JPH0331179B2 - - Google Patents

Description

[Detailed description of the invention]

The present invention relates to an antithrombotic agent whose main component is a protein polysaccharide derived from a basidiomycete belonging to the genus Corsicolor, and more particularly to an antithrombotic agent comprising crestin. Krestin has already been provided to society as an antitumor agent, has extremely low toxicity, and can be administered for a long period of time without worrying about disturbance of intestinal flora. Also,
It does not affect mutagenicity or allergic reactions, and therefore does not cause teratogenesis in healthy people.
It is an extremely safe substance with no risk of allergic reactions. In recent years, the frequency of thrombosis has been increasing in Japan due to changes in lifestyle and eating habits, and thrombus formation is a problem as a factor in the progression of pathological conditions, including cerebral infarction, myocardial infarction, and even nephritis. Therefore, countermeasures are required. Antithrombotic therapy includes anticoagulant therapy, thrombolytic therapy, and antiplatelet therapy. Heparin and warfarin are used as anticoagulant drugs, and urokinase is used as a thrombolytic agent, and these drugs are effective against some thromboses. Although these drugs are effective in preventing coagulation and dissolving thrombus in secondary thrombus formation, they are almost ineffective against primary thrombi caused by platelet aggregation. Most arterial thromboses such as myocardial infarction and cerebral thrombosis are primary thrombi caused by platelet aggregation.
In recent years, treatment for this problem has become more important. Antiplatelet therapy agents such as ticlopidine and aspirin are used, but they have side effects such as liver damage and bleeding, making it difficult to administer them safely over a long period of time. The present inventors have discovered that the protein polysaccharide of the present invention has an antithrombotic pharmacological effect in addition to an antitumor effect, leading to the present invention. The protein polysaccharide which is the active ingredient of the antithrombotic agent of the present invention is disclosed in, for example, Japanese Patent Publication No. 46-17149, Japanese Patent Publication No. 51-1987,
Publication No. 36322, Special Publication No. 14274, Special Publication No. 1983
It is a known substance described in Publication No. 14276, Japanese Patent Publication No. 56-39288, etc., and is obtained by culturing basidiomycetes belonging to the genus Corsicolor in hot water of mycelium, broth, or fruiting body. Or an extract with an alkaline aqueous solution, containing about 18 to 38% protein and having a molecular weight of 5,000 to 300,000 (ultracentrifugation measurement). Among the protein polysaccharides of the present invention, C. versicolor mycelium [FERM-P2412
(ATCC20547)] is commercially available under the trade name Krestin (Recent New Drugs Vol. 28, pp. 14-16), as mentioned above.
1977 and 29th edition, pages 96-101, 1978, Pharmaceutical Directory, page 1346, May 1978, 6th edition, published by Yakugyo Jihosha, Medical Drugs Japan Pharmaceutical Collection, 7th edition, page 240, 1983, It is also called PS-K (Published by Yakugyo Jihosha), and some of its properties are as follows. The sugar moiety of the main fraction is β-D-glucan, and the structure of this glucan moiety is 1→3, 1→4 and 1→
It has a branched structure containing 6 bonds, and the amino acids that make up the protein are mostly acidic amino acids such as aspartic acid and glutamic acid, and neutral amino acids such as valine and leucine, and few basic amino acids such as lysine and arginine. Soluble in water, almost insoluble in methanol, pyridine, chloroform, benzene, and hexane. Gradually decomposes from about 120℃. The protein polysaccharide of the present invention is known to be an extremely safe substance for living organisms, having extremely low toxicity and causing almost no side effects. The acute toxicity values of the protein polysaccharide of the present invention are shown in Table 1 below.

