Application of itaconic acid compound in preparation of medicine for treating retinal artery occlusion

Abstract

The invention provides application of an itaconic acid compound in preparation of a medicine for treating retinal artery occlusion, and belongs to the technical field of treatment of fundus diseases. In particular to new application of itaconic acid and derivatives thereof in preparation of drugs for treating or preventing retinal artery occlusion, the technical cognition of itaconic acid compounds in the traditional application field is broken through, a new small molecule candidate drug is provided for prevention and treatment of retinal ischemic diseases, and the application value is definite. The damage to the structure of the retina in retinal artery occlusion can be relieved, and the visual function is effectively improved; the retina microglial cells can be promoted to be polarized to M2 type, the generation of anti-inflammatory factors IL-10 and TGF beta can be increased, and a protective effect is achieved in the inflammatory response of the retina; the effect of protecting retinal ganglion cells from apoptosis is also achieved. The compound can be used for related diseases such as retinal ischemic diseases, fundus retina inflammation injury and optic nerve injury, and has a wide application prospect for fundus diseases.

Description

Technical Field

[0001] This invention relates to the field of fundus disease treatment technology, and in particular to the use of itaconic acid compounds in the preparation of drugs for treating retinal artery occlusion. Background Technology

[0002] Retinal artery occlusion (RAO), commonly known as "eye stroke," is a clinical condition caused by the interruption of blood flow to the retinal arteries, leading to retinal ischemia and potentially sudden, catastrophic vision loss or even blindness. RAO progresses rapidly, has a poor prognosis, involves numerous risk factors, and lacks a standard treatment regimen. However, RAO has a rapid onset and a short treatment window; studies have shown that irreversible death of retinal ganglion cells (RGCs) can occur within 90 minutes of RAO onset. Even after treatment restores blood flow to the retinal arteries, the surge in glucose and oxygen supply exacerbates tissue oxidation, inducing an mitochondrial "oxidative burst." This results in an overload of reactive oxygen species (ROS) in the mitochondria, causing mitochondrial oxidative damage, further damaging nerve cells and leading to retinal ischemia-reperfusion injury (RIRI). Current clinical treatment options include traditional therapies such as lowering intraocular pressure, high-flow oxygen therapy, vasodilation, and antiplatelet aggregation, as well as more aggressive treatments such as thrombolysis, hyperbaric oxygen therapy, and arterial intervention. The goal of treatment is to improve and restore retinal circulation and prevent retinal necrosis. However, the treatment window for retinopathy of retinopathy (RAO) is short, and even after thrombolysis, patients still face the challenge of retinopathy-associated retinal injury (RIRI). It has been reported that only 20% of patients experience limited visual recovery after treatment. How to maximize the salvage of ischemic but not yet infarcted tissue after RAO surgery, while simultaneously reducing oxidative damage caused by RIRI and restoring the function of the ganglion cell layer, has become a critical challenge in RAO treatment. Furthermore, there is currently a lack of specific targeted therapies specifically addressing the retinal structural and ganglion cell damage caused by RAO. Summary of the Invention

[0003] The purpose of this invention is to address the above-mentioned deficiencies of the prior art by providing the use of itaconic acid or its derivatives or pharmaceutically acceptable salts in the preparation of drugs for the treatment or prevention of retinal artery occlusion, thereby solving the problem of a lack of clinically available drugs for the treatment of RAO.

[0004] To achieve the above objectives, the present invention adopts the following technical solution: A first aspect of the present invention is to provide the use of itaconic acid compounds or pharmaceutically acceptable salts thereof in the preparation of medicaments for the treatment or prevention of retinal artery occlusion, wherein the itaconic acid compound is itaconic acid or an itaconic acid derivative having a structure as shown in Formula I: CH2=C(COOR1)- CHR3- COOR2 (Formula I) Among them, R1 and R2 are each individually selected from: hydrogen, C1-C 20 Straight-chain or branched alkyl groups, C3-C 20 cycloalkyl, C6-C 20 aryl or C7-C 20 Aryl alkyl groups; and R1 and R2 are not both hydrogen.

[0005] Furthermore, R3 is C4-C 12 n-alkyl groups.

[0006] Furthermore, the itaconic acid derivative is 4-octylitaconic acid.

[0007] Furthermore, the treatment includes improvement of the subject's disease symptoms or improvement of their health status.

[0008] Furthermore, the treatment includes reducing or delaying the occurrence of abnormal changes in retinal tissue structure during retinal artery occlusion.

[0009] Furthermore, the treatment includes promoting the M2 polarization of retinal microglia.

[0010] Furthermore, the treatment includes modulating the retinal inflammatory response.

[0011] Furthermore, the treatment includes inhibiting damage or death of retinal ganglion cells.

