Fluorination-modified aflibercept nano-particles and application thereof in eye drops

Abstract

The invention relates to fluorinated modified aflibercept nano-particles and application thereof in eye drops, and belongs to the technical field of nano-drugs. The fluoridation modified aflibercept nano-particles disclosed by the invention are obtained by coupling a perfluoroalkane chain containing a maleimide group and reduced aflibercept. And due to the hydrophobic and oleophobic characteristics of the fluoroalkane chain and the chemical and biological inertness, the modified aflibercept can quickly cross an eye barrier after being dropped into eyes, and the content of VEGF in the eyes is reduced, so that the non-invasive and efficient treatment of CNV is realized. According to the invention, the administration route of aflibercept is innovated, the treatment of choroidal neovascularization can be realized through a non-invasive route in an eye drop administration mode, and the compliance of a patient is greatly improved.

Description

Technical Field

[0001] This invention relates to the field of nanomedicine technology, and in particular to a fluorinated aflibercept nanoparticle and its application in eye drops. Background Technology

[0002] Choroidal neovascularization (CNV) is a leading cause of severe vision loss worldwide. It is estimated that approximately 288 million people globally will be affected by CNV by 2040. CNV is characterized by macular pigment deposition and progressive degeneration of photoreceptor cells and the retinal pigment epithelium (RPE). This leads to an imbalance between limited blood supply to photoreceptor cells in the macular region and high oxygen demand, resulting in RPE hypoxia and upregulation of vascular endothelial growth factor (VEGF).

[0003] The standard treatment for CNV is intravitreal injection of anti-VEGF, but patient compliance is poor, complications are frequent, and recurrence rates are high. Therefore, developing non-invasive and safe treatment strategies is crucial for treating CNV. Ocular drops have become a major treatment for ocular surface diseases due to their convenience and non-invasiveness, but successful delivery of large molecule drugs to the posterior segment of the eye still faces significant challenges from multiple ocular barriers.

[0004] Drug treatment for choroidal neovascularization primarily focuses on inhibiting the growth of new blood vessels under the retina and reducing or eliminating inflammatory responses; there are currently no specific drugs for treatment. Treatment effects can be achieved through photodynamic therapy and intravitreal injection of anti-VEGF drugs.

[0005] Photodynamic therapy (PDT) primarily involves intravenously injecting photosensitizing drugs into patients with idiopathic choroidal neovascularization (ICD). These photosensitizing drugs have a high affinity for choroidal neovascularization, accumulating locally within the neovascularized vessels. Low-intensity laser irradiation of the affected area then activates the photosensitizing drugs, promoting a local reaction that leads to the coagulation and necrosis of the neovascularized vessels. PDT can be applied to various subfoveal choroidal neovascularization diseases, but it also presents certain challenges, such as high treatment costs, high repeat treatment rates, short-term vision loss, and insufficient long-term efficacy stability.

[0006] With the discovery of VEGF's crucial role in neovascularization, intravitreal injection of anti-VEGF drugs such as conbercept, aflibercept, and ranibizumab has become the primary clinical treatment for CNV. However, because these drugs are difficult to target retinal lesions with sustained concentration, clinical treatment often requires multiple injections over a short period, and recurrence is common. Frequent retinal injections can also cause serious complications such as ocular inflammation, subconjunctival hemorrhage, lens damage, and retinal detachment. Furthermore, while topical ocular drops are convenient and inexpensive, drug delivery to the posterior segment of the eye still requires overcoming multiple ocular barriers. Frequent retinal injections can also lead to serious complications such as ocular inflammation, subconjunctival hemorrhage, lens damage, and retinal detachment. Summary of the Invention

[0007] Existing technologies struggle to overcome multiple intraocular barriers and efficiently deliver aflibercept to lesions in the posterior segment of the eye. This invention provides fluorinated aflibercept nanoparticles and their application in eye drops. By coupling reduced aflibercept with a perfluoroalkane chain containing a maleimide group, the modified aflibercept can rapidly cross the ocular barrier after administration, reducing intraocular VEGF levels due to the hydrophobic and oleophobic properties of the fluoroalkane chain and its chemical and biological inertness. This achieves non-invasive and highly effective treatment of CNV.

[0008] This invention is achieved through the following technical solution:

[0009] The first objective of this invention is to provide fluorinated aflibercept nanoparticles, which are obtained by coupling a perfluoroalkane chain containing a maleimide group with reduced aflibercept.

