JPH07258092A - Agent for treatment of diseases causing abnormal accumulation of collagen - Google Patents

Abstract

PURPOSE:To prepare a drug or a quasi-drug effective for specifically suppressing abnormal synthesis of collagen without exerting undesirable influence on the synthesis of proteins necessary for the restoration of various organs. CONSTITUTION:This agent for the prevention or treatment of diseases or morbid state caused by the abnormal synthesis of collagen contains at least one kind of substance selected from saikosapopin a, saikosaponin b1, saikosaponin b2, saikosaponin c and saikosaponin d as an active component.

Description

Detailed Description of the Invention

[0001]

TECHNICAL FIELD The present invention relates to a preventive or therapeutic agent for diseases or pathological conditions caused by abnormal synthesis of collagen containing saikosaponins. Such a therapeutic or prophylactic agent can be used not only as a drug but also as a quasi drug.

[0002]

BACKGROUND OF THE INVENTION Psychosaponins are known as components derived from roots of Saiko or Budhirum falcatum L., and the roots are Chinese herbs for the purpose of lower fever, sedation and detoxification. Has been blended since ancient times.
Furthermore, saikosaponins have the effects of healing skin wounds, preventing and improving rough skin, and preventing aging to prevent sagging and loss of skin, and are incorporated in pharmaceuticals, cosmetics and the like.

Further, the extracted saikosaponin was separated into saikosaponin a, saikosaponin b ₁ , saikosaponin b ₂ , saikosaponin c and saikosaponin d, and the above effects were enhanced by using each component alone. A technique is also proposed (Japanese Patent Publication No. 4-36137).

As described above, saikosaponins show various biological actions, and are also interested in their use for the development of new applications.

[0005]

As described above, the inventors of the present invention have conducted research to find out new useful effects of saikosaponin and a method of utilizing the saikosaponin, and as a result, the saikosaponins significantly inhibit fibroblast collagen synthesis promotion. The present inventors have found that they have effects and can prevent or treat diseases caused by excessive synthesis of collagen, and completed the present invention.

Therefore, according to the present invention, at least one selected from the group consisting of saikosaponin a, saikosaponin b ₁ , saikosaponin b ₂ , saikosaponin c and saikosaponin d is contained as an active ingredient. Provided is a therapeutic or prophylactic agent for a disease or pathological condition caused by the characterized abnormal synthesis of collagen.

The saikosaponins used in the present invention are
The extract may be extracted and isolated from Sasaiko by a method known per se, or may be extracted and isolated from a tissue culture of a plant piece of Mishima Psychophyllum. The thus-isolated saikosaponins, specifically, saikosaponin a, saikosaponin b ₁ , saikosaponin b ₂ , saikosaponin c, and saikosaponin c, are used alone or in combination of two or more thereof. can do.

Such saikosaponins are not limited by their mechanism of action, but for example, in a fibroblast culture system derived from normal human skin, Transforming
Growth factor-β1 (hereinafter referred to as TGF-β1) specifically inhibits the promotion of collagen synthesis in fibroblasts, and more preferably, it does not affect other protein synthesis. Can be advantageously used for the prevention or treatment of diseases or pathological conditions caused by abnormal synthesis of Such specificity is also revealed by shortening the healing days as compared with the model of delayed wound healing using an aged rat.

It has been already known that certain steroid drugs have an inhibitory effect on the proliferation ability of fibroblasts and collagen synthesis. For example triamcinolone (tr
iamcinolone) suppresses abnormal proliferation of fibroblasts in a human keloid-derived fibroblast culture system and exhibits an inhibitory effect on collagen oversynthesis. However, triamcinolone is known to have an inhibitory effect not only on the proliferation of human normal skin-derived fibroblasts but also on collagen synthesis, and further delays the number of healing days in a wound model, and is a therapeutic agent for hypertrophic scars. Is not preferable.

In comparison with such a conventional technique,
The usefulness of the present invention will be obvious. By the way, to explain it more specifically, the above-mentioned saikosaponins alone do not act on the collagen synthesis of normal human skin fibroblasts, and as described above, they do not act like fibroblasts stimulated by TGF-β1 or the like. It has not been described in the prior art document that a specific inhibitory effect on the promotion of collagen synthesis in cells in which collagen synthesis is promoted is exhibited. On the other hand, TGF-β1
It is also unknown at all that it does not show an inhibitory effect on the promotion of protein synthesis in cells such as fibroblasts stimulated by the cells which also promote the synthesis of proteins other than collagen. It is extremely surprising that the psychosaponins exhibit the above-mentioned selectivity of action.