[0012] Furthermore, the drug is administered via intraperitoneal injection, intravitreal administration, intravenous injection, or oral administration.

[0013] Furthermore, in animal models, the drug is administered intravitreal at a dose of 0.5–5 μg per animal, or intraperitoneally at a dose of 0.1–50 mg / kg.

[0014] Furthermore, the dosage form of the drug is one or more of the following: capsules, tablets, oral preparations, microcapsule preparations, injections, eye drops, eye washes, ophthalmic creams, and ophthalmic gels.

[0015] Furthermore, the drug is administered before retinal artery occlusion occurs.

[0016] Compared with the prior art, the beneficial effects of the present invention are: (1) This invention provides a new use of itaconic acid and its derivatives in the preparation of drugs for the treatment or prevention of retinal artery occlusion, which breaks through the technical understanding of the traditional application fields of itaconic acid compounds and provides a new class of small molecule candidate drugs for the prevention and treatment of retinal ischemic diseases, with clear application value.

[0017] (2) By selecting alkyl substituents with a C4-C12 carbon chain length on the basis of the itaconic acid molecular skeleton, and having the carboxyl group exist in the form of free acid or esterification, the itaconic acid derivative has significantly improved physicochemical properties, bioavailability and in vivo effects compared to itaconic acid, thus making it more suitable for the treatment or prevention of retinal artery occlusion.

[0018] (3) The itaconic acid derivatives provided by the present invention all have a clear general structural formula, and the pharmacological activity can be optimized by adjusting the carbon chain length of R3 and the substitution form of R1 and R2. They have good structural tunability and extensibility, which is beneficial to subsequent drug screening and dosage form development, and significantly improves practicality and industrialization potential.

[0019] (4) In a representative embodiment of the present invention, 4-octylitaconic acid exhibits particularly prominent improvement effects related to retinal artery occlusion. The constructed animal model demonstrates that 4-octylitaconic acid can alleviate retinal structural damage in retinal artery occlusion and effectively improve visual function. Simultaneously, 4-octylitaconic acid can promote the M2 polarization of retinal microglia and increase the production of anti-inflammatory factors IL-10 and TGFβ, playing a protective role in retinal inflammatory responses. It also protects retinal ganglion cells from apoptosis. 4-octylitaconic acid can serve as a representative compound to verify the feasibility of the technical solution of the present invention, while providing reliable technical support for the application of other itaconic acid derivatives within the same structural range. It can be used for retinal ischemic diseases, fundus retinal inflammation, optic nerve damage, and other related diseases, showing broad application prospects in fundus diseases. Attached Figure Description

[0020] Figure 1 The results of fundus photography and optical coherence tomography (OCT) of UPOAO model mice 3 days after vitrectomy intervention; Figure 2 The results show the retinal ERG function of UPOAO model mice 7 days after 4-OI cavity injection intervention. Wave a in the figure shows no difference, while wave b shows a significant decrease. Figure 3 A schematic diagram showing the number of RGCs stained in the retina of UPOAO model mice 7 days after 4-OI cavity injection intervention; Figure 4 The results of retinal RGC counts in UPOAO model mice 7 days after 4-OI cavity injection intervention; Figure 5 The results of M2 polarization in retinal microglia of UPOAO model mice 7 days after 4-OI cavity injection intervention; Figure 6 The results of fundus photography and optical coherence tomography (OCT) of UPOAO model mice 3 days after intraperitoneal injection of 4-OI; Figure 7 The results of retinal ERG visual function testing in UPOAO model mice 7 days after intraperitoneal injection of 4-OI showed no difference in wave a, but a significant decrease in wave b. Figure 8 A schematic diagram of the number of RGCs stained in the retina of UPOAO model mice 7 days after intraperitoneal injection of 4-OI. Figure 9 The results of retinal RGC counts in UPOAO model mice 7 days after intraperitoneal injection of 4-OI; Figure 10 The results of M2 polarization in retinal microglia of UPOAO model mice 7 days after intraperitoneal injection of 4-OI; Figure 11 The changes in the level of the anti-inflammatory factor IL-10 secreted by BV2 cells after in vitro intervention with 4-OI; Figure 12 The changes in the level of the anti-inflammatory factor TGFβ secreted by BV2 cells after in vitro intervention with 4-OI. Detailed Implementation

[0021] To make the objectives, technical solutions, and advantages of the present invention clearer, embodiments of the present invention are described in detail below. Examples of these embodiments are shown in the accompanying drawings, wherein the same or similar reference numerals denote the same or similar elements or elements having the same or similar functions throughout. The embodiments described below with reference to the accompanying drawings are exemplary and are only used to explain the present invention, and should not be construed as limiting the present invention.

