JPH0383923A - Migration inhibitor for medial smooth muscle cell - Google Patents

Abstract

PURPOSE:To obtain a migration inhibitor for medial smooth muscle cells, containing an ethanolamine derivative as an active ingredient and useful for preventing and treating arteriosclerosis. CONSTITUTION:The objective substance obtained by mixing an ethanolamine derivative expressed by the formula (R<1> is H or lower alkyl; R<2> is H or acyl derived from nicotinic acid, triene higher fatty acid or pentaene higher fatty acid; R<3> is pyridylmethyl, except that both R<1> and R<2> are H) with a pharmaceutical carrier or excipient and preparing a pharmaceutical according to a conventional method. The aforementioned inhibitor can be prepared in the form of tablet, powder, capsule, granule, oily suspension, syrup, etc. The dose thereof is 100-1500mg for an adult per day administered in 1-3 divided portions. The compound expressed by the formula is capable of preventing and treating thickening and atherogenesis of blood vessels in arteriosclerosis or preventing and treating angiostenosis in transplantation of artificial blood vessels by inhibiting migration of medial smooth muscle cells.

Description

DETAILED DESCRIPTION OF THE INVENTION [Industrial Field of Application] The present invention relates to a fugue inhibitor of smooth muscle cells media (SMC) and an agent for preventing or treating arteriosclerosis, which contains an ethanolamine derivative as an active ingredient.

[Prior art and its problems] Focusing on the high incidence of arteriosclerosis in patients with hyperlipidemia, antihyperlipidemic drugs that lower blood cholesterol and triglycerides have been developed as preventive and therapeutic agents for arteriosclerosis. Antihyperemic agents (eg, clofibrate, probucol, cholestyramine) have been used extensively. However, it has been difficult to treat arteriosclerosis itself with conventional drugs.

However, as the mechanism of arteriosclerosis has been elucidated in recent years, the possibility of developing therapeutic agents for arteriosclerosis itself has been suggested. That is, in arteriosclerosis, thickening and atherosclerosis of blood vessels are observed, migration of SMCs, proliferation of SMCs by migrating light, and accompanying accumulation of cholesterol esters and SMCs.
Foaming is an important risk factor, so SM
Fugue inhibitor C can serve as a new type of arteriosclerosis therapeutic agent.

Furthermore, when an artificial blood vessel is transplanted, it is said that fugue of SMC occurs at the joint site between the blood vessel and the artificial blood vessel, resulting in stenosis of the blood vessel. A similar phenomenon occurs when P T CA (
Pcrcutaneous Transluminal
Coronary angloplasty (percutaneous coronary artery dilatation surgery) also occurs. Therefore, SMC fugue inhibitors can also serve as preventive or therapeutic agents for vascular disorders during artificial blood vessel transplantation and after PTCA surgery.

[Means for Solving the Problem] As a result of intensive research to develop an SMC fugue inhibitor from the above viewpoint, the present inventors learned that a specific ethanolamine derivative has the above-mentioned excellent inhibitory effect, and developed the present invention. Completed the invention.

The present invention is a fugue inhibitory agent for medial smooth muscle cells containing an ethanolamine derivative having the general formula as an active ingredient, and a prophylactic or therapeutic agent for arteriosclerosis containing the above-mentioned ethanolamine derivative as an active ingredient.

In the above formula, R1 represents a hydrogen atom or a lower alkyl group,
Examples of lower alkyl groups include methyl, ethyl, n-propyl, l5o-propyl, n-butyl, l5o-butyl, tert-butyl.

R2 represents a hydrogen atom or represents an acyl group derived from nicotinic acid, triene higher fatty acid or pentaene higher fatty acid, and preferred examples of the acyl group include:
Nicotinic acid, 9.12.15-octadecatrienoic acid (
α-lyllunic acid), 8.9.12-octadecatrienoic acid (γ-lyllunic acid), 8.11.14-eicosatrienoic acid (dihomoγ-lyllunic acid), 5.8.11.1
4.17-Nicosapencitric acid and 7.10.13
．． 16.19-Docosapentaene 3.

Acid can be given. R represents a pyridylmethyl group, and a preferred example is a 3-pyridylmethyl group. However, this excludes the case where both R and R2 in the above formula represent a hydrogen atom.

The ethanolamine derivative having the formula (I) is a known compound described in JP-A-81-189252 and has an inhibitory effect on platelet aggregation, and is useful for preventing diseases caused by it, such as thrombosis and cancer metastasis. It is known to be useful as a drug.

Compound (I) of the present invention is produced according to the method described in the above-mentioned publication.

That is, first, nicotinic acid, triene higher fatty acids, pentaene higher fatty acids, or their reactive derivatives are condensed with the amine of ethanolamine. As the condensing agent, for example, ethyl chloroformate is preferably used. Examples of the above-mentioned reactive derivatives include thiazolidine thionamide derivatives of carboxylic acids. The thus obtained acylated ethanolamine and 3-chloromethylpyridine were combined in the presence of a base such as sodium hydride with benzene,
The reaction is carried out in an aprotic solvent such as toluene.

