JPH08301761A - Agent for treatment and prevention of intestinal disease - Google Patents

Abstract

PURPOSE: To obtain the subject agent containing an alkaloid originated from a specific plant as an active component, effective for suppressing the inflammation in intestinal diseases, especially enteritis to ameliorate the symptom and, accordingly, useful for the treatment and/or prevention of ulcerative colitis, Crohn's disease, etc. CONSTITUTION: An alkaloid originated from a plant of the genus stephania, family Menispermaceae [preferably cepharanthine of formula (R1 and R4 are each CH3 ; R2 and R3 together form CH2 ) originated from Stephania cepharantha Hayata] is used as an active component of the subject agent. The alkaloid can be produced e.g. by extracting a plant of the genus Stephania with a solvent such as methanol, concentrating the extract, dissolving the concentrated product in an acidic aqueous solution such as dilute hydrochloric acid, alkalifying the solution and collecting the produced precipitate. The daily dose of the agent is preferably 1-300mg in terms of the alkaloid component in the case of oral administration.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to an agent for treating and / or preventing intestinal diseases comprising an alkaloid as an active ingredient. More specifically, the present invention relates to an agent for treating and / or preventing intestinal diseases, which comprises an alkaloid derived from a plant belonging to the genus Stefania of the family Araceae.

[0002]

2. Description of the Related Art Plants belonging to the genus Stefania belonging to the genus Scarabaeaceae, such as Tamatsuki-lahudji and Tokou-dabu-laji, contain plant alkaloids such as cepharanthin, isotetrandrine, berbamine, cicleanine, homoallomoline and cephalanolin. Pharmaceutical preparations containing alkaloids extracted from plants of the genus Stefania, in particular, Tamasaki tsurugi, have anti-allergic effects, immunomodulatory effects,
Has an antiplatelet aggregation effect, and an anticancer drug resistance overcoming effect,
It is widely used clinically as a safe drug with few side effects for leukopenia, alopecia areata, exudative otitis media, viper bite and the like.

[0003]

An object of the present invention is to provide a novel pharmaceutical use of alkaloids derived from plants of the genus Stefania.

[0004]

The present inventors have conducted research to achieve the above object, and as a result, have found that alkaloids derived from plants of the genus Stefania are useful as active ingredients for agents for treating and / or preventing intestinal diseases, particularly enteritis. And completed the present invention.

[0005] The present invention is a therapeutic / prophylactic agent for intestinal diseases, particularly enteritis, containing an alkaloid derived from a plant belonging to the genus Stefania of the family Aphididae as an active ingredient. The alkaloid, which is an active ingredient of the present invention, can be extracted from a plant belonging to the genus Stefania (for example, Tamasaki luffy, Koutou luffy, etc.) according to a conventional method. Preferably, alkaloids derived from Tamasaki diphthium are used. As an active ingredient of the present invention, an extract obtained by concentrating an extract from a plant belonging to the genus Stefania, a precipitate produced when an acidic solution of the extract is made alkaline, an alkaloid-containing fraction further separated therefrom, and a conventional method Crystals obtained by separation and purification can be used. For example, a Stefania plant (a rhizome, a stem, a seed, a leaf, etc. can be used, but not limited to these parts) is extracted with a solvent such as methanol, ethanol, acetone, ethyl acetate, benzene, etc., and the extract is concentrated. The alkaloid fraction can be separated by dissolving the concentrate in an acidic aqueous solution such as dilute hydrochloric acid, dilute sulfuric acid, citric acid aqueous solution, oxalic acid aqueous solution, making the solution alkaline, and collecting the resulting precipitate. The obtained fraction may be further purified by known means such as various types of chromatography and recrystallization.

[0006] Alkaloids derived from plants of the genus Stefania include cepharanthine, isotetrandrine, berbamine.
, Cycleanine, homoaromolyn (homoar
omoline), cepharanoline (cepharanoline), aromorine (aromoline), obamedin (obamegine), norcycleanine (norcycleanine), 2-norcepharanthin (2-n
orcepharanthine), 2-norcepharolin
anoline), 2-norberbamine, secocepharanthine, obabeline
rine), 2-norisotetrandri
ne), oxyacanthine, oxyacanthine, stephibaberine, and bisbenzylisoquinoline alkaloids such as thalrugosine; coclaurine, reticuline, laudanidine, protosinome
nine), benzylisoquinoline alkaloids such as N-methylcoclaurine; FK-300
0, sinomenine, cephamonin
ne), tannagine, cephamuline
line) and other morphinan alkaloids; last avilin (lastourvilline), isocoridine (isocorydine),
Aporphine alkaloids such as corydine;
Proaporphine alkaloids such as stepharine; cepharamine, acnadinine
(aknadinine), and hasbasan alkaloids such as aknadilactam. The therapeutic / prophylactic agent for intestinal diseases of the present invention may contain at least one alkaloid, and may contain a mixture of two or more alkaloids.

