WO2025078308A1

WO2025078308A1 - Gel formulation for topical ocular use

Info

Publication number: WO2025078308A1
Application number: PCT/EP2024/078144
Authority: WO
Inventors: Per WIKSTRÖM
Original assignee: Glucox Biotech AB
Current assignee: Glucox Biotech AB
Priority date: 2023-10-10
Filing date: 2024-10-07
Publication date: 2025-04-17
Anticipated expiration: 2026-04-10

Abstract

A gel formulation for ocular topical use, comprising a compound of formula (I); (I). The formulation is useful for the treatment of ocular disorders.

Description

GEL FORMULATION FOR TOPICAL OCULAR USE

FIELD OF THE INVENTION

The present invention relates to a pharmaceutically useful formulation and the use thereof in the treatment of ocular disorders. More particularly, the invention relates to a gel formulation for topical ocular use containing a therapeutically active benzenesulfonamide derivative, and the use of such formulation in the treatment of ocular disorders.

BACKGROUND

Throughout this disclosure, various non-patent publications are referred to by first author and year of publication. Full citations for these publications are presented in a references section immediately before the claims. The disclosures of these documents and of any patent publications referred to herein are hereby incorporated in their entireties by reference.

International application No. PCT/EP2019/061950, published as WO 2019/215291 A1 , discloses certain benzenesulfonamide derivatives and their use in therapy, in particular in the treatment of conditions or disorders associated with nicotinamide adenine dinucleotide phosphate oxidase 4 or 2 (NOX4 or NOX2). The application discloses the activity of such derivatives in the modulation of various conditions affecting the eye, such as diseases involving lens epithelial-to-mesenchymal transition and retinopathy.

Oxidative stress has been shown to be involved in various ocular conditions, including conditions affecting the anterior segment and/or posterior segment of the human eye (Shu, 2023). Thus, generation of reactive oxygen species (ROS) and oxidative stress can contribute to the development and progression of eye-related disorders such as age-related macular degeneration, glaucoma, retinopathy, retinal ischemia-reperfusion injury, cataract, and corneal conditions and diseases. Oxidative stress has also been shown to be generally involved in inflammation linked to ocular diseases (Dammak, 2021), where the imbalance between the production of reactive oxygen species (ROS) and their elimination by protective mechanisms may lead to chronic inflammation.

There is ample evidence that involves the ROS-generating enzyme NOX, in particular NOX4 but also NOX2, in the pathophysiological mechanisms contributing to the progression of eye diseases. An interplay among NOX, oxidative stress, inflammation, and hypertensive ocular disease has been shown, underlining the importance of NOX in the oxidative and inflammatory pathogenesis of various eye diseases, including age-related macular degeneration, glaucoma, retinopathy, retinal ischemia-reperfusion injury, cataract, and corneal conditions and diseases and in retinal neurodegeneration.

Age-Related Macular Degeneration (AMD)

Age-related macular degeneration (AMD) is a multifactorial disease and the leading cause of irreversible blindness in the elderly population. It is generally divided into two groups according to the presence or absence of neovascularization, viz. on the one hand, dry or non-neo- vascular AMD and, on the other hand, wet or neovascular AMD (nAMD).

Oxidative stress and choroidal vascular dysfunction have been suggested to be critically involved in AMD pathogenesis, including dry AMD pathogenesis. The retina is one of the highest oxygen-consuming tissues in the human body, and local oxygen metabolic environment in the retina plays an essential role in keeping retinal homeostasis between the supply and consumption of retinal oxygen. During the retinal process of transforming light into vision, reactive oxygen species (ROS) are generated as normal metabolic byproducts. However, when the generation of ROS exceeds the capacity of the antioxidant systems, the balance of redox homeostasis is disrupted, and oxidative stress occurs. NOX4 has been found in retinal pigment epithelial (RPE) cells and has been associated with increased ROS production in these cells. Therefore, excessive ROS production by NOX4 in the RPE may contribute to the oxidative damage seen in AMD, including the formation of drusen and the degeneration of photoreceptor cells. Evidence that ROS and vascular dysfunction may contribute together to the pathology of wet AMD has also been presented and NOX4 has been demonstrated as one connection between and vascular endothelial growth factor (VEGF) and ROS in human choroidal endothelial cells (Ruan, 2021).

There is presently no effective treatment for dry AMD, though, for example, antioxidant treatment has been suggested as a protection against oxidative damage. Treatment of wet AMD, on the other hand, includes treatment by anti-VEGF drugs, generally by ocular injection. Such injections are, however, both painful and associated with potential degeneration of healthy endothelium.

Glaucoma

Glaucoma is a group of eye diseases characterized by elevated intraocular pressure and damage to the optic nerve, which, similar to AMD, can lead to vision loss. While the disease can occur at any age, it is more common in the elderly and is one of the leading causes of blindness for people over the age of 60.

There are two major types of glaucoma, viz. open-angle glaucoma (or primary open-angle glaucoma), and angle-closure glaucoma (also referred to as “closed-angle glaucoma” or “narrow-angle glaucoma”). Open-angle glaucoma is the most common type of glaucoma and occurs when deficient drainage of aqueous humor from the eye causes damage to the optic nerve due to an increase in the intraocular pressure. Angle-closure glaucoma occurs by the iris blocking the drainage angle of the eye, causing an abrupt increase of the intraocular pressure, which, if not urgently treated, may cause blindness.

Increasing evidence has been indicates that reactive oxygen species (ROS) play a key role in the pathogenesis of glaucoma, cf. for example (Izzotti, 2006) and (Fan Gaskin, 2021). Thus, it has been suggested that the vascular alterations often associated with glaucoma could contribute to the generation of oxidative damage, and oxidative stress occurring also in retinal cells has been suggested to be involved in the neuronal cell death affecting the optic nerve in, especially, open angle glaucoma. Kimura, 2017, has reported that drugs with anti- oxidative properties prevent glaucomatous retinal degeneration in mouse models of glaucoma and optic neuritis.

NOX4 has been found to be one of the main sources of ROS in glaucoma. Thus, in a study on acute ocular hypertension (AOH)-induced retinal ischemia/hypoxia in mice, NOX4 was shown to be highly expressed in the mouse retina, in particular in the retinal ganglion cell layer (GCL) (Liao, 2023). It was shown that NOX4 inhibition reduced ROS overproduction, inhibited inflammatory factor release, suppressed glial cell activation and hyperplasia, inhibited leukocyte infiltration, reduced retinal cell senescence and apoptosis in damaged areas, reduced retinal degeneration and improved retinal function. Based on the observed results, the authors concluded that targeted inhibition of NOX4 may offer a new treatment of acute glaucoma.

Retinopathy

There are 4 major types of retinopathy, viz. retinopathy of prematurity, hypertensive retinopathy, central serous retinopathy and, the most common type, diabetic retinopathy. In diabetic patients, diabetic retinopathy is a common complication of diabetes that affects the retina due to a sustained high blood sugar level. Studies have demonstrated that oxidative stress is a critical contributor to the pathogenesis of diabetic retinopathy (Kang, 2020). NOX4 has been found in the retinas of diabetic patients and in animal models of diabetes, and NOX4- generated ROS are thought to contribute to retinal vascular dysfunction and damage in diabetic retinopathy (Dionysopoulou, 2023). Therefore, NOX4 is as a potential target also for the therapy of retinopathy, in particular diabetic retinopathy. Evidence of the importance of NOX4 as a major ROS-generating enzyme system in the oxidative and inflammatory processes surrounding hypertensive eye diseases, including hypertensive retinopathy, has also presented (Santana-Garrido, 2021).

Besides hypertensive retinopathy, hypertensive eye diseases also include hypertensive choroidopathy and hypertensive optic neuropathy, for which, therefore, NOX4 inhibition has a potential usefulness.

Increasing evidence supports the role of neurodegeneration and inflammation in ischemic retinopathies (Barber, 1998; Krady, 2005; Antonetti, 2012), and causative factors implicated in their development of ischemic retinopathies, include oxidative stress and glutamate exci- totoxicity. Thus, retinal ischemia has been linked to increased levels of glutamate, which leads to over activation of glutamate receptors (Louzada-Junior, 1992; Osborne, 2004), thus making excitotoxicity a significant pathological mechanism implicated in ischemic retinal disorders. Ischemia induced release of glutamate and activation of N-methyl-D-aspartate (NMDA) receptor leads to an increase of intracellular Ca²⁺ levels that lead to the death of retinal ganglion cells and axons (Choi, 1988), with subsequent damage to the optic nerve and impairment of vision. However, loss of amacrine cells has also been observed in ischemic retinal disease, such as diabetic retinopathy (Gastinger, 2006) and is believed to represent the early events of retinal ischemia. Dionysopoulou, 2020, disclosed that intravitreal administration of (RS)-a-amino-3-hydroxy-5-methyl-4- isoxazolepropionic acid hydrobromide (AMPA) increased ROS levels in rat retina, that the increase was NOX-mediated, and that NOX inhibition may provide providing neuroprotective and/or anti-inflammatory actions in the retinal model of AMPA excitotoxicity. However, Dionysopoulou, 2020, taught that topical treatment with a NOX4 selective inhibitor did not protect bNOS expressing amacrine cells in retina and suggested that this could be due to a due to the lack of expression of NOX4 in bNOS positive amacrine cells.

The most common form of retinopathy, viz., diabetic retinopathy, has been divided into an early, non-proliferative stage involving both vascular and neural elements and a later, proliferative stage involving formation of neovessels. The retinal vascular changes include increased permeability, pericyte and endothelial cell loss, vessel tortuosity, leukostasis and capillary blockage, dropout and hemorrhage. The neural changes include dysregulation of cell functions in both glial (Muller cells) and neuronal cells (inner retinal layer). There is still no accurate animal model for diabetic retinopathy to describe the pathological process. However, the excess formation of VEGF is an important pathological impact in the process.

VEGF is a very potent vascular permeability factor and analyses of diabetic retinae and vitreous indicate that the upregulation of VEGF is correlated with the presence of diabetic macular edema. Studies in animal models support a central role for VEGF in diabetes-associated blood-retinal barrier breakdown. VEGF was injected into the eyes of cynomolgus monkeys, rabbits or rats and as a result, vessel dilatation and tortuosity, vascular retinal permeability developed (Tolentino, 1996; Edelman, 2005; Ishida, 2003). Transgenic mice have also been produced to overexpress VEGF in the photoreceptor (Lai, 2005). In these animals, pathological changes consistent with those seen in early stages of non-proliferative diabetic retinopathy were observed (microaneurysm, retinal vascular leakage and tortuous vessels).

Retinal ischemia-reperfusion injury

The retina is known to be sensitive to ischemia-reperfusion (l/R) injury due to its high oxygen consumption. Conditions such as retinal artery or vein occlusion can lead to ischemia (lack of blood flow) in the retina, followed by reperfusion injury when blood flow is restored, i.e., retinal ischemia-reperfusion injury. NOX4-derived ROS have been implicated in the oxidative stress and retinal damage associated with ischemia-reperfusion injury (Chen 2018), and NOX4 is as a potential target also for the therapy of retinal ischemia-reperfusion injury.

Cataract

Cataract is a complete or a partial opacification on or in the human lens, or in the capsule, that impairs the vision. There are several types of cataracts, including cataracts affecting the center of the lens (nuclear cataracts), cataracts that affect the edges of the lens (cortical cataracts), cataracts that affect the back of the lens (posterior subcapsular cataracts), and congenital cataracts. Although congenital cataracts may have serious visual consequences, early-onset cataracts contribute to a relatively small percentage of visual disability, whereas age-related cataracts account for nearly half of all cases of blindness worldwide. Cataracts may be treated by eye surgery, but re-opacification of the lens post cataract surgery is not uncommon. It has been shown that oxidative stress in the eye plays an important role in the onset and the progression of the cataract (Kaur, 2012), and oxidative damage is a major cause of, in particular, cortical and nuclear cataracts, the most common types of age-related cataracts (Beebe, 2010). The involvement of NOX in cataract pathogenesis has been shown e.g., by Santana-Garrido, 2021.

