Method for reducing corneal fluorescein staining with TRPM8 agonists

Abstract

The present invention provides a method for effectively reducing corneal fluorescein staining (CFS) or improving corneal health, which includes administering a therapeutically effective amount of a TRPM8 agonist to a subject in need thereof, wherein the TRPM8 agonist comprises a DIPA compound (e.g., DIPA-9). The novel therapeutic use of the TRPM8 formulations according to the present invention may demonstrate improved efficacy through reduced CFS or SANDE.

Description

[0001] METHOD FOR REDUCING CORNEAL FLUORESCEIN STAINING WITH TRPM8

[0002] AGONISTS

[0003] CROSS-REFERENCE TO REL ATED APPLICATIONS

[0004] [1] This application claims priority to U. S. Application No. 63 / 744,367, filed on January 13, 2025, and U. S. Application No. 63 / 747,272, filed on January 20, 2025, the contents of which are incorporated herein by reference in their entirety.

[0005] BACKGROUND OF THE INVENTION

[0006] [2] Corneal fluorescein staining (CFS) highlights damaged eye surface cells, indicating various diseases like contact lens issues, infections, dry eye disease (DED), and systemic conditions such as Sjogren's syndrome, diabetes, and neurotrophic keratopathy, by revealing epithelial defects, inflammation, or erosion. It is a key diagnostic tool for assessing severity in DED, revealing patterns like superficial punctate keratopathy (SPK) or patchy staining that show compromised tear film or cellular adhesion, affecting vision.

[0007] [3] Common conditions associated with CFS include DED, contact lens wear, trauma / erosions, neurotrophic keratopathy. In patients with DED, staining (SPK) often appears in the lower or interpalpebral (between eyelids) areas, reflecting tear deficiency or instability. Contact lens wear can cause focal staining from mechanical stress or poor fit. Viral (herpes), bacterial, or fungal infections can cause characteristic staining patterns. In patients with trauma / erosions, injury or conditions like Epithelial Basement Membrane Dystrophy (EBMD) cause cells to detach, allowing dye to pool. As to neurotrophic keratopathy, nerve damage reduces sensation, leading to poor healing and increased permeability.

[0008] [4] Systemic diseases with corneal staining incudes Sjogren's syndrome (an autoimmune disorder causing severe dry eye, often with significant corneal staining), diabetes (which can lead to corneal damage, affecting epithelial health), autoimmune conditions (such as lupus, rheumatoid arthritis, sarcoidosis), and neurological disorders (such as multiple sclerosis which affects comeal nerves).

[0009] [5] CFS reliably reveals dye uptake (i.e., dye entering damaged or missing epithelial cells), superficial punctate keratopathy (i.e., tiny dots of staining from desquamated cells), and patchy staining (with wider areas of damage, often seen in DED).[6] TRPM8 is part of a family of transient receptor potential (TRP) cation channels, which are associated with the perception of sensations such as temperature change. See D D. McKemy et al., Nature, 2002, 416, 52-58. TRPM8 is a cold thermoreceptor, while others of the TRP family found on ocular surfaces such as TRP VI handle other sensations such as heat, irritation, and pain. See J. M. Yang et al., Pharmaceuticals (Basel, Switzerland), 2018, 77(4), 125; and J. M Yang et al., BMC Ophthalmol., 2017, 77(1), 101. Accordingly, selective activation of TRPM8 without concurrent activation of nociceptive channels such as TRPV1 has been identified as a desirable characteristic for pharmaceutical candidates intended to treat DED.

[0010] [7] Among known TRPM8 agonists, several compounds have been found to cause undesirable side effects or formulation limitations, and thus are unsuitable for development as eyedrop administration. For example, menthol may cause ocular irritation or stinging, and icilin lacks adequate aqueous solubility for incorporation into ophthalmic delivery vehicles. TRPM8 antagonism has also been considered for DED treatment, because evaporative cooling and hyperosmotic stimuli may trigger dry eye pain. See J M Yang et al., BMC Ophthalmol., 2017, 77(1), 101. However, this pathway may reduce tear secretion which would also be undesirable.

[0011] [8] Like most therapeutic agents targeting tear secretion pathways, TRPM8 agonists have primarily demonstrated efficacy in improving tear production, as measured by Schirmer’s Test, with minimal or inconsistent effects on comeal staining. Accordingly, improvements in corneal health and reductions in comeal staining have generally not been expected outcomes for agents acting primarily through tear secretion or sensory pathways. Unexpectedly, clinical evaluation of formulations containing DIPA-9 (i.e., l-di(isopropyl)phosphinoyl-nonane), a novel TRPM8 agonist, demonstrated not only improvements in tear production and symptom relief, but also significant reductions in corneal fluorescein staining and improvements in corneal health. These effects were surprising in view of the known mechanism of action of TRPM8 agonists and prior clinical experience with related therapies, which had not shown meaningful improvements in corneal staining.

[0012] [9] The present invention offers novel and surprisingly effective methods for reducing corneal fluorescein staining, in association with dry eye disease, and methods for improving corneal health with a TRPM8 agonist such as DIPA-9.

[0013] BRIEF SUMMARY OF THE INVENTION

[0010] The present invention provides a therapeutic use of formulations containing a TRPM8 agonist for effectively and rapidly reducing corneal fluorescein staining, which demonstrates improved efficacy in a variety of clinical endpoints, with significant and rapid improvement in symptom assessment in dry eye questionnaire (SANDE) and corneal fluorescein staining (CFS) scores. Given its mechanism of action (MOA), the invention was unexpectedly effective in reducing corneal fluorescein staining, improving clinical symptoms or signs of conditions associated with corneal fluorescein staining including, e.g., DED, contact lens wear, trauma / erosions, and neurotrophic keratopathy. As an example, patients with DED show multiple signs and symptoms outside of tear production, unlike other products with similar MOA which primarily improved tear production only. The invention is particularly effective at about 0.2% concentration of the TR. PM8 agonist, which consistently demonstrated superior efficacy across all measured endpoints in clinical testing

[0014]

[0011] Notably, the method of the present invention has achieved not only significant but also rapid reduction of corneal fluorescence staining (CFS), as well as corneal health. This was totally unexpected based on prior studies, especially the rapidness and magnitude of the reduction of CFS (e.g., reduction of total CFS by at least 0.7 points relative to baseline), even starting from week 1.

[0015]

[0012] More specifically, the present invention offers methods for reducing corneal fluorescein staining, thereby treating CFS-associated conditions (e.g., DED, contact lens wear, trauma / erosions, and neurotrophic keratopathy), especially by, among other things,, comprising administering an effective amount of a TRPM8 agonist - e.g., a 1 -diisopropyl phosphi noy 1 alkane (DIP A) compound formulation - to a subject in need thereof in order to effectively reduce corneal fluorescein staining (e.g., by at least 0.7 points relative to baseline), even starting from week 1. The present invention also offers methods for improving corneal health, by administration of a TRPM8 agonist (e.g., in the form of a DIP A compound formulation) to a subject in need of such improvement.

[0016]

[0013] Examples of the DIP A compounds include but are not limited to the following: /

[0017] Formula /

[0018] Code Chemical Name ole ula / \

[0019] M c r Chemical S tincture Weight

[0020] 1 -di (i sopropy l)-ph osphi noyl - C12H27OP

[0021] DIPA-6.2 v

[0022] hexane 218.32 V O- / / z

[0023] /

[0024] l-di(isopropyl)-phosphinoyl- C13H29OP

[0025] DIPA-7

[0026] heptane 232.34

[0027] l-di(isopropyl)phosphinoyl- C14H31OP

[0028] DIPA-8

[0029] octane 246.37

[0030] 1 -di (i sopropyl)phosphinoyl - C15H33OP

[0031] DIPA-9 Ap “ / ,

[0032] nonane 260.40 ( “0 —

[0033] / \ ' <

[0034]

[0035] /

[0036]

[0014] An analog, derivative, and / or pharmaceutically acceptable salt of any of the above¬ identified exemplary DIPA compounds may also be used in the methods of this in <ven \tion.

