Sitagliptin for use in retinal diseases with neovascularization

Abstract

The present invention relates to sitagliptin or a pharmaceutically or veterinary acceptable salt thereof for use in the local eye treatment of neovascular retinal diseases. The invention also encompasses pharmaceutical or veterinary compositions for use in the local treatment of these diseases.

Description

[0001] Sitagliptin for use in retinal diseases with neovascularization

[0002] This application claims the benefit of European Patent Application 24383099.9 filed October 9th, 2024.

[0003] Technical Field

[0004] The present invention relates to the field of medical approaches for ocular diseases that may lead to partial or total blindness. The invention provides a treatment with sitagliptin or a pharmaceutically or veterinary acceptable salt thereof to be applied locally in the eyes.

[0005] Background Art

[0006] Diabetic retinopathy (DR hereinafter) is one of the most frequent chronic complications of diabetes and still represents the leading cause of preventable blindness in working-age population. This retina disease is caused by chronic hyperglycemia (high blood sugar) in diabetic patients, which triggers metabolic pathways that damage the retinal blood vessels, leading to the slow progression of the DR. DR can be differentiated in the early and the advanced stages.

[0007] In the early stages of diabetic retinopathy (ESDR hereinafter), the eye is characterized by the presence of mild retinal lesions such as microvascular abnormalities like microaneurysms (weak spots in blood vessels), microhemorrhages (tiny bleeds), and hard exudates (protein deposits), and the visual acuity is generally preserved.

[0008] At present, the common clinical recommended options for the treatment of ESDR are the optimization of the metabolic control by, for example, a strict glycemic control, and the maintenance of a tight control of blood pressure. There are pharmacological options for the glycemic control such as dipeptidyl peptidase-IV (DPP-IV) inhibitors, which are a class of drugs used by their blood sugar control effect when systemically administered, and which may accordingly have a positive effect on ESDR. However, as shown in WO2017186934A1, it has been recently proposed DPP-IV inhibitors to have a direct and beneficial effect in the treatment of ESDR when locally administered on the eye topically, without altering the blood glucose levels.

[0009] The ESDR tends to progress towards the advanced stages of diabetic retinopathy (ASDR hereinafter) in which new blood vessels abnormally grow, the structural lesions are significant and the visual acuity is compromised. ASDR can be differentiated in two stages, which are not mutually exclusive. One of them is the diabetic macular edema (DME hereinafter), the other is the proliferative diabetic retinopathy (PDR hereinafter).

[0010] Tight control of blood glucose levels and blood pressure is also recommended in ASDR, but the beneficial effects are less evident than when these general measures are implemented in the beginning of the disease or when ESDR appears. But, by contrast, there are indeed options in clinical practice for the treatment of ASDR. These include laser photocoagulation, intravitreal injections of corticosteroids, intravitreal injections of anti- vascular endothelial growth factor (herein VEGF) agents, and vitrectomy (surgery to remove vitreous gel from the eye). However, these treatments are aggressive, invasive, expensive, and with significant adverse effects.

[0011] The topical administration of DPP-IV inhibitors as indicated in the above-mentioned patent document is demonstrated to be useful for treating diseases in which neurodegeneration plays an essential role. Since neurodegeneration is crucial in the pathogenesis of ESDR there is no doubt of the potential role of DPP-IV inhibitors administered in eye drops in these early stages. However, when DR progresses towards ASDR such as PDR in which ischemia and neovascularization are the main pathogenic drivers, the role of DPP-IV inhibitors was expected to be completely marginal or, considering the reported angiogenic / neovascularization effect of DPP-IV inhibitors such as sitagliptin, even negative or detrimental (Kolibabka, M. et al. (2018); Gongalves, A., et al. (2018); Lee, OS., et al. (2016)).

[0012] In fact, anti-angiogenic drugs are promising for treating the ASDR, as they help reduce existing fragile vessels and prevent new ones from forming. Thus, they reduce the risk of bleeding, reduce inflammation of the macula and protect against vision or visual acuity loss.

[0013] For example, anti-angiogenic drugs slow the growth of abnormal blood vessels and reduce the risk of particular complications like vitreous hemorrhage (bleeding in the eye) and / or fractional retinal detachment, thus they are interesting for treating PDR. In addition, as these treatments prevent the growth of neovessels, which are fragile and leaking, they are also useful for the treatment of DME, as well as other retinal diseases in which angiogenesis plays a predominant role, such as the ‘wet’ stage of age-related macular degeneration.

[0014] However, there still remain a significant unmet medical need for the treatment of neovascular retinal diseases, and the development of novel and less aggressive and invasive therapeutic strategies is crucial for improving the quality of life and preventing vision loss in patients. Summary of Invention

[0015] Surprisingly, the inventors found that sitagliptin or a pharmaceutically or veterinary acceptable salt thereof, contrary to the teachings of the above-mentioned prior art teachings, presents anti-angiogenic activity when administered locally in the eye.

[0016] Neovascular diseases are intrinsically linked to angiogenesis, the process of new blood vessel formation.

[0017] As shown in Example 1 , the inventors demonstrated in in vitro experiments that sitagliptin exerted anti-angiogenic effects by decreasing the cell migration rate and tube formation in HRECs.

[0018] The inventors furthermore demonstrated in Example 2 that sitagliptin was able to inhibit and regress angiogenesis (neovascular sprouts) in retinal explants obtained from diabetic rats treated with sitagliptin eye drops before euthanasia, as well as when sitagliptin was administered directly to retinal explants, mimicking intravitreal injection.

[0019] The inventors furthermore demonstrated in Example 3 that topical administration of sitagliptin reduces laser-induced choroidal neovascularization (LI-CNV) in both diabetic and non-diabetic mice. Also, it is demonstrated that the neovascular area was significantly greater in LI-CNV vehicle-treated mice compared to LI-CNV sitagliptin-treated mice in both diabetic and non-diabetic mice.

[0020] Accordingly, and as shown below in the examples, the inventors herein present sitagliptin or a pharmaceutically or veterinary acceptable salt thereof as an anti-angiogenic drug, useful in the treatment of retinal neovascular diseases.

[0021] Thus, a first aspect of the invention is sitagliptin or a pharmaceutically or veterinary acceptable salt thereof, for use in the local eye treatment of retinal neovascular diseases.

[0022] Of note is that the present invention in all of its aspects is directed specifically to the treatment of established neovascular retinal disease (i.e. it is not directed to its prevention).

[0023] The invention provides, in a second aspect, a pharmaceutical or veterinary composition for use in the local eye treatment of a neovascular retinal disease, which comprises a therapeutically effective amount of sitagliptin or a pharmaceutically or veterinary acceptable salt thereof as defined in the first aspect, together with one or more pharmaceutically or veterinary acceptable excipients.

[0024] Brief Description of Drawings

[0025] Fig. 1A shows the kinetic scratch wound healing migration of HRECs cells (human retinal endothelial cells, hereinafter).

[0026] Representative images of endothelial cell migration in HRECs treated with sitagliptin (S) 200 pM or vehicle (C) in normoglucose 5 mM (NG) conditions at baseline (TO), 6 (T6 hs), 10 (T10 hs) and 12 (T12 hs) hours of treatment.

[0027] Fig. 1 B shows the kinetic scratch wound healing migration of HRECs cells. Representative images of endothelial cell migration in HRECs treated with sitagliptin (S) 200 pM or vehicle (C) in high glucose 30 mM (HG) conditions at baseline (TO), 6 (T6 hs), 10 (T10 hs) and 12 (T12 hs) hours of treatment.

[0028] Fig. 1C shows the kinetic scratch wound healing migration of HRECs cells. Quantification of endothelial cell migration in HRECs in 5 mM glucose (G) conditions at 6, 10 and 12 hours of treatment. X-axis represents treatment duration in hours. Y-axis represents HREC migration cells (% over control). White bars: sitagliptin (S) 200 pM treatment; Black bars: vehicle (C). *p < 0.05 control vs. sitagliptin; ** p < 0.01 control vs. sitagliptin.

[0029] Fig. 1 D shows the kinetic scratch wound healing migration of HRECs cells. Quantification of endothelial cell migration in HRECs in 30 mM glucose (G) conditions at 6, 10 and 12 hours of treatment. X-axis represents treatment duration in hours. Y-axis represents HREC migration cells (% over control). White bars: sitagliptin (S) 200 pM treatment; Black bars: vehicle (C). *p < 0.05 control vs. sitagliptin; ** p < 0.01 control vs. sitagliptin.

[0030] Fig. 2A shows the quantification of tube formation in HRECs under the exposure to different treatments.

[0031] Y-axis represents number (Nb) of junctions I mm2. X-axis represents treatment duration in hours.

[0032] Black bars: HRECs treated with VEGF; Dark Gray bars: HRECs treated with VEGF + sitagliptin; Light Gray bars: HRECs treated with control (without VEGF) + sitagliptin; White bars: HRECs treated with vehicle. For each treatment duration, from left to right, bars are Black, Dark Gray, Light Gray and White.

[0033] Imaged software with the Angiogenesis Analyzer tool (version 1 ,53p) was used to measure angiogenic parameters. Error bars indicate standard deviation. * p< 0.05; ** p< 0.01; *** p< 0.001.