Corneal diseases and conditions

Corneal diseases may be generally classified into 3 main types, viz. keratitis, corneal ectasia and corneal dystrophy.

Keratitis is an inflammation of the cornea that can either be infectious or noninfectious. Infectious keratitis is mainly bacterial, though there are also corneal inflammations caused by viruses, fungi and parasites. Noninfectious keratitis may be caused by, for example, eye injuries and conditions that dry out the surface of the eye.

Corneal ectasia is group of non-inflammatory disorders of the eye that involve the bilateral thinning of the cornea. Keratoconus is a specific type of corneal ectasia in which the cornea thins and weakens, leading to bulging and distortion. Corneal ectasia sometimes occurs as a complication of certain surgeries, including laser-assisted in situ keratomileusis (LASIK) eye surgery and corneal transplant.

Corneal dystrophy is a group of genetic disorders involving abnormal deposits of proteins, fluid or other materials in one or more layers of the cornea. Fuchs dystrophy is the most common type of corneal dystrophy. Other types include epithelial basement membrane dystrophy, lattice corneal dystrophy and granular corneal dystrophy.

Further corneal conditions and diseases include bullous keratopathy, corneal abrasion, herpetic eye disease, iridocorneal endothelial syndrome, keratoconjunctivitis, and pterygium.

It is known that oxidative stress contributes to the causes of various corneal and ocular surface (or corneal-endothelial) conditions and diseases, including various conditions belonging to the above-mentioned main groups (keratitis, corneal ectasia and corneal dystrophy), such as corneal inflammation, keratoconus, Fuchs' endothelial corneal dystrophy, as well as, for example, pterygium, and dry eyes (Cejka, 2015; Yin, 2018). Increased NOX4 expression in corneal-endothelial diseases compared to a normal cornea has been reported (Matthaei, 2014) and regulation of NOX4 has been suggested as a therapeutic strategy for regulating the homeostasis of corneal-endothelial cells and treating corneal diseases.

Chemical injuries due to alkali agents, such as NH₃, KOH, NaOH etc., or acid agents, such as H2SO4, H2SO3, HF etc., may cause ocular traumas, including corneal burns and injuries. While the first line treatment generally consists of irrigation of the eye to remove the offending substance and restore the physiologic pH, subsequent medical treatment may be needed to enhance recovery of the corneal epithelium and augment collagen synthesis, while also minimizing collagen breakdown and controlling inflammation. One serious complication after corneal injury is neovascularization, a condition that is characterized by the invasion of new blood vessels into the cornea caused by an imbalance between angiogenic and antiangio- genic factors that preserve corneal transparency. While corneal neovascularization may be due to, for example, the above mentioned chemical injuries, other risk factors implicated in the pathogenesis of the disease include contact lens wear, ocular surface disease, previous surgery and herpes. It has been shown that the expression of NOX is increased in corneal epithelial cells after alkali burn injury and that NOX inhibition effectively attenuates alkali burn-induced ROS generation and reduce corneal neovascularization due to alkali burns. Li, 2022, discloses that NOX4 inhibition can attenuate the inflammatory response and reduce corneal neovascularization by scavenging excess ROS. Thus, NOX inhibition, in particular NOX4 inhibition, may offer a new therapeutic approach to treat corneal neovascularization diseases.

There remains a need for an improved treatment of ocular diseases linked to NOX4 and/or NOX2 activity. There further remains a need for a convenient and effective means of ocular topical delivery of a medicament useful in the treatment of ocular diseases.

SUMMARY OF THE INVENTION

A first aspect relates to a gel formulation for ocular topical use, comprising a compound of formula (I)

or a pharmaceutically acceptable salt thereof, wherein

Ri is selected from C1-C6 alkyl, C3-C6 cycloalkyl, C1-C6 alkoxy, hydroxy, and halogen; each Ria is independently selected from C1-C6 alkyl, C1-C6 alkoxy, hydroxy, and halogen; R₂ is selected from C1-C6 alkyl, C1-C6 alkoxy, halogen, and hydroxy;

R3, R4, Rs, and R₆ are independently selected from H and F; and any alkyl is optionally substituted with one or more halogens.

A further aspect relates to the gel formulation as defined herein, for use in the prevention or treatment, (e.g., the treatment), of an eye disorder in a mammal patient.

A further aspect relates to the gel formulation as defined herein, for use in the prevention or treatment, (e.g., the treatment), for which disorder a beneficial effect is achieved by inhibition of NADPH oxidase in the eye, e.g. NOX2 or NOX4, in particular NOX4.

A further aspect relates to the gel formulation as defined herein, for use in the prevention or treatment, (e.g., the treatment), for which disorder a beneficial effect is achieved by inhibition of VEGF in the eye.

A further aspect relates to the gel formulation as defined herein, for use in the prevention or treatment, of an ocular condition or disorder, such as a disorder as mentioned herein above (cf. under the heading “Background of the invention”), for example, an eye disorder selected from age-related macular degeneration, glaucoma, retinopathy, retinal ischemia-reperfusion injury, cataract, corneal diseases, bullous keratopathy, corneal abrasion, herpetic eye disease, iridocorneal endothelial syndrome, keratoconjunctivitis, pterygium, and eye inflammation.

A still further aspect relates to the use of a compound of formula (I) as defined herein in the manufacture of a gel formulation for use in the prevention or treatment (e.g., treatment) of an ocular condition or disorder, such as a disorder as mentioned herein above (cf. under the heading “Background of the invention”), for example, an eye disorder selected from AMD, glaucoma, retinopathy, retinal ischemia-reperfusion injury, cataract, corneal diseases, bullous keratopathy, corneal abrasion, herpetic eye disease, iridocorneal endothelial syndrome, keratoconjunctivitis, pterygium, and eye inflammation.

A still further aspect relates to a method for the prevention or treatment (e.g., treatment) of an ocular condition or disorder in a mammal patient, e.g., a condition or disorder as mentioned herein above (cf. under the heading “Background of the invention”), for example, an eye disorder selected from AMD, glaucoma, retinopathy, retinal ischemia-reperfusion injury, cataract, corneal diseases, bullous keratopathy, corneal abrasion, corneal alkali or acid burn, neovascularization of the eye, for example, corneal or retinal neovascularization, herpetic eye disease, iridocorneal endothelial syndrome, keratoconjunctivitis, pterygium, and eye inflammation.

A still further aspect relates to a dosage container useful for topical ocular administration containing a gel formulation as defined herein.

Further aspects of the invention will become apparent from the reading of the detailed description and the examples.

BRIEF DESCRIPTION OF THE DRAWINGS

FIGURE 1 is a graph showing the number of bNOS expressing amacrine cells (% of control) in eye tissue from diabetic rats treated by topical ocular administration of 20 pl/eye of a gel formulation of EXAMPLE 1 (Diabetic + Ex. 1), and from healthy (Control) or diabetic (Diabetic) rats treated by topical ocular administration of a gel formulation (20 pl/eye) similar to that of EXAMPLE 1 but for containing no API.

FIGURE 2 is a graph showing the number of bNOS expressing amacrine cells (% of control) in eye tissue from diabetic rats treated with the same API as in EXAMPLE 1 , in DMSO as a vehicle (10 mg/ml) (Diabetic + API in DMSO), and in eye tissue from healthy (Control) or diabetic (Diabetic) rats, treated with the DMSO vehicle only.

FIGURE 3 is a graph showing the mean Gray value for NFL immunoreactivity (% of control) in eye tissue from diabetic rats treated by topical ocular administration of 20 pl/eye of a gel formulation of EXAMPLE 1 (Diabetic + Ex. 1), and from healthy (Control) or diabetic (Diabetic) rats treated by topical ocular administration of a gel formulation (20 pl/eye) similar to that of EXAMPLE 1 but for containing no API.

FIGURE 4 is a graph showing the mean Gray value for NFL immunoreactivity (% of control) in eye tissue from diabetic rats treated with the same API as in EXAMPLE 1 , in DMSO as a vehicle (10 mg/ml) (Diabetic + API in DMSO), and in eye tissue from healthy (Control) or diabetic (Diabetic) rats, treated with the DMSO vehicle only. FIGURE 5 is a graph showing the number of caspase-3 positive cells/area (pm²) (% of control) in eye tissue from diabetic rats treated by topical ocular administration of 20 pl/eye of a gel formulation of EXAMPLE 1 (Diabetic + Ex. 1), and from healthy (Control) or diabetic (Diabetic) rats treated by topical ocular administration of a gel formulation (20 pl/eye) similar to that of EXAMPLE 1 but for containing no API.

FIGURE 6 is a graph showing the number of caspase-3 positive cells/area (pm²) (% of control) in eye tissue from diabetic rats treated with the same API as in EXAMPLE 1 , in DMSO as a vehicle (10 mg/ml) (Diabetic + API in DMSO), and in eye tissue from healthy (Control) or diabetic (Diabetic) rats, treated with the DMSO vehicle only.

FIGURE 7 is a graph showing the mean Gray value for GFAP immunoreactivity (% of control) in eye tissue from diabetic rats treated by topical ocular administration of 20 pl/eye of a gel formulation of EXAMPLE 1 (Diabetic + Ex. 1), and from healthy (Control) or diabetic (Diabetic) rats treated by topical ocular administration of a gel formulation (20 pl/eye) similar to that of EXAMPLE 1 but for containing no API.

FIGURE 8 is a graph showing the mean Gray value for GFAP immunoreactivity (% of control) in eye tissue from diabetic rats treated with the same API as in EXAMPLE 1 , in DMSO as a vehicle (10 mg/ml) (Diabetic + API in DMSO), and in eye tissue from healthy (Control) or diabetic (Diabetic) rats, treated with the DMSO vehicle only.

FIGURE 9 is a graph showing the number of reactive lba-1 positive cells/area (pm²) (% of control) in eye tissue from diabetic rats treated by topical ocular administration of 20 pl/eye of a gel formulation of EXAMPLE 1 (Diabetic + Ex. 1), and from healthy (Control) or diabetic (Diabetic) rats treated by topical ocular administration of a gel formulation (20 pl/eye) similar to that of EXAMPLE 1 but for containing no API.

FIGURE 10 is a graph showing the number of reactive lba-1 positive cells /area (pm²) (% of control) in eye tissue from diabetic rats treated with the same API as in EXAMPLE 1 , in DMSO as a vehicle (10 mg/ml) (Diabetic + API in DMSO), and in eye tissue from healthy (Control) or diabetic (Diabetic) rats, treated with the DMSO vehicle only.

FIGURE 11 is a graph showing the TNF-a level (pg/mg of total protein) in eye tissue from diabetic rats treated by topical ocular administration of 20 pl/eye of a gel formulation of EXAMPLE 1 (Diabetic + Ex. 1), and from diabetic (Diabetic) rats treated by topical ocular administration of a gel formulation (20 pl/eye) similar to that of EXAMPLE 1 but for containing no API.

FIGURE 12 is a graph showing the TNF-a level (pg/mg of total protein) in eye tissue from diabetic rats treated with the same API as in EXAMPLE 1 , in DMSO as a vehicle (10 mg/ml) (Diabetic + API in DMSO), and in eye tissue from diabetic (Diabetic) rats, treated with the DMSO vehicle only.

FIGURE 13 is a graph showing the VEGF level (pg/mg of total protein) in eye tissue from diabetic rats treated by topical ocular administration of 20 pl/eye of a gel formulation of EXAMPLE 1 (Diabetic + Ex. 1), and from diabetic (Diabetic) rats treated by topical ocular administration of a gel formulation (20 pl/eye) similar to that of EXAMPLE 1 but for containing no API.

FIGURE 14 is a graph showing the VEGF level (pg/mg of total protein) in eye tissue from diabetic rats treated with the same API as in EXAMPLE 1 , in DMSO as a vehicle (10 mg/ml) (Diabetic + API in DMSO), and in eye tissue from healthy (Control) or diabetic (Diabetic) rats, treated with the DMSO vehicle only.

FIGURE 15 is a graph showing the number of bNOS expressing amacrine cells (% of control) in eye tissue from rats treated by intravitreal injection of PBS (50 mM) (Control), AMPA, 8.4 mM (AMPA), or AMPA (8.4 mM) and compound AF (0.1 mM) (AMPA+ comp. AF).

