioside, GM2 ganglioside, GM3 ganglioside, 1,2- dipalmitoyl-sn-glycero-3 -phosphocholine (DPPC), l,2-dioleoyl-sn-glycero-3-phospho-L- serine (DOPS), l,2-dioleoyl-sn-glycero-3 -phosphocholine (DOPC), a glycosphingolipid, a sphingolipid, phosphatidylinositol 4, 5 -bisphosphate (PIP2), a ceramide, cholesterol, ergosterol, phytosterol, a hopanoid, a steroid, fluorinated-GMl, fluorinated-GM2, and fluorinated-GM3. In some embodiments, the lipid entity can be a-linolenic acid. In some embodiments, the lipid entity can be y-linolenic acid. In some embodiments, the lipid entitycan 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.

[0062] The attachment of a lipid entity to a polypeptide is referred to as lipidation. In some embodiments, lipidation may comprise N-myristoylation. As used herein, “N-myristoylation” refers to the attachment of a myristate to an N-terminal glycine.

[0063] In some embodiments, lipidation may comprise palmitoylation. As used herein “palmitoylation” refers to the creation of a thioester linkage of long-chain fatty acids on one or more cysteine residues present in a peptide or protein.

[0064] In some embodiments, lipidation comprises GPI-anchor addition. As used herein “GPI-anchor addition” refers to the linkage of glycosyl-phosphatidylinositol (GPI) to the C- terminus of a protein.

[0065] In some embodiments, lipidation comprises prenylation. As used herein “prenylation” refers to the creation of a thioether linkage of an isoprenoid lipid (e.g., famesyl (C-15) or geranylgeranyl (C-20)) to a cysteine present in a peptide or protein. In some embodiments, lipidation comprises geranylation. In some embodiments, lipidation includes geranylgeranylation. In some embodiments, lipidation comprises the association of a ligand entity with any compound that is soluble in a cellular membrane (e.g., 10: 1 in equilibrium constant K assoc ^10).

[0066] In some embodiments, lipidation may comprise one or more of the following: attachment of diacylglycerol to the side chain of an N-terminal cysteine of a peptide or protein via the sulfur atom; attachment of O-octanoyl to a serine or threonine of a peptide or protein; and attachment of S-archaeol to a cysteine of a peptide or protein. In some embodiments, lipidation may occur, for example, at any lysine, glutamic acid, aspartic acid, serine, threonine, cysteine, and / or tyrosine. In some embodiments where a chemerin analog comprises one or more ornithine, lipidation may occur at any ornithine.

[0067] In some embodiments, the lipid entity can be linked at or near the N-terminus of chemerin or fragment or analog thereof. In some embodiments, the lipid entity can be linked at or near the C-terminus of chemerin or fragment or analog thereof.

[0068] In some embodiments, lipidation may include fluorination. Fluorination can include the addition of one or more CeFi3 chains. Without wishing to be bound by theory, it is thought that the presence of one or more CeFi3 chains may allow a lipid entity to segregate from 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).

[0069] In some embodiments, the presence of at least one alkene in the structure of a lipid entity provides increased fluidity in a membrane (z.e., greater ability to move within the membrane) as compared to similar lipid entities lacking at least one alkene. In some embodiments, a lipid entity with greater fluidity is able to provide enhanced activity towards targets (e.g., receptors, ion channels, or enzymes) with a low density in a membrane. Without wishing to be bound by theory, it is possible that lipid entities with increased ability to move within a membrane are able to encounter a low density target faster than a lipid entity with less mobility within a membrane.

[0070] The composition of the present disclosure can optionally comprise a linker that links the lipid entity to chemerin or the fragment or analog thereof. For example, the linker can have a length of between about 2 A and 175 A, inclusive. In some embodiments, a linker is between 30 A and 150 A, inclusive.

[0071] In some embodiments, the linker can comprise a peptide. In some embodiments, a peptide linker is between about 2 and 20 amino acid residues in length. In some embodiments, a peptide linker is between about 5 and 10 amino acid residues in length. According to various embodiments, peptide linkers can be designed such that one or more a- helices are formed between chemerin or a fragment or analog thereof and a lipid entity. In some embodiments, a peptide linker may comprise a plurality of a-helices. In some embodiments, the plurality of a-helices is consecutive. In some embodiments, a plurality of a-helices is 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, or more a-helices.

[0072] In some embodiments, a peptide linker can comprise repeating units, for example a plurality of repeating glycine-asparagine (GN) units. In some embodiments, a peptide linker can comprise an epitope tag (e.g., a c-Myc tag) or other marker to allow for identification and / or characterization of provided agents and their fate in vitro and / or in vivo.

[0073] In some embodiments, the linker can comprise a non-peptide entity. In some embodiments, non-peptide linkers may be a synthetic polymer. According to various embodiments, the synthetic polymer may be any of a variety of lengths. In some embodiments, a linker comprising a synthetic polymer comprises a monomeric unit of the polymer. In some embodiments, a linker 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).

[0074] In some embodiments, a linker can comprise at least one molecule of polyethylene glycol (PEG). Specific, non-limiting examples of suitable polymeric linkers include linkers with one or more monomeric units according to one of the following formulas:where n represents an integer greater than or equal to 1. In some embodiments, n is an integer between 2 and 50, 4 and 24, and / or 8 and 24, inclusive.

[0075] In some embodiments, the linker can have any one of the following structures:

[0076] In some embodiments, a linker can comprise l-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-Pyridyldthio)ethyl)-4-Azidosalicylamide), Succinimidyl trans-4-(maleimidylmethyl)cyclohexane- 1 -carboxylate (SMCC), and / or N-(t-BOC)-aminooxyacetic acid. Those of skill in the art will be able to identify additional candidate linkers according to known methods.

[0077] In some embodiments, a linker can comprise both a peptide and a non-peptide entity.

[0078] In some embodiments, a linker is formed, at least in part, as a result of a click reaction as further described below. In some embodiments, the click reaction is an azide-alkyne Huisgen cycloaddition reaction.

[0079] Additional examples of lipid entities, linkers, and methods of lipidation can be found, without limitation, at US20160052982, the contents of which are incorporated herein by reference.Formulations

[0080] In some embodiments, the pharmaceutical composition can be formulated for topical administration, e.g., ophthalmic topical administration. In some embodiments, the pharmaceutical compositions provided herein are formulated for ophthalmic topical administration and comprise: a) about 0.1% to about 0.5% w / v of NaCl; (b) about 25 mM to about 100 mM of phosphate buffered saline; and (c) about 0.025% to about 0.1% w / v of a lipidated chemerin composition that includes a chemerin fragment consisting of the sequence of Y-F-P-G-Q-F-A-F-S (SEQ ID NO: 2) or a chemerin analog consisting of the sequence of Y*-F-L-P-S*-Q-F-A*-Tic-S (SEQ ID NO: 3), wherein * denotes D amino acids and Tic represents 1,2, 3, 4 tetrahydroisoquinoline-3-carboxylic acid, the chemerin fragment or chemerin analog being linked to a lipid entity via a linker; wherein the formulation has a pH of about 6.5 to about 8.5, and an osmolality of about 200 to about 450 mOsm / Kg.

[0081] In some embodiments, a formulation of the present disclosure comprises 0.1% to 0.5% w / v of NaCl (e.g., 0.1% to 0.2%, 0.2% to 0.3%, 0.3% to 0.4% or o.4% to 0.5% w / v of NaCl). In some embodiments, the formulation of the present disclosure comprises 0.3% w / v of NaCl.

