JPH0623107B2 - Pharmaceutical composition containing prostacyclin as an active ingredient - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION <Industrial field of application> The present invention relates to a pharmaceutical composition containing prostacyclins as an active ingredient. More specifically, the present invention provides that the oxygen atom at the 6,9-position of prostaglandin I ₁ is a methine group, ie, —H.
The present invention relates to a pharmaceutical composition containing a novel C = -substituted prostacyclin as an active ingredient.

<Prior Art> Prostacyclin is a local hormone mainly produced mainly in the inner wall of the blood vessel of an artery, and is an important factor that regulates the cell function of a living body by its strong physiological activities such as platelet aggregation inhibitory activity and vasodilatory activity. There is an attempt to use this product directly as a medicine (PJLew
is & JOGrady “Clinical Pharmacology of Prosta-cy
clin ”Raven Press, NY, 1981).

However, since natural prostacyclin has an enol ether bond which is very easily hydrolyzed in the molecule, it is easily inactivated under neutral or acidic conditions, and it cannot be said to be a preferable compound as a drug because of its chemical instability. Therefore, a chemically stable synthetic prostacyclin derivative having physiological activity similar to that of natural prostacyclin has been intensively studied both inside and outside.

Among them, a derivative obtained by substituting the oxygen atom at the 6,9-position of prostacyclin with a methylene group, that is, 9 (0) -methanoprostacyclin (carbacycline) is known as a prostacyclin that sufficiently satisfies chemical stability. Ori (DR Mortons “Prostacyclin” JRVane and S. Bergs
See Tom, Eds, Raven Press, NY, 1979, pp31-41) Expected as a drug. However, the biological activity of 6,9 (0) -methanoprostacyclin is weaker than that of natural prostacyclin, and its action selectivity cannot be said to be specific, so that it is not necessarily a preferable compound. On the other hand, as stable prostacyclins, the oxygen atom at the 6,9-position is -N
Derivatives substituted with the = group, namely nitriloprostacyclin, are known and their biological activity is known to be comparable to natural prostacyclins (GL Bundy et al., Tetr.
ahedron Letter, 1371 (1978) and W. Bartmann et al., Tetrahe.
dron Letter, 23, 3467 (1982)).

<Object of the Invention> An object of the present invention is to provide a pharmaceutical composition containing a novel prostacyclin, which is chemically stable and has an excellent pharmacological action, as an active ingredient.

<Structure and Effect of the Invention> The present inventors focused on the chemical structure of the above-described stabilized prostacyclin, and newly obtained a derivative in which an oxygen atom at the 6,9-position was replaced with a methine group, that is, a —CH═ group. The heading has arrived at the present invention. That is, the present invention provides the following formula [I] With prostacyclin represented by as an active ingredient,
A pharmaceutical composition for lowering blood pressure or suppressing platelet aggregation, which is contained together with a pharmaceutically acceptable carrier.

Here, R ⁵ is a hydrogen atom or a methyl group. As other reference examples, a C _{2 to} C ₁₀ alkyl group, a substituted or unsubstituted phenyl group, a substituted or unsubstituted alicyclic group, a substituted or unsubstituted phenyl (C _{1 to} C ₂ ) alkyl group, Examples include tri (C ₁ -C ₇ ) hydrocarbon-silyl groups, or one equivalent of cations. The alkyl group of C ₂ -C _10, for example, ethyl, n- propyl, iso- propyl, n- butyl, sec- butyl, tert- butyl, n- pentyl, n- hexyl, n- heptyl, n- octyl ，
Examples thereof include linear or branched ones such as n-nonyl and n-decyl.

Examples of the substituent of the substituted or unsubstituted phenyl group include a halogen atom, a hydroxy group, a C _{2 to} C ₇ acyloxy group, and a C _{1 to} C ₄ which may be substituted with a halogen atom.
Alkyl group, C ₁ which may be substituted with halogen atom
-C ₄ alkoxy group, a nitrile group, a carboxyl group or a _(C 1 ~C ₆₎ alkoxycarbonyl group are preferable.
Here, the halogen atom is fluorine, chlorine, bromine, or the like.
Particularly, fluorine or chlorine is preferable. Examples of the C _{2 to} C ₇ acyloxy group include acetoxy, propionyloxy, n
-Butyryloxy, iso-butyryloxy, n-valeryloxy, iso-valeryloxy, caproyloxy,
Enanthyloxy, benzoyloxy, etc. can be mentioned.