FIGURE 16 is a graph showing the mean Gray value for GFAP immunoreactivity (% of control) in eye tissue from rats treated by intravitreal injection of PBS (50 mM) (Control); AMPA, 8.4 mM (AMPA); or AMPA (8.4 mM) and compound AF (0.1 mM) (AMPA+ comp. AF).

FIGURE 17 is a graph showing the number of lba-1 positive cells/area (pm²) (% of control) in eye tissue from rats treated by intravitreal injection of PBS (50 mM) (Control); AMPA, 8.4 mM (AMPA); or AMPA (8.4 mM) and compound AF (0.1 mM) (AMPA+ comp. AF).

FIGURE 18 is a graph showing the mean Gray value for NFL immunoreactivity (% of control) in eye tissue from untreated rats (Control); rats treated by intravitreal injection of streptozoto- cin (70 mg/kg in citric buffer) (Diabetic); and from rats treated by intravitreal injection of streptozotocin (70 mg/kg in citric buffer, day 0) followed by topical ocular treatment with a DMSO solution of compound AF (10 mg/ml, 20 pl/eye) daily for 14 days, starting on day 1 (Diabetic + comp. AF).

FIGURE 19 is a graph showing the number of bNOS expressing amacrine cells (% of control) in eye tissue from untreated rats (Control); rats treated by intravitreal injection of streptozotocin (70 mg/kg in citric buffer) (Diabetic); and from rats treated by intravitreal injection of streptozotocin (70 mg/kg in citric buffer, day 0) followed by topical ocular treatment with a DMSO solution of compound AF (10 mg/ml, 20 pl/eye) daily for 14 days, starting on day 1 (Diabetic + comp. AF).

FIGURE 20 is a graph showing the Bcl-2/GAPDH ratio in eye tissue from rats treated by intravitreal injection of streptozotocin (70 mg/kg in citric buffer, day 0) followed by topical ocular treatment with a DMSO solution of compound AF (10 mg/ml, 20 pl/eye) daily for 14 days, starting on day 1 (Diabetic + comp. AF).

FIGURE 21 is a graph showing the mean Gray value for GFAP immunoreactivity (% of control) in eye tissue from untreated rats (Control); rats treated by intravitreal injection of streptozotocin (70 mg/kg in citric buffer) (Diabetic); and from rats treated by intravitreal injection of streptozotocin (70 mg/kg in citric buffer, day 0) followed by topical ocular treatment with a DMSO solution of compound AF (10 mg/ml, 20 pl/eye) daily for 14 days, starting on day 1 (Diabetic + comp. AF).

FIGURE 22 is a graph showing the number of lba-1 positive cells /area (pm²) (% of control) in eye tissue from untreated rats (Control); rats treated by intravitreal injection of streptozotocin (70 mg/kg in citric buffer) (Diabetic); and from rats treated by intravitreal injection of streptozotocin (70 mg/kg in citric buffer, day 0) followed by topical ocular treatment with a DMSO solution of compound AF (10 mg/ml, 20 pl/eye) daily for 14 days, starting on day 1 (Diabetic + comp. AF).

FIGURE 23 is a graph showing the vitroretinal fluorescence (a.u X step) measured in eyes of 4 groups of rabbits having received an induction of vascular leakage by intravitreal injection rhVEGF165, and treatment with topical application of EXAMPLE 1 (1) or a corresponding gel formulation containing no API (2), or with intravitreal injection of Eylea® (3), or Kenacort® Retard (4). DETAILED DESCRIPTION OF THE INVENTION

Definitions

In general, any term used herein shall be given its normal meaning as accepted within the field to which the present invention belongs. For the sake of clarity, however, some definitions will be given herein below, and shall apply throughout the specification and the appended claims, unless otherwise specified or apparent from the context.

Unless otherwise specified or apparent from the context, the articles “a” and “an” are used herein to refer to one or to more than one (i.e., to at least one) of the grammatical object of the article. By way of example, “an element” generally means one element or more than one element.

"Ameliorating" or "alleviating" a condition or state as used herein shall mean to relieve or lessen the symptoms of that condition or state.

As used herein, the term “antioxidant” refers to an agent capable of protecting other compounds (e.g., an API) from oxidation. A pharmaceutically useful antioxidant should be chemically and pharmacologically essentially inert and non-toxic at the amounts used.

As used herein, “API” stands for “active pharmaceutical ingredient”, which in connection with the present disclosure is a compound of formula (I) as defined herein or a pharmaceutically acceptable salt of thereof.

The term “disease or disorder affecting the eye” includes, for example, cataract including diabetic cataract, re-opacification of the lens post cataract surgery, diabetic and other forms of retinopathy.

The term “dosage container” as used herein refers to a container, such as a bottle, vial, tube, flask etc, suitable to contain a volume of the formulation as provided herein, either a volume corresponding to a unit (single) dose, or a volume corresponding to more than one dose (multi-dose). The dosage container may include means allowing for application of a suitable amount of the formulation to an eye of a patient, or such means may be provided separately from the container. As used herein, "effective" as in an amount effective to achieve an end, i.e. , "therapeutically effective amount", means the quantity of a component that is sufficient to yield an indicated therapeutic response without undue adverse side effects (such as toxicity, irritation, or allergic response) commensurate with a reasonable benefit/risk ratio when used in the manner of this disclosure. An effective amount may vary according to factors known in the art, such as the disease state, age, sex, and weight of the human or animal being treated.

The term “excipient” refers to a pharmaceutically acceptable chemical, such as known to those of ordinary skill in the art of pharmacy to aid in the administration of the medicinal agent. It is a compound that is useful in preparing a pharmaceutical composition, generally safe, non-toxic and neither biologically nor otherwise undesirable, and includes excipients that are acceptable for veterinary use as well as human pharmaceutical use.

As used herein, the term “eye disease” (which term is considered herein to be synonymous with “ophthalmic disorder”, “ophthalmic disease”, “ocular disorder”, “ocular disease”, or “eye disorder” etc) refers to a disease affecting the eye of a mammal subject, i.e., an animal or a human, preferably a human.

The term “gel” as used herein refers to as a substantially dilute cross-linked system containing a liquid phase, which system exhibits no flow when in a steady state. It is a solid or semisolid system of at least two constituents, consisting of a condensed mass enclosing and interpenetrated by a liquid phase.

The term “humectant” refers to a hydrophilic compound capable of retaining moisture in a formulation.

As used herein, the term "inhibition" or “inhibiting” etc when used in respect of a disease progression or disease complication in a subject means preventing or reducing the disease progression and/or disease complication in the subject.

The term “inhibitor” used in the context of the invention is defined as a molecule that inhibits completely or partially the activity of another molecule, e.g., an enzyme. The term “mammal” refers to a human or any mammalian animal, e.g., a primate, a farm animal, a pet animal, or a laboratory animal. Examples of such animals are monkeys, cows, sheep, horses, pigs, dogs, cats, rabbits, mice, rats etc. Preferably, the mammal is a human.

As used herein, the term “ocular”, refers to the eye, and, for example, the expression “ocular formulation” refers to a formulation adapted and suitable for application to an eye of a subject, such as a human.

"Optional" or "optionally" means that the subsequently described event or circumstance may or may not occur, and that the description includes instances where the event or circumstance occurs and instances where it does not.

The term “osmolality” refers to the number of osmotically active solute particles dissolved in a kilogram of a solvent (generally water).

“Pharmaceutically acceptable” means being useful in preparing a pharmaceutical composition that is generally safe, non-toxic and neither biologically nor otherwise undesirable and includes being useful for veterinary use as well as human pharmaceutical use.

A "preservative", as used herein, refers to an additive which inhibits both microbial growth and kills microorganisms that contaminate a formulation exposed to the surroundings.

The term “solubilizing agent” (or alternatively “solubilizer”) refers to a compound that, when added to a solvent phase or formulation, is capable of increasing the solubility of another compound in said solvent phase or formulation.

The term "solubilizing effective amount" of a substance ("solubilizer") within a formulation refers to an amount of the substance sufficient to solubilize another component of the composition. For example, an "API-solubilizing effective amount" is an amount sufficient to solubilize an API (which in the present case is a compound of formula (I) or a pharmaceutically acceptable salt thereof).

The term “subject” as used herein refers to a mammal individual, unless otherwise specified or apparent from the context. The term “surfactant” refers to a chemical compound capable of lowering the surface tension (or interfacial tension) between two liquids, between a gas and a liquid, or between a liquid and a solid. A surfactant is an amphiphilic compound, i.e. , a compound that contains a hydro- phobic moiety (“the hydrophobic tail”) as well as a hydrophilic moiety (the “hydrophilic head” or “polar head”). Most commonly, surfactants are classified according to the hydrophilic head. A “non-ionic surfactant” has no electrically charged groups in its head; the hydrophilic head of a “cationic surfactant” carries a net positive electrical charge, and the hydrophilic head of an “anionic surfactant” carries a net negative electrical charge.

The term “tonicity adjusting agent” (or alternatively “tonicity agent”), as used herein, refers to a compound that contributes to the osmolality of a solution. The osmolality of an ocular formulation is preferably adjusted to minimize discomfort to the patient upon ocular administration.

The term “topical” as used in “topical application” refers to the local application (of a medicament) to a part of the surface of the body of subject, including mucosal surfaces of the body. For example, “topical ocular application” refers to the application to an eye of a subject. A “topical ocular formulation” refers to a formulation suitable for topical ocular application.

As used herein, "treating" encompasses, e.g., inducing inhibition, regression, or stasis of a disease, disorder or condition, or ameliorating or alleviating a symptom of a disease, disorder or condition.

The term "unit dose" as used herein is the amount of the inventive formulation to be administered to the subject in a single administration, or the amount of a compound of formula (I), or salt of thereof contained in said amount of the inventive formulation. The unit dose disclosed herein can be administered once only, or periodically, such as once daily, twice daily, three times daily, four times daily, five times daily, every other day, weekly, twice weekly, three times weekly, four times weekly, five times weekly or six times weekly, for example.

The term “viscosity” as used herein refers to the dynamic viscosity.

The term “viscosity agent” (which in the technical field of the invention may also be referred to as viscosity enhancing agent, viscosity enhancer, viscosity modifying agent, viscosity imparting agent, thickening agent, thickener etc.) refers to an agent capable of increasing the viscosity of a liquid when admixed with the liquid.

The term “alkyl” refers to straight or branched chain alkyl of the general formula C_nH_2n+i-

The term “cycloalkyl” refers to a cyclic alkyl group of the general formula C_nH2n-i.

The expression “Cm-Cn” in connection with a moiety such as, for example, an alkyl or cycloalkyl, indicates that the moiety contains a number of carbon atoms ranging from m to n, and where n is higher than m.

The term “Cm-Cn alkyl” refers to an alkyl containing from m to n carbon atoms, wherein n is an integer higher than m, and m is at least 1. For example, methyl is a C1 alkyl.

The term “alkoxy” refers to a moiety of formula

wherein R is an alkyl group.

The term “Cm-Cn alkoxy” refers to an alkoxy group wherein the alkyl group is Cm-Cn alkyl. For example, methoxy is a C1 alkoxy.

The term “halogen” as used herein generally refers to fluoro (F), chloro (Cl), bromo (Br), or iodo (I), preferably fluoro (F), chloro (Cl), or bromo (Br).

The term “hydroxy” refers to the moiety HO-.

The compound of formula (I)

A first aspect relates to a gel formulation for use by ocular topical application, comprising a compound of formula (I) as defined herein.

In a compound of formula (I), Ri is selected from C1-C6 alkyl, C3-C6 cycloalkyl, C1-C6 alkoxy, hydroxy, and halogen.

In some embodiments, Ri is selected from C1-C6 alkyl, C3-C6 cycloalkyl, C1-C6 alkoxy, and halogen. In some embodiments, Ri is selected from C1-C6 alkyl, C3-C6 cycloalkyl, and halogen. In some embodiments, Ri is selected from C1-C6 alkyl, and halogen. In some embodiments, Ri is selected from C1-C6 alkyl. In some embodiments, Ri is selected from halogen.