[0022] Terms and definitions in this invention The term "treatment or prevention" refers to a method aimed at achieving a beneficial or desired clinical outcome. For the purposes of this invention, beneficial or desired clinical outcomes include, but are not limited to, relieving symptoms, reducing disease severity, stabilizing the condition (i.e., preventing deterioration), delaying or slowing disease progression, improving or alleviating the condition, and achieving detectable or undetectable partial or complete remission. Another meaning of "treatment" is extending a patient's survival compared to the expected survival rate without treatment. Therefore, "treatment" is an intervention aimed at altering the pathological process of a disease. Specifically, treatment can directly prevent, delay, or reduce pathological retinal tissue degeneration or damage.

[0023] The term "therapeutic or preventative effective amount" refers to an amount of an active agent sufficient to induce a specific biological state, effect, and / or response. Specifically, in this invention, it refers to an amount sufficient to achieve the desired outcome, such as an effective treatment of sepsis, when administered to a subject, including mammals, such as humans. The effective amount of the agents described herein can vary depending on factors such as the subject's disease state, age, sex, and weight. As will be understood by those skilled in the art, dosages or treatment regimens can be adjusted to provide an optimal therapeutic response. For example, in this invention, the drug is in unit dose form, wherein each unit dose form contains 0.1 to 1000 mg of itaconic acid or a derivative thereof or a pharmaceutically acceptable salt thereof.

[0024] Furthermore, the effective dose of a treatment regimen for a subject may consist of a single administration or include a series of applications. The length of the treatment period depends on various factors, such as disease severity, the subject's age, reagent concentration, patient responsiveness to the reagent, or combinations thereof. It will also be understood that the effective dose of the reagent used for treatment may be increased or decreased during a specific treatment regimen. Dosage changes can be produced and become apparent through standard diagnostic analyses known in the art. In one aspect, the reagents of the present invention may be administered before, during, or after treatment with conventional therapies for the diseases or conditions discussed, such as retinal artery occlusion.

[0025] The term "subject" refers to any member of the animal kingdom, typically a mammal. The term "mammal" refers to any animal classified as a mammal, including humans, other higher primates, domestic and agricultural animals, as well as zoo, sport, or pet animals such as dogs, cats, cows, horses, sheep, pigs, goats, rabbits, etc. Typically, the mammal is human.

[0026] This invention provides the use of a compound or a pharmaceutically acceptable salt thereof in the preparation of a medicament for treating or preventing retinal artery occlusion. The compound is selected from itaconic acid or itaconic acid derivatives. Itaconic acid derivatives have the structure shown in Formula I: CH2=C(COOR1)- CHR3- COOR2 (Formula I) Among them, R1 and R2 are each individually selected from: hydrogen, C1-C 20 Straight-chain or branched alkyl groups. In some embodiments, R1 and R2 are each individually selected from: hydrogen, C1-C2, C2-C2, C3-C4, C4-C5, C4-C5, C6 ... 10 The alkyl group; in a more preferred embodiment, it is selected from hydrogen or C1-C6 alkyl groups (e.g., methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, pentyl, or hexyl). R3 is selected from C4-C6 alkyl groups. 12 Straight-chain or branched alkyl groups. Specifically, more preferably C4-C6. 12The alkyl group, such as n-butyl, n-hexyl, n-heptyl, n-octyl, n-nonyl, n-decyl, n-undecyl, or n-dodecyl. In some embodiments, R3 is preferably n-octyl.

[0027] To better illustrate the applicability of the itaconic acid derivatives provided by this invention, the following specific examples are provided: When R1=R2=H, the straight-chain R3 can be n-butyl, n-pentyl, n-hexyl, n-heptyl, n-octyl, n-decyl, or n-dodecyl; when R1=R2=H, the branched R3 can be isobutyl, isopentyl, isohexyl, isooctyl, or isodecyl; when R1=H and R2=methyl, ethyl, or n-propyl, R3 can be n-butyl, n-hexyl, n-octyl, or n-decyl, and the corresponding branched form of isobutyl, isohexyl, or isooctyl; when R1=methyl, ethyl, or n-propyl and R2=H... When R1 = methyl, ethyl, or n-propyl, and R2 = methyl, ethyl, or n-propyl, R3 can be n-butyl, n-hexyl, n-octyl, or n-decyl, as well as branched isobutyl, isohexyl, or isooctyl, etc.

[0028] This invention reveals that itaconic acid, an endogenous metabolite with an unsaturated dicarboxylic acid structure, exhibits a significant regulatory role in the pathological process of retinal ischemic diseases. Specifically, itaconic acid can mitigate or delay the occurrence of abnormal changes in retinal tissue structure during retinal artery occlusion, promote the M2 polarization of retinal microglia, regulate retinal inflammatory responses, and inhibit retinal ganglion cell damage or death through multiple mechanisms, thus positively influencing the occurrence and development of retinal artery occlusion.

