Ocular composition

Abstract

The present invention provides an ocular composition comprising hyaluronic acid nanoparticles with a size of about 1 to 100 nm, which exhibits improved intraocular retention, wherein the hyaluronic acid nanoparticles are cross-linked between hyaluronic acid molecules to form a network structure.

Description

composition for the eye

[0001] The present invention relates to an ocular composition with improved retention within the eye by including hyaluronic acid nanoparticles.

[0002] Eye drops are liquid medications administered into the eye to protect the eye and to prevent and treat eye diseases. Because eye drops can be easily administered to the eye using a non-invasive method, they are used as a treatment for dry eye syndrome, glaucoma, allergies, and various other eye conditions.

[0003] On the other hand, eye drops can cause problems regarding tear drainage and loss of the ocular surface barrier; furthermore, they present the inconvenience of requiring regular and consistent repeated application at predetermined times. In other words, the medicinal effect of the substances contained in conventional eye drops may be reduced due to the limitations of contact force on the ocular surface.

[0004] To overcome these drawbacks, various methods have been proposed to increase viscosity by including specific substances, thereby improving retention on the ocular surface. In particular, hyaluronic acid, a biocompatible material, is the most widely used due to its high water retention and viscosity, which allow it to remain on the ocular surface for a relatively long time. However, conventional hyaluronic acid has a linear bonding structure, which limits its ability to improve adhesion and retention time on the actual ocular surface.

[0005] The background description of the invention is provided to facilitate a better understanding of the present invention. The matters described in the background description should not be construed as an acknowledgment that they exist as prior art.

[0006] Meanwhile, the inventors of the present invention anticipated that the retention of eye drops on the ocular surface could be improved if the structural limitations of conventional hyaluronic acid were overcome; accordingly, the inventors focused on hyaluronic acid with a reticular structure rather than hyaluronic acid with a linear structure.

[0007] More specifically, hyaluronic acid (HA) is a complex negatively charged polysaccharide and a high-molecular-weight compound composed of N-acetylglucosamine and glucuronic acid, which is present in connective tissue, subcutaneous tissue, nerve tissue, or joint synovial fluid in the body. Furthermore, due to its excellent biocompatibility, HA is a natural polymer widely utilized in various fields, such as cosmetic or medical purposes. In its natural state, HA primarily exists in the extracellular matrix and plays a role in stabilizing tissue structures by forming matrix fluid through mixing with fibrous tissue and collagen. As a substance found in the tissues of all vertebrates, it is an ideal chemical substance for use as a component for tissue repair purposes due to its lack of immunogenicity.

[0008] These HAs can be converted into gel-type polymers through crosslinking, which enhances viscosity and can extend in vivo degradation and retention time. For example, to increase the chemical structural stability of hyaluronic acid and slow down its degradation in vivo to maintain its physical properties for a longer period, technologies have been developed to stabilize hyaluronic acid by adding crosslinking agents such as 1,4-butanediol diglycidyl ether (BDDE). Among these, the maximum in vivo retention period of existing hyaluronic acid crosslinkers and hyaluronic acid derivative crosslinkers is known to be about 6 to 24 months.

[0009] However, high-viscosity hyaluronic acid crosslinks obtained by simply increasing the amount of crosslinking agent to enhance the in vivo retention time of such hyaluronic acid have a problem in that they may degrade in vivo, or unreacted residual crosslinking agent components may cause toxicity and trigger inflammatory reactions. Furthermore, removing these residual crosslinking agents requires an additional removal process, which poses a problem of economic inefficiency in the hyaluronic acid production process.

[0010] Accordingly, the inventors of the present invention focused on a method of HA crosslinking through a physical method. More specifically, the inventors of the present invention discovered that when HA is irradiated with radiation as a physical crosslinking method, HA undergoes a crosslinking reaction without the need for a separate crosslinking agent, thereby forming a network-structured HA gel. In other words, when using HA with a network structure produced through a physical crosslinking method, problems regarding crosslinking agents that may cause toxicity and side effects can be prevented, and it can be economical as no additional processes are required.

[0011] Meanwhile, hyaluronic acid has been used in ocular and ocular eye drops, but in conventional ocular and ocular eye drops, hyaluronic acid with a linear structure and a particle size of 200 to 1000 nm has been mainly used. However, conventional hyaluronic acid has not been able to provide sufficient moisture and adhesion to additives in ocular eye drops. Accordingly, the inventors of the present invention recognized that if the aforementioned HA with a network structure is used in eye drops, the limitations of conventional eye drops containing hyaluronic acid can be overcome.