[0082] In some embodiments, a formulation of the present disclosure comprises about 0.025% to about 0.1% w / v of a chemerin composition, i.e., a chemerin composition comprising a human chemerin fragment having the sequence of YFPGQFAFS (SEQ ID NO: 2) or comprising a chemerin analog having the sequence of Y*FLPS*QFA*-Tic-S (SEQ ID NO: 3), wherein the * denotes D-amino acids, and wherein Tic represents (S)-l, 2,3,4- tetrahydroisoquinoline-3-carboxylic acid. In some embodiments, a formulation of the presentdisclosure comprises about 0.05% to about 0.1% w / v of a chemerin composition, i.e., a chemerin composition comprising a human chemerin fragment having the sequence of YFPGQFAFS (SEQ ID NO: 2) or comprising a chemerin analog having the sequence of Y*FLPS*QFA*-Tic-S (SEQ ID NO: 3), wherein the * denotes D-amino acids, and wherein Tic represents (S)-l,2,3,4-tetrahydroisoquinoline-3-carboxylic acid.

[0083] In some embodiments, the pharmaceutical compositions provided herein are formulated for ophthalmic topical administration and comprise: a) about 0.1% to about 0.5% w / v of NaCl; (b) about 25 mM to about 100 mM of phosphate buffered saline; and (c) about 0.05% w / v of a lipidated chemerin composition that includes a chemerin fragment consisting of the sequence of Y-F-P-G-Q-F-A-F-S (SEQ ID NO: 2) or a chemerin analog consisting of the sequence of Y*-F-L-P-S*-Q-F-A*-Tic-S (SEQ ID NO: 3), wherein * denotes D amino acids and Tic represents 1,2, 3, 4 tetrahydroisoquinoline-3-carboxylic acid, the chemerin fragment or chemerin analog being linked to a lipid entity via a linker; wherein the formulation has a pH of about 6.5 to about 8.5, and an osmolality of about 200 to about 450 mOsm / Kg.

[0084] In some embodiments, the pharmaceutical compositions provided herein are formulated for ophthalmic topical administration and comprise: a) about 0.1% to about 0.5% w / v of NaCl; (b) about 25 mM to about 100 mM of phosphate buffered saline; and (c) about 0.1% w / v of a lipidated chemerin composition that includes a chemerin fragment consisting of the sequence of Y-F-P-G-Q-F-A-F-S (SEQ ID NO: 2) or a chemerin analog consisting of the sequence of Y*-F-L-P-S*-Q-F-A*-Tic-S (SEQ ID NO: 3), wherein * denotes D amino acids and Tic represents 1,2, 3, 4 tetrahydroisoquinoline-3-carboxylic acid, the chemerin fragment or chemerin analog being linked to a lipid entity via a linker; wherein the formulation has a pH of about 6.5 to about 8.5, and an osmolality of about 200 to about 450 mOsm / Kg.

[0085] In some embodiments, a formulation of the present disclosure comprises about 0.025% to about 0.05%; or about 0.05% to about 0.075%; or about 0.075% to about 0.1% w / v of a chemerin composition disclosed herein. In some embodiments, the formulation of the present disclosure comprises about 0.05% of a chemerin composition disclosed herein. In some embodiments, the formulation of the present disclosure comprises about 0.1% of a chemerin composition disclosed herein.

[0086] In some embodiments, a formulation of the present disclosure comprises about 0.01%, about 0.015%, about 0.02%, about 0.025%, about 0.03%, about 0.035%, about 0.04%, about 0.045%, about 0.5%, about 0.055%, about 0.06%, about 0.065%, about 0.07%, about0.075%, about 0.08%, about 0.085%, about 0.09%, about 0.095%, about 0.1%, about 0.125%, about 0.15%, about 0.175%, about 0.2%, about 0.225%, about 0.25%, or about 0.3% of a chemerin composition disclosed herein.

[0087] In some embodiments, a formulation of the present disclosure comprises 25 mM to 50 mM (e.g., 25 mM to 30 mM, 30 mM to 35 mM, 35 mM to 40mM, 40 mM to 45 mM or 45 mM to 50 mM,) phosphate buffered saline. In some embodiments, the formulation of the present disclosure comprises 50 mM phosphate buffered saline. In some embodiments, the formulation of the present disclosure comprises 100 mM phosphate buffered saline.

[0088] In some embodiments, a formulation of the present disclosure has a pH of 6.5 to 8.5 (e.g., 6.5 to 7.0, 7.0 to 7.5, 7.5 to 8.0 or 8.0 to 8.5). In some embodiments, the formulation of the present disclosure has a pH of about 7.2 to about 7.4. In some embodiments, the formulation of the present disclosure has a pH of about 7.4 to 7.6. In some embodiments, the formulation of the present disclosure has a pH of about 7.5.

[0089] In some embodiments, a formulation of the present disclosure comprises an osmolality of 200 to 450 mOsm / Kg (e.g., 200 to 250, 250 to 300, 300 to 350, 350 to 400 or 400 to 450 mOsm / kg). In some embodiments, the formulation has an osmolality of about 290 to about 320 mOsm / kg (e.g., 290 to 300, 300 to 310, 310 to 315, 315 to 320, 320 to 325, 325 to 330, 330 to 335, 335 to 340, 340 to 345 or 345 to 350 mOsm / Kg). In some embodiments, the formulation of the present disclosure comprises an osmolality of 290 to 325 mOsm / Kg (e.g., 290 to 295, 295 to 300, 300 to 305, 305 to 310, 310 to 315, 315 to 325 mOsm / Kg). In some embodiments, the formulation of the present disclosure comprises an osmolality of 313 to 322 mOsm / Kg (e.g., 313 to 315, 315 to 317, 317 to 319 or 319 to 322 mOsm / Kg). In some embodiments, the formulation of the present disclosure comprises an osmolality of 314 to 319 mOsm / Kg (e.g., 314 to 315, 315 to 316, 316 to 317 or 318 to 319 mOsm / Kg).Kits

[0090] In some embodiments, provided herein are kits for treatment of ocular neuropathic pain. The kits may comprise the soluble lipidated agent formulated in an ophthalmic formulation as single-use ampoules, for administration as ophthalmic drops. The kits may contain, for example and without limitation, 30 ampoules for a 1-week supply. The kits may be packaged in 4 cartons for a 4-week supply.Methods of Treatment

[0091] The compositions and methods described herein may be used to treat ocular pain and / or inflammation, e.g., ocular neuropathic pain, e.g., neuropathic corneal pain (NCP). One aspect of the present disclosure relates to methods of administering soluble lipidated ligand agents, e.g., formulated as an ophthalmic formulation, comprising human chemerin fragments or chemerin analogs for the treatment of ocular pain and / or ocular inflammation, e.g., neuropathic ocular pain.

[0092] Ocular inflammation may be caused by a microbial infection of the eye. Such infection may be fungal, viral, or bacterial. Ocular inflammation can also be caused by trauma, burn, autoimmune disease, chemical injury, contact lens, or other external stimuli. Methods of diagnosing ocular inflammation can be found in Teoh and Dick, “Diagnostic techniques for inflammatory eye disease: past, present and future: a review,” BMC Ophthalmology 2013, 13:41, the contents of which are incorporated herein by reference.

[0093] Ocular neuropathic pain may be associated with dry eye disease (DED), refractive surgery, herpes simplex keratitis, herpes zoster ophthalmicus, Sjogren’s syndrome, or chemical burns. Neuropathic ocular pain may also be systemic in origin, e.g., ocular neuropathic pain may be associated with small fiber polyneuropathy or fibromyalgia.

[0094] Ocular neuropathic pain can also be caused by inflammation. Neuropathic pain has typical symptoms like dysesthesias (spontaneous or evoked burning pain, often with a superimposed lancinating component), but the pain may also be deep and aching. Other sensations like hyperesthesia, hyperalgesia, allodynia (pain due to a normoxious stimulus), and hyperpathia (particularly unpleasant, exaggerated pain response) may also occur. In some embodiments, the ocular inflammation and / or ocular pain to be treated by the methods of the disclosure is associated with DED. DED is primarily caused by the break-down of the preocular tear film which results in dehydration of the exposed outer surface. People with DED may experience irritated, gritty, scratchy or burning eyes; a feeling of something in their eyes; excess watering; and blurred vision. When tear production decreases or tears evaporate too quickly from the eyes, symptoms of dry eye can develop. Tears are made up of three layers: oil, water and mucus. Each component protects and nourishes the front surface of the eye. A smooth oil layer helps prevent evaporation of the water layer, while the mucin layer spreads the tears evenly over the surface of the eye. If the tears evaporate too quickly or do not spread evenly over the cornea due to deficiencies with any of the three tear layers, dry eye symptoms can develop. A common form of dry eyes occurs when the water layer of tears is inadequate. This condition is also called keratoconjunctivitis sicca (KCS). The definition andclassification of DED can be found at “The Definition and Classification of Dry Eye Disease: Report of the Definition and Classification Subcommittee of the International Dry Eye Workshop (2007),” the Ocular Surface 2007, Vol. 5, 75-92, the contents of which are incorporated herein by reference.

