Patent Application
20070293556
1. Field of the Invention
The present invention relates to new use of a hydantoin derivative, in particular, (2S,4S)-6-fluoro-2′,5′-dioxospiro [chroman-4,4′-imidazolidine]-2-carboxamide, as a pharmaceutical preparation.
2. Description of the Related Art
The number of patients with diabetes mellitus as a life style-related disease is increasing, and in a survey on diabetes mellitus in 2002 conducted by the Ministry of Health, Labor and Welfare, the number of patients with diabetes mellitus in Japan is estimated to be 7.4 millions. In recent epidemiological study on 913 cases of non-insulin-dependent diabetes mellitus, about 8% (about 600,000 patients) of patients with diabetes mellitus are reported to have maculopathy. It is estimated that as the number of patients with diabetes mellitus is increased, the number of patients with diabetic maculopathy is also increased.
Diabetic maculopathy, together with diabetic retinopathy, is considered important as one of retinal diseases in patients with diabetes mellitus. Diabetic maculopathy is classified into macular edema, ischemic maculopathy, retinal pigment epitheliopathy and macular traction. The object of diabetic retinopathy therapy is to prevent blindness (loss of visual acuity), while the object of diabetic maculopathy therapy is to prevent and ameliorate deterioration of visual acuity. Macula lutea are significantly different in form from retinas so as to attain high central visual acuity (sharpest and high visual acuity), and have a special structure (absent from an inner plexiform layer and an inner nuclear layer) with extremely fewer tissues other than visual cells. Accordingly, clinically problematic deterioration of visual acuity is due to maculopathy. Development of photocoagulation and vitrectomy came to enable blindness attributable to retinopathy to be almost prevented, but is not satisfactory for maculopathy, so therapy different from retinopathy therapy is needed for maculopathy. This is also important in light of treatment of not a few patients having maculopathy only without having retinopathy. Especially, recent increase in pan-photocoagulation for diabetic retinopathy is estimated to worsen macular edema in diabetic maculopathy, to cause further deterioration of visual acuity. Accordingly, the main point of therapy is shifting toward improvement of quality of life (QOL) of patients by maintaining and improving visual acuity.
Macular edema caused by breakage of a blood-retinal barrier in a retinal vascular endothelial cell or a retinal pigment epithelial cell accounts for about 90% of maculopathy and is a major cause for deterioration of visual acuity in maculopathy. This deterioration of visual acuity does not lead to blindness but causes extreme deterioration of visual acuity referred to as social blindness making usual living difficult. On one hand, the average life span becomes increasing longer owing to advancement of medical technology, and thus such deterioration of visual acuity is a serious problem that cannot be neglected in consideration of QOL. Major therapy conducted for preventing or ameliorating deterioration of visual acuity includes photocoagulation, vitrectomy and chemotherapy. Under the present circumstances, photocoagulation and vitrectomy are examined for their effectiveness in clinical study, and effectiveness and safety for macular edema are still not established. There are cases where complications of neovascular glaucoma and worsening edema occur, and thus there is an earnest desire for the advent of effective and safe chemotherapy. In present chemotherapy, steroids and carbonate dehydratase inhibitors with anti-inflammatory action as major efficacy are used in symptomatic therapy, but their effectiveness is not established and their administration for a long time leads to occurrence of side effects, and thus continuous use thereof in chronic diseases such as diabetes mellitus is difficult under the present circumstance.
(2S, 4S)-6-Fluoro-2′,5′-dioxospiro [chroman-4,4′-imidazolidine]-2-carboxamide (hereinafter, referred to as SNK-860) found by the present applicant was developed as a compound which has a strong inhibitory activity on aldose reductase and is highly safe even in administration for a long time, and clinical test thereof as a therapeutic agent for diabetic neuropathy is advancing worldwide at present.
With respect to hydantoin derivatives including SNK-860, use thereof for diabetic neuropathy is described in JP-A 61-200991 (1986), use thereof for diseases in circulatory organs in JP-A 04-173791 (1992), use thereof for various diseases accompanying aging in JP-A 06-135968 (1994), use thereof for simple diabetic retinopathy in JP-A 07-242547 (1995), and use thereof for diabetic keratopathy in JP-A 08-231549 (1996). However, the effectiveness of the hydantoin derivatives for diabetic maculopathy has not been reported.