The above-mentioned bisbenzylisoquinoline alkaloid is a compound having the following structure.

[0008]

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[0009]

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[0010]

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[0011]

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The benzyl isoquinoline alkaloid is a compound having the following structure.

[0013]

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The therapeutic / prophylactic agent for intestinal diseases of the present invention preferably contains at least one alkaloid selected from cepharanthin, isotetrandrine, berbamine, cicleanine, homoallomoline and cephalanolin, and more preferably contains cepharanthin. The alkaloid may be a pharmaceutically acceptable acid addition salt.
Pharmaceutically acceptable acid addition salts include hydrochloric acid, hydrobromic acid, sulfuric acid, phosphoric acid, addition salts of inorganic acids such as nitric acid, acetic acid,
Succinic, malic, tartaric, citric, maleic,
Addition salts of organic acids such as fumaric acid, methanesulfonic acid, p-toluenesulfonic acid and the like can be mentioned.

[0015] Pharmaceutical preparations containing alkaloids derived from plants of the genus Stefania include Cepharanthi (Cepharanthi), an alkaloid preparation extracted from T. serrata.
n, registered trademark, Kaken Seiyaku Co., Ltd.) is commercially available.

Cepharanthin (registered trademark) is an alkaloid such as cepharanthin, isotetrandrine, berbamine, cicleanine, homoallomoline, cephalanoline, allomoline, obamedin, norcycleanin, 2-norcepharanthin, 2-norcephalanolin, 2-norberbamine, secocepharanthin , Obaverine, 2-norisotetrandrine, oxyacanthin and tallugocin. Among these alkaloids, the major alkaloid components of Cepharanthin® are cepharanthin, isotetrandrine, berbamine, cicleanine, homoallomoline and cephalanolin. In this specification, the term “cepharanthin (registered trademark)” refers to a commercial product of an alkaloid preparation (commercially available from Kaken Seiyaku Co., Ltd.) extracted from Tamasaki diphthida, and “cepharanthin” When described, the structural formula (I) (wherein R ₁ is CH ₃ ,
R ₂ and R ₃ form —CH ₂ —, and R ₄ is CH ₃ ).

The therapeutic / prophylactic agent for intestinal diseases of the present invention may use alkaloid components as they are, but usually, pharmaceutically acceptable additives such as excipients, binders, lubricants, solvents, It is formulated into tablets, powders, granules, capsules, injections, liquids, suppositories and the like by adding diluents, stabilizers, isotonic agents and the like. As excipients, for example, starch, lactose, methylcellulose, crystalline cellulose, synthetic aluminum silicate, etc., as binders, for example, hydroxypropylcellulose, polyvinylpyrrolidone, etc., as lubricants, for example, talc, magnesium stearate, calcium stearate Sodium chloride, glucose, glycerol, mannitol and the like are used as isotonic agents.

The therapeutic / prophylactic agent for intestinal diseases of the present invention is
It can be administered by intravenous, subcutaneous, intraperitoneal or rectal administration. Preferably, it is administered by oral, intravenous or subcutaneous administration. The dosage of the therapeutic / prophylactic agent for intestinal disease of the present invention varies depending on the patient's age, weight, medical condition, and the like.
Usually, in the case of oral administration, 0.1 to 1000 mg, preferably 1 to 300 mg of alkaloid component per day is used.
mg.

The therapeutic / prophylactic agent for intestinal diseases of the present invention has an effect of suppressing inflammation and improving symptoms of intestinal diseases, particularly enteritis, and is useful for treating and / or preventing intestinal diseases. . In the present invention, enteritis means inflammatory bowel disease in the small intestine (including the duodenum, jejunum and ileum) or the large intestine (including the cecum, colon and rectum). Enteritis includes infectious enteritis with obvious causes, ischemic enteritis, radiation enteritis, drug-induced enteritis, irritable bowel syndrome, and other intractable inflammatory bowel diseases whose cause is still unknown. Sexual bowel disease includes ulcerative colitis, Crohn's disease, and intestinal Behcet's disease, as well as enteritis whose cause is not clear and whose disease unit is not yet clear.