[0034] Fig. 2B shows the quantification of tube formation in HRECs under the exposure to different treatments.

[0035] Y-axis represents number (Nb) of segments I mm2. X-axis represents treatment duration in hours.

[0036] Black bars: HRECs treated with VEGF; Dark Gray bars: HRECs treated with VEGF + sitagliptin; Light Gray bars: HRECs treated with control (without VEGF) + sitagliptin; White bars: HRECs treated with vehicle. For each treatment duration, from left to right, bars are Black, Dark Gray, Light Gray and White.

[0037] Imaged software with the Angiogenesis Analyzer tool (version 1 ,53p) was used to measure angiogenic parameters. Error bars indicate standard deviation. * p< 0.05; ** p< 0.01; *** p< 0.001.

[0038] Fig. 2C shows the quantification of tube formation in HRECs under the exposure to different treatments.

[0039] Y-axis represents total branching length (mm). X-axis represents treatment duration in hours.

[0040] Black bars: HRECs treated with VEGF; Dark Gray bars: HRECs treated with VEGF + sitagliptin; Light Gray bars: HRECs treated with control (without VEGF) + sitagliptin; White bars: HRECs treated with vehicle. For each treatment duration, from left to right, bars are Black, Dark Gray, Light Gray and White.

[0041] Imaged software with the Angiogenesis Analyzer tool (version 1 ,53p) was used to measure angiogenic parameters. Error bars indicate standard deviation. * p< 0.05; ** p< 0.01; *** p< 0.001.

[0042] Fig. 2D shows the quantification of tube formation in HRECs under the exposure to different treatments.

[0043] Y-axis represents total meshes I mm2. X-axis represents treatment duration in hours. Black bars: HRECs treated with VEGF; Dark Gray bars: HRECs treated with VEGF + sitagliptin; Light Gray bars: HRECs treated with control (without VEGF) + sitagliptin; White bars: HRECs treated with vehicle. For each treatment duration, from left to right, bars are Black, Dark Gray, Light Gray and White.

[0044] Imaged software with the Angiogenesis Analyzer tool (version 1 ,53p) was used to measure angiogenic parameters. Error bars indicate standard deviation. * p< 0.05; ** p< 0.01; *** p< 0.001.

[0045] Fig. 3A shows how sitagliptin mitigates choroid-retinal pigment epithelium (RPE) sprouting in ex-vivo culture. Retinal explants were obtained from non-diabetic rats and cultured under high glucose (HG) 30 mM conditions with and without sitagliptin (S) 100 pM or under normoglucose (NG) 5 mM conditions. Representative images of choroid sprouting for each condition assessed by fluorescent stain. The white arrow points to Sprouts. The scale bar of the bottom right corner represents 500 pm.

[0046] Fig. 3B shows how sitagliptin mitigates choroid-retinal pigment epithelium (RPE) sprouting in ex-vivo culture. Retinal explants were obtained from non-diabetic rats and cultured under high glucose (HG) 30 mM conditions with and without sitagliptin (S) 100 pM or under normoglucose (NG) 5 mM conditions. Y-axis quantifies the area of choroidal sprouting (mm2). X-axis represents the different condition assessed. Calculated with Imaged 1.53p software (National Institute of Health, USA). *: p< 0.05.

[0047] Fig. 4 shows how sitagliptin mitigates choroid-retinal pigment epithelium (RPE) sprouting in ex-vivo culture. Retinal explants were obtained from non-diabetic rats and cultured under high glucose (HG) 30 mM conditions with and without sitagliptin (S) 100 pM or under normoglucose (NG) 5 mM conditions. Representative brightfield images of choroid sprouting for each condition obtained using a PRG-300HDS high-definition camera (World Precision Instruments).*: p< 0.05. The white arrow points to Sprouts. The scale bar of the bottom right corner represents 1000 pm.

[0048] Fig. 5 shows how sitagliptin reduces choroid-retinal pigment epithelium (RPE) sprouting in ex-vivo cultures. Retinal explants were obtained from streptozotocin-induced diabetes (STZ)-induced diabetic rats (D-STZ) treated with vehicle (eye drops: twice a day for 2 weeks), STZ-induced diabetic rats treated with sitagliptin 100 pM (eye drops; twice a day) for 2 weeks (D-STZ-S 100 pM), and non-diabetic rats (C). Representative images of choroid sprouting for each condition assessed by fluorescent stain. The scale bar of the bottom right corner represents 500 pm.

[0049] Fig. 6 shows how sitagliptin reduces choroid-retinal pigment epithelium (RPE) sprouting in ex-vivo cultures. Retinal explants were obtained from streptozotocin-induced diabetes (STZ)-induced diabetic rats (D-STZ) treated with vehicle (eye drops: twice a day for 2 weeks), STZ-induced diabetic rats treated with sitagliptin 100 pM (eye drops; twice a day) for 2 weeks (D-STZ-S 100 pM), and non-diabetic rats (C). Y-axis quantifies the sprouting area (mm2). X-axis represents the different condition assessed. Calculated with Imaged 1.53p software (National Institute of Health, USA). *: p<0.05.

[0050] Fig. 7 shows how sitagliptin reduces choroid-retinal pigment epithelium (RPE) sprouting in ex-vivo cultures. Retinal explants were obtained from streptozotocin-induced diabetes (STZ)-induced diabetic rats (D-STZ) treated with vehicle (eye drops: twice a day for 2 weeks), STZ-induced diabetic rats treated with sitagliptin 100 M (eye drops; twice a day) for 2 weeks (D-STZ-S 100 M), and non-diabetic rats (C). Representative brightfield images of choroid sprouting for each condition obtained using a PRO-300HDS high- definition camera (World Precision Instruments). *: p<0.05. The scale bar of the bottom right corner represents 1000 pm.

[0051] Fig. 8A shows how sitagliptin reduces the area of choroidal neovascularization induced by laser (LI-CNV) in diabetic and non-diabetic mice. Fluorescence angiography obtained at day 1 and day 7 post-laser of a representative mouse from each experimental group are shown. CNV lesions are hyperfluorescent areas. The arrows are pointing to one lesion from each representative fluorescence angiography at day 1 and at day 7 post-laser. “DMV” stands for Diabetic Mouse Vehicle. “DMS” stands for Diabetic Mouse Sitagliptin. “N-DMV” stands for Non-Diabetic Mouse Vehicle. “N-DMS” stands for Non-Diabetic Mouse Sitaglitpin. “D1” stands for Day 1. “D7” stands for Day 7.

[0052] Fig. 8B shows the quantification analysis performed by measuring hyperfluorescent areas. Error bars indicate standard deviation. *: p <0.001. “ACL” stands for Area of CNV lesions. “A.U.” stands for arbitrary units. “V” stands for Vehicle. “D1” stands for Day 1 . “D7” stands for Day 7. “S” stands for Sitagliptin. “DM” stands for Diabetic Mice. “N-DM” stands for NonDiabetic Mice.

[0053] Fig. 9A shows how sitagliptin reduces the area of choroidal neovascularization induced by laser (LI-CNV) in diabetic and non-diabetic mice. Images of flat-mounted (RPE-choroid- sclera) eyecups at day 7 post-laser of a representative mouse from each experimental group are shown. Endothelial cells of vessels emerging from the choroid, stained with isolectin B4, are displayed in white color. . “V” stands for Vehicle. “S” stands for Sitagliptin. “C” stands for Control. “DM” stands for Diabetic Mice. “N-DM” stands for Non-Diabetic Mice.

[0054] Fig. 9B shows the quantification of LI-CNV lesions in the experimental groups. Error bars indicate standard deviation. Y-axis represents the area of neovascularization measured using IB4 staining in pm2. *: p<0.01. “AC” stands for Area of CNV. “V” stands for Vehicle. “S” stands for Sitagliptin. “C” stands for Control. “DM” stands for Diabetic Mice. “N-DM” stands for Non-Diabetic Mice.

[0055] Detailed description of the invention All terms as used herein in this application, unless otherwise stated, shall be understood in their ordinary meaning as known in the art. Other more specific definitions for certain terms as used in the present application are as set forth below and are intended to apply throughout the description and claims.

[0056] The term "about" or “around” as used herein refers to a range of values ± 10% of a specified value. For example, the expression "about 10" or “around 10” includes ± 10% of 10, i.e., from 9 to 11.

[0057] For “neovascular retinal diseases” is to be understood as the retinal disorders which course with neovascularization, or in other words retinal disorders with neovascularization, characterized by the abnormal growth of new blood vessels (angiogenesis) within the retina. This includes for example wet Age-Related Macular Degeneration (wARMD), ASDR, retinopathy of prematurity, retinal vein occlusion, rubeosis iridis and corneal neovascularization.

[0058] For “early stages of diabetic retinopathy” is to be understood as the stages of DR in which, due to the presence of diabetes, functional abnormalities can be detected in the eye (i.e. chromatic discrimination, contrast sensitivity and electroretinography abnormalities), but the pattern of microvascular changes of DR has not yet been fully established.