[0095] DED can be diagnosed, for example and without limitation, through a comprehensive eye examination. Testing, with emphasis on the evaluation of the quantity and quality of tears produced by the eyes, may include: (a) patient history to determine the patient's symptoms and to note any general health problems, medications or environmental factors that may be contributing to the dry eye problem; (b) external examination of the eye, including lid structure and blink dynamics; (c) evaluation of the eyelids and cornea using bright light and magnification; and (d) measurement of the quantity and quality of tears for any abnormalities. Special dyes may be put in the eyes to better observe tear flow and to highlight any changes to the outer surface of the eye caused by insufficient tears.

[0096] In some embodiments, the compositions and methods of the disclosure reduce the symptoms of ocular inflammation and / or ocular pain. In some embodiments, the compositions and methods of the disclosure reduce burning and / or stinging eyes, e.g., associated with dry eye disease. In some embodiments, the compositions and methods of the disclosure reduce the level of fluorescein staining associated with dry eye disease. In some embodiments, the compositions and methods of the disclosure reduce the tear film break-up of the eye associated with dry eye disease.

[0097] Ocular inflammation may also be present as uveitis. Symptoms of uveitis commonly include redness, blurry vision, pain, light sensitivity, and floaters and flashes. In some embodiments, the compositions and methods of the disclosure treat uveitis.

[0098] In some embodiments, the compositions and methods of the disclosure may be used to treat inflammation resulting from an auto-immune disease, retinal inflammation, allergic conjunctivitis, or inflammation resulting from nerve injury or nerve degeneration. In some embodiments, the compositions and methods of the disclosure may be used to treat ocular pain, e.g., chronic ocular pain, or pain resulting from chemotherapy or radiation, pain resulting from nerve injury or nerve degeneration, or pain resulting from dysesthesia or allodynia. In some embodiments, the compositions and methods of the disclosure may be used to treat pain resulting from ocular keratitis, acute angle-closure glaucoma, or intraocular inflammation.

[0099] In some embodiments, the ocular neuropathic pain is not associated with dry eye disease (DED).

[0100] In some embodiments, the ocular neuropathic pain is not associated with ocular inflammation.

[0101] In some embodiments, the ocular neuropathic pain is not associated with ocular inflammation. In some embodiments, the ocular neuropathic pain is not associated with uveitis.

[0102] In some embodiments, a subject has visual analog scale (VAS) pain, and does not have neuropathic ocular pain.

[0103] In some embodiments, a subject has positive in vivo confocal microscopy (IVCM) findings, e.g., as evaluated by an ophthalmologist, and does not have neuropathic ocular pain. The subject may have evidence of one or more microneuromas. The subject may have decreased nerve density.

[0104] In some embodiments, the method of treating ocular inflammation and / or ocular pain, e.g., ocular neuropathic pain, e.g., neuropathic corneal pain (NCP), comprises administering to the subject a therapeutically effective amount of a soluble lipidated ligand agent comprising: a ligand entity comprising: (a) a human chemerin fragment having the sequence of YFPGQFAFS (SEQ ID NO: 2) or (b) a chemerin analog having the sequence of Y*FLPS*QFA*-Tic-S(SEQ ID NO: 3), wherein the * denotes D-amino acids, and wherein Tic represents (S)-l,2,3,4-tetrahydroisoquinoline-3-carboxylic acid; at least one lipid entity covalently linked to the ligand entity at or near the N-terminus, at or near the C-terminus, or at or near both the N-terminus and C-terminus; and an optional linker entity connecting the ligand entity to the at least one lipid entity; wherein the soluble lipidated ligand agent is formulated in a concentration of about 0.025% to about 0.1% w / v; and wherein the soluble lipidated agent is administered about 4 times daily. In some embodiments, the soluble lipidated agent is formulated as an ophthalmic formulation as provided herein.

[0105] In some embodiments, the method of treating ocular inflammation and / or ocular pain, e.g., ocular neuropathic pain, e.g., neuropathic corneal pain (NCP), comprises administering to the subject a therapeutically effective amount of a soluble lipidated ligand agent comprising: a ligand entity comprising: (a) a human chemerin fragment having the sequence of YFPGQFAFS (SEQ ID NO: 2) or (b) a chemerin analog having the sequence of Y*FLPS*QFA*-Tic-S(SEQ ID NO: 3), wherein the * denotes D-amino acids, and wherein Tic represents (S)-l,2,3,4-tetrahydroisoquinoline-3-carboxylic acid; at least one lipid entity covalently linked to the ligand entity at or near the N-terminus, at or near the C-terminus, or at or near both the N-terminus and C-terminus; and an optional linker entity connecting the ligand entity to the at least one lipid entity; wherein the soluble lipidated ligand agent isformulated in a concentration of about 0.05% or about 0.1% w / v; and wherein the soluble lipidated agent is administered about 4 times daily.

[0106] In some embodiments, the methods of treatment of the disclosure induce nerve regeneration in the subject. In some embodiments, the method of treatment reduces microneuromas in the cornea of the subject, increases the sensitivity of the cornea of the subject, and / or increases dendritiform cells in the cornea of the subject.

[0107] In some embodiments, the method comprises administering the soluble lipidated agent once a day, twice a day, three times, four times a day, five times a day, six times a day, seven times a day, or eight times a day to the subject. In some embodiments, the soluble lipidated agent is administered four times a day to the subject.

[0108] In some embodiments, the soluble lipidated agent is formulated as an ophthalmic formulation and is administered four times a day to the subject.

[0109] In some embodiments, the method comprises administering the soluble lipidated agents provided herein, e.g., formulated as ophthalmic formulations, as topical drop formulations. In some embodiments, a soluble lipidated agent formulated as an ophthalmic formulation is administered at the same time every day to the subject.

[0110] In some embodiments, one drop of an ophthalmic formulation is administered to each eye of the subject. In some embodiments, the drops are self-administered by the subject.

[0111] In some embodiments, a soluble lipidated agent provided herein formulated as an ophthalmic formulation is administered at four-hour intervals throughout the day, and up to four times a day to the subject.

[0112] In some embodiments, the method of treatment of the disclosure has a duration of at least 1 month, at least 2 months, at least 3 months, at least 4 months, at least 5 months, at least 6 months, or at least 1 year.

[0113] In some embodiments, the subject is a human.

[0114] In some embodiments, a pharmaceutical composition of the disclosure can be administered in combination with an anti-cytokine or anti-inflammatory agent for treating ocular neuropathic pain, e.g., dry eye disease (DED). Anti-cytokine or anti-inflammatory agents include, but are not limited to, NF Kappa B inhibitors, for example corticosteroids, glucocorticoids such as flucinolonone; nonsteroidal anti-inflammatory drugs (NSAIDs) such as sulindac and tepoxalin; antioxidants such as dithiocarbamate; and other compounds such as sulfasalazine [2-hydroxy-5-[-4-[C2-pyridinylamino)sulfonyl]azo]benzoic acid], clonidine, and autologous blood-derived products such as Orthokine.Methods of Measuring Efficacy of the Treatment

[0115] The present disclosure provides example methods of measuring the efficacy of the treatment of ocular neuropathic pain. Other methods of measurement known in the art may also be employed. In some embodiments, a physician or health care provider may assess the efficacy of a method described herein for the purpose of determining the continuation and / or duration of the treatment.