[0037]

[0015] The method of this invention may be earned out by administering a TRPM8 ag \onist by > using a wiping, brushing, or spraying method, or finger application to apply a therapeutic7ally effective amount of a TRPM8 agonist (e.g., DIPA-9) to the eyelid of the subject, or a traditional eye drop method to apply a therapeutically effective amount of a TRPM8 agonist (e.g., DIP / X-9) to the cornea.

[0038]

[0016] According to the present invention, the wipe and / or a pad material may include but are not limited to cotton, rayon (known as viscose), lyocelL cellulose, chemically modified cellulose, polyester, polypropylene, polyethylene terephthalate (PET), nylon, velour / velvet, microfiber (blend of PET and Nylon), plant-based fibers like wood / bamboo pulp, and / or any blend of these materials.

[0039]

[0017] The formulation according to the present invention can be in the form of liquid, solid, semisolid, gel, cream, or ointment. For instance, the formulation may be an aqueous formulation. The formulation may include a pharmaceutically acceptable amount of surfactants / solubilizers, and / orgeiling agents, and / or gel adjusting agents, and / or pH adjusting agents, and / or osmolarity adjusting agents, and / or preservatives.

[0040]

[0018] One aspect of the present invention provides a method for treating dry eye disease comprising administering an effective amount of a TRPM8 agonist to a subject in need thereof to effectively reduce corneal fluorescein staining, wherein the TRPM8 agonist comprises a DIP A compound.

[0041]

[0019] Another aspect of the present invention provides a method for reducing corneal fluorescein staining in association with dry eye disease, comprising a step of administering an effective amount of a TRPM8 agonist to a subject in need thereof, wherein the TRPM8 agonist comprises a DIPA compound. Examples of the DIPA compound may include but are not limited to DIPA-6, DIPA-7, DIPA-8, or DIPA-9, or an analog, derivative, and / or pharmaceutically acceptable salt thereof

[0020] In some embodiments, the TRPM8 agonist includes DIPA-9.

[0042]

[0021] In some embodiments, the TRMP8 agonist is administered in the form of a pharmaceutical formulation comprising the TRPM8 agonist and a pharmaceutically acceptable carrier.

[0043]

[0022] In some embodiments, the pharmaceutical formulation is in the form of liquid, solid, semi¬ solid, gel, cream, or ointment. For instance, the formulation may be an aqueous formulation.

[0044]

[0023] In some embodiments, the TRPM8 agonist is contained in the pharmaceutical formulation at a concentration ranging from about 0.01% (w / w) to about 0.4% (w / w). In some embodiments, the TRPM8 agonist is contained in the pharmaceutical formulation at a concentration ranging from about 0.2% (w / w) to about 0.4% (w / w).

[0045]

[0024] In some embodiments, the TRPM8 agonist comprise DIPA-9 at a concentration of about 0.2% (w / w).

[0046]

[0025] In some embodiments, the TRPM8 agonist is administered topically using a wiping, brushing, or spraying method, or finger application, or a traditional eye drop method, to ocular surface or cornea of the subject.

[0047]

[0026] In some embodiments, the pharmaceutical formulation further comprises a surfactant or solubilizer, a gelling agent, a gel adjusting agent, a pH adjusting agent, an osmolarity adjusting agent, or a preservative.

[0048]

[0027] A gel adjusting agent is a substance used to modify the properties of a gel formulation, including its viscosity, texture, stability, and release characteristics. For instance, cationic ions (such as calcium ions, sodium ions, or other positively charged ions) are gel adjusting agent forsodium alginate, since they increase viscosity of sodium alginate gel by forming ionic cross¬ linking bonds with the negatively charged carboxylate groups in sodium alginate chains.

[0049]

[0028] Examples of the surfactant or solubilizer include but are not limited to an organic solvent selected from ethanol, isopropyl alcohol, or denatured alcohol; a non-ionic surfactant or solubilizer such as polysorbate 80, polysorbate 60, polysorbate 20, polyethylene glycol (PEG), ethoxylated fatty alcohol, PEG-40 hydrogenated castor oil, polyoxyl castor oil, and polyoxyl hydrogenated castor oil; and an ionic surfactant such as benzalkonium chloride and cetylpyridinium chloride.

[0050]

[0029] Examples of the gelling agent include but are not limited to sodium hyaluronate, deacetylated gellan gum, sodium alginate, xanthan gum, carrageenan, pol oxamer, carboxymethyl cellulose sodium, hydroxypropyl methylcellulose), hydroxyethyl cellulose, and any combination thereof;

[0051]

[0030] Examples of the preservative include but are not limited to sorbic acid, benzalkonium chloride, benzalkonium bromide, methyl hydroxybenzoate, ethyl hydroxybenzoate, EDTA-2Na (ethylenediaminetetraacetic acid disodium salt) or any combination thereof.

[0052]

[0031] In some embodiments, the pharmaceutical formulation includes a surfactant or solubilizer at a concentration ranging from about 0.01wt% to about 5.0 wt%.

[0053]

[0032] In some embodiments, the pharmaceutical formulation includes a gelling agent at a concentration ranging from about 0.01 wt% to about 20 vvt%, or from about 0.3 wt% to about 6 wt%.

[0054]

[0033] In some embodiments, the pharmaceutical formulation has a pH ranging from about 3.5 to about 8.0, or from about 4.0 to about 7.0.

[0055]

[0034] In some embodiments, administering the TRPM8 agonist (e.g., DIPA-9) achieves rapid reduction of corneal fluorescein staining, e.g., starting from week 1 or after one-week treatment.

[0056]

[0035] In some embodiments, the formulation significantly improves tear production as measured by Schirmer’s Test.

[0057]

[0036] In some embodiments, the formulation improves tear quality, stability and quantity, thereby inducing statistically significant reductions in corneal fluorescein staining, both in individual zones such as the inferior zone and across the entire corneal surface.

[0058]

[0037] In some embodiments, the formulation alleviates ocular discomfort symptoms, including pain, burning and / or stinging.

[0038] In some embodiments, the formulation induces a cooling sensory signal. This cooling sensation counteracts discomfort sensations commonly associated with dy' eye disease, thereby- providing rapid subjective symptom relief.

[0059]

[0039] In some embodiments, the formulation results in statistically significant improvements in patient-reported symptoms, as measured by the Eye Dryness Score, SANDE frequency, and SANDE severity. For instance, the method may reduce eye dryness score, or SANDE frequency and severity rapidly, starting within week 1 of administration of the formulation.

[0060]

[0040] In some embodiments, the formulation induces trends in ocular cooling sensations posttreatment, suggesting enhanced patient comfort.

[0061]

[0041] In some embodiments, the method reduces total corneal fluorescein staining by at least 0.7 points relative to baseline (e.g., within week 1 or after one-week treatment).

[0062]

[0042] In some embodiments, the therapeutically effective amount of a TRPM8 agonist is administrated to surface(s) of ocular margin of the subject, including the eyelid and eyelid margin, wherein the ocular margin is the primary target site for delivery of the TRPM8 agonist. For instance, the TRPM8 agonist is delivered at a higher amount to the primary target site than to the cornea, while remaining tolerable and non-irritating.

[0063]

[0043] In some embodiments, the surfaces of the ocular margin include receptive fields located on a supraorbital region, an infraorbital region, a nasal region, a zygomatic region, or any combination thereof.

[0064]

[0044] For instance, the supraorbital region may include upper eyelid, lower eyelid, medial canthus, lateral canthus, and / or superior orbital rim.

[0065]

[0045] In some further embodiments, the upper eyelid may include eyelid margin near eye lashes; while the lower eyelid may include eyelid margin near eyelashes.

[0066]

[0046] In some embodiments, the TRPM8 agonist is topically applied onto the ocular margin with a finger, a cotton swab, a wipe, a pad, a brush, or a spray nozzle.