[0020]

[Examples and test examples]

Test Example 1 Cytokine Production Suppressing Effect The inhibitory effect of the agent of the present invention [Cepharanthin (registered trademark)] on cytokine production of peripheral blood monocytic-macrophage cells (Mo-Mφ cells) of enteritis patients was measured. Sulfasalazine and prednisolone were used as control drugs. 3 patients with active ulcerative colitis,
3 patients with Crohn's disease and 1 patient with intestinal Behcet's disease
Approximately 30 ml of the heparin-supplemented peripheral blood of each example was collected, and Ficoll-Lonray Perc
oll) to collect monosites (Mo) and macrophages (Mφ). This was added to a Rosewell Park Memorial Institute medium (R) supplemented with 10% fetal calf serum (FCS).
It was adjusted to 1 × 10 ⁴ cells / ml using PMI-1640). A test drug of a predetermined concentration [cepharanthin (registered trademark, Kaken Seiyaku Co., Ltd.): 1 μg / m 2
1, 10 μg / ml, sulfasalazine: 10 ⁻⁴ M, prednisolone: 10 μg / ml], and simultaneously 10 μg / m of lipopolysaccharide (Lipopolysaccharide) as a stimulant.
After addition of 1, the mixture was incubated at 37 ° C. for 3 hours under carbon dioxide gas. Cytokines in the supernatant [tumor necrosis factor α (TNFα), interleukin 6 (IL-
6) and interleukin 8 (IL-8)] were measured by enzyme-linked immunosorbent assay (Enzyme-Linked Immunosorbent Assay). The results are shown in Tables 1 to 3.

[0021]

[Table 1]

[0022]

[Table 2]

[0023]

[Table 3]

As is clear from the above results, the drug of the present invention was obtained from peripheral blood Mo-M of ulcerative colitis patients and Crohn's disease patients.
has a significant inhibitory effect on TNFα production of φ-system cells,
In addition, TNFα production of peripheral blood Mo-Mφ cells of patients with intestinal Behcet's disease had a tendency to suppress, and an inhibitory effect equivalent to or higher than that of sulfasalazine and prednisolone was observed. Furthermore, the agent of the present invention significantly suppresses the production of inflammatory cytokines (IL-6 and IL-8) in peripheral blood Mo-Mφ cells of patients with ulcerative colitis and patients with Crohn's disease, and is equivalent to sulfasalazine and prednisolone. An inhibitory effect was observed. The agent of the present invention suppresses cytokine production of peripheral blood Mo-Mφ cells of enteritis patients, and is effective in suppressing inflammation of enteritis.

Test Example 2 Effect on Mouse Colitis Model TNFα and dietary fatty acids, particularly ω6 unsaturated fatty acids such as linoleic acid, are considered to be important factors in inflammatory bowel disease. Mice were bred on a diet to which safflower oil, which is a fat and oil rich in linoleic acid, was added, and then TNFα was administered to prepare a colitis model. Specifically, it is d with safflower oil 10% added food.
After pre-breeding dy mice for 10 to 60 days, TNFα was transanally administered at 2,500 to 10,000 units / head to prepare a colitis model. The large intestine was collected two days after the administration of TNFα. In the pathological section of the colon, images similar to ulcerative colitis that did not extend to the muscular layer, such as lymph folliculitis and decreased glandular duct structure, were observed. This model was used as a mouse model similar to ulcerative colitis.

[0026] A 10% ddy mouse was fed on a diet containing 10% safflower oil.
After pre-breeding for up to 60 days,
Transdermal administration was performed at 000 units / head, and the colon was collected two days after TNFα administration. In the drug administration group, Cepharanthin (registered trademark) 5 mg
/ Kg / day was administered orally. The pathological section image of the large intestine in each group was observed. In the group to which Cepharanthin (registered trademark) was not administered, lymph folliculitis and a decrease in duct structure were observed. In contrast, in the group to which Cepharanthin (registered trademark) was administered, attenuated lymph folliculitis was observed.