Preferable examples of the C ₁ -C ₄ alkyl group which may be substituted with halogen include methyl, ethyl, n-propyl, iso-propyl, n-butyl, chloromethyl, dichloromethyl, trifluoromethyl and the like. You can As the C ₁ -C ₄ alkoxy group which may be substituted with halogen, for example, methoxy, ethoxy, n-
Preferred examples include propoxy, iso-propoxy, n-butoxy, chloromethoxy, dichloromethoxy, trifluoromethoxy and the like. (C _{1 to} C
₆ ) Examples of the alkoxycarbonyl group include methoxycarbonyl, ethoxycarbonyl, butoxycarbonyl, hexyloxycarbonyl and the like.

The substituted phenyl group can have 1 to 3 substituents, preferably 1 substituent as described above.

The substituted or unsubstituted alicyclic group is substituted or unsubstituted with the same substituents as described above, saturated or unsaturated C _{5 to} C ₈ , preferably C _{5 to} C ₈ , particularly preferably C ₆ groups such as cyclopentyl, cyclohexyl, cyclohexenyl, cycloheptyl, cyclooctyl and the like can be mentioned.

Examples of the substituted or unsubstituted phenyl (C ₁ -C ₂ ) alkyl group include benzyl, α-phenethyl, β-phenethyl, etc. in which the phenyl group is substituted with the same substituent as described above or unsubstituted. .

Examples of the tri (C ₁ -C ₇ ) hydrocarbon-silyl group include trimethylsilyl, triethylsilyl, t-butyldimethylsilyl group-containing tri (C ₁ -C ₄ ) alkylsilyl, and t-butyldiphenylsilyl group-containing diphenyl group. Preferable ones are (C ₁ -C ₄ ) alkylsilyl, tribenzylsilyl group, dimethyl- (2,4,6-tri-t-butylphenoxy) silyl group and the like. As one equivalent of cations, for example, alkali metal cations such as Na ⁺ and K ⁺ ; divalent or trivalent metal cations such as 1 / 2Ca ²⁺ , 1 / 2Mg ²⁺ and 1 / 3Al ³⁺ ; ammonium ion, Examples thereof include ammonium cations such as tetramethylammonium ion.

R ⁵ is particularly preferably a methyl group or a hydrogen atom.

R ¹ is a hydrogen atom, but another reference example is a methyl group.

R ² is an alkyl group of unsubstituted _C 5 -C _8. As another reference example, a substituted C ₁ -C ₅ alkyl group substituted with a phenyl group, a phenoxy group, a C ₁ -C ₈ alkoxy group or a C ₅ -C ₈ cycloalkyl group which may be substituted; or Examples thereof include a substituted or unsubstituted alicyclic group. C ₅ The unsubstituted alkyl group -C _8, it may be straight chain or branched, for example n- pentyl, n- hexyl, 2-methyl-1-hexyl, 2-methyl-2 -Hexyl, n-heptyl, n-octyl, etc.,
Preferably n-pentyl, n-hexyl, 2-methyl-
1-hexyl, 2-methyl-2-hexyl and the like can be mentioned. The alkyl group of the substituted C ₁ -C ₅ alkyl group may be linear or branched,
Examples thereof include methyl, ethyl, n-propyl, iso-propyl, n-butyl, sec-butyl, t-butyl, n-pentyl and the like. These alkyl groups are
Phenyl group; phenoxy group; methoxy, ethoxy, n-
C ₁ -C ₆ alkoxy group such as propoxy, iso-propoxy, n-butoxy, iso-butoxy, t-butoxy, n-pentoxy, n-hexoxy; substituted with C ₅ -C ₆ cycloalkyl group such as cyclopentyl and cyclohexyl Has been done. These substituents may be further substituted with the substituents mentioned as the substituents for the substituted phenyl group for R ⁵ .

Examples of the substituted C _{1 to} C ₅ alkyl group include C _{1 to} C ₂ substituted by a phenoxy group or a phenyl group which may be substituted with a fluorine atom, a chlorine atom, a methyl, ethyl or trifluoromethyl group among these. Alkyl group or propoxymethyl, ethoxyethyl, propoxyethyl, butoxymethyl, methoxypropyl, 2-
Ethoxy-1,1-dimethylethyl, propoxydimethylmethyl, cyclohexylmethyl, cyclohexylethyl, cyclohexyldimethylmethyl, 2-cyclohexyl-1,1-dimethylethyl and the like are preferable.