When Ri is C1-C6 alkyl, Ri more particularly may be C1-C4 alkyl, or C1-C3 alkyl. In some embodiments, when Ri is C1-C6 alkyl, Ri more particularly is methyl or isopropyl. In some embodiments, when Ri is C1-C6 alkyl, Ri more particularly is methyl. In some of these embodiments, any such alkyl group is halogenated, e.g., Ri is selected from methyl and trifluoromethyl.

When Ri is C1-C6 alkoxy, Ri more particularly may be C1-C4 alkoxy, or C1-C3 alkoxy. In some embodiments, when Ri is C1-C6 alkoxy, Ri more particularly is methoxy. In some of these embodiments, any such alkoxy group is halogenated, e.g., Ri is selected from methoxy and trifluoromethoxy.

When Ri is halogen, such halogen e.g., may be selected from F, Cl and Br. In some embodiments, any such halogen is selected from Cl and Br, e.g., Br. In some embodiments, Ri is Br.

In some embodiments, Ri is selected from C1-C3 alkyl, cyclopropyl, C1-C3 alkoxy, hydroxy, and halogen. In some embodiments, Ri is selected from methyl, trifluoromethyl, ethyl, isopropyl, cyclopropyl, methoxy, hydroxy, and halogen; e.g., from methyl, trifluoromethyl, isopropyl, cyclopropyl, methoxy, hydroxy, Cl, and Br.

In a compound of formula (I), each Ri_a is independently selected from C1-C6 alkyl, C1-C6 alkoxy, hydroxy, and halogen. In some embodiments, each Ri_a is independently selected from C1-C6 alkyl, hydroxy, and halogen. In some embodiments, each Ri_a is independently selected from C1-C6 alkyl, and halogen. In some embodiments, each Ri_a is independently selected from C1-C6 alkyl. In some embodiments, each Ri_a is independently selected from halogen. In some embodiments, at least one Ri_a is selected from halogen.

When any Ri_a is C1-C6 alkyl, such Ri_a more particularly may be C1-C4 alkyl, or C1-C3 alkyl. In some embodiments, when any Ri_a is C1-C6 alkyl, such Ri_a more particularly is methyl. In some of these embodiments, any such alkyl group is halogenated, e.g., fluorinated, such as in trifluoromethyl. When any Ri_a is C1-C6 alkoxy, such Ri_a more particularly may be C1-C4 alkoxy, or C1-C3 alkoxy. In some embodiments, when any Ri_a is C1-C6 alkoxy, such Ri_a more particularly is methoxy. In some of these embodiments, any such alkyl group is halogenated, e.g., fluorinated, such as in trifluoromethoxy.

When any Ri_a is halogen, such halogen e.g., may be selected from F, Cl and Br. In some embodiments, any such halogen is selected from Cl and Br, e.g., Cl. In some embodiments, both Ri_a are Cl.

In some embodiments, each Ri_a is independently selected from methyl, trifluoromethyl, hydroxy, and Cl; e.g., methyl, hydroxy, and Cl, or from methyl and Cl, e.g., from Cl.

In a compound of formula (I), R2 is selected from C1-C6 alkyl, C1-C6 alkoxy, halogen, and hydroxy. In some embodiments, R2 is selected from C1-C6 alkyl, halogen, and hydroxy. In some embodiments, R2 is selected from C1-C6 alkyl, and halogen. In some embodiments, R2 is selected from C1-C6 alkyl. In some embodiments, R2 is selected from halogen.

When R2 is C1-C6 alkyl, R2 more particularly may be C1-C4 alkyl, or C1-C3 alkyl. In some embodiments, when R₂ is C1-C6 alkyl, R₂ more particularly is methyl. In some of these embodiments, any such alkyl group is halogenated, e.g., R₂ is selected from methyl and trifluoromethyl.

When R₂ is C1-C6 alkoxy, R₂ more particularly may be C1-C4 alkoxy, or C1-C3 alkoxy. In some embodiments, when R₂ is C1-C6 alkoxy, R2 more particularly is methoxy.

When R2 is halogen, such halogen e.g., may be selected from F, Cl and Br. In some embodiments, any such halogen is selected from F and Cl. In some embodiments, any such halogen is selected from Cl and Br. In some embodiments, any such halogen is Cl. In some embodiments, any such halogen is Br.

In some embodiments, R2 is selected from methyl, trifluoromethyl, methoxy, trifluoromethoxy, hydroxy, F, Cl, and Br.

As mentioned herein, any alkyl group (including as part of another moiety, such as an alkoxy group) may optionally be substituted by one or more halogens, e.g., one or more halogens selected from F and Cl, in particular F. In some embodiments, any reference to, for example, a methyl group or methoxy group, also includes the corresponding halogenated, e.g., trifluorinated group.

In some embodiments, Ri is selected from C1-C3 alkyl, C1-C3 alkoxy, hydroxy, and halogen; and each Ri_a is independently selected from C1-C3 alkyl, and halogen.

In some embodiments, Ri is selected from C1-C3 alkyl, and halogen; and each Ri_a is independently selected from C1-C3 alkyl, and halogen.

In some embodiments, Ri is selected from halogen; and each Ri_a is independently selected from halogen.

In some embodiments, Ri is Br; and each Ri_a is Cl.

In some embodiments, Ri is selected from C1-C3 alkyl, C1-C3 alkoxy, hydroxy, and halogen; each Ri_a is independently selected from C1-C3 alkyl, and halogen; and R2 is selected from C1-C3 alkyl, C1-C6 alkoxy, halogen, and hydroxy.

In some embodiments, Ri is selected from C1-C3 alkyl, and halogen; each Ri_a is independently selected from C1-C3 alkyl, and halogen; and R₂ is selected from C1-C3 alkyl, halogen, and hydroxy.

In some embodiments, Ri is selected from halogen; each Ri_a is independently selected from halogen; and R₂ is selected from C1-C3 alkyl.

In some embodiments, Ri is Br; each Ri_a is Cl; and R₂ is methyl.

In a compound of formula (I) each one of R3, R4, Rs, and Re is independently selected from H and F. In some embodiments, at least two of R3, R4, Rs, and Re are H. In some embodiments, at least three of R3, R4, Rs, and Re are H. In some embodiments, R3 and R4 are H. In some embodiments, R3, R4, and Rs are H. In some embodiments, R3, R4, Rs, and Re are all H. In some particular embodiments, R3 and R4 are H, and Rs and Re are F. In some further particular embodiments, R3, R4, and Rs are H, and Re is F. In still further embodiments, at least one of R₃, R4, Rs, and R₆ is F; e.g., at least one of R₅ and R₆ is F. In still further embodiments, R₃ and R₄ are H; and R₅ and R₆ are selected from H and F.

For example, in some embodiments, R1 is Br; each Ri_a is Cl; R2 is as defined herein, e.g., R₂ is methyl; R₃ and R₄ are H, and R₅ and R₆ are independently selected from H and F.

In some embodiments, the compound of formula (I) is selected from any of the compounds shown in TABLE 1 , which compounds may be referred to herein as “compounds A-AF”. TABLE 1

The compounds shown in TABLE 1 have been described in WO 2019/215291 A1 , also disclosing their NOX2 and/or NOX 4 inhibiting activity. In some embodiments, the compound of formula (I) is 4-bromo-2,6-dichloro-N-[2-(2- methylphenyl)ethyl]benzene-1-sulfonamide (compound J). In some embodiments, the compound of formula (I) is 4-hydroxy-/V-[2-(2-hydroxyphenyl)ethyl]-2,6-dimethyl- benzenesulfonamide (compound AF). Compound J is a highly selective NOX4 inhibitor, whereas compound AF is a highly selective NOX2 inhibitor.

The term pharmaceutically acceptable salt of a compound refers to a salt that is pharmaceutically acceptable, as defined herein, and that possesses the desired pharmacological activity of the parent compound. Pharmaceutically acceptable salts include acid addition salts formed with inorganic acids, e.g., hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid; or formed with organic acids, e.g., acetic acid, benzenesulfonic acid, benzoic acid, camphorsulfonic acid, citric acid, ethanesulfonic acid, fumaric acid, glucohep- tonic acid, gluconic acid, glutamic acid, glycolic acid, hydroxynaphtoic acid, 2-hydroxye- thanesulfonic acid, lactic acid, maleic acid, malic acid, malonic acid, mandelic acid, methanesulfonic acid, muconic acid, 2-naphthalenesulfonic acid, propionic acid, salicylic acid, succinic acid, tartaric acid, p-toluenesulfonic acid, trimethylacetic acid; or salts formed when an acidic proton present in the parent compound either is replaced by a metal ion, e.g., an alkali metal ion, an alkaline earth ion, or an aluminum ion; or coordinates with an organic or inorganic base. Acceptable organic bases include e.g., diethanolamine, ethanolamine, N- methylglucamine, triethanolamine, morpholine, and tromethamine. Acceptable inorganic bases include e.g., ammonia, aluminum hydroxide, calcium hydroxide, potassium hydroxide, sodium carbonate and sodium hydroxide.

The gel formulation

A gel formulation according to the invention generally comprises a therapeutically effective amount of a compound of formula (I) as defined herein or a pharmaceutically acceptable salt thereof, in a carrier in the form of a gel.

The topical ocular formulation provided herein is a pharmaceutical gel formulation and comprises a compound of formula (I), as defined herein, or a pharmaceutically acceptable salt thereof (which compound or salt may be collectively referred to herein as an API) at a concentration suitable to provide a therapeutically effective amount of said API by topical administration to the eye (ocular administration) of a suitable volume of said gel, e.g., 1-5 drops of the gel, or 1-3 drops of the gel, or 1 drop (or droplet) of the gel into the eye of a treated subject. In some embodiments, the concentration of the API in the formulation is in the range of about 0.1 % (w/w) of the formulation (i.e., 100 g of the formulation contains 0.1 g of compound of formula (I) in free base form, or a corresponding amount of a salt thereof) to about 10 % (w/w); e.g., about 0.2 to about 10 % (w/w), about 0.5 to about 10 % (w/w), about 1 to about 10 % (w/w), about 2 to about 10 % (w/w), or about 5 to about 10 % (w/w),

In some further embodiments, the concentration of the API in the formulation is in the range of about 0.1 % (w/w) of the formulation to about 5 % (w/w); e.g., about 0.2 to about 5 % (w/w), about 0.5 to about 5 % (w/w), about 1 to about 5 % (w/w), or about 2 to about 5 % (w/w).

In still some further embodiments, the concentration of the API in the formulation is in the range of about 0.1 % (w/w) of the formulation to about 2 % (w/w); e.g., about 0.2 to about 2 % (w/w), about 0.5 to about 2 % (w/w), or about 1 to about 2 % (w/w).

In some embodiments, the gel formulation provided herein contains an API as defined herein in the form of a suspension in the gel carrier. In such embodiments, an advantageous feature of the gel formulation of the invention is the high homogeneity of the suspension throughout the gel formulation and its high stability over time against sedimentation.

In some embodiments, the gel formulation of the invention has pH in the range of 6.8 to 8.5, an osmolality in the range of 200 to 600 mOsm/kg H₂O, and a dynamic viscosity, measured at 100 rpm and 25 °C, in the range of 500 to 2000 mPa.s.

The pH

The formulation provided herein has a pH of from about 6.8 to about 8.5. In some embodiments, the pH is at most 8.4. For example, in some embodiments the formulation has a pH of from 6.8 to 8.4, from 7.0 to 8.4, from 7.2 to 8.4, from 7.4 to 8.4, or from 8.0 to 8.4. In some embodiments, the pH is at most 8.0. For example, in some embodiments the formulation has a pH of from 6.8 to 8.0, from 7.0 to 8.0, from 7.2 to 8.0, or from 7.4 to 8.0. In some further embodiments, the formulation has a pH in the range of 6.8 to 7.8, or 6.8 to 7.6, or 6.8 to 7.4. In some further embodiments, the formulation has a pH in the range of 7.0 to 7.8, or 7.0 to 7.6, or 7.0 to 7.4. In some further embodiments, the formulation has a pH in the range of 7.2 to 7.8, or 7.2 to 7.6, or 7.2 to 7.4. In some embodiments, the formulation has a pH of about

7.4.