[0067]

[0047] In some embodiments, the TRPM8 agonist include a DIP A compound, or an analog, derivative, or pharmaceutically acceptable salt thereof,

[0068]

[0048] A further aspect of the present invention provides a method for improving corneal health, comprising a step of administering an effective amount of a TRPM8 agonist to a subject in need thereof, wherein the TR. PM8 agonist comprises DIPA-9. In some embodiments, the method improves corneal health rapidly (e.g, starting within week 1 or after one-week treatment).

[0049] In some embodiments, the TRPM8 agonist is administered in the form of a pharmaceutical formulation comprising DIPA-9 at a concentration ranging from about 0.01% (w / w) to about 0.4% (w / w) and a pharmaceutically acceptable carrier.

[0069]

[0050] In some embodiments, the TRPM8 agonist is administered topically using a wiping, brushing, or spraying method, or finger application, or a traditional eye drop method, to ocular surface or cornea of the subject.

[0070]

[0051] In some embodiments, the formulation significantly improves tear production as measured by Schirmer’s Test.

[0071]

[0052] In some embodiments, the formulation improves tear quality, stability and / or quantity, thereby inducing statistically significant reductions in corneal fluorescein staining, both in individual zones such as the inferior zone and across the entire corneal surface.

[0072]

[0053] In some embodiments, the formulation improves tear quality, stability and / or quantity, thereby inducing statistically significant reductions in corneal fluorescein staining rapidly, starting within week 1, both in individual zones such as the inferior zone and across the entire corneal surface.

[0073]

[0054] In some embodiments, the formulation alleviates ocular discomfort symptoms, including pain, burning and / or stinging.

[0074]

[0055] In some embodiments, the formulation induces a cooling sensory signal. This cooling sensation counteracts discomfort sensations commonly associated with dry eye disease, thereby providing rapid subjective symptom relief

[0075]

[0056] In some embodiments, the formulation results in statistically significant improvements in patient-reported symptoms, as measured by the eye dryness score, and / or SANDE frequency, and SANDE severity.

[0076]

[0057] In some embodiments, the formulation results in statistically significant improvements in patient-reported symptoms, rapidly, starting from 1 week, as measured by the Eye Dryness Score, and / or SANDE frequency, and SANDE severity.

[0077]

[0058] In some embodiments, the formulation induces trends in ocular cooling sensations posttreatment, suggesting enhanced patient comfort.

[0078]

[0059] In some embodiments, the method reduces total corneal fluorescein staining by at least 0.7 points relative to baseline (e.g., within week 1 or after one-week treatment).

[0060] As used herein, the term “of” may also imply “and,” unless such an interpretation results in a contradiction or infeasibility.

[0079]

[0061] As used herein, the terms “treating,” “treatment,” “therapeutic,” or “therapy” do not necessarily mean total cure or abolition of the disease or condition. Any alleviation of any undesired signs or symptoms of a disease or condition, to any extent can be considered treatment and / or therapy.

[0080]

[0062] As used herein, the terms "individual," "patient," or "subject" are used interchangeably. None of the terms require or are limited to situation characterized by the supervision (e.g., constant or intermittent) of a health care worker (e.g., a doctor, a registered nurse, a nurse practitioner, a physician's assistant, an orderly, or a hospice worker).

[0081]

[0063] As used herein, a "therapeutically effective amount" refers to a sufficient amount of TRPM8 agonists, at a reasonable benefit / risk ratio applicable to treating ocular disorders in a subject in need thereof. It is understood, however, that the total daily usage of TRPM8 agonists may be decided by the attending physician or personal coach within the scope of sound medical judgment. The specific effective dose level for any particular subject will depend upon a variety of factors including the other disorder being treated and the severity of the disorder; the specific composition employed, the age, body weight, general health, sex and diet of the subject; the time of administration and route of administration; the duration of the administration; drugs used in combination or coincidental with TRPM8 agonists; and like factors well known in the medical arts or sports science. In addition, a "therapeutically effective amount" is the amount that will elicit the biological or medical response of a tissue, system, or subject that is being sought by a researcher or clinician.

[0082]

[0064] One of skill in the art recognizes that an amount may be considered "effective" even if the condition is not totally eradicated or prevented, but it or its symptoms and / or effects are improved or alleviated partially in the subject. Various indicators for determining the effectiveness of a method are known to those skilled in the art of treating ocular disorders in a subject in need thereof.

[0083]

[0065] Unless otherwise defined herein, scientific and technical terms used in connection with the present application shall have the meanings that are commonly understood by those of ordinary skill in the art to which this disclosure belongs. It should be understood that this invention is not limited to the particular methodology, protocols, and reagents, etc., described herein and as suchcan vary Other terms are defined herein within the description of the various aspects of the invention.

[0084] BRIEF DESCRIPTION OF THE DRAWINGS

[0085]

[0066] Fig. la, Fig. lb, Fig. 1c, Fig. 1 d and Fig. le illustrate data for the sensation of coolness and changes in ocular parameters after a single application of vehicle or DIP A- 9 (also referred to herein as Cryosim-3 or C3), 2 mg / mL.

[0086]

[0067] Fig. 2a, Fig. 2b, Fig. 2c, Fig. 2d, Fig. 2e and Fig. 2f illustrate changes in ocular parameters after four times a day application of vehicle or DIP -9, 2mg / mL for 2-weeks.

[0087]

[0068] Fig. 3a and Fig. 3b illustrate change from baseline in unanesthetized Schirmer’s test score at week 4, which shows: A. Study eye; B. Fellow (Non-study) eye.

[0088]

[0069] Fig. 4a and Fig. 4b illustrate change from baseline in anesthetized Schirmer’s test score at week 4, which shows: A. Study eye; B. Fellow (Non-study) eye.

[0089]

[0070] Fig. 5a and Fig. 5b illustrate change from baseline in total corneal fluorescein staining score across visits, which shows: A Study eye; B. Fellow (Non-study) eye.

[0090]

[0071] Fig. 6a, Fig. 6b and Fig. 6c illustrate change from baseline in SANDE scores across visits, which shows: A. Frequency of symptoms score; B. Severity of symptoms score; C. Global score.

[0091]

[0072] Fig. 7 illustrates change from baseline in Eye Dryness Score across visits.

[0092]

[0073] Fig. 8 illustrates mean change from baseline in CFS score in periods 1 and 2 for total CFS and inferior CFS.

[0093]

[0074] Fig. 9 illustrates mean change from baseline in SANDE score in periods 1 and 2.

[0094]

[0075] Fig. 10 illustrates mean change from baseline in Eye Dryness Score in periods 1 and 2.

[0095]

[0076] Fig. 11 illustrates mean change from baseline in Ocular Discomfort Score in periods 1 and

[0096]

[0077] Fig. 12 illustrates mean change from baseline in tear break-up time in periods 1 and 2.

[0097]

[0078] Fig. 13 illustrates mean change from baseline in unanesthetized Schirmer’s Tear Test scores in periods 1 and 2.

[0098]

[0079] Fig. 14 illustrates mean change from baseline in anesthetized Schirmer’s Tear Test scores in periods 1 and 2.

[0099] DETAILED DESCRIPTION OF THE INVENTION

[0080] Reference will now be made in detail to the preferred embodiments of the invention, examples of which are further illustrated. While the invention will be described in conjunction with the preferred embodiments, it will be understood that they are not intended to limit the invention to these embodiments. To the contrary, the invention is intended to cover alternatives, modifications and equivalents, which may be included within the spirit and scope of the invention as defined by the claims. Furthermore, in the detailed description of the present invention, numerous specific details are set forth in order to provide a thorough understanding of the present invention. However, it will be obvious to one of ordinary skill in the art that the present invention may be practiced without these specific details In other instances, well known methods, procedures, components, and other features have not been described in detail as not to unnecessarily obscure aspects of the present invention.