[0010] Furthermore, the fluorinated aflibercept nanoparticles have a particle size of 150 nm-300 nm.

[0011] Furthermore, the molar ratio of the perfluoroalkane chain containing the maleimide group to the reduced aflibercept is 1:1 to 1:4.

[0012] Furthermore, the perfluoroalkane chain containing maleimide groups is obtained by mixing and purifying an active ester solution containing maleimide groups with a perfluoroalkylamine solution.

[0013] Further, the active ester containing the maleimide group is 5-maleimide valerate-active ester, 6-(maleimide)hexanoate succinimide ester, maleimide-diethylene glycol-succinimide ester, or 4-(N-maleimide methyl)cyclohexane-1-carboxylic acid succinimide ester.

[0014] Further, the perfluoroalkylamine is 2,2,3,3,3-pentafluoropropylamine, 1H,1H-perfluoropentanamine, 1H,1H-perfluoroheptylamine, or 1H,1H-perfluorononamine.

[0015] Furthermore, the concentration of the active ester solution containing maleimide groups is 1-10 mg / mL;

[0016] And / or, the concentration of the perfluoroalkylamine solution is 1-10 mg / mL.

[0017] Furthermore, the reduced aflibercept is obtained by mixing and stirring a reducing agent with aflibercept in a buffer solution and then filtering.

[0018] Furthermore, the reducing agent is dithiothreitol, tris(2-carboxyethyl)phosphine, or 2-mercaptoethylamine.

[0019] Furthermore, the buffer solution is a PBS buffer containing boric acid, a HEPES buffer containing boric acid, or a carbonate buffer containing boric acid.

[0020] A second objective of this invention is to provide the application of the fluorinated aflibercept nanoparticles in the preparation of eye drops.

[0021] The technical solution of the present invention has the following advantages compared with the prior art:

[0022] In clinical treatment, intervention and inhibition of neovascularization are the main methods to slow the progression of CNV. The posterior segment of the eye is the primary site of neovascularization-related eye diseases, characterized by irreplaceable tissue and irreversible degeneration. Existing treatments include intravitreal injection of anti-VEGF monoclonal antibodies (such as aflibercept and conbercept), which reduces elevated VEGF levels at the site of neovascularization, inhibiting further vascular expansion and temporarily restoring retinal thickness and function. However, this therapy requires long-term intravitreal injections, easily leading to drug resistance and relapse. Another approach is photodynamic therapy, which involves intravenous injection of photosensitizing drugs. Under laser irradiation, the photosensitizing drugs generate reactive oxygen species, destroying neovascularization and slowing the rate of vision loss. However, this method of destroying blood vessels also has significant side effects, potentially causing secondary subretinal hemorrhage, retinal pigment epithelium tears, and choroidal ischemia, leading to further visual deterioration. Therefore, changing the administration method and strategy, and developing a safe and efficient drug delivery system to address the shortcomings of traditional treatments is imperative.

[0023] This invention provides fluorinated aflibercept nanoparticles and their application in eye drops. The invention designs a fluorinated aflibercept delivery system and prepares fluorinated 13F / AFL for the treatment of CNV. 13F / AFL can efficiently cross multiple ocular barriers (tear film barrier, corneal barrier, conjunctival barrier, and scleral barrier) to reach the posterior segment of the eye. It reduces ocular VEGF levels, inhibits angiogenesis in a non-invasive manner, reduces laser spot area, and achieves CNV treatment.

[0024] This invention innovates the administration route of aflibercept, allowing for non-invasive treatment of choroidal neovascularization via eye drops, thus greatly improving patient compliance. Attached Figure Description

[0025] To make the content of this invention easier to understand, the invention will be further described in detail below with reference to specific embodiments and accompanying drawings, wherein:

[0026] Figure 1 The particle size and PDI diagram of the nanoparticles in Comparative Example 1 of the present invention are shown.

