JP4228195B2 - Subconjunctival drug delivery system - Google Patents

Description

【００３６】
４．脈絡膜血管新生阻害試験
ベタメサゾン含有マイクロスフェアー注射剤の脈絡膜血管新生阻害効果について、レーザー誘発ラット脈絡膜血管新生モデルを用いて下記の方法で検討を行った。対照として溶媒（０．４％（ｗ／ｖ） ポリソルベート８０／２．６％（ｗ／ｖ）グリセリン水溶液）のみを含有させたマイクロスフェアー注射剤を用い、下記のように操作を行った。
【００３７】
１）ラットに５％（ｗ／ｖ）塩酸ケタミン注射液および２％（ｗ／ｖ）塩酸キシラジン注射液の混合液（７：１）１ｍｌ／ｋｇを筋肉内投与することによりラットを全身麻酔し、０．５％（ｗ／ｖ）トロピカミド／０．５％（ｗ／ｖ）塩酸フェニレフリン点眼液を両眼に点眼して散瞳させた後、クリプトンレーザー光凝固装置により光凝固を行った。光凝固は、焦点を網膜深層に合わせて太い網膜血管を避け、1眼につき８ヶ所散在状に実施した（凝固条件：スポットサイズ１００μｍ、出力１００ｍＷ、凝固時間０．１秒）。光凝固後、眼底撮影を行い、レーザー照射部位を確認した。
【００３８】
２）光凝固直後に３０Ｇ針を装着したマイクロシリンジを用い、ベタメサゾン含有マイクロスフェアー注射剤をラット上部結膜下に一眼当たり５０μｌ投与した。対照群には、溶媒（０．４％（ｗ／ｖ） ポリソルベート８０／２．６％（ｗ／ｖ）グリセリン水溶液）のみを含有させたマイクロスフェアー注射剤を上部結膜下に一眼当たり５０μＬ投与した。
【００３９】
３）光凝固１４および２８日後に１０％（ｗ／ｖ）フルオレセイン水溶液０．１ｍｌを尾静脈から注入して、蛍光眼底撮影を行った。蛍光眼底撮影で蛍光漏出が認められなかったスポットを陰性、蛍光漏出が認められたスポットを陽性と判定した。下記の計算式に従い、レーザー照射８ヶ所のスポットに対する陽性スポット数の割合から新生血管発現率（％）を算出した。なお、軽度過蛍光を示すものは２スポットの発生で１カウントの陽性とした。
【００４０】
【式１】
新生血管発現率（％）＝
（蛍光漏出スポット数／レーザー照射スポット数）×１００
得られた結果は平均値±標準誤差で表示した。統計解析はスチューデントｔ検定を用いた。有意水準は両側５％とした。
【００４１】
ベタメサゾン含有マイクロスフェアーの脈絡膜血管新生阻害効果を表２に示す。光凝固１４日後の対照群の血管新生発現率６０．９±４．４％に対し、ベタメサゾン含有マイクロスフェアー群の血管新生発現率は１２．５±２．４％であり、統計学的に有意な脈絡膜血管新生阻害作用を示した。また、光凝固２８日後においても対照群の血管新生発現率７３．４±６．０％に対し、ベタメサゾン含有マイクロスフェアー群の血管新生発現率は１２．５±２．４％であり、統計学的に有意な脈絡膜血管新生阻害作用を示した。以上の結果より、ベタメサゾン含有マイクロスフェアーは結膜下投与により、投与１４および２８日後においても脈絡膜血管新生阻害作用を示すことが判明した。
【００４２】
【表２】

【００４３】
【発明の効果】
本発明により、結膜下投与による後眼部への優れたドラッグデリバリーシステムを提供することができる。[0001]
BACKGROUND OF THE INVENTION
The present invention relates to a drug delivery system to the posterior eye segment such as the retina, choroid, and optic nerve.
[0002]
[Prior art]
There are many intractable diseases as diseases in the posterior segment of the eye such as the retina, choroid and optic nerve, and the development of an effective treatment method is desired. For eye diseases, it is most common to administer the drug by instillation, but the drug hardly transfers to the posterior segment of the eye such as the retina, choroid and optic nerve. In addition, it is extremely difficult to maintain the drug concentration in the tissue even after migration.
[0003]
Therefore, intravenous injection, oral administration, and vitreous injection have been attempted as methods for administering drugs for posterior ocular diseases. In intravenous injection or oral administration, the amount of drug transferred to the posterior eye region, which is the target site, is extremely small, and an unexpected systemic action (side effect) of the drug may appear strongly.