The method for pH measurement

The pH has been measured using a Hanna HI 8424, Mettler Toledo InLab® Micro electrode.

The osmolality

Ideally, a pharmaceutical formulation for ocular administration should preferably have an osmolality within a range of 200 to 600 mOsm/kg H2O, in order not to cause discomfort on application to the eye, though values somewhat outside this range may be tolerated in case the amount of formulation applied to the eye is small.

In some embodiments, the gel formulation of the invention has an osmolality in the range of about 200 to about 550 mOsm/kg H2O, about 200 to about 500 mOsm/kg H2O, about 200 to about 450 mOsm/kg H2O, or about 200 to about 400 mOsm/kg H2O, or about 200 to about 350 mOsm/kg H2O.

In some embodiments, the gel formulation of the invention has an osmolality in the range of about 300 to about 600 mOsm/kg H₂O, about 300 to about 550 mOsm/kg H₂O, about 300 to about 500 mOsm/kg H₂O, about 300 to about 450 mOsm/kg H₂O, about 300 to about 400 mOsm/kg H₂O, or about 300 to about 350 mOsm/kg H₂O.

In some embodiments, the gel formulation of the invention has an osmolality in the range of about 400 to about 600 mOsm/kg H2O, about 400 to about 550 mOsm/kg H2O, about 400 to about 500 mOsm/kg H2O, or about 400 to about 450 mOsm/kg H2O.

In some embodiments, the gel formulation of the invention has an osmolality in the range of about 450 to about 600 mOsm/kg H2O, about 450 to about 550 mOsm/kg H2O, or about 450 to about 500 mOsm/kg H2O.

In some embodiments, the gel formulation of the invention has an osmolality in the range of about 500 to about 600 mOsm/kg H2O, or about 500 to about 550 mOsm/kg H2O.

In some embodiments, the gel formulation of the invention has an osmolality in the range of about 250 to about 500 mOsm/kg H2O, about 250 to about 450 mOsm/kg H2O, about 250 to about 400 mOsm/kg H₂O, about 250 to about 375 mOsm/kg H₂O, about 250 to about 350 mOsm/kg H₂O, or about 250 to about 325 mOsm/kg H₂O.

In some embodiments, the gel formulation of the invention has an osmolality in the range of about 260 to about 375 mOsm/kg H₂O, about 270 to about 375 mOsm/kg H₂O, about 280 to about 375 mOsm/kg H₂O, or about 290 to about 375 mOsm/kg H₂O.

In some embodiments, the gel formulation of the invention has an osmolality in the range of about 260 to about 350 mOsm/kg H₂O, about 270 to about 350 mOsm/kg H₂O, about 280 to about 350 mOsm/kg H₂O, or about 290 to about 350 mOsm/kg H₂O.

In some embodiments, the gel formulation of the invention has an osmolality in the range of about 260 to about 320 mOsm/kg H₂O, about 270 to about 320 mOsm/kg H₂O, or about 280 to about 320 mOsm/kg H₂O, or about 285 to about 315 mOsm/kg H₂O, or about 290 to about 310 mOsm/kg H₂O, or about 295 to about 305 mOsm/kg H₂O, e.g., about 300 mOsm/kg H₂O.

It is noted that, some of the excipients used to prepare the gel formulations, for example, the solubilizing agents employed and the buffering agent, also may function as osmolality (tonicity) agents. However, if necessary, one or more further tonicity agents, such as, for example, mannitol, sorbitol, glycerol, polyethylene glycol (PEG), polypropylene glycol (PPG), or sorbitol, may be added to the gel formulation, in an amount sufficient to adjust the osmolality thereof to withing a desired range, though in some embodiments, such additional agents are not considered necessary. The person of ordinary skill in the art will be well capable of measuring and, if necessary, adjusting the osmolality of the gel formulation.

Method for osmolality measurement

The osmolality was measured using a Roebling Type 13/13DR osmometer by following the manufacturer’s instructions.

The viscosity

The gel formulation provided herein has a dynamic viscosity in the range of about 500 mPa.s to about 2000 mPa.s when measured at 100 rpm, at a temperature of 25 °C using a method as described herein. In some embodiments, the viscosity is in a range of about 500 mPa.s to about 1900 mPa.s, about 500 mPa.s to about 1800 mPa.s, about 500 mPa.s to about 1700 mPa.s, about 500 mPa.s to about 1600 mPa.s, about 500 mPa.s to about 1500 mPa.s, about 500 mPa.s to about 1400 mPa.s, about 500 mPa.s to about 1300 mPa.s, or about 500 mPa.s to about 1200 mPa.s.

In some embodiments, the viscosity is in a range of about 600 mPa.s to about 2000 mPa.s, about 600 mPa.s to about 1900 mPa.s, about 600 mPa.s to about 1800 mPa.s, about 600 mPa.s to about 1700 mPa.s, about 600 mPa.s to about 1600 mPa.s, about 600 mPa.s to about 1500 mPa.s, about 600 mPa.s to about 1400 mPa.s, about 600 mPa.s to about 1300 mPa.s, or about 600 mPa.s to about 1200 mPa.s.

In some embodiments, the viscosity is in a range of about 700 mPa.s to about 2000 mPa.s, about 700 mPa.s to about 1900 mPa.s, about 700 mPa.s to about 1800 mPa.s, about 700 mPa.s to about 1700 mPa.s, about 700 mPa.s to about 1600 mPa.s, about 700 mPa.s to about 1500 mPa.s, about 700 mPa.s to about 1400 mPa.s, about 700 mPa.s to about 1300 mPa.s, or about 700 mPa.s to about 1200 mPa.s.

In some embodiments, the viscosity is in a range of about 800 mPa.s to about 2000 mPa.s, about 800 mPa.s to about 1900 mPa.s, about 800 mPa.s to about 1800 mPa.s, about 800 mPa.s to about 1700 mPa.s, about 800 mPa.s to about 1600 mPa.s, about 800 mPa.s to about 1500 mPa.s, about 800 mPa.s to about 1400 mPa.s, about 800 mPa.s to about 1300 mPa.s, or about 800 mPa.s to about 1200 mPa.s.

In some embodiments, the viscosity is in a range of about 900 mPa.s to about 2000 mPa.s, about 900 mPa.s to about 1900 mPa.s, about 900 mPa.s to about 1800 mPa.s, about 900 mPa.s to about 1700 mPa.s, about 900 mPa.s to about 1600 mPa.s, about 900 mPa.s to about 1500 mPa.s, about 900 mPa.s to about 1400 mPa.s, about 900 mPa.s to about 1300 mPa.s, or about 900 mPa.s to about 1200 mPa.s.

In some embodiments, the viscosity is in a range of about 1000 mPa.s to about 2000 mPa.s, from about 1000 mPa.s to about 1900 mPa.s, from about 1000 mPa.s to about 1800 mPa.s, from about 1000 mPa.s to about 1700 mPa.s, from about 1000 mPa.s to about 1600 mPa.s, from about 1000 mPa.s to about 1500 mPa.s, from about 1000 mPa.s to about 1400 mPa.s, from about 1000 mPa.s to about 1300 mPa.s, or from about 1000 mPa.s to about 1200 mPa.s. Method for viscosity measurement

As referred to herein, the viscosity is a dynamic viscosity and has been measured at 25°C and 100 rpm using a Brookfield DV-II+ viscosimeter with bucket/needle SC4-16/8R.

In some embodiments, the gel formulation of the invention has pH in the range of 6.8 to 8.5, an osmolality in the range of 200 to 600 mOsm/kg H2O, and a dynamic viscosity, measured at 100 rpm and 25 °C, in the range of 500 to 2000 mPa.s.

In some further embodiments, the gel formulation provided herein has a pH in the range of

6.8 to 7.5, for example 6.8 to 7.4, or 6.8 to 7.2; an osmolality in the range of 250 to 400 mOsm/kg; and a dynamic viscosity in the range of 500 to 1700 mPa.s.

6.8 to 7.2, an osmolality in the range of 250 to 400 mOsm/kg, and a dynamic viscosity in the range of 500 to 1500 mPa.s, for example, 600 to 1500 mPa.s, such as in the range of 700 to 1500 mPa.s, e.g., in the range of 800 to 1500 mPa.s.

6.8 to 7.2, an osmolality in the range of 250 to 400 mOsm/kg, and a dynamic viscosity in the range of 700 to 1700 mPa.s, for example, 700 to 1500 mPa.s, such as in the range of 700 to 1200 mPa.s, e.g., in the range of 700 to 1000 mPa.s, e.g., at least 800 mPa.s.

In some exemplary embodiments, the gel formulation provided herein has a pH in the range of 6.8 to 7.5, for example 6.8 to 7.4, or 6.8 to 7.2; an osmolality in the range of 250 to 400 mOsm/kg; and a dynamic viscosity in the range of 500 to 1700 mPa.s.

6.8 to 7.2, an osmolality in the range of 250 to 400 mOsm/kg, and a dynamic viscosity in the range of 700 to 1700 mPa.s, for example, 700 to 1500 mPa.s, such as in the range of 700 to 1200 mPa.s, e.g., in the range of 700 to 1000 mPa.s, e.g., at least 800 mPa.s. The gel formulation for ocular topical use as provided herein comprises a compound of formula (I) as defined herein above, or a pharmaceutically acceptable salt thereof, and one or more excipients.

The topical ocular gel formulation provided herein will generally contain a compound of formula (I) as defined herein in the form of a solution or suspension in an aqueous gel vehicle, comprising one or more pharmaceutically acceptable excipients, Generally, the excipients will include one or more solubilizers, one or more buffers, and one or more viscosity agents.

The solubilizer

The gel formulation will generally contain at least one solubilizer for the compound of formula (I) or the pharmaceutically acceptable salt thereof. The at least one solubilizer may be selected from, for example, dimethyl sulfoxide (DMSO) and surfactants, such as non-ionic surfactants, for example, a surfactant from the Tween® series, e.g. Tween® 20 (CAS number 9005-64-5), 40 (CAS number 9005-66-7), or 80 (CAS number 9005-65-6). For example, the surfactant may be Tween® 80 (polyoxyethylene (80) sorbitan monooleate), also referred to as polysorbate 80, Kolliphor® 80 etc. In some embodiments, the gel formulation includes at least one of DMSO and a non-ionic surfactant, such as a Tween® surfactant, e.g., Tween® 80, optionally in combination with one or more further solubilizers, such as a cyclodextrine derivative (for example, hydroxypropyl-p-cyclodextrin), tetraethylene glycol, ethanol, or polyvinylalcohol). Optionally, the solubilizer may include an oil such as PEG300 or PEG400 (CAS number 25322-68-3), or silicone oil (CAS number 63148-62-9).

In some embodiments, the solubilizer comprises at least one of DMSO and a polyoxyethylene sorbitan monooleate, such as Tween® 80, e.g., a mixture of DMSO and a polyoxyethylene sorbitan monooleate. For example, the solubilizer may comprise a mixture of DMSO and a non-ionic surfactant containing the two components at a weight ratio of 1 :10 to 10:1 , e.g., 1 :8 to 8:1 , 1 :5 to 5:1. In some embodiments, the solubilizer is a mixture containing DMSO and polyoxyethylene sorbitan monooleate at a weight ratio of DMSO to polyoxyethylene sorbitan monooleate of from 1 :10 to 1 :1 , e.g., 1 :10 to 1 :2, or 1 :10 to 1 :3, or 1 :10 to 1 :4. In some embodiments, the solubilizer is a mixture containing DMSO and polyoxyethylene sorbitan monooleate at a weight ratio of DMSO to polyoxyethylene sorbitan monooleate of from 1 :8 to 1 :1 , e.g., 1 :8 to 1 :2, or 1 :8 to 1 :3, or 1 :8 to 1 :4; for example, 1 :6 to 1 :1 , e.g., 1 :6 to 1 :2, or 1 :6 to 1 :3, or 1 :6 to 1 :4. In some of these embodiments, the non-ionic surfactant is a polyoxyethylene sorbitan monooleate, such as polyoxyethylene (80) sorbitan monooleate.