[0059] For “advanced stages of diabetic retinopathy” is to be understood as the stages of DR in which the structural lesions are significant and the visual acuity is compromised. Both by the exaggerated vascular leakage which results in the diabetic macular edema (DME), and the hypoxia / ischemia which induces an unbalance between angiogenic and anti- angiogenic factors leading to neovascularization and the so-called proliferative diabetic retinopathy (PDR).

[0060] For “proliferative diabetic retinopathy” (PDR) is to be understood as an advanced stage of diabetic retinopathy characterized by the growth of abnormal blood vessels (neovascularization) within the retina. These new vessels are fragile owing to architectural weakness, are prone to bleeding and tend to grow in the vitreous body in which they are eventually anchored by means of fibrovascular tissue. This fibrotic tissue may contract, and this can lead to advanced DR and fractional retinal detachment, accompanied by severe vision loss. Retinal detachment and vitreous haemorrhage constitute the advanced stage of DR and are sight-threatening conditions (Wong, TY, et al. (2016). Neovascularization in the retina signals the onset of PDR, with a risk of vitreous hemorrhage and traction retinal detachment, as before commented, as well as anterior segment neovascularization of the iris (rubeosis) or anterior chamber angle with intraocular pressure rise (neovascular glaucoma). It can also be referred to as “neovascular retinopathy”, “diabetic retinopathy with neovascularization” or “diabetic retinopathy with abnormal blood vessel growth”.

[0061] The presence of neovascularization within the optic disc or close to it (where the optic nerve connects to the retina), or in any area of the retina other than the optic disc is indicative of PDR. Also, vitreous or preretinal hemorrhage, i.e. bleeding from fragile new vessels in the vitreous (gel-like substance within the eye) or in front of the retina, are indicative of PDR. Additional factors supporting PDR diagnosis are severe intraretinal microvascular abnormalities (dilated, irregular, or newly formed blood vessels within the retina); venous beading (abnormal variations in the width of retinal veins appearing like a string of beads); and retinal detachment (a serious complication where scarring from abnormal blood vessels pulls the retina away from its normal position).

[0062] A PDR diagnosis is determined if neovessels and / or vitreous hemorrhage are present according to the International Clinical Diabetic Retinopathy (ICDR) Severity Scale, or the level of DR is higher than 60 in the Early Treatment of Diabetic Retinopathy Study (ETDRS) classification. The ETDRS classification is the “gold standard” for research and clinical trials (Early photocoagulation for diabetic retinopathy. ETDRS report number 9. Early Treatment Diabetic Retinopathy Study Research Group. (1991). Ophthalmology, 98(5 Suppl), 766-785). In everyday clinical practice, the ICDR Severity Scale, which in essence is a simplified ETDRS system, is a commonly used classification system (Wilkinson, C. P., Ferris, F. L., 3rd, Klein, R. E., Lee, P. P., Agardh, C. D., Davis, M., Dills, D., Kampik, A., Pararajasegaram, R., Verdaguer, J. T., & Global Diabetic Retinopathy Project Group (2003). Proposed international clinical diabetic retinopathy and diabetic macular edema disease severity scales. Ophthalmology, 110(9), 1677-1682. https: / / doi.Org / 10.1016 / S0161 -6420(03)00475-5).

[0063] For “diabetic macular edema” (DME) is to be understood as an advanced stage of diabetic retinopathy characterized by fluid accumulation in the macula, the central part of the retina responsible for detailed vision or visual acuity, causing swelling (edema), causing blurred vision, distorted vision, and loss of central vision. Macular edema is caused by leakage from damaged blood vessels in the eye. DME is diagnosed when the following three criteria are met: retinal thickening at or within 500 pm of the center of the macula; hard exudates at or within 500 pm of the center of the macula, if associated with thickening of the adjacent retina; and / or a zone (or zones) of retinal thickening one disc area in size at least part of which is within one disc diameter of the center

[0064] The term "wet age-related macular degeneration" (wARMD) refers to an advanced form of age-related macular degeneration characterized by the abnormal growth of choroidal blood vessels beneath the retina which is also known as “choroidal neovascularization”, specifically in the macula. These new vessels are prone to leaking fluid and blood, leading to retinal damage, scarring, and a rapid decline in central vision. wARMD is responsible for the majority of severe vision loss associated with age-related macular degeneration and is a result of angiogenesis and subretinal neovascularization.

[0065] The term "angiogenesis" refers to the process through which new blood vessels are formed from pre-existing vessels. This process involves the proliferation, migration, and organization of endothelial cells. Agents that promote or stimulate angiogenesis are considered angiogenic or pro-angiogenic, while those that inhibit and regress it are considered anti-angiogenic. Synonyms for "angiogenesis" include “neovascularization”, “vasculogenesis”, and terms emphasizing “vessel growth or formation”.

[0066] The term "regress" refers to blood vessels and / or vasculature (including neovasculature and / or neovessels), is used herein to mean to retract or shrink.

[0067] In the context of the invention, the term “neuroprotection” means any kind of treatment or prophylactic method that can be used in order that neurons constituting the neuroretina remain preserved and in a physiological state corresponding to the one of a health subject animal (including humans). The “neuroretina” is the part of the retina including the neurons and glial cells without the retinal pigment epithelium. Neuroretina is the responsible of the visual cycle.

[0068] The expression "therapeutically effective amount" as used herein, refers to the amount of a compound that, when administered, is sufficient to prevent development of, or alleviate to some extent, one or more of the symptoms of the disease which is addressed. The particular dose of compound administered according to this invention will of course be determined by the particular circumstances surrounding the case, including the compound administered, the route of administration, the particular condition being treated, and the similar considerations.

[0069] The term "pharmaceutically or veterinary acceptable" as used herein pertains to compounds, materials, compositions, and / or dosage forms which are, within the scope of sound medical and veterinary judgment, suitable for use in contact with the tissues of a subject (e.g. human or any other animal) without significant toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit / risk ratio. Each excipient, etc., must also be “acceptable" in the sense of being compatible with the other ingredients of the pharmaceutical composition. It must also be suitable for use in contact with the tissue or organ of humans and animals without excessive toxicity, irritation, allergic response, immunogenicity or other problems or complications commensurate with a reasonable benefit / risk ratio. Suitable excipients, etc. can be found in standard pharmaceutical texts, and include, as a way of example preservatives, agglutinants, humectants, emollients, and antioxidants.

[0070] The term “pharmaceutically or veterinary acceptable salts” encompasses any salt formed from pharmaceutically or veterinary acceptable non-toxic acids including inorganic or organic acids such as for example acetic, trifluoroacetic, benzenesulfonic, benzoic, camphorsulfonic, citric, ethansulfonic, oxalic, fumaric, gluconic, glutamic, hydrobromic, hydrochloric, orthophosphoric, lactic, maleic, malic, mandelic, methanesulfonic, phosphoric, succinic, sulfuric, tartaric, p-toluenesulfonic, glutamic, aspartic acid, and the like. There is no limitation regarding the salts, except that if used for therapeutic purposes, they must be pharmaceutically or veterinary acceptable.

[0071] “Administered locally in the eye” or “local eye administration” or “ocular administration” refers to the direct administration or application of a substance or composition in the eye. Non-limiting examples of local eye administration include “Topical eye administration”, “injectable ocular administration”, and any other administration that deliver the substance or composition in the ocular tissues. “Topical eye administration” or “topical administration in the eye” or “topical ocular administration” or “administered topically in the eye” are to be understood as the administration of a substance or composition on the surface of the eye. The surface of the eye includes the cornea, sclera, conjunctival fornix, conjunctiva, eyelids, and associated structures such as the lacrimal glands and tear ducts. “Injectable administration” or “injectable eye administration” or “injectable ocular administration” or “injected in the eye” are to be understood as the administration of a substance or composition by injection in the eye. Non-limiting examples of “injectable ocular administration” include “intravitreal injectable administration” or “subconjunctival injectable administration”. Terms related to local administration exclude systemic administration of a substance or a composition which may reach the eye through systemic circulation, such as oral medications or intravenous injections. In fact, the focus of the invention is on the local or direct administration to ensure targeted and effective treatment of ocular conditions.

[0072] For “local eye treatment” or “ocular treatment” is to be understood as the local eye administration, as referred in the previous paragraph, with an intended therapeutic effect. Examples of “local eye treatments” include, but are not limited to, “topical eye treatment”, “intravitreal injectable treatment” and “subconjunctival injectable treatment”. For “topical eye treatment" is to be understood the direct administration of a substance or composition on the surface of the eye with an intended therapeutic effect.

[0073] For “intravitreal injection” or “intravitreal injectable treatment” is to be understood the administration of a substance or composition directly in the vitreous humor of the eye with an intended therapeutic effect. The vitreous humor is the clear, gel-like substance that fills the space between the lens and the retina.