[0100]

[0081] Generally speaking, the present invention provides a novel therapeutic use of formulations containing TRPM8 agonists for the treatment of dry eye disease, which demonstrates improved efficacy in a variety of clinical endpoints, with significant improvement in SANDE and Corneal Fluorescein Staining (CFS) scores. The present invention offers methods for reducing corneal fluorescein staining (e.g., by at least 0.7 points relative to baseline, even within one-week of treatment) and / or improving corneal health, by administration of a TRPM8 agonist (e.g., DIPA-9 formulation). The present invention has demonstrated unexpected effectiveness (e.g., both intensity and speed) in reducing corneal fluorescein staining, improving clinical symptoms or signs of conditions associated with corneal fluorescein staining including, eg., DED, contact lens wear, trauma / erosions, and neurotrophic keratopathy.

[0101]

[0082] For instance, the invention is particularly effective at about 0.2% concentration of the TRPM8 agonist (e.g., DIPA-9), which consistently demonstrated superior efficacy across all measured endpoints in clinical testing.

[0102]

[0083] In some embodiments, drug-delivery method uses a wiping, or brushing, or spraying, or finger application method to deliver a TRPM8 agonist to targets on the ocular surfaces, which is more effective and efficient as compared to the traditional method using eye drops, or uses a drop method to deliver a TRPM8 agonist to ocular surfaces.

[0103]

[0084] In some embodiments, the formulation according to the present invention can be an aqueous formulation, wherein the aqueous formulation can be used to deliver both water-soluble, water-insoluble, and / or oil-soluble TRPM8 agonists.

[0085] In some embodiments, the TRPM8 agonist-containing formulation can be applied 1-4 times a day to the ocular margin surface of the subject for at least one week, preferably 2 times a day.

[0104]

[0086] In some embodiments, the concentration of TRPM8 agonists in the formulation may be 0.01 mg / mL to 20 mg / mL, preferably ranging from 0.5 mg / mL to 10 mg / mL.

[0105]

[0087] In some embodiments, the pH range of the formulation is 3.5-8.0, preferably ranging from 4.0-7.0.

[0106]

[0088] In some embodiments, the formulation induces a cooling sensory signal. This cooling sensation counteracts discomfort sensations commonly associated with dry eye disease, thereby providing rapid subjective symptom relief.

[0107]

[0089] In some embodiments, the formulation attenuates activity of pain-transmitting neurons and dampens the hyperexcitability of pain nerves, thereby providing sustained subjective symptom relief.

[0108]

[0090] In some embodiments, the formulation results in statistically significant improvements in patient-reported symptoms, as measured by the Eye Dryness Score, and / or SANDE frequency, and SANDE severity.

[0109]

[0091] In some embodiments, the formulation results in statistically significant improvements in patient-reported symptoms rapidly, starting from week 1, as measured by the Eye Dryness Score, and / or SANDE frequency, and SANDE severity.

[0110]

[0092] In some embodiments, the formulation improves tear quality, stability and / or quantity, directly protecting the epithelium from desiccation and friction, thereby resulting in reduced corneal fluorescein staining, both in individual zones such as the inferior zone and across the entire corneal surface.

[0111]

[0093] In some embodiments, the formulation improves tear quality, stability and / or quantity, directly protecting the epithelium from desiccation and friction, resulting in reduced corneal fluorescein staining rapidly, starting from week 1, both in individual zones such as the inferior zone and across the entire corneal surface.

[0112]

[0094] In some embodiments, the formulation significantly improves tear production as measured by Schirmer’s Test.

[0113] Example 1. Clinical study for efficacy

[0114]

[0095] In clinical DIPA-9 studies conducted in Korea in subjects with DED, 60 subjects (n=30 pergroup) participated in a single-dose study, and 40 subjects (n=20 per group) completed a 2 -week, repeat-dose study, DIPA-9 or vehicle (water) was applied with a cotton gauze pad to upper eyelids of subjects with DED (n=30). Cooling sensation, tear film break-up time (TBUT), basal tear secretion, and corneal staining were evaluated. DIPA-9 was then applied 4 times daily (QID) for 2 weeks to subjects using a pre-loaded single-unit applicator containing DIPA-9 2 mg / mL (0.2%) in water (n=20) or water only. After 2 weeks of QID dosing with DIPA-9 0.2% as an eyelid wipe, subjects experienced improved signs and symptoms relative to the control (water) group with no reports of irritation or pain. See, e.g., J. M Yang et al., BMC Ophthalmol., 2017, 77(1), 101.

[0115]

[0096] Fig. la, Fig. lb, Fig. 1c, Fig. Id and Fig. le show data for the sensation of coolness and changes in ocular parameters after a single application of vehicle or DIPA-9, 2 mg / mL.

[0116]

[0097] More specifically, DIPA-9 increased the VAS for ocular cooling score within 5 min after application, and lasted for an average of 46 ± 2 min (Fig la) The DIPA-9 treatment scored higher than the vehicle at every time point and there was a clear-cut pharmacological effect. The sensations reported were of refreshing and dynamic cooling., with an energizing effect. None of patients reported sensing ocular pain or irritation after topical application of vehicle or DIPA-9. The dry eye symptom scores improved significantly after DIPA-9 compared to baselines and was not seen with the vehicle controls (Fig. lb). The TBUT was significantly elevated above baseline at 30 min and 40 min after DIPA-9 (Fig. 1c). The intergroup comparison of the TBUT did not show a significant difference. The basal tear secretion significantly increased at 20 min, 40 min, and 60 min after DIPA-9, but not in the vehicle control group (Fig. Id). The intergroup comparison was highly significant for the 3 time points tested. However, no differences were observed in corneal staining score (CSS) scores of the 2 treatment groups (Fig. le).

[0117]

[0098] Fig. 2a, Fig. 2b, Fig. 2c, Fig. 2d, Fig. 2e and Fig. 2f illustrate changes in ocular parameters after four times a day application of vehicle or DIPA-9, 2mg / mL for 2 -weeks.

[0118]

[0099] The total of 40 patients (n = 20 per group) who completed the 2-week course of study did not differ in age, sex, and baseline symptoms, or in ocular surface parameters and were young adults with a mild degree of DED. The weight of the applicators used in this study decreased by 80 ± 5 mg (Week 1) and 83 ± 4 mg (Week 2) for the vehicle group and 77 ± 7 mg (Week 1) and 81 ± 4 mg (Week 2) for the DIPA-9 group, respectively. As the recommended usage was for 4 times a day the average volume per application off-loaded each time the applicator was used was about 20 µL of water or 40 µg of DIPA-9 for both eyes for a 2 mg / mL solution. The basal tearsecretion was significantly increased in the DIPA-9 group at 1 and 2 weeks when compared to baseline and when compared to the vehicle (Fig. 2b), However, the changes in ocular parameters after vehicle or DIPA-9 showed no significant changes in TBUT and corneal staining score at Week 1 or Week 2 (Fig. 2a, Fig. 2c). The changes in ocular symptoms scores, assessed by 3 questionnaires, showed that the severity of symptoms assessed by VAS score or by the total OSDI score were significantly improved at Week 2 but not at Week 1 after DIPA-9 when compared with the vehicle (Fig. 2d, Fig. 2e). The computer vision syndrome (CVS) type of symptoms was significantly improved at both 1 and 2 weeks after DIPA-9 (Fig. 2f). No significant adverse effect such as ocular pain, irritation, or discomfort was reported from both groups during the 2 weeks study.

[0119] Example 2. Clinical study for treatment of neuropathic ocular pain

[0120]

[0100] Methods: A prospective pilot study of 15 patients with dry eye (DE)-associated neuropathic ocular pain (NOP) was conducted. These patients applied topical DIPA-9, 2 mg / mL to their eyelid, 4 times / day for 1 month. The patients underwent clinical examinations They also completed the Ocular Pain Assessment Survey (OPAS), which is a validated questionnaire for neuropathic ocular pain (NOP), at baseline, 1 week, and 1 month after treatment. See, e.g., HJ. Yoon et al., Topical TRPM8 Agonist for Relieving Neuropathic Ocular Pain in Patients with Dry Eye: A Pilot Study. Journal of clinical medicine, 2021, 10(2), 250.