[0029] Building upon this, the present invention further discovers that itaconic acid derivatives obtained by rationally modifying the itaconic acid core structure can also exert similar or superior therapeutic effects on retinal artery occlusion as itaconic acid. Such rational structural modifications include, but are not limited to, introducing substituents onto the itaconic acid backbone, substituting adjacent carbons of double bonds, and salting or derivatizing carboxyl groups. None of these modifications destroy the key pharmacologically active structure of the itaconic acid core. As a specific embodiment of the present invention, the itaconic acid derivative is 4-octylitaconic acid. Experimental data show that 4-octylitaconic acid or its salts have a significant effect on improving retinal artery occlusion.

[0030] This invention argues that the reason itaconic acid derivatives retain their therapeutic effect on retinal artery occlusion through the aforementioned structural modifications is primarily due to two factors. Firstly, they preserve the core structural unit of itaconic acid and its related biological mechanisms of action. Simultaneously, structural modifications improve the compound's lipid solubility, cell membrane permeability, bioavailability, and targeted tissue distribution characteristics, thereby achieving more favorable pharmacodynamic performance in vivo. Secondly, these derivatives have a clear structural and functional inheritance relationship with the itaconic acid core, and their technical effects are predictable to those skilled in the art. Therefore, any itaconic acid derivatives obtained through structural modifications based on the itaconic acid core that conform to the conventional understanding of those skilled in the art should be considered equivalent or alternative embodiments of the technical solution of this invention.

[0031] The itaconic acid and its derivatives involved in this invention have their chemical structures clearly defined by general formula I, wherein the ranges of substituents R1, R2, and R3 are clearly defined in the specification, enabling those skilled in the art to accurately understand and identify the range of compounds covered by this invention. Itaconic acid, as a known compound, has its preparation methods and physicochemical properties well-documented in the prior art. Corresponding itaconic acid derivatives, especially compounds formed through alkyl substitution or esterification, can also be prepared using well-known organic synthesis methods in the art.

[0032] It should be understood that the present invention is not limited to the specific examples of itaconic acid derivatives listed in the specification. All itaconic acid derivatives within the structural range shown in Formula I can achieve the same or similar technical effects as 4-octyl itaconic acid in the specific embodiments, and are reasonable technical variations of the present invention, and are all within the protection scope of the present invention.

[0033] In this invention, pharmaceutically acceptable salts may include, but are not limited to: alkali metal salts (such as sodium salts, potassium salts, lithium salts); alkaline earth metal salts (such as magnesium salts, calcium salts); ammonium salts or organic amine salts (such as triethanolamine salts, amino acid salts, etc.).

[0034] The drugs or compositions of the present invention can be formulated into suitable forms with commonly used pharmaceutically acceptable carriers. “Pharmaceutically acceptable” means physiologically acceptable and generally does not cause allergic reactions or similar reactions, such as rashes and dizziness, when administered to humans.

[0035] In some embodiments, the drugs or compositions of the present invention can be administered via systemic routes, including but not limited to injection (e.g., intravitreal injection, intraperitoneal injection, intravenous injection). After systemic administration, the drugs or compositions can enter the systemic circulation and, under certain conditions, cross the blood-retinal barrier or exert their effects by modulating signaling pathways related to retinal inflammation and apoptosis, thereby producing a regulatory effect under retinal-related pathological conditions. It should be understood that different routes of administration, dosages, and frequencies of administration may affect the distribution characteristics and extent of action of the substances in the body. The present invention does not limit the drugs or compositions to exert their effects through a specific mechanism or pathway.

[0036] In some embodiments, the drug or composition is formulated for different routes of administration, including injectable or oral formulations. For oral administration, it can indirectly affect retinal tissue status by systemically modulating inflammatory responses or apoptosis-related processes; for injectable administration, the formulation can enhance the regulatory effect by improving bioavailability or shortening the onset time. The dosage form can be one or more of capsules, tablets, oral dosage forms, microcapsule formulations, injections, eye drops, eye washes, intraocular injection solutions, ophthalmic creams, and ophthalmic gels. Preferably, it is formulated into a dosage form suitable for the treatment of retinal artery occlusion.

[0037] In some embodiments, pharmaceutically acceptable excipients include, but are not limited to: fillers (such as microcrystalline cellulose, lactose, mannitol, etc.), binders (such as hydroxypropyl cellulose, polyvinylpyrrolidone, etc.), disintegrants (such as croscarmellose sodium, croscarmellose, etc.), lubricants (such as magnesium stearate, talc, etc.), glidants, antioxidants, preservatives, colorants, flavoring agents, surfactants, controlled-release materials, or coating materials, etc.