Thus, the preparation of the present invention containing a saikosaponin as an active ingredient can be advantageously used for preventing or treating a disease or a pathological condition caused by abnormal synthesis of collagen. Diseases caused by abnormal synthesis of collagen include, but are not limited to, various types of fibrosis. Specific examples of these fibrosis include diseases associated with fibrosis of various organs, such as scleroderma, liver fibrosis, pulmonary fibrosis, and renal fibrosis. For the etiology, diagnosis, treatment, etc. of fibrosis, refer to the latest information, Vol. 42, No. 10, page 2052 et seq.

It will be understood that the formulation of the present invention can be used not only as a drug but also as a quasi drug, considering the onset site of the disease and its characteristics.

The preparation of the present invention can be prepared as a pharmaceutical composition or a quasi drug composition, and can be provided in various dosage forms depending on the respective usage. Examples of such dosage forms include powders, granules, fine granules, dry syrups, tablets, capsules and other oral preparations, ointments, plasters, liniments, lotions (solutions, suspensions, emulsions), creams. Agent,
Examples thereof include external preparations such as patches, injections, suppositories and the like.

These pharmaceutical compositions or quasi-drug compositions contain, in addition to the above-mentioned essential ingredients, appropriate pharmaceutically acceptable excipients depending on the above-mentioned dosage forms as long as the effects of the present invention are not impaired. Drugs or quasi drugs such as excipients, disintegrants, binders, lubricants, aqueous components, powder components, oils, surfactants, moisturizers, thickeners, preservatives, antioxidants, fragrances, colorants, etc. Additives for goods can be blended.

For example, the powder is prepared by adding suitable excipients, lubricants and the like to the above-mentioned saikosaponins if necessary and mixing them well. Tablets are prepared by adding suitable excipients, disintegrating agents, binders, lubricants, etc. to saikosaponins as necessary and compressing them according to a conventional method. If necessary, the tablets may be coated to give film-coated tablets, sugar-coated tablets, enteric-coated tablets and the like.

The capsules are prepared by, for example, adding appropriate excipients, lubricants and the like to the saponins, if necessary, and mixing them well.
Fill into a suitable capsule to make a capsule. Further, it may be post-filled as granules or fine particles by a conventional method.

When the composition of the present invention is used for the actual prevention or treatment of each of the above diseases, among the active ingredients, saikosaponin a, saikosaponin b ₁ , saikosaponin b ₂ , saikosaponin c, saikosaponin d At least one of
The dose of the species is appropriately determined depending on the body weight, age, sex, degree of disease, etc. of the patient, but is generally within the range of 0.1 mg to 10000 mg per adult per day for oral administration, and per adult per day for transdermal administration. In the range of 0.01 mg to 1000 mg, and 0.1% per day for adults for other routes of administration.
It is administered in the range of mg to 1000 mg.

[0018]

EXAMPLES Next, the present invention will be described in detail with reference to examples, but it goes without saying that the technical scope of the present invention is not limited to these examples.

Example 1: Collagen of normal human skin fibroblasts
Study of action on gene synthesis 42,000 normal human dermal fibroblasts at 7th passage /
Seed each well into a 6-well plate (Falcon 3046) and
After culturing in DMEM medium containing 0% fetal bovine serum for 4 days to reach confluency, the cells were subjected to an experiment.

After culturing for 24 hours in a DMEM medium (containing 10% defatted fetal bovine serum) containing each test substance and 10 μg / ml L-ascorbic acid, 50 mM β-aminopropionitrile (10 μl / well) and L -[2,3
-3H] Proline (20 μCi / well) was added, and the cells were further cultured for 24 hours.

After completion of the culture, the collagenous and non-collagen proteins in the culture medium and cell homogenate were measured by the following procedure by the bacterial collagenase digestion method. After removing free radioactive proline by dialysis, N
Collagenase (2000 unit / ml) was allowed to act at 37 ° C. for 90 minutes in the presence of ethylmaleimide and calcium chloride. After completion of the reaction, 10% trichloroacetic acid-0.
5% tannic acid was added and the mixture was centrifuged, and the radioactivity in the supernatant was measured as collagenous protein. In addition, it is 0 for precipitation.
After adding 5% sodium dodecyl sulfate-5 mM dithiothreitol and boiling and heating, the radioactivity of the non-collagen protein was measured.

