JP2001261556A - Smooth muscle anomalous shrinkage inhibitor - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a calcium ion-independent smooth muscle anomalous shrinkage inhibitor that is uneffective on the calcium ion-independent physiological smooth muscle shrinkage, an injection composition for treating the cardiovascular diseases caused by smooth muscle anomalous shrinkage, a prophylactic and/or therapeutic agent for black out and narrowed visual field, reduction of the side-effect of the chemical-inducing orthostatic hypotension and Rho- kinase signal transduction system inhibitor. SOLUTION: These calcium ion-independent smooth muscle anomalous shrinkage inhibitor that is uneffective on the calcium ion-independent physiological smooth muscle shrinkage, an injection composition for treating the cardiovascular diseases caused by smooth muscle anomalous shrinkage, a prophylactic and/or therapeutic agent for black out and narrowed visual field, reduction of the side-effect of the chemical-inducing orthostatic hypotension and Rho- kinase signal transduction system inhibitor include, as an active ingredient, at least one selected from the group consisting icosapentaenoic acid, salts and esters thereof.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

[0001] The present invention relates to a novel inhibitor of abnormal contraction of smooth muscle, specifically, at least one active ingredient selected from the group consisting of icosapentic acid (hereinafter abbreviated as EPA), salts and esters thereof. As an agent for suppressing abnormal contraction of smooth muscle. Further, the present invention provides a method for treating a circulatory system disease caused by abnormal contraction of vascular smooth muscle, comprising as an active ingredient at least one selected from the group consisting of EPA, pharmaceutically acceptable salts and esters thereof. And compositions for injection. In particular, the present invention relates to an injectable composition wherein the cardiovascular disease is cerebral vasospasm or coronary vasospasm. Further, the present invention provides at least one selected from the group consisting of EPA, salts and esters thereof.
And Rho-kinase signaling system inhibitors. The EPA in the present invention is all-cis-5,8,
11,14,17-icosapentic acid (all-cis-
5,8,11,14,17-icoscaptain
oiacid).

[0002]

2. Description of the Related Art Unlike striated muscles such as skeletal muscle and myocardium, smooth muscle contraction involves a contraction mechanism that does not involve a change in membrane potential due to vasoconstrictor substances such as angiotensin II and epinephrine, in addition to contraction involving a change in membrane potential. Exists. These vasoconstrictors (hereinafter abbreviated as Ca ²⁺⁾ cytoplasmic calcium ion increasing concentrations, the Ca ²⁺ binds to calmodulin, Ca ²⁺ - myosin light by activated myosin light chain kinase by calmodulin It is believed that chain phosphorylation causes contraction. Such smooth muscle contraction occurs at an intensity depending on the degree of increase in cytoplasmic Ca ²⁺ concentration. In other words, Ca ²⁺ -dependent smooth muscle contraction
It is considered a normal contraction.

On the other hand, contraction by some vasoconstrictors increases the sensitivity of Ca ²⁺ via a low molecular weight GTP-binding protein (hereinafter abbreviated as G protein) of the cell membrane, and increases the concentration of cytoplasmic Ca ^2+. It is known that the above contraction or contraction independent of Ca ²⁺ concentration is caused. That is, Ca ²⁺ -independent smooth muscle contraction, which is considered to be an abnormal pathological contraction. For example, sphingosylphosphorylcholine (hereinafter abbreviated as SPC) causes rabbit rectal smooth muscle to be a pathway independent of the pathway mediated by inositol triphosphate-Ca ²⁺ -calmodulin (at least Mito).
gen-activated protein kinase (including MAPK))
Is known to shrink via (American
Journal of Physiology (American)
JournalofPhysiology), 32 volumes,
G370-377, 1995).

[0004] Rho kinase whose activity is enhanced by RhoA, one of G proteins, is a type of protein kinase. Rho kinase directly phosphorylates myosin light chain to increase Ca ²⁺ sensitivity or to contract smooth muscle in a Ca ²⁺ -independent manner, while inhibiting its activity by phosphorylating myosin phosphatase. It is thought that it suppresses dephosphorylation of myosin light chain. This Ca ²⁺ -independent Rho kinase signal transduction system is a so-called pathological abnormal contraction, and is considered to play an important role in the pathological condition of vascular smooth muscle abnormalities such as hypertension (anesthesia) , Vol. 47, No. 5,
530-540, 1998).