[0012] Furthermore, the inventors of the present invention recognized that if the HA gel with a network structure is converted to a specific nano-particle size, it can be included in an eye drop, i.e., an ocular composition, and the retention force of the composition within the eye can be further improved.

[0013] Ultimately, the inventors of the present invention developed an ocular composition comprising hyaluronic acid (HA) nanoparticles having a network structure and a size of about 1 to 100 nm.

[0014] Accordingly, the problem that the present invention aims to solve is to provide an ocular composition with improved retention within the eye by including hyaluronic acid nanoparticles.

[0015] In addition, another problem that the present invention aims to solve is to provide an ocular drug delivery system comprising hyaluronic acid nanoparticles having a size of about 10 to 100 nm.

[0016] The problems of the present invention are not limited to those mentioned above, and other unmentioned problems will be clearly understood by those skilled in the art from the description below.

[0017] In order to solve the problem described above, the present invention provides an ocular composition with improved retention force in the eye, comprising hyaluronic acid (HA) nanoparticles having a size of about 1 to 100 nm, wherein the hyaluronic acid nanoparticles are cross-linked between hyaluronic acid molecules to form a network structure.

[0018] According to the features of the present invention, the residual force may include the residual time and residual amount within the cornea.

[0019] According to another feature of the present invention, the retention time of the ocular composition in the cornea can be improved by about 60 minutes or more compared to a composition containing hyaluronic acid, but is not limited thereto.

[0020] According to another feature of the present invention, the residual amount of the ocular composition in the cornea can be improved by about 20 times or more compared to a composition containing hyaluronic acid, but is not limited thereto.

[0021] According to another feature of the present invention, after 60 minutes of administration, the residual amount of the ocular composition in the cornea can be improved by about 1.5 times or more compared to a composition containing hyaluronic acid, but is not limited thereto.

[0022] According to another feature of the present invention, the size of the hyaluronic acid nanoparticles is disclosed to be about 1 to 100 nm, but is not limited thereto, and preferably can be about 3 to 70 nm.

[0023] In order to solve other problems as described above, the present invention provides an ocular drug delivery system comprising hyaluronic acid (HA) nanoparticles having a size of about 1 to 100 nm, wherein the hyaluronic acid nanoparticles are cross-linked between hyaluronic acid molecules to form a network structure.

[0024] According to a feature of the present invention, the ocular drug delivery system may induce a pharmacologically active substance to be bound within the hyaluronic acid nanoparticles or retained within the network structure of the nanoparticles so as to adhere to the eye.

[0025] According to another feature of the present invention, the pharmacologically active substance may be an active ingredient effective in preventing or treating ophthalmic diseases.

[0026] The present invention can provide an ocular composition with improved retention within the eye. More specifically, hyaluronic acid in conventional ocular compositions has mainly been used as linear hyaluronic acid with a particle size of 200 nm or more, which has not provided sufficient moisture retention and adhesive strength of additives in the ocular composition. Meanwhile, the inventors of the present invention recognized and discovered that the limitations of conventional ocular compositions could be overcome by using nano-sized hyaluronic acid in the ocular composition. Accordingly, the present invention, by including hyaluronic acid nanoparticles, can improve the viscosity of the composition and enhance adhesion within the cornea. Furthermore, the hyaluronic acid nanoparticles according to the present invention can retain moisture due to their structural characteristic of a network structure, and accordingly, the present invention can provide a moisturizing effect within the eye.

[0027] Furthermore, depending on the structural characteristics of the hyaluronic acid nanoparticles according to the present invention, the retention of additional additive substances within the eye can be enhanced, thereby further improving the pharmacological effects of the additive substances. That is, the present invention can provide a drug delivery system capable of sustaining the pharmacological effects of additives within the eye for a long period of time and efficiently delivering them.

[0028] Ultimately, the present invention can provide an improved eye drop that can promote eye health.

[0029] The effects according to the present invention are not limited to those exemplified above, and various other effects are included in this specification.

[0030] FIG. 1 is a flowchart of a method for manufacturing an ocular composition according to one embodiment of the present invention.

[0031] Figure 2 is the result of the particle size distribution of HA nanoparticles of an ocular composition according to one embodiment of the present invention.

[0032] Figures 3a and 3b show the results of ocular mucosal adhesion according to the size of nanoparticles used in an ocular composition according to one embodiment of the present invention.