As described above, establishment of effective and highly safe therapy for diabetic maculopathy is strongly desired in the medical field. Under the present circumstances, the advent of highly safe chemotherapy enabling administration for a long time is strongly desired because of the problem of safety in treatment by ophthalmologic operation. However, even a model for evaluating experimental diabetic maculopathy, which is important for development of such therapeutic agents, is absent at present, and establishment of an experimental model for development of pharmaceutical preparations is an urgent task.
The present invention has been made in consideration of the background described above, and an object of the present invention is to provide a prophylactic or therapeutic agent for diabetic maculopathy, which can be administered for a long time and exhibits efficacy in a mechanism different from that of existing medicines, as well as an experimental animal model which can be used in evaluation of medicines for diabetic maculopathy.
The present inventors should first establish an experimental animal model for diabetic maculopathy. That is, simplicidentata such as rats have no macula lutea, and there is no report on edema at a site outside of a retina such as a visual cell layer, that is, at a site corresponding to macula lutea, and whether its severeness is increased or decreased by diabetes mellitus is not reported either. Accordingly, the present inventors studied its pathologic condition using an animal, and as a result we found that when a diabetic rat was allowed to be in an intraocular ischemic state and then subjected to reperfusion, edema was expressed on a visual cell layer. In this experimental model, it is suggested that an increase in oxidation stress, such as excessive production of free radicals occurring in the eye by ischemia and reperfusion, promotes vascular permeability by breaking an inner blood-retinal barrier (barrier regulating the transfer of a substance from a retinal blood vessel to the outside of the blood vessel) and an outer blood-retinal barrier (barrier regulating the transfer of a substance from a choroid to retina). Accordingly, it is estimated that the edema was expressed by this promotion of vascular permeability in addition to the promotion of retinal vascular permeability by diabetes mellitus. The present model thus expressing edema in the visual cell layer has an onset mechanism very similar to that of macular edema in human diabetic maculopathy and can be said to be a model suitable for evaluation of diabetic maculopathy.
Then, the present inventors examined whether edema was expressed or not in a macula lutea in a diabetic monkey by using an evaluation system established in a rat. As a result, it was confirmed that edema is observed in a macular central fovea participating most in central visual acuity. This can be said to further evidence that the edema-expressing model established in a rat is suitable in evaluation of diabetic maculopathy.
When the present inventors used the above-mentioned experimental model to evaluate SNK-860, it was revealed that SNK-860 is effective for edema in a retinal visual cell layer having a central role in maintaining visual acuity or for edema in a macula lutea (particularly macular central fovea). By further conducting clinical evaluation, it was revealed that the present compound is not only effective for edema in a macula lutea but also exhibits an effect of improving visual acuity. That is, the present invention relates to a prophylactic or therapeutic agent for diabetic maculopathy, which comprises, as an active ingredient, a hydantoin derivative represented by the following general formula, preferably (2S,4S)-6-fluoro-2′,5′-dioxospiro [chroman-4,4′-imidazolidine]-2-carboxamide (SNK-860).
(In the formula, X represents a halogen or a hydrogen atom, R1 and R2 concurrently or differently represent a hydrogen atom or an optionally substituted C1 to C6 alkyl group, or R1 and R2, together with a nitrogen atom bound thereto and optionally another nitrogen atom or an oxygen atom, are combined to form a 5- to 6-membered heterocycle, and the halogen represented by X is preferably fluorine, and the C1 to C6 alkyl group is preferably a methyl group.)
Examples of the diabetic maculopathy include macular edema, and retinal pigment epitheliopathy. Examples of the diabetic macular edema include local macular edema and diffuse macular edema. The prophylactic or therapeutic agent for diabetic maculopathy according to the present invention is preferably in the form of an oral agent.
The present invention also relates to an experimental animal model with diabetic maculopathy, which uses an animal such as simplicidentata or primates other than humans. This model is an animal model with diabetic maculopathy produced by subjecting a diabetic animal to intraocular ischemia/reperfusion to express edema in a retinal visual cell layer or in a macula lutea (particularly in macular central fovea). As the animal with diabetes mellitus, it is possible to use not only animals having diabetes mellitus induced for example by administering a pharmacological agent such as streptozotocin or alloxan into a rat (normal rat) or a monkey (normal monkey), but also animals with hereditary diabetes mellitus.