When the ethanolamine derivative (I) is used as a migration inhibitor of medial smooth muscle cells or as a prophylactic or therapeutic agent for arteriosclerosis, the dosage for adults is approximately 100 to 150 ml per mouthful.
The dose is 0.0 cm, and the dose is divided into 1 to 3 doses as necessary.

The ethanolamine derivative of the present invention is mixed with a pharmaceutical carrier or excipient in a conventional manner and formulated into tablets, powders, capsules, and granules. Examples of carriers or excipients include calcium carbonate, calcium phosphate, starch, sucrose, lactose, talc, magnesium stearate, and the like. In addition to the above-mentioned solid formulations, the compounds of the present invention can also be formulated into liquid formulations such as oily suspensions and syrups.

The compound of the present invention can also be stabilized by inclusion with cyclodextrin.

EXAMPLES Next, the present invention will be explained in more detail with reference to Examples and Test Examples, but the present invention is not limited thereto.

Formulation Example 1: Capsule [Formulation] Main drug: (N-5,8,11,14,17-eicosa (Compound A) Excipient: Corn starch lubricant Magnesium nistearate Total amount (per capsule) 198.5 ■ 3.5 ■ 400.0 ■ Excipients are added to Compound A, which is the upper drug, and it is made into powder or granules. Then, a lubricant is added and mixed evenly, and then it is put into a hard capsule. Fill.

Formulation Example 2: Tablet [Prescription] Main drug: Compound A 500.0■
Excipient: Crystalline cellulose 67.0■〃
: Corn starch 89.0■〃 :
Lactose 44.0 mg Disintegrant: Calcium carboxymethyl cellulose 25.0 ■ Binder: Hydroxyprobyl cellulose 62.5 ■ Add excipient, disintegrant and binder to Compound A, the main drug, and mix evenly, then form granules. Then, a lubricant is added and a compressed tablet is formed. Further, if necessary, the resulting tablets may be coated with a suitable coating (eg, hydroxypropylmethylcellulose, shellac, etc.).

Test Example (I) Establishment of a cultured smooth muscle cell fugue inhibition assay system using Chcmotactic Chamber (trade name, NERO-PROBE) (I) Principle The fugue of smooth muscle cells (SMC) referred to here is based on Chem.
otaxls, which refers to the movement of cells toward the concentration gradient of fugetactic factors.

SMC fugetactic factors include LTB4, which is produced by macrophages and leukocytes, and platelets, which are released and produced by endothelial cells (Platelate DerivedGro).
wth factor (PDGF)) and 12-METE produced by platelets, both of which are considered risk factors for the formation of arteriosclerosis.

The migration mechanism of SMCs is thought to be due to Ca influx into the cells after stimulation by chemotactic factors, which activates the acto-myosin system via the Ca-calmodulin system. Antagonists are potent inhibitors thereof (IC5o.

10”~10’M).

The principle of SMC migration measurement is to use a chamber separated into two layers with a bore filter, with SMC in the upper layer.

Add migratory factors and inhibitors to the bottom layer.

The cells are cultured in a CO incubator for a certain period of time, and the number of cells passing through the bore and fugeting to the lower surface of the filter is measured.

(Left below) (2> Reagents and experimental conditions/cells Porcine aortic media smooth muscle cells (P-SMC) IXIO'
/getelf ・Fugetactic factor/inhibitor (positive control) Verapasll (I0M-10-7M)
Wako Pure Chemical 10 ・Medium DME + 1O%F CS 01800 company ・
F CS Lot No, 300070208Ir
v1ne 5clent1flc Chamber Micro Chemotaxis Chaaber,
48vell Neuro probe filter pore size 8 m, NMP-8-25
80■ Academica/Dyeing Solution Crystal Violet/Fixing Solution
Formalin buffer (I0% formalin/PBS(-)) (3) Experimental method Medium 25 containing fugetactic factors and inhibitors in the lower chamber
Add μQ↓ Cover with a bore filter. At this time, be careful not to introduce air bubbles ↓ Place the chamber on the upper layer and fix it ↓ Sprinkle 150 μg of P-SMCI ↓ Peel off the cells on the upper layer of the filter and add phosphate buffer 0.
Wash with (a) migrated cells should be on the lower layer side ↓ Fix the filter with formalin ↓ 30m1n, at r, t.

Wash 3 times with phosphate buffer 0 and distilled water↓ crystal violet staining ↓　80mIn, atr, t.

Wash 3 times with distilled water ↓ Measure the number of cells on the filter under a stereomicroscope (4) Results and discussion ■ Migration of P-SMC by LTB4 LTB4 is produced by macrophages, leukocytes, etc., and is associated with smooth muscle cells and leukocyte fugue. Indicates the action, etc. At the early stage of arteriosclerosis, microphages infiltrate beneath the vascular endothelial cells, phagocytose denatured LDL, and accumulate it to form foam.