[0004]
In vitreous injection, since the drug is directly injected into the eye, the amount of drug transferred to the posterior segment is larger than that of intravenous injection or oral administration. Non-patent literature 1 summarizes the posterior segment delivery by vitreous injection. However, vitreous injection is an administration method that requires advanced techniques and involves considerable pain, so the burden on the patient is large and multiple administrations are extremely difficult.
[0005]
In contrast to these administration methods, subconjunctival injection has a relatively simple procedure, causes less damage to the ocular tissue and less burden on the patient than vitreous injection. Although drug transferability to the posterior segment after subconjunctival injection has also been reported (see Non-Patent Document 2), the half-life is very short, It was difficult. Thus, frequent administration is required to maintain the drug concentration in the tissue, but frequent administration increases the burden on the patient.
[0006]
In order to avoid frequent administration, as a method of maintaining the drug concentration in the eye, a method of administering a drug-polymer conjugate intravenously (see Non-Patent Document 3), a microsphere containing a drug, A method of injecting into the vitreous body (see Patent Document 1) is known.
[0007]
[Non-Patent Document 1]
Jounal of ocular pharmacology and therapeutics, (2001) 17/4, 393-401
[0008]
[Non-Patent Document 2]
Invest. Ophthalmol. Visual Sci. 18 (3) 250-255, 1979
[0009]
[Non-Patent Document 3]
Invest. Ophthalmol. Visual Sci. 40 (11), 2690-2696, 1999
[0010]
[Patent Document 1]
JP2000-247871
[0011]
[Problems to be solved by the invention]
As described above, in the conventional technique, it is difficult to maintain the tissue concentration of the drug injected subconjunctivally, and it has been desired to develop a continuous drug delivery system to the posterior eye region by subconjunctival injection.
[0012]
[Means for Solving the Problems]
Thus, as a result of intensive studies, the present inventors have found that if sustained-release fine particles containing a drug are administered under the conjunctiva, they are very useful as a sustained drug delivery system to the posterior segment of the eye.
That is, the invention according to claim 1 is a continuous drug delivery system to the posterior eye part, characterized by administering fine particles containing a drug under the conjunctiva,
The particle diameter of the fine particles is 50 nm to 75 μm,
A drug delivery system in which the microparticles are formed of a biodegradable or biosoluble polymer.
The invention according to claim 2 is an injection containing fine particles containing a drug, which is a subconjunctival injection that enables continuous drug transfer to the posterior eye part,
The particle diameter of the fine particles is 50 nm to 75 μm,
It is an injection in which the fine particles are formed of a biodegradable or biosoluble polymer.
The invention according to claim 3 is the drug delivery system according to claim 1 or the subconjunctival injection according to claim 2, wherein the posterior eye portion is the retina, choroid, optic nerve, vitreous body or crystalline lens.
The invention according to claim 4 is the drug delivery system according to claim 1 or the subconjunctival injection according to claim 2, wherein the drug is a drug for the treatment or prevention of a retina, choroid, optic nerve, vitreous or lens disease. is there.
In the invention according to claim 5, the drug is an anti-inflammatory agent, immunosuppressive agent, antiviral agent, anticancer agent, angiogenesis inhibitor, antithrombotic agent, optic neuroprotective agent, antibacterial agent or antifungal agent. A drug delivery system or the subconjunctival injection according to claim 2.
[0013]
DETAILED DESCRIPTION OF THE INVENTION
The present invention relates to a drug delivery system for the posterior eye segment used for administering fine particles containing a drug under the conjunctiva. The present invention also relates to an injectable preparation containing fine particles containing a drug, which allows transfer of the drug to the posterior eye segment. By administering the drug-containing microparticles subconjunctivally, the transfer to the posterior eye segment is better than that by intravenous injection or oral administration, and there are few side effects on the whole body. Compared to vitreous injection, the procedure is simple and the burden on the patient is small. Furthermore, the drug concentration in the target tissue can be maintained for a long time by containing the drug in the fine particles.