[0005] Commercially available antihypertensive agents represented by calcium antagonists that suppress Ca ²⁺ channels include the aforementioned C ²⁺
In order to suppress a2 ⁺ -dependent normal smooth muscle contraction, during treatment of hypertension, for example, normal reflex contraction is inhibited, and side effects of orthostatic hypotension appear. In addition, hypertension in which a calcium antagonist is ineffective is known. Therefore, a substance that suppresses only Ca ²⁺ -independent abnormal smooth muscle contraction without affecting physiological and normal smooth muscle contraction is desired.
However, at present, there is no satisfactory inhibitor that selectively suppresses abnormal contraction of smooth muscle.

[0006] Among cardiovascular diseases caused by pathological vascular smooth muscle contraction, particularly those requiring immediate treatment, such as cerebral vasospasm or coronary vasospasm,
Various trials have been conducted, including calcium antagonists, but no sufficient therapeutic effect has been obtained. A therapeutic agent that does not affect physiological and normal smooth muscle contraction at normal sites and suppresses only abnormal smooth muscle contraction at the site of a contracted diseased body is desired, but at present it is completely unsatisfactory. is there.

[0007] Further, narrowing of the visual field and dark vision are considered to be symptoms caused by inadequate blood flow to ocular tissues due to abnormal contraction of ocular blood vessels caused by various causes. At present, some calcium antagonists and nitroglycerin are used as therapeutic agents, but these also have insufficient therapeutic effects, and there are concerns about side effects such as a decrease in systemic blood pressure.

[0008] In terms of therapeutic results, the situation is as described above. Looking at the compounds that suppress these abnormal contractions by focusing on the basics, almost no substance that specifically inhibits RhoA or Rho kinase is known. In fact, fasudilic acid (hereinafter abbreviated as HA-1077) or Y-27632 and its derivatives have only been reported as inhibitors of Rho kinase (Nature, 389, October 30, 2009). No., 990
-994, 1997).

On the other hand, EPA is known to have a serum lipid-lowering effect, a platelet aggregation-suppressing effect, and the like, and is commercially available in Japan as a therapeutic agent for obstructive arteriosclerosis and hyperlipidemia. However, there is no knowledge about the inhibitory action of EPA on abnormal smooth muscle contraction, the effect on circulatory diseases caused by abnormal contraction of vascular smooth muscle, or the inhibitory action of Rho kinase signaling system.

[0010]

The existing inhibitors of smooth muscle contraction, such as calcium antagonists, completely inhibit Ca ²⁺ -dependent physiological and normal smooth muscle contraction in a dose-dependent manner in vitro. However, the effect of suppressing abnormal smooth muscle contraction is not always sufficient, and there are many cases of ineffective cases in conditions such as spasm and hypertension. Select the invention, for example, SPC specifically suppressed by the Ca ²⁺ -independent smooth muscle abnormality contraction elicited by, that does not affect the Ca ²⁺ dependent physiological normal contraction such as by depolarization To provide a highly inhibitory agent.

The present invention also relates to a cardiovascular disease caused by abnormal contraction of vascular smooth muscle, particularly a disease requiring immediate treatment, such as cerebral vasospasm or coronary vasospasm, pulmonary vasospasm,
It is an object of the present invention to provide an injectable composition for treating mesenteric artery spasm or finger vasospasm. In particular, by suppressing only the abnormal contraction of Ca ²⁺ -independent smooth muscle at the diseased site without affecting the physiological and normal smooth muscle contraction at the normal site, a rapid effect and high side effects are suppressed. It is an object of the present invention to provide an injectable composition for therapeutic use. Another object of the present invention is to provide a highly specific inhibitor of the Rho kinase signaling system, which is considered to be a cause of abnormal smooth muscle contraction.

[0012]

Means for Solving the Problems The present inventors have conducted intensive studies on abnormal contraction of smooth muscle, and found that EPA
Finding the fact that it had never been known before, it has an inhibitory effect on the Rho kinase signaling system, has an inhibitory effect on abnormal smooth muscle contraction, and has no effect on smooth muscle normal contraction, The present invention has been completed.