[0033] FIGS. 4a to 4c are ex vivo microscopic images of the corneal surface for an ocular composition according to one embodiment of the present invention and the results of quantification thereof.

[0034] Figures 5a and 5b are in vivo microscopic images of the surface of a living cornea for an ocular composition according to one embodiment of the present invention and the results of quantification thereof.

[0035] FIGS. 6a to 6c are in vivo microscopic images of the surface of an excised cornea for an ocular composition according to one embodiment of the present invention and the results of quantification thereof.

[0036] The advantages and features of the present invention and the methods for achieving them will become clear by referring to the embodiments described below in detail together with the accompanying drawings. However, the present invention is not limited to the embodiments disclosed below but may be implemented in various different forms. These embodiments are provided merely to ensure that the disclosure of the present invention is complete and to fully inform those skilled in the art of the scope of the invention, and the present invention is defined only by the scope of the claims.

[0037] In this specification, expressions such as “have,” “may have,” “include,” or “may include” indicate the presence of such features (e.g., numerical values, functions, operations, or components such as parts) and do not exclude the presence of additional features.

[0038] In this specification, expressions such as “A or B,” “at least one of A or / and B,” or “one or more of A or / and B” may include all possible combinations of items listed together. For example, “A or B,” “at least one of A and B,” or “at least one of A or B” may refer to cases including (1) at least one A, (2) at least one B, or (3) both at least one A and at least one B.

[0039] The expression “configured to” as used in this specification may be replaced, depending on the context, with, for example, “suitable for,” “having the capacity to,” “designed to,” “adapted to,” “made to,” or “capable of.”

[0040] As used in this specification, the term "or" means "and / or" unless otherwise stated.

[0041] The term "prevention and treatment of ophthalmic diseases" as used in the present invention refers to the prevention and treatment of corneal and conjunctival epithelial damage caused by endogenous diseases such as Sjögren's syndrome, Stevens-Johnson syndrome, or dry eye syndrome, or exogenous diseases caused by medication after surgery, trauma, or the use of hard contact lenses. The above ophthalmic diseases are not necessarily limited thereto.

[0042] The term 'pharmaceuticalally acceptable salt' as used in the present invention refers to an acidic or basic salt that may be present in the hyaluronic acid and carboxymethylcellulose of the present invention, unless otherwise indicated. For example, the pharmaceutically acceptable salt includes sodium salts, potassium salts, calcium salts, etc., and preferably includes sodium salts.

[0043] As used herein, the term “about” refers to a normal margin of error for each value that is readily known to those skilled in the art. In this specification, the designation of an “about” value or parameter includes an example relating to the value or parameter itself. Furthermore, unless otherwise stated or otherwise evident from the context, the term “about” indicates a range of values ​​corresponding to within 10% in either direction (greater than or less than) a mentioned reference value.

[0044] As used herein, the term “patient or individual” refers interchangeably to any single animal requiring treatment, more preferably a mammal (including such non-human animals, e.g., cats, dogs, horses, rabbits, zoo animals, cattle, pigs, sheep, and non-human primates). In various embodiments of this specification, the patient referred to may be a human.

[0045] The term “Drug Delivery System (DDS)” as used in this invention refers to a system designed to efficiently deliver a drug to a specific site in the body, minimize side effects, and maximize drug efficacy.

[0046] As used in this invention, the term “drug” refers to a substance administered to an individual to treat, prevent, or suppress diseases, injuries, or other physical abnormalities. Drugs can be used by methods such as ingestion, topical application, or injection, and are chemical substances that exhibit pharmacological biological effects.

[0047] The term “synthetic compound” as used in this invention refers to a compound manufactured through artificial chemical reactions or processes, and refers to a chemical substance synthesized under laboratory or industrial conditions rather than originating directly from nature.

[0048] The term “natural compound” as used in this invention refers to a compound derived from natural products, meaning a chemical substance found in nature or produced by living organisms. Such natural compounds originate from various living organisms, such as plants, animals, and microorganisms, and may possess physiological or pharmacological activity.

[0049] The term “adjuvant” as used in this invention refers to a substance used to enhance the efficacy of a specific drug or treatment, and may be used in conjunction with drug therapy. This invention includes all adjuvant substances used to alleviate inflammatory responses to reduce pain and minimize damage caused by inflammation.

[0050] The term “nanoparticle” as used in the present invention refers to various particles ranging in size from 1 to 100 nanometers (nm) used to deliver pharmaceuticals or proteins to specific tissues or cells within the body.