Examples of the substituted or unsubstituted alicyclic group are the same as those listed for R ⁵ . R ² is preferably n-pentyl, 2-methyl-1-hexyl.

R ³ and R ⁴ represent a hydrogen atom. As another reference example, the same or different, C ₂ -C ₇ acyl group, tri _(C 1 ~C ₇₎ hydrocarbon - a group forming an acetal bond together with a silyl group, or a hydroxyl oxygen atom.

The C ₂ -C ₇ acyl group, for example, may be mentioned acetyl, propionyl, n- butyryl, iso- butyryl, n- valeryl, iso- valeryl, caproyl, enanthylic, benzoyl and the like.

Of _these, C 2 -C ₆ aliphatic acyl group such as acetyl, n- or iso- butyryl, caproyl, or benzoyl are preferred.

As the tri (C ₁ -C ₇ ) hydrocarbon-silyl group, the same groups as those mentioned for R ⁵ can be mentioned.

Examples of the group that forms an acetal bond with the oxygen atom of the hydroxyl group include methoxymethyl, 1-ethoxyethyl,
2-methoxy-2-propyl, 2-ethoxy-2-propyl, (2-methoxyethoxy) methyl, benzyloxymethyl, 2-tetrahydropyranyl, 2-tetrahydrofuranyl, 4- (4-methoxy-tetrahydropyranyl) Group or 6,6-dimethyl-3-oxa-2-oxo-
A bicyclo [3.1.0] hex-4-yl group may be mentioned. Of these, 2-tetrahydropyranyl, 2
-Tetrahydrofuranyl, 1-ethoxyethyl, 2-methoxy-2-propyl, (2-methoxyethoxy) methyl, 4- (4-methoxytetrahydropyranyl) group, 6,
6-Dimethyl-3-oxa-2-oxo-bicyclo [3.
1.0] hex-4-yl group or dimethyl (2,4,6-tri-
The t-butylphenyloxy) silyl group is particularly preferred.

Among the reference examples of R ³ or R ⁴ , t-butyldimethylsilyl group, 2-tetrahydropyranyl group, acetyl group, 1-methoxy-1-methylethyl group, 4- (4-methoxytetrahydropyranyl) group , 6,6-Dimethyl-3
-Oxa-2-oxo-bicyclo [3.1.0] hex-4
An -yl group and a dimethyl (2,4,6-tri-t-butylphenyloxy) silyl group are preferred.

Specific examples of the prostacyclins provided by the present invention include the following (1), (4), (5) and (6) for them.
There is one. In addition, the following others are given as reference examples.

(1) 9 (0) methano-Δ ^{6 (9)} prostaglandin I ₁ (2) 16,17,18,19,20-pentanor-15-cyclopentyl-9 (0) methano-Δ ^{6 (9)} prostaglandin Glandin I ₁ (3) 16,17,18,19,20-Pentanol-15-cyclohexyl-9 (0) methano-Δ ^{6 (9)} Prostaglandin I ₁ (4) 17,20-Dimethyl-9 ( 0) Methano-Δ ^{6 (9)} prostaglandin I ₁ (5) 15-methyl-9 (0) methano-Δ ^{6 (9)} prostaglandin I ₁ (6) (1) to (5) methyl ester (7) Ethyl ester of (1) to (5) (8) (6) 11,15-bis-t-butyldimethylsilyl ether (9) (6) 11-position is methoxyisopropyl group, 15-position is t -Compound protected with butyldimethylsilyl group (10) (6) 11-position of t-butyldiphenylsilyl group, 15-position protected with t-butyldimethylsilyl group (11) 11-position of (6) Is 4- (4-methoxytetrahydropyranyl) group, 15-position is t-bu Compound protected with dimethylsilyl group (12) (6) is protected with dimethyl (2,4,6-tri-t-butylphenyloxy) silyl group at position 11 and with t-butyldimethylsilyl group at position 15 Compounds (13) Sodium salts, ammonium salts, and potassium salts of carboxylic acids of (1) to (5) The prostacyclins of the present invention and those mentioned as reference examples above are represented by the following formula [II] It is produced by treating a hydroxyprostacyclin represented by the formula (4) with a sulfonic acid anhydride in the presence of a base, and subjecting it to a deprotection reaction, a hydrolysis reaction, and a salt formation reaction, if necessary.