[0013]

DESCRIPTION OF THE PREFERRED EMBODIMENTS The present invention will be described below in detail.
A first aspect of the present invention is an agent for suppressing abnormal contraction of smooth muscle, comprising as an active ingredient at least one selected from the group consisting of EPA, a salt thereof and an ester thereof. Smooth muscles are all included regardless of organs or sites, and particularly include vascular smooth muscle, digestive smooth muscle, and respiratory smooth muscle. Although the EPA content ratio and the aspect in the total fatty acids are not particularly limited as long as the inhibitory effect is obtained,
The EPA content ratio in all fatty acids is at least 80% by mass or more, preferably 90% by mass or more, and more preferably 95% by mass or more.
% Or more can be used, and 97% or more is more preferable. EPA is preferably EPA, EPA ethyl ester (hereinafter abbreviated as EPA-E), EPA sodium salt (hereinafter abbreviated as EPA-Na), or EPA glyceride (hereinafter abbreviated as EPA-G).
A-Na is more preferred.

[0014] A second aspect of the present invention relates to icosapentic acid,
An injectable composition for treating a circulatory disease caused by abnormal contraction of vascular smooth muscle, comprising as an active ingredient at least one selected from the group consisting of pharmaceutically acceptable salts and esters thereof. is there. The circulatory system disease is a disease in which immediate action is particularly desired, and examples thereof include cerebral vasospasm or coronary vasospasm, pulmonary vasospasm, mesenteric artery vasospasm, and finger vasospasm. The EPA content ratio in all fatty acids and the embodiment are not particularly limited as long as a therapeutic effect is obtained, but the EPA content ratio in all fatty acids is at least 80% by mass or more, preferably 90% by mass or more, and more preferably 95% by mass or more. What can be used, and what is 97 mass% or more is more preferable. Embodiments of EPA are EPA-E, E
PA-Na or EPA-G is preferred, and EPA-N
a is more preferred. As an administration method, rapid administration is preferably performed once or several times intravenously or intraarterially, but continuous administration over several hours to several days using an infusion pump or an infusion pump or the like is also possible. .

[0015] A third aspect of the present invention is an inhibitor of a Rho-kinase signaling system containing at least one selected from the group consisting of EPA, salts and esters thereof. Preferred are inhibitors of the Rho kinase signaling system in smooth muscle contraction, and also inhibitors of the SPC-Rho kinase signaling system. It is also an apparent Rho kinase inhibitor. As long as the inhibitory effect is obtained, the EPA content ratio and the mode in the total fatty acids are not limited.
PA and EPA-Na are preferred, and EPA-Na is more preferred.

[0016] A fourth aspect of the present invention relates to icosapentic acid,
Prevention of dark vision and visual field narrowing characterized by containing as an active ingredient at least one selected from the group consisting of pharmaceutically acceptable salts and esters thereof.
Or a therapeutic agent. The EPA content ratio in all fatty acids and the embodiment are not particularly limited as long as a therapeutic effect is obtained, but the EPA content ratio in all fatty acids is at least 80% by mass or more, preferably 90% by mass or more, and more preferably 95% by mass or more. What can be used, and what is 97 mass% or more is more preferable. As for the mode of EPA, EPA-E, EPA-Na and EPA-G are preferable. The administration route is not particularly limited as long as a therapeutic effect can be obtained, but oral administration, intravenous administration or eye drops are preferable. Further, a composition for oral administration containing EPA-E as an active ingredient or an injection composition containing EPA-Na as an active ingredient is preferred.

According to a fifth aspect of the present invention, there is provided a drug-induced orthostatic hypotension comprising, as an active ingredient, at least one selected from the group consisting of EPA, pharmaceutically acceptable salts and esters thereof. And a drug for treating hypertension, particularly a calcium antagonist, an angiotensin II converting enzyme inhibitor or a nitro compound. The EPA content ratio in all fatty acids and the embodiment are not particularly limited as long as a therapeutic effect is obtained, but the EPA content ratio in all fatty acids is at least 80% by mass or more, preferably 90% by mass or more, and more preferably 95% by mass or more. What can be used, and what is 97 mass% or more is more preferable. As for the mode of EPA, EPA-E, EPA-Na and EPA-G are preferable, and the administration route is not particularly limited as long as a reducing effect is obtained. Oral administration or intravenous administration is preferred, and oral administration is more preferred. Further, a composition for oral administration containing EPA-E as an active ingredient or an injection composition containing EPA-Na as an active ingredient is preferred. EP
A mode in which A is administered simultaneously with a drug that induces side effects and a mode in which A is separately administered are provided. If administered separately,
EPA can be administered before or after the agent that induces side effects.