[0074] For “subconjunctival injection” or “subconjunctival injectable treatment” is to be understood the administration of a substance or composition directly in the subconjunctival space of the eye with an intended therapeutic effect. The subconjunctival space is the area beneath the conjunctiva, the thin, transparent membrane that covers the white part of the eye (sclera) and the inner surface of the eyelids

[0075] The term "delivery agent" is to be understood as a pharmaceutically acceptable vehicle. The delivery agent can be organic, inorganic, or both. Suitable delivery agents are well known to those of skill in the art and include, without limitation, large, slowly metabolized macromolecules such as proteins, polysaccharides, polylactic acids, polyglycolic acids, polymeric amino acids, amino acid copolymers, lipid aggregates (such as oil droplets or liposomes) and inactive virus particles. Delivery agents may also include, saline, buffer, dextrose, water, glycerol, ethanol, and the combinations thereof. Non-limiting examples of the delivery agent include a polycationic polymer, a polymer, a viral particle, a vesicle, a nanovesicle, a liposome, or a nanoparticle.

[0076] As indicated above, in a first aspect the invention provides sitagliptin or a pharmaceutically or veterinary acceptable salt thereof, for use in the local eye treatment of a neovascular retinal disease. In other words, the sitagliptin or a pharmaceutically or veterinary acceptable salt thereof is for use in the treatment of a neovascular retinal disease, wherein the sitagliptin or a pharmaceutically or veterinary acceptable thereof is administered locally in the eye.

[0077] The chemical formula of sitagliptin is shown herewith.

[0078] Salts, solvates and prodrugs

[0079] In particular embodiments, the pharmaceutically acceptable salt is formed from pharmaceutically acceptable non-toxic acids including inorganic or organic acids selected from the group consisting of acetic, trifluoroacetic, benzenesulfonic, benzoic, camphorsulfonic, citric, ethansulfonic, oxalic, fumaric, gluconic, glutamic, hydrobromic, hydrochloric, orthophosphoric, lactic, maleic, malic, mandelic, methanesulfonic, phosphoric, succinic, sulfuric, tartaric, p-toluenesulfonic, glutamic, aspartic acid.

[0080] The preparation of pharmaceutically acceptable salts of Sitagliptin can be carried out by methods known in the art. For instance, they can be prepared by reacting Sitagliptin with a stoichiometric amount of the appropriate pharmaceutically acceptable acid in water, in an organic solvent or in a mixture of them. Sitagliptin and their salts may differ in some physical properties, but they are equivalent for the purposes of the present invention.

[0081] Sitagliptin and its salts may be in crystalline form either as free solvation compound or as solvate (e.g., hydrate). All these forms are within the scope of the present invention. Methods of solvation are generally known within the art. In general, the solvated forms with pharmaceutically acceptable solvents such as water, ethanol and the like are equivalent to the unsolvated form for the purposes of the invention.

[0082] In a particular embodiment, the sitagliptin pharmaceutically acceptable salt is sitagliptin phosphate. In particular sitagliptin phosphate monohydrate.

[0083] Sitagliptin and its prodrugs are also within the scope of the present invention. Thus, the first aspect of the invention encompasses a sitagliptin prodrug for the local eye treatment of neovascular retinal diseases. Combination with further treatments

[0084] Also, in a particular embodiment, the first aspect of the invention refers to the sitagliptin or the pharmaceutically or veterinary acceptable salt in combination with a further active ingredient.

[0085] In particular embodiments, the further active ingredient is selected from the group consisting of anti-VEGF agents, corticosteroids, and mixtures thereof. Non-limiting examples of anti-VEGF agents include ranibizumab, bevacizumab, aflibercept, brolucizumab and conbercept. Non-limiting examples of corticosteroids include triamcinolone, dexamethasone, fluocinolone, prednisolone, and acceptable salts thereof.

[0086] In particular embodiments, the further active ingredient is administered simultaneously with sitagliptin or a pharmaceutically or veterinary acceptable salt thereof. In another particular embodiment, the further active ingredient is administered separately, in any order, within a therapeutically effective interval.

[0087] Also, in a particular embodiment, the first aspect of the invention refers to the sitagliptin or the pharmaceutically or veterinary acceptable salt in combination with a further treatment.

[0088] In particular embodiments, the further treatment is selected from the group of intravitreal injection of a further active ingredient or laser photocoagulation. In particular embodiments, the further active ingredient is selected from the group consisting of anti- VEGF agents, corticosteroids, and mixtures thereof.

[0089] In particular embodiments, the further treatment is performed simultaneously with sitagliptin or a pharmaceutically or veterinary acceptable salt thereof. In another particular embodiment, the further treatment is performed separately, in any order, within a therapeutically effective interval.

[0090] Delivery

[0091] The sitagliptin or salt thereof described herein may be provided on its own or together with a delivery agent, or together with other molecules which contribute to the desired therapeutic effect. The inclusion of the sitagliptin or salt thereof disclosed herein within a delivery agent may be convenient, for example, in order to target the desired cells or tissues, and to enter into them an interact with its target, to increase the treatment efficacy and / or to improve degradation resistance. Thus, in a particular embodiment of the first aspect, the sitagliptin or a pharmaceutically or veterinary salt thereof is included within a delivery agent. In a more particular embodiment, the delivery agent is selected from the group consisting of polycationic polymer, a polymer, a viral particle, a vesicle, a nanovesicle, a liposome and a nanoparticle.

[0092] Treatment of neovascular retinal diseases

[0093] For the purpose of the invention the treatment of a neovascular retinal disease encompasses the treatment of the neovascular retinal disease itself, and complications thereof that course with or are caused by neovascularization in the eye.

[0094] It is noted that treatment does not include prevention. In this regard it is noted that other conditions such as the dry age-related macular degeneration, which involves retinal thinning and drusen formation without neovascularization; the ESDR, which is marked by retinal microaneurysms and hemorrhages but lacks the abnormal vessel growth seen in the advanced stages of DR; the glaucoma; and the retinitis pigmentosa are not considered as neovascular retinal diseases even though they could eventually evolve or progress towards it. Therefore, treatment of such other conditions is not herein included.

[0095] In a more particular embodiment, the neovascular retinal disease is selected from the group consisting of ASDR, wARMD, retinopathy of prematurity, retinal vein occlusion, rubeosis iridis and corneal neovascularization. More in particular, it is selected from the group consisting of ASDR and wARMD.

[0096] In ASDR, chronic high blood sugar levels damage existing vessels in the retina. This oxygen deprivation triggers the release of signals promoting abnormal, fragile blood vessel growth. This neovascularization is the defining characteristic of PDR, leading to complications such as bleeding within the eye and potentially retinal detachment. Additionally, the leaky nature of these new vessels contributes to DME, a swelling of the central retina that threatens vision or visual acuity.

[0097] More in particular it is selected from the group consisting of PDR, DME and wARMD. More in particular it is selected from the group consisting of PDR and wARMD. More in particular it is selected from the group consisting of DME and wARMD.

[0098] In a more particular embodiment, the neovascular disease is ASDR. The treatment of the ASDR encompasses the treatment of the ASDR itself, and complications thereof that course with or are caused by neovascularization in the eye. In a particular embodiment of the first aspect, the neovascular retinal disease is selected from the group consisting of PDR and DME.

[0099] In a particular embodiment, the neovascular retinal disease is PDR.

[0100] In another particular embodiment, the neovascular retinal disease is DME.

[0101] In another particular embodiment, the neovascular retinal disease is PDR and DME.

[0102] In another particular embodiment, the neovascular retinal disease is wARMD.

[0103] Topical administration

[0104] The first aspect of the invention also provides sitagliptin or a pharmaceutically or veterinary acceptable salt thereof for use in the topical eye treatment of a neovascular retinal disease. In other words, the sitagliptin or a pharmaceutically or veterinary acceptable salt thereof is for use in the treatment of a neovascular retinal disease, wherein the sitagliptin or a pharmaceutically or veterinary acceptable thereof is administered topically in the eye.

[0105] Injectable administration

[0106] In another embodiment, the sitagliptin or a pharmaceutically or veterinary acceptable salt thereof as defined in the first aspect is for use in the injectable eye treatment of a neovascular retinal disease. In other words, the sitagliptin or a pharmaceutically or veterinary acceptable salt thereof is for use in the treatment of a neovascular retinal disease, wherein the sitagliptin or a pharmaceutically or veterinary acceptable thereof is injected in the eye.

[0107] In a more particular embodiment, it is for use in the intravitreal injectable treatment of a neovascular retinal disease. In other words, the sitagliptin or a pharmaceutically or veterinary acceptable salt thereof is for use in the treatment of a neovascular retinal disease, wherein the sitagliptin or a pharmaceutically or veterinary acceptable thereof is intravitreally injected in the eye.

[0108] As above exposed, a second aspect of the invention is a pharmaceutical or veterinary composition for use in the local eye treatment of a neovascular retinal disease which comprises a therapeutically effective amount of sitagliptin or a pharmaceutically or veterinary acceptable salt thereof as defined in the first aspect, together with one or more topical pharmaceutically or veterinary acceptable excipients. In other words, the composition is for use in the treatment of a neovascular retinal disease, wherein the composition is administered locally in the eye.

[0109] The compositions of the invention can be applied on the surface of the eye and allow sitagliptin or a pharmaceutically or veterinary acceptable salt thereof release to reach the retina. In fact, the retina is reached by the transscleral route (i.e. , sitagliptin reaches the retina by crossing the sclera).