The results are shown in FIGS. 1 and 2. 10 μg /
ml saikosaponin b ₁ (indicated as SSb _{1 in the} figure)
Showed no significant effect on the production of collagenous and non-collagenous proteins. On the other hand, 3 ng /
TGF-β1 in ml showed a significant stimulatory effect on the production of collagenous and non-collagenous proteins. Therefore, when both of them are combined, the production of collagenous protein is suppressed to the same extent as that of Psychosaponin b ₁ alone, whereas the production of non-collagenous protein is suppressed by TGF-β 1 alone. Maintained about the same as. The inhibitory action on the production of collagenous protein by saikosaponin is not due to the non-specific reaction due to the inhibition of the production of all proteins, but is highly specific, and the inhibitory action against this production is TGF-β.
Since it was observed only in the case where the production of collagen was promoted by 1., it is considered that it is very useful when Saikosaponin is used as a therapeutic agent for a disease associated with abnormal accumulation of collagen. Example 2: Examination of effect on wound healing 1 10-week-old or 70-week-old Wistar rat was used.
Each group consisted of 4 animals, and their backs were shaved and used for the test. Rats are anesthetized with pentobarpital and 2 cm to the left and right from the midline
An incision of about 2.5 cm was made along the midline of the skin. Immediately thereafter, the incision was sutured by Mitsuheru. The creams of the following Examples and Comparative Examples were applied once a day in 100 μl aliquots for 8 days. Remove the suture needle on the 9th day and cross section 1 cm
A skin section was prepared so that The cutting tension of this section was measured using a Toyo Sokki Co., Ltd. Tensilon UTM-4.

Example 3, Example 4 and Comparative Example 1 Creams were prepared by selecting the compositions of these examples as shown in Table 1 below.

Table 1 --------------------------------------------- Composition (each composition is shown by weight%) Example 3 Example 4 Comparative Example 1 ------------------ (Oil Phase) Psychosaponin b ₁ 0.1 1.0-cetostearyl alcohol 3.5 3.5 3.5 squalane 38.0 38.0 38.0 beeswax 3.0 3.0 3.0 reduced lanolin 5.0 5.0 5.0 stearic acid monoglyceride 2.0 2.0 2.0 polyoxyethylene (20) sorbitan 2.0 2.0 2.0 monopalmitic acid ester ethylparaben 0.3 0.3 0.3 ( Aqueous phase) 1,3-butylene glycol acetate 7.0 7.0 7.0 Glycerin 5.0 5.0 5.0 Adjusted to 100% by weight with ion-exchanged water Residual residue Residual residue --------------------------------- −−−−−−−−−−−−−− Heat the prepared oil phase Maintained at 75 ° C and added to the aqueous phase warmed to 75 ° C with stirring. Next, the mixture is uniformly emulsified with a homomixer and then rapidly cooled with stirring to obtain a cream.

Results The average tension of the 70 week old rat treated with Comparative Example 1 was 37.
The average tension of the 70-week-old rats treated with Example 3 or Example 4 was 5 g / cm, respectively, while that of 0.5 g / cm.
The values were significantly higher at 49.9 g / cm and 524.3 g / cm. Further, the average tension of the 10-week-old rat treated in Comparative Example 1 was 533.6 g / cm. From the above results, it is found that the rat's wound-healing ability decreases with age, and that saikosaponins have a decreased old rat (7).
It was found that the wound healing ability at 0 weeks of age was restored to the level of a young rat (10 weeks old). Generally, it is said that collagen synthesis is indispensable in the process of wound healing. However, in the present example, since the saikosaponins promoted the wound healing rather than delaying the wound healing, the saikosaponins did not inhibit the collagen synthesis necessary for the living body, and the collagen promoted abnormally. It is considered to suppress only synthesis. That is,
Although healing of the surgical site or wound is promoted, it is considered that hyperplasia such as scarring does not occur, which suggests that psychosaponins are extremely useful as healing-improving agents for surgical site or wound. Is clear.

Examples of the formulations of the present invention will be given below by way of examples, but these formulations can also obtain the same effects as the above.