As used herein, the term “abnormal smooth muscle contraction” refers to smooth muscle contraction that occurs independently of Ca ²⁺ concentration.
It is a smooth muscle contraction caused by a chemical substance represented by PC. The action mechanism, a smooth muscle contraction occurring Ca ²⁺ independent occurring Ca ²⁺ sensitivity of smooth muscle via the G protein by vasoconstrictor is enhanced. Specifically, RhoA of smooth muscle cells, Ca ²⁺ through Rho kinase signal transduction system
Smooth muscle contraction that occurs independently. Abnormal smooth muscle contraction
For example, it can be confirmed as a Ca ²⁺ -independent contraction caused by SPC in an isolated blood vessel specimen or a skinned blood vessel specimen and suppressed by a Rho kinase inhibitor (Y-27632, HA-1077). At this time, intracellular Ca ²⁺ such as fura-2
Monitoring the Ca ²⁺ concentration using a concentration indicator is a contraction characterized by the absence of an increase in Ca ²⁺ concentration associated with contraction. It does not include Ca ²⁺ -dependent physiological smooth muscle normal contraction in which contraction occurs at an intensity depending on the degree of increase in cytoplasmic Ca ²⁺ concentration due to membrane depolarization.

[0019] In pathological conditions, abnormal smooth muscle contraction seems to play an important role in diseases in which abnormal vascular smooth muscle tone is considered to be involved, such as hypertension, cerebral vasospasm or coronary vasospasm. That is, when abnormal smooth muscle contraction occurs in vascular smooth muscle, it may cause circulatory diseases such as coronary artery spasm and cerebral artery spasm. When abnormal smooth muscle contraction occurs in ocular blood vessels, it may cause dark vision or narrowing of the visual field.

When abnormal contraction of smooth muscle occurs in digestive smooth muscle, it may cause digestive disorders such as diarrhea or abdominal pain due to esophageal spasm, gastric spasm, hyperintestinal motility, spastic ileus or abdominal angina. Can be. When abnormal smooth muscle contraction occurs in the respiratory smooth muscle, it may cause respiratory diseases such as dyspnea or bronchial asthma.

Furthermore, the inhibitor of abnormal smooth muscle contraction of the present invention is useful as an inhibitor in an experiment on abnormal smooth muscle contraction using an isolated blood vessel sample or a skinned blood vessel sample. Further, the inhibitor of abnormal contraction of smooth muscle of the present invention can be applied to treatment, research, and the like for diseases caused by abnormal contraction of smooth muscle.

Injectable compositions for the treatment of circulatory diseases caused by abnormal contraction of vascular smooth muscle are provided in humans and animals in which the above-mentioned various symptoms requiring suppression of abnormal contraction of smooth muscle are promptly and low in orthostatic. It is useful as a therapeutic agent with few side effects such as blood pressure.

As used herein, the term “inhibitor of the Rho kinase signaling system” refers to the activation of RhoA in smooth muscle by, for example, binding of GTP to RhoA by SPC stimulation.
A series of pathways in which Rho kinase is activated by activated RhoA and protein phosphorylation by Rho kinase is enhanced (Biochemistry, Vol. 69, No. 1, 16-29)
P. 1997).

As used herein, the term “Rho kinase” is a type of protein kinase that binds to and activates activated RhoA. Specifically, the term “Rho kinase” directly phosphorylates myosin light chain, It phosphorylates myosin phosphatase and suppresses its activity. Rho kinase (also called ROKα or ROCKII) and p160ROCK (Rhoassociatedcoi)
ledcoilkinase, ROCKKI or RO
Kβ). Inhibitors of the Rho kinase signaling system are apparent Rho kinase inhibitors. Therefore, the inhibitor of the Rho kinase signaling system of the present invention is useful for studying the Rho kinase signaling system.

The EPA used in the present invention, its salts and esters are commercially available as EPA having a purity of 99% by mass or more,
PA-Na purity of about 99% by mass and EPA-E purity of 98% by mass or more can be obtained. Further, it can be obtained by purifying fish oil or EPA-producing bacteria and a culture solution thereof by a known method, for example, a continuous distillation method, a urea addition method, a liquid chromatography method, a supercritical fluid chromatography method, or a combination thereof. If necessary, an esterification treatment can be performed to obtain an alkyl ester such as ethyl ester or an ester such as glyceride. Also, inorganic bases such as sodium salt and potassium salt or benzylamine salt,
Organic bases such as diethylamine salts or arginine salts,
It may be a salt with a basic amino acid such as a lysine salt. In the present invention, EPA means, unless otherwise specified,
In addition to free fatty acids, salts and esters as described above are included. When administered to humans or animals,
Pharmaceutically acceptable ones are preferred.