The hydroxyprostacyclins of the above formula [II], which are original compounds, are novel compounds and are produced by the production method described below.

In the above formula [II], R ³ ′ and R ⁴ ′ are the same or different and are a C _{2 to} C ₇ acyl group, or a group having an acetal bond together with an oxygen atom of a hydroxyl group, and specific examples of such a group are as described above. Is. ^{R 8, R 8 ', R} 8 " is an alkyl group of _C 1 -C ₇ identical or different, aryl group or an alkyl group _.C 1 -C ₇ representing an aryl alkyl group, are, for example, methyl, ethyl, n -Propyl, iso-propyl, n-butyl, sec-butyl, tert-butyl, n-
Pentyl, n-hexyl, n-heptyl and the like can be mentioned. Examples of the aryl group include a phenyl group, and examples of the arylalkyl group include a benzyl group,
Examples thereof include a phenethyl group.

Hydroxyprostacyclins are first dissolved in an excess of base in the medium and then treated with sulfonic anhydride. The reaction solvent is preferably a halogenated hydrocarbon such as methylene chloride, dichloroethane, chloroform, carbon tetrachloride,
Methylene chloride is particularly preferably used. The base used is preferably a pyridine base such as pyridine, picoline, collidine, ruticine, 4-dimethylaminopyridine,
Pyridine is particularly preferably used, and 4-dimethylaminopyridine is preferably used together with pyridine. The amount of the base used is 2 to 100 equivalents, preferably 10 to 30 equivalents relative to the raw material compound, and 4-dimethylaminopyridine may be used in a catalytic amount when used together with pyridine. Use 5% equivalents.

The sulfonic acid anhydride is preferably trifluoromethanesulfonic acid, methanesulfonic acid, p-toluenesulfonic acid anhydride, or a mixed acid anhydride of C ₁ -C ₃ alkylsulfonic acid and trifluoromethanesulfonic acid, particularly trifluoromethane. Sulfonic anhydride is preferred. The sulfonic acid anhydride used is 2-100 equivalents, preferably 5-30 equivalents, based on the starting compound. The reaction temperature is -30 ° C 50 ° C, preferably -10 ° C to 20 ° C. The reaction time is confirmed by the disappearance of the starting compound by TLC.

The reaction solution thus obtained is treated with saturated N 2
After sufficiently adding aHCO ₃ water, post-treatment may be carried out according to a method usually performed. For example, a sparingly soluble organic solvent in water such as hexane, pentane, petroleum ether, or ethyl ether may be added, or the reaction mixture may be directly condensed under reduced pressure, and the same procedure as described above may be performed to wash the resulting organic mixture with brine to remove anhydrous sulfuric acid. After drying with a drying agent such as magnesium, anhydrous sodium sulfate, or anhydrous potassium carbonate, the organic medium is removed under reduced pressure to obtain a crude product. The crude product can be purified by a purification means such as column chromatography, thin layer chromatography, liquid chromatography or the like, if desired. The product thus obtained can be further subjected to a deprotection reaction, a hydrolysis reaction and a salt formation reaction, if necessary.

When the protecting group is a group which forms an acetal bond together with the oxygen atom of the hydroxyl group, for example, acetic acid, a pyridinium salt of p-toluenesulfonic acid or a cation exchange resin is used as a catalyst to remove the protecting group of the hydroxyl group. , Tetrahydrofuran, ethyl ether, dioxane, acetone, acetonitrile and the like are preferably used as a reaction solvent. The reaction is usually performed in the temperature range of −78 ° C. to + 30 ° C. for about 10 minutes to 3 days. When the protective group is an acyl group, it is hydrolyzed in, for example, an aqueous solution of caustic soda, caustic potash, calcium hydroxide or a water-alcohol mixed solution, or a methanol or ethanol solution containing sodium methoxide, potassium methoxide or sodium ethoxide. It can be carried out by disassembling.

Hydrolysis reaction of the ester group of the carboxyl group, for example, using an enzyme such as lipase, in a solvent containing water -10
It is carried out in the temperature range of ℃ to +60 ℃ for 1 minute to 24 hours.

The product after the deprotection reaction or hydrolysis reaction can be purified by the same purification means as described above.