[0027] Figure 2 This is an experimental diagram showing the results of the eye drops' penetration of the tear film barrier in vitro in Test Example 1 of this invention;

[0028] Figure 3 This is an experimental diagram showing the results of the eye drops' penetration of the corneal barrier in vitro in Test Example 1 of this invention;

[0029] Figure 4 This is an experimental diagram showing the results of the eye drops' penetration of the conjunctival barrier in vitro in Test Example 1 of this invention;

[0030] Figure 5 This is an experimental diagram showing the results of the eye drops' penetration of the scleral barrier in vitro in Test Example 1 of this invention;

[0031] Figure 6 This is a graph showing the ELISA results of the eye drops on RPE cells in Test Example 2 of this invention;

[0032] Figure 7 The pharmacokinetics of the eye drops in mouse eyes in Test Example 3 of this invention (n = 4);

[0033] Figure 8 The distribution of the eye drops in the mouse cornea in Test Example 3 of this invention (n = 4);

[0034] Figure 9 The distribution of the eye drops in the lens of mice in Test Example 3 of this invention (n = 4);

[0035] Figure 10The distribution of the eye drops in the mouse retina in Test Example 3 of this invention (n = 4);

[0036] Figure 11 This is the drug distribution of the eye drops in the mouse choroid in Test Example 3 of the present invention (n = 4).

[0037] Figure 12 The figure shows the experimental results of VEGF expression level in the RPE-choroid-sclera complex of mice in test example 3 of this invention (n = 6).

[0038] Figure 13 This is a representative FFA image obtained 14 days after topical eye drops were applied to mice with choroidal neovascularization in Test Example 3 of this invention; circles indicate the location of vascular leakage; circles point to the CNV location;

[0039] Figure 14 This is a representative OCT image obtained 14 days after topical eye drops were applied to mice with choroidal neovascularization in Test Example 3 of this invention; the arrow points to the CNV location. Detailed Implementation

[0040] The present invention will be further described below with reference to the accompanying drawings and specific embodiments, so that those skilled in the art can better understand and implement the present invention. However, the embodiments described are not intended to limit the present invention.

[0041] Unless otherwise specified, the experimental methods used in the following examples are conventional methods, and the materials and reagents used are commercially available.

[0042] 5-Maleimide valerate-active ester solution (Maclean, M992657), 1H,1H-perfluoroheptylamine (Adamas, 56519A), aflibercept (Meilun Biotechnology, MB2806).

[0043] Example

[0044] A solution of 5-maleimide valerate-active ester (10 mg / mL, dissolved in DMSO, 1.1 mol) was added to a solution of 1H,1H-perfluoroheptylamine (10 mg / mL, dissolved in DMSO, 1 mol). The mixture was stirred at room temperature for 12 hours. The crude product was purified by silica gel column chromatography (HEX / EA = 1 / 1, v / v, as eluent) to obtain the fluorinated monomer PFA-Mal containing the maleimide group. Dithiothreitol (10 mM) was added to aflibercept solution (5 mg / mL, dissolved in PBS containing 50 mM boric acid). The mixture was stirred at 37 °C for 30 minutes. After filtration through a desalting column (G-25, PBS containing 1 mM DTPA), the reduced aflibercept was obtained. Subsequently, PFA-Mal (5 mg / mL dissolved in DMSO, 1.1 equivolate / SH group) was added to the reduced aflibercept solution (100 μg / mL), stirred overnight at room temperature, ultrafiltered (MWCO = 10 kDa), and lyophilized to obtain fluorinated aflibercept 13F / AFL.

[0045] FITC solution (1 mg / mL in DMSO, 5 µL) was added to aflibercept solution (100 µg / mL dissolved in 0.1 M NaHCO3 solution, 1 mL). The mixture was stirred overnight at room temperature in the dark. After ultrafiltration purification (MWCO = 10 kDa), the FITC-labeled aflibercept was obtained by lyophilization. FITC AFL).

[0046] FITC-labeled 13F / AFL ( FITC 13F / AFL) was prepared according to the method described in the examples.

[0047] Comparative Example

[0048] A solution of 5-maleimide valerate-active ester (10 mg / mL, dissolved in DMSO, 1.1 mol) was added to a solution of 1-decylamine (10 mg / mL, dissolved in DMSO, 1 mol). The mixture was stirred at room temperature for 12 hours. The product was purified by silica gel column chromatography (HEX / EA = 1 / 1, v / v, as eluent) to obtain the fluorine-free monomer DA-Mal containing a maleimide group. Dithiothreitol (10 mM) was added to an aflibercept solution (5 mg / mL, dissolved in PBS containing 50 mM boric acid). The mixture was stirred at 37 °C for 30 minutes. After filtration through a desalting column (G-25, PBS containing 1 mM DTPA), reduced aflibercept was obtained. Subsequently, DA-Mal (5 mg / mL dissolved in DMSO, 1.1 equivolate / SH group) was added to the reduced aflibercept solution (100 μg / mL), stirred overnight at room temperature, ultrafiltered (MWCO = 10 kDa), and lyophilized to obtain non-fluorinated aflibercept 13C / AFL.