[0116] Visual Analog Questionnaire (VAS) Scores for Symptoms of Ocular Discomfort and Dryness: symptoms of ocular comfort and dryness are graded for each eye on a scale of 0-10, where 0=extremely uncomfortable, extremely dry and 10=excellent comfort, no dryness. In some embodiments, an average rating may be provided for morning, afternoon and evening. Three sets of copies of this questionnaire may be used, before and after an in- office pain test (hyperosmolar and / or proparacaine tests).

[0117] In some embodiments, improvement in pain (VAS) intensity relative to baseline for patients treated with OK-101 or a composition comprising the same is at least 10 units, e.g., at least 10 units, at least 15 units, at least 20 units, at least 25 units, or at least 30 units. In some embodiments, the patient has a starting VAS intensity of at least 30 units, or moderate pain. In some embodiments, the patient has a starting VAS intensity of at least 40 units. In some embodiments, the patient has a starting VAS intensity of at least 50 units.

[0118] Ocular Pain Assessment Survey (OP AS): a twenty-seven item questionnaire specifically designed to assess symptoms and quality of life in ocular pain.

[0119] In some embodiments, improvement in pain (OPAS) for patients treated with OK- 101 or a composition comprising the same is at least 1 point, at least 2 points, at least 3 points, at least 4 points, at least 5 points, at least 6 points, at least 7 points, at least 8 points, at least 9 points, or 10 points.

[0120] Ocular Surface Disease Index (OSDI): a twelve-item questionnaire to assess symptoms of ocular irritation consistent with dry eye and their impact on vision-related functioning.

[0121] In some embodiments, improvement OSDI for patients treated with OK-101 or a composition comprising the same is at least at least 10 units, at least 15 units, at least 20 units, at least 25 units, at least 30 units, at least 35 units, at least 40 units, at least 45 units, at least 50 units, at least 55 units, at least 60 units, at least 65 units, at least 70 units, at least 75 units, at least 80 units, at least 85 units, at least 90 units, or at least 95 units.

[0122] Best Corrective Distance Visual Acuity (VA): using the Snellen test, the right eye is documented in 20 / XXX format. Any subject with VA worse than 20 / 200 is documented as>20 / 200. The method of correction used for VA testing is documented. The same steps are followed for the left eye.

[0123] Slit lamp Examination (SLE): the eyelids, lashes, conjunctiva, cornea, lens, iris, and anterior chamber are assessed. The Slit lamp Examination (SLE) is performed before any study drug instillation or staining test and is performed on both eyes.

[0124] Tear Break-Up Time (TBUT): TBUT is performed using fluorescein strips. After application of one drop of saline on the strip, it touches the inferior tarsal conjunctiva. After 30 seconds and several blinks, the tear film is examined using a slit lamp with blue illumination. The time lapse between the last blink and the appearance of the first randomly distributed dark discontinuity in the fluorescein-stained tear film is measured three times and the mean value of the measurements is calculated.

[0125] In some embodiments, improvement in TBUT relative to baseline levels for patients treated with OK-101 or a composition comprising the same is at least 0.2, e.g., at least 0.2, at least 0.3, at least 0.35, or at least 0.4-fold.

[0126] Corneal Fluorescein Staining (CFS): fluorescein is instilled, and the entire cornea is examined using slit-lamp evaluation with blue illumination after 2 minutes. Staining is graded using the National Eye Institute (NEI) grading scheme, with a total range of 0-15.

[0127] In some embodiments, improvement in CLS relative to baseline levels for patients treated with OK-101 or a composition comprising the same is at least 1, at least 1.2, at least 1.4-fold.

[0128] Conjunctival Lissamine Green Staining (CLS): grading of conjunctival lissamine green staining is performed after 1 minute of application of a lissamine green drop. The nasal and temporal conjunctiva are examined using slit lamp evaluation with white light. Staining is graded using the NEI grading scheme, with a total range of 0-18.

[0129] In some embodiments, improvement in CLS relative to baseline levels for patients treated with OK-101 or a composition comprising the same is at least 1, at least 1.2, at least 1.4-fold.

[0130] Corneal Sensitivity: measured using a Cochet Bonnet aesthesiometer before the instillation of any dilating or anesthetic eye drops. Corneal sensitivity is assessed in the qualifying NEI zones of each eye. This instrument is constituted by a retractable 6 cm length filament of 0.12 mm in diameter, and it is generally considered the gold standard for corneal sensation assessment. The monofilament is gently pressed against the cornea, and is shortened in steps of 1.0 cm if a negative response is obtained. If a positive response isobtained, the monofilament is advanced by 0.5 cm, and the corneal sensitivity is measured by the longest filament length necessary to obtain a positive response.

[0131] In some embodiments, corneal sensitivity is decreased relative to baseline levels for patients treated with OK-101 or a composition comprising the same by at least 1 cm, at least 2 cm, at least 3 cm, at least 4 cm, or at least 5 cm.

[0132] Hyperosmolar Saline Test: a sodium chloride hypertonic drop (Muro 128) is instilled in each eye, and after 20 seconds any change in pain is graded according to the VAS score.

[0133] Proparacaine Challenge Test a single drop of proparacaine hydrochloride ophthalmic solution, (Alcaine®, 0.5%) is used on each eye, respectively, and any change in pain or other symptoms are noted after 90 seconds (to account for initial discomfort from the preservative in proparacaine solution). Any remaining ocular pain may be attributed to the central component. Pain is generally graded according to the VAS score.

[0134] In some embodiments, improvement in pain (VAS) intensity relative to baseline for patients treated with OK-101 or a composition comprising the same is at least 10 units, e.g., at least 10 units, at least 15 units, at least 20 units, at least 25 units, or at least 30 units.

[0135] Schirmer’s Test with Anesthesia (type II): performed 5 minutes after a drop of topical anesthesia with 0.5% proparacaine (Alcon Inc., Ft Worth TX) is applied to the eye. The Schirmer’s test is performed by placing a narrow filter-paper strip (5 x 35 mm strip of Whatman #41 filter paper) in the inferior temporal cul-de-sac. This test is conducted in a dimly lit room. The subject should gently close his or her eyes until five minutes have elapsed and the strips are removed. Since the tear front continues advancing a few millimeters after it has been removed from the eyes, it is important to mark the tear front at precisely five minutes. Aqueous tear production is measured by the length in millimeters of the wet part of the strip produced by the 5 -minute test.

[0136] In some embodiments, tear production is increased relative to baseline levels for patients treated with OK-101 or a composition comprising the same by at least 1 mm, at least 2 mm, at least 3 mm, at least 4 mm, or at least 5 mm.

[0137] In Vivo Confocal Microscopy (IVCM): aims to gain anatomical images of the sub- basal nerve plexus of the cornea. Generally, four sets of images are taken from the central cornea using a special microscope that gently contacts the cornea. An anesthetic drop is placed into the eye of the subject so that he or she does not feel any pain during the procedure. Hydroxypropyl methylcellulose 2.5% (Genteal Gel, Novartis Ophthalmics or similar) may be placed into the subject’s eyes during the procedure to act as a cushioningagent to minimize subject discomfort. The subject is asked to look at a fixation light during the procedure, and one or more trained technician(s) may capture the images.

[0138] The images of IVCM described herein may be analyzed. The three best representative images of each eye from the IVCM measurements are selected for nerve analysis at screening, week 4, week 8 and week 16. Two masked observers perform the IVCM images analysis as previously described (Muller et al., 2015). Briefly, corneal subbasal nerve plexus will be quantified using NeuronJ (www.imagescience.org / meijering / software / neuronj / ), a semiautomated tracing plug-in of ImageJ (www.imagescience.org / meijering / software). Nerves are categorized as main trunks (not branching from other nerves) and branches (nerves emerging from main trunks). The number of total nerves is defined as the sum of both main trunks and branches in one image and are reported as frame per nerve. The total nerve density is assessed as the total nerve length in micrometers per frame area (0.16 mm2), and then the value will be converted into mm / mm2. The nerve number is counted and reported as nerve per frame. Microneuromas are evaluated as described in Moein HR et al., Visualization of microneuromas by using in vivo confocal microscopy: An objective biomarker for the diagnosis of neuropathic corneal pain? Ocul Surf. 2020 Oct;18(4):651-656. Epub 2020 Jul 11. Nerve regeneration rate per month is calculated: as (nerve density at baseline - nerve density at follow up) / months of follow-up. Dendritiform cells density and size are assessed as density of cells / mm2.