[0110] Thus, in a particular embodiment, the pharmaceutical or veterinary composition is for use in the topical eye treatment of a neovascular retinal disease. In other words, the composition is for topical administration in the eye.

[0111] Thus, in a particular embodiment of the invention, the pharmaceutical and veterinary composition of the second aspect is a topical composition. More in particular, it is a topical ocular composition; or, in other words, the composition is a topical ophtalmic composition.

[0112] Formulation and administration methods of

[0113] Various formulations exist for the topical compositions of the invention, providing flexibility in treatment. Non-limiting examples include include solutions, suspensions, emulsions, semi-solid preparations. Non limiting examples of semi-solid preparartion include lotions, creams, unguents, gels and ointments.

[0114] - Solutions: clear liquids where the active ingredient is dissolved in a suitable solvent.

[0115] - Creams, lotions, and unguents: semi-solid preparations with varying consistencies, used for longer contact time on the eye surface.

[0116] Emulsions: mixtures of oil and water-based components, often used for delivering poorly soluble drugs.

[0117] - Gels: semi-solid viscous preparations that provide sustained release of medication.

[0118] - Ointments: semi-solid greasy preparations offering extended drug contact time.

[0119] - Suspensions: contain fine particles of undissolved drug.

[0120] The choice of formulation depends on the specific drug, the desired therapeutic effect, and patient factors and may be easily determined by a person skilled in the art.

[0121] For example, solutions and lotions can be prepared at the time of administration from powders, supplied in a dry sterile form to be dissolved or suspended in an appropriate liquid vehicle.

[0122] Thus, in a particular embodiment of the second aspect, the pharmaceutical or veterinary topical composition is selected from the group consisting of a solution, a cream, a lotion, an unguent, an emulsion, a gel, an ointment and a suspension. The pharmaceutical or veterinary topical compositions of the invention, in particular embodiments, are topical compositions applicable to the cornea, sclera, conjunctival fornix, conjunctiva, eyelids.

[0123] Topical compositions of the invention can be administered through several methods, each offering advantages for specific situations and suitable depending on the formulation, which the skilled person knows. Non-limiting examples of administration methods include eye drops, direct application in the eyelid, aerosol and non-aerosol sprays and direct irrigation.

[0124] Eye drops: the most common method for solutions. Drops can, for example, be instilled directly in the lower conjunctival sac (the pocket formed by the lower eyelid) or in the cornea.

[0125] - Creams, lotions, ointments: applied directly in the eyelid margin or lower conjunctival sac, often with a gentle massage to spread the medication.

[0126] - Aerosol and non-aerosol sprays: provide a fine mist for application to the closed eye, offering convenience and potential for less wastage.

[0127] Direct irrigation: used for flushing the eye with large volumes of solution, often to remove irritants or contaminants.

[0128] The choice of administration method depends on the formulation, the target site of action within the eye, and patient-specific factors such as comfort and ease of use. And the skilled person knows it and would accordingly choose the appropriate administration method.

[0129] In a particular embodiment, the topical composition of the invention is a solution. In a more particular embodiment, the topical composition is a solution in the form of eye drops. In other words, an eye drop solution. Thus, in a particular embodiment, the topical composition is an eye drop solution. The administration in the form of eye drops implies the great advantage of being easy to be used by the subject in need thereof, and nonuncomfortable.

[0130] These pharmaceutical or veterinary topical compositions also relate to solid or semi-solid matrices or supports, in particular bioerodible and / or biodegradable polymer matrices for the delivery of sitagliptin or a pharmaceutically or veterinary acceptable salt thereof, that are comprised in the matrices.

[0131] Excipients for topical compositions

[0132] Additionally, the compositions of the present invention may contain other ingredients, such as colorants, and other components known in the state of the art for use in topical ocular compositions.

[0133] Topical compositions of the present invention can be prepared according to methods well known in the state of the art. The appropriate excipients, and their amounts, can readily be determined by those skilled in the art according to the type of formulation being prepared.

[0134] Non-limiting examples of excipients to include in the topical composition of the invention include humectants, colorants, pH buffers, surfactants, solvents (organic and inorganic solvents; i.e. water), viscosity agents, preservatives, agglutinants, emollients, and antioxidants, isotonifying and / or isoosmozing agents, mucoadhesive polymers, and / or agents enhancing absorption of the active principle (i.e.: sitagliptin or a pharmaceutically or veterinary acceptable salt thereof).

[0135] Non-limiting examples of surfactants include glycerides, polysorbates, sodium lauryl sulphate, phospholipids, (such as phosphatidyl choline or phosphatidyl glycerol), polyoxyethylene fatty acids, mono-di and triglycerides, optionally polyoxyethylene substituted, and mixtures thereof.

[0136] Non-limiting examples of particular humectants (also named humectant solvents) include polyethylene glycol (PEG of general formula H(OCH2CH2)nOH; wherein n is the mean of oxyethylene groups in the polymer), propylene glycol, glycerin, and mixtures thereof. In the context of the invention, the humectant is a compound having solvent and humectant properties.

[0137] PEG of different molecular weights are widely used in pharmaceutical compositions (being topical, parenteral, ophthalmicals, orals and rectal compositions). Appropriate PEGs to be used in the topical compositions for use according to the invention has a molecular weight from 300 to 35000 g / mol, more in particular from 600 to 20000 g / mol, even more in particular from 1000 to 8000 g / mol, more particularly from 3000 to 6000 g / mol, and preferably about 4000 g / mol. In a particular embodiment of the compositions for use according to the invention, the humectant is comprised in an amount from 1% to 49% in weight / volume in relation to the total volume of the composition. More in particular from 5% to 40%, even from 10% to 30%, and even more in particular from 15% to 25%.

[0138] Excipients used as pH buffers are those allowing a pH from 4.5 to 9.0, more in particular from 4.5 to 8.5, even more in particular from 4.5 to 8.0, even more in particular from 4.5 to 7.5, even more in particular from 5.0 to 7.5, even more in particular from 5.0 to 7.0.

[0139] Examples of pH buffers include citrate salts (citric acid / citrate buffer), phosphate salts (phosporic acid / phosphate buffer), borate salts (boric acid / borate buffer), and mixtures thereof, all salts being those pharmaceuticaly accepatble. pH buffers may in additional comprise amino acids, in particular arginine, lysine, and an amine-derived compound selected from methylglucamine and trometamol, and mixtures thereof.

[0140] Excipients, more appropriate for lipophilic compositions (which means compositions non miscible with water at 15-35 °C) include synthetic or semisynthetic lipophilic excipients comprising cocoa butter, vegetal hydrogenated oils and solid semisynthetic glycerides.

[0141] In a particular embodiment, the organic solvents used in the composition are selected from the group consisting of ricin oil, PEG, poloxamers, polysorbates, glycerin, triglycerides with C6-C10 carbon atoms fatty acids, and mixtures thereof.

[0142] The viscosity agents are in particular polyvynil alcohol, compounds derived from cellulose such as methylecellulose and hydroxypropylmethylcellulose, carbomers, PEG and mixtures thereof. The preservatives are in particular boric acid, benzalconium chloride, benzoic acid, p-hydroxybenzoic esters of C1-C4-alkyl chains, chlorobuthanol, benzyl alcohol, and mixtures thereof.

[0143] Isotonifying and / or isoosmozing agents, are in particular sodium chloride, dextrose, trehalose, mannitol, amino acids and mixtures thereof. Agents enhancing absorption of the active principle include saponin, fatty acids, pyrrolidone, polyvinylpyrrolidone, pyruvic acid and mixtures thereof. The mucoadhesive polymers (used commonly as gelling agents) are in particular hyaluronic acid, polygalacturonic acid, polyacrylic acid, chondroitin sulphate, methylcellulose, hydroxypropylmethylcellulose, gelatine, methylcellulose, xanthan gum, sodium carboxymethylcellulose, chitosan, carbopol, gellan gum, pectin, alginates, carrageenans, and mixtures thereof.

[0144] Emulsion and microemulsion bases are in particular fatty acid esters of glycerin, polyoxyethylene alcohols, ricin oil, triglycerides with C6-C10 carbon atoms fatty acids, and mixtures thereof.

[0145] In a particular embodiment, cream and ointment bases are selected from the group consisting of Vaseline, paraffin, PEG, silicones and mixtures thereof.

[0146] In another particular embodiment of the second aspect, the topical composition further comprises cyclodextrins, hyaluronic acid or mixtures thereof.

[0147] In another particular embodiment, the pharmaceutical or veterinary topical composition has a dynamic viscosity from 5.0 x 10'4Pa.s to 300 Pa.s, and a pH from 4.5 to 9.0.

[0148] Indeed, the pharmaceutical or veterinary topical compositions of the invention are liquid compositions or semi-solid compositions having a consistency of that of a cream or an unguent.

[0149] When in this description it is indicated that a composition has a particular viscosity within a range, it is related to the dynamic viscosity. Thus, the pharmaceutical or veterinary topical compositions of the invention have a dynamic viscosity from 5.0 x 1 O'4Pa.s to 300 Pa.s, at room temperature and normal atmospheric pressure. In a particular embodiment of this second aspect, the dynamic viscosity of the pharmaceutical or veterinary topical composition is from 8.9 x 10'4Pa.s to 100 Pa.s.