[0014]
In the present invention, the material forming the fine particles is preferably a biodegradable or biosoluble polymer. Specific examples include polylactic acid, lactic acid-glycolic acid copolymer, lactic acid-caprolactone copolymer, polyanhydride, poly Biodegradable polymers such as orthoester, polyepsilon caprolactone, polyacryl cyanoacrylate, polyhydroxyalkanoate, polyphosphoester, polyamino acid, poly α-hydroxy acid; gelatin, collagen, hyaluronic acid, dextran, starch, alginic acid Natural polymers such as sodium, agar, pullulan, albumin, carrageenan, pectin, xanthan gum, gellan gum, casein, chitosan, fibrinogen; methacrylic acid copolymer, polyvinyl alcohol, hydroxypropi Methylcellulose, hydroxypropyl methyl cellulose acetate, hydroxyethyl cellulose, carboxymethyl cellulose, methyl cellulose, polyvinylpyrrolidone, polyethylene glycol, synthetic polymer poly N- alkyl acrylamide.
[0015]
The molecular weight of these polymer substances is not particularly limited, and can be appropriately selected depending on the type of drug contained in the fine particles, the effective therapeutic concentration of the drug, the drug release period, and the like.
[0016]
The particle diameter of the fine particles in the present invention is preferably 50 nm to 150 μm. Fine particles having a particle size of 50 nm or less are difficult to produce, and particles having a particle size of 150 μm or more are not preferable as an injection because the particles are too large. A more preferable particle diameter is 200 nm to 75 μm.
[0017]
The drug delivery system of the present invention is used for the treatment or prevention of diseases of the retina, choroid and optic nerve. Specific examples of the disease include inflammation caused by various causes, viral and bacterial infections, diseases caused by retinal choroidal neovascularization, diseases caused by retinal ischemia, and optic neuropathy caused by glaucoma. More specifically, uveitis, cytomegalovirus retinitis, age-related macular degeneration, diabetic retinopathy, proliferative vitreoretinopathy, retinal detachment, retinitis pigmentosa, central retinal vein occlusion, Examples include central retinal artery occlusion.
[0018]
There is no restriction | limiting in particular about the drug contained in microparticles | fine-particles, The drug suitable for an object disease can be selected. Specifically, steroidal agents such as betamethasone, dexamethasone, triamcinolone, prednisolone, fluorometholone, hydrocortisone, progesterone or their derivatives; anti-inflammatory agents such as bromofenac, diclofenac; TNF-α inhibitors, PDE-IV inhibitors Cytokine inhibitors such as cyclosporine and tacrolimus; antiviral agents such as ganciclovir, acyclovir and interferon β; antibacterial agents such as ofloxacin, clarithromycin and erythromycin; fluorouracil, methotrexate and MMP inhibitors Anti-cancer agents such as: endostatin, VEGF inhibitor, antisense oligonucleotide, PKC inhibitor, adhesion factor inhibitor, angiogenesis inhibitors such as vasostatic steroids; MK-801, timolol, creatine, taurine, BDNF, etc. Neuroprotective agents, neurotrophic factors, carbonic anhydrase inhibitors such as acetazolamide, and thrombolytic agents such as urokinase. The drug-containing fine particles are preferably a matrix type uniformly dispersed in the fine particles, or a capsule type in which a drug is encapsulated with fine particles as a core.
[0019]
The amount of drug contained in the microparticles may be appropriately increased or decreased according to the type of drug, effective therapeutic concentration, drug release period, symptoms, and the like. The content of the drug is 0.01 to 95% by weight of the fine particles, preferably 0.1 to 20% by weight.
[0020]
Fine particles in the present invention can be produced using a known milling method, phase separation method (coacervation method), spray drying method, supercritical fluid method, interface deposition method, interface reaction method, It is not limited to these. More specifically, the in-liquid drying method (J. Control. Release, 2,343-352, (1985)), which is an interfacial deposition method, and the interfacial polymerization method (Int. J. Pharm., 28, 125-132), which is an interfacial reaction method. (1986)), self-emulsifying solvent diffusion method (J. Control. Release, 25, 89-98 (1993)) and the like. From these production methods, an appropriate production method may be selected as appropriate in consideration of the particle size of the fine particles, the kind, properties, and content of the drug to be contained.
[0021]
As a specific production example of the fine particles, a production example of the drug-containing fine particles containing betamethasone which is an anti-inflammatory agent as a drug and using polylactic acid as a material of the fine particles will be described in Examples described later.
[0022]
The effect of the present invention will be described in detail in the section of the reticulochoroid drug concentration measurement test described later. Was measured, and it was found that the drug concentration in the retina choroid was sustained.
[0023]
The microparticles in the drug delivery system of the present invention are administered subconjunctivally. As a subconjunctival administration method, a conventional subconjunctival injection may be used. As described in the section of the prior art, subconjunctival injection has a relatively simple procedure and a low burden on the patient.