Further, the present invention encompasses a method of evaluating a pharmacological agent for diabetic maculopathy, which comprises using the model animal described above. That is, the method of the present invention is a method of evaluating the effectiveness of a pharmacological agent on edema, which comprises administering a pharmacological agent to be evaluated into the model animal and measuring the thickness of a retinal visual cell layer or the thickness and/or volume of a macula lutea.
The present invention provides not only a therapeutic agent for diabetic maculopathy, which can be administered for a long time, but also an experimental model animal necessary for searching for a therapeutic agent for diabetic maculopathy.
Hereinafter, the present invention is described in more detail.
Hydantoin derivatives (particularly SNK-860) can be orally administered for example as tablets, capsules, powder, granules, liquid or syrup or parenterally as an injection and suppositories, which were formed by usual pharmaceutical manufacturing techniques. Pharmaceutically acceptable excipients in pharmaceutical manufacturing, for example starch, lactose, refined white sugar, glucose, crystalline cellulose, carboxy cellulose, carboxymethyl cellulose, carboxyethyl cellulose, calcium phosphate, magnesium stearate and gum arabic can be used in the solid preparation, and if necessary a lubricant, a binder, a disintegrating agent, a coating agent, a coloring agent etc. can be incorporated into the solid preparation. In the liquid preparation, a stabilizer, a solubilizer, a suspending agent, an emulsifying agent, a buffer agent, a preservative etc. can be used. The dose varies depending on symptoms, age, administration method, preparation form etc., but preferably the compound described above is administered usually in the range of 1 to 200 mg, preferably 1 to 100 mg, into an adult all at once or in divided portions per day for consecutive days.
In the model animals with diabetic maculopathy according to the present invention, animals with diabetes mellitus by treating normal animals with a pharmacological agent such as streptozotocin (STZ), or alloxan or animals with hereditary diabetes mellitus, can be used as diabetic animals. As the type of the animals, simplicidentata such as rats, nonhuman primates such as monkeys, duplicidentata such as rabbits, and carnivorous animals such as canines can be used. When the simplicidentata, duplicidentata or carnivorous animals that inherently do not have macula lutea are used, edema is expressed in a retinal visual cell layer and the thickness of the retinal visual cell layer can be used in evaluation. In the nonhuman primates, on the other hand, there are usually macula lutea, so edema is expressed in a macula lutea and the thickness and/or volume of the macula lutea is used in evaluation. The thickness etc. of the macula lutea are evaluated preferably in the site of macular central fovea. Intraocular ischemia/reperfusion treatment can be easily carried out by stopping a retinal blood stream by increasing the intraocular pressure and then relieving the intraocular pressure to allow reperfusion. The thickness of the retinal visual cell layer or the macula lutea varies significantly depending on individuals, so a treated eye and untreated eye are set preferably in the same individual by subjecting only one eye to intraocular ischemia/reperfusion treatment. By so doing, relative evaluation of “thickness of a treated eye/thickness of an untreated eye” can be carried out on the basis of the untreated eye in each animal. A pharmacological agent to be examined is administered into the model animal with diabetic maculopathy according to the present invention and then evaluated for the effectiveness of the pharmacological agent for edema as described above, whereby the effectiveness of the pharmacological agent for diabetic maculopathy can be evaluated. The method of administering the pharmacological agent is not particularly limited, and administration of the pharmacological agent is also carried out after intraocular ischemia/reperfusion treatment thereby clarifying the therapeutic effect.
1. Test Method
Diabetes mellitus was induced in male Sprague Dawley rats (8-week-old) weighing about 250 g by injecting streptozotocin (STZ manufactured by Sigma) intravenously into their tail at the dose of 60 mg/kg. One week after the treatment with STZ, serum glucose was measured, and rats with at least 300 mg/dl glucose were then used in the experiment as diabetic rats. The set groups were the following 3 groups, and after 2 weeks after the treatment with STZ, 5% gum arabic solution or SNK-860 solution was orally administered once a day.
After administration for 2 weeks, intraocular ischemia was caused by a treatment described below. After the treatment was finished, the animals were maintained as usual for 2 days, and thereafter, the eyeballs were excised and histologically evaluated. Administration of the pharmacological agent was also conducted for the period (2 days) of reperfusion after ischemic treatment.