[0051] The terms used in this document are used merely to describe specific embodiments and are not intended to limit the scope of other embodiments. Singular expressions may include plural expressions unless the context clearly indicates otherwise. Terms used herein, including technical or scientific terms, may have the same meaning as generally understood by those skilled in the art described in this document. Terms used in this document that are defined in general dictionaries may be interpreted as having the same or similar meaning as they have in the context of the relevant technology, and are not to be interpreted in an ideal or overly formal sense unless explicitly defined in this document. In some cases, even terms defined in this document may not be interpreted to exclude the embodiments of this document.

[0052] The features of each of the various embodiments of the present invention may be combined or combined with one another, either partially or wholly, and as will be fully understood by those skilled in the art, various technical interlocking and operation are possible, and each embodiment may be implemented independently of one another or together in an interrelationship.

[0053] ocular composition and method of manufacturing the same

[0054] Hereinafter, with reference to FIGS. 1 and 2, a method for manufacturing an ocular composition according to one embodiment of the present invention will be described in detail.

[0055] FIG. 1 is a flowchart illustrating a method for manufacturing an ocular composition according to one embodiment of the present invention. For convenience of explanation, the description will be explained with reference to FIG. 2.

[0056] A method for preparing an ocular composition according to one embodiment of the present invention may include a step (S110) of including hyaluronic acid (HA) nanoparticles.

[0057] The HA nanoparticles in the step (S110) containing nanoparticles may refer to nanoparticles having a size of about 1 to 100 nm, which have a three-dimensional network structure formed by cross-linking between hyaluronic acid molecules. At this time, physical methods may be used for cross-linking between hyaluronic acid molecules, for example, the HA nanoparticles in the ocular composition according to one embodiment of the present invention may originate from an HA gel formed by cross-linking through irradiation with radiation. At this time, the radiation may include electron beams, gamma rays, and X-rays, and the HA nanoparticles in the ocular composition according to one embodiment of the present invention may be formed through irradiation from at least one of electron beams, gamma rays, and X-rays.

[0058] In this regard, referring to FIG. 2, the particle size distribution results of HA nanoparticles of an ocular composition according to one embodiment of the present invention are shown.

[0059] First, the HA nanoparticles of the ocular composition according to one embodiment of the present invention are shown to have a distribution of 1 to 100 nm, and more preferably, about 3 to 70 nm.

[0060] Next, it is shown that the average size (Z-Average) of the HA nanoparticles in the ocular composition according to one embodiment of the present invention is 7.955.

[0061] Next, it is found that the size of the particle most abundantly contained within the HA nanoparticles of the ocular composition according to one embodiment of the present invention is 10.42 nm.

[0062] Accordingly, referring again to FIG. 1, the HA nanoparticles of the ocular composition according to one embodiment of the present invention have a size of about 3 to 70 nm, and by including HA particles of such a specific size in the ocular composition, the ocular composition of the present invention may have improved retention power in the eye. At this time, the retention power may include the retention time and retention amount in the cornea.

[0063] Ultimately, since the retention time in the cornea of ​​the ocular composition according to one embodiment of the present invention formed by the aforementioned process can be improved by about 60 minutes or more compared to a composition containing hyaluronic acid that is not three-dimensionally cross-linked, the ocular composition according to one embodiment of the present invention containing HA nanoparticles can have an improved retention time in the cornea compared to conventional ocular compositions.

[0064] Furthermore, as the residual amount in the cornea of ​​the ocular composition according to one embodiment of the present invention formed by the aforementioned process can be improved by about 20 times or more compared to a composition containing hyaluronic acid that is not three-dimensionally cross-linked, the ocular composition according to one embodiment of the present invention containing HA nanoparticles can have an improved residual amount in the cornea compared to conventional ocular compositions.

[0065] Meanwhile, for the therapeutic and preventive functions of the ocular composition, the method for preparing an ocular composition may further include, but is not limited to, antibacterial agents, antiviral agents, steroid agents, antihistamine agents, mast cell stabilizers, artificial tears, analgesic and anti-inflammatory agents, immunosuppressants, carbonic anhydrase inhibitors, prostaglandin derivatives, cholinergics, α-agonists, and β-blockers, and may further include all additive substances used as eye drops for the treatment and prevention of the eye.