The inhibitor of abnormal contraction of smooth muscle of the present invention, the injectable composition for treating cardiovascular diseases caused by abnormal contraction of vascular smooth muscle, and the fatty acid mixture in which the inhibitor of the Rho kinase signal transduction system is not an essential component. When it is contained, it is desired that the EPA content in the total fatty acids is large, and at least 80% by mass.
Or more, preferably 90% by mass or more, more preferably 9% by mass.
5% by mass or more can be used, and 97% by mass or more is more preferable. It is preferable that other long-chain fatty acids are small, and it is particularly desirable that the arachidonic acid content is small.
It is preferably less than 1% by mass, more preferably less than 1% by mass, and even more preferably less than 0.5% by mass. Fish oil or a simple concentrate of fish oil is not preferred because fatty acids other than EPA, such as arachidonic acid, docosahexaenoic acid, and docosapentaenoic acid, are contained in 50% by mass or more of the total fatty acids.

The active ingredients are EPA-E, EP
A-Na or EPA-G is preferred, EPA-E is more preferred for oral administration, and EPA-Na or EPA-G is preferred for injections.
A-Na is more preferred. Examples of EPA-G include triicosapentaenoyl glyceride (hereinafter abbreviated as EPA-TG), 1,2-di (icosapentaenoyl) glyceride, 1,3-di (icosapentaenoyl) glyceride, 1-icosapentaenoyl glyceride, 2-icosapentaenoyl glyceride, mixed glycerides in which hydroxyl groups other than EPA groups are substituted with medium-chain fatty acid groups other than EPA, and mixtures thereof. Among these, EPA-TG, diicosapentaenoyl glyceride or diicosapentaenoyl mixed glyceride is preferred, and EPA-TG is more preferred. The ratio of the EPA group to the total amount of the long-chain fatty acid residues of glyceride is at least 80% by mass, preferably 90% by mass or more, more preferably 95% by mass or more, and even more preferably 97% by mass or more.

The inhibitor of the abnormal contraction of smooth muscle and the injectable composition for treating circulatory diseases caused by abnormal contraction of vascular smooth muscle according to the present invention comprise an active ingredient comprising a compound (which may be inevitably contained during purification). ) May be administered alone or in a class of suitable carriers or vehicles commonly used, such as excipients, binders, lubricants, coloring agents, flavoring agents,
If necessary, sterile water or vegetable oil, furthermore, a harmless organic solvent or a harmless solubilizing agent (eg, glycerin, propylene glycol), an emulsifier, a suspending agent (eg, Tween 80, gum arabic solution), an isotonic agent, A suitable pharmaceutical preparation can be prepared by appropriately selecting and combining with a pH adjuster, a stabilizer, a soothing agent and the like.

Because EPA is highly unsaturated, the above formulations may further comprise antioxidants such as butylated hydroxytoluene, butylated hydroxyanisole, propylgallate, gallic acid, pharmaceutically acceptable quinones and α-tocopherol. Is desirably contained in an effective amount.

The dosage form of the preparation includes tablets, capsules,
Microcapsules, granules, fine granules, powders, oral liquid preparations, suppositories, syrups, inhalants, eye drops, ointments, injections (emulsion, suspension, non-aqueous), or use It is administered to patients irrespective of oral or intravenous or intraarterial, inhalation, ophthalmic, rectal, vaginal or external use in the form of a solid injection used as an emulsion or suspension, especially capsules such as soft capsules and Oral administration in microcapsules is preferred. Injectables (emulsion, suspension,
Non-aqueous) or intravenous or intraarterial administration as a solid injection used in an emulsion or suspension at the time of use is preferred. Of the various symptoms requiring suppression of abnormal smooth muscle contraction, oral administration is preferred when a prophylactic effect is desired, and intravenous or intraarterial administration is preferred when relatively rapid efficacy is desired.
When it is necessary to suppress abnormal contraction of respiratory smooth muscle such as bronchial asthma, it can be administered using a known nebulizer as an inhalant.

It should be noted that high-purity EPA-E soft capsules and microcapsules, Epadel and Epadel S (both manufactured by Mochida Pharmaceutical Co., Ltd.) are safe treatments for obstructive arteriosclerosis and hyperlipidemia with few side effects. It is already marketed in Japan as a drug.