[0150] In particular embodiments, the dynamic viscosity is measured at a shear rate of 1 s-1at 20°C. Alternatively, it can be measured according to standard methods known by the skilled person.

[0151] In particular embodiments, the pharmaceutical or veterinary composition, wherein the composition has a dynamic viscosity from 5.0 x 10'4Pa.s to 300 Pa.s measured at a shear rate of 1 s-1at 20 °C, and a pH from 4.5 to 9.0.

[0152] The term “dynamic viscosity” or “viscosity coefficient q” referes to the tangencial force per unit surface, known as shearing stress T and expressed in pascals, necessary to move, parallel to the sliding plane, a layer of liquid of 1 square meter at a rate (v) of 1 meter per second relative to a parallel layer at a distance (x) of 1 meter. The ratio dv / dx is a speed gradient giving the rate of shear D expressed in reciprocal seconds (s-1), so that q=T / D. The unit of dynamic viscosity is the pascal second (Pa.s). In another particular embodiment the pH of the pharmaceutically or veterinary topical compositions of the invention, is from 5.5 to 7.5, more particularly about 7.0.

[0153] In yet another particular embodiment of the pharmaceutically or veterinary topical compositions of the invention, the sitagliptin or a pharmaceutically or veterinary acceptable salt thereof is in a concentration from 1 mg / ml to 200 mg / ml, from 1 mg / ml to 150 mg / ml, from 1 mg / ml to 120 mg / ml, from 1 mg / ml to 100 mg / ml or from 1 mg / ml to 80 mg / ml in relation to the total volume of the composition. In another particular embodiment it is in a concentration from 5 mg / ml to 200 mg / ml, from 5 mg / ml to 150 mg / ml, from 5 mg / ml to 120 mg / ml, from 5 mg / ml to 100 mg / ml, from 5 mg / ml to 80 mg / ml in relation to the total volume of the composition. In another particular embodiment it is in a concentration from 50 mg / ml to 200 mg / ml, from 50 mg / ml to 150 mg / ml, from 50 mg / ml to 120 mg / ml, from 50 mg / ml to 100 mg / ml or from 50 mg / ml to 80 mg / ml in relation to the total volume of the composition. Particularly, it is about 40 mg / ml, 41 mg / ml, about 42 mg / ml, about 43 mg / ml, about 44 mg / ml, about 45 mg / ml, about 46 mg / ml, about 47 mg / ml, about 48 mg / ml, about 49 mg / ml, about 50 mg / ml, about 51 mg / ml, about 52 mg / ml, about 53 mg / ml, about 54 mg / ml, about 55 mg / ml, about 56 mg / ml, about 57 mg / ml, about 58 mg / ml, about 59 mg / ml, or about 60 mg / ml in relation to the total volume of the composition.

[0154] More in particular, the pharmaceutical or veterinary topical composition has a dynamic viscosity from 5.0 x 10'4Pa.s to 300 Pa.s at 20 °C, a pH from 4.5 to 9.0, and wherein the sitagliptin or a pharmaceutically or veterinary acceptable salt thereof is in a concentration from 1 mg / ml to 200 mg / ml in relation to the total volume of the composition.

[0155] More in particular, the pharmaceutical or veterinary topical composition has a dynamic viscosity from 5.0 x 10'4Pa.s to 300 Pa.s measured at a shear rate of 1 s-1at 20 °C, a pH from 4.5 to 9.0, and wherein the sitagliptin or a pharmaceutically or veterinary acceptable salt thereof is in a concentration from 1 mg / ml to 200 mg / ml in relation to the total volume of the composition.

[0156] More in particular, the pharmaceutical or veterinary topical composition has a dynamic viscosity from 5.0 x 10'4Pa.s to 300 Pa.s at 20 °C, a pH from 4.5 to 9.0, and wherein the sitagliptin or a pharmaceutically or veterinary acceptable salt thereof is in a concentration from 5 mg / ml to 100 mg / ml in relation to the total volume of the composition.

[0157] More in particular, the pharmaceutical or veterinary topical composition has a dynamic viscosity from 5.0 x 10'4Pa.s to 300 Pa.s measured at a shear rate of 1 s-1at 20 °C, a pH from 4.5 to 9.0, and wherein the sitagliptin or a pharmaceutically or veterinary acceptable salt thereof is in a concentration from 5 mg / ml to 100 mg / ml in relation to the total volume of the composition.

[0158] Injectable compositions

[0159] Although topical administration is preferred, other forms are possible, such as injectable administration. Therefore, the composition containing the effective amount of the sitagliptin or pharmaceutically or vetereinary acceptable salt thereof can be administered as an injectable solution or suspension, preferably an intravitreal injectable liquid. In a particular embodiment of the second aspect, the composition is for use in the injectable eye treatment of a neovascular retinal disease. More in particular, the composition is for use in the injectable intravitreal treatment. In other words, the composition is for injection. More in particular it is for ocular injection. More in particular it is for subconjuctival injection or intravitreal injection. More in particular it is for intravitreal injection.

[0160] In particular embodiments, the composition of the second aspect is an injectable composition. More in particular it is an ocular injectable composition. More in particular, it is an subconjuctival injectable or intravitreal injectable composition. More in particular, it is an intravitreal injectable composition.

[0161] Excipients for injectable compositions

[0162] Selection of the excipients and the most appropriate methods for formulation in view of the particular purpose of the composition (topical or injectable) is within the scope of ordinary persons skilled in the art of pharmaceutical technology.

[0163] In this regard, pharmaceutical composition delivered via intravitreal injection may typically be formulated in a sterile saline solution, balanced salt solution (BSS), or other appropriate mediums. The formulation may also include additional components such as preservatives, stabilizers, biodegradable polymers for sustained release, nanoparticles for targeted delivery, or pH-responsive materials to optimize therapeutic efficacy and patient outcomes.

[0164] In a particular embodiment, the injectable compositions further comprise cyclodextrins and hyaluronic acid.

[0165] Physical parameters of injectable compositions The physical parameters of the injectable compositions of the invention are carefully controlled to ensure biocompatibility and effectiveness. These parameters include the pH range (generally 6.8 to 7.4), osmolarity (typically isotonic at approximately 300 ± 10 mOsm / kg), and viscosity (similar to that of the natural vitreous humor). The injection volume may range from 0.01 to 0.5 ml, more in particular from 0.05 to 0.1 ml.

[0166] The formulation of injectable compositions of the invention, are designed to provide targeted, localized treatment with minimal systemic exposure and adverse effects.

[0167] Sustained release

[0168] The compositions for use according to the invention, both for topical and injectable administration, are, in particular embodiments, sustained-release compositions. That is, the compositions are formulated as sustained-release delivery systems allowing the delivery of the active principle (i.e.: sitagliptin or a pharmaceutically or veterinary acceptable salt thereof) at a predetermined rate in order to maintain a drug concentration for a specific period of time with minimum side effects.

[0169] Particular formulations for the sustained-release delivery comprise nanoparticles and microparticles encapsulating the sitagliptin or a pharmaceutically or veterinary acceptable salt thereof, liposomes and niosomes, all of them comprising a compound selected from polylactic acid, poly(lactic-co-glycolic) acid, polystyrenes, chitosan, albumin, lectins, gelatins, acrylates and methacrylates, polycaprolactones, polyacrylamides, dextranes, agarose, sorbitan, cholesterol, and mixtures thereof. Other particular formulations for the sustained-release delivery comprise a polymer conjugated with the sitagliptin or a pharmaceutically or veterinary acceptable salt thereof constituting hydrogels.

[0170] Kit

[0171] In a particular embodiment of the second aspect, the composition, either topical and injectable, is provided as part of a kit. In a particular, embodiment, the kit further comprises a composition comprising a further active ingredient; and instructions for administering the compositions. In a particular embodiment, the instructions are for administering the compositions in the form of eye drops.

[0172] In another particular embodiment, the composition is provided as part of a kit, which comprises a container A comprising the composition and a container B comprising a further active ingredient, or a composition comprising a further active ingredient; and optionally instructions for administering the compositions. In another particular embodiment, both the container A and B contain sterile solutions.

[0173] In another particular embodiment, the further active ingredient is selected from the group consisting of anti-VEGF agents, corticosteroids and mixtures thereof. More in particular, the anti-VEGF agent is selected from the group consisting of ranibizumab, bevacizumab, aflibercept, brolucizumab and conbercept. More in particular, the corticosteroid is selected from the group consisting of triamcinolone, dexamethasone, fluocinolone, prednisolone, and acceptable salts thereof.

[0174] Treatment of neovascular retinal diseases

[0175] In a particular embodiment of the second aspect of the invention, either for topical and injectable compositions, the neovascular retinal disease is selected from the group consisting of ASDR, wARMD, retinopathy of prematurity, retinal vein occlusion, rubeosis iridis and corneal neovascularization. In a more particular embodiment, the neovascular retinal disease is selected from the group consisting of ASDR and wARMD. In a more particular embodiment, the neovascular retinal disease is selected from the group consisting of PDR, DME and wARMD.

