Hypotonic gel-forming formulations with enhanced rheological properties - Patents.com

Abstract

Improved hypotonic gelling vehicles are described as solubilizing agents for drugs and as a means of providing sustained drug delivery to the eye. The addition of small amounts of low molecular weight PEG to hypotonic gel-forming compositions enhances the viscosity of the solution on the ocular surface, prolongs tear film breakup time, and increases ocular comfort. It has been discovered that while solubilizing drugs at higher concentrations enhances drug penetration into body tissues, hypotonic gelling vehicles further improve the distribution of drugs over a larger surface area for increased absorption and sustained release for reduced side effects and longer duration of action.

Description

[Technical field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims priority to U.S. Provisional Patent Application No. 63 / 287,415, filed December 8, 2021, by Laura Ensign, Tung Heng Henry Hsueh and Justin Hanes ("Hypotonic Gel-forming Formulations with Enhanced Rheological Properties"), the disclosures of which are incorporated by reference herein in their entirety.

[0002] STATEMENT REGARDING FEDERALLY SPONSORED RESEARCH OR DEVELOPMENT This invention was made with Government support under Grant No. EY031041 awarded by the National Institutes of Health. The Government has certain rights in this invention.

[0003] FIELD OF THEINVENTION The present invention is generally in the field of formulations for enhanced drug delivery, particularly at epithelial surfaces such as the ocular surface. [Background technology]

[0004] 2. Background of the Invention The eye is a highly complex structure with unique anatomy and physiology. The internal anatomy of the eye includes the anterior (front) segment, which occupies approximately 1 / 3 of the total area of ​​the eye, and the posterior (rear) segment, which constitutes the remaining 2 / 3 of the area. Tissues such as the cornea, conjunctiva, aqueous humor, iris, ciliary body, and lens make up the anterior segment, while the posterior segment includes the sclera, choroid, retinal pigment epithelium, neural retina, optic nerve, and vitreous humor.

[0005] Both the anterior and posterior segments of the eye are affected by a variety of ocular diseases and disorders, many of which are vision-threatening, including glaucoma, allergic conjunctivitis, anterior uveitis and cataracts, age-related macular degeneration (AMD), and diabetic retinopathy. Although many therapeutic agents are available for treating ocular diseases, delivery of effective amounts of drugs to the interior of the eye often requires uncomfortable and potentially dangerous invasive techniques (e.g., intraocular injections).

[0006] Sustained delivery of drugs across the mucosal surface of the eye has the potential to improve the treatment and prevention of many ocular diseases, including infectious, inflammatory, and degenerative ocular conditions. However, achieving sustained prophylactic or therapeutic drug concentrations using traditional soluble dosage forms remains difficult due to drug degradation, rapid excretion, and rapid systemic absorption.

[0007] One mechanism for the delivery of therapeutic agents to the eye is by application of topical agents onto the ocular surface, for example by the formation of an artificial tear film. The tear film (a liquid layer bathing the cornea and conjunctiva) is responsible for the comfort, mechanical, environmental and immune protection, and epithelial health of the ocular surface; it forms a smooth refractive surface for vision. Tear prevents drying by coating the ocular surface, and also protects the eye from external stimuli. Since there are no blood vessels on the ocular surface, oxygen and nutrients are transported to the surface cells by the tear. In addition, foreign bodies that enter the eye are washed away by the tear.

[0008] Recent approaches to drug delivery to the anterior segment include the adjustment of traditional topical solutions with permeation and viscosity enhancers, as well as the development of traditional topical formulations (e.g., suspensions, emulsions, and ointments). The use of gels to coat the eye for drug delivery has also been envisioned (see, e.g., U.S. Publication Nos. 2021 / 0196837 and 2021 / 0177751). Various nanoformulations have also been introduced for anterior ocular drug delivery, and drug release devices and nanoformulations are being developed for posterior ocular delivery, for example, to treat chronic vitreoretinal diseases. However, many topical formulations have poor rheological properties (e.g., high adhesion and poor viscosity), resulting in discomfort and gel breakdown upon blinking.

[0009] In particular, there remains a pressing need for enhanced delivery systems that provide effective drug delivery through mucosal surfaces to epithelial cells without invasive, vision-disruptive, or uncomfortable procedures for repeated or long-term treatment of chronic diseases and disorders. There is also an unmet need for compositions for mucosal delivery that provide retention and sustained release of prophylactic, therapeutic, or diagnostic agents at mucosal surfaces.

[0010] It is therefore an object of the present invention to provide a drug delivery composition for increased ocular comfort and lubrication with reduced stickiness.

[0011] It is also an object of the present invention to provide a composition for delivery of active agents to the eye that provides a physical barrier to pathogen entry. It is a further object of the present invention to provide improved compositions for delivery to epithelial tissues across mucosal surfaces, which allow for the retention and sustained release of prophylactic, therapeutic or diagnostic agents across the mucosal surface to the epithelial tissues. [Prior art documents] [Patent documents]

[0012] [Patent Document 1] US Patent Application Publication No. 2021 / 0196837 [Patent Document 2] US Patent Application Publication No. 2021 / 0177751 Summary of the Invention [Means for solving the problem]

[0013] Summary of the Invention It has been discovered that the addition of small amounts of low molecular weight polyethylene glycol ("PEG") to a hypotonic gel-forming composition enhances the viscosity of the solution at the ocular surface, prolongs the tear break-up time, and increases ocular comfort. A composition has been developed with improved properties for effective delivery of drugs to epithelial tissues having mucosal surfaces, particularly to the eye, to treat ocular diseases and disorders. The composition comprises a hypotonic gel-forming polymer in combination with low molecular weight PEG, which has reduced adhesion and low tack to the ocular surface while maintaining the low viscosity required for ocular application in the form of eye drops. The formulations include a therapeutic, prophylactic, nutraceutical, and / or diagnostic agent, a gel-forming polymer for application to a mucosal or epithelial surface formulated to be at a concentration below the critical gel concentration (CGC) of the polymer under isotonic conditions and at a temperature between room and body temperature (about 25 to about 37° C.), a low molecular weight PEG in an amount of between about 0.1% to about 1.0%, inclusive, by total weight / solution (w / v), and excipients to form a pharma- ceutically acceptable hypotonic formulation of the polymer suitable for delivery to a mucosal or epithelial surface of an individual in need thereof.

[0014] Preferably, the gel-forming polymer is between more than 10% and less than 18% in the aqueous vehicle. In a preferred embodiment, the gel-forming polymer is a poloxamer, such as poly(ethylene glycol)-block-poly(propylene glycol)-block-poly(ethylene glycol) (Pluronic® F127). In some embodiments, the gel-forming polymer is between 10% and 16% Pluronic® F127. Preferred PEGs are PEG 100, PEG 200, PEG 300, PEG 400, PEG 600, PEG 800, PEG 1,000, PEG 2,000, and most preferably PEG 400. In a preferred embodiment, the PEG is in an amount between about 0.01% w / v and about 1% w / v inclusive, preferably 0.4% w / v.

[0015] In some embodiments, the therapeutic, prophylactic, nutritional supplement, or diagnostic agent to be delivered is water soluble. In other embodiments, the therapeutic, prophylactic, nutritional supplement, or diagnostic agent to be delivered is poorly water soluble. In some embodiments, the agent is a protein or peptide, a small molecule, a sugar or polysaccharide, a lipid, a glycolipid, a glycoprotein, a nucleic acid, an oligomer or polymer thereof, or a small molecule. Preferred agents for delivery include steroids, glaucoma agents, tyrosine kinase inhibitors, immunosuppressants, anti-fibrotic agents, anti-infective agents, hormones, or chemotherapeutic agents.

[0016] The formulation releases the therapeutic, prophylactic or diagnostic agent at the mucosal or epithelial surface for a period of at least 1 hour, preferably at least 24 hours. Preferably, the gel-forming polymer forms a uniformly thick layer upon administration to the mucosal or epithelial surface.

[0017] Exemplary mucosal or epithelial surfaces include ocular, oral, pharyngeal, esophageal, pulmonary, auricular, nasal, intraoral, lingual, vaginal, cervical, urogenital, alimentary, and anorectal surfaces. In a preferred embodiment, the mucosal or epithelial surface is the ocular surface of the eye, and the formulation maintains an effective concentration of the therapeutic, prophylactic, or diagnostic agent in the epithelial tissues inside the eye for more than one week, for example, in any one of the epithelial tissues such as the cornea, aqueous humor, sclera, conjunctiva, iris, lens, retina, and retinal pigment epithelium.

[0018] Generally, the formulation for administration is in the form of a dry powder, gel, or liquid. In some embodiments, the formulation is provided in a single dosage unit or multiple dosage units for administration.

[0019] Methods of administering the hypotonic gel-forming formulation to, or in need of, an epithelial surface, preferably a mucosal surface, are also described. The methods are particularly suitable for administration to the ocular surface of the eye to treat or prevent one or more diseases or disorders of the eye. Exemplary diseases or disorders include glaucoma, dry eye syndrome (DES), macular degeneration, diabetic retinopathy, scleroderma, and cancer. When administered to the eye, the formulation is administered as eye drops to the subject's eye, preferably in an amount of between about 10 μl and about 100 μl of the formulation, inclusive. In some embodiments, the method is repeated once or more, for example, hourly, daily, every 2, 3, 4, 5, 6 days, weekly, biweekly, or less frequently, as needed. The formulation typically forms a gel at the ocular surface having a thickness of between about 0.01 mm and 2 mm, inclusive, or between about 0.5 mm and 1.5 mm, inclusive. In some embodiments, the formulation prior to forming a gel on the ocular surface has a tonicity of between about 50 mOsm / L and about 280 mOsm / L, inclusive. In some embodiments, the gel formed from the formulation has a tonicity that approaches isotonic after water absorption has occurred and equilibrium has been re-established. [Brief description of the drawings]

[0020] [Figure 1] 1A-1B are bar graphs of the rheological properties of 16% (w / v) Pluronic® F127 with 0-1% glycerin added, showing adhesion (0-1.2 J / m2) (FIG. 1A) and maximum tack (0-1.0 N) (FIG. 1B) (n=3-6).

[0021] [Diagram 2] Figures 2A-2B are bar graphs of the rheological properties of 16% (w / v) Pluronic® F127 spiked with 0-1% PEG 300, showing adhesion (0-1.2 J / m2) (Figure 2A) and maximum tack (0-1.0 N) (Figure 2B) (n=3-6). *p<0.05.

[0022] [Diagram 3] Figures 3A-3B are bar graphs of the rheological properties of 16% (w / v) Pluronic® F127 spiked with 0-1% PEG 400, showing adhesion (0-1.2 J / m2) (Figure 3A) and maximum tack (0-0.8 N) (Figure 3B) (n=3-6). *p<0.05.