[0139] In some embodiments, the subject to be treated has positive IVCM findings. Positive IVCM findings may be evaluated by an ophthalmologist or other healthcare professional. Positive IVCM findings include, for example and without limitation, decreased nerve density and evidence of microneuromas. In some embodiments, the subject has both decreased nerve density and evidence of microneuromas.

[0140] In some embodiments, the subject displays at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, or at least 90% decreased nerve density relative to a healthy subject.

[0141] In some embodiments, evidence of microneuromas is the presence of at least one microneuroma. In some embodiments, the microneuroma is at least 25 mm in diameter.Definitions

[0142] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which the invention pertains. Although other methods and materials similar, or equivalent, to thosedescribed 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.

[0143] The terms “peptide,” “polypeptide,” and “protein” are used interchangeably herein and typically refer to a molecule comprising 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 mimetic). Peptides may be naturally occurring, synthetically produced, or recombinantly expressed. Peptides may also comprise additional groups modifying the amino acid chain, for example, functional groups added via post- translational modification. Examples of post-translational modifications include, but are not limited to, acetylation, alkylation (including methylation), biotinylation, glutamyl ati on, glycylation, glycosylation, isoprenylation, lipoylation, phosphopantetheinylation, phosphorylation, selenation, and C-terminal amidation. The term “peptide” also includes peptides comprising modifications of the amino terminus and / or the carboxyl terminus. Modifications of the terminal amino group include, but are not limited to, des-amino, 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., wherein lower alkyl is C1-C4 alkyl). The term peptide also includes modifications, such as but not limited to those described above, of amino acids falling between the amino and carboxy termini. The term peptide can also include peptides modified to include one or more detectable labels.

[0144] The phrase “amino acid residue” as used herein refers to an amino acid that is incorporated into a peptide by an amide bond or an amide bond mimetic.

[0145] The terminal amino acid at one end of the peptide chain typically has a free amino group (z.e., the amino terminus or N terminus). The terminal amino acid at the other end of the chain typically has a free carboxyl group (z.e., the carboxy terminus or C terminus). Typically, the amino acids making up a peptide are numbered in order, starting at the amino terminus and increasing in the direction of the carboxy terminus of the peptide.

[0146] As used herein, the term “analog” refers to a variant or mutant polypeptide having one or more amino acid modifications compared to the wild type.

[0147] As used herein, an “amino acid modification” refers to a change in the amino acid sequence of a predetermined amino acid sequence. Exemplary modifications include an amino acid substitution, insertion and / or deletion. An “amino acid modification at” a specified position, e.g. of chemerin or a fragment thereof, refers to the substitution or deletionof the specified residue, or the insertion of at least one amino acid residue adjacent the specified residue. By insertion “adjacent” a specified residue is meant insertion within one to two residues thereof. The insertion may be N-terminal or C-terminal to the specified residue.

[0148] An “amino acid substitution” refers to the replacement of at least one existing amino acid residue in a predetermined amino acid sequence with another different “replacement” amino acid residue. The replacement residue or residues may be “naturally occurring amino acid residues” (i.e. encoded by the genetic code) and selected from the group consisting of: alanine (Ala); arginine (Arg); asparagine (Asn); aspartic acid (Asp); cysteine (Cys); glutamine (Gin); glutamic acid (Glu); glycine (Gly); histidine (His); isoleucine (lie): leucine (Leu); lysine (Lys); methionine (Met); phenylalanine (Phe); proline (Pro); serine (Ser); threonine (Thr); tryptophan (Trp); tyrosine (Tyr); and valine (Vai). Substitution with one or more non-naturally occurring amino acid residues is also encompassed by the definition of an amino acid substitution herein. A “non-naturally occurring amino acid residue” refers to a residue, other than those naturally occurring amino acid residues listed above, which is able to covalently bind adjacent amino acid residues(s) in a polypeptide chain. Examples of non-naturally occurring amino acid residues include norleucine, ornithine, norvaline, homoserine and other amino acid residue analogues such as those described in Ellman et al. Meth. Enzym. 202:301-336 (1991). To generate such non-naturally occurring amino acid residues, the procedures of Noren et al. Science 244: 182 (1989) and Ellman et al., supra, can be used. Briefly, these procedures involve chemically activating a suppressor tRNA with a non-naturally occurring amino acid residue followed by in vitro transcription and translation of the RNA. In some embodiments, an L-amino acid can also be substituted by a D-amino acid.

[0149] An “amino acid insertion” refers to the incorporation of at least one amino acid into a predetermined amino acid sequence. While the insertion will usually consist of the insertion of one or two amino acid residues, the present application contemplates larger “peptide insertions”, e.g. insertion of about three to about five or even up to about ten amino acid residues. The inserted residue(s) may be naturally occurring or non-naturally occurring as disclosed above.

[0150] An “amino acid deletion” refers to the removal of at least one amino acid residue from a predetermined amino acid sequence.

[0151] The term “pharmaceutical composition” refers to a mixture of a compound disclosed herein with other chemical components, such as diluents or carriers. The pharmaceutical composition can facilitate administration of the compound to an organism.Pharmaceutical compositions can also be obtained by reacting compounds with inorganic or organic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, salicylic acid and the like.

[0152] As used herein, the term “pharmaceutically acceptable carrier” refers to any of the standard pharmaceutical carriers, such as a phosphate buffered saline solution, water, emulsions (e.g., such as an oil / water or water / oil emulsions), and various types of wetting agents. The compositions also can include stabilizers, preservatives, and adjuvants.

[0153] As used herein, the terms “treat,” “treating,” “treatment,” and the like refer to reducing or ameliorating a disorder and / or a symptom associated therewith. It will be appreciated that, although not precluded, treating a disorder or condition does not require that the disorder or symptom associated therewith be completely eliminated. The terms “treat,” “treating,” or “treatment,” do not include prevention.

[0154] The term “therapeutically effective amount” refers to the amount of a compound that, when administered, is sufficient to prevent development of, or alleviate to some extent, one or more of the symptoms of the disorder, disease, or condition being treated. The term “therapeutically effective amount” also refers to the amount of a compound that is sufficient to elicit the biological or medical response of a cell, tissue, system, animal, or human that is being sought by a researcher, veterinarian, medical doctor, or clinician.

[0155] As used herein, a “subject” can be any mammal, e.g., a human, a non-human primate, mouse, rat, dog, cat, cow, horse, pig, sheep, goat, camel. In a preferred embodiment, the subject is a human.

[0156] As used herein, a “subject in need thereof’ is a subject having neuropathic ocular pain.

[0157] 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, and the like. Unless specifically stated or obvious from context, as used herein, the term “or” is understood to be inclusive and covers both “or” and “and.”

[0158] Throughout the specification the word “comprising,” or variations such as “comprises” or “comprise,” will be understood to imply the inclusion of a stated element,integer or step, or group of elements, integers or steps, but not the exclusion of any other element, integer or step, or group of elements, integers or steps.

[0159] Unless specifically stated or obvious from context, as used herein, the term “about” when used in conjunction with numerical values and / or ranges generally refers to those numerical values and / or ranges near to a recited numerical value and / or range. In some instances, the term “about” can mean within ± 10% of the recited value. For example, in some instances, “about 100 [units]” can mean within ± 10% of 100 (e.g., from 90 to 110).