[0176] In a more particular embodiment, the neovascular retinal disease is ASDR. In a more particular embodiment, the ASDR is selected from the group consisting of PDR and DME.

[0177] In particular embodiments, the ASDR is PDR.

[0178] In other particular embodiments, the ASDR is DME.

[0179] In other particular embodiments, the ASDR is PDR and DME.

[0180] In a particular embodiment, the neovascular retinal disease is wARMD.

[0181] Combination with further treatments

[0182] Also, in particular embodiments, the second aspect of the invention refers to the either topical and injectable compositions further comprising a further active ingredient.

[0183] In particular embodiments, the further active ingredient is selected from the group consisting of anti-VEGF agents, corticosteroids, and mixtures thereof. Non-limiting examples of anti-VEGF agents include ranibizumab, bevacizumab, aflibercept, brolucizumab and conbercept. Non-limiting examples of corticosteroids include triamcinolone, dexamethasone, fluocinolone, prednisolone, and acceptable salts thereof.

[0184] Also, in a particular embodiment, the second aspect of the invention refers to the composition in combination with a further treatment.

[0185] In particular embodiments, the further treatment is selected from the group of intravitreal injection of a further active ingredient or laser photocoagulation. In particular embodiments, the further active ingredient is selected from the group consisting of anti- VEGF agents, corticosteroids, and mixtures thereof.

[0186] In particular embodiments, the further treatment is performed simultaneously with the administration of composition. In another particular embodiment, the further treatment is performed separately, in any order, within a therapeutically effective interval.

[0187] The first aspect can also be formulated as the use of the sitagliptin or a pharmaceutically or veterinary acceptable salt thereof as defined in the first aspect for the manufacture of a medicament for treating a neovascular retinal disease. Alternatively, this can be worded as the use of the sitagliptin or a pharmaceutically or veterinary acceptable salt thereof as defined in the first aspect for the manufacture of pharmaceutical or veterinary composition as defined in the second aspect for the treatment of a neovascular retinal disease.

[0188] The first aspect can also be formulated as a method for the treatment of a neovascular retinal disease, the method comprising administering a therapeutically effective amount of the sitagliptin or a pharmaceutically or veterinary acceptable salt thereof as defined in the first aspect, together with pharmaceutical or veterinary acceptable excipients, in a subject in need thereof.

[0189] Throughout the description and claims the word "comprise" and variations of the word, are not intended to exclude other technical features, additives, components, or steps. Furthermore, the word “comprise” encompasses the case of “consisting of”. Additional objects, advantages and features of the invention will become apparent to those skilled in the art upon examination of the description or may be learned by practice of the invention. The following examples and drawings are provided by way of illustration, and they are not intended to be limiting of the present invention. Furthermore, the present invention covers all possible combinations of particular and preferred embodiments described herein.

[0190] Examples

[0191] Example 1: In vitro Experiments The effect of sitagliptin in key steps of the angiogenic process, including cell migration and cell tube formation using Human Retinal Endothelial Cells (HRECs), was assessed.

[0192] Methods

[0193] Human Retinal Endothelial Cell Culture

[0194] To evaluate the effect of sitagliptin on angiogenesis in vitro, the commercially-purchased cell line Immortalized Human Retinal Endothelial Cells (HRECs) was used to perform scratch wound healing and tube formation techniques. HRECs were purchased from Innoprot (Spain). The cells were grown at 37°C in a humidified atmosphere of 95% air and 5% CO2, with Endothelial Cell Medium kit (EC medium) containing 5% fetal bovine serum (FBS), 1% endothelial cell growth supplement and 1% penicillin / streptomycin solution (Innoprot, Spain), on flasks or plates coated with attachment factor (15 pg / ml fibronectin protein, Millipore Sigma, USA). The HRECs were split at -90% confluence using passage reagent Tryp-LE Express (ThermoFisher Set., USA), and culture media was replaced every 2-3 days. Cells between passages 3-4 were used in this study.

[0195] Scratch Wound Healing

[0196] HRECs were seeded in a fibronectin-coated 48-well plate at 5 x 104cells / well and were allowed to make a confluent monolayer in a complete medium with normal glucose (NG) (5 mM) or high glucose (HG) (30 mM) by adding D-(+)-Glucose (Millipore Sigma, USA), during 48 hours. The medium was then replaced by an EC medium containing 1% FBS (Innoprot, Spain) in which the cells were maintained for 24 hours in NG or HG conditions. Wounds were made by cutting the cell monolayers with a pipette tip. After washing twice with sterile Phosphate-buffered saline (PBS), media containing sitagliptin (Sitagliptin Phosphate Monohydrate; Selleckchem 654671-77-9), or vehicle, in the presence of 25 ng / mL VEGF (rhVEGF165, Biotechne R&D Systems, USA), were added to the wells. Each condition was done in quadruplicate. The HRECs were cultured at 37°C and 5% CO2 and visualized under an inverted phase contrast microscope (CKX41 Olympus Corporation, Japan) at different time points to monitor the healing of the wounds. Photographs were taken with a 4X objective and quantified with Imaged 1 ,53p software (National Institute of Health, USA).

[0197] Endothelial Cell Tube Formation Assay

[0198] To study tube formation in response to sitagliptin, we pretreated HRECs in separate 25 cm2flasks for three days, either with sitagliptin (200 mM) or vehicle, in NG medium and in the presence or absence of 25 ng / mL VEGF (rhVEGF165, Bio-techne R&D Systems, USA). Tube formation studies were conducted using an in vitro angiogenesis assay in a 96-well format. Briefly, Matrigel (Corning, USA) was thawed in ice overnight, and then 60 pl of Matrigel were added into each well of a 96-well plate and incubated for 45 min at 37°C to gel. HRECs were then harvested from the 25 cm2flasks, counted, and single-cell suspension at 1 x 105cells / mL was prepared in separate tubes using a medium with corresponding treatments. The cells were seeded in the Matrigel pretreated plate (1 x 104cells per well, 100 pl), and incubated at 37°C and 5% CO2. The tube formation was visualized directly through an inverted phase contrast microscope (CKX41 , Olympus Corporation, Japan). Photomicrographs were acquired at 2h, 4h, 6h, and 8h with a 4X objective, and analyzed and quantified using the Angiogenesis Analyzer for Imaged 1.53p software (National Institute of Health, USA). The area of each image, or the user selection concerned by the analysis, was first defined. The parameters studied were «Nb junctions»: number of junctions in the analyzed area; «Nb segments)): number of segments in the analyzed area; «Nb meshes»: number of meshes in the analyzed area; and «Tot. Branching length»: sum of the length of the trees composed from segments and branches in the analyzed area.

[0199] Results

[0200] The cell migration rate was significantly decreased when HRECs were treated with sitagliptin (Figures 1A, 1 B, 1C and 1 D). In vitro formation of capillary-like tubes by endothelial cells on a basement membrane matrix is a powerful method to screen for drugs that inhibit and regress angiogenesis. It was demonstrated that VEGF-induction promoted the formation of tubular structures of HRECs in Matrigel, whereas this effect was abolished when cells were treated with sitagliptin (Figures 2A, 2B, 2C and 2D). In conclusion, sitagliptin treatment abolish the formation of capillary-like tubes by endothelial cells, which is indicative of anti-angiogenic effect, and so of a therapeutic effect against neovascular retinal diseases. In particular against ASDR and wARMD, and more in particular against PDR and wARMD. More in particular against PDR.

[0201] Example 2: Ex vivo experiments

[0202] Study of the antiangiogenic effect of sitagliptin in retinal explants obtained from rats. Two ex vivo experimental approaches were performed to evaluate diabetes-induced angiogenesis:

[0203] 1) Explants obtained from non-diabetic rats (Long Evans rats) and cultured under high or normal glucose conditions.

[0204] 2) Explants obtained from rats with streptozotocin-induced diabetes (D-STZ) and from non-diabetic rats were used as controls.

[0205] After euthanasia, the eyes were immediately enucleated and maintained in sterile PBS pH 7.4 before dissection. After removing the cornea and lens from the anterior part of the eye, the posterior segment formed by the sclera-choroid-RPE complex was separated from the retina and cut into approximately 1 mm x 1 mm pieces. The posterior segment fragments were isolated with forceps and placed in 40 pl Matrigel (Corning, USA) drops in 24-well glass bottom plates (Cellvis, USA). The plates were incubated without medium in a cell culture incubator at 37°C for 30 minutes. Then, 500 pl of EBM-2 Basal Medium with EGM- 2 SingleQuots Supplements kit (Lonza, Switzerland) were added to each well and incubated at 37°C with 5% CO2 for 24 h before the respective treatments. For the HG condition, glucose was added to a final concentration of 30 mM, and 5 mM corresponded to the NG condition. VEGF from the supplement kit was replaced with 5 ng / ml rrVEGF164 (Biotechne R&D Systems, USA), or no VEGF was added. The explants obtained from non-diabetic rats were then treated for 5 days with either sitagliptin (Sitagliptin Phosphate Monohydrate; Selleckchem 654671-77-9) or vehicle. Sprouting was monitored daily with an inverted phase contrast microscope (CKX41 , Olympus Corporation, Japan). On the fifth day, the wells were washed with PBS, and a fluorescent probe, CellTracker Green CMFDA Dye (ThermoFisher Set., USA), was added. After 45 minutes of incubation at 37°C, photographs were taken with a Leica DM IRBE inverted fluorescence microscope at 2.5X (BP450-490 excitation filter and LP515 emission filter) for fluorescence imaging. A PZMTI I l-M I stereomicroscope with a PRO-300HDS high-definition camera (World Precision Instruments) was also used at 2X to take brightfield images. RPE-choroidal sprouting areas were quantified with Imaged 1.53p software (National Institute of Health, USA).