[0023] [Figure 4] 4A-4B are bar graphs of rheological properties showing adhesion (0-1.2 J / m2) (FIG. 4A) and maximum tack (0-1.0 N) (FIG. 4B) of 16% (w / v) Pluronic® F127 without additives or with the following additives: 0.4% PEG 300, 0.4% PEG 400, 0.4% PEG 400 + 0.3% polyglycerol ("PG"), dextran 70, 1% carboxymethylcellulose ("CMC"), 0.5% polyoxyethylene (20) sorbitan monooleate ("PS80"), and 0.4% PG samples (n=3-6), respectively. *p<0.05.

[0024] [Diagram 5]FIG. 5 is a line graph showing viscosity (101-105 mPa.s) over temperature (15-42° C.) for 16% Pluronic® F127 (line with open squares) and for 16% Pluronic® F127 + 0.4% PEG 400 (line with filled circles), respectively.

[0025] [Figure 6] 6A-6B are bar graphs of the force required to expel the first drop from the bottle, showing the squeezing force (0-20 N) for samples containing saline, 12% Pluronic® F127, 12% F127 + 0.3% PEG400, and 18% Pluronic® F127 at room temperature (FIG. 6A) and 33° C. (FIG. 6B), respectively. *At 33° C., 18% F127 formed a gel and was extruded as a gel band rather than a drop.

[0026] [Figure 7] FIG. 7 is a bar graph of the force required to expel the first drop from the bottle, showing the squeezing force (0-25 N) for samples including saline, 12% Pluronic® F127, 12% Pluronic® F127 + 0.3% PEG 300, 12% Pluronic® F127 + 0.3% PEG 400, BROMSITE® ("Bromfenac Ophthalmic Solution, 0.075%"), and SYSTANE® Ultra (Alcon lubricating eye drops, in which the active ingredients are polyethylene glycol 400 (0.04%) and propylene glycol (0.3%), respectively). *Bromfenac Ophthalmic Solution, 0.075% ("BROMSITE®") required significantly more squeezing force than saline (p<0.05).

[0027] [Figure 8]FIG. 8 is a bar graph of tear breakup time (TBUT) as measured in New Zealand White rabbit eyes, showing the TBUT (0-60 seconds) for each sample including saline, SYSTANE®, GENTEAL® (lubricating eye drops containing carboxymethylcellulose, dextran, glycerin, hypromellose, polyethylene glycol 400 (PEG 400), polysorbate, polyvinyl alcohol, povidone, or propylene glycol), and 12% F127 + 0.4% PEG400, respectively. n=4 different eyes per group with two independent measurements performed by a masked observer; *p<0.05 compared to saline, #p<0.05 compared to all other formulations.

[0028] [Figure 9] 9A-9H are graphs of preservative-free OcuGel (n=3) stability at room temperature for up to 6 months. (9A) pH, (9B) absorbance (AU), (9C) viscosity (mPa*s, shear rate 100 s-1), (9D) osmolality (mOsm / kg), (9E) BAK concentration (% w / v), and (9F) Poloxamer 407 molecular weight (Mw), (9G) polydispersity and (9H) % w / v measurements in OcuGel formulations.

[0029] [Figure 10] 10A-10G are graphs of OcuGel (n=3) stability at room temperature for up to 3 months. (10A) pH, (10B) absorbance (AU), (10C) viscosity (mPa*s, shear rate 100 s), (10D) osmolality (mOsm / kg), and (10E) Poloxamer 407 molecular weight (Mw), (10F) polydispersity and (10G) % w / v measurements for the above formulations.

[0030] [Figure 11]Figure 11 is a graph of fluorescein-based tear breakup time (TBUT) measured 90 minutes after administration of OcuGel (12% Poloxamer 407, 0.4% PEG400, 1 mM borate buffer, 0.01% BAK) with or without the addition of 0.2% HA at various osmolalities (72-300 mOsm / kg) in NZW rabbits compared to no treatment, saline, and SYSTANE® HYDRATION PF. *p<0.05 compared to OcuGel at 72 mOsm / kg. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0031] Detailed Description of the Invention I. Definition As generally used herein, "pharmacologically acceptable" refers to compounds, materials, compositions, and / or dosage forms that are suitable, within the scope of sound medical judgment, for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit / risk ratio.

[0032] "Biocompatible" and "biologically compatible," as used herein, generally refer to a substance that is generally non-toxic to and does not cause any significant adverse effects to a recipient, along with any metabolic or breakdown products thereof. Generally, a biocompatible substance is one that does not induce a significant inflammatory, immune, or toxic response when administered to an individual.

[0033] The terms "gel" and "hydrogel", as used herein, refer to a swollen, water-containing network of finely dispersed polymer chains that are insoluble in water, where the polymer molecules are in the external or dispersed phase and water (or an aqueous solution) forms the internal or dispersed phase. The chains can be chemically crosslinked (chemical gels) or physically crosslinked (physical gels). Chemical gels have polymer chains that are connected through covalent bonds, whereas physical gels have polymer chains linked by non-covalent bonds or cohesive forces (e.g., van der Waals interactions, ionic interactions, hydrogen bonds, or hydrophobic interactions).

[0034] The polymer chains are typically hydrophilic or contain hydrophilic polymer blocks. "Gel-forming polymer" is used to describe any biocompatible polymer, including homopolymers, copolymers, and combinations thereof, that can form a physical hydrogel in an aqueous medium when present at or above the critical gel concentration (CGC).

[0035] "Critical gel concentration" or "CGC" as used herein refers to the minimum concentration of a gel-forming polymer required for gel formation (e.g., the concentration at which a solution to gel (sol-gel) transition occurs). The critical gel concentration may depend on many factors, including the particular polymer composition, molecular weight, temperature, and / or the presence of other polymers or excipients.

[0036] The term "thermosensitive gel-forming polymer" refers to a gel-forming polymer that exhibits one or more property changes with a change in temperature. For example, some thermosensitive gel-forming polymers are water-soluble below a certain temperature, but become water-insoluble as the temperature increases. The term "lower critical solution temperature (LCST)" refers to the temperature below which the gel-forming polymer and the solvent are completely miscible and form a single phase. For example, the "LCST of a polymer solution" means that the polymer is uniformly dispersed in the solution at or below that temperature (i.e., the LCST), but aggregates and forms a second phase when the solution temperature is increased above the LCST.

[0037] "Hydrophilic", as used herein, refers to a molecule that has a greater affinity for water, and therefore a greater solubility in water, when compared to organic solvents. The hydrophilicity of a compound can be quantified by measuring its partition coefficient between water (or a buffered aqueous solution) and a water-immiscible organic solvent (e.g., octanol, ethyl acetate, methylene chloride, or methyl tert-butyl ether). A compound is considered hydrophilic if, after equilibration, there is a greater concentration of the compound in water than in the organic solvent.

[0038] "Hydrophobic" as used herein refers to a molecule that has a greater affinity for, and therefore a greater solubility in, organic solvents compared to water. The hydrophobicity of a compound can be quantified by measuring its partition coefficient between water (or a buffered aqueous solution) and a water-immiscible organic solvent (e.g., octanol, ethyl acetate, methylene chloride, or methyl tert-butyl ether). A compound is considered hydrophobic if, after equilibration, there is a greater concentration of the compound in the organic solvent than in water.

[0039] As used herein, "ocugel" refers to 12% F127, 0.4% PEG400, and 1 mM borate buffer, formulated with or without a preservative (e.g., 0.01% benzalkonium chloride ("BAK")) at either 72 mOsm or 150 mOsm.

[0040] As used herein, the term "treating" includes inhibiting, alleviating, preventing, or eliminating one or more symptoms or side effects associated with the disease, condition, or disorder being treated.

[0041] The terms "reduce", "inhibit", "alleviate" or "decrease" are used relative to a control (either no other treatment or a treatment with a known degree of effectiveness). Those skilled in the art will easily identify the appropriate control to use for each experiment. For example, the reduction in response in subjects or cells treated with a compound is compared to the response in subjects or cells not treated with the compound.

[0042] As used herein, the term "effective amount" or "therapeutically effective amount" refers to a dosage sufficient to treat, inhibit, or alleviate one or more symptoms of the disease condition being treated, or otherwise provide the desired pharmacological and / or physiological effect. The exact dosage will vary depending on various factors, such as subject-dependent variables (e.g., age, immune system health, etc.), the disease or disorder, and the treatment being administered. An effective amount can be relative to a control. Such controls are known in the art and discussed herein, and can be, for example, the state of the subject before or in the absence of administration of a drug or drug combination, or in the case of a drug combination, the effect of the combination can be compared to the effect of administration of only one of the drugs.

[0043] "Excipient" is used herein to include compounds that are not therapeutically or biologically active compounds. Thus, an excipient should be pharma- ceutically or biologically acceptable or relevant, e.g., an excipient should generally be non-toxic to the subject. "Excipient" includes a single such compound, and is intended to include multiple compounds.

[0044] The term "osmolarity" as generally used herein refers to the total number of dissolved components per liter. Osmolarity is similar to molar concentration, but includes the total number of moles of dissolved species in a solution. An osmolarity of 1 Osm / L means that there is one mole of dissolved component per liter of solution. Some solutes (e.g., ionic solutes that are dissociated in solution) contribute more than one mole of dissolved component per mole of solute in the solution. For example, NaCl contributes more than one mole of dissolved component per mole of solute in the solution. + and Cl - thus providing two moles of dissolved component per mole of dissolved NaCl in solution. Physiological osmolarity is typically in the range of about 280 to about 310 mOsm / L.

[0045] The term "tonicity" as generally used herein refers to the osmotic pressure gradient resulting from the separation of two solutions by a semipermeable membrane. In particular, tonicity is used to describe the osmotic pressure created across a cell membrane when a cell is exposed to an external solution. Solutes that can cross the cell membrane do not contribute to the final osmotic pressure gradient. Only dissolved species that do not cross the cell membrane contribute to the osmotic pressure difference, and therefore tonicity. The term "hypertonic" as generally used herein refers to a solution that has a higher concentration of solutes present inside the cell. When a cell is immersed in a hypertonic solution, water tends to flow outward from the cell to balance the concentration of solutes. The term "hypotonic" as generally used herein refers to a solution that has a lower concentration of solutes present inside the cell. When a cell is immersed in a hypotonic solution, water flows into the cell to balance the concentration of solutes. The term "isotonic," as generally used herein, refers to a solution in which the osmotic pressure gradient across the cell membrane is essentially balanced. An isotonic formulation is one that has essentially the same osmotic pressure as human blood. Isotonic formulations generally have an osmotic pressure of about 250-350 mOsm.

[0046] "Gel-forming concentration" refers to the concentration of a polymer that undergoes a phase shift from a solution to a gel. The critical gelation concentration (CGC) corresponds to the minimum gelling agent concentration required for gelation.