In this way, it is highly conceivable that LTB4 causes fugue movement of smooth muscle cells at the arteriosclerotic site, so LTB4 may be a fugetogenic factor.
The study was conducted using 4. The results are shown in Figure 1.

As is clear from Figure 1, LTB4 was to -11 g/ml for P-SMC at reaction time IQ -20 hr.
It promoted migration in a more concentration-dependent manner, and the effect was maximum at 5XIO'g/ml. However, at 10'g/ml, it showed almost no fugetactic effect. This fact that the fugetactic factor does not exhibit fugetactic effects at high concentrations is due to the fact that other fugetactic factors (I2-HETE).

15-HETE) and is considered to be a characteristic of fugetactic factors.

It is currently unclear why such concentration dependence is exhibited. Based on the above results, the concentration of LT84 used was increased to 10'.
It was set as g/ml.

Next, the reaction time between LTB4 and P-SMC was investigated.

SMC migration occurs when the reaction time with migration factors is too short.
No significant migration occurs, and if the duration is too long, the cells themselves will migrate to some extent even under normal conditions, resulting in too high a fugue rate in the control group.

Therefore, the report of J., NAKAO et al. (Atheroscl.
erosls, 4B (I983), 309-319
), the reaction time of LTB4 was 8 hr, 20 hr
We conducted a study. The results are shown in Figure 2. As is clear from FIG. 2, P-SMC migrated depending on the reaction time with LTB4, and showed significant fugue migration at 20 hr. Therefore, the reaction time between LTB4 and p-SMC was set to 20 hours.

■ Setting the concentration of Verapamll used as a positive control for inhibiting SMC fuguetaxis (I) As mentioned in the principle, SMC fuguetaxis is strongly inhibited by Ca antagonists. Verapaml is as shown in Figure 3,
LTB-induced migration was almost 100% inhibited at IO'M, but no concentration-dependent inhibition was observed from IO to lo-8M10. IC5° is 5
It was XIO'M. This result was 10 times higher than the experimental result using rabbit SMC.

This difference seemed to be due to differences in the rice and wheat animals and the culture conditions. Since the fugetaxis inhibitory effect of Verapamyl at 10-7 M was reproducible, this concentration was used as a positive control.

(II) SMC Migration Inhibitory Effect (I) Reagents and Experimental Methods An experiment on SMC fuge-inhibitory effect was conducted under the conditions and method set in the previous section. The results are shown in Figure 4.

・Fugage factor LTB lo'g/m1 ・Positive control of inhibitor Varapail110'
M n-7~8 ・Reaction time 20 hr ・Measurement sample compound A (purity 90%) 10'M n-3~4 (2
) Results and Discussion Compound A at 10'M significantly inhibited LTB-induced SMC fugue. The suppression rate is approximately 100%, Ver.
apaml was 88%.

This indicates that Compound A has an SMC fugetaxis inhibitory effect, although at a higher concentration than a Ca antagonist.

Acute Toxicity An acute toxicity test was conducted by oral administration using ICR male mice (4 weeks old). LD5o of the compound of the present invention
All values were over 500 μ/kg, confirming high safety.

[Operations and Effects of the Invention] According to the present invention ζ, an inhibitor of fugue of smooth muscle cells media (SMC) containing an ethanolamine derivative as an active ingredient is provided. The ethanolamine derivative of the present invention is SMC
By inhibiting the fugue movement of blood vessels, it is possible to prevent or treat thickening and atherosclerosis of blood vessels in arteriosclerosis, or to prevent or treat blood vessel stenosis during transplantation of artificial blood vessels.

[Brief explanation of drawings]

FIG. 1 is a graph showing the relationship between LTB4 concentration and the number of migrated P- and SMC. Figure 2 shows the contact time with LTB and migrated P-SMC.
It is a graph showing the relationship between the numbers of. Figure 3 shows Verapamll concentration and migrated P-SM.
It is a graph showing the relationship between the numbers of C. FIG. 4 is a graph showing the relationship between the concentration of Vcrapa... or Compound A and the number of migrated P-SMC.

Claims (1)

[Claims] 1) General formula (I) ▲There are mathematical formulas, chemical formulas, tables, etc.▼(I) (In the formula, R^1 represents a hydrogen atom or a lower alkyl group,
R^2 represents a hydrogen atom or an acyl group derived from nicotinic acid, triene higher fatty acid or pentaene higher fatty acid, and R^3 represents a pyridylmethyl group. However, this excludes the case where R^1 and R^2 both represent hydrogen atoms. ) A migration inhibitor of medial smooth muscle cells containing an ethanolamine derivative having the following as an active ingredient. 2) A prophylactic or therapeutic agent for arteriosclerosis, which contains the ethanolamine derivative according to claim 1 as an active ingredient.