The dose of the inhibitor of abnormal contraction of smooth muscle of the present invention and the composition for injection for treatment of circulatory diseases caused by abnormal contraction of vascular smooth muscle is sufficient to exhibit the target action. However, the dose can be appropriately adjusted depending on the dosage form, administration method, number of administrations per day, degree of symptoms, body weight, age and the like. In the case of intravenous or intraarterial administration for which rapid action is desired, 1 to 200 mg, preferably 5 to 100 mg, and more preferably 10 to 50 mg of EPA is used.
The administration may be performed once or several times, and if necessary, the administration may be continued for several hours to several days using an infusion or an infusion pump. In the case of oral administration, EPA is 0.1 to 9 g / day, preferably 0.5 to 6 g / day, and more preferably 1 to 3 g / day.
The dose is divided into three times a day, but the whole dose may be divided into one or several times as needed.

(Experimental Examples) The effects of the present invention will be described in detail with reference to experimental examples in order to explain the present invention in detail, but the present invention is not limited to these examples.

(Experimental Example 1: Effect of EPA on SPC contraction of bovine cerebral artery and porcine coronary artery) A bovine cerebral artery blood vessel or a porcine coronary artery blood vessel was separated, a peripheral tissue and endothelium were removed, and a blood vessel specimen (bovine cerebral artery blood vessel) : Width 5 mm, length 1
5 mm, porcine coronary artery: width 3 mm, length 10 mm). Vascular specimens under ventilation of 95 vol% O ₂ -5 vol% CO ₂ mixed gas, Krebs-HEPES buffer at ^{37 ℃ (Na + 137.4mM, K} + 5.9mM, Ca 2+
1.2 mM, Mg ²⁺ 1.2 mM, Cl ⁻ 148.1 m
M, glucose 11.5 mM, HEPES 11.6 mM
(PH 7.3)) and suspended in a Magnus tube, and the isometric tension was measured with a tension transducer. About 3g
After a steady state was reached with a tension load of, the following experiment was performed.

Depolarized by K ⁺ 118 mM and Ca
The tension at the time of 2 ⁺ -dependent contraction was measured, and this was regarded as normal contraction. After washing K ⁺ , SPC, which is a Ca ²⁺ -independent abnormal vasoconstrictor, was added at 40 μM, and the tension during abnormal contraction was measured. When the tension during abnormal contraction becomes constant,
The change in tension was measured by adding 60 μM of the test substance. When the tension became constant, the change in tension was measured as K ⁺ 118 mM. FIG. 1 shows the change in tension measured using a bovine cerebral artery blood vessel specimen as the test substance, EPA-Na.

Normal contraction was caused by K ⁺ 118 mM, and abnormal contraction comparable to normal contraction was observed by SPC. This abnormal contraction was almost completely suppressed by EPA-Na, but when K ⁺ was again adjusted to 118 mM, the same contraction as normal contraction was observed. That is, EPA-N
a suppresses abnormal contraction due to SPC, but K ⁺ 118
It can be seen that the normal contraction by mM is not affected.

The test substance EPA-E, EPA-TG, sodium salt docosahexaenoic acid, when the docosapentaenoic acid sodium salt or sodium salt arachidonic acid, a similar experiment was performed, a normal by the abnormality contraction and K ⁺ 118 mM by SPC It did not affect any of the contractions.

The above results were similarly obtained for bovine cerebral artery blood vessels and porcine coronary artery blood vessels. From the above results, EPA-
Na was found to specifically suppress Ca ²⁺ -independent abnormal vasoconstriction by SPC without affecting normal contraction by K ⁺ 118 mM.

(Experimental Example 2: Inhibitory effect of EPA-Na on shrinkage of skinned fiber SPC) A small piece (width 0.1 mm, length 1.5 mm) of a porcine coronary artery blood vessel sample was cut out, and a bubble containing Krebs-HEPES buffer was cut out. -Isometric tension was measured with a tension transducer in the wells on the plate. The contraction caused by K ⁺ 118 mM was measured and defined as normal contraction. Then, the relaxation solution (M
g-ATP 4.5 mM, EGTA 2 mM, calmodulin 2.7 μM, carbonyl cyanide p-trifluoromethoxyphenylhydrazone 1 μM, leupeptin 1 μM)
In 20 μM β-escin for 20 minutes at 25 ° C.

With the Ca ²⁺ concentration kept constant (pCa 6.3), 40 μM of SPC was added in the presence or absence of 60 μM of EPA-Na, and the change in tension was measured.
As a result, in the absence of EPA-Na, abnormal contraction stronger than normal contraction was observed by SPC.
Abnormal contraction due to SPC was almost completely suppressed in the presence of Na. When the same experiment was performed in the presence of HA-107730 μM which is a Rho kinase inhibitor instead of EPA-Na, contraction by SPC was almost completely suppressed.