[0049] FITC-labeled 13C / AFL ( FITC 13C / AFL) was prepared according to the method described in the example.

[0050] Figure 1 The figures show the particle size and polydispersity index (PDI) of the nanoparticles in the examples and comparative examples. The particle size is approximately 240 nm, and the PDI is approximately 0.2. It can be seen that the present invention can synthesize nanoparticles with controllable size.

[0051] Test Example 1

[0052] A transwell (0.4 μm pore size) was used to construct an in vitro tear film barrier model by sequentially adding 1% w / v PBS solution containing mucin (100 μL) and artificial tears (50 μL) to the upper chamber, and PBS (pH 7.4, 1 mL) to the lower chamber. For measurements, FITC-labeled 13F / AFL and 13C / AFL (20 μg Aflibercept, 50 μL) were added to the upper chamber, respectively. At specific time points, the mixture from the lower chamber (100 μL) was collected and the same volume of fresh PBS was added. The fluorescence intensity (λ) of Aflibercept in the lower chamber mixture was determined by fluorescence spectrophotometry. ex = 488 nm, λ em = 535 nm). The penetration efficiency (%) of the comparative example and the embodiment to the tear film barrier = (cumulative content of Aflibercept in the lower chamber × 100) / content of added Aflibercept.

[0053] HCECs were sized at a concentration of 1.0–1.5 × 10⁻⁶. 5 cells / cm 2The culture medium was inoculated into the upper chamber of a Transwell (0.4 μm pore size) at a specific density, and the medium was changed every 2 days for 23-25 ​​days. The transmembrane resistance of HCECs was measured using a transmembrane resistivity meter. When the transmembrane resistance reached 400 Ω·cm... 2 At a certain time point, HCECs monolayer formation was observed, successfully establishing an in vitro corneal barrier model. During the assay, the Transwell medium was replaced with fresh serum-free medium. FITC-labeled 13F / AFL and 13C / AFL (20 μg Aflibercept, 50 μL) were added to the upper chamber, and incubated at 37 °C. At specific time points, 100 μL of the lower chamber medium was collected and the same volume of fresh serum-free medium was added. The fluorescence intensity (λ) of Aflibercept in the medium was measured by fluorescence spectrophotometry. ex = 488nm, λ em = 535 nm). The permeation efficiency (%) of the comparative example and the embodiment to the HCEC monolayer = (cumulative content of Aflibercept in the lower chamber) × 100 / initial content of Aflibercept added).

[0054] HConEpiCs were set at 1.0-1.5 × 10 5 cells / cm 2 The HConEpiCs were inoculated into the upper chamber of a Transwell substrate at a density of 0.4 μm, with the culture medium changed every 2 days, and cultured for 23-25 ​​days. The transmembrane resistance of the HConEpiCs was measured using a transmembrane resistivity meter. When the transmembrane resistance reached 1000 Ω·cm... 2 At a certain time point, HConEpiCs monolayer formation was observed, successfully constructing an in vitro corneal barrier model. During the assay, the Transwell medium was replaced with fresh serum-free medium. FITC-labeled 13F / AFL and 13C / AFL (20 μg Aflibercept, 50 μL) were added to the upper chamber, and incubated at 37 °C. At specific time points, 100 μL of the lower chamber medium was collected and the same volume of fresh serum-free medium was added. The fluorescence intensity (λ) of Aflibercept in the medium was measured by fluorescence spectrophotometry. ex = 488 nm, λ em = 535 nm). The permeation efficiency (%) of the comparative example and the embodiment of HConEpiC monolayer = (cumulative content of Aflibercept in the lower chamber × 100) / initial content of Aflibercept added.