Retinal Ischemia by Increasing the Intraocular Pressure
A drip infusion set (Terufusion Drip Infusion Set manufactured by Terumo) was connected to a bottle containing an intraocular perfusion solution (Opeguard MA manufactured by Senjyu Seiyaku), and an extension tube (Terumo) to which a three-way stopcock had been attached was connected thereto. A needle (30 G×½, manufactured by Nippon Becton Dickinton) was fitted to the end of the tube. The bottle containing an intraocular perfusion solution was fixed to a certain height with a stand. The rats were anesthetized by administering sodium pentobarbital (Somunopentyl manufactured by Schering-Plough Animal Health) intraperitoneally in a ratio of 50 mg/kg, and then a mydriatic (Mydrin P manufactured by Santen Pharmaceutical) and a local anesthetic (Benoxyl eye drop 0.4%, Santen Pharmaceutical) were dropped onto the right eye. The anesthetic was additionally administered when necessary. Thereafter, a needle was stuck into an anterior chamber of the rat right eye and the intraocular pressure load was performed by manipulating the three-way stopcock (the intraocular pressure was increased to 130 mmHg or more for 60 minutes). Because the ocular fundus in the Sprague Dawley rat turns from red to white by stopping a retinal blood stream by increasing the intraocular pressure, achievement of retinal ischemia can be easily observed. After the intraocular pressure was increased for the predetermined time, the needle was removed to relieve the intraocular pressure to allow reperfusion, and an antibacterial eye drop (tarivit eye ointment manufactured by Santen Pharmaceutical) was applied onto the right eye.
Histological Evaluation
Two days after the ischemia treatment (two days after the reperfusion), the rat left and right eyeballs were excised under anesthesia with ether. The excised eyeballs were placed in an ice-cold fixing solution (phosphate buffer solution containing 3% glutaraldehyde) and fixed therein for 2 days. Thereafter, the eyeballs were washed for 1 day with a phosphate buffer solution. The eyeballs were embedded in a usual manner into paraffin to prepare a transverse section containing a bundle of optic nerves. The section was stained with hematoxylin-eosin. The histological evaluation was conducted by each 2 visual fields in left and right side (4 visual fields/rat) in the vicinity of the bundle of optic nerves, from an optical microscope to an image analyzer (IPAP-WIN, Sumika Techno Service). In each of the resulting retinal images, the thickness of the visual cell layer was measured. The degree of edema was expressed in percentage by dividing the thickness of the visual cell layer of the ischemic/re-perfused eyeball (right eye) by the thickness of the visual cell layer of the untreated eyeball (left eye) in the same individual. As an indicator of retinal cell functions, nuclei in the inner retinal layer (ganglion cell layer) were counted, and the degree of loss of nuclei was evaluated relative to the ratio of the number of nuclei occurring per unit area.
2. Results and Discussion
The effect of SNK-860 on edema is shown in
Next, loss of nuclei from ganglion cells is described. As a result of examination of the degree of loss of nuclei from cells, no loss of nuclei was recognized in 5 rats in the normal control group. In the diabetic control group, evident loss of nuclei occurred in 3 of 7 rats, among which 2 rats showed loss of 50% or more nuclei. In the diabetic group given 32 mg/kg SNK-860, loss of nuclei was not observed in all 4 rats.
These results reveal that SNK-860 inhibits edema formation under diabetes in a visual cell layer and also prevents disturbances in functions of retinal cells.
1. Test Method
The test was carried out in accordance with Efficacy Pharmacological Test Example 1. The set groups were the following 4 groups, and from 2 weeks after the treatment with STZ, 5% gum arabic solution or SNK-860 solution was orally administered once a day.
Retinal ischemia produced by increasing the intraocular pressure was in accordance with Efficacy Pharmaceutical Test Example 1. Histological evaluation was also in accordance with Efficacy Pharmaceutical Test Example 1.
2. Results and Discussion
The effect of SNK-860 on edema is shown in
1. Method
Diabetes mellitus was induced in male monkeys (Macaca fascicularis) (3-year-old) weighing about 2.1 to 2.4 kg by intravenously injecting STZ into their foreleg vein at the dose of 80 mg/kg. Two days after the treatment with STZ, blood glucose level was measured, and monkeys with at least 200 mg/dl glucose were then used in the experiment as diabetic monkeys. Insulin was administered subcutaneously once or twice per day into monkeys showing a blood glucose level of 300 mg/dl. The set groups were the following 3 groups, and from 2 weeks after the treatment with STZ, 5% gum arabic solution or SNK-860 solution was orally administered once a day.