[0066] For example, the HA nanoparticles of the ocular composition according to one embodiment of the present invention may have the same structure as the HA gel, as they are generated by splitting from a cross-linked gel. That is, the HA nanoparticles of the ocular composition according to one embodiment of the present invention, like the HA gel, may have a limited liquid retention capacity or degree of swelling until saturation, and thus can absorb and retain a predetermined amount of moisture (water) or substances. Accordingly, the aforementioned additives as eye drops may be bound within the HA nanoparticles of the ocular composition according to one embodiment of the present invention or retained within the structure of the nanoparticles and adhered to the eye, so the ocular composition according to one embodiment of the present invention can be used as a drug delivery system. Furthermore, since the ocular composition according to one embodiment of the present invention includes HA nanoparticles, the retention force within the eye may be improved, and the aforementioned additives may also have their retention force within the eye improved by the ocular composition according to one embodiment of the present invention.

[0067] Ultimately, the ocular composition according to one embodiment of the present invention can enhance the retention of additive substances within the eye as eye drops, thereby more effectively improving the pharmacological effects of the additive substances. Furthermore, since the ocular composition according to one embodiment of the present invention can retain moisture (water) in addition to the aforementioned additive substances, it can provide a moisturizing effect on the cornea. Therefore, the ocular composition according to one embodiment of the present invention can simultaneously include an enhanced drug delivery system and a moisture retention function.

[0068] According to the above process, the method for manufacturing an ocular composition according to one embodiment of the present invention can provide an ocular composition with improved residual force within the eye.

[0069] ocular drug delivery system

[0070] An ocular drug delivery system according to one embodiment of the present invention comprises hyaluronic acid (HA) nanoparticles having a size of about 1 to 100 nm, and the hyaluronic acid nanoparticles may be cross-linked between hyaluronic acid molecules to form a network structure, but are not limited thereto.

[0071] In one embodiment of the present invention, the ocular drug delivery system may induce a pharmacologically active substance to be bound within the hyaluronic acid nanoparticles or retained within the network structure of the nanoparticles so as to adhere to the eye, but is not limited thereto.

[0072] In one embodiment of the present invention, the pharmacologically active substance refers to any active ingredient that exhibits a pharmacological effect directly or indirectly in vivo, and the active ingredient may include not only a substance that directly exhibits a pharmacological effect but also an adjuvant that assists or enhances the effect. In the present invention, it refers to an active ingredient that is effective in the prevention or treatment of ophthalmic diseases.

[0073] At this time, the pharmacologically active substance may include not only active ingredients in the form of compounds such as synthetic compounds and natural compounds, but also active ingredients in the form of non-compounds such as inorganic substances (calcium, minerals, etc.), peptides, and extracts, and may include any substance that can be supported within the hyaluronic acid nanoparticles used in the present invention without limitation.

[0074] The present invention will be explained in more detail below through examples. However, since these examples are merely illustrative of the present invention, the scope of the present invention should not be interpreted as being limited by these examples.

[0075] Confirmation and verification of an ocular composition according to one embodiment of the present invention

[0076] Hereinafter, with reference to FIGS. 3a to 6c, an ocular composition according to one embodiment of the present invention will be identified and verified.

[0077] Figure 3a is an image showing the results of ocular mucosal adhesion according to the size of nanoparticles used in an ocular composition according to one embodiment of the present invention. In this case, Figures 3a and 3b, which will be described later, are methods for verifying ocular adhesion in an in vivo environment, which were performed in C57BL / 6 mice. More specifically, Comparative Example 1, Comparative Example 2, Comparative Example 3, and Example were administered to the eyes of mice pre-anesthetized with isoflurane. In this case, Example (3D hyaluronic acid 10 nm) is an ocular composition according to one embodiment of the present invention, containing 1000 μg / ml HA nanoparticles, Comparative Example 1 (hyaluronic acid) is a composition containing 1000 μg / ml of conventional linear hyaluronic acid, and Comparative Examples 2 and 3 are compositions containing 1000 μg / ml of hyaluronic acid particles having sizes of 200 nm and 600 nm, respectively. At this time, the hyaluronic acid particles included in Comparative Examples 2 and 3 are hyaluronic acid particles having the same structure (network structure) as the HA nanoparticles of the ocular composition according to one embodiment of the present invention, but differ in size. Furthermore, after adding FITC to the Examples, Comparative Examples 1, 2, and 3 (conjugation), the respective residual amounts in the cornea were observed and measured through FITC fluorescence analysis. At this time, the residual fluorescence of the ocular composition was observed using an in vivo fluorescence spectrometer (IVIS Lumina LT). The Examples, Comparative Examples 1, 2, and 3 were administered to the corneas of mice for approximately 5 minutes and approximately 60 minutes.