From the above results, it is found that SPC causes Ca ²⁺ -independent abnormal contraction of smooth muscle in a pathway mediated by Rho kinase, and EPA-Na completely suppresses abnormal contraction by this SPC-Rho kinase signal transduction system. I understood.

(Experimental example 3: Inhibitory effect of EPA-Na on SPC contraction of human mesenteric artery)
The mesenteric artery was isolated from the gastrointestinal and surrounding tissues removed from the patient. The surrounding tissue and endothelium of the separated arterial blood vessel were removed to obtain a blood vessel specimen (1 mm in width and 5 mm in length). The blood vessel specimen was suspended in a Magnus tube filled with Krebs-HEPES buffer at 37 ° C under aeration of 95% by volume O ₂ -5% by volume CO ₂ mixed gas, and isometric tension was measured with a tension transducer. did. After a steady state was reached with a tension load of about 0.5 g, the following experiment was performed.

Depolarized by K ⁺ 118 mM and Ca
The tension at the time of 2 ⁺ -dependent contraction was measured, and this was regarded as normal contraction. After washing K ⁺ , 40 μM of SPC was added and the tension at the time of abnormal shrinkage was measured. As a result, the same degree of normal contraction was observed with K ⁺ 118 mM in the blood vessel specimens of all patients. In blood vessel samples of patients with a serum total cholesterol value of 220 mg / dl or more, abnormal contraction almost equivalent to normal contraction was observed by SPC, but in blood vessel samples of patients with a serum total cholesterol value of less than 220 mg / dl, SPC
Almost no shrinkage was observed. This abnormal contraction is E
Almost completely suppressed by the addition of PA-Na 60 μM. From the above results, the serum total cholesterol level was 220 mg / d
It was found that Ca ²⁺ -independent abnormal vasoconstriction occurred in vascular specimens of 1 or more patients, which was suppressed by EPA-Na.

From the above results, EPA selectively inhibits Ca ²⁺ -independent abnormal contraction of smooth muscle,
^{2 +} -dependent physiological smooth muscle normal contraction is not affected, so it is a highly specific inhibitor of abnormal smooth muscle contraction. An injectable composition for treating circulatory diseases, preventing and / or treating dark vision and narrowing of the visual field or an agent for reducing side effects of drug-induced orthostatic hypotension;
It has also been found that the inhibitor is a highly specific inhibitor of the Rho kinase signaling system, which is considered to be the cause of abnormal smooth muscle contraction.

[0045]

EXAMPLES Examples of the inhibitor of the present invention will be shown below.
The present invention is not limited to these. (Example 1) Injection EPA-Na 1 g Glucose 5 g The above components were weighed, and sterilized water for injection was added to make the total amount 10
The volume was adjusted to 0 ml, the pH was adjusted with sodium hydroxide, sterilized by filtration, and then dispensed into sterile vials in 5 ml portions.

(Example 2) Injection EPA-Na 2 g Sodium chloride 0.9 g Gallic acid 0.2 g The above components were weighed, and sterile water for injection was added to make the total amount 100.
ml, adjusted to pH with sodium hydroxide, sterilized by filtration, and dispensed into sterile vials 5 ml each.

(Example 3) Emulsion EPA-TG 100 g Concentrated glycerin 25 g Egg yolk phospholipid 12 g α-tocopherol 3 g The above components were weighed, and distilled water for injection was added to make 1 L.
The mixture was dispersed with a homomixer. This was emulsified with a high-pressure injection emulsifier to prepare a fat emulsion. 100m of the fat emulsion
After dispensing 1 l into plastic bottles,
It was sterilized in an autoclave for 0 minutes to obtain a fat infusion. After sterilization, it was vacuum-packaged with an OV-film (manufactured by Unitika).

(Example 4) Fat infusion EPA-E 50 g Refined soybean oil 950 g Refined egg yolk lecithin 120 g Oleic acid 5 g Concentrated glycerin 250 g
Then, the mixture was dispersed with a homomixer, and distilled water for injection was added to make a total volume of 10 L. This was emulsified with a high-pressure injection emulsifier to prepare a fat emulsion. 200 ml of the fat emulsion
After dispensing into plastic bottles,
It was sterilized in an autoclave for 20 minutes to obtain a fat infusion.
After sterilization, it was vacuum-packaged with an OV-film (manufactured by Unitika).