[0055] Pig eyeballs were obtained from a nearby slaughterhouse. After collection, the periorbital tissue was removed and the sclera was separated, then washed three times with PBS. The sclera was fixed between the supply and receiving chambers of a Franz transdermal diffusion cell (Φ = 9 mm), with its outer surface facing the supply chamber, to construct an in vitro scleral barrier model. For measurement, FITC-labeled 13F / AFL and 13C / AFL (200 μg Aflibercept, 400 μL) were added to the supply chamber, respectively. Simultaneously, the lower receiving chamber was filled with PBS and incubated at 37 °C. At specific time points, the receiving chamber mixture (200 μL) was collected and the same volume of fresh PBS was added. The Aflibercept content (λ) in the receiving chamber mixture was determined by fluorescence spectrophotometry. ex = 488 nm, λ em = 535 nm). The penetration efficiency (%) of the comparative example and the embodiment to the scleral barrier = (cumulative content of Aflibercept in the receiving chamber × 100) / content of Aflibercept added in the supply chamber.

[0056] Figures 2-5 The results show the penetration of the eye drops obtained in the Examples and Comparative Examples on the in vitro tear film barrier, in vitro corneal barrier, in vitro conjunctival barrier, and in vitro scleral barrier, respectively. It can be seen that the nanoparticles of Example 1 have better penetration effects on the in vitro tear film barrier, corneal barrier, conjunctival barrier, and scleral barrier than the nanoparticles of Comparative Example 1, and have more opportunities to cross the ocular barrier and accumulate in the eye.

[0057] Test Example 2

[0058] RPE cells were loaded at 1 × 10 5 Cells were seeded at a density of 10 cells / well in 6-well plates and cultured for 24 hours. Fresh serum-free medium was then added, along with nanoparticles (10 μg Aflibercept / mL) from the comparative and example studies, and incubated at 37 °C for 24 hours. Cells were then collected. The concentration of VEGF protein in the culture medium was determined using an ELISA kit. Results are shown below. Figure 6 After treatment in the example, the VEGF protein content in the RPE cell supernatant decreased by about 50%, approaching that of free aflibercept, indicating that the example has a good ability to inhibit VEGF expression in vitro.

[0059] Test Example 3

[0060] A mouse CNV model was established using laser photocoagulation. C57BL / 6 mice were anesthetized, and tropicamide eye drops were instilled onto the ocular surface to dilate the pupils. Four burn spots (50 μm in diameter) were created around the optic nerve using a laser (wavelength 532 nm, power 475 mW, 0.05 s). Successful penetration of the Bruch's membrane resulted in bubble-like burns, which were considered effective laser spots. To prevent intraocular infection, erythromycin ointment was applied to the ocular surface of the mice postoperatively.

[0061] The pharmacokinetics of FITC-labeled 13F / AFL and 13C / AFL were evaluated by topical eye drops in normal mice. Figure 7 Pharmacokinetics of the eye drops in mouse eyes; Figures 8-11 These are drug distribution diagrams of the eye drops in the mouse cornea, mouse lens, mouse retina, and mouse choroid, respectively; Figures 7-11 It can be seen that, compared with 13C / AFL, 13F / AFL reaches its peak concentration (C0.05) in the mouse eye more quickly. max ), and C max The concentration of 13F / AFL was twice that of 13C / AFL, indicating that fluorinated aflibercept can promote drug retention in the eye. The drug concentrations of FITC-labeled 13F / AFL and 13C / AFL in the cornea, lens, retina, and choroid of mice after topical instillation were measured to further evaluate the distribution of 13F / AFL in the eye. The overall concentration in all tissues of the eye of mice treated with 13F / AFL was significantly higher than that of mice treated with 13C / AFL. Furthermore, the drug concentration of 13F / AFL in the posterior segment of the retina and choroid was three times that of 13C / AFL, mainly attributed to the rapid and efficient crossing of the ocular barrier by fluorinated aflibercept, leading to accumulation in the posterior segment of the eye.