[0047] II. Enhanced Hypotonic Gel-Forming Compositions A hypotonic formulation of hydrogel-forming polymers, preferably including poloxamers, with improved rheological properties has been developed for enhanced delivery of therapeutic, diagnostic, prophylactic or other agents to epithelial tissues. The formulation is particularly suitable for enhanced delivery of therapeutic, diagnostic, prophylactic or other agents to the eye. Other suitable epithelial tissues include oral cavity surfaces, pharyngeal surfaces, esophageal surfaces, pulmonary surfaces, ocular surfaces, auricular surfaces, nasal surfaces, oral surfaces, lingual surfaces, vaginal surfaces, cervical surfaces, urogenital surfaces, gastrointestinal surfaces, anorectal surfaces, and / or skin surfaces. In a preferred embodiment, the epithelial tissue or surface is vaginal and rectal surfaces. In many of these tissues, epithelial cells are under a mucous membrane coating.

[0048] The polymers are administered at concentrations less than their normal critical gelling concentration (CGC). Poloxamer gel administered at concentrations equal to or greater than its CGC onto the surface of a mucosal surface such as the eye assembles into a bolus on the surface. On the ocular surface, the gel bolus is rapidly removed by blinking. In contrast, fluid from a poloxamer solution administered hypotonically forms a uniform thin gel coating over the entire ocular surface when the poloxamer is at a concentration below its CGC, thereby maintaining the drug reservoir in intimate association with the mucosal surface and enhancing and facilitating delivery of the drug to the eye. The presence of additional low molecular weight polyoxyethylene oxide polymers (e.g., PEG 300 or PEG 400) reduces adhesion to the ocular surface, improves ocular comfort and lubrication, and provides enhanced resistance to breakage by blinking and tearing. As water is absorbed into the tissue, the poloxamer concentrates and forms a gel near the epithelial tissue surface, thereby trapping the drug molecules in a sustained-release gel on the tissue surface (rather than a bolus of gel forming primarily in the lumen, as occurs with traditional thermal gelation methods where gelling polymers are administered at or above their CGC concentrations). Endogenous mucin sugar polymers affect the gelling properties of the hypotonic gelling agent, including the concentration of gelling agent required to gel and the pore structure of the resulting gel / mucin mixture. Following mucosal application, the hypotonic gelling vehicle coats the epithelium, including the inside of the fold or eyelid.

[0049] The examples clearly show that the addition of a polymer such as low molecular weight polyethylene glycol (e.g., approximately 0.1%-1.0% w / w PEG300 or PEG400) rather than high molecular weight PEG (1000 D or greater, or 2000 D or greater) reduces adhesion of the gel to the mucosal surface of the eye. The reduced adhesion prolongs the tear film breakup time (retention) of the formulation administered to the eye while maintaining viscosity, compared to a comparable gel in the absence of low molecular weight PEG. Thus, the addition of low molecular weight PEG to a hypotonic formulation of a hydrogel-forming polymer for ocular administration enhances the residence time of the resulting hydrogel at the ocular surface by providing improved flexibility and / or reduced adhesion.

[0050] Improved hypotonic formulations of hydrogel-forming polymers that can form a uniform gel coating on epithelial surfaces with reduced adhesion but that do not gel under storage conditions comprise one or more gel-forming polymers, low molecular weight PEG, and optionally one or more additional excipients and / or one or more therapeutic, prophylactic, or diagnostic agents in a hypotonic carrier.

[0051] Typically, the hydrogel forms a thin coating covering the entire ocular surface. The size and thickness of the coating depends on the amount of material applied and the size / shape of the ocular surface. In some embodiments, the hydrogel forms a thin gel on the ocular surface, which has a thickness of about 0.01 mm to about 2 mm, inclusive, or about 0.5 mm to 1.5 mm, inclusive; for example, a thickness of about 0.01 mm, 0.05 mm, 0.1 mm, 0.15 mm, 0.2 mm, 0.3 mm, 0.4 mm, 0.5 mm, 0.6 mm, 0.7 mm, 0. mm, 0.9 mm, 1.0 mm, 1.5 mm, or 2 mm, or a thickness of more than 2 mm. In some embodiments, the gel forms a film with a uniform thickness that coats the entire ocular surface. In other embodiments, the gel coats the ocular surface with a film with a non-uniform thickness, for example, a concave or convex conformation. In some embodiments, the gel film on the ocular surface prevents pathogens from contacting the ocular surface, for example, in some embodiments, the gel film prevents bacterial, viral, fungal or protozoan pathogens from contacting the eye.

[0052] A. Hydrogel-Forming Polymers The hypotonic gel-forming composition includes one or more gel-forming polymers that are utilized at a concentration below the polymer's normal critical gel concentration (CGC) (e.g., the concentration at which the polymer solution gels in a test tube when warmed to 37° C.).

[0053] Temperature-sensitive or thermoresponsive hydrogels form solutions that undergo a sol-gel transition when the following criteria are met: 1) at or above the critical gelling concentration (CGC), and 2) At or above the critical gel temperature.

[0054] Thermosensitive gelators used for biomedical applications (at or above their CGC) are liquid at room temperature but form gels at body temperature. An increase in temperature induces rearrangement and alignment of the polymer chains, resulting in gelation into a three-dimensional structure. This phenomenon is generally governed by the ratio of hydrophilic to hydrophobic portions of the polymer chain. A common feature is the presence of hydrophobic methyl, ethyl, or propyl groups.

[0055] Temperature-sensitive polymers meeting these criteria can be administered topically to mucosal and / or epithelial tissue in a hypotonic solution at a range of concentrations below their CGC to form a uniform gel coating in vitro.

[0056] Examples of temperature-sensitive gel-forming polymers that can be used include, but are not limited to, polyoxyethylene-polyoxypropylene-polyoxyethylene triblock copolymers (e.g., those designated by the CTFA designations Poloxamer 407 (CAS 9003-11-6, molecular weight 9,840-14,600 g / mol, weight percentage of polyoxyethylene approximately 70%; available from BASF as LUTROL® F127) (F127)) and Poloxamer 188 (CAS 9003-11-6, molecular weight 7680-9510 g / mol, weight percentage of polyoxyethylene approximately 80%; available from BASF as LUTROL® F68)); poloxamers are also known by the trade name Pluronic® (e.g., Pluronic® F98 (CAS 9003-11-6, molecular weight 13000 g / mol, weight percentage of polyoxyethylene approximately 80%; available from BASF); Tetronics tetrafunctional block copolymers based on ethylene oxide and propylene oxide available from BASF as TETRONIC®; poly(N,N-diethylacrylamide); poly(N,N-dimethylacrylamide); poly(N-vinylcaprolactam); poly(N-alkylacrylamide); poly(N-vinylalkylamide); poly(N-isopropylacrylamide); polyethylene oxide methacrylate polymers; poly(lactic acid-co-glycolic acid) (PLGA)-polyethylene glycol triblock copolymers (PLGA-PEG-PLGA and PEG-PLGA-PEG); polycaprolactone (PCL)-polyethylene glycol triblock copolymers (PCL-PEG-PCL and PEG-PCL-PEG); chitosan; and combinations thereof.

[0057] The hydrogels can be formed from individual gel-forming polymers or as a combination of gel-forming agents. For example, a poloxamer and another gel-forming polymer (e.g., a tetronic polymer) can be used in combination to achieve desired characteristics. Furthermore, different forms of the same gel-forming agent (e.g., poloxamer 188 and poloxamer 407) can be combined to achieve desired characteristics.

[0058] The polymer is provided at a concentration lower than that in an aqueous solution that forms a gel in a test tube when heated to 37°C. The concentration must be high enough for equilibrium, but below the CGC, to absorb enough fluid for the CGC to be achieved in vivo. Thus, gelation may occur preferentially on / near mucosal epithelial surfaces, e.g., the surface of the eye. The range of time it takes for gelation to occur depends on the mucosal surface (capacity and rate of water absorption), the tonicity of the administered solution (a more hypotonic solution drives more rapid fluid absorption), and the concentration of the administered polymer (if the polymer concentration is too low, not enough fluid absorption occurs to concentrate the polymer into its CGC). Gelation generally occurs within 1-20 seconds (inclusive) upon administration, e.g., as an eye drop, onto the surface of the eye. For example, in some embodiments, gelation occurs within less than 1 second, or within 1-2 seconds, 2-3 seconds, 3-4 seconds, 4-5 seconds, 5-6 seconds, 6-7 seconds, 7-8 seconds, 8-9 seconds, 9-10 seconds, or within less than 5 seconds, or within less than 10 seconds after administration onto the ocular surface.

[0059] The concentration of the polymer and the presence of additional components (e.g., endogenous mucin) affect the coverage and the rate and extent of gelation. Previous studies have shown that 18% Pluronic® F127 gel mixed with purified porcine gastric mucin (1%) or human cervical mucus (1:1 ratio) does not capture virus-sized (approximately 100 nm) nanoparticles (polystyrene nanoparticles coated with polyethylene glycol, PSPEG) as effectively as 18% F127 gel alone. In contrast, 24% F98 gel captured PSPEG particles more effectively when mixed with mucin or human cervical mucus. However, in vivo virus capture with hypotonic gelators was more effective at capturing viruses, including human immunodeficiency virus (HIV, approximately 120 nm) and herpes simplex virus (HSV, approximately 180 nm). Administration of a hypotonic solution containing 18% Pluronic® F98 and 24% CGC results in effective capture of subsequently administered HIV in the vagina. Similarly, hypotonic solutions containing 10% and 15% F127 and 18% CGC were also effective in reducing the MSD of HIV, including capture. Furthermore, both 15% Pluronic® F127 and 18% Pluronic® F98 reduced the spread of subsequently administered HSV in mouse vaginal mucus. The distribution of individual virus MSDs on a 1 second timescale demonstrated that viral capture (shift to the left) was more uniform in the gel formed by the hypotonic 18% Pluronic® F98 vehicle compared to 15% Pluronic® F127. Further testing of virus capture by hypotonic gelling agents in the colorectum found that 12% Pluronic® F98 (CGC 24%) did not effectively capture PSPEG nanoparticles administered 30 minutes after the gelling vehicle, while 18% Pluronic® F98 was effective in capturing PSPEG nanoparticles in the mouse colorectum. Importantly, these examples illustrate the difference in the gels that form when hypotonic gelling agents are administered to different mucosal surfaces, in this case mixing with vaginal mucus compared to colorectal mucus prior to gelling.

[0060] Thus, in some embodiments, the gel-forming polymer is present in the formulation in an amount of between 1% and 50%, e.g., between 5% and 20%, inclusive, e.g., 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19% or 20%. Typically, formulations for administration to the ocular surface contain an amount of hydrogel-forming polymer that is a concentration below the critical gel concentration (CGC) of the polymer under isotonic conditions and at temperatures between room temperature and body temperature (about 25° C. to about 37° C., inclusive).