After administration for 2 weeks, intraocular ischemic treatment was carried out as described below, and after the treatment was finished, the animals were maintained as usual for 7 days. Before the ischemic treatment and 7 days after treatment, the thickness and volume of the macular central fovea (in the diameter range of 1 mm from the center of the macula lutea) were measured by an OCT scanner (Stratus OCT, Carl Zeiss). Administration of the pharmacological agent was also conducted for the period (7 days) of reperfusion after the ischemic treatment.
Retinal ischemia produced by increasing intraocular pressure was in accordance with Efficacy Pharmaceutical Test Example 1. However, the size of a needle used was 25 G×½ (Terumo). After a mydriatic (Mydrin P manufactured by Santen Pharmaceutical) was dropped onto the right eye, the monkey was anesthetized by intramuscularly administering ketaral (Sankyo Life Tech). Subsequently, a local anesthetic (Benoxyl eye drop 0.4%) was dropped onto the eye, and the monkey was prevented from blinking with an eyelid speculum. Anesthesia with ketaral was additionally carried out when necessary.
The thickness and volume of the macular central fovea were measured in the following manner. After a mydriatic (Mydrin P) was dropped onto the right eye of the monkey to dilate the pupil of the eye sufficiently, the monkey was anesthetized by intramuscularly administering Ketaral. Thereafter, the monkey was allowed to sit on a monkey chair and the head was fixed. The inside of the eye was observed with an OCT scanner to identify the macula lutea, followed by scanning. On the basis of the resulting cross-sectional macular image, the thickness and volume of the macular central fovea were analyzed.
**P < 0.01 vs. “value before ischemia”.
2. Results
The results are shown in Table 1 and
1. Method
Among patients with diabetic maculopathy, 10 patients with diabetic macular edema having a retinal thickening or a hard exudates in a posterior pole of the retina were subjects. SNK-860 was orally administered in a dose of 30 mg (2 tablets each containing 15 mg SNK-860) once a day before breakfast for 8 weeks. During this test period, simultaneous use of eparlestat, intravitreal injection and sub-Tenon injection of an adrenal cortical hormone, and photocoagulation and vitrectomy were prohibited. Basic therapy of diabetes mellitus was carried out so as to give good blood glucose control throughout the test period.
Evaluation was carried out in terms of the thickness of the macular central fovea (in the diameter range of 1 mm from the center of macula lutea) and the thickness at the center of the central fovea measured by optical coherence tomography (OCT, Carl Zeiss), as well as corrected visual acuity (Log MAR).
Log MAR (Log Minimum Angle of Resolution) is one kind of logarithmic visual acuity, which is visual acuity expressed in logarithmic minimum angle of resolution. Decimal visual acuity 1.0 used frequently in Japan is 0.0 in terms of Log MAR, and decimal visual acuity 0.1 is 1.0 in Log MAR. Alog MAR visual acuity of 0.1 to 0.5 corresponds to a decimal visual acuity of 0.8 to 0.32.
2. Results
12 eyes in the 10 cases were evaluated. When the test was initiated, the thickness of the macular central fovea (in the diameter range of 1 mm) was 324.3 μm on average, and the thickness at the center of the central fovea was 323.1 μm on average. After 8 weeks, these were reduced 300.4 μm and 298.7 μm respectively (Table 2,
Out of the 12 eyes, 2-stage development with respect to corrected visual acuity was recognized in 3 eyes, 1-stage development in 2 eyes, and deterioration in only 1 eye (Table 3). The corrected visual acuity (Log MAR) on average was improved from 0.30 to 0.24 (
Diabetic maculopathy according to conventional findings is a gradually worsening disease that is considered hardly ameliorated. In contrast, the results of the present examples indicate that SNK-860 is effective for diabetic maculopathy. In respect of safety, no particularly problematic side effect was recognized.
| Number | Date | Country | Kind |
|---|---|---|---|
| 2004-022547 | Jan 2004 | JP | national |
| Filing Document | Filing Date | Country | Kind | 371c Date |
|---|---|---|---|---|
| PCT/JP05/01187 | 1/28/2005 | WO | 5/10/2007 |