[0160] The following Examples are meant to be demonstrative of the compositions and methods provided herein and are not meant to be limiting in any way.EXAMPLESExample 1: A Phase 2a, Randomized, Double-Masked, Placebo-Controlled Study Assessing the Safety and Efficacy of OK-101 Treatment in Subjects with Neuropathic Corneal Pain

[0161] OK-101 is a lipid-conjugated 12-mer chemerin peptide, Y*FLPS*QFA*-Tic-S (SEQ ID NO: 3), wherein the * denotes D-amino acids, and wherein Tic represents (S)- l,2,3,4-tetrahydroisoquinoline-3-carboxylic acid. The primary objective of this study in subjects with Neuropathic Corneal Pain is to evaluate the efficacy and safety of OK-101 in concentrations of 0.05% and 0.1%, as compared to placebo, instilled 4 times / day, as assessed by visual analogue scale (VAS).

[0162] OK-101 is designed to bind with high affinity to ChemR23 receptors. OK-101 was shown to exhibit potent ocular pain-reducing activity in a mouse model of corneal neuropathic pain. OK-101 was developed using a membrane-anchored-peptide (MAP) technology to produce a novel long-acting drug candidate. OK-101 is also designed to combat washout through the inclusion of a lipid ‘anchor’ within its molecular structure to enhance residence time of OK-101 within the ocular environment. OK-101 has also been shown to resolve the inflammation in animal models of asthma and modulate the inflammation environment in autoimmune diseases by recruiting regulatory T cells (Tregs), as well as attenuating neuropathic pain in mice.

[0163] Baseline signs and symptoms of subjects enrolled in the clinical trial are provided in Table A.Table A.Primary Objectives

[0164] The primary objectives of this study in subjects with neuropathic corneal pain were:

[0165] Objective 1 : Efficacy and safety of OK-101 in concentrations of 0.05% and 0.1%, as compared to placebo, instilled 4 times / day in subjects with neuropathic corneal pain, as assessed by visual analogue scale (VAS), were evaluated.Secondary Objectives

[0131] The secondary objectives of this study in subjects with non-active neuropathic corneal pain were:

[0132] Objective 1 : Assess Ocular Pain by OSDI and OPAS scores

[0133] Objective 2: Measure QoL improvement by OPAS.

[0134] Objective s: Assess drop comfortExploratory Objectives

[0135] Exploratory Objective 1 : Evaluated the efficacy of topical OK-101 on corneal sensation.

[0136] Exploratory Objective 2: Evaluated the efficacy of topical OK-101 on corneal nerve regeneration.

[0137] Exploratory Objective 3: Evaluated the efficacy of topical OK-101 on change in microneuromas.

[0138] Exploratory Objective 4: Evaluated the efficacy of topical OK-101 on change in dendritiform cells.Study Design

[0139] A randomized, double-masked, placebo-controlled trial to investigate the efficacy of OK-101 study drug in treating the symptoms of NCP at baseline (Visit 1) through the end of study (16 weeks) in NCP subjects was conducted.

[0140] All subjects provided written informed consent prior to any study procedures at the day of Screening Visit (Day 0).

[0141] Subjects were evaluated for clinical symptoms of NCP at screening visit (Day -8), Visit 1 (Day 0), Visit 2 (Day 28±2, Week 4), Visit 3 (Day 56±2, Week 8) and Visit 4 (Day 112±7, Week 16).Dosage Regimen

[0142] QID refers to 4x daily administration. Approximately 48 subjects were randomized in a 1 : 1 : 1 ratio to 0.05% QID (Arm B), 0.1% QID (Arm B), and placebo QID (Arm C), respectively (i.e., approximately 32 subjects received the total active treatment group, and 16 subjects will receive the placebo).

[0143] Recruited subjects started the randomized study drug in the study eye, 4 times daily at 4-hour intervals (up to 16 hours per day), for four weeks. In case of bilateral NCP, the drug was instilled in both eyes.

[0144] OK-101, in a concentration of 0.05%: one drop of study drug (0.05%) were instilled in the affected eye(s), four times daily; and

[0145] OK-101, in a concentration of 0.1%: one drop of study drug (0.1%) were instilled in the affected eye(s), four times daily; and

[0146] Placebo: one drop of placebo were instilled in affected eye(s), four times daily.

[0147] During the treatment period (Day 0 to Week 12), only the study drug was permitted as a topical treatment. At any time during the 12 weeks of treatment, subjects were seen for an unscheduled visit to evaluate for safety in case of reported ocular side effects.Dosage Form

[0148] One drop of OK-101 or placebo was instilled in each affected eye(s) four times daily (every 4 hours) for 12 weeks, unless the dosing regimen was changed during the followup visits. The subject used the ophthalmic drops at the same time every day.Efficacy Analysis ConsiderationsVisual Analog Questionnaire (VAS) Scores for Symptoms of Ocular Discomfort and Dryness

[0149] The Wilcoxon Rank Sum test was used to analyze the difference in the mean Visual Analog Questionnaire (VAS) scores for symptoms of ocular discomfort and dryness between the different treatment arms at each post-baseline visit on the intention to treat (ITT) population. The primary endpoint was the difference at visit 5 (week 16) for the different treatment arms. Additionally, for each post-baseline visit, the following statistics were reported for each treatment arm: mean, difference in means (Arms A & B vs. Arm C), standard error (SE), minimum, maximum, median, and two-sided Wilcoxon Rank Sum p- values (Arms A & B vs. Arm C).

[0150] The change-from-baseline (CFB) in VAS for symptoms of ocular discomfort and dryness was analyzed in a similar fashion.Ocular Pain Assessment Survey (OP AS)

[0151] The Ocular Pain Assessment Survey (OPAS) data were analyzed and summarized similarly to the VAS data as described above.Ocular Surface Disease Index (OSDI)

[0152] The Ocular Surface Disease Index (OSDI) twelve-item questionnaire assesses symptoms of ocular irritation consistent with dry eye disease and its impact on vision-related functioning.Ocular Pain Assessment Survey (OP AS)

[0153] The Ocular Pain Assessment Survey (OPAS) is a twenty-seven item questionnaire specifically designed to assess symptoms and quality of life in ocular pain.Best Corrective Distance Visual Acuity ( A)

[0154] The best-corrected distance visual acuity (VA) was recorded for each participant using the Snellen test. The right eye was documented in 20 / XXX format. Any subject with VA worse than 20 / 200 was documented as >20 / 200. The method of correction used for VA testing was documented. The same steps were followed for the left eye.Visual Analogue Questionnaire (VAS)

[0155] The Visual Analogue Questionnaire (VAS) measures symptoms of ocular comfort and dryness, graded for each eye on a scale of 0-10, where 0=extremely uncomfortable, extremely dry and 10=excellent comfort with no dryness. An average rating was provided for the morning, afternoon and evening. Three sets of copies of this questionnaire were used, before and after each in-office pain test (hyperosmolar and proparacaine tests).Slit lamp Examination (SLE)

[0156] The eyelids, lashes, conjunctiva, cornea, lens, iris, and anterior chamber were assessed. The Slit lamp Examination (SLE) is performed before any study drug instillation or staining test and is performed on both eyes.Tear Break-Up Time (TBUT)

[0157] Tear Break-Up Time (TBUT) was performed using fluorescein strips. After application of one drop of saline on the strip, it touched the inferior tarsal conjunctiva. After 30 seconds and several blinks, the tear film was examined using a slit lamp with blue illumination. The time lapse between the last blink and the appearance of the first randomly distributed dark discontinuity in the fluorescein-stained tear film was measured three times and the mean value of the measurements is calculated.Corneal Fluorescein Staining (CFS)

[0158] For Corneal Fluorescein Staining (CFS), fluorescein was instilled, and the entire cornea was examined using slit-lamp evaluation with blue illumination after 2 minutes. Staining was graded using the national eye institute (NEI) grading scheme, with a total range of 0-15.Conjunctival Lissamine Green Staining (CLS)

[0159] For Conjunctival Lissamine Green Staining (CLS), grading of conjunctival lissamine green staining was performed after 1 minute of application of a lissamine greendrop. The nasal and temporal conjunctiva were examined using slit lamp evaluation with white light. Staining was graded using the NEI grading scheme, with a total range of 0-18. Corneal Sensitivity