The compound having a carboxyl group, which has been purified by the reaction for removing a protecting group as described above, is then optionally subjected to a salt purification reaction to give a corresponding carboxylic acid salt. The salt-forming reaction is known per se, and neutralizes approximately the same amount of a carboxylic acid and a basic compound such as sodium hydroxide, potassium hydroxide and sodium carbonate, or ammonia, trimethylamine, monoethanolamine and morpholine by a usual method. It is carried out by reacting.

The starting material compound [II] used in the production method of the present invention can be obtained by the following reaction.

The compound of the above formula [III] is a known compound, and can be produced by the method described in P409 (1982) of the 102nd Annual Meeting of the Pharmaceutical Society of Japan.

The compound [IV] is obtained by isomerizing the exo double bond of the compound [III] and, if desired, converting the protecting group. Isomerization of the exo double bond is accomplished by heating with ruthenium trichloride.trihydrate and anhydrous potassium carbonate in methanol. Conversion of the protecting group is carried out by the method described above. Then, silylation is introduced into this product [IV] to derive the compound [V]. A silylating agent such as hexamethyldisilazane was used for the introduction of the silyl group, and tetrabutylammonium fluoride was used as an auxiliary agent, preferably 0.05 to 0.50 equivalent, while confirming the disappearance of the starting compound [IV]. Will be achieved.

The carbonyl group of the obtained ketone body [V] can be reduced to the compound [II] which is the starting material of the present invention.
As the reducing agent, an ordinary carbonyl reducing agent such as sodium borohydride, lithium hydride, aluminum, etc. is preferably used, and the reaction condition is also selected as an ordinary mild condition.
Thus, the desired hydroxyprostacyclin represented by the formula [II] is obtained.

The novel prostacyclins represented by the formula [I] provided by the present invention surprisingly have a very strong biological activity. For example, 9 (0) methano-Δ ^{6 (9)} prostaglandin I ₁ induces ADP-induced rabbit platelet aggregation by IC.
Not only can it be suppressed at ₅₀ 0.034 / ml, but it also has a cytoprotective effect on rabbit bone epithelial cells at pH 3 of 10 ^-6 M. On the other hand, leukemia cell growth inhibitory action of L 1210 was observed at an IC _{50 of} about 2 to 4 μg / ml.

The active compounds of the present invention can be administered to warm-blooded animals, such as humans or non-human animals, in need of controlling vascular movement. The active compound of the present invention can be administered to warm-blooded animals in need of controlling vascular system movement for prevention or treatment.

The active compounds of the present invention can be applied to patients, for example, antiangina, vasodilation, hypotension, antithrombotic, antiarteriosclerosis, antianginal, antimyocardial infarction, antipulmonary arterial hypertension, antistroke, antitransient ischemic attack. (Transient Ischemic Attac
k), can be administered for patients with anti-deep vein thrombosis or anti-peripheral vasculopathy. As a reference, the active compound of the present invention can also be administered to suppress malignant tumors or metastasis of malignant tumors in addition to anti-endotoxin shock and anti-thrombocytopenic purpura. The compounds of the invention can also be used in organ transplantation, vascular surgery or extracorporeal circulation. For example, blood, blood products, blood substitutes, and other isolated body parts (eg, limbs or organs; if attached to the original body and also stored separately or prepared for transplantation) It can also be used as an additive to fluids used for artificial extracorporeal circulation and perfusion (including when attached to a new body). During such circulation and perfusion, condensed platelets tend to occlude blood vessels and circulatory organs. The presence of the active compounds according to the invention avoids this blockage. For this purpose, the active compounds according to the invention are gradually or in one or several portions divided into circulating blood, blood of blood-donating animals, parts of the body to be perfused (attached to or separated from the recipient). ), Or two or all of these at 0.1-1 μg / kg-body weight / min. The above-mentioned reference example also exhibits the same operation.

For these purposes, these compounds may be administered orally or parenterally, for example, rectally, subcutaneously, intramuscularly or intravenously, preferably orally or intravenously.

For oral administration, it can be a solid preparation or a liquid lumber. Examples of solid formulations include tablets, pills,
There are powders and granules. In such solid dosage forms one or more active substances is at least one pharmaceutically acceptable carrier, such as the commonly used sodium bicarbonate, calcium carbonate, potato starch,
It is mixed with sucrose, mannitol, carboxymethylcellulose, etc. Formulation operation is carried out according to a conventional method,
It may contain additives for formulation other than the above, for example, lubricants such as calcium stearate, magnesium stearate and glycerin.