[0038] In some embodiments, in animal model studies, the drug or composition is administered intravitreally at a dose of 0.5 to 5 μg per animal, or intraperitoneally at a dose of 0.1 to 50 mg / kg.

[0039] The invention has now been generally described, and will be more readily understood by referring to the following embodiments, which are provided by way of example and not by way of limitation.

[0040] All experimental animals used in this invention, specifically C57BL / 6 mice, were purchased from Liaoning Changsheng Biotechnology Co., Ltd. Male C57BL / 6 mice aged 8-12 weeks and weighing 21g ± 1g were selected.

[0041] The silicone suture plugs (diameter 0.22±0.01mm, length 6.0±0.1mm) used in this embodiment of the invention were ordered from Pingdingshan Yushun Biotechnology Co., Ltd. The suture plugs were placed in a 35mm sterile petri dish, soaked in 75% alcohol for 1 hour, and then air-dried in a biosafety cabinet for later use.

[0042] The 4-octyl itaconate (4-OI) powder used in this embodiment of the invention was purchased from Target Mol, catalog number T4580. It was dissolved using dimethyl sulfoxide (DMSO) to obtain a 100 mg / mL 4-octyl itaconate stock solution, which was stored at -20°C for later use. The molecular formula of 4-octyl itaconate (4-OI) is C1. 13 H 22 O4, molecular weight 242.31, CAS number 3133-16-2, structure as follows: .

[0043] Preparation of the 4-octylitaconic acid intraperitoneal injection working solution: A mixture of polyethylene glycol 12-hydroxystearate (Solutol HS-15, TargetMol), polyethylene glycol 400 (PEG400, TargetMol), and double-distilled water (ddH2O) at a volume ratio of 2:4:13 was prepared. One volume of the 4-octylitaconic acid stock solution (100 mg / mL) was added to 19 volumes of diluent, and diluted 20-fold to obtain the working solution with a final concentration of 2.5 mg / mL. The control solution was prepared by mixing one volume of DMSO with 19 volumes of diluent. All solutions were sonicated until clear. Mice were weighed before administration, and the dosage volume was adjusted daily based on body weight, with the final intraperitoneal dose being 50 mg / kg. All drug preparations were performed in a biosafety cabinet.

[0044] Preparation of working solution for intravitreal injection of 4-octylitaconic acid: Prepare a 2.5 mg / mL working solution from a 100 mg / mL stock solution of 4-octylitaconic acid using the same method as described for intraperitoneal injection. The control solution is prepared using flux + DMSO. The injection volume per eyeball is 1 μL.

[0045] Preparation of 4-octylitaconic acid working solution for cell experiments: 100 mg / mL of 4-octylitaconic acid stock solution was prepared into a 60 μg / mL working solution using complete DMEM medium.

[0046] The drugs and reagents used in the embodiments of this invention: isoflurane and oxygen / nitrous oxide mixture were purchased from Shenzhen Ruiwode Technology Co., Ltd. Polyethylene glycol 12-hydroxystearate and polyethylene glycol 400 were purchased from TargetMol. Other conventional antibodies and reagents were commercially available products.

[0047] The instruments and equipment used in the embodiments of this invention are as follows: the surgical microscope is an Olympus SZX7 from Japan; the electroretinogram (ERG) system is a RetiMINER 4.0 from Chongqing Airsys; the fiber injection system for vitrectomy is a Nanoject III from the USA; the fluorescence confocal microscope is a TCS-SP8 from Germany; and the inverted fluorescence microscope is an Olympus CellSens from Japan. All other instruments are common commercially available products.