[0160] Corneal Sensitivity was measured using a Cochet Bonnet aesthesiometer before the instillation of any dilating or anesthetic eye drops. Corneal sensitivity was assessed in the qualifying NEI zones of each eye. This instrument was constituted by a retractable 6 cm length filament of 0.12 mm in diameter, and it was considered the gold standard for corneal sensation assessment. The monofilament was gently pressed against the cornea, and was shortened in steps of 1.0 cm if a negative response was obtained. If a positive response was obtained, the monofilament was advanced by 0.5 cm, and the corneal sensitivity was measured by the longest filament length necessary to obtain a positive response.Hyperosmolar Saline Test

[0161] For the Hyperosmolar Saline Test, a sodium chloride hypertonic drop (Muro 128) was instilled in each eye, and after 20 seconds any change in pain was graded according to the VAS score.Proparacaine Challenge Test

[0162] For the Proparacaine Challenge Test, a single drop of proparacaine hydrochloride ophthalmic solution, (Alcaine®, 0.5%) was used on each eye respectively and any changes in pain or other symptoms were noted after 90 seconds (to account for initial discomfort from the preservative in proparacaine solution). Any remaining ocular pain was generally attributed to the central component. Pain was graded according to the VAS score.Schirmer ’s Test with Anesthesia (type II)

[0163] The Schirmer’s Test with Anesthesia (type II) was performed 5 minutes after a drop of topical anesthesia with 0.5% proparacaine (Alcon Inc., Ft Worth TX) was applied to the eye. The Schirmer’s test was performed by placing a narrow filter-paper strip (5 x 35 mm strip of Whatman #41 filter paper) in the inferior temporal cul-de-sac. This test was conducted in a dimly lit room. The subject gently closed their eyes until five minutes have elapsed and the strips were removed. Since the tear front continues advancing a few millimeters after it has been removed from the eyes, it was important to mark the tear front at precisely five minutes. Aqueous tear production was measured by the length in millimeters of the wet part of the strip produced by the 5 -minute test.In Vivo Confocal Microscopy (IVCM)

[0164] The / / / Vivo Confocal Microscopy (IVCM) test aims to gain anatomical images of the sub-basal nerve plexus of the cornea. Four sets of images were taken from the centralcornea using a special microscope that gently contact the cornea. An anesthetic drop was placed into the eye of the subject so that he or she did not feel any pain during the procedure. Hydroxypropyl methylcellulose 2.5% (Genteal Gel, Novartis Ophthalmics or similar) was placed into the subject’s eyes during the procedure to act as a cushioning agent to minimize subject discomfort. The subject was asked to look at a fixation light during the procedure, and one or more trained technician(s) captured the images. The images were analyzed by the investigators.IVCM Analysis

[0165] Three best representative images of each eye from the IVCM measurements were selected for nerve analysis at screening, week 4, week 8 and week 16. Two masked observers performed the IVCM images analysis as previously described (Muller et al., 2015). Briefly, corneal subbasal nerve plexus was quantified using NeuronJ (http: / / www.imagescience.org / meijering / software / neuronj / ), a semiautomated tracing plug-in of ImageJ (http: / / www.imagescience.org / meijering / software). Nerves were categorized as main trunks (not branching from other nerves) and branches (nerves emerging from main trunks). The number of total nerves was defined as the sum of both main trunks and branches in one image and reported as frame per nerve. The total nerve density was assessed as the total nerve length in micrometers per frame area (0.16 mm2), and then the value converted into mm / mm2. Nerve number was counted, and reported as nerve per frame. Microneuromas were evaluated as described in Moein HR et al., Visualization of microneuromas by using in vivo confocal microscopy: An objective biomarker for the diagnosis of neuropathic corneal pain? Ocul Surf. 2020 Oct;18(4):651-656. Epub 2020 Jul 11. Nerve regeneration rate per month was calculated as nerve density at baseline - nerve density at follow up / months of follow-up. Dendritiform cells density and size were assessed as density of cells / mm2as previously described.Randomization and Visit Schedule

[0166] The first study treatment was administered on the same day as randomization (i.e., at the Baseline / Visit 1 / Day 0 visit). Subjects were instructed to contact the study site at any time if they have health-related concerns. All study visits were scheduled 28 (+ / - 7) days relative to the randomization date. Each active arm (Arm A and Arm B) was individually compared against the placebo arm (Arm C).Inclusion Criteria:

[0167] Individuals who met the following criteria were eligible for inclusion in the study:

[0168] Male or female aged > 18 years.

[0169] Symptoms of neuropathic corneal pain for at least 3 months, such as burning, stinging, light sensitivity, discomfort, or pain.

[0170] Positive IVCM findings evaluated by an experienced ophthalmologist: decreased nerve density and evidence of microneuromas.

[0171] Females of childbearing potential must have a negative pregnancy test.

[0172] Best corrected distance visual acuity (BCDVA), using corrective lenses, if necessary, in the study eye of at least +1.0 Log MAR (Snellen <20 / 200).

[0173] Satisfying all Informed Consent requirements.

[0174] Ability and willingness to comply with study procedures.

[0175] Ability to speak and understand the English language sufficient to understand the study, provide written consent, and allow completion of all study assessments.Exclusion Criteria:

[0176] Subjects who met any of the following criteria at Screening were excluded from the study:

[0177] Evidence of any active ocular infection.

[0178] Evidence of any intraocular inflammation.

[0179] Evidence of any persistent epithelial defect / ulcer.

[0180] Evidence of any corneal scar / comeal edema.

[0181] Presence of any other ocular conditions that require topical medications during the treatment phase.

[0182] History of severe systemic allergy or severe ocular allergy.

[0183] Inability to suspend topical medications 8 days prior to the starting date.

[0184] Inability to continue oral medications for NCP without changes during the study duration.

[0185] No changes or less than 50% improvement in VAS score after topical 0.5% proparacaine hydrochloride (Alcaine, Alcon, Fort Worth, TX).

[0186] History of any ocular surgery within three months before study Visit l(day 0).

[0187] Ocular surgery expected during the 16 weeks of the trial.

[0188] Use of refractive / therapeutic contact lenses during the study period.

[0189] Female subjects who are pregnant / have a positive pregnancy test result or are breastfeeding or intend to become pregnant during the study treatment period.

[0190] Drug addiction / alcohol abuse within the last year.

[0191] Participation in another clinical trial concurrently.Study Design

[0192] The study was a phase 2a, randomized, double-masked, placebo-controlled trial assessing the safety and efficacy of OK-101 treatment in subjects with neuropathic corneal pain.

[0193] Subjects were evaluated for clinical symptoms of NCP at Screening Visit (Day -8 - Day 0), Visit 1 (Day 0), Visit 2 (Day 28±2, Week 4), Visit 3 (Day 56±2, Week 8). Visit 4 (Day 84±2, Week 12), and Visit 5 (Day 112±7, Week 16).

[0194] This lasted a total of 16 weeks (approximately 4 months) after completion of the screening period, which lasted for up to 8 days (i.e., the screening visit can be done anytime from Day -8 to Day 0). During this time, the subject attended 5 visits in total. Subjects were seen for an unscheduled visit to evaluate for safety in case of reported ocular side effects.

[0195] From Visit 1 (Day 0) to Visit 4 (Week 12), the subject was asked to self-administer the study drug 4 times per day (one drop in both eyes every 4 hours, for example at 9:00 am, 1 :00 pm, 5:00 pm and 9:00 pm). The subject returned to clinic for Visit 2 (Week 4) and Visit 3 (Week 8), where they were evaluated for resolution or change in NCP symptoms and continue the dosing.

[0196] At Visit 4 (Week 12) the subject stopped the study drug but continued in the study for an additional 4 weeks to monitor any potential eye drop effects on the subject’s health.

[0197] Study design is summarized in FIG. 11.Control Product, Dose, and Mode of Administration

[0198] The medication or placebo was provided in single-use ampoules. Together with the monthly box, subjects were provided with enough ampoules for the administration.