[0024]
Furthermore, when the system of the present invention is used, the drug can be efficiently delivered to the posterior eye part such as the retina, choroid or optic nerve, which is the target site, so that the dose of the drug can be reduced and the side effect can be expected to be reduced.
[0025]
Since the microparticles used in the drug delivery system of the present invention are administered subconjunctivally, the dosage form is preferably an injection. An injection can be prepared using a widely used injection formulation technique. For example, commonly used additives such as osmotic pressure adjusting agents such as sodium chloride, buffering agents such as sodium phosphate, surfactants such as polysorbate 80, thickeners such as methylcellulose, and fine particles are added to distilled water for injection. What is necessary is just to prepare a formulation. If a high-pressure syringe without a needle is used, it is possible to administer the microparticles as they are without using an injection.
[0026]
The results of the fine particle production example, preparation preparation example, drug concentration measurement test and choroidal neovascularization inhibition test are shown below.
[0027]
【Example】
1. Method for Producing Drug-Containing Fine Particles An example of producing fine particles that can be used in the drug delivery system of the present invention is shown below.
[0028]
Betamethasone (0.025 g) and polylactic acid (0.25 g) having a weight average molecular weight of 20000 were dissolved in benzyl alcohol (1.5 mL), and the resulting solution was used as a drug / polymer solution. A 2.0% (w / v) aqueous polyvinyl alcohol solution (30 mL) was homogenized with a homogenizer (5000 rpm), and the drug / polymer solution was added dropwise thereto. This mixture was homogenized for 5 minutes after completion of the dropwise addition to prepare an O / W emulsion. Ultrapure water (300 mL) was stirred (300 rpm) using a stirrer, and the prepared O / W emulsion was added dropwise thereto, followed by stirring for 1 hour after the end of plucking. After completion of the stirring, the obtained suspension was centrifuged, and the supernatant was removed. In order to wash the precipitate, ultrapure water (30 mL) was added to disperse the precipitate, and centrifuged again to remove the supernatant. This operation was performed once again. The washed precipitate was sieved to obtain particles having a particle diameter of 50 nm to 75 μm. The obtained particles were freeze-dried to obtain betamethasone-containing microspheres.
[0029]
2. Formulation Preparation Method Betamethasone-containing microsphere powder (442 mg) was dispersed in 4 mL of a solvent (0.4% (w / v) polysorbate 80 / 2.6% (w / v) glycerin aqueous solution), and the resulting dispersion was obtained. The liquid was a betamethasone-containing microsphere injection.
[0030]
3. Intrareticular Drug Concentration Measurement Test Betamethasone-containing microsphere injection was used to measure the concentration of betamethasone in the retina-choroid according to the following method. Concentration measurement was performed as follows using a betamethasone suspension as a control. The concentration of betamethasone in the retina choroid was compared between the microsphere administration group and the suspension administration group. In the betamethasone suspension, betamethasone in a solvent (0.4% (w / v) polysorbate 80 / 2.6% (w / v) glycerin aqueous solution) so that the concentration of betamethasone is 1% (w / v). Suspended.
[0031]
1) Oxybuprocaine hydrochloride (0.5% (w / v)) ophthalmic solution was instilled into both eyes of Japanese white rabbits to anesthetize the ocular surface.
[0032]
2) Using a 27G needle syringe, betamethasone-containing microsphere injection was administered 100 μL per eye under the upper conjunctiva. Since the betamethasone content in the microspheres is about 4.6% (w / v), the dose of betamethasone is about 500 μg. In the control group, a 27G needle syringe was used, and 1% (w / v) betamethasone suspension was administered at 50 μL per eye under the upper conjunctiva.
[0033]
3) The rabbits were sacrificed 2, 7, 14, 21, and 28 days after administration, and after resection of each eyeball, the retina choroid was recovered, and the betamethasone concentration in the retina choroid was measured by high performance liquid chromatography.
[0034]
The results of changes in drug concentration over time are shown in Table 1. As is clear from Table 1, in the betamethasone suspension, the concentration of betamethasone in the retina choroid was about 0.96 μg / g tissue after 7 days, but was below the detection limit after 14 days. On the other hand, in the betamethasone-containing microsphere, the concentration of betamethasone in the retina choroid was about 0.09 μg / g tissue even after 28 days, and the drug concentration in the retina choroid was maintained.