[0078] Referring to FIG. 3a, an example of an ocular composition according to one embodiment of the present invention is shown to emit the highest fluorescence intensity of white, and the example is shown to remain continuously adhered within the eyeball regardless of the passage of time. Furthermore, the example is shown to have no difference in fluorescence intensity even after 60 minutes or more have elapsed, which means that the ocular composition according to one embodiment of the present invention can be stably adhered within the eyeball for a long time.

[0079] On the other hand, Comparative Example 1 is shown to emit the lowest fluorescence intensity within the eye, and after 60 minutes, there is almost no fluorescence within the eye.

[0080] Furthermore, Comparative Examples 2 and 3 also appear to emit fluorescence with lower intensity than the Examples, and the fluorescence of the composition is shown to be emitted not only in the eyes but also in the nose of the mouse. That is, in the case of HA with a network structure that does not have a nanoparticle size of 1 to 100 nm, it cannot be stably adhered to the inside of the eyes and flows down into the nasal cavity, which may reduce its effectiveness. In addition, Comparative Examples 2 and 3, like Comparative Example 1, appear to have a decrease in fluorescence intensity over time (60 minutes or more).

[0081] In this regard, referring to FIG. 3b, a quantitative graph for FIG. 3a described above is shown. At this time, since the measurement of fluorescence intensity was calculated by targeting the intraocular region where fluorescence emission is most distinct, the adhesive area affected by the composition and the overall average intensity were not considered.

[0082] The fluorescence intensity for each composition at 5 minutes was found to be the same at 100%, but the fluorescence intensity for each composition at 60 minutes was found to be highest in the embodiment of the present invention.

[0083] In addition, an example of an ocular composition according to one embodiment of the present invention shows that its fluorescence intensity remains the same at 100% even after 60 minutes have elapsed. That is, the ocular composition according to one embodiment of the present invention has higher adhesiveness than conventional compositions containing HA and HA particles of different sizes, and this may mean that the effect can be sustained for a long time without loss.

[0084] Ultimately, according to the results of FIGS. 3a and 3b, the hyaluronic acid particles with a network structure are shown to have heterogeneously enhanced adhesion at a specific size, and accordingly, the size of the HA nanoparticles in the ocular composition according to one embodiment of the present invention may preferably be 1 to 100 nm.

[0085] FIG. 4a is an ex vivo microscopic image of the corneal surface of an ocular composition according to one embodiment of the present invention. FIG. 4a to 4c, to be described later, is a method for verifying ocular adhesion in an ex vivo environment, which was performed using ocular tissue of euthanized rabbits. More specifically, after separating the cornea from the excised ocular tissue, it was fixed with 4% formaldehyde, and a polydimethylsiloxane (PDMS) mold was used to prevent fluid flow. In the comparative examples and examples, a diluent containing DMSO and physiological saline in a 1:2 ratio was used. Furthermore, Comparative Example 1 (Not treated) is a corneal state in which nothing other than the diluent was treated, Comparative Example 2 (control) is a control group comprising a composition containing a human antibody conjugated with FITC, Comparative Example 3 (hyaluronic acid) is a composition containing 100 μg / ml of conventional linear hyaluronic acid, and the example is an ocular composition according to one embodiment of the present invention containing 100 μg / ml HA nanoparticles. Comparative Example 2 (FITC conjugated mouse antibody), Comparative Example 3 (FITC conjugated hyaluronic acid), and the example (FITC conjugated 3D hyaluronic acid) were conjugated with FITC, and the remaining amounts of Comparative Example 2, Comparative Example 3, and the example in the cornea were observed and measured through FITC fluorescence analysis. At this time, as a control, when only FITC was included in the diluent, FITC was bound to the cornea due to chemical binding; therefore, to compensate for this, FITC bound to an antibody was used as a control (Comparative Example 2). These Comparative Examples 1, 2, 3, and the example were administered to the cornea for about 2 hours, and after the administration was completed, microscopic imaging was performed after washing three times with a buffer solution.Furthermore, after analyzing the residual amount on the corneal surface using a fluorescence microscope, the corneal tissue was embedded in an OCT compound and sectioned to a thickness of 10 μm, and the residual amount in the corneal cross-section was analyzed using a fluorescence microscope. More specifically, the residual amount within the corneal cross-section was confirmed using a fluorescence microscope and quantified using ImageJ.

[0086] Referring to Fig. 4a, the green fluorescence (FITC) intensity is shown to be highest in the example, while Comparative Example 1 and Comparative Example 2 show almost no green fluorescence.