[0199] Each carton contained: 30 ampoules for a 1-week supply. The Delivery System Kit contained the following: 4 cartons for a 4-week supply.Results

[0200] 384 subjects were enrolled in the study. 144 subjects failed screening, and 240 subjects were randomized to treatment or placebo arms.

[0201] Subjects treated with OK-101 displayed significantly improved signs of pain, as demonstrated by Conjunctival Sum Staining (FIGs 1, 2A-2B, 3, 4, 9, 12A-12B), tear film break up time (FIGs.5 and 13), and significantly improved symptoms of pain, including burning / stinging (FIGs.7A-7B, 10, and 17B), blurred vision (FIG. 15B), itching (FIG. 17C), eye dryness (FIG. 17D), and pain on the VAS intensity scale (FIGs. 6 and 14). Self-reported improvements in symptoms are reported in FIG. 16. Percentages of responders with these improvements are summarized in Tables B, C, and D.Table B. Percentage of Responders with Improvement in Signs at Day 85, Pre-CAETable C. Percentage of Responders with Improvement in Symptoms at Day 85, Intent-to-Treat Population, Pre-CAETable D. Responders with Improvement in Both Signs and Symptoms at Day 85, ITT Population

[0202] Treatment emergent adverse events (TEAEs) were observed to be similar to the placebo-treated group. No severe drug related ocular TEAEs were seen. Possible drug-related TEAEs were observed in one patient in the OK-101 0.05% treatment group and 3 patients in the placebo-treated group, again, highlighting the favorable safety profile of OK-101 (FIG. 18).Conclusions

[0203] Significant drug effects were observed in multiple signs (conjunctival staining, TFBUT) and symptoms (ocular pain, burning / stinging and blurred vision) of DED as early as the 15-days after dosing. Drug effect was durable throughout the trial for a number of endpoints. Significant improvements were observed across multiple symptoms as measured in a daily symptom diary including pain, eye dryness and itching within the first two weeks of treatment. OK-101 exhibited excellent drop comfort, comparable to that of artificial tears, with a favorable adverse event profile and no drug-related serious adverse events. These observed endpoints support a proposed mechanism-of-action of restoring goblet cell loss by OK-101 as demonstrated in preclinical animal models.

[0204] Thus, OK-101 reduces inflammatory markers, reduces pain, burning / stinging, blurred vision, and eye dryness, and normalizes goblet cell loss.

Claims

CLAIMSWhat is claimed is:

1. A method of treating neuropathic corneal pain in a subject in need thereof, the method comprising: administering to the subject a therapeutically effective amount of a soluble lipidated ligand agent comprising: a ligand entity comprising:(a) a human chemerin fragment having the sequence of YFPGQFAFS (SEQID NO: 2) or(b) a chemerin analog having the sequence of Y*FLPS*QFA*-Tic-S (SEQ ID NO: 3), wherein the * denotes D-amino acids, and wherein Tic represents (S)-l, 2,3,4- tetrahydroisoquinoline-3-carboxylic acid; at least one lipid entity covalently linked to the ligand entity at or near the N-terminus, at or near the C-terminus, or at or near both the N-terminus and C-terminus; and an optional linker entity connecting the ligand entity to the at least one lipid entity; wherein the soluble lipidated ligand agent is formulated in a concentration of about 0.05% or about 0.1% w / v; and wherein the soluble lipidated agent is topically administered about 4 times daily to the eye of the subject.

2. The method of claim 1, wherein the subject in need thereof has a Visual Analog Scale (VAS) pain intensity of at least 30 units, and wherein the method decreases VAS pain intensity by at least 10 units.

3. The method of claim 2, wherein the method decreases VAS pain intensity by at least 20 units.

4. The method of claim 2 or 3, wherein the method decreases VAS pain intensity by at least 30 units.

5. The method of claim 1, wherein the subject in need thereof has positive In Vivo Confocal Microscopy (IVCM) findings, wherein positive IVCM findings are decreased nerve density and evidence of microneuromas.

6. The method of any one of claims 1-5, wherein the subject does not have dry eye disease or is not diagnosed as having dry eye disease.

7. The method of any one of claims 1-6, wherein the soluble lipidated agent is administered in a formulation comprising about 0.05% w / v.

8. The method of any one of claims 1-6, wherein the soluble lipidated agent is administered in a formulation comprising about 0.1% w / v.

9. The method of any one of claims 1-8, wherein the lipid entity is linked to the chemerin fragment or chemerin analog through a linker.

10. The method of claim 9, wherein the linker comprises polyethylene glycol, a peptide, or a combination thereof.

11. The method of claim 10, wherein the linker is selected from the group consisting of:

12. The method of any one of claims 1-11, wherein the chemerin fragment or chemerin analog comprises at least 10 amino acids, at least 15 amino acids, at least 20 amino acids, at least 30 amino acids, at least 40 amino acids, at least 50 amino acids, at least 60 amino acids, at least 70 amino acids, at least 80 amino acids, at least 90 amino acids, at least 100 aminoacids, at least 110 amino acids, at least 120 amino acids, at least 130 amino acids, at least 140 amino acids, or at least 150 amino acids.

13. The method of any one of claims 1-12, wherein the chemerin fragment consists of the sequence YFPGQFAFS (SEQ ID NO: 2).

14. The method of any one of claims 1-13, wherein the chemerin analog consists of the sequence Y*FLPS*QFA*-Tic-S (SEQ ID NO: 3), wherein the * denotes D-amino acids, and wherein Tic represents (S)-l,2,3,4-tetrahydroisoquinoline-3-carboxylic acid.

15. The method of any one of claims 1-14, wherein the chemerin fragment or chemerin analog is resistant to proteolysis.

16. The method of any one of claims 1-15, wherein the lipid entity is selected from the group consisting of a-linolenic acid, y-linolenic acid, stearidonic acid, eicosapentaenoic acid, docosahexaenoic acid, linoleic acid, dihomo-y-linolenic acid, arachidonic acid, docosatetraenoic acid, palmitoleic acid, vaccenic acid, paullinic acid, oleic acid, elaidic acid, gondoic acid, erucic acid, nervonic acid, mead acid, myristic acid, palmitic acid, stearic acid, l,2-dipalmitoyl-sn-glycero-3 -phosphoethanolamine (DPPE), GM1 ganglioside, GM2 ganglioside, GM3 ganglioside, l,2-dipalmitoyl-sn-glycero-3 -phosphocholine (DPPC), 1,2- dioleoyl-sn-glycero-3-phospho-L-serine (DOPS), l,2-dioleoyl-sn-glycero-3 -phosphocholine (DOPC), a glycosphingolipid, a sphingolipid, phosphatidylinositol 4,5-bisphosphate (PIP2), a ceramide, cholesterol, ergosterol, phytosterol, a hopanoid, and a steroid.

17. The method of claim 16, wherein the lipid entity is selected from the group consisting of a-linolenic acid, y-linolenic acid, palmitic acid, vaccenic acid, oleic acid, and elaidic acid.

18. The method of any one of claims 1-17, wherein the lipid entity is linked at or near the N-terminus of the chemerin fragment or chemerin analog.

19. The method of any one of claims 1-18, wherein the lipid entity is linked at or near the C-terminus of the chemerin fragment or chemerin analog.

20. The method of any one of claims 1-19, wherein the chemerin fragment or chemerin analog has the structure21. The method of any one of claims 1-20, wherein the subject is a human.

22. The method of any one of claims 1-21, wherein the method induces nerve regeneration in the subject, optionally wherein the method reduces microneuromas in thecornea of the subject, increases the sensitivity of the cornea of the subject, and / or increases dendritiform cells in the cornea of the subject.

23. The method of any one of claims 1-22, wherein the soluble lipidated agent is formulated as an ophthalmic formulation comprising: a) about 0.1% to about 0.5% w / v of NaCl; (b) about 25 mM to about 100 mM of phosphate buffered saline; and (c) about 0.05% or about 0.1% w / v of the soluble lipidated agent; wherein the formulation has a pH of about 6.5 to about 8.5, and an osmolality of about 200 to about 450 mOsm / Kg.

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