The ddy mice were used in groups A, B, C and D
The animals were divided into groups and fed with a safflower oil-supplemented diet and preliminarily reared for 18 to 20 days. After preliminary breeding, TNF was added to groups C and D.
α was administered transanally at 10,000 units / head. T
The colon was collected 2 days after NFα administration. Group B and Group D were orally administered Cepharanthin (registered trademark) 5 mg / kg / day for three consecutive days before colon collection. Four hours prior to colon collection, 0.1 g / kg of Evans Blue was injected intravenously.
Evans blue colorectal 4 hours after injection was extracted, and the amount of Evans blue leaked was measured as an index of inflammation by an absorption method. Table 4 shows the results.

[0028]

[Table 4]

As a result, the drug of the present invention attenuated lymph folliculitis in a pathological section image of the colon in a mouse model similar to ulcerative colitis, and significantly suppressed the amount of Evans blue leaked, which is an indicator of inflammation. On the other hand, the drug of the present invention is TNF
Physiological levels of Evans blue leakage not due to alpha stimulation were not suppressed.

Test Example 3 Cepharanthin (cepharant) against mouse colitis model
Effect of hine) After pre-breeding ddy mice for 10 to 60 days on a diet containing 10% safflower oil, TNFα was transanally administered at 2,500 to 10,000 units / head, and two days after TNFα administration, Was collected. In the drug administration group, cepharanthine 5 mg / kg / day
The day was administered orally. The pathological section image of the large intestine in each group was observed. In the group to which cepharanthin was not administered, lymph folliculitis and decreased duct structure were observed. On the other hand, in the cepharanthin administration group, attenuated lymph folliculitis was observed. As a result, cepharanthin attenuated lymphoid folliculitis in a pathological section of the large intestine in a mouse model similar to ulcerative colitis.

Test Example 4 Clinical Effect on Patients with Ulcerative Colitis Cepharanthin (registered trademark) 6 m in 7 cases of ulcerative colitis
g to 15 mg / day were orally administered three times a day. There were 4 males and 3 females, ages 24 to 57.
I was old. Of these, sulfasalazine 3 g / day is 3 g
2 months after oral administration for 2 months did not improve
In two cases, improvement was not observed even when 3 g / day of sulfasalazine and 3 g / day of camostat mesylate were administered orally, and three cases were the first cases. The results are shown in Tables 5 and 6.

[0032]

[Table 5]

[0033]

[Table 6]

As a result, the effect of the drug of the present invention was 2
In 5 cases, the efficacy was confirmed. In addition, two cases in which improvement was not observed even after administration of sulfasalazine 3 g / day for 3 months were compared with Cepharanthin (registered trademark) 6
Administration of mg to 15 mg / day for 3 months all showed significant effects. In addition, administration of cepharanthin (registered trademark) 6 mg to 15 mg / day for 0.5 to 4 months in two cases in which improvement was not observed even after administration of 3 g / day of sulfasalazine and 3 g / day of camostat mesylate for 3 months. All were found to be effective. It is clear that the agent of the present invention has a better therapeutic effect than sulfasalazine.

Test Example 5 Clinical Effect on Crohn's Disease Patients Cepharanthin (registered trademark) 15 m
g / day was orally administered three times a day. The case is male 3
For example, one woman was 22 to 43 years old. Of these, two cases showed no improvement even after oral administration of sulfasalazine 3 g / day for 3 months. Improvement was observed even after oral administration of sulfasalazine 3 g / day and camostat mesylate 3 g / day for 3 months. One case was not recognized, and one case was the first case. Table 7 shows the results.

[0036]

[Table 7]

As a result, the effect of the agent of the present invention was markedly 1
In three cases, the efficacy was 3 cases. Furthermore, two cases in which improvement was not observed even after administration of sulfasalazine 3 g / day for 3 months were compared with Cepharanthin (registered trademark) 1
When 5 mg / day was administered for 0.5 to 4 months, the efficacy and efficacy were one by one. 3 g of sulfasalazine
/ Day and 3 g / day of camostat mesylate did not improve even after 3 months administration. In cases where no improvement was observed, administration of Cepharanthin (registered trademark) 15 mg / day for 6 months was effective. It is clear that the agent of the present invention has a better therapeutic effect than sulfasalazine.

Test Example 6 Clinical Effect on Patients with Intestinal Behcet's Disease One patient with intestinal Behcet's disease was orally administered 15 mg / day of Cepharanthin (registered trademark) three times a day. The case was male and was 24 years old. In this case, 3 g of sulfasalazine / day and 3 g of camostat mesylate /
In this case, no improvement was observed even after oral administration for 3 months. Table 8 shows the results.