[0061] A preferred hydrogel-forming polymer is poly(ethylene glycol)-block-poly(propylene glycol)-block-poly(ethylene glycol) (Pluronic® F127; "F127"). In some embodiments, formulations for ocular administration contain Pluronic® F127 at a concentration below the CGC, e.g., between 5% and 18%, inclusive, e.g., between about 8% and about 15%, between about 10% and about 14%, or between about 11% and about 13%, inclusive. In certain embodiments, the Pluronic® F127 is present in an amount of approximately 12%.

[0062] B. Hypotonic Carriers The gel-forming composition includes a hypotonic carrier. The hypotonic carrier is typically a biocompatible carrier that preferably causes little or no signs of irritation when administered to the eye of a human subject. The carrier may be naturally occurring or non-naturally occurring, including synthetic and semi-synthetic carriers. The preferred carrier is water-based. Other solutions, including sugar-based (e.g., glucose, mannitol) solutions and various buffers (phosphate buffer, Tris buffer, HEPES), can also be used.

[0063] When a hypotonic solution is applied to an epithelial surface (e.g., the surface of the eye), a fluid shift occurs, moving water into the epithelial tissue. This can cause the epithelial cells to swell. In some cases, if the osmotic pressure difference is too great, the epithelial cells can rupture, causing tissue irritation or destruction of the epithelial membrane.

[0064] A hypotonic solution refers to a solution that causes water absorption by the epithelial surface to which it is administered. Examples of hypotonic solutions include, but are not limited to, Tris[hydroxymethyl]-aminomethane hydrochloride (Tris HCl, 10-100 mM, pH 6-8), (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES, 10-100 mM, pH 6-8), and dilute solutions of PBS (e.g., a solution containing 0.2 g KCl, 0.2 g KH2PO4, 8 g NaCl, and 2.16 g Na2HPO4·7H2O in 1000 ml H2O).

[0065] The hypotonic carrier concentrates the dissolved gel-forming polymer on the epithelial surface, resulting in uniform gel formation on the surface. The hypotonic carrier usually contains water as a major component. The hypotonic carrier can be water, but a mixture of water and water-miscible organic solvents can also be used. Suitable water-miscible organic solvents include alcohols (e.g., ethanol, isopropanol); ketones (e.g., acetone); ethers (e.g., dioxane); and esters (e.g., ethyl acetate).

[0066] Thus, in some embodiments, the hypotonic carrier is distilled water with one or more osmolarity-modifying excipients. Sodium chloride is the excipient most frequently used to adjust osmolarity when the solution is hypotonic. Other excipients used to adjust hypotonic solutions include glucose, mannitol, glycerol, propylene glycol, and sodium sulfate. Osmolarity-modifying excipients can include pharma- ceutically acceptable salts, such as sodium chloride, sodium sulfate, potassium chloride, and other salts to make buffers, such as disodium hydrogen phosphate, potassium dihydrogen phosphate, calcium chloride, and magnesium sulfate. Other excipients used to adjust tonicity can include glucose, mannitol, glycerol, or propylene glycol.

[0067] In some embodiments, the hypotonic carrier has an osmolality lower than the effective isotonicity point (the concentration at which fluid is neither absorbed nor secreted by the underlying tissue) at the mucosal surface. The isotonicity point varies for different mucosal surfaces and different buffers depending on the active ion transport at the epithelial surface. For example, a 0.9% solution of NaCl (saline) is isotonic with blood and tears. Human tears have an osmolality in the range of 280-320 mOsm / L, while the osmolality of the aqueous layer of the precorneal tear film is approximately 300 mOsm / L. Thus, in some embodiments, the solution has a tonicity of 280 mOsm / L or less, e.g., 50 mOsm / L to 280 mOsm / L, 100 mOsm / L to 280 mOsm / L, 150 mOsm / L to 250 mOsm / L, 200 mOsm / L to 250 mOsm / L, 220 mOsm / L to 250 mOsm / L, 220 mOsm / L to 260 mOsm / L, 220 mOsm / L to 270 mOsm / L, or 220 mOsm / L to 280 mOsm / L.

[0068] The isotonicity point in the vagina for sodium-based solutions is about 300 mOsm / L, while in the colorectum it is about 450 mOsm / L. In some embodiments, the solution is hypotonic at the mucosal surface of the colorectum, having a tonicity of 400 mOsm / L or less, e.g., 50 mOsm / L to 400 mOsm / L; 350 mOsm / L or less, e.g., 50 mOsm / L to 350 mOsm / L; or 300 mOsm / L or less, e.g., 50 mOsm / L to 280 mOsm / L. In some embodiments, the solution is hypotonic at the mucosal surface of the vagina, having a tonicity of 300 mOsm / L or less, e.g., 50 mOsm / L to 280 mOsm / L.

[0069] The hypotonic carrier may include one or more pharma- ceutically acceptable acids, one or more pharma-ceutically acceptable bases, or their salts. Pharmaceutically acceptable acids include hydrobromic acid, hydrochloric acid, and sulfuric acid, and organic acids such as methanesulfonic acid, tartaric acid, and malic acid. Pharmaceutically acceptable bases include hydroxides of alkali metals such as sodium or potassium and alkaline earth metals such as calcium or magnesium, and organic bases such as pharma-ceutically acceptable amines. The hypotonic carrier may include a pharma-ceutically acceptable buffer such as a citrate buffer or a phosphate buffer.

[0070] C. Polyoxyalkylene glycol (PEO or PEG) The improved hypotonic gel-forming compositions include one or more additional PEGs as shear-thinning polymers. In a preferred embodiment, the one or more additional PEGs are low molecular weight PEGs.

[0071] Increased viscosity without shear results in increased residence time on the mucosal surface of the eye, while shear thinning upon blinking maximizes comfort and lubrication. Thus, a gelling material with these properties has advantageous lubricating properties on the ocular surface while resisting removal upon blinking. In some examples, one or more low immunogenic and biocompatible polymers are incorporated into the gelling vehicle to increase viscosity without shear and / or shear thinning upon blinking. In some examples, a shear thinning polymer in a concentration range that conventionally exhibits shear thinning properties is also included in the composition. The shear thinning polymer typically comprises a small amount (0.01% to 2%, inclusive) of a water-soluble low immunogenic and biocompatible polymer. A preferred shear thinning polymer is low molecular weight polyethylene glycol (PEG). Exemplary PEGs have a molecular weight of about 100 Da to about 10,000 Da inclusive (e.g., PEG 200, PEG 300, PEG 400, PEG 600, PEG 800, PEG 900, and PEG 1,000). In preferred embodiments, the PEG has a molecular weight of about 100 Da to about 5,000 Da inclusive; about 100 Da to about 2,000 Da inclusive; about 100 Da to about 1,000 Da inclusive; about 100 Da to about 800 Da inclusive; and about 200 Da to about 500 Da inclusive. In some embodiments, one or more PEGs are present in the improved hypotonic gel-forming compositions in an amount between 0.1% and 1% total weight:volume (w:v), inclusive, such as 0.1%, 0.2%, 0.3%, 0.4%, 0.5%, 0.6%, 0.7%, 0.8%, 0.9% and 1.0%. In a preferred embodiment, the low molecular weight PEG is present in an amount of 0.4% total weight:volume (w:v).

[0072] In some embodiments, the formulation is for use as an eye drop that is dispensed directly from a bottle or other dispenser onto the eye. Typically, a formulation for use as an eye drop contains a type and amount of lubricant that does not increase the squeezing force required for the ejection of a drop, such that the eye drop can be ejected, for example, in the form of a drop, for administration directly to the eye from a bottle of solution, using a squeezing force similar to that required to eject a similarly sized drop of saline or another isotonic solution from the same bottle. For example, in some embodiments, the squeezing force required to eject a drop is between 10 Newtons (N) and 20 N, for example, about 11 N, 12 N, 13 N, 14 N, 15 N, 16 N, 17 N, 18 N, 19 N, or about 20 N. In a preferred embodiment, the squeezing force required to eject a drop of the hypotonic gel-forming formulation from the bottle is between approximately 15 N and 17 N, inclusive.

[0073] The addition of low molecular weight PEG to the formulation also increases the tear film breakup time (TBUT) upon administration of the formulation to the ocular surface. Generally, >10 seconds is considered to be a normal TBUT in human subjects (e.g., 10 seconds, 11 seconds, 12 seconds), 5-10 seconds is marginal, and <5 seconds is considered a short time for TBUT. A short tear film breakup time is indicative of an inadequate tear film, and the longer it takes, the more stable the tear film. Thus, in some embodiments, the addition of PEG to the formulation increases the tear film breakup time (TBUT) by between 1 second (S) and 10 S, inclusive, compared to the TBUT associated with administering a commercial formulation without PEG. For example, in some embodiments, the TBUT after administration in vivo is at or about 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100% greater than normal. A normal TBUT is generally >10 seconds in humans. In preferred embodiments, the TBUT is extended to be between 15-60 seconds, between 20-50 seconds, between 25-45 seconds, between 30-40 seconds (inclusive) following administration of the formulation to the eye in vivo.

[0074] D. Additional Medications Typically, the hypotonic gel-forming composition includes one or more agents to be delivered to the eye. Examples include therapeutic agents, prophylactic agents, and / or diagnostic agents. Biologically active agents are substances used for treatment (e.g., therapeutic agents), prevention (e.g., prophylactic agents), diagnosis (e.g., diagnostic agents), or to effect a cure or alleviation of a disease or illness, or to alter the structure or function of the body, or prodrugs (which become biologically active or more active after being placed in a given physiological environment). These can be small molecule drugs (e.g., molecular weights less than 2000, 1500, 1000, 750, or 500 atomic mass units (amu)), peptides or proteins, sugars or polysaccharides, nucleotides or oligonucleotides (e.g., aptamers, siRNAs, and miRNAs), lipids, glycoproteins, lipoproteins, or combinations thereof.

[0075] The agents may include one or more of those listed in Martindale: The Complete Drug Reference, 37th Edition (Pharmaceutical Press, London, 2011).

[0076] In one embodiment, the drug to be delivered is poorly soluble in water but soluble in a carrier that includes a gelling polymer. In another embodiment, the drug is water-soluble. Data also show that water-soluble drugs, such as brimonidine tartrate, which is soluble up to approximately 2-3 mg / mL, can be beneficial.

[0077] The hypotonic gel-forming formulation may include a therapeutically effective amount of a therapeutic agent to treat, inhibit, or alleviate one or more symptoms of the disease state being treated. The hypotonic gel-forming composition may include an effective amount of a prophylactic agent to prevent one or more symptoms of a disease or disorder.