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[Table] The acute toxicity values shown in Table 1 above were determined using the following test method. Mice are ICR-JCL strain, 4-5 weeks old, weight 21~
A 24 g sample was used, and the rats were 4- to 5-week-old, weighing 100 to 150 g. The protein polysaccharide of the present invention was administered through four routes: intravenous, subcutaneous, intraperitoneal, and oral. The protein polysaccharide of the present invention was dissolved in physiological saline and administered, and the animals were observed for general symptoms, death, and body weight for 7 days, and after the observation period, they were sacrificed and autopsied. As shown in Table 1, no cases of death were observed in both rats and mice even at the maximum dose that could be administered, and the protein _{polysaccharide} of the present invention is extremely safe for living organisms. Next, the antithrombotic effect of the protein polysaccharide of the present invention will be explained. Platelet aggregation inhibitory effect When collagen under the endothelium enters the bloodstream due to arteriosclerosis or damage to the vascular endothelium, platelet adhesion and subsequent aggregation occur there. Collagen-induced aggregation causes the release of adenosine diphosphate (ADP) and serotonin from platelet dense bodies. ADP has a strong platelet aggregation effect,
Causes irreversible secondary aggregation. The substance of the present invention had an effect of suppressing platelet aggregation (see Example 1). Thromboembolism inhibitory effect When intravenously administered platelet aggregating substance ADP or soluble collagen to mice, platelets aggregate in the bloodstream, clogging capillaries, and ultimately cause pulmonary embolism, resulting in death within 5 minutes. When the active substance of the invention was administered before the injection of platelet aggregation substances, a thromboembolic protective effect was observed (see Example 2). That is, the protein polysaccharide of the present invention has extremely low acute toxicity, is a safe drug, and is useful as an antithrombotic agent since it exhibits platelet aggregation inhibiting action and thromboembolic death inhibiting action. The protein polysaccharide of the present invention can be made into any dosage form when used as an antithrombotic agent. Moreover, administration is also performed by each route. In addition, when used in combination with conventional antithrombotic agents, the effect is expected to be enhanced. In the case of oral administration, the tablets applied thereto;
Granules, powders, capsules, etc. contain binders, encapsulating agents,
It may contain additives such as excipients, lubricants, disintegrants, and wetting agents, and when used as oral liquid preparations, oral solutions, shaken mixtures, suspensions, emulsions, It may be in the form of a syrup or as a dry product which is redissolved before use. Additionally, such liquid formulations may contain any commonly used additives and preservatives. When used for injection, the composition may include stabilizers,
They may also contain additives such as buffers, preservatives, tonicity agents, and are presented in unit-dose ampoules or multi-dose containers. It should be noted that the compositions may be in the form of aqueous solutions, suspensions, solutions, emulsions in oily or aqueous vehicles, wherein the active ingredient is dissolved in a suitable vehicle, e.g., a pyrogen-free, sterile vehicle, before use. It may also be a powder that is redissolved in water. The antithrombotic agent of the present invention can be administered orally or parenterally to humans and animals, but oral administration is preferred. Oral administration includes sublingual administration. Parenteral administration includes injections such as subcutaneous, intramuscular, intravenous, infusion, and the like. The dosage of the antithrombotic agent of the present invention depends on whether it is an animal or a human being, and is influenced by age, individual differences, medical conditions, etc., so in some cases it may be necessary to administer an amount outside the range shown below, but in general, it may be administered to humans. The oral dosage of the active substance of the present invention is 1 kg body weight per day.
10-1000mg, preferably 20-600mg once to three times
Administer in divided doses. Example 1 A platelet aggregation inhibition test was conducted using Krestin. Tetrate solution for blood sedimentation was used as an anticoagulant, and 9 samples of human venous blood were collected from each sample. This was centrifuged at 400×g for 6 minutes, the supernatant was collected, and human PRP (Platelet Rich Plasma) was prepared. The residue after collecting the supernatant was further collected at 700×g for 20
Centrifuge for 1 minute and remove the supernatant from PPP (Platelet Poor).
Plasma). Changes in the transmittance of PRP were measured using an agglomerometer (manufactured by Biodata, PAP-3).
The degree of platelet aggregation was measured using the following method. Arachidonic acid (1.64 mM), collagen (0.26 mg/ml), and ADP (50 μM) were used as flocculants, and Krestin was added to PRP 2 minutes before adding the flocculant. As a result, Krestin showed an inhibitory effect on platelet aggregation against all aggregating agents. The results are expressed as the aggregation inhibition rate relative to the maximum aggregation degree of the control (physiological saline added instead of Krestin), and are shown in Table 2.
Shown below. Example 2 A test for suppressing thromboembolic death was conducted using Crestin. 10 ddY mice per group (male, weight 20-22 g)
Orally administered Krestin 1g/Kg, and 3 hours later
ADP400mg/Kg amount or soluble collagen (Sigma
0.21 ml/animal was administered intravenously, and the mortality rate was measured 10 minutes later. As a control, distilled water was orally administered instead of Krestin. As a result, as shown in Table 3, Krestin showed an effect of preventing thromboembolism death. Example 3 Using a pressure-type automatic filling machine, 330 mg of Crestin was filled into No. 0 hard capsules to produce capsules.

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Claims (1)

[Scope of Claims] 1. A hot water or alkaline aqueous extract of mycelia or fruiting bodies obtained by culturing Basidiomycetes belonging to the genus Corsicolor, containing about 18 to 38% protein and having a molecular weight of 5000. ~300000 (Ultracentrifugation measurement method)
An antithrombotic agent whose active ingredient is a protein polysaccharide. 2. The antithrombotic agent according to claim 1, wherein the protein polysaccharide is obtained from Coriolus versicolor.