[0121]

[0101] Results: This study enrolled 20 patients with DE accompanying NOP features. Five patients (25.0%) discontinued the treatment because of drug ineffectiveness or intolerance. The remaining 15 patients (75.0%) were included in the analysis. Five patients had a history of intraocular surgery, and one patient had a history of ocular trauma. At 1 week after treatment, eye pain intensity, Quality of Life (QoL, driving / watching TV, general activity, sleep, and enjoying life / relations with other people), and associated factors (burning sensation, light sensitivity, and tearing) were improved. The total OPAS scores of eye pain intensity, QoL (sleep), and associated factors (burning sensation and light sensitivity) remained improved at 1 month. However, the score of non-eye pain and aggravating factors did not change after treatment (see Table 1). Among the clinical DE parameters, OSDI and Schirmer test score were improved at 1 month after treatment (see Table 2). There w'ere no significant differences in pain scores according to previous medications.Table 1. Changes in the Ocular Pain Assessment Survey scores after the application of DIPA-9 for 1 month

[0122] Baselinea1 Weekb1 Monthcp-Value * a vs. b a vs. c b vs. c Eye pain intensity (0-60) 30.60 ± 12.84 26.47 ± 11.45 21.53 ± 10.84 0.009 0.015 0.073 Non-eye pain (0-20) 7.67 ± 6.22 6.73 ± 6.18 5.47 ± 5.62 0.999 0.435 0.409 Quality of life (total 0-60) 33.53 ± 14.24 27.60 ± 15.49 27. 17 ± 16.06 0.003 0.022 0.743 Reading and / or computer use (0-10) 7.79 ± 1.76 7.14 ± 2.48 6.93 ± 2.59 0.120 0.054 0.272 Driving and / or watching TV (0-10) 6.80 ± 2.31 5.27 ± 2.52 5.60 ± 2.90 0.002 0.070 0.417 General activity (walking, etc.) (0-10) 4.00 ± 3.18 3.27 ± 2.71 3.20 ± 2.86 0.016 0.138 0.843

[0123] Mood (0-10) 5.40 ± 2.77 4.53 ± 2.50 4.40 ± 2.47 0.121 0.177 0.769 Sleep (0-10) 4.27 ± 3.81 2.93 ± 3.67 2.73 ± 3.81 0.027 0.049 0.486 Enjoying life / relations with other 5.07 ± 2.84 4.33 ± 2.97 4.27 ± 3.03 0.036 0.068 0.806 people (0- 10)

[0124] Aggravating factors (total 0-2) 1.11 ± 0.49 0.87 ± 0.56 0.88 ± 0.57 0.113 0.132 0.077 Mechanical stimuli (0-1) 0.63 ± 0.29 0.47 ± 0.25 0.47 ± 0.26 0.068 0.086 0.999 Chemical stimuli (0-1) 0.47 ± 0.35 0.41 ± 0.35 0.41 ± 0.32 0.363 0.432 0.872 Associated factors (total 0-4) 2.09 ± 0.76 1.55 ± 0.85 1.58 ± 0.93 0.006 0.046 0.835 Redness (0–1) 0.41 ± 0.32 0.41 ± 0.30 0.39 ± 0.30 0.094 0.104 0.080 Burning sensation (0-1) 0.57 ± 0.37 0.40 ± 0.33 0.29 ± 0.29 0.007 0.002 0.015 Sensitivity to light (0-1) 0.76 ± 0.24 0.57 ± 0.26 0.59 ± 0.28 0.005 0.030 0.663 Tearing (0-1) 0.36 ± 0.29 0.17 ± 0.18 0.21 ± 0.27 0.013 0.197 0.578

[0125] All values are presented as mean ± SD * Compared using repeated measures analysis of variance with Bonferroni ’s post hoc. test, (a) means “baseline,” (b) means “1-week”, and (c) means “ 1 -month''Table 2. Changes in clinical parameters after the application of DIPA-9 for 1 month.

[0126] BASELINE 1 Month Z p- Value Ocular surface

[0127] 57.5 ± 13.8 40.2 ± 12.6 -3.41 0.001 disease index

[0128] Tear break-up time

[0129] 4 13 ± 0.83 4.00 ± 0.85 -0.82 0.414 (s)

[0130] Schirmer test score

[0131] 7.07 ± 2.76 8.47 ± 2.80 -3.02 0.003 (mm)

[0132] Corneal staining

[0133] 0.60 ± 0.91 0.13 ± 0.35 -1.82 0.068 score (0-9)

[0134] All values are presented as mean ± SD. Compared using the Wilcoxon signed rank test.

[0135]

[0136] Example 3. Phase 2b clinical trial study of WS-12 formulation

[0137]

[0102] WS-12 is a selective TRPM8 receptor agonist compound that activates cold-sensitive sensory neurons to stimulate lacrimation and modulate neurosensory signaling, which has the chemical structure of (lR,2S,5R)-2-isopropyl-N-(4-methoxyphenyl)-5-methyl cyclohexane-1-carboxamide

[0138]

[0103] Study design: At the end of the Screening Visit, all qualified subjects were assigned to administer WS-12 vehicle twice a day to both eyes for 14 days (vehicle run-in period). After the vehicle run-in period, subjects were re-evaluated for signs and symptoms of Dry Eye Disease (DED). Subjects who requalify based on inclusion / exclusion criteria will be randomized in a 1:1:1 ratio to receive WS-12 0.0014% (lower dose), WS-12 0.003% (higher dose) or WS-12 vehicle administered as 1 drop in each eye twice daily for 84 days. This study utilized a Controlled Adverse Environment (CAE) chamber. Efficacy endpoints were evaluated pre- and post- exposure to the CAE.

[0139]

[0104] Results: The results showed that 0.003% WS-12 (n = 122) was associated with early and sustained improvements in unanesthetized Schirmer score (Days 1 and 14, p < 0.0001), improvements in ocular surface staining (measured with total corneal and total conjunctival staining, Days 14 and 84, p ≤ 0.0365) and hyperemia (Day 84, p < 0.0215). Statistically significant improvements in symptoms were observed for the 0.003% concentration on SANDE (Days 14, 28, and 84, p ≤ 0.0254), ODS-VAS (Day 84, p = 0.0281), Eye Dryness-VAS (Day 84, p = 0.0302), and multiple QoL measures (Days 14, 28, and 84, p < 0.05). See, e.g., Wirta, David L., et al, The Ocular Surface, 2022, 26, 166-173. There are no effective data or results for WS-12 at week 1.Example 4. Phase 3 pivotal studies (COMET-2 and COMET-3) of WS-12 formulation

[0105] Study design: To evaluate the safety and efficacy of the transient receptor potential melastatin 8 agonist WS-12 on signs and symptoms of dry eye disease (DED), two identical randomized, multicenter, double-masked, vehicle-controlled phase 3 studies were conducted. Patients (COMET-2, N = 465; COMET-3, N = 466) were randomized 1:1 to 0.003% (WS-12 formulation) or vehicle twice daily for 90 days.

[0140]

[0106] Results: The primary end point (the proportion of patients achieving a 10-mm or greater increase in unanesthetized Schirmer test score from baseline) was met in both studies, with more patients receiving WS-12 formulation achieving a 10-mm or greater increase in Unanesthetized Schirmer Test on day 14 (WS-12 formulation vs. vehicle: COMET-2, 42.6% vs. 8.2%; COMET-3, 53.2% vs. 14.4%; P < 0.0001 for both).