[0078] 1. Therapeutic Agents In some embodiments, the formulation comprises one or more therapeutic agents. The therapeutic agents to be delivered may include anti-infective agents (antibiotics, antiviral agents, antifungal agents), agents for treating eye disorders (glaucoma, dry eye), anti-inflammatory agents, inhibitors of neovascularization and / or fibrosis, neuroactive agents, or chemotherapeutic agents for treating diseases (e.g., abnormal neovascularization or cancer). Exemplary agents include brinzolamide, cyclosporine A, brimonidine tartrate, moxifloxacin, budesonide, sunitinib, and acriflavine.

[0079] In some embodiments, the formulation comprises one or more protein therapeutics. In a preferred embodiment, the formulation comprises one or more short therapeutic peptides. Examples of useful proteins include hormones (e.g., insulin, growth hormones (including somatomedins), and reproductive hormones. Examples of other useful drugs include neurotransmitters (e.g., L-DOPA), antihypertensives or saluretic diuretics, e.g., metolazone from Searle Pharmaceuticals, carbonic anhydrase inhibitors, e.g., acetazolamide from Lederle Pharmaceuticals, insulin-like drugs such as glyburide, blood glucose lowering drugs of the sulfonylurea class, synthetic hormones (e.g., Android F and ICN from Brown Pharmaceuticals, Representative antiproliferative agents include TESTRED® (methyltestosterone) from SymBio Pharmaceuticals. Representative antiproliferative agents include angiogenesis inhibitors, anti-VEGF compounds; receptor tyrosine kinase (RTK) inhibitors such as sunitinib (Sutent®); tyrosine kinase inhibitors such as sorafenib (Nexavar®), erlotinib (Tarceva®), pazopanib, axitinib, and lapatinib; transforming growth factor-α or transforming growth factor-β inhibitors. In one embodiment, the active agent is sunitinib.

[0080] In some embodiments, the drug is a drug for the treatment or prevention of retinal diseases (e.g., degenerative retinal diseases). Exemplary drug types include antioxidant molecules that remove and prevent oxidative and nitrosative damage, anti-infective agents, corticosteroids, analgesics, and nutritional supplements. In a preferred embodiment, the drug is a small molecule drug for treating macular degeneration. Exemplary drugs include xanthophyll (lutein), verteporfin (visudyne), natamycin (natacin), sulfacetamide ophthalmic (Bleph-10), pegaptanib (macugen), cephalosporins (ceftriaxone), and corticotropins.

[0081] 2. Diagnostic Agents In some embodiments, the preparation comprises a diagnostic agent. These agents can also be used prophylactically. Examples of diagnostic agents include paramagnetic molecules, fluorescent compounds, magnetic molecules, and radionuclides, x-ray imaging agents, and contrast agents. Examples of other suitable contrast agents include radiopaque gases or gas-emitting compounds.

[0082] Exemplary diagnostic agents include dyes, such as fluorescent dyes and near infrared dyes, SPECT imaging agents, PET imaging agents and radioisotopes.

[0083] III. METHODS AND PREPARATIONS OF IMPROVED HYPOTNIC GEL-FORMING COMPOSITIONS Hypotonic gel-forming compositions containing low molecular weight PEG can be prepared as liquids for administration onto mucosal and / or epithelial surfaces. Such formulations are particularly suitable for enhanced delivery of therapeutic, diagnostic, prophylactic, or other agents to the eye.

[0084] Drug solubilization offers potential advantages including enhanced physical stability during storage, increased drug permeation into the body, and more reproducible drug dosing when administered to patients.

[0085] Epithelial tissue or surface

[0086] Water-insoluble drugs are particularly difficult to deliver to mucosal surfaces (such as the surface of the eye) due to lack of absorption. The above formulation is particularly suitable for delivering therapeutic agents that are poorly soluble in water. Water-soluble drugs are also difficult to deliver to mucosal surfaces in a sustained manner. Therefore, gelling substances can also be used as vehicles to improve mucosal delivery of water-soluble drugs, for example, by providing more sustained drug absorption, which can reduce side effects and provide longer-term effectiveness.

[0087] Furthermore, mucosal surfaces are rapidly cleared and regenerated as a normal defense mechanism against infection and foreign particulate matter. Improved drug solubilization can improve mucosal penetration, and improving the distribution, penetration, and retention of hydrophobic drugs is a promising strategy for improving therapeutic efficacy. Methods for solubilizing various drugs and drug complexes with low water solubility into materials that also have temperature-sensitive gelation properties are described in U.S. Publication Nos. 2021 / 0196837 and 2021 / 0177751.

[0088] In some embodiments, the formulations can be prepared as liquids for administration onto the ocular surface. Gel-forming liquids or polymers solubilize insoluble drugs by forming micelles. Powders can be made by lyophilization and reconstituted at the time of use.

[0089] The formulation may also include pharma- ceutically acceptable diluents, preservatives, solubilizers, stabilizers, emulsifiers, adjuvants, and / or carriers. Stabilizers (e.g., SPAN® 20 (sorbitan laurate, CAS number 1338-39-2) may promote dissolution and prevent reaggregation. Other exemplary stabilizers include polysorbates or TWEENS®, such as polysorbate 20, polysorbate 60, polysorbate 65, and polysorbate 80, and polyglycerol esters (PGE), polyoxyethylene alkyl ethers, polyoxyl stearates, fatty acids (e.g., oleic acid), and propylene glycol monostearate (PGMS). In some cases, the composition includes one or more stabilizers.

[0090] Dosage formulations are typically prepared as single or multiple liquid or dry dosage units in a suitable applicator (e.g., eye drop dispenser). Those skilled in the art are aware of many options for drug storage and application, such as dual chamber devices that can be used to keep various components separate during storage. Multiple dosage units typically include a barrel loaded with the powder and a plunger with dosage increments thereon. These are typically sterilized and packaged in sealed, sterile packaging for storage and distribution. See also Remington: The Science and Practice of Pharmacy, 22nd Edition.

[0091] The dosage unit administrator is designed to fit the anatomical location where the drug is to be delivered (e.g., intraocular administration with one or more eye drops). In an exemplary embodiment, the formulation is a liquid having a total volume of between about 0.01 ml and about 2.5 ml, inclusive, to expel each drop (e.g., 50 μl / drop) of between about 10 μl and about 200 μl, inclusive.

[0092] IV. Methods of Using the Improved Hypotonic Gel-Forming Compositions The hypotonic gel-forming composition can in principle be applied to any water-absorbing surface, including the ocular surface, and other mucosal tissues to form a gel. Preferably, the formulation is applied as a liquid to a mucosal coating on an epithelial surface (e.g., the ocular surface) of a subject in need of a therapeutic, prophylactic, diagnostic, or nutritional effect. The gel-forming composition can be applied to an epithelial surface (e.g., the ocular surface) in any of a number of ways known to those skilled in the art, so long as the hypotonic solution, or the reagents that form the hypotonic solution, contact the surface. Suitable epithelial surfaces include ocular surfaces, oral surfaces, pharyngeal surfaces, esophageal surfaces, pulmonary surfaces, aural surfaces, nasal surfaces, intraoral surfaces, lingual surfaces, vaginal surfaces, cervical surfaces, urogenital surfaces, gastrointestinal surfaces, anorectal surfaces, and / or skin surfaces.

[0093] By applying the gel-forming composition as a hypotonic formulation, water is absorbed into the epithelial tissue. Water absorption provides concentration of the gel-forming polymer at the surface, resulting in uniform gel formation at the ocular surface. The gel can act as a barrier, a reservoir, or a combination thereof. Drugs or excipients in the gel-forming composition can be trapped in the gel and released at or into the ocular surface beneath the gel.

[0094] Exemplary epithelial surfaces to which the compositions may be applied include ocular surfaces, as well as oral, pharyngeal, esophageal, pulmonary, auricular, nasal, intraoral, lingual, vaginal, cervical, urogenital, gastrointestinal, anorectal, and / or dermal surfaces.

[0095] In some examples, the hypotonic gel-forming composition maintains an effective concentration of one or more active agents at or near the application site for an extended period of time, e.g., greater than 20 seconds, greater than 30 seconds, 40 seconds, 50 seconds, 60 seconds, 70 seconds, 80 seconds, 90 seconds, 100 seconds, or greater than 100 seconds, e.g., 2 minutes, 3 minutes, 4 minutes, 5 minutes, 6 minutes, 7 minutes, 8 minutes, 9 minutes, or 10 minutes, up to or greater than 1 hour, e.g., 2 hours, greater than 2 hours, greater than 1 day, greater than 2 days, greater than 3 days, greater than 4 days, greater than 5 days, greater than 6 days, or greater than 1 week. In some embodiments, the extended intraocular residence time of one or more active agents is around or within one of many cell types in the eye, at or near the application site. In one embodiment, the extended intraocular residence time of one or more active agents is about 1 week.

[0096] In some cases, the hypotonic gel-forming compositions increase the concentration of one or more active agents at or near the site of application by 2-fold, 3-fold, 4-fold, 5-fold, 6-fold, 7-fold, 8-fold, 9-fold, 10-fold, 20-fold, 30-fold, 50-fold, 100-fold, or more than 100-fold, compared to the active agents delivered without a gel-forming vehicle (e.g., in a saline solution).

[0097] A. Method of Administration When the hypotonic gel-forming composition is applied to the eye, mucosal sites having an increased concentration of active agent include one or more of the cornea, aqueous humor, sclera, conjunctiva, iris, lens, retina, and retinal pigment epithelium.

[0098] The method can be used to treat or prevent one or more diseases or disorders, or treat or prevent one or more symptoms of one or more diseases or disorders in a subject in need.In some embodiments, the method delivers an effective amount of agent to achieve a desired physiological goal or change in a subject.Exemplary physiological changes include the fluctuation of the amount of one or more biomarkers in a subject, for example, in the eye of a subject.The change and / or the desired outcome of treatment can be monitored to evaluate the effectiveness of treatment and to determine the amount and degree of treatment required at any given point.

[0099] In some embodiments, the improved hypotonic gel-forming compositions deliver active agents (e.g., acriflavine and sunitinib malate) to the retina and / or choroid in an amount effective to reduce retinal and / or choroidal neovascularization by 10%, 20%, 30%, 40%, 50%, or greater than 50% compared to the active agents delivered without a gel-forming vehicle (e.g., in a saline solution).

[0100] In other examples, the hypotonic gel-forming compositions deliver an active neuroprotective agent (e.g., sunitinib malate) to the retina in an amount effective to increase survival of retinal ganglion cells following optic nerve injury and / or increase expression of gamma-synuclein and / or betaIII tubulin in retinal ganglion cells following optic nerve damage by 2-fold, 3-fold, 4-fold, 5-fold, or more than 5-fold compared to the active agent delivered without a gel-forming vehicle (e.g., in a saline solution).