[0087] Furthermore, referring to FIG. 4b, an ex vivo microscopic image of a cross-section of the cornea for an ocular composition according to one embodiment of the present invention is shown. Similar to FIG. 4a, the FITC intensity of the example is shown to be the highest, while Comparative Examples 1 and 2 show almost no FITC emission. Furthermore, Comparative Example 3 shows FITC emission, but at a very low intensity.

[0088] In this regard, referring to FIG. 4c, a quantitative graph for FIG. 4b is shown, and the mean fluorescence intensity of the example according to one embodiment of the present invention is about 25, which is about 20 times higher than that of Comparative Example 3, which has a mean fluorescence intensity of 1.

[0089] Accordingly, an ocular composition according to one embodiment of the present invention can have a residual amount in the cornea improved by about 20 times or more compared to a conventional composition containing hyaluronic acid that is not three-dimensionally cross-linked. That is, by including HA nanoparticles, the ocular composition according to one embodiment of the present invention can improve the residual amount in the eye, that is, in the cornea.

[0090] FIG. 5a is an in vivo microscopic image of the surface of a living cornea for an ocular composition according to one embodiment of the present invention. At this time, 5a to 6c, to be described below, is a method for verifying ocular adhesion in an in vivo environment, which was performed on 12 C57BL / 6 mice. More specifically, Comparative Example 2, Comparative Example 3, and Example were administered to the eyes of mice pre-anesthetized with isoflurane, wherein the diluent was the same as that described in Fig. 3a above, Comparative Example 1 (Not treated) was the eye of a mouse that was not treated with anything other than the diluent, Comparative Example 2 (control) was a control group comprising a composition containing a human antibody conjugated with FITC, Comparative Example 3 (hyaluronic acid) was a composition containing 100 μg / ml of conventional linear hyaluronic acid, and Example was an ocular composition according to one embodiment of the present invention containing 100 μg / ml HA nanoparticles, and Comparative Example 2 (FITC conjugated human antibody), Comparative Example 3 (FITC conjugated hyaluronic acid), and Example (FITC conjugated 3D hyaluronic acid) were conjugated with FITC, and the remaining amounts of Comparative Example 2, Comparative Example 3, and Example in the cornea were observed and measured through FITC fluorescence analysis. Fluorescence intensity was measured using IVIS LUMINA III (Perkin Elmer), and fluorescence intensity was measured at various time points ranging from approximately 5 to 120 minutes after administration. Furthermore, for histological evaluation, mice were euthanized, both eyes were excised, and the corneas were isolated from the excised ocular tissues. The residual fluorescence in the isolated corneas was measured using IVIS LUMINA III (Perkin Elmer), and the corneas with the measured residual fluorescence were embedded in an OCT compound, sectioned to a thickness of 10 μm, and the residual fluorescence in the cross-sections of the corneas was measured using a fluorescence microscope.Furthermore, Figures 5a and 5b show the results of measuring the fluorescence residue of the entire ocular tissue of a mouse, and verification and quantification were performed using IVIS.

[0091] Referring to Fig. 5a, Comparative Example 1 is shown to not emit fluorescent light at all.

[0092] Next, Comparative Examples 2 and 3 were found to remain in the mouse's eyeball for up to 5 minutes after administration. However, since Comparative Examples 2 and 3 did not emit fluorescent light at all after 60 minutes, it may mean that Comparative Examples 2 and 3 did not remain in the eyeball after 60 minutes.

[0093] On the other hand, an embodiment containing HA nanoparticles with a network structure according to one embodiment of the present invention is shown to remain in the eye of a mouse for up to 60 minutes.

[0094] In this regard, referring to FIG. 5b, a quantitative graph for FIG. 5a is shown, and the fluorescence intensity at 5 min is similar for Comparative Example 2, Comparative Example 3, and the Example without statistical difference; however, the fluorescence intensity at 60 min is about 6 for Example 3, which is about 6 times higher than that of Comparative Example 1, Comparative Example 2, and Comparative Example 3, which have a fluorescence intensity of about 1. At this time, since Comparative Example 2 and Comparative Example 3 have the same fluorescence intensity as Comparative Example 1, which was not treated at all, it may mean that Comparative Example 2 and Comparative Example 3 were not retained in the living eye and were completely lost.