[0141]

[0107] Reduction in global SANDE score by day 28 (key secondary end point) favored WS-12 formulation in both studies, and was statistically significant in COMET-2, but not in COMET-3. Reductions with WS-12 formulation were observed in total CFS on days 28 and 90, but not on Day 14; and in total conjunctival staining at all time points (nominal significance due to hierarchical testing). The most common adverse event was instillation-site pain (burning or stinging), reported in approximately 51—53% of WS-12 formulation-treated patients. See, e.g., Pattar, Guruprasad R., et al., Ophthalmology, 2025. There are no effective data or results for WS-12 at week 1.

[0142] Example 5. U. S. Phase 2a Clinical trial study of a DIPA-9 formulation

[0143]

[0108] A randomized, multicenter, parallel, vehicle-controlled, double-masked Phase 1 / 2a study (NCT06400459) was performed to evaluate the safety, tolerability, and efficacy of DIPA-9 Ophthalmic Eyelid Wipes in subjects with DED at 10 investigative sites throughout the United States. Three groups were compared: vehicle (placebo), DIPA-9 0.1% (low dose), and DIPA-9 0.2% (high dose). The study was conducted in accordance with Good Clinical Practice guidelines and International Organization for Standardization (ISO) 14155, adhered to the tenets of the Declaration of Helsinki, and each study site received prospective Institutional Review Board (IRB) approval from Sterling IRB (Atlanta, GA). Informed consent was obtained from each subject prior to the initiation of any study-specific procedures.

[0144]

[0109] Study design: After screening, subjects participated in a single (subject)-masked run-in period, in which all subjects dosed BID for 7 days with DIPA-9 Ophthalmic Eyelid Wipe Placebo(vehicle, containing no active ingredient) by administering the eyelid wipe to the upper eyelid margin of both study eye and fellow eye. After this on Day 1, eligible subjects entered a doublemasked treatment period and were randomized independently at each site in a 1:1:1 manner to DIPA-9 Ophthalmic Eyelid Wipe 0.2% (high dose), DIPA-9 Ophthalmic Eyelid Wipe 0.1% (low dose), or vehicle. Subjects were randomized centrally through an interactive web response system. Randomization was implemented using blocks of 3 and was stratified by study site.

[0145]

[0110] Subjects dosed investigational product (IP) BID to both eyes for 28 days. Subjects attended weekly visits: Baseline (Day 1), Week 1 (Day 8), Week 2 (Day 15), Week 3 (Day 22), and Week 4 (Day 29).

[0146]

[0111] Outcomes: The efficacy endpoints included the mean change from baseline in unanesthetized Schirmer’s test score in the study eye at Week 4 (primary) and mean change from baseline in unanesthetized Schirmer’s test score at Weeks 1, 2, and 3; in total CFS and inferior CFS scores at all 4 visits; and in anesthetized Schirmer’s test at Week 4. Exploratory efficacy endpoints of both eyes together at all 4 visits included Symptom Assessment in Dry Eye (SANDE) Questionnaire, Eye Dryness Score (EDS) Visual Analog Scale (VAS) (0-100), Ocular Discomfort (ODS) VAS (0-100), and investigational product (IP) comfort VAS (American Society of Heating, Refrigerating and Air-Conditioning Engineers [ASHRAE]; 7-point scale).

[0147]

[0112] Results: Efficacy-DED signs

[0148]

[0113] The Schirmer’ s test was performed by Investigators for each eye separately, both with and without anesthetizing the eye. Due to a large vehicle effect, which is common in DED trials, comparisons of DIPA-9 0.2% to vehicle did not achieve statistical significance. Fig. 3a and Fig.

[0149] 3b illustrate change from baseline in unanesthetized Schirmer’s test score at week 4. In the study eye at Week 4 (Fig. 3a), the LS mean (standard error [SE] ) change from baseline in unanesthetized Schirmer’s tests score was 0.94 (0.87) mm in the vehicle arm, 1.49 (0.84) mm in the DIPA-90.1% arm, and 1.79 (0.88) mm in the 0.2% arm, showing no statistically significant differences in the DIPA-9 arm compared with vehicle. The fellow (non-study) eye also did not demonstrate a significant difference in the DIPA-9 arm compared with vehicle (Fig. 3b). There was a trend toward improvement in the 0.2% arm compared with the 0.1% arm (p=0.0511), suggesting potential dose-dependent efficacy.

[0150]

[0114] The anesthetized Schirmer’s test, which was performed only at Baseline and Week 4, produced similar results. Fig. 4a and Fig 4b illustrate change from baseline in anesthetizedSchirmer’s test score at week 4. In the study eye (Fig. 4a), the LS mean (SE) change from baseline in anesthetized Schirmer’s test score was 0.24 (0.74) mm in the vehicle arm, 0.35 (0.70) mm in the DIPA-9 0.1% arm, and 1.38 (0.74) mm in the 0.2% arm, showing no statistically significant differences in the DIPA-9 arms compared with vehicle. The fellow (non-study) eye also did not demonstrate a significant difference in the DIPA-9 arms compared with vehicle (Fig. 4b). There was an evident trend toward improvement in the 0.2% arm compared with the 0.1% arm and the vehicle, suggesting potential dose-dependent efficacy.

[0151]

[0115] Total Corneal Fluorescence Staining (CFS) change from baseline (CFB) scores are presented in Fig. 5a and Fig. 5b In the study eye (Fig. 5a), the total CFS score of the DIPA-90.2% arm was statistically significantly reduced (i.e., improved) compared with the vehicle arm at Week 4 (p<0.05). In the fellow eye (Fig. 5b), the DIPA-9 0.2% arm had significantly reduced scores compared with vehicle at all time points, starting from week 1 (p<0.05), and the 0 1% arm had significantly reduced scores compared with vehicle at Week 1 (p<0.05).

[0152]

[0116] These results indicate significant reduction of CFS and improvement in corneal health, which were unexpected based on prior studies, especially rapid corneal fluorescence staining improvement, starting from week 1.

[0153]

[0117] For inferior CFS change from baseline scores, For the fellow eye, at Week 2 and Week 4, the DIPA-9 0.2% score was significantly reduced (i.e., improved) compared with both vehicle (p=0.0056 and p=0.0318, respectively) and 0.1% (p=0.0034 and p=0.0128, respectively). These results indicate significant improved corneal health, which was unexpected based on prior studies

[0118] DED symptoms: The SANDE subject questionnaire examined the frequency and severity of DED symptoms. These 2 outcomes, as well as a combined global score, were presented. Fig.

[0154] 6a, Fig. 6b and Fig. 6c illustrate change from baseline in SANDE scores across visits. The DIPA-9 0.2% arm had statistically significantly reduced scores (i.e., improvements) compared with both vehicle and 0.1% at each visit (Weeks 1, 2, 3, and 4) and for each outcome (frequency [Fig. 6a], severity [Fig. 6b], and global scores [Fig. 6c]; p<0.05). These results indicate significant improved symptom scores, especially rapid symptomatic improvement, starting from week 1, which was unexpected based on prior studies.

[0155]

[0119] For unanesthetized Schirmer’s score, DIPA-9 0.2% increased tear production (+2.1 mm vs. +0.9 mm for vehicle), showing a promising trend. For anesthetized Schirmer’s score, DIPA-9 0.2% significantly improved basal tear production (+1.7 mm vs. 0 mm for vehicle).

[0120] Eye dryness (both eyes assessed together) was reported by subjects using a 0 to 100 VAS Fig. 7 illustrates change from baseline in Eye Dryness Score (EDS) across visits. Statistically significant differences in least-squares (LS) mean change from baseline EDS scores were observed at Week 4, when the DIPA-9 0.2% arm showed significantly larger reductions (improvements) compared with both vehicle and 0.1% (p<0.05; Fig. 7). This improvement highlights the efficacy of DIPA-9 in improving multi patient-reported dryness symptoms with statistical significance, including both SANDE and EDS. These results indicate unexpected consistent and statistically significant symptom improvement when administering DIPA-90.2% formulation through eyelid wiping on upper eyelid.