In some embodiments, the composition comprises solubilizer (including one or further solubilizing agents) and API at a weight ratio of solubilizer to API of 1 :1 to 15:1 , or 2:1 to 15:1 , or 3:1 to 15:1 , or 4:1 to 15:1 , or 5:1 to 15:1 ; or 1 :1 to 10:1 , or 2:1 to 10:1 , or 3:1 to 10:1 , or 4:1 to 10:1 , or 5:1 to 10:1 ; or 5:1 to 8:1 ; or 1 :1 to 8:1 , or 2:1 to 8:1 , or 3:1 to 8:1 , or 4:1 to 8:1 , or 5:1 to 8:1.

The viscosity agent

The gel formulation will generally contain at least one viscosity agent. Suitable viscosity agents for use herein include viscosity agents providing a gel formulation comprising a compound of formula (I) and having a viscosity, expressed as dynamic viscosity, in the range as defined herein above. Such a viscosity agent can be selected from, for example, different cellulose polymer derivatives, such as hydroxyethyl cellulose (HEC), (hydroxypropyl)methyl cellulose, methylcellulose, and carboxymethyl cellulose derivatives, optionally in combination with one or more further viscosity agents, e.g., selected from the group consisting of polyvinyl alcohol, poly(acrylic acid) homo- or copolymers (carbomers), and polyvinylpyrrolidone. The viscosity agent preferably is a high viscosity cellulose derivative, such as a high viscosity carboxymethyl cellulose derivate, for example, a high viscosity carboxymethylcellulose sodium salt (CAS No. 9004-32-4), having a viscosity of, for example, 1500 to 3000 mPa.s (as a 1% solution).

The viscosity agent (which term may refer to either one particular viscosity agent or a mixture of such agents) is present in a total amount sufficient to provide the desired viscosity in the formulation, i.e. a viscosity within the ranges as mentioned herein above. As will be apparent to the person of ordinary skill in the art, the exact amount will vary as a function of the particular viscosity agent(s) selected, and furthermore will depend on the other ingredients in the formulation, and the concentrations of those other ingredients. The person of ordinary skill in the art will be able to determine the required amount of viscosity agent in light of the present description and illustrating examples provided herein.

In some embodiments, the gel formulation provided herein comprises a viscosity agent, such as a high viscosity carboxymethylcellulose, in an amount ranging from 0.5 % w/w to 5 % w/w (based on the total weight of the formulation), e.g., 1 to 5 % w/w, or 1 to 4 % w/w, or 1 to 3 % w/w, or 1 to 2 % w/w, or 1 to 1 .5 % w/w.

The buffering agent

The gel formulation will generally contain at least one pH buffer, to maintain the pH of the formulation in the range indicated herein.

For example, phosphate buffer solutions at different pH values are commercially available, such in the form of tablets, that may be used to prepare phosphate-buffered isotonic saline (PBS) of different concentrations, for example, a 1X PBS solution containing 137 mM NaCI, 2.7 mM KCI, 10 mM Na2HPC>4, and 1.8 mM KH2PO4, which may be used to prepare 0.5X PBS, by 1 :2 dilution.

Additionally, it is well within the knowledge of the person of ordinary skill in the art to prepare a phosphate buffer solution having a pH within a desired range by admixing the suitable salt ingredients with deionized water and optionally adjusting the pH with a strong acid such as HCI or a strong base, such as NaOH.

In some embodiments, the buffering agent is a phosphate buffer, such as mentioned herein.

The gel formulation may optionally one or more further active ingredients in addition to the compound of formula (I) or the salt thereof and/or one or more further excipients in addition to those mentioned herein above, such as antioxidants, e.g., EDTA or similar agents, preservatives, e.g., benzalkonium chloride or similar agents, humectants, e.g., glycerol or similar agents, tonicity adjusting agents, e.g., mannitol or similar agents, additional pH regulating agents, e.g., a strong acid (such as HCI) or a strong base (such as NaOH), etc.

In some embodiments, the gel formulation provided herein is contains an API as defined herein, as a suspension in the liquid phase of the gel carrier. In such embodiments, an advantageous feature of the gel formulation of the invention is the homogeneity of the obtained suspension and high stability against sedimentation.

A further advantage of the gel formulation provided herein is the generally very high stability against chemical degradation of the API in said formulation. In some embodiments, the gel formulation provided herein contains a compound of formula (I) or a pharmaceutically acceptable salt thereof, in a gel vehicle comprising a solubilizer agent such as DMSO in combination with Tween® 80 or a similar non-ionic surfactant, a high viscosity carboxymethyl cellulose, and phosphate buffer saline.

In some embodiments, the gel formulation provided herein contains about 1 to 5 % by weight of a compound of formula (I) or a pharmaceutically acceptable salt thereof, about 5 to 10 % by weight of a solubilizer agent such as DMSO in combination with Tween® 80 or a similar non-ionic surfactant, about 1 to 5 % by weight of a high viscosity carboxymethyl cellulose, and phosphate buffer saline, optionally as the only further ingredient (i.e. , constituting about 80 to 93 % by weight).

The eye condition or disease

A further aspect relates to a gel formulation comprising a compound of formula (I) as defined herein, for use in the treatment of an ocular condition or disease, viz. a condition affecting the eye of a subject, by ocular topical application of said gel to the subject.

In some embodiments, the ocular condition or disease is linked to, e.g., driven or aggravated by, oxidative stress in the eye. In some embodiments, the disorder is caused or driven by elevated NOX4 and/or NOX2 activity in the eye. In some embodiments, the disorder is caused or driven by elevated NOX4 activity in the eye. In some embodiments, the disorder is caused or driven by elevated NOX2 activity in the eye.

The ocular condition (or disorder or disease) may be selected from for example, AMD, glaucoma, retinopathy, retinal ischemia-reperfusion injury, cataract, corneal diseases and conditions, bullous keratopathy, corneal abrasion, herpetic eye disease, iridocorneal endothelial syndrome, keratoconjunctivitis, and pterygium.

In some embodiments, the ocular condition is the result of an insult or trauma to the eye, such as alkali or acid burn or any other form of injury (cut or other forces).

In some embodiments, the ocular condition is neovascularization of the eye, e.g. corneal or retinal neovascularization. In some embodiments, the ocular condition is corneal neovascularization, such as due to, for example, contact lens wear, ocular surface disease, previous eye surgery, ocular trauma, ocular acid burn, ocular alkali burn, or herpes infection.

In some embodiments, the ocular condition is scar formation, e.g., as a result of an insult or trauma to the eye, such as alkali or acid burn or any other form of injury (cut or other forces).

Thus, in some embodiments, the treatment is preventive after, for example, an alkali burn or other insult to the eye, to avoid or reduce neovascularization and/or scar formation in the eye.

In some embodiments, the ocular condition is selected from AMD, glaucoma, retinopathy, retinal ischemia-reperfusion injury, cataract (including post-operative capsular opacification), and corneal diseases and conditions.

In some embodiments, the ocular conditions is AMD. In some embodiments, the ocular condition is glaucoma. In some embodiments, the ocular condition is retinopathy. In some embodiments, the ocular condition is retinal ischemia-reperfusion injury. In some embodiments, the ocular condition is cataract (including post-operative capsular opacification). In some embodiments, the ocular condition is a corneal disease or condition. In some embodiments, the disorder is an ocular inflammatory disease, e.g., a chronic inflammation of the eye. In some embodiments, the disorder is a hypertensive eye disease, such as hypertensive retinopathy.

The AMD may be dry or wet AMD. In some embodiments, the AMD is dry AMD. In some embodiments, the AMD is wet AMD.

The glaucoma may be open-angle glaucoma or angle-closure glaucoma. In some embodiments, the glaucoma is open-angle glaucoma.

The retinopathy may be, for example, retinopathy of prematurity, hypertensive retinopathy, central serous retinopathy, or diabetic retinopathy. In some embodiments, the retinopathy is hypertensive retinopathy or diabetic retinopathy. In some embodiments, the retinopathy is hypertensive retinopathy. In some embodiments, the retinopathy is diabetic retinopathy. The cataract may be, for example, a nuclear cataract, a cortical cataract, a posterior subcap- sular cataract, or a congenital cataract. In some embodiments, the cataract is a nuclear cataract, a cortical cataract, or a posterior subcapsular cataract. In some embodiments, the cataract is a nuclear cataract, or a cortical cataract. In some embodiments, the cataract is a nuclear cataract. In some embodiments, the cataract is a cortical cataract.

Cataracts also may develop as a secondary condition in patients suffering from glaucoma or diabetes, and in some embodiments, the treated patient is one suffering from glaucoma and/or diabetes. In some further embodiment, the cataract is a re-opacification of the lens post cataract surgery.

The corneal disease or condition may be, for example, keratitis (infectious or non-infectious), corneal ectasia, such keratoconus, corneal dystrophy, such as Fuchs dystrophy, epithelial basement membrane dystrophy, lattice corneal dystrophy or granular corneal dystrophy. In some embodiments, the corneal disease or condition is selected from keratitis, and corneal dystrophy. In some embodiments, the corneal disease is keratitis. In some embodiments, the keratitis is non-infectious. In some embodiments, the corneal disease is corneal dystrophy. In some embodiments, the corneal dystrophy is Fuchs dystrophy. In some embodiments, the corneal disease is corneal ectasia. In some embodiments, the corneal ectasia is keratoconus.

Use of the gel formulation

The gel formulation provided herein will be topically administered to the eye of a subject, e.g., from a dosage container allowing for applying a small volume of the formulation to the eye, to allow for administration to an eye of e.g., 1-10 drops (or droplets) having a drop/drop- let volume of 10-100 pl, or 10-50 pl, or 10-40 pl, e.g., 20-40 pl. In some embodiments, the formulation is applied from a single-dose container, or a single-use container. In some embodiments, the formulation is applied from a multidose container.

The therapeutically effective amount of the API may lie within the range of from 0.05-4.0 mg per administration (or an equivalent amount of a pharmaceutically acceptable salt of the API). In some embodiments, the therapeutically effective amount of the API is 0.05-2.0 mg per administration. In some embodiments, the therapeutically effective amount of the API is about 0.05 mg per administration. In some embodiments, the therapeutically effective amount of the API is about 0.1 mg per administration. In some embodiments, the therapeutically effective amount of is about 0.5 mg per administration. In some embodiments, the therapeutically effective amount of the API is at least 0.05 mg/day.

Preferably, the formulation provided herein will be periodically administered 1-6 times a day, e.g., 1-5 times a day, 1-3 times a day, or 1-2 times a day. In some embodiments, the periodic administration is once a day. In some embodiments, the periodic administration is twice a day. In some embodiments, the periodic administration is three times a day. In some embodiments, the periodic administration is once every 2 days. In some embodiments, the formulation is administered once a week.

In some embodiments, the formulation is administered once daily for a period of 2 to 14 days, or for a longer period, e.g., for a period of 1 month to 6 months, 2-6 months, or 3-6 months, e.g., up to 12 months, up to 2 years, up to 5 years, or even up to 10 years, or even longer. In some embodiments, the formulation is administered once a day for a period of 3 days. In some embodiments, the formulation is administered once daily for a period of 5 to 14 days. In some embodiments, the formulation is administered once daily for a period of 10 to 14 days. In some embodiments, the formulation is administered once daily for about 7 days. In some embodiments, the formulation is administered for a period of 1-12 months, or for a period of 1-6 months, or for a period of 1-3 months, e.g., once a week for a period of 3-6 months. The precise dosage regiment and length of the treatment period however will normally be decided upon by the treating physician.

A further aspect is a dosage container, containing the gel formulation as provided herein. The dosage container may be such as to include integral means to allow for administering a suitable dosage of the formulation to the eye of a patient, or such means may be provided separately. In some embodiments, the dosage container is a multidose container, allowing for the application of appropriate doses of the formulation to the eye of a patient. For example, in some embodiments, the dosage container is a bottle of the type sold by Nemera. In some embodiments, the dosage container is a Novelia® PFMD bottle or a bottle of similar type. In some of these embodiments, the formulation provided herein is preservative-free.