Example 5 Soft Capsules Soft gelatin capsules (about 0.5 ml volume) were sterilized, ethylated and purified from fish oil, ie, EPA-E90.
0.2% by mass of α-tocopherol in a composition containing 6% by mass, 2.3% by mass of ethyl arachidonic acid, 2.2% by mass of ethyl octadecatetraenoate and 0.7% by mass of ethyl ω-3 icosatetraenoate In addition, EPA-
E was filled to 300 mg, and then sealed.

[0050]

EFFECTS OF THE INVENTION The inhibitor of abnormal contraction of smooth muscle according to the present invention, which comprises EPA as an active ingredient, is an inhibitor in an experiment on abnormal contraction of smooth muscle using an isolated blood vessel specimen or a skinned blood vessel specimen. Useful as Further, the inhibitor of abnormal contraction of smooth muscle of the present invention can be applied to treatment, research, and the like for diseases caused by abnormal contraction of smooth muscle.

Injectable compositions for treating circulatory diseases caused by abnormal contraction of vascular smooth muscle are used for diseases requiring suppression of abnormal contraction of smooth muscle in humans and animals, for example, blood vessels such as cerebral arteries and coronary arteries. Cardiovascular diseases such as spasm, dark vision or narrowed visual field, esophageal spasm, gastric spasm, diarrhea or abdominal pain due to hyperintestinal motility, digestive system diseases such as spastic ileus or abdominal angina, or dyspnea or bronchial asthma It is useful as a therapeutic agent with few side effects such as orthostatic hypotension for respiratory diseases such as. In particular, for diseases for which a rapid effect of a therapeutic agent is desired, such as cerebral vasospasm or coronary vasospasm, Ca ²⁺ at a diseased site is not affected without affecting physiological and normal smooth muscle contraction at a normal site. By suppressing only the independent abnormal contraction of smooth muscle, a therapeutic injection composition having high immediate effect and few side effects can be obtained.

The EPA content ratio in the total fatty acids of the inhibitor of the abnormal contraction of smooth muscle of the present invention and the injectable composition for treating circulatory diseases caused by abnormal contraction of vascular smooth muscle is higher than that of fish oil and its simple concentrate. Because of the high specificity of the suppression effect,
Alternatively, it has an extremely excellent property with a high therapeutic effect and few side effects such as nausea. Further, the inhibitor of the Rho kinase signaling system of the present invention has high inhibitory activity and specificity, and is useful for studying the function of the Rho kinase signaling system or studying the development of a therapeutic agent for diseases involving abnormal smooth muscle contraction. It is.

[Brief description of the drawings]

FIG. 1. Effect of E on SPC contraction in bovine cerebral artery vascular preparations
It is a record of a tension change when observing the influence of PA-Na.

──────────────────────────────────────────────────の Continued on the front page (51) Int.Cl. ⁷ Identification symbol FI Theme coat ゛ (Reference) A61P 43/00 111 A61P 43/00 111 C12N 9/99 C12N 9/99

Claims (8)

[Claims] 1. An agent for suppressing abnormal contraction of smooth muscle, comprising as an active ingredient at least one selected from the group consisting of icosapentic acid, salts and esters thereof. 2. The inhibitor for abnormal contraction of smooth muscle according to claim 1, wherein the active ingredient is contained in a fatty acid mixture, and the content ratio of icosapentic acid, its salt and ester to all fatty acids is 80% by mass or more. . 3. The inhibitor for abnormal contraction of smooth muscle according to claim 1, which comprises icosapentic acid and / or sodium icosapentate as an active ingredient. 4. At least one member selected from the group consisting of icosapentic acid, pharmaceutically acceptable salts and esters thereof.
An injectable composition for treating a circulatory disease caused by abnormal contraction of vascular smooth muscle, comprising: 5. The injectable composition according to claim 4, wherein the active ingredient is contained in a fatty acid mixture, and the content ratio of icosapentic acid, its salt and ester to all fatty acids is 80% by mass or more. 6. The composition for injection according to claim 4, which comprises icosapentic acid and / or sodium icosapentate as an active ingredient. 7. The composition for injection according to claim 4, wherein the circulatory system disease is cerebral vasospasm, coronary vasospasm, pulmonary vasospasm, mesenteric artery vasospasm or finger vasospasm. 8. Rho containing at least one selected from the group consisting of icosapentic acid, salts and esters thereof.
An inhibitor of the kinase signaling system.