[0101] In further examples, the hypotonic gel-forming compositions deliver an active agent (e.g., brinzolamide) to the eye in an amount effective to reduce intraocular pressure (IOP) by 10%, 20%, 30%, 40%, 50%, or more than 50% in less than 2 hours, less than 4 hours, less than 6 hours, less than 8 hours, less than 10 hours, less than 12 hours, or less than 24 hours compared to delivery without a gel-forming vehicle (e.g., in a saline solution).

[0102] In other examples, the hypotonic gel-forming compositions deliver an active agent (e.g., cyclosporine A) to the eye in an amount effective to increase tear production by 10%, 20%, 30%, 40%, 50%, or more than 50% within less than 2 hours, less than 4 hours, less than 6 hours, less than 8 hours, less than 10 hours, less than 12 hours, or less than 24 hours compared to delivery without a gel-forming vehicle (e.g., in a saline solution).

[0103] In some embodiments, the formulation of the hypotonic gel-forming composition comprising the therapeutic agent, preventive agent, nutritional supplement or diagnostic agent is administered to the eye of a subject as an eye drop.In some embodiments, the administration to the eye is repeated one or more times as part of a treatment regimen, for example, to provide a sufficient concentration of the drug in the eye.Thus, in some embodiments, the administration to the eye is repeated at a time selected from every hour, every day, every 2 days, every 3 days, every 4 days, every 5 days, every 6 days, every week, every 2 weeks, or less frequently.

[0104] B. The disease to be treated In some embodiments, the hypotonic gel-forming composition is administered to one or more mucosal epithelial surfaces for delivery of one or more active agents across the epithelium for treatment or prevention of one or more diseases or disorders. Typically, the hypotonic gel-forming composition comprising one or more active agents is administered to the ocular surface of a subject having a disease or disorder to form a hydrogel film across the ocular surface for delivery of one or more active agents to the eye for treatment of the disease or disorder.

[0105] In some embodiments, the method treats or prevents a disease or disorder including glaucoma, dry eye syndrome (DES), macular degeneration, diabetic retinopathy, scleroderma, and cancer. A preferred method treats or prevents macular degeneration in the eye of a subject.

[0106] 1. Macular degeneration In some embodiments, the hypotonic gel-forming composition is administered to the ocular surface of a subject for the treatment of macular degeneration in the eye of a subject.

[0107] Age-related macular degeneration (AMD) is a common condition that affects the central field of vision. It usually first affects people in their 50s and 60s or older. There are two basic types of macular degeneration: "dry" and "wet." Approximately 85%-90% of cases of macular degeneration are "dry" (atrophic), while 10-15% are "wet" (exudative).

[0108] In some embodiments, the hypotonic gel-forming composition is administered to the ocular surface of a subject for the treatment of dry macular degeneration in the eye of the subject.Dry macular degeneration is a common eye disorder among people over 50 years old.It causes blurred or reduced central vision due to the thinning of the macula.The macula is the part of the retina that is responsible for clear vision in direct line of sight.

[0109] Treatment of early dry AMD is generally nutritional therapy with a healthy diet rich in antioxidants to support macular cells.When AMD progresses further but is still dry, supplements are prescribed to add higher amounts of certain vitamins and minerals that can increase healthy pigments and support cell structure.Therefore, in some embodiments, the hypotonic gel-forming composition that comprises one or more nutritional supplements is administered to the ocular surface of a subject for the treatment of dry macular degeneration in the eye of the subject.

[0110] In other embodiments, the hypotonic gel-forming composition is administered to the ocular surface of a subject for the treatment of wet macular degeneration in the subject's eye. In the "wet" form of macular degeneration, abnormal blood vessels (known as choroidal neovascularization or CNV) grow under the retina and macula. These new blood vessels can then bleed and leak fluid, causing the macula to bulge or lift from its normally flat position, distorting or destroying central vision. Under these circumstances, vision loss can be rapid and severe. VEGF is an acronym. Currently, the most common and effective clinical treatment for wet age-related macular degeneration is anti-vascular endothelial growth factor (anti-VEGF therapy), which uses an inhibitor of VEGF (anti-VEGF). VEGF is a molecule that supports the growth of new blood vessels. In the case of wet AMD, VEGF promotes the growth of new weak blood vessels in the choroid layer behind the retina, and these blood vessels leak blood, lipids, and serum into the retina layer. This leakage (bleeding) causes scarring in the retina and kills macular cells, including photoreceptor rods and cones. Thus, in some embodiments, the hypotonic gel-forming composition comprising one or more anti-VEGF agents is administered to the surface of the eye of a subject to treat wet macular degeneration in the eye of the subject.

[0111] In other embodiments, the hypotonic gel-forming composition is administered to the ocular surface of a subject for the treatment of Stargardt Disease, a form of macular degeneration found in young people caused by a recessive gene.

[0112] 2. Dry Eye Syndrome (DES) In some embodiments, the hypotonic gel-forming composition is administered to the ocular surface of a subject for the treatment of dry eye syndrome (DES) in the eye of a subject.

[0113] Dry eye syndrome (DES), also known as keratoconjunctivitis sicca (KCS), or dry eye disease, is a common condition that occurs when the eyes do not make enough tears or the tears evaporate too quickly. Other associated symptoms include irritation, redness, discharge, and easily fatigued eyes. Blurred vision may also occur. Symptoms range from mild and occasional to severe and persistent. Corneal scarring can occur if left untreated. The cornea comprises the clear outer dome of the eye that captures light and focuses it through the lens onto the retina. The cornea is an avascular tissue and receives most of its nutrients from tears, air, and fluids inside the eye.

[0114] Dry eye occurs when the eye does not produce enough tears or when tears evaporate too quickly. It can result from contact lens use, meibomian gland dysfunction, pregnancy, Sjogren's syndrome, vitamin A deficiency, omega-3 fatty acid deficiency, LASIK surgery, and certain medications (e.g., antihistamines), certain blood pressure medications, hormone replacement therapy, and antidepressants. Chronic conjunctivitis (e.g., from exposure to cigarette smoke or infection) can also result in the condition. Diagnosis is mostly based on the symptoms, but many other tests can be used.

[0115] Treatment options include drugs that reduce eyelid inflammation (e.g., antibiotics), drugs that control corneal inflammation (e.g., the immunosuppressant cyclosporine (Restasis) or corticosteroids), or tear-stimulating drugs (e.g., cholinergic agents (pilocarpine, cevimeline)). Thus, in some embodiments, the hypotonic gel-forming compositions comprising one or more antibiotics, anti-inflammatory agents, or cholinergic agents are administered to the ocular surface of a subject for treatment of DES in the subject's eye.

[0116] The invention will be further understood by reference to the following non-limiting examples. EXAMPLES

[0117] Example 1: Addition of PEG to a gel-forming eye drop formulation increases tear breakup time Methods and Materials material Endotoxin-free ultrapure water, poly(ethylene glycol)-block-poly(propylene glycol)-block-poly(ethylene glycol) (F127), sodium carboxymethylcellulose (CMC) USP reference standard, sodium fluorescein, polyethylene glycol 400 (PEG400) USP reference standard, polyethylene glycol 300 (PEG300) USP reference standard (PEG400), propylene glycol (PG), and polysorbate 80 (PS80) were purchased from Sigma Aldrich. Saline (0.9% NaCl), Systane® ULTRA eye drops lubricant high performance, and GenTeal® Tears Lubricant eye ointment were purchased from Alcon.

[0118] Rheology Samples For sample preparation for rheological testing, 16% (w / v) F127 and other additives (0.025-1% PEG300, PEG400, and glycerin; 0.4% PG, 0.5% PS80, 1% CMC) were mixed together and mixed with purified water prior to reconstitution. Note that 16% F127 was used for in vitro testing to allow characterization in the gel state, whereas in vivo testing used a hypotonic 12% gel-forming formulation.

[0119] Tear Break-Up Time (TBUT) Sample For tear film breakup time (TBUT) testing, 5 mg / mL sodium fluorescein powder was added to the various formulations (saline, GenTeal® Tears, Systane® ULTRA, with 12% F127, 0.4% PEG400) before mixing. All formulations were freshly prepared for each experiment by mixing for 3 days on a Benchmark Scientific waving rocker (30° inclination angle and 30 rpm speed, 4° C.). All formulations were kept packaged to avoid exposure to light and stored at 4° C. when not in use.

[0120] Eye drop bottle strength measurement The formulations were prepared in 7 mL volumes and transferred to 10 mL eye dropper bottles (Steri-Dropper). An FG-3005 Digital Force Gauge system (Nidac) with 50 N capacity was used to measure the bottle squeeze force. The force gauge was attached to a syringe pump (NE300), which provided a constant forward displacement to squeeze the bottle. The force probe was placed vertically 1.5 cm from the bottom of the inverted bottle, which was loosely fixed by a metal clamp. The force probe was moved at a constant speed of 1 mm / s to squeeze the bottle. The force was recorded when the first drop of liquid was dispensed from the tip of the bottle. For measurements taken at 33° C. and 37° C., the entire system was placed in a heated oven and allowed to equilibrate for 15 minutes before measurements were taken.

[0121] Rheological Testing Anton Paar cone and plate rheometer (model MCR 302) with a PP25 probe was used to measure maximum adhesion and adhesion. The probe was placed in contact with the sample and then moved perpendicularly away from the sample at a controlled rate to measure normal force. The peak force on the curve was considered the maximum adhesion, while adhesion is the area under the force curve. Samples (200 μL) were transferred to the rheometer sample holder using a Drummond Wiretrol 200 μL wire plunger pipette. Samples were equilibrated at the specified temperature for 60 seconds with a 1 mm gap between the probe and the sample holder. A total of 100 points were measured in linear gradient mode at 60 second intervals (initial set as -0.005 mm / s and final set as -0.5 mm / s). Unless specified, n=3-6 were measured for each sample group, and each sample was prepared individually. Final data was exported from Antron Parr RheoCompass. Maximum adhesive force and adhesion were calculated with GraphPad Prism 9. For temperature gradient viscosity experiments, the loading gap was set to 0.3 mm. A total of 26 points with a temperature change of 0.25 min (linear gradient mode) were selected with the additional parameters gamma = 1 (1 / s) and a temperature gradient from 15 to 40 °C (n = 3). Raw data including temperature (°C) and viscosity (mPa*s) measurements were exported from the Anton Parr RheoCompass software and analyzed / plotted with GraphPad Prism 9. Statistical analysis of two groups was performed using a two-tailed Student's t-test. For comparison of multiple groups, one-way ANOVA with Dunnett's multiple comparison test was used. Statistical analysis was performed using GraphPad Prism 9.