[0048] This invention uses a mouse model of retinal ischemia-reperfusion injury caused by retinal artery occlusion. The model uses a silicone suture to block blood flow from the pterygopalatine artery to the ophthalmic artery in mice, simulating the pathological process of retinal ischemia-reperfusion in retinal artery occlusion. Establishment of a mouse model of retinal artery occlusion: Healthy male C57BL / 6 mice aged 8-12 weeks, weighing 21g±1g, were selected and housed for one week in an IVC system environment with a temperature of 24-17℃, relative humidity of 60%, and free access to food and water. Mice were randomly divided into two groups to establish a unilateral pterygopalatine artery and ophthalmic artery occlusion (UPOAO) model. Specifically, 8-week-old male C57BL / 6 mice were general anesthetized, and their necks were shaved and disinfected. The left common carotid artery, internal carotid artery, and external carotid artery were isolated by exposing the neck. The distal end of the external carotid artery was ligated with 8-0 silk suture, and another 8-0 silk suture was inserted into the external carotid artery, with a slipknot tied near the bifurcation of the common carotid artery. To maintain reperfusion and avoid hemorrhage and death during the procedure, a specially designed silicone suture plug was inserted through the external carotid artery, rather than through the common carotid artery. Using ophthalmic scissors, a small inverted "V" shaped incision is made between the two sutures on the external carotid artery. A sterile silicone suture (size 6021) is inserted 1 mm below the ligation line of the external carotid artery. The suture is gently pushed with forceps until the silicone end of the suture is fully inserted into the pterygopalatine artery. The external carotid artery is cut at the small incision. The suture is pulled back, flipped, and inserted into the internal carotid artery, then inserted outward into the pterygopalatine artery. The insertion depth of the suture is approximately 6 ± 1 mm from the bifurcation of the common carotid artery, with the silicone end of the suture roughly positioned at the bifurcation. A slipknot is slightly tightened. The ligation of the internal carotid artery is then removed, and the previously cut external carotid artery is severed. The silicone suture is then pulled back to the bifurcation of the common carotid artery, rotated counterclockwise, and inserted into the internal carotid artery, further into the pterygopalatine artery. The silicone end of the suture is positioned near the bifurcation of the common carotid artery, effectively occluding the ophthalmic artery. The slipknot is then tightened, and the skin is sutured. Two hours after the ischemic embolism, the silicone suture is carefully removed from the pterygopalatine artery to avoid significant bleeding. The sutures in the common carotid artery were removed to restore arterial reperfusion. Finally, the skin wound was sutured, and the mice were routinely fed during reperfusion to obtain the UPOAO model mice.

[0049] Example 1 Evaluation of the therapeutic efficacy of intracavitary injection of 4-octylitaconic acid (hereinafter referred to as 4-OI) for RAO.

[0050] The experimental C57BL / 6 mice were randomly divided into 3 groups.

[0051] 4-OI treatment group (UPOAO+4-OI): Mice were injected intravitreally with 2.5 mg / mL of 4-OI working solution one day before UPOAO modeling. 1 μL was injected into each eye. Control mice were injected with an equal volume of control solution. Unilateral UPOAO modeling was performed one day after injection.

[0052] Model control group (UPOAO): Mice were fed a regular diet daily to induce UPOAO in one eye.

[0053] Healthy group (Con): Mice that underwent sham surgery were fed a regular diet daily and received intravitreal injections of the same volume of control solvent as the mice in the 4-OI intervention group.

[0054] Three days after modeling, general anesthesia was administered via intraperitoneal injection of 2% sodium pentobarbital at a dose of 0.3 mL per 100g of body weight. Compound topiramate eye drops were then instilled for 5 minutes to dilate the pupils. Fundus and retinal structures were assessed using optical coherence tomography (OCT). Figure 1 As shown, compared with the healthy group (Con), the mice in the model control group (UPOAO) showed retinal pallor and white lesions in the fundus after retinal ischemia, and OCT images showed significant edema and thickening of the inner retinal structure. In contrast, the mice in the 4-OI treatment group (UPOAO+4-OI) did not show retinal pallor, white lesions in the fundus, and OCT results did not show significant edema and thickening of the inner retinal layer. A comparison between the 4-OI treatment group (UPOAO+4-OI) and the model control group (UPOAO) revealed that the retinal structure was more intact after 4-OI treatment. This indicates that intravitreal injection of 4-OI has a significant protective effect on retinal structure.

[0055] Seven days after modeling, changes in visual function in mice were evaluated using electroretinography (ERG). Mice were generally anesthetized by intraperitoneal injection of 0.3 mL of 2% sodium pentobarbital per 100 g body weight. After the anesthesia stabilized, compound topiramate eye drops were instilled to dilate the pupils for 15 minutes. The dilated mice were then placed on an ERG control table, and the amplitude changes of the a and b waves were recorded using different light intensities. Figure 2 As shown, compared with the healthy group (Con), the model control group (UPOAO) showed no difference in wave a, but wave b was significantly decreased. However, after 4-OI treatment, compared with the model control group (UPOAO+4-OI), the 4-OI treatment group (UPOAO+4-OI) showed no statistically significant difference in wave a under different stimulation intensities, but wave b was significantly increased. P < 0.05 P < 0.01 P < 0.001 The result (P<0.0001) indicates that intravitreal injection of 4-OI has a protective effect on visual function in UPOAO model mice.

[0056] Example 2 The protective effect of 4-OI on retinal ganglion cells (RGCs).