[0156]

[0121] DIPA-9 0.2% reduced ocular discomfort scores (-17.2) more than vehicle (-12.0), demonstrating symptom relief that complements its efficacy in improving ocular surface health.

[0157]

[0122] DIPA-9 0.2% showed trends toward increased ocular cooling sensations within 5-30 minutes post-dose, with 39.6% of participants reporting a ‘‘slightly cool” sensation compared to 30.6% in the vehicle group at 5 minutes post-dose.

[0158]

[0123] Unlike similar agents targeting dry eye disease, DIPA-9 formulations showed statistically significant improvements in corneal fluorescein staining (CFS), SANDE scores, and thermal sensations. This was particularly surprising given the known MOA, which primarily suggested efficacy in improving tear production (as evidenced by Schirmer’s Test). Prior agents with similar MOAs demonstrated little to no impact on corneal staining, making the present findings non-obvious and inventive. Collectively, the data presented herein support the inventive use of DIPA-9 formulations, particularly at the 0.2% concentration, for the treatment of signs and symptoms of dry eye disease. The combination of improvements across multiple endpoints, including the unexpected and significant reductions in corneal fluorescein staining, represents a novel therapeutic approach to addressing both symptoms and underlying ocular surface damage, through targeted delivery of TRPM8 agonists and demonstrates unexpected clinical meaningful benefits beyond tear production alone.

[0159]

[0124] Also, none of the prior studies known to the inventors disclose such rapid reduction of for CFS and improvement in corneal health (e.g., starting within week 1). The previously approved drugs for the treatment of dry eye, such as Restasis or Cequa, which are eye drop formulations containing cyclosporin, an anti-inflammatory agent, and they reduced Corneal Fluorescence staining (CFS) score after 6 or 3 months, respectively Surprisingly, the method according to thepresent invention has achieved not only significant but also rapid improvements of Corneal Fluorescence staining (CFS), as well as corneal health. This was totally unexpected based on prior studies, especially the rapidness and intensity of corneal fluorescence staining improvement (e.g., reduction of total CFS by 0.7 points relative to baseline), starting from week 1.

[0160] Example 6. U. S. Phase 2b Clinical trial study of DIPA-9 formulations

[0161]

[0125] A randomized, single center, double-masked Phase 2b study (NCT07161011) was performed to evaluate the safety, tolerability, and efficacy of DIPA-9 Ophthalmic Eyelid Wipe dosing techniques and select optimal dosing concentration in subjects with DED. In Period 1, three dosing techniques were compared: wiping upper eyelid skin slight away from the eyelash line, wiping upper eyelid skin directly at the lash margin covering the eyelash line, and laying the wipe on the upper eyelid skin for 5 seconds and then wiping directly at the lash margin covering the eyelash line. In Period 2, two groups were compared: DIPA-9 0.2% and DIPA-9 0.4%.

[0162]

[0126] Study design: In Period 1 subjects participated in a 50-day, double-masked three dosing technique comparisons period using DIPA-9 Ophthalmic Eyelid Wipe 0.2% followed by a 14-day between period washout. In Period 2 subjects participated in a 7-day, single-masked treatment dosing technique period using either DIPA-9 Ophthalmic Eyelid Wipe 0.2% or 0.4%.

[0163]

[0127] Each endpoint was presented descriptively by visit (absolute values and changes from baseline as appropriate). The changes from baseline were over the four 7-day dosing periods where each dosing period will have its own baseline. Continuous variables will be summarized by descriptive statistics (number of subjects, mean, SD, median, minimum, and maximum), and categorical variables were summarized using the count and percentage of subjects within each category'.

[0164]

[0128] Results: Total and inferior zone CFS: Mean change from baseline (CFB) in total corneal fluorescein staining (“tCFS”) and inferior-zone CFS (“inCFS”) was summarized across administration techniques and study periods in Fig. 8.

[0165]

[0129] During Period 1, all patients received DIPA-9 0.2%. Techniques 1 and 2 demonstrated comparable reductions in tCFS (mean CFB approximately —0.7), whereas Technique 3 produced a greater reduction (mean CFB approximately -1.0). Within-group analysis showed a statistically significant improvement for Technique 3 (p = 0.0307); however, no statistically significant differences were observed in the between-group analysis (Technique 1 vs Techniques 2 and 3: p = 0.9480 and p = 0.5410; Technique 2 vs Technique 3: p = 0. 3924). Dosing Techniques 1 and 2showed similar results, while Dosing Technique 3 was numerically better. Technique 2 was selected as the optimal delivery method for Phase 3; it maintains clinical efficacy while reducing chances of direct ocular surface contact.

[0166]

[0130] In Period 2, DIPA-9 0.2% demonstrated a greater reduction in tCFS compared with DIPA- 9 0.4%. The within-group change for the DIPA-9 0.2% group was statistically significant (p = 0.0165), whereas the between-group comparison did not reach statistical significance (p = 0.480) with a small sample size of 15 patients at each group, indicating that the 0.2% concentration was sufficient to effectively reduce tCFS, while the 0.4% concentration did not provide additional benefits. In both Period 1 and 2, DIPA-9 0.2% demonstrated consistent and stable reduction in Corneal Fluorescein Staining (CFS), validating DIPA-9 ’s impact on ocular surface health.

[0167]

[0131] Similar trends were observed for inferior-zone CFS. In Period 2, DIPA-9 0.2% produced a greater reduction in inferior-zone CFS (mean CFB approximately -0.4) compared with DIPA-9 0.4% (mean CFB approximately -0.2). The reduction observed with DIPA-9 0.2% was statistically significant in the within-group analysis relative to baseline (p = 0.0150).

[0168]

[0132] SANDE: Mean change from baseline (CFB) in SANDE Frequency, Severity, and Global scores was summarized across administration techniques and treatment periods. Fig. 9 illustrates mean change from baseline in SANDE score in periods 1 and 2. Across all conditions, negative mean CFB values were observed, indicating improvement from baseline.

[0169]

[0133] During Period 1, in which all subjects received DIPA-9 0.2%, Technique 1 produced numerically greater reductions across all SANDE domains (mean CFB approximately -11 to -12) compared with Technique 2 (approximately ~5 to ~8) and Technique 3 (approximately -9 to -10).

[0170]

[0134] In Period 2, overall improvements were slightly smaller than those observed in Period 1. DIPA-9 0.4% demonstrated numerically greater reductions in SANDE scores (mean CFB approximately 7 to 8) compared with DIPA-9 0.2% (mean CFB approximately 1 to 6) without significant difference between groups (p = 0.3772 for frequency; p = 0.9293 for severity; p = 0. 5881 for global score). Reliable symptomatic relief confirmed in both Period 1 and 2 and SANDE scores across multiple trials confirmed that DIPA-9 formulations delivered reproducible reductions in both symptom frequency and severity.

[0171]

[0135] EDS: Eye Drymess Score (EDS) was assessed using a visual analog scale (VAS) ranging from 0 to 100. Fig. 10 depicts the change from baseline in EDS across study visits. In Period 1, statistically significant reductions in mean change from baseline (CFB) EDS scores were observedwithin group (p = 0.0034, 0.0005, 0.0109 for Technique 1, 2, and 3, respectively). Similarly, in Period 2, both DIPA-9 0.2% and 0.4% demonstrated reductions in mean CFB EDS scores, reflecting an improvement in patient-reported eye dryness symptoms. Consistency in EDS improvements across study periods were validated.

[0172]

[0136] ODS: Ocular Discomfort Score (ODS) was assessed using a 0-100 visual analog scale (VAS). Fig. 11 illustrates the change from baseline (CFB) in ODS across study visits. Reductions in mean CFB ODS were observed in both Period 1 and Period 2, showing improvement of in the following symptoms including burning / stinging, itching, pain, sandy / gritty sensation, and sensitivity to light. Positive Ocular Discomfort Score (ODS) trends further support the 0.2% dose as the optimized dose for comprehensive symptom management.