A further aspect is a kit (which may also be referred to as a kit-of-parts), comprising a dosage container as disclosed herein, and instructions for use. In some embodiments, such a kit also includes one or more additional containers, containing further appliances or materials useful in connection with the administration of the formulation, such as rinses, wipes, separate dosage means etc.

The ocular formulation disclosed herein is useful for the treatment of ocular diseases as mentioned herein, e.g., ocular diseases linked to N0X4 and/or N0X2 activity.

EXAMPLES

The invention is illustrated by the following non-limiting examples. Biological test results are illustrated in FIGURES 1-23, with significance indicated as follows: * p < 0.05; ** p < 0.01 ; *** p < 0.001 , compared to control tissues; # p < 0.05; ## p < 0.01 ; ### p < 0.001 , compared to AM PA or diabetic non-treated tissues.

EXAMPLE 1

A gel formulation (50 g) was prepared having the composition indicated in TABLE 2.

TABLE 2

One (1) tablet of PBS was added to 400 ml of distilled water followed by mixing with an agitator until complete dissolution of the tablet, to obtain a 0.5X PBS solution (a solution containing 68.5 mM NaCI, 1.35 mM KOI, 5 mM Na₂HPO₄, and 0.9 mM KH₂PO₄).

Separately, the API (4-bromo-2,6-dichloro-/V-[2-(2-methylphenyl)ethyl]benzene-1 -sulfonamide, compound J) (500 mg), in the form of a powder, was admixed with 500 pl of DMSO and stirred until a colourless solution was obtained. To the solution, Tween® 80 (2.5 ml) was added and the solution was stirred for additionally 10 minutes. PBS 0.5X (40 ml) was added followed by stirring until complete homogenization. To the homogeneous suspension (no visible particles), CMC HV (600 mg) was added slowly followed by stirring for 10-15 minutes until complete homogenization. Finally, a sufficient volume of PBS 0.5X was added to bring the total weight of the solution to 50 g, and stirring was performed for about 2.5-3 h at room temperature (RT) until a homogenous, white gel was obtained. The pH, osmolality, dynamic viscosity, and amount of API (as a % of theoretical value) were determined at the day of preparation of the gel (day 0), and after 7 and 32 days of storage of the gel formulation, at 5 ± 3 °C and 20 ± 5 °C (RT), respectively. The results are reported in TABLE 3.

TABLE 3

representative due to error in handling.

The concentration of the API was determined by Reverse Phase LC-UV using an HPLC Agilent Eclipse 3.5 pm x 2.1 x 50 mm column, at the following conditions: flow: 0.9 ml/min, column temperature: 25 °C, stop time: 4 min, mean pressure 1.2 x 10⁴ kPa (min. 0 kPa, max. 4 x 10⁴ kPa), rack temperature: 20 °C, injection volume: 10 pl and, as mobile phase: A. trifluoroacetic acid 0.05 % (aq.), B. acetonitrile I trifluoroacetic acid (99.95 /0.05), (A:B 50:50). For injection, 1 g of the gel formulation was diluted in 20 ml of a 50:50 mixture of A and B and mixed for 45 minutes, whereafter 1 ml of the mixture was further diluted in 10 ml of the 50:50 mixture of A and B. UV detection was performed at 254 nm. The results of the analysis indicate that the gel formulation is stable for at least 32 days, at both 5 ± 3 °C and room temperature, and the measured values of the parameters osmolality and dynamic viscosity remain acceptable during the entire test period. No precipitation of the API was observed during the entire test period.

In some embodiments, the gel formulation provided herein contains a compound of formula (I) as defined herein or a pharmaceutically acceptable salt thereof, as a suspension in a gel carrier. In such embodiments, an advantageous feature of the gel formulation of the invention is the homogeneity of the obtained suspension and high stability against sedimentation. EXAMPLE 2

A gel formulation (50 g) was prepared using the ingredients listed in TABLE 4.

TABLE 4

The API was admixed with DMSO and the mixture was subjected to magnetic stirring for 15 minutes, to give a colorless solution. Tween® 80 was added to the solution followed by magnetic stirring for 10 minutes, to give an oily and light yellowish solution. The PBS solution (about 80 % of the final volume) was added, followed by further magnetic stirring for 10 minutes, giving a white oily suspension, with no visible particles. The CMC HV was admixed, with magnetic stirring for 15 minutes, to give a white oily gel. To the gel, the remainder of the PBS solution was added, followed by magnetic stirring for 1 h. A white oily gel with no visible particles was obtained, having a pH of 7.09, an osmolality of 348 mOsm/kg H2O and a dynamic viscosity of 588.8 mPa.s (at 25 °C and 100 rpm).

EXAMPLE 3

A gel formulation (50 g) was prepared using the ingredients listed in TABLE 5.

TABLE 5

The same procedure as in EXAMPLE 2 was followed, except for the last stirring step, which was performed for 3 h. A white oily gel with no visible particles was obtained, having a pH of 7.07, an osmolality of 355 mOsm/kg H₂O and a dynamic viscosity of 998.4 mPa.s (at 25 °C and 100 rpm).

EXAMPLE 4

New Zealand White (albino) rabbits, aged about 2-3 months, having a body weight of 2.0-2.5 kg (18 males), were obtained from “Charles River”, Granja San Bernardo S.L., in Spain. All animals were ear-tagged at their arrival and the identification numbers were also marked in ears using indelible ink following the inclusion examination. The animals were held in observation for 7 days following their arrival and were observed each day for signs of illness, whereby particular attention was paid to their eyes. The animals were housed individually or two by two in standard cages. All animals were housed under identical environmental conditions at a temperature of 18 ± 3°C and a relative humidity of 45-80%, and continuous ventilation. Animals were continuously exposed (in-cage) to 10-200 lx light in a 12-hour light (from 7:00 a.m. to 7:00 p.m.) and darkness-controlled cycles. Throughout the study, animals had free access to food (approximately 90 g/day) and water.

At the day of the experiment, 15 animals were selected based on good health and homogeneous body weight (mean body weight ± 20 %) and having no visible ocular defects. The animals were randomized into the study groups based on body weight. A single droplet (50 pl) of gel formulation of EXAMPLE 1 was instilled in each eye of the animals at time point t = 0 h and at the time points indicated in TABLE 6, the animals were euthanized by intravenous injection of overdosed pentobarbital following a sedation.

TABLE 6

Immediately after euthanasia, the aqueous humor and retina from both eyes were collected, weighed, internal standard (4-bromo-2,6-dichloro-/V-[2-(2-hydroxyphenyl)ethyl]benzenesul- fonamide, compound Z) was added in samples, the samples were vortexed for 10 seconds and frozen and stored at a temperature of -80°C ± 15°C until assay. The content of the API was determined in samples of aqueous humor and retina by rapid resolution liquid chromatography/tandem mass spectrometry (RRLC-MS/MS), using a HALO® 2.7 pm C8 column (inner diameter 2.1 mm, length 100 mm), at the following conditions: flow: 0.2 ml/min, column temperature: 20 °C, stop time: 9 min, mean pressure 1.28 x 10⁴ kPa (min. 10³ kPa, max. 4 x 10⁴ kPa), rack temperature: 20 °C, injection volume: 10 pl, and gradient elution (mobile phase: A. formic acid 0.1 (aq.), B. acetonitrile, A:B 10:90 to 50:50). The Agilent G6410B Triple Quadrupole System was used for the detection, with the parameters as indicate in TABLE 7.

TABLE 7

Calculation of C_max (the highest mean value measured (ng/g for retina or ng/g of aqueous humor) and Tmax (time-point of the highest mean value) was performed using the mean data of each treatment group. Results are shown in TABLE 8.

TABLE 8

The API was quantified in the aqueous humor until 8 h post dose. At this timepoint, 3 values out of 6 were still above the lower limit of quantification (LLOQ), 5 ng/g of aqueous humor. The Tmax observed was at 0.50 h with a corresponding Cmax of 236 ng/g of aqueous humor. In one (1) eye out of 30, the measured value was higher than the upper limit of quantification (IILOQ), 523 ng/g of aqueous humor, viz. 4315 ng/g at time-point 0.5h. This value considered to be an atypical value and was not included in the calculations.

In the retina, the API remained higher than the LLOQ (17 ng/g of retina) in all samples at 8 h post dose. The T_max observed was at 0.50 h with a corresponding C_max of 1561 ng/g of retina. For 3 eyes out of 30, measured values were higher than the IILOQ (1 709 ng/g of retina) but were relatively close to the IILOQ value and therefore were extrapolated. The T_max observed was at 0.50 h with a corresponding C_max of 915 ng/g of aqueous humor.

EXAMPLE 5

An experimental protocol essentially as described by Dionysopoulou, 2023, was followed. Briefly, diabetes induced alterations in rat retina were obtained by administration of strepto- zotocin (STZ). Starting 48 hours after the administration of the STZ, the gel formulation of EXAMPLE 1 was topically administrated (20 pl) to the rats, once daily, for 14 days. Control animals, as well as diabetic non treated animals, received 20 pl of a similar gel formulation, except for containing no API. The test results of EXAMPLE 5 were compared to those of similar tests, performed using the API (10 mg/ml) in DMSO as a vehicle.

In both series of tests, tissue was collected 24 hours after the final day of treatment and were further processed either for immunohistochemical studies or for ELISA analysis. The immunohistochemical studies included the use of antibodies raised against brain nitric oxide synthase (bNOS), neurofilament (NFL), cleaved caspase-3, glial fibrillary acidic protein (GFAP), and ionized calcium-binding adapter molecule 1 (lba-1). Additionally, ELISA analysis was performed for the determination of the protein levels of TNF-a and VEGF in rat retina. The results obtained in EXAMPLE 5 and in corresponding tests using the DMSO vehicle, are illustrated in FIGS. 1-14.

The results show that the API, when administered to the eye of the rats, by topical administration of either a gel according to the invention or a solution in DMSO, was able to limit the diabetes induced oxidative damage, apoptosis, loss retinal ganglion cells axons, activation of macro/microglia, as well as the expression of the pro-inflammatory cytokine TNF-a (FIGS. 1- 12). The API also reduced loss of amacrine cells, which was surprising, since previously published studies suggested that NOX4 inhibition did not have any effect on amacrine cells in retina (Dionysopoulou, 2020). The protective effect on amacrine cells was improved when the API was administered in gel formulation (FIG. 1), compared to administration as a DMSO solution (FIG. 2). Moreover, the results showed that the API had a substantial effect to reduce the VEGF expression when administered by use of the gel formulation of the invention (FIG. 13), whereas the effect of the DMSO formulation was negligible (FIG. 14). This difference is surprising and remarkable, especially since VEGF has been recently identified as a primary initiator of proliferative diabetic retinopathy, and as a potential mediator of non-prolif- erative retinopathy (Aiello, 2000).

EXAMPLE 6

A gel formulation (50 g) is prepared having the composition indicated in TABLE 9.

TABLE 9

The gel formulation is prepared essentially as described in EXAMPLE 1, but substituting 4- hydroxy-/V-[2-(2-hydroxyphenyl)ethyl]-2,6-dimethyl-benzenesulfonamide (compound AF) for compound J.

EXAMPLE 7

Adult Sprague-Dawley rats were employed in an in vivo model of diabetic retinopathy as described by Dionysopoulou (2020), by intravitreal injection of (RS)-a-amino-3-hydroxy-5-me- thyl-4-isoxazolepropionic acid hydrobromide (AMPA) with or without 4-hydroxy-/V-[2-(2- hydroxyphenyl)ethyl]-2,6-dimethyl-benzenesulfonamide (Compound AF). At the end of the 24-hour test period, the animals were euthanized, and retinal tissues were collected and used in immunohistochemical studies (brain nitric oxidase synthetase (bNOS), glial fibrillary acidic protein (GFAP), and ionized calcium-binding adaptor molecule-1 (I ba-1 )). Quantification was performed using two-tailed unpaired t-test or one-way ANOVA analysis, followed by Newman-Keuls post hoc analysis. The result show that compound AF protected the bNOS positive amacrine cells against AMPA induced excitotoxicity (FIG. 15) and reversed the AMPA induced overactivation of both macroglia (FIG. 16) and microglia (FIG. 17). EXAMPLE 8

Adult Sprague-Dawley rats were employed in an in vivo model of diabetic retinopathy as described by Dionysopoulou (2020), by intravitreal administration of streptozotocin, followed by daily topical ocular administration (as eye drops) of a DMSO solution of compound AF (10 mg/ml, 20 pl/eye) for 14 days. At the end of the 14-day test period, the animals were euthanized, and retinal tissues were collected and used in immunohistochemical studies (bNOS, GFAP, lba-1), and Western blot analysis. Quantification was performed using two- tailed unpaired t-test or one-way ANOVA analysis, followed by Newman-Keuls post hoc analysis.