[0206] It was evaluated whether sitagliptin was able to inhibit and regress angiogenesis in retinal explants obtained from non-diabetic rats exposed to normal or HG conditions. Explants incubated in the HG medium had more positive sprouts than explants incubated in the NG medium. The number of neurovascular sprouts in explants incubated with sitagliptin in HG was quantified, and it was observed that this treatment significantly reduced the area of neurovascular sprouts compared to the explants without sitagliptin treatment (Figures 3A, 3B and 4). Similar results were obtained in retinal explants obtained from STZ-induced diabetic rats treated with sitagliptin eye drops (twice at day) for 2 weeks prior the obtention of retinal explants (Figures 5, 6 and 7). In conclusion, sitagliptin reduces the neurovascular sprouts in explants, which is indicative of the inhibition and regression of angiogenesis (i.e. anti-angiogenic activity). This is indicative of the therapeutic potential against neovascular retinal diseases. In particular against ASDR, and more in particular against PDR.

[0207] Example 3: In vivo experiment

[0208] This study aimed to evaluate the efficacy of topical administration of sitagliptin for the treatment of wARMD in diabetic and non-diabetic mice.

[0209] Methods A laser-induced choroidal neovascularization (LI-CNV) model was used. This is an established method to evaluate therapies for wARMD. Diabetic (db / db, n=12) and nondiabetic mice (C57BL / 6J, n=12) with LI-CNV were included. The db / db mouse carries a mutation in the leptin receptor gene and is a model for obesity-induced type 2 diabetes. A group of 4 C57BL / 6J mice in whom choroidal neovascularization was not induced served as a control group.

[0210] The choroidal neovascularization was induced in mice’s retinas by applying a green laser to rupture Bruch’s membrane. Briefly, once the animal was anesthetized, pupils were dilated with Tropicamide eyedrops and the animal was placed in the Micron III platform (Phoenix Research Labs., Pleasanton, CA, USA), putting the lens in contact with the cornea. Bruch’s membrane rupture was achieved by applying an argon laser of 352 nm, a power of 250 mW during 100 ms, and a fixed diameter of 50 pm by using the image- guided laser system. Animals received 4 burns per eye at clock positions 3, 6, 9, and 12 equidistant from the optic nerve.

[0211] Diabetic and non-diabetic mice received topical ocular administration of sitagliptin [sitagliptin phosphate monohydrate (Y0001812, Merck KGaA, Darmstadt, Germany), concentrated 10 mg / mL (n= 6 animals per group), or vehicle eye drops [phosphate buffered saline (PBS), n= 6 animals per group] for a week (starting 4 hours before laser injury). Eye drops (twice per day) were randomly administered with the aid of a micropipette (5 pL) onto the superior corneal surfaces.

[0212] Animals were subjected to fundus fluorescence angiography (FA) at days 1 and 7 postlaser. Flat-mounted RPE-choroid-sclera eyecups were obtained after euthanasia at day 7 post-laser. The area of CNV was assessed by immunofluorescence using isolectin B4 for labeling endothelial cells emerging from the choroid.

[0213] Results

[0214] It was found that sitagliptin eye drops reduce CNV induced by laser in diabetic (db / db), as well as in non-diabetic mice, in the examinations performed in vivo (Figures 8A) and postmortem (9A). The neovascular area, measured in fluorescence angiography as well as in retinal flat-mounted samples, was significantly greater in LI-CNV vehicle-treated mice compared to LI-CNV sitagliptin-treated mice in both groups: diabetic and non-diabetic mice (Figure 8B and 9B).

[0215] Citation List

[0216] Patent Literature

[0217] WO2017186934A1 - Dipeptidyl peptidase-4 inhibitors for topical eye treatment of retinal neurodegenerative diseases

[0218] Non-Patent Literature

[0219] Kolibabka, M. et al. Anti-angiogenic effects of the DPP-4 inhibitor linagliptin via inhibition of VEGFR signalling in the mouse model of oxygen-induced retinopathy. Diabetologia 61, 2412-2421 (2018). https: / / doi.org / 10.1007 / sQ0125-018-4701-4

[0220] Gongalves, A., et al. (2018). The dipeptidyl peptidase-4 (DPP-4) inhibitor sitagliptin ameliorates retinal endothelial cell dysfunction triggered by inflammation. Biomedicine & pharmacotherapy, 102, 833-838. https: / / doi.Org / 10.1016 / j.biopha.2018.03.144

[0221] Lee, CS., et al. (2016). Dipeptidyl Peptidase-4 Inhibitor Increases Vascular Leakage in Retina through VE-cadherin Phosphorylation. Sci Rep 6, 29393. https: / / doi.Org / 10.1038 / srep29393

[0222] Wong, T. Y., Cheung, C. M., Larsen, M., Sharma, S., & Simo, R. (2016). Diabetic retinopathy. Nature reviews. Disease primers, 2, 16012. https: / / doi.org / 10.1038 / nrdp.2016.12

[0223] Early photocoagulation for diabetic retinopathy. ETDRS report number 9. Early Treatment Diabetic Retinopathy Study Research Group. (1991). Ophthalmology, 98(5 Suppl), 766- 785. https: / / doi.org / 10.1016 / S0161-6420(13)38011-7

[0224] Wilkinson, C. P., Ferris, F. L., 3rd, Klein, R. E., Lee, P. P., Agardh, C. D., Davis, M., Dills, D., Kampik, A., Pararajasegaram, R., Verdaguer, J. T., & Global Diabetic Retinopathy Project Group (2003). Proposed international clinical diabetic retinopathy and diabetic macular edema disease severity scales. Ophthalmology, 110(9), 1677-1682. https: / / doi.Org / 10.1016 / S0161 -6420(03)00475-5

Claims

Claims1 Sitagliptin or a pharmaceutically or veterinary acceptable salt thereof, for use in the local eye treatment of a neovascular retinal disease.2.- The sitagliptin or the pharmaceutically or veterinary acceptable salt thereof for use according to claim 1 , in combination with a further active ingredient.3.- A pharmaceutical or veterinary composition for use in the local eye treatment of a neovascular retinal disease, which comprises a therapeutically effective amount of sitagliptin or a pharmaceutically or veterinary acceptable salt thereof, together with one or more pharmaceutically or veterinary acceptable excipients.4.- The pharmaceutical or veterinary composition for use according to claim 3, further comprising a further active ingredient.5.- The pharmaceutical or veterinary composition for use according to any one of claims 3-4, which is a topical composition.6.- The pharmaceutical or veterinary composition for use according to claim 5, which is selected from the group consisting of a solution, a cream, a lotion, an unguent, an emulsion, an ointment, a gel, and a suspension.7.- The pharmaceutical or veterinary composition for use according to claim 6, which is an eye drop solution.8.- The pharmaceutical or veterinary composition for use according to any one of claims 3-7, wherein sitagliptin or the pharmaceutically or veterinary acceptable salt thereof is in a concentration from 1 mg / ml to 200 mg / ml in relation to the total volume of the composition.9.- The pharmaceutical or veterinary composition for use according to any one of claims 1-8, wherein sitagliptin or the pharmaceutically or veterinary acceptable salt thereof is in a concentration of about 5-50 mg / ml in relation to the total volume of the composition.10.- The pharmaceutical or veterinary composition for use according to any one of claims 3-4, which is an injectable composition.11.- The pharmaceutical or veterinary composition for use according to claim 10, which is an intravitreal injectable composition.12.- The pharmaceutical or veterinary composition for use according to any one of claims 3-11, which is a sustained-release composition.13.- The sitagliptin or the pharmaceutically or veterinary acceptable salt thereof for use according to any one of claims 1-2, or the pharmaceutical or veterinary topical composition for use according to any one of claims 3-12, wherein the neovascular retinal disease is selected from the group consisting of advanced stage of diabetic retinopathy, wet age-related macular degeneration, retinopathy of prematurity, retinal vein occlusion, rubeosis iridis, and corneal neovascularization.14.- The sitagliptin or the pharmaceutically or veterinary acceptable salt thereof for use according to any one of claims 1-2, or the pharmaceutical or veterinary topical composition for use according to any one of claims 3-12, wherein the neovascular retinal disease is selected from the group consisting of an advanced stage of diabetic retinopathy and wet age-related macular degeneration.15.- The sitagliptin or the pharmaceutically or veterinary acceptable salt thereof for use according to any one of claims 1-2, or the pharmaceutical or veterinary topical composition for use according to any one of claims 3-12, wherein the neovascular retinal disease is proliferative diabetic retinopathy.

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