[0035]
[Table 1]

[0036]
4). Choroidal neovascularization inhibition test The choroidal neovascularization inhibitory effect of betamethasone-containing microsphere injection was examined by the following method using a laser-induced rat choroidal neovascularization model. A microsphere injection containing only a solvent (0.4% (w / v) polysorbate 80 / 2.6% (w / v) glycerin aqueous solution) was used as a control, and the operation was performed as follows.
[0037]
1) The rats were anesthetized by intramuscular administration of 1 ml / kg of a mixed solution (7: 1) of 5% (w / v) ketamine hydrochloride injection and 2% (w / v) xylazine hydrochloride injection. Then, 0.5% (w / v) tropicamide / 0.5% (w / v) phenylephrine hydrochloride ophthalmic solution was instilled into both eyes to make mydriatics, and then photocoagulated with a krypton laser photocoagulator. The photocoagulation was carried out at 8 spots per eye while focusing on the deep retina and avoiding thick retinal blood vessels (coagulation conditions: spot size 100 μm, output 100 mW, coagulation time 0.1 seconds). After photocoagulation, fundus photography was performed to confirm the laser irradiation site.
[0038]
2) Using a microsyringe equipped with a 30G needle immediately after photocoagulation, 50 μl of a betamethasone-containing microsphere injection was administered per eye under the rat upper conjunctiva. In the control group, a microsphere injection containing only a solvent (0.4% (w / v) polysorbate 80 / 2.6% (w / v) glycerin aqueous solution) was administered at 50 μL per eye under the upper conjunctiva. did.
[0039]
3) After 14 and 28 days of photocoagulation, 0.1 ml of 10% (w / v) fluorescein aqueous solution was injected from the tail vein, and fluorescence fundus photography was performed. Spots where fluorescence leakage was not observed in fluorescence fundus photography were determined to be negative, and spots where fluorescence leakage was observed were determined to be positive. According to the following calculation formula, the neovascularization rate (%) was calculated from the ratio of the number of positive spots to 8 spots irradiated with laser. In addition, the thing which shows mild hyperfluorescence was made into 1 count positive by generation | occurrence | production of 2 spots.
[0040]
[Formula 1]
Neovascular expression rate (%) =
(Number of fluorescent leakage spots / number of laser irradiation spots) × 100
The obtained results were expressed as an average value ± standard error. Statistical analysis used Student's t test. The significance level was 5% on both sides.
[0041]
Table 2 shows the choroidal neovascularization inhibitory effect of betamethasone-containing microspheres. The angiogenesis rate of the betamethasone-containing microsphere group was 12.5 ± 2.4% compared with the control group angiogenesis rate of 60.9 ± 4.4% after 14 days of photocoagulation. It showed a significant choroidal neovascularization inhibitory effect. In addition, even after 28 days of photocoagulation, the angiogenesis expression rate of the betamethasone-containing microsphere group was 12.5 ± 2.4%, whereas the angiogenesis expression rate of the control group was 73.4 ± 6.0%. It showed a clinically significant inhibitory effect on choroidal neovascularization. From the above results, it was found that betamethasone-containing microspheres exhibited choroidal neovascularization inhibitory effect even 14 and 28 days after administration by subconjunctival administration.
[0042]
[Table 2]

[0043]
【The invention's effect】
According to the present invention, an excellent drug delivery system for the posterior eye segment by subconjunctival administration can be provided.

Claims (5)

A sustained drug delivery system for the posterior segment of the eye, characterized by administering fine particles containing a drug under the conjunctiva ,
The particle diameter of the fine particles is 50 nm to 75 μm,
A drug delivery system in which the fine particles are formed of a biodegradable or biosoluble polymer . An injection containing fine particles containing a drug, and a subconjunctival injection that enables continuous drug transfer to the posterior eye ,
The particle diameter of the fine particles is 50 nm to 75 μm,
An injection wherein the fine particles are formed of a biodegradable or biosoluble polymer . The drug delivery system according to claim 1 or the subconjunctival injection according to claim 2, wherein the posterior eye portion is a retina, choroid, optic nerve, vitreous body or crystalline lens. The drug delivery system according to claim 1 or the subconjunctival injection according to claim 2, wherein the drug is a drug for treating or preventing a retina, choroid, optic nerve, vitreous body or lens disease. The drug delivery system according to claim 1 or claim 2, wherein the drug is an anti-inflammatory agent, immunosuppressive agent, antiviral agent, anticancer agent, angiogenesis inhibitor, antithrombotic agent, optic neuroprotective agent, antibacterial agent or antifungal agent. Subconjunctival injection.