[0062] All C57BL / 6 mice were randomly divided into four groups: From day 7 to 21, mice were given daily topical instillations of PBS eye drops (5 μL), 13F / AFL, and 13C / AFL eye drops (10 μg Aflibercept / eye, 5 μL). The Normal group consisted of normal, untreated mice. On day 22, mice were sacrificed, and eyeballs were collected. Lysis was performed using RIPA lysis buffer (200 μL) containing protease inhibitors (30 min), followed by centrifugation (12000 g, 15 min, 4 ℃), and the supernatant was collected. VEGF concentration was determined using an ELISA kit. Experimental results are shown below. Figure 12After CNV modeling, compared with mice treated with PBS eye drops, mice treated with 13C / AFL eye drops showed a decrease of approximately 26% in VEGF protein levels in the eyes, while mice treated with 13F / AFL eye drops showed a decrease of approximately 50% in VEGF protein levels, with expression levels returning to normal. This indicates that angiogenesis was effectively inhibited. Compared with the comparative eye drops, the eye drops in this example showed a higher ability to inhibit VEGF expression in the mouse choroidal angiogenesis model, thus providing better relief for choroidal angiogenesis.

[0063] like Figure 13 As shown, the intensity and area of ​​CNV leakage were measured using fluorescein sodium (FFA) to assess the treatment effect. After CNV modeling, neovascularization occurs as new blood vessels branch out from existing vessels, cross the Bruch membrane, and reach the choroid and retina, increasing microvascular permeability and causing pathological vascular leakage. Compared with the PBS group, the laser spot and leakage intensity were significantly reduced in the 13F / AFL eye drop treatment group, indicating that neovascularization was inhibited.

[0064] Fundus angiography reflects the lesion in a planar direction, and further OCT is used along the transverse section to display the lesion location and analyze the treatment effect of AF-NPs. For example... Figure 14 As shown, after laser modeling, the subepithelial retinal neuroepithelial mass was spindle-shaped, and the striped RPE layer was irregularly swollen (arrow). After treatment with 13C / AFL and 13F / AFL, the degree of swelling was significantly reduced, and the retinal structure of mice in the 13F / AFL group was basically restored to the normal level.

[0065] Obviously, the above embodiments are merely illustrative examples for clear explanation and are not intended to limit the implementation. Those skilled in the art will recognize that other variations or modifications can be made based on the above description. It is neither necessary nor possible to exhaustively list all possible implementations here. However, obvious variations or modifications derived therefrom are still within the scope of protection of this invention.

Claims

1. A fluorinated modified aflibercept nanoparticle, characterized in that, The fluorinated modified aflibercept nanoparticles are coupled with the reduced aflibercept through a perfluoroalkane chain containing a maleimide group.

2. The aflibercept nanoparticle of claim 1, wherein, The particle size of the fluorinated modified aflibercept nanoparticles is 150 nm-300 nm.

3. The aflibercept nanoparticle of claim 1, wherein, The molar ratio of the perfluoroalkane chain containing a maleimide group to the reduced aflibercept is 1:1-1:

4.

4. The aflibercept nanoparticle of claim 1, wherein, The perfluoroalkane chain containing a maleimide group is obtained by mixing, stirring and purifying a solution of an active ester containing a maleimide group with a solution of a perfluoroalkylamine.

5. The aflibercept nanoparticle of claim 4, wherein the ratio of the amount of the first polymer to the amount of the second polymer is about 1 :

1. The active ester containing a maleimide group is 5-maleimide valeric acid-active ester, 6-(maleimido) hexanoic acid succinimidyl ester, maleimide-diglycol-succinimidyl ester or 4-(N-maleimido methyl) cyclohexane-1-carboxylic acid succinimidyl ester. ​ And / or, the concentration of the solution of the active ester containing a maleimide group is 1-10 mg / mL.

6. The aflibercept nanoparticle of claim 4, wherein, The perfluoroalkylamine is 2,2,3,3,3-pentafluoropropylamine, 1H,1H-perfluoropentylamine, 1H,1H-perfluoroheptylamine or 1H,1H-perfluorononylamine. And / or, the concentration of the solution of the perfluoroalkylamine is 1-10 mg / mL.

7. The aflibercept nanoparticle of claim 1, wherein, The reduced aflibercept is obtained by mixing, stirring and filtering a reducing agent with aflibercept in a buffer solution.

8. The aflibercept nanoparticle of claim 7, wherein, The reducing agent is dithiothreitol, tris(2-carboxyethyl)phosphine or 2-mercaptoethylamine.

9. The aflibercept nanoparticle of claim 7, wherein, The buffer solution is a PBS buffer solution containing boric acid, a HEPES buffer solution containing boric acid or a carbonate buffer solution containing boric acid.

10. Use of the fluorinated modified aflibercept nanoparticles of any one of claims 1-9 in the preparation of eye drops.

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