[0057] The number of random globulins (RGCs) was characterized and statistically analyzed. Retinal samples from three groups of mice were fixed with 4% paraformaldehyde for 60 minutes. The retina was dissected and intact, and the retina was immersed in 5% BSAT solution and sealed overnight at 4°C. The samples were incubated with Brn3a goat serum primary antibody at 4°C for 48 hours, rinsed in PBS buffer, and then incubated overnight at 4°C with Alexa Fluor 594 donkey anti-goat fluorescent secondary antibody. After rinsing in PBS buffer, the slides were mounted with anti-fluorescence quenching mounting medium and photographed using an inverted microscope for RGC counting. The results are shown below. Figure 3 As shown, the number of RGCs decreased in the model control group (UPOAO) compared to the healthy group (Con), while the number of RGCs increased significantly in the 4-OI treatment group (UPOAO+4-OI) compared to the model control group (UPOAO). Figure 4 The results show that the number of RGCs was statistically significant in the 4-OI treatment group (UPOAO+4-OI). P < 0.05 P < 0.01 P < 0.001 The result (P<0.0001) indicates that intravitreal injection of 4-OI significantly reduced apoptosis of RGCs in UPOAO model mice and had a significant protective effect on RGCs.

[0058] Example 3 Effects of intravitreal injection of 4-OI on retinal microglial cell polarization.

[0059] Retinal samples were collected from three groups of mice, fixed with 4% paraformaldehyde for 60 minutes, and then dissected for histological examination. The fixed samples were subjected to Iba1 / CD206 immunofluorescence double labeling to assess the effect of 4-OI on the number and polarization status of microglia. Results are as follows: Figure 5 As shown, there was no difference in M2 microglia between the model control group (UPOAO) and the healthy group (Con), while the 4-OI treatment group (UPOAO+4-OI) showed a significant increase in M2 microglia compared to the model control group (UPOAO). This indicates that intravitreal injection of 4-OI can promote the polarization of microglia towards the anti-inflammatory M2 type, thus playing a protective role in RAO damage.

[0060] Example 4 Evaluation of the therapeutic efficacy of intraperitoneal injection of 4-OI for RAO.

[0061] The experimental C57BL / 6 mice were randomly divided into 3 groups.

[0062] 4-OI treatment group (UPOAO+4-OI): Mice were intraperitoneally injected with 2.5 mg / mL of 4-OI working solution 6 hours before UPOAO modeling, with a final intraperitoneal dose of 50 mg / kg. Unilateral UPOAO modeling was performed 6 hours after injection. One day after UPOAO modeling, a second injection at the same dose of 50 mg / kg was administered.

[0063] Model control group (UPOAO): Mice were fed a regular diet daily to induce UPOAO in one eye.

[0064] Healthy group (Con): Mice undergoing sham surgery were fed a regular diet daily and received an intraperitoneal injection of the same volume of control solvent as the 4-OI intervention group. The injection time was the same as the 4-OI treatment group.

[0065] Three days after modeling, the fundus and retinal structures were assessed using the fundus and optical coherence tomography (OCT) module, following the same method as above. Figure 6 As shown, compared with the healthy group (Con), the mice in the model control group (UPOAO) showed retinal pallor and white lesions in the fundus after retinal ischemia, and OCT images showed significant edema and thickening of the inner retinal structure. In contrast, the mice in the 4-OI treatment group (UPOAO+4-OI) did not show retinal pallor, white lesions in the fundus, and OCT results did not show significant edema and thickening of the inner retinal layer. A comparison between the 4-OI treatment group (UPOAO+4-OI) and the model control group (UPOAO) revealed that the retinal structure was more intact after 4-OI treatment. This indicates that intraperitoneal injection of 4-OI also has a significant protective effect on retinal structure.

[0066] Seven days after modeling, changes in visual function in mice were evaluated using electroretinography (ERG), following the same intravitreal procedure. Results were as follows: Figure 7 As shown, compared with the healthy group (Con), the model control group (UPOAO) showed no difference in wave a, but wave b was significantly decreased. However, after 4-OI treatment, compared with the model control group (UPOAO+4-OI), the 4-OI treatment group (UPOAO+4-OI) showed no statistically significant difference in wave a under different stimulation intensities, but wave b was significantly increased. P < 0.05 P < 0.01 P < 0.001 The result (P<0.0001) indicates that intraperitoneal injection of 4-OI also has a protective effect on visual function in UPOAO model mice.

[0067] Example 5 Study on the protective effect of intraperitoneal injection of 4-OI on retinal ganglion cells (RGCs).

[0068] The method is the same as the intravitreal injection method described above. The results are as follows: Figure 8 As shown, the number of RGCs decreased in the model control group (UPOAO) compared to the healthy group (Con), while the number of RGCs increased significantly in the 4-OI treatment group (UPOAO+4-OI) compared to the model control group (UPOAO). Figure 9 The results show that the number of RGCs was statistically significant in the 4-OI treatment group (UPOAO+4-OI). P < 0.05 P < 0.01 P < 0.001 The result (P<0.0001) indicates that intraperitoneal injection of 4-OI can also significantly reduce the apoptosis of RGCs in UPOAO model mice, and has a significant protective effect on RGCs.