[0122] Tear breakup time (TBUT) Pluronic® F127 was hypotonically formulated at 12% w / v with 0.4% PEG400. Several commercially available eye drops (including saline and lubricating eye drops from the SYSTANE and GENTEAL product lines), as well as 12% F127 with 0.4% PEG400, had 0.5 mg / mL sodium fluorescein added for visualization purposes. Tear breakup time (TBUT) was performed in healthy New Zealand White rabbits (n=4 different eyes per group, with two independent measurements performed by a blinded observer). Eye drops containing fluorescein were administered in a 50 μL volume to conscious rabbits gently wrapped in a towel. After administration, the eyelids were manually closed twice to allow for distribution of the tear film and drug. The eyes were then gently held open and TBUT was measured and recorded. TBUT was scored by two individuals who agreed and timed in a blinded manner without knowing the identity of the formulation. As shown, the hypotonic 12% F127 with 0.4% PEG400 clearly showed a significant increase in TBUT compared to the best performing commercial lubricant eye drop formulation (*p<0.05). For multiple group comparison, one-way ANOVA and Dunnett's multiple comparison test were used. Statistical analysis was performed using GraphPad Prism 9.

[0123] result Effect of glycerin on maximum adhesive strength Pluronic® F127 was formulated at 16% w / v to be held above the critical gel concentration (CGC) to facilitate in vitro gelation at 37° C. and evaluation of gel rheological properties. Various amounts of glycerin (0.2-1%, 0% represents F127 with no additive) were added to characterize the effect on (FIG. 1A) adhesion and (FIG. 1B) maximum adhesive force of F127 gels (n=3-6). The addition of glycerin had no measurable effect.

[0124] Effect of PEG300 on maximum adhesion Pluronic® F127 was formulated at 16% w / v to be held above the critical gel concentration (CGC) to facilitate in vitro gelation at 37°C and evaluation of the rheological properties of the gels. Various amounts of PEG300 (0.2-1%, 0% represents F127 with no additive) were added to characterize the effect on (Figure 2A) adhesion and (Figure 2B) maximum adhesive force of F127 gels (n=3-6). Maximum adhesive force was significantly reduced with the addition of 0.4-1% PEG300.

[0125] Effect of PEG400 on maximum adhesion Pluronic® F127 was formulated at 16% w / v to be held above the critical gel concentration (CGC) to facilitate in vitro gelation at 37°C and evaluation of gel rheological properties. Various amounts of PEG400 (0.025-0.1%, 0% represents F127 with no additive) were added to characterize the effect on (Figure 3A) adhesion and (Figure 3B) maximum adhesive force of F127 gels (n=3-6). Maximum adhesive force was significantly reduced with the addition of 0.1%-1% PEG400.

[0126] Effect of PEG300 and PEG400 on maximum adhesion force Pluronic® F127 was formulated at 16% w / v to be held above the critical gel concentration (CGC) to facilitate in vitro gelation at 37° C. and evaluation of gel rheological properties. Various lubricants and demulcents were added to characterize the effect on (FIG. 4A) adhesion and (FIG. 4B) maximum adhesion of F127 gels (n=3-6). The addition of PEG300 and PEG400 reduced maximum adhesion, with the most pronounced effect observed with PEG400.

[0127] Viscosity change Pluronic® F127 was formulated at 16% w / v to be held above the critical gel concentration (CGC) to facilitate evaluation of in vitro gelation and gel rheological properties. A constant stress temperature gradient was performed over 15-40 °C to evaluate the change in viscosity with temperature with and without the addition of 0.4% PEG400. The viscosity profile as a function of temperature was not significantly affected by the presence of 0.4% PEG400 (Figure 5). Thus, the reduction in maximum adhesion with the addition of PEG400 does not affect the thermoreversible gelation behavior of F127.

[0128] Effect on squeezing force Saline, 12% F127, 12% F127 with 0.3% PEG400, and 18% F127 were added to a standard eye dropper bottle to measure the squeezing force required to expel the first drop at (Figure 6A) room temperature or (Figure 6B) 33°C, which mimics storage in a clothing pocket. *At 33°C, 18% F127 formed a gel and was extruded as a gel band rather than a droplet. In contrast, formulation with 12% F127 (below the CGC) prevented premature gelation.

[0129] Saline, 12% F127, 12% F127 with 0.3% PEG300, 12% F127 with 0.3% PEG400, BromSite, and Systane ULTRA were added to a standard eye dropper bottle and the squeezing force required to expel the first drop at room temperature was measured (Figure 7). Only BromSite required significantly more squeezing force than saline (p<0.05), whereas the F127 formulation was indistinguishable from saline.

[0130] Effect of PEG400 on tear film breakup time Pluronic® F127 was hypotonically formulated at 12% w / v (a concentration below the critical gel concentration (CGC) shown to provide a thin uniform gel layer in vivo (but not form a gel in vitro). Several commercial eye drops (including saline, and lubricating eye drops from the Systane and GenTeal product lines), as well as 12% F127 with 0.4% PEG400, had 0.5 mg / mL sodium fluorescein added for visualization purposes. Tear film breakup time (TBUT) was performed in healthy New Zealand White rabbits (n=4 different eyes per group with two independent measurements by a blinded observer). As shown in FIG. 8, hypotonic 12% F127 with 0.4% PEG400 clearly showed a significant increase in TBUT compared to the best performing commercial lubricating eye drop formulation.

[0131] Example 2: Stability and sterility testing material and method OcuGel, containing 12% F127, 0.4% PEG400, and 1 mM borate buffer, was formulated at either 72 mOsm or 150 mOsm, with or without 0.01% benzalkonium chloride ("BAK") For pH measurements, 300 μL of each sample was transferred to a 1.5 mL Eppendorf® and measured with a Mettler Toledo EL20 pH meter with a microelectrode.

[0132] For absorbance measurements, 200 μL of each sample was added to a Nunc TM MicroWell TMTransferred to a 96-well microplate and absorbance at λ=490 nm was measured with an absorbance plate reader (Synergy MTX reader). Sample viscosity (mPa*s) was measured using a Modular Compact Rheometer (MCR302) with a PP25 parallel plate measurement system (Anton Parr). 200 μL of sample was dispersed on a 37° C. platform using a Drummond WIRETROL® II 200 μL with a wire plunger. The location of the PP25 probe was set at 0.5 mm with 1 minute of temperature equilibration between the formulation and the probe. A linear gradient mode with an initial 1 (1 / s) and an end 100 (1 / s) was set with a total of 10 measurement points (each with a measurement period of 10 seconds). The viscosity value (mPa*s) at 100 (1 / s) was then reported at each time point for stability comparison.

[0133] For osmolality measurements, a Vapro osmometer system (ELITechGroup) was used to measure the osmolality of the samples.

[0134] BAK concentration, when present in the formulation, was measured by high performance liquid chromatography (HPLC, Prominence LC2030, Shimadzu) and a LUNA® 5 μm C18 100 Å, 00G-422-E0 column (Phenomenex). Acetonitrile and water were used as the mobile phase in a ratio of 75:25. Samples were eluted isocratically through a C18 reversed-phase column at a flow rate of 1 mL / min at 40° C. UV absorbance was monitored at 205 nm and the area under the curve (AUC) was used to calculate the BAK concentration.

[0135] To measure F127 molecular weight (Mw), 100 μL of the above formulation was frozen at −80° C. and lyophilized. DMF with 0.1% LiBr solution (1.2 mL) was added to the sample and vortexed to dissolve. A gel permeation chromatography system (1260 Infinity II series, Agilent Technologies) with an organic column (Agilent, 10 aem MIXED-B column) was used to separate and detect F127 at a flow rate of 1 mL / min and a pressure set around 33 bar. Mw and polydispersity (PD) values ​​were calculated using Alginate software (version 1.4).

[0136] OcuGel with and without preservatives was formulated in small batches, sterile filtered, and aliquoted into sterile eye drop bottles in a biosafety cabinet. The eye drop bottles were capped, wrapped in heat shrink wrap sealer, and stored in a biosafety cabinet at room temperature. Ten bottles per time point (0, 3 months) were stored in a light-tight cabinet at room temperature. For each time point, the ten bottles were<USP 71> For pooling and sterility testing according to guidelines, Pace Analytical (登録商標) The material was shipped to Life Sciences.

[0137] Safety Testing Acute: OcuGel (12% Poloxamer 407, 0.4% PEG400, 1 mM Borate Buffer, 0.01% BAK, pH 7.4) was formulated in small batches, sterile filtered, and aliquoted into sterile eye drop bottles in a biosafety cabinet.

[0138] All animals were handled and treated in accordance with the Association for Research in Vision and Ophthalmology Statement for the Use of Animals in Ophthalmic and Vision Research. Equal numbers of male and female New Zealand White rabbits were used in the local acute toxicity studies.

[0139] For acute dosing studies, n=4 rabbits were dosed hourly with 50 μL of OcuGel in the right eye and 50 μL of SYSTANE® ULTRA in the left eye for 8 hours. After the last dose, the rabbits were anesthetized and an ocular evaluation was performed by a board-certified ophthalmologist.

[0140] Corneal neovascularization was graded on a 0-2 scale (0 indicates normal function and 2 indicates severe injury or inflammation). Pupillary light reflex, conjunctival injection, and conjunctival discharge were graded on a 0-3 scale (where 0 indicates normal function and 3 indicates severe injury or inflammation). Conjunctival swelling, corneal opacity (severity), corneal opacity (area), anterior chamber cells, iris involvement, anterior vitreous cells, eyelid discharge, eyelid swelling, eyelid vascularity, meibomian gland function, and fluorescein staining were graded on a 0-4 scale (0 indicates normal function and 4 indicates severe injury or inflammation). After the examination, rabbits were closely monitored in their home cages until they were fully awake.

[0141] Chronic: OcuGel (12% Poloxamer 407, 0.4% PEG400, 1 mM borate buffer, 0.01% BAK, pH 7.4) was formulated in small batches, sterile filtered, and aliquoted into sterile eye drop bottles in a biosafety cabinet. All animals were handled and treated in accordance with the Association for Research in Vision and Ophthalmology Statement for the Use of Animals in Ophthalmic and Vision Research. Equal numbers of male and female New Zealand White rabbits were used in local acute toxicity studies. For acute dosing studies, n=4 rabbits were administered 50 μL of OcuGel in the right eye and 50 μL of SYSTANE® ULTRA in the left eye, three times per day for 14 days. After the last dose on days 7 and 14, the rabbits were anesthetized and ocular evaluations were performed by a certified ophthalmologist. Corneal neovascularization was graded on a 0-2 scale (0 indicates normal function and 2 indicates severe injury or inflammation). Pupillary light reflex, conjunctival injection, and conjunctival secretions were graded on a 0-3 scale (where 0 indicates normal function and 3 indicates severe injury or inflammation). Conjunctival swelling, corneal opacity (severity), corneal opacity (area), anterior chamber cells, iris involvement, anterior vitreous cells, eyelid secretions, eyelid swelling, eyelid vascularity, meibomian gland function, and fluorescein staining were graded on a 0-4 scale (0 indicates normal function and 4 indicates severe injury or inflammation). After the examination, rabbits were closely monitored in their home cages until they were fully awake.