Example 5: Composition of emulsion lotion composition wt% Saikosaponin b ₂ 0.01 Ethanol 3.0 Cetyl alcohol 1.0 Beeswax 0.5 Vaseline 2.0 Squalane 6.0 Dimethylpolysiloxane 2.0 Glycerin 4.0 1,3-Butylene glycol 4.0 Polyoxyethylene (10) 1.0 Monooleic acid ester Glycerol monostearic acid ester 1.0 Ethylparaben 0.3 Quinceseed extract (5% extract) 20.0 Ion-exchanged water Psychosaponin b ₂ is added to the remaining prepared ethanol and dissolved to form an alcohol phase. On the other hand, an aqueous phase is prepared by adding glycerin and 1,3-butylene glycol to ion-exchanged water and heating and dissolving at 70 ° C. Other components except the quince seed extract are mixed and heated to 70 ° C to prepare.
This is added to the previous aqueous phase and pre-emulsification is performed. After adding an alcohol phase and a quince seed extract to this emulsion, and homogenizing the emulsion particles with a stirrer and a homomixer, degassing and filtering,
Cool down.

Example 6 Solution Lotion Composition Weight% Psychosaponin a 0.5 Ethanol 10.0 Glycerin 4.0 1,3-Butylene glycol 4.0 Oleyl alcohol 0.1 Polyoxyethylene (20) sorbitan 0.5 Monolaurate Polyoxyethylene (15) lauryl 0.5 Alcohol ether Ethyl paraben 0.3 Ion-exchanged water Glycerin and 1,3-butylene glycol are dissolved in the residual prepared ion-exchanged water at room temperature to prepare an aqueous phase. Other components such as saikosaponin a are dissolved in ethanol. This is mixed and solubilized in the above aqueous phase.

Example 7 Ointment Composition Weight% Psychosaponin c 10.0 Glycerin monoethyl ether diacetate 3.0 Vaseline 40.0 Stearyl alcohol 15.0 Mokurou 20.0 Polyoxyethylene (10) 0.25 monooleate glycerin monostearate 0.25 ethylparaben 0.3 ion Exchanged water To the residual- prepared 70 ° C. ion-exchanged water, other components mixed and dissolved at 70 ° C. were added, uniformly emulsified with a homomixer, cooled after emulsification to obtain an ointment.

[0030] Example 8: powders prepared saikosaponins d500 lactose G500g (total volume 100
0 g) is mixed well to prepare a powder.

Example 9: Tablet preparation 200 g saikosaponin b ₁ , lactose g 360 g, 6% H
Lactose (360 g), valerycyo starch (65 g), and talc stearate (15 g) (total amount: 1000 g) are mixed well and tabletted to produce 1000 tablets.

Example 10: Capsule preparation 150 g of saikosaponin b _2, 150 g of lactose, 165 g of valleycyo starch and 15 g of talc stearate.
(Total amount of 1000 g) is mixed well and filled into hard capsules to produce 1000 capsules.

[0033]

INDUSTRIAL APPLICABILITY According to the present invention, collagen is contained by containing at least one selected from the group consisting of saikosaponin a, saikosaponin b ₁ , saikosaponin b ₂ , saikosaponin c and saikosaponin as an active ingredient. It becomes possible to provide a pharmaceutical composition or a quasi drug composition which can be extremely utilized as a preventive or therapeutic agent for a disease caused by abnormal synthesis of a.

[Brief description of drawings]

FIG. 1 is a graph showing the action of psychosaponin b _{1 on} collagen synthesis in human fibroblasts.

FIG. 2 is a graph showing the effect of saikosaponin b _{1 on} the non-collagen protein synthesis of human fibroblasts.

─────────────────────────────────────────────────── ─── Continuation of the front page (51) Int.Cl. ⁶ Identification code Internal reference number FI technical display area A61K 35/78 ADA N 8217-4C

Claims (2)

[Claims] 1. Abnormal synthesis of collagen comprising at least one selected from the group consisting of saikosaponin a, saikosaponin b ₁ , saikosaponin b ₂ , saikosaponin c and saikosaponin d as an active ingredient. A preventive or therapeutic agent for a disease or pathological condition caused by. 2. The preventive or therapeutic agent according to claim 1, wherein the disease or pathological condition is scleroderma, hepatic fibrosis and pulmonary fibrosis, and hypertrophic scar and keloid at the wound site.