The results show that compound AF did not afford protection to the axons of ganglion cells against the diabetic insult (FIG. 18), but limited the diabetes induced loss of bNOS positive amacrine cells (FIG. 19), caused the upregulation of Bcl-2 protein in the diabetic treated retinas (FIG. 20), and reduced the diabetes induced activation of macroglia (FIG. 21) and microglia (FIG. 22), respectively.

EXAMPLE 9

The protocol described in EXAMPLE 8 is repeated, using a daily topical ocular administration (as a 20-pl eye drop) of the formulation of EXAMPLE 6, instead of the DMSO solution.

EXAMPLE 10

The gel formulation of EXAMPLE 1 was used in a rabbit model of blood retinal barrier breakdown by VEGF-induced vascular leakage. The gel formulation was topically administered (instillation) to the rabbit eyes and, for comparative purposes, a glucocorticoid Kenacort® Retard or a fusion protein aflibercept (Eylea®) was administered by intravitreal injection. Kenacort® Retard is a general anti-inflammatory agent having several medicinal functions to lower inflammatory responses, whereas Eylea® binds to circulating VEGFs like a VEGF trap. Both Kenacort® Retard and Eylea® are in clinical use for the treatment of diabetic retinopathy.

Throughout the procedure, animals were anesthetized by an intramuscular injection of a mix Rompun® (xylazine) and Imalgene® (ketamine) before intravitreal administration, induction, fluorophotometry or euthanasia. A topical anesthesia (a drop of 0.4% oxybuprocaine solution) was instilled on the cornea for local anesthesia before intravitreal administration, induction, or fluorophotometry. Eye pupils were dilated by instillation of one drop of 0.5% tropicamide and of 10% phenylephrine before intravitreal administration, induction, or fluorophotometry.

The animals received treatment with the gel formulation of the invention (EXAMPLE 1 , a corresponding gel formulation without API, Kenacort® Retard, and Eylea®, respectively) as outlined in TABLE 10.

TABLE 10

Increase in retinal vascular permeability was induced on Day 0 for all groups by a single 50 pl intravitreal injection of 500 ng rhVEGF165 with carrier protein (diluted in PBS) into the right eye, using a Hamilton syringe (30-G needle). The injection was performed using a microscope on anesthetized animals.

On Day 2, sodium fluorescein (10% in saline solution 0.9%, 50 mg/kg) was injected via the marginal ear vein in test animals. Following fluorescein injection, animals were anesthetized, and pupils were dilated. Measurement of ocular fluorescence was performed with an FM-2 Fluorotron™ Master ocular fluorophotometer in both eyes. A series of 148 scans with a step size of 0.25 mm were recorded from the cornea to the retina along the optical axis. The results are illustrated in FIG. 23. REFERENCE LIST

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Claims

1. A gel formulation for ocular topical use, comprising a compound of formula (I)

or a pharmaceutically acceptable salt thereof, wherein

Ri is selected from C1-C6 alkyl, C3-C6 cycloalkyl, C1-C6 alkoxy, hydroxy, and halogen; each is independently selected from C1-C6 alkyl, C1-C6 alkoxy, hydroxy, and halogen;

R2 is selected from C1-C6 alkyl, C1-C6 alkoxy, halogen, hydroxy, and hydroxy-C1-C3 alkyl;

R3, R4, Rs, and Re are independently selected from H and F; and any alkyl is optionally substituted with one or more halogens.

2. The gel formulation according to claim 1 , wherein

R1 is selected from C1-C3 alkyl, C1-C3 alkoxy, hydroxy, and halogen; each Ria is independently selected from C1-C3 alkyl, and halogen;

R2 is selected from C1-C3 alkyl, halogen, and hydroxy;

R3, R4, Rs, and Re are independently selected from H and F; and any alkyl is optionally substituted with one or more fluoro.

3. The gel formulation according to claim 1 , wherein the compound of formula (I) is selected from:

/V-[2-(2-methoxyphenyl)ethyl]-2,4,6-trimethylbenzene-1 -sulfonamide;

/V- [2-(2-fluorophenyl)ethyl]-2, 4, 6-trimethylbenzene-1 -sulfonamide;

4-bromo-2,6-dichloro-/V-[2-(2-methoxyphenyl)ethyl]benzene-1 -sulfonamide;

4-bromo-2,6-dichloro-/V-[2-(2-fluorophenyl)ethyl]benzene-1 -sulfonamide;

/V-[2-(2-chlorophenyl)ethyl]-2,4,6-trimethylbenzene-1-sulfonamide;

/V- [2-(2-bromophenyl)ethyl]-2, 4, 6-trimethylbenzene-1 -sulfonamide;

2.4.6- trimethyl-/V-{2-[2-(trifluoromethyl)phenyl]ethyl}benzene-1 -sulfonamide;

2.4.6-trimethyl-/V-[2-(2-methylphenyl)ethyl]benzene-1-sulfonamide;

2.4.6-trimethyl-/V-{2-[2-(trifluoromethoxy)phenyl]ethyl}benzene-1 -sulfonamide;

4-bromo-2,6-dichloro-/V-[2-(2-methylphenyl)ethyl]benzene-1 -sulfonamide;

4-bromo-2,6-dichloro-/V-{2-[2-(trifluoromethyl)phenyl]ethyl}benzene-1 -sulfonamide;

4-bromo-2,6-dichloro-/V-[2-(2-chlorophenyl)ethyl]benzene-1 -sulfonamide;

2.6-dichloro-4-cyclopropyl-/V-[2-(2-fluorophenyl)ethyl]benzene-1-sulfonamide; 2.6-dichloro-/V-[2-(2-chlorophenyl)ethyl]-4-cyclopropylbenzene-1 -sulfonamide;

2.6-dichloro-/V-[2-(2-chlorophenyl)ethyl]-4-(trifluoromethyl)benzene-1 -sulfonamide;

N-[2, 2-difluoro-2-(2-methylphenyl)ethyl]-2, 4, 6-tri methyl benzene- 1 -sulfonamide;

4-bromo-2,6-dichloro-/V-[2,2-difluoro-2-(2-methylphenyl)ethyl]benzene-1 -sulfonamide;

/\/-[2-(2-chlorophenyl)-2, 2-difluoroethyl]-2, 4, 6-trimethylbenzene-1 -sulfonamide;

4-bromo-2,6-dichloro-/V-[2-(2-chlorophenyl)-2,2-difluoroethyl]benzene-1 -sulfonamide; /\/-[2-(2-chlorophenyl)ethyl]-2,6-dimethyl-4-(propan-2-yl)benzene-1 -sulfonamide;

2.6-dimethyl-/V-[2-(2-methylphenyl)ethyl]-4-(propan-2-yl)benzene-1-sulfonamide; /V-[2-fluoro-2-(2-methylphenyl)ethyl]-2,4,6-trimethylbenzene-1-sulfonamide;

4-bromo-2,6-dichloro-/V-[2-fluoro-2-(2-methylphenyl)ethyl]benzene-1 -sulfonamide; /V-[2-fluoro-2-(2-methylphenyl)ethyl]-2,6-dimethyl-4-(propan-2-yl)benzene-1 -sulfonamide; /\/-[2-(2-hydroxyphenyl)ethyl]-2, 4, 6-trimethylbenzene-1 -sulfonamide;

4-bromo-2,6-dichloro-/V-[2-(2-hydroxyphenyl)ethyl]benzenesulfonamide;

2.6-dichloro-/V-[2-(2-hydroxyphenyl)ethyl]-4-(trifluoromethyl)benzenesulfonamide;

2.4-dichloro-6-hydroxy-/V-[2-(2-hydroxyphenyl)ethyl]benzenesulfonamide;

2.4-dichloro-6-hydroxy-/V-[2-(o-tolyl)ethyl]benzenesulfonamide;

/V-[2-(2-chlorophenyl)ethyl]-4-methoxy-2,6-dimethyl-benzenesulfonamide;

/V-[2-(2-hydroxyphenyl)ethyl]-4-methoxy-2,6-dimethyl-benzenesulfonamide; and 4-hydroxy-/V-[2-(2-hydroxyphenyl)ethyl]-2,6-dimethyl-benzenesulfonamide.

4. The gel formulation according to claim 3, wherein the compound of formula (I) is 4-bromo-

2.6-dichloro-N-[2-(2-methylphenyl)ethyl]benzene-1 -sulfonamide.

5. The gel formulation according to any one of claims 1 to 4, having a pH in the range of 6.8 to 8.0, an osmolality in the range of 200 to 600 mOsm/kg H2O, and a dynamic viscosity, measured at 100 rpm and 25 °C, in the range of 500 to 2000 mPa.s,

6. The gel formulation according to claim 5, wherein the pH is in the range of 6.8 to 7.5, the osmolality is in the range of 250 to 400 mOsm/kg H2O, and/or the dynamic viscosity is in the range of 700 to 1700 mPa.s.

7. The gel formulation according to any one of claims 1 to 6, wherein the compound of formula (I), or the pharmaceutically acceptable salt thereof, is present at a concentration of from 0.1 to 5 % by weight of the formulation.

8. The gel formulation according to any one of claims 1 to 7, comprising a solubilizer, a buffer, and a viscosity agent.

9. The gel formulation according to claim 8, wherein the solubilizer comprises dimethylsulfoxide and a non-ionic surfactant, the buffer is phosphate-buffered saline, and/or the viscosity agent is a cellulose derivate.

10. The gel formulation according to claim 9, wherein the non-ionic surfactant is a polyoxyethylene sorbitan monooleate.

11. The gel formulation according to any one of claims 8 to 10, wherein the viscosity agent is carboxymethyl cellulose.

12. The gel formulation as defined in any one of claims 1 to 11 , for use in the prevention or treatment of an ocular disorder in a mammal patient.

13. The gel formulation for use according to claim 12, wherein the ocular disorder is selected from age-related macular degeneration, glaucoma, retinopathy, retinal ischemia-reperfusion injury, cataract, corneal diseases, bullous keratopathy, corneal abrasion, corneal alkali or acid burn, corneal neovascularization, herpetic eye disease, iridocorneal endothelial syndrome, keratoconjunctivitis, pterygium, and eye inflammation.

14. The gel formulation for use according to claim 12 or 13, wherein the treatment comprises periodic dropwise application of the gel into an eye of the mammal patient.

15. A dosage container for topical ocular administration containing a gel formulation as defined in any one of claims 1 to 14.

16. Use of a gel formulation as defined in any one of claims 1 to 14, in the manufacture of a medicament for prevention or treatment of an ocular disorder in a mammal patient.

17. The use according to claim 16, wherein the ocular disorder is selected from age-related macular degeneration, glaucoma, retinopathy, retinal ischemia-reperfusion injury, cataract, corneal diseases, bullous keratopathy, corneal abrasion, corneal alkali or acid burn, corneal neovascularization, herpetic eye disease, iridocorneal endothelial syndrome, keratoconjunctivitis, pterygium, and eye inflammation.

18. A method for prevention or treatment of an ocular disorder by administering, to a mam- mal in need of such prevention or treatment, a therapeutically effective amount of a gel formulation as defined in any one of claims 1 to 14.

19. The method according to claim 18, wherein the ocular disorder is selected from age-related macular degeneration, glaucoma, retinopathy, retinal ischemia-reperfusion injury, cata- ract, corneal diseases, bullous keratopathy, corneal abrasion, corneal alkali or acid burn, corneal neovascularization, herpetic eye disease, iridocorneal endothelial syndrome, keratoconjunctivitis, pterygium, and eye inflammation.