[0069] Example 6 Effects of intraperitoneal injection of 4-OI on retinal microglia polarization.

[0070] The method is the same as the intravitreal injection described above. The results are as follows: Figure 10 As shown, there was no difference in M2 microglia between the model control group (UPOAO) and the healthy group (Con), while the 4-OI treatment group (UPOAO+4-OI) showed a significant increase in M2 microglia compared to the model control group (UPOAO). This indicates that intraperitoneal injection of 4-OI can also promote the polarization of microglia towards the anti-inflammatory M2 type, thus playing a protective role in RAO damage.

[0071] Example 7 Effects of 4-OI on inflammatory factors in microglia.

[0072] Mouse microglia (BV2) cell line were cultured in vitro, and an in vitro oxygen-glucose deprivation (OGD) model was constructed using a three-gas incubator to simulate the ischemia-reperfusion pathological process of model cells in recurrent oxygen reflux (RAO). Specifically, cells were plated during the logarithmic growth phase and subjected to OGD for 6 hours, followed by reoxygenation for 24 hours. Cells were divided into three groups: a normal control group (Con), a model control group (OGD / R), and a 4-OI intervention group (OGD / R+4-OI). In the 4-OI intervention group (OGD / R+4-OI), 100 mg / mL of 4-OI stock solution was prepared into a 60 μg / mL working solution using complete DMEM medium to stimulate cells. The control group (Con) was stimulated with the same volume of solvent.

[0073] After 6 hours of oxygen and glucose deprivation and 24 hours of reoxygenation, the supernatants of three groups of cells were collected, and the levels of IL-10 and TGFβ were detected using an ELISA kit. Figure 11 The results for IL-10 are shown in the figure. Compared with the control group (Con), the OGD / R control group (OGD / R) showed a significant decrease in the inhibition of the inflammatory factor IL-10. P < 0.05 P < 0.01 P < 0.001 P < 0.0001. The 4-OI intervention group (OGD / R + 4-OI) showed a significantly higher IL-10 level compared to the model control group (OGD / R). P < 0.05 P < 0.01 P < 0.001 (P < 0.0001). Figure 12 The results for TGFβ are shown in the figure. Compared with the control group (Con), the control group (OGD / R) showed a significant decrease in the inhibition of the inflammatory factor TGFβ. P < 0.05 P < 0.01 P < 0.001 P < 0.0001. The 4-OI intervention group (OGD / R + 4-OI) showed a significantly higher IL-10 level compared to the model control group (OGD / R). P < 0.05 P < 0.01 P < 0.001 (P<0.0001). The above results indicate that 4-OI can significantly increase the expression of anti-inflammatory factors IL-10 and TGFβ, and play a protective role in RAO.

[0074] For any points not covered above, existing technologies shall apply.

[0075] Although specific embodiments of the present invention have been described in detail by way of examples, those skilled in the art should understand that the above examples are for illustrative purposes only and are not intended to limit the scope of the invention. Those skilled in the art can make various modifications or additions to the described specific embodiments or use similar methods to replace them, without departing from the direction of the invention or exceeding the scope defined by the appended claims. Those skilled in the art should understand that any modifications, equivalent substitutions, improvements, etc., made to the above embodiments based on the technical essence of the present invention should be included within the protection scope of the present invention.

Claims

1. Use of an itaconic acid compound or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating retinal artery occlusion, wherein, The itaconic acid compound has a structure as shown below: 。 2. Use according to claim 1, characterized in that, The treatment includes improvement of disease symptoms or improvement of health conditions of the subject.

3. Use according to claim 1, characterized in that, The use includes alleviating or delaying the extent of abnormal changes in retinal tissue structure in retinal artery occlusion.

4. Use according to claim 1, characterized in that, The use includes inhibiting damage or death of retinal ganglion cells.

5. Use according to claim 1, characterized in that, The use includes regulating retinal inflammatory response associated with retinal artery occlusion.

6. Use according to claim 5, characterized in that, The retinal inflammatory response includes promoting M2-type polarization of retinal microglia.

7. Use according to claim 1, characterized in that, The drug is administered by intraperitoneal injection, intravitreal administration, intravenous injection or oral administration.

8. Use according to claim 7, characterized in that, In animal models, the drug is administered by intravitreal administration at a dose of 0.5-5 μg per animal, or by intraperitoneal injection at a dose of 0.1-50 mg / kg.

9. Use according to claim 7, characterized in that, The dosage form of the drug is one or more of a capsule, a tablet, an oral agent, a microcapsule preparation, an injection, an eye drop, an eye wash, an eye cream, and an eye gel.

10. Use according to any one of claims 1 to 9, characterized in that, The drug is administered before the occurrence of retinal artery occlusion.

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