[0039]

[Table 8]

As a result, the effect of the drug of the present invention was effective.

Test Example 7 Acute toxicity test LD50 (m) in an acute toxicity test using male mice
g / kg) are shown in Table 9.

[0042]

[Table 9]

Formulation Example 1 500 mg of cepharanthin hydrochloride, 3.0 g of lactose, 1.28 g of corn starch, 200 mg of hydroxypropylcellulose and 20 mg of magnesium stearate are mixed well, granulated, and then tabletted to give 100 m / tablet.
g tablets.

Formulation Example 2 500 mg of the alkaloid fraction of Aspergillus vulgaris, 2.5 g of lactose, 1.75 g of potato starch, 240 mg of crystalline cellulose and 10 mg of calcium stearate are mixed well, and the mixture is filled into capsules, and the alkaloid component in one capsule Capsules containing 10 mg.

Formulation Example 3 500 mg of Alkaloid Fraction of Tamasaki Daffodil is dissolved in diluted hydrochloric acid, distilled water for injection is added, and the solution is made isotonic with common salt, and the total volume is made up to 100 ml. This solution was filtered through a 0.2 μm membrane filter, dispensed and sealed in a 10 ml ampule, and then sterilized by heating to obtain an injection.

Production Example Production of Cepharanthin (registered trademark) A methanol extract of the tuberous root of a plant belonging to the genus Stefania genus Tamasakitsurafuji was dissolved in dilute hydrochloric acid, and this solution was made alkaline with sodium hydroxide, and the resulting precipitate was collected by filtration. . The resulting precipitate is washed with a dilute aqueous sodium hydroxide solution, extracted with ether, and the extract is concentrated under reduced pressure to produce the precipitate.

[0047]

EFFECTS OF THE INVENTION The therapeutic / prophylactic agent for intestinal diseases of the present invention has an effect of suppressing inflammation and improving symptoms of intestinal diseases, particularly enteritis. It is useful for treating and / or preventing intestinal diseases such as ulcerative colitis, Crohn's disease, and intestinal Behcet's disease.

Claims (9)

[Claims] 1. A therapeutic or preventive agent for intestinal diseases, comprising as an active ingredient an alkaloid derived from a plant belonging to the genus Stefania of the family Aphididae. 2. A method for treating enteritis comprising an alkaloid derived from a plant belonging to the genus Stefania of the family Aphididae as an active ingredient.
Prophylactic agent. 3. The therapeutic / preventive agent according to claim 2, wherein the plant belonging to the genus Stefania is Tamasaki tsutsuji. 4. Cepharanthin, isotetrandrine,
Berbamine, cicleanine, homoalomoline, cephalanolin, aromolin, obamedin, norcycleanine,
2-norcepharanthin, 2-norcephalanolin, 2-norberbamine, secocepharanthin, obaverin, 2-norisotetrandrine, oxyacanthin, stefiveaverine, tallugocin, cokraurin, reticulin, rowanidin, protosinomenin, N-methylcoclaurin, FK -3000, at least one alkaloid selected from the group consisting of sinomenine, cephamonin, tannadine, cefamulin, lastabavirin, isocollidine, collidine, stephalin, cephalamin, acnadinine and acnadilactam, or a pharmaceutically acceptable acid addition salt thereof is effective. Treatment of enteritis, which is contained as an ingredient
Prophylactic agent. 5. A cepharanthin, an isotetrandrine,
An agent for treating or preventing enteritis, comprising as an active ingredient at least one alkaloid selected from the group consisting of berbamine, cicleanine, homoallomoline and cephalanolin or a pharmaceutically acceptable acid addition salt thereof. 6. A treatment for enteritis comprising cepharanthin or a pharmaceutically acceptable acid addition salt thereof as an active ingredient.
Prophylactic agent. 7. The therapeutic / prophylactic agent according to any one of claims 2 to 6, which is a therapeutic / prophylactic agent for ulcerative colitis. 8. The therapeutic / prophylactic agent according to claim 2, which is a therapeutic / prophylactic agent for Crohn's disease. 9. The therapeutic / prophylactic agent according to any one of claims 2 to 6, which is a therapeutic / prophylactic agent for intestinal Behcet's disease.