[0173]

[0137] TBUT: Tear Break-Up Time measures the time (in seconds) it takes for the tear film to start breaking up after a blink. As illustrated in Fig 12, the positive mean CFB of TBUT Statistically significant within-group improvement with DIPA-9 0.2% (p = 0.0456 for Technique 2 in Period 1 and p = 0.0203 for DIPA-9 0.2% in Period 2), indicating enhanced tear film stability over the study period. The TBUT Data confirmed enhanced Tear Film Stability and measurement of TBUT at 0.2% confirms a significant therapeutic impact on the mechanical stability of the tear film. These findings were consistent with the observed reductions in CFS for ocular surface health improvement and patient-reported eye dryness and ocular discomfort,

[0174]

[0138] uSTT and aSTT: The Schirmer tear test was performed with (aSTT) and without (uSTT) topical anesthesia to assess basal tear secretion alone and total tear production (including reflex tearing), respectively. As illustrated in Fig. 13 and Fig. 14, the positive mean CFB of aSTT and uSTT was observed in DIPA-9 0.2% for Technique 2, indicating improvement in tear production. However, the Schirmer's test was a blunt instrument with high variability. The test measured tear production but did not evaluate tear quality or stability. Technique variations, such as the use of anesthetics which aims to differentiate basal from reflex tearing, indicate poor repeatability, sensitivity and specificity. See, e.g., Wolffsohn, James S.. et al., The ocular surface, 2017, 75(3), 539-574. Despite intrinsic test variability, numerical trends in tear secretion confirmed the TRPM8 -mediated stimulation of natural lacrimation.

[0175]

[0139] Conclusion: Across both study periods, DIPA-9 demonstrated consistent improvements in objective signs and patient-reported symptoms. In Period 1, all administration techniques with DIPA-9 0.2% reduced total and inferior corneal fluorescein staining, SANDE scores, EDS, ODS,

[0176] 73and improved TBUT; Technique 2 was selected for Period 2 based on comparable efficacy and greater convenience. In Period 2, DIPA-9 0.2% showed greater results in corneal staining and TBUT, while symptom improvements (SANDE, EDS, ODS) were observed for both concentrations.

[0177]

[0140] Compared to clinical trials of other TRPM8 agonists such as WS-12, the clinical trials using DIPA-9 demonstrated promising results in quickly improving CFS and SANDE scores with no reported installation-site pain through eyelid delivery. Statistically significant improvements in both CFS were seen in one week during phase 2b clinical trials. The rapidness and magnitude of such CFS improvements were unexpected, compared to the WS-12 phase 3 trials which only saw definitive improvements at two weeks or later. SANDE, similarly, showed quicker improvements with DIPA-9 at one week compared to WS-12 where improvements appeared two weeks or later.

[0178]

[0141] Although specific embodiments and examples of this invention have been illustrated herein, it will be appreciated by those skilled in the art that any modifications and variations can be made without departing from the spirit of the invention. The examples and illustrations above are not intended to limit the scope of this invention. Any combination of embodiments of this invention, along with any obvious extensions or analogs, are within the scope of this invention. Further, it is intended that this invention encompass any arrangement, which is calculated to achieve that same purpose, and all such variations and modifications fall within the scope of the appended claims.

[0179]

[0142] All the features disclosed in this specification (including any accompanying claims, abstract and drawings) may be replaced by alternative features serving the same, equivalent or similar purpose, unless expressly stated otherwise. Thus, unless expressly stated otherwise, each feature disclosed is one example of a generic series of equivalent or similar features.

Claims

What Is Claimed Is:

1. A method for reducing corneal fluorescein staining, comprising a step of administering an effective amount of a TRPM8 agonist to a subject in need thereof, wherein the TRPM8 agonist comprises a 1-diisopropylphosphinoylalkane (DIPA) compound.

2. The method of claim 1, wherein the DIPA compound is DIPA-9.

3. The method of claim 1 or 2, wherein the TRPM8 agonist is administered in the form of a pharmaceutical formulation comprising the TRPM8 agonist and a pharmaceutically acceptable carrier.

4. The method of claim 3, wherein the pharmaceutical formulation is in the form of liquid, solid, semi-solid, gel, cream, or ointment.

5. The method of claim 3 or 4, w'herein the TRPM8 agonist is contained in the pharmaceutical formulation at a concentration ranging from about 0.01% (w / w) to about 0.4% (w / w).

6. The method of claim 5, wherein the TRPM8 agonist comprise DIPA-9 contained in the pharmaceutical composition at a concentration of about 0.2% (w / w).

7. The method of any one of claims 1 to 6, wherein the TRPM8 agonist is administered topically using a wiping, brushing, or spraying method, or finger application, or a traditional eye drop method, to the ocular surface or cornea of the subject.

8. The method of claim 7, wherein the topical administration of the TRPM8 agonist is on the skin surface around or near eye(s) of the subject.

9. The method of any one of claims 3 to 8, wherein the pharmaceutical formulation further comprises a surfactant or solubilizer, a gelling agent, a gel adjusting agent, a pH adjusting agent, an osmolarity adjusting agent, or a preservative.

10. The method of claim 9, wherein the pharmaceutical formulation comprises the surfactant or solubilizer at a concentration ranging from about 0.01wt% to about 5.0 wt%; or the pharmaceutical formulation comprises the gelling agent at a concentration ranging from about 0.01 wt% to about 20 wt%, or from about 0.3 wt% to about 6 wt%.

11. The method of any one of claims 3 to 10, wherein the pharmaceutical formulation has a pH ranging from about 3.5 to about 8.0, or from about 4.0 to about 7.0.

12. The method of any one of claims 1 to 11, wherein administering the TRPM8 agonist achieves rapid reduction of corneal fluorescein staining starting from week 1.

13. The method of any one of claims 1 to 12, wherein the administration of the TRPM8 agonist improves tear production as measured by Schirmer’s Test.

14. The method of any one of claims 1 to 13, wherein the method induces a cooling sensation and alleviates ocular discomfort symptoms.

15. The method of any one of claims 1 to 14, wherein the method reduces Eye Dryness Score, or SANDE frequency and severity.

16. The method of any one of claims 1 to 15, wherein the method reduces Eye Dryness Score, or SANDE frequency and severity rapidly, starting from week 1.

17. The method of any one of claims 1 to 16, wherein the method reduces total comeal fluorescein staining by at least 0.7 points relative to baseline.

18. A method for treating an ocular disorder associated with corneal fluorescein staining in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a TRPM8 agonist to reduce corneal fluorescein staining in the subject.

19. The method of claim 18, wherein the TRPM8 agonist is DIPA-9.

20. The method of claim 18 or 19, wherein the ocular disorder is erosion or contact lens wear of the corneal, infection, inflammation, dry eye disease (DED), Sjogren's syndrome, diabetes, or neurotrophic keratopathy.

21. A method for improving corneal health, comprising a step of administering a therapeutically effective amount of a TRPM8 agonist to a subject in need thereof, wherein the TRPM8 agonist comprises DIPA-922. The method of claim 21, wherein the TRPM8 agonist is administered in the form of a pharmaceutical formulation comprising DIPA-9 at a concentration ranging from about 0.01% (w / w) to about 0.4% (w / w) and a pharmaceutically acceptable carrier.

23. The method of claim 21 or 22, wherein the method improves tear production as measured by Schirmer’s Test; induces a cooling sensation and alleviates ocular discomfort symptoms; or reduces Eye Dryness Score, or SANDE frequency and severity.

24. The method of any one of claims 21 to 23, wherein the method reduces total corneal fluorescein staining by at least 0.7 points relative to baseline.

25. Use of a TRPM8 agonist for manufacture of a medicament for reducing corneal fluorescein staining or improving corneal health in a subject in need thereof.

26. The use of claim 25, wherein the TRPM8 agonist is a DIP A compound.

27. The use of claim 26, wherein the DIP A compound is DIPA-9

Images