[0142] Tear film breakup time (TBUT) measurement OcuGel, containing 12% F127, 0.4% PEG400, 1 mM borate buffer, and 0.01% BAK, was formulated with or without 0.2% HA (2 MDa) at various osmolalities (100 mOsm / kg, 150 mOsm / kg, 200 mOsm / kg, 250 mOsm / kg, 300 mOsm / kg) and adjusted to pH 7-7.4. Saline and Systane® Hydration PF were used as comparators. TBUT was performed in healthy New Zealand White rabbits (n=3). The various formulations were first administered topically (50 μL). After 90 minutes, 50 μL of 2% disodium fluorescein (pH 7.4) was administered to the treated eye. After administration, the eyelids were manually closed twice to allow for distribution of the tear film and drug.The eyes were then gently held open and TBUT was measured and recorded.TBUT was scored by two individuals (n=3 independent animals) who agreed in a blinded manner and confirmed the time, without knowing the identity of the formulation.Data are mean±SD.Statistical analysis was performed by one-way ANOVA with multiple comparisons for the OcuGel 72mOsm / kg group.

[0143] result Room Temperature Stability and Sterility Testing OcuGel (12% Poloxamer 407, 0.4% PEG400, 1 mM Borate Buffer, 0.01% BAK) was formulated in small batches, sterile filtered, and aliquoted into sterile eye drop bottles in a biosafety cabinet. The eye drop bottles were capped, wrapped in heat shrink wrap sealer, and stored in a biosafety cabinet at room temperature. Three bottles per time point (0, 1 month, 3 months, 6 months) were stored in a light-tight cabinet at room temperature. For each time point, samples were characterized for pH, absorbance, viscosity, osmolality, BAK concentration, and polymer molecular weight.

[0144] 9A-9H are graphs of the stability of preservative-free OcuGel (n=3) at room temperature for up to 3 months. (9A) pH, (9B) absorbance (AU), (9C) viscosity (mPa*s, shear rate 100 s-1), (9D) osmolality (mOsm / kg), (9E) BAK concentration (% w / v), and (9F) Poloxamer 407 molecular weight (Mw), (9G) polydispersity and (9H) % w / v measurements for OcuGel formulations.

[0145] 10A-10G are graphs of the stability of OcuGel (n=3) at room temperature for up to 6 months. (10A) pH, (10B) absorbance (AU), (10C) viscosity (mPa*s, shear rate 100 s-1), (10D) osmolality (mOsm / kg), and (10E) Poloxamer 407 molecular weight (Mw), (10F) polydispersity, and (10G) % w / v measurements for the above formulations.

[0146] The pH remained within the range of 7.0–7.4 (Figures 9A, 10A) and the absorbance was 0.04–0.06. The AU remained in the range of 0.009-0.011% (w / v) (Figure 9E), the poloxamer 407 molecular weight (Mw) remained in the range of 12,000-13,000 Da (Figure 9F, 10E), the poloxamer 407 polydispersity (PD) remained in the range of 1.0-1.2 (Figure 9G, 10F), and the poloxamer 407 concentration remained in the range of 11.5-12.5% ​​(w / v) (Figure 9H, 10G).

[0147] sterility Table 1 shows the effect of OcuGel with BAK preservative at (A) 0 and (B) months.<USP 71> (C) Met the sterility requirements for 3 months and that preservative-free OcuGel<USP 71> All OcuGel formulations tested met the sterility requirements of USP <71> The product passed the sterility test according to the standard.

[0148] Preclinical Stability Studies Grading presented for the acute toxicity studies (Table 2), as well as in the chronic studies at 7 days (Table 3) and 14 days (Table 4), showed essentially no significant evidence of ocular irritation with either treatment regimen.

[0149] Tear breakup time (TBUT) OcuGel (12% poloxamer 407, 0.4% PEG400, 1 mM borate buffer, 0.01% BAK) was formulated with and without 0.2% HA at various osmolalities (72-300 mOsm / kg) and administered to New Zealand White rabbits. Fluorescein-based TBUT was assessed 90 min after administration.

[0150] Figure 11 is a graph of OcuGel (12% Poloxamer 407, 0.4% PEG400, 1 mM Borate Buffer, 0.01% BAK) administered with or without the addition of 0.2% HA at various osmolalities (72-300 mOsm / kg) to NZW rabbits, measured as fluorescein-based tear breakup time (TBUT) 90 minutes after administration, compared to no treatment, saline, and SYSTANE® HYDRATION PF. *p<0.05 compared to OcuGel at 72 mOsm / kg.

[0151] As shown in FIG. 11, TBUT returned to the baseline level of untreated rabbits in the saline and Systane® Hydration PF groups. In contrast, all OcuGel-treated animals had increased TBUT compared to untreated animals at 90 minutes, except for the OcuGel+HA 250mOsm / kg group. However, HA formulations with higher osmolality (including 250mOsm / kg and 300mOsm / kg) had reduced TBUT compared to OcuGel at 72mOsm. The data supported the long-lasting surface lubricating effect, including when additional lubricating polymers such as HA were added, and the importance of hypotonic formulations to achieve these benefits.

[0152] Tables 1-4 clearly demonstrate the sterility and safety of the above formulations.

[0153] [Table 1]

[0154] [Table 2]

[0155] [Table 3]

[0156] [Table 4]

[0157] Those skilled in the art will recognize, or be able to ascertain using no more than routine experimentation, equivalents to the specific embodiments of the invention described herein which equivalents are intended to be encompassed by the following claims.

Claims

1. 1. A formulation for delivering a therapeutic, prophylactic, diagnostic or nutritional supplement to the eye, said formulation comprising:

1. A gel-forming polymer for application to a mucosal or epithelial surface, the gel-forming polymer being formulated to be at a concentration below the critical gel concentration (CGC) of said polymer under isotonic conditions and at a temperature between room temperature and body temperature (about 25 to about 37° C.); low molecular weight polyethylene glycol (PEG) in an amount of between about 0.1% and about 1.0%, inclusive, by total weight / volume (w / v); excipients for forming a pharma- ceutically acceptable hypotonic formulation of said polymer suitable for delivery to a mucosal or epithelial surface of an individual in need thereof; Including, Optionally, including therapeutic, prophylactic, nutritional, or diagnostic agents; formulation.

2. The formulation of claim 1 , wherein the gel-forming polymer is a temperature-sensitive gel-forming polymer.

3. 3. The formulation of claim 2, wherein the temperature sensitive gel-forming polymer has a lower critical solution temperature that is less than 30°C, preferably less than 21°C.

4. The formulation of any one of claims 1 to 3, wherein the gel-forming polymer is between greater than 10% and less than 18% in the aqueous vehicle.

5. The formulation of any one of claims 1 to 4, wherein the gel-forming polymer is a poloxamer.

6. 6. The formulation of claim 5, wherein the gel-forming polymer is poly(ethylene glycol)-block-poly(propylene glycol)-block-poly(ethylene glycol) (Pluronic® F127).

7. The gel forming polymer is between 10% and 16% F127. The formulation according to any one of claims 1 to 6.

8. 8. The formulation of any one of claims 1 to 7, wherein the PEG is one or more selected from the group consisting of PEG 100, PEG 200, PEG 300, PEG 400, PEG 600, PEG 800, and PEG 1,000.

9. The formulation of any one of claims 1 to 8, wherein the PEG is PEG 400.

10. 10. The formulation of any one of claims 1 to 9, wherein the PEG is in an amount of between about 0.01% w / v and about 1% w / v, inclusive.

11. The formulation of any one of claims 1 to 10, wherein the PEG is in an amount of about 0.4% w / v.

12. A formulation according to any one of claims 1 to 11, comprising poloxamer F127, PEG 400 and borate buffer formulated with or without a preservative.

13. 13. The formulation of claim 12, comprising approximately 12% poloxamer F127, 0.4% PEG 400, and 1 mM borate buffer, formulated with or without a preservative, such as 0.01% benzalkonium chloride ("BAK"), at either 72 mOsm or 150 mOsm.

14. The formulation of any one of claims 1 to 13, wherein the formulation releases the therapeutic, prophylactic or diagnostic agent at the mucosal or epithelial surface for a period of at least 1 hour.

15. 15. The formulation of claim 14, wherein the formulation releases the therapeutic, prophylactic, or diagnostic agent at the mucosal or epithelial surface for a period of at least 24 hours.

16. The formulation of any one of claims 1 to 15, wherein the gel-forming polymer forms a uniformly thick layer upon administration onto the mucosal or epithelial surface.

18. The formulation of any one of claims 1 to 17, wherein the mucosal or epithelial surface is the ocular surface of the eye.

19. 20. The formulation of claim 18, wherein the formulation maintains an effective concentration of a therapeutic, prophylactic, or diagnostic agent in the interior epithelial tissue of the eye for greater than one week.

20. 20. The formulation of claim 19, wherein the epithelial tissue inside the eye is selected from the group consisting of the cornea, aqueous humor, sclera, conjunctiva, iris, lens, retina, and retinal pigment epithelium.

21. A formulation according to any one of claims 1 to 20 for administration in the form of a dry powder, a gel or a liquid, preferably eye drops.

22. 22. The formulation of claim 21, wherein the formulation is provided in single or multiple dosage units for administration.

23. including therapeutic, prophylactic, nutritional, or diagnostic agents; 23. The formulation of any one of claims 1 to 22, wherein the agent is a protein or peptide, a small molecule, a sugar or polysaccharide, a lipid, a glycolipid, a glycoprotein, a nucleic acid, an oligomer or polymer thereof, or a small molecule.

24. 24. The formulation of any one of claims 1 to 23, wherein the drug is selected from the group consisting of steroids, glaucoma drugs, tyrosine kinase inhibitors, immunosuppressants, antifibrotic drugs, anti-infective drugs, antioxidants that prevent oxidative and / or nitrosative damage, hormones and chemotherapeutic drugs.

25. 25. A method for administering a medicament to a mucosal or epithelial surface of a subject, said method comprising administering a formulation according to any of claims 1 to 24 to a mucosal or epithelial surface of said subject in need thereof.

26. 26. The method of claim 25, wherein the mucosal or epithelial surface is an ocular surface of the eye.

27. 27. The method of claim 26, wherein the method treats or prevents one or more diseases or disorders of the eye.

28. 28. The method of claim 27, wherein the disease or disorder is selected from the group consisting of glaucoma, dry eye syndrome (DES), macular degeneration, diabetic retinopathy, scleroderma, and cancer.

29. 29. The method of any one of claims 26 to 28, wherein the formulation is administered as eye drops to the eye of the subject, preferably wherein the formulation forms a gel on the surface of the eye with a thickness of between about 0.01 mm and 2 mm, inclusive, and preferably has a tonicity of between about 50 mOsm / L and about 280 mOsm / L, inclusive.

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