[0095] Accordingly, an ocular composition according to one embodiment of the present invention can have a corneal retention time improved by about 60 minutes or more compared to a conventional composition containing hyaluronic acid that is not three-dimensionally cross-linked. That is, by including HA nanoparticles, the ocular composition according to one embodiment of the present invention can improve the retention time within the eye, that is, in the cornea.

[0096] In this regard, FIG. 6a is an in vivo microscopic image of the excised corneal surface for an ocular composition according to one embodiment of the present invention. The cornea photographed under the microscope is the cornea excised 60 minutes after Comparative Example 2, Comparative Example 3, and the Example were treated. Furthermore, the residual fluorescence amount in the cornea at the microscope level was confirmed using a fluorescence microscope and quantified using ImageJ.

[0097] Referring to Fig. 6a, after 60 minutes, the fluorescence intensity in the cornea is shown to be highest in the example, and the same fluorescence intensity is maintained in both corneas. Furthermore, Comparative Examples 1, 2, and 3 each have different fluorescence intensities in both corneas and are shown to have lower fluorescence intensities than the example.

[0098] Furthermore, referring to Fig. 6b, an in vivo microscopic image of an excised corneal cross-section for an ocular composition according to one embodiment of the present invention is shown, and in the corneal cross-section, as in Fig. 6a, the FITC intensity of the example is shown to be the highest, and Comparative Examples 1, 2, and 3 show that FITC hardly emits light.

[0099] In this regard, referring to FIG. 6c, a quantitative graph for FIG. 6b is shown, and the relative fluorescence intensity of the embodiment according to one embodiment of the present invention is about 310 or higher, which is about 1.5 times higher than that of Comparative Example 1, Comparative Example 2, and Comparative Example 3, which have a fluorescence intensity of about 210.

[0100] Accordingly, an ocular composition according to one embodiment of the present invention can have a residual amount in the cornea improved by about 1.5 times or more compared to a conventional composition containing hyaluronic acid that is not three-dimensionally cross-linked, under conditions of administration for 60 minutes or more. That is, since the ocular composition according to one embodiment of the present invention contains HA nanoparticles, it can maintain a residual amount in the eye, that is, in the cornea, for a long period of time of 60 minutes or more.

[0101]

[0102] [National R&D projects that supported this invention]

[0103] [Project ID] 1465040345

[0104] [Project No.] HI23C0689000023

[0105] [Ministry Name] Ministry of Health and Welfare

[0106] [Name of Project Management (Specialized) Agency] Korea Health Industry Development Institute

[0107] [Research Project Name] Drug Delivery Therapy Technology Development Project

[0108] [Project Title] Development of Mucosal-Adhesive Hyaluronic Acid Nanocarriers for Effective Delivery of Glaucoma Treatment Drugs

[0109] [Name of Project Performing Organization] Dongguk University Industry-Academic Cooperation Foundation

[0110] [Research Period] April 1, 2023 ~ December 31, 2027

Claims

1. An ocular composition with improved retention force within the eye, It comprises hyaluronic acid (HA) nanoparticles having a size of about 1 to 100 nm, and The above hyaluronic acid nanoparticles are, An ocular composition having a network structure formed by cross-linking between hyaluronic acid molecules.

2. In Paragraph 1, The above residual force is, An ocular composition comprising a retention time and amount in the cornea.

3. In Paragraph 2, The retention time of the above ocular composition in the cornea is An ocular composition that is improved by about 60 minutes or more compared to a composition containing hyaluronic acid.

4. In Paragraph 2, The residual amount of the above ocular composition in the cornea is, An ocular composition that is improved by more than 20 times compared to a composition containing hyaluronic acid.

5. In Paragraph 2, 60 minutes after administration, The residual amount of the above ocular composition in the cornea is, An ocular composition that is improved by about 1.5 times or more compared to a composition containing hyaluronic acid.

6. In Paragraph 1, The size of the above hyaluronic acid nanoparticles is, An ocular composition having a length of about 3 to 70 nm.

7. As an ocular drug delivery system, It comprises hyaluronic acid (HA) nanoparticles having a size of about 1 to 100 nm, and The above hyaluronic acid nanoparticles are, An ocular drug delivery system in which hyaluronic acid molecules are cross-linked to form a network structure.

8. In Paragraph 7, The above ocular drug delivery system is an ocular drug delivery system that induces a pharmacologically active substance to be bound within the hyaluronic acid nanoparticles or retained within the network structure of the nanoparticles so as to adhere to the eye.

9. In Paragraph 8, The above pharmacologically active substance is an ocular drug delivery system that is an active ingredient effective in the prevention or treatment of ophthalmic diseases.

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