Liquid formulations for oral administration include, for example, emulsions, solutions,
Includes suspensions, syrups or xyls. These formulations contain a commonly used pharmaceutically acceptable carrier such as water or liquid paraffin.

Enteric-coated preparations for oral administration include solid preparations such as those mentioned above, such as cellulose acetate phthalate, hydroxypropylmethyl cellulose phthalate, polyvinyl alcohol phthalate, styrene maleic anhydride copolymer or methacrylic acid, methyl methacrylate copolymer. It is produced by spraying an organic solvent or water solution of an enteric substance to form an enteric coating. Enteric coated solid preparations such as powders and granules can be encapsulated.

The pharmaceutically acceptable carrier in the present specification includes other auxiliary agents, fragrances, stabilizers, or preservatives which are usually used as necessary.

The liquid formulation may also be administered in capsules made of an absorbable material such as gelatin.

In addition, the prostacyclins of the present invention can be administered intranasally, and as such nasal solution for intranasal administration, for example, hydroxypropylcellulose, a base such as lactose, and prostacyclins in saline or isotonic glucose solution. Those dissolved or emulsified are preferably used.

Solid form preparations for rectal administration include suppositories which contain one or more active substances and are manufactured by a method known per se.

Formulations for parenteral administration are provided as sterile aqueous or non-aqueous solutions, suspensions, or emulsions. Non-aqueous solutions or suspensions include vegetable oils such as propyl glycol, polyethylene glycol or olive oil,
An injectable organic ester such as ethyl oleate is a pharmaceutically acceptable carrier. Such formulations may also contain adjuvants such as preservatives, wetting agents, emulsifying agents, dispersing agents and stabilizing agents. These solutions, suspensions, and emulsions can be sterilized by, for example, appropriately passing through a bacteria-retaining filter, blending a bactericide, or irradiating. In addition, a sterile solid preparation can be produced and dissolved in sterile water or a sterile injection solvent immediately before use before use.

The compound of the present invention can also be used by forming an inclusion compound with α, β or γ-cyclodextrin or methylated cyclodextrin.

The dose of the compound of the present invention depends on the condition of the subject receiving the administration,
Usually depends on age, sex, weight, route of administration, etc.
It can be administered in an amount of 0.02 μg to 100 mg / kg-body weight / day. Such dosage may be once or several times daily, for example 2
It is also possible to administer the drug in 6 divided doses.

<Example> Hereinafter, the present invention will be described in more detail with reference to Examples.

Reference example 1 A solution of alcohol 1 (66 mg, 0.11 mM) in methylene chloride (2 ml) was cooled to 0 ° C. and pyridine (20 equivalents), 4-
DMAP (4-dimethylaminopyridine, catalytic amount) and trifluoromethanesulfonic anhydride (10 equivalents) are added in this order. TLC (ether: n-hexane = 1: 1
1) When the disappearance of the raw material is confirmed, saturated NaHC
An O ₃ aqueous solution is added, and the mixture is extracted with ether. The ether layer is further washed with saturated aqueous NaCl, dried over anhydrous MgSO ₄ , and the solvent is distilled off. The residue is subjected to silica gel column chromatography (ether: n-hexane = 1: 3).
Separating purified by olefin Another object is 20 mg (35
%) And 18 mg (24%) of silyl ether 3 was by-produced.

Product; IR (neat) ν: 2925,2850,1735,1440 cm ^-1 NMR (δ, CDCl ₃ ): 5.45 (m, 2H), 5.25 (br.s.1H), 4.68 (m, 2H), 4.20 -3.70 (m, 4
H), 3.68 (s, 3H), 3.65-3.25 (m, 2H), 2.95 (m, 1H), 0.90 (m, 3
H). Mass (m / e): 501 (M = OCH ₃ ), 345m / e 501.3538 (calcd for C
₃₁ H ₄₉ O ₅ , 501.3567, M-OCH ₃ ) Reference example 2 Olefin 2 (20 mg, 0.038 mM) was added to AcOH: H ₂ O: TH
Heat at 65 ° C. in a mixed solvent of F = 3: 1: 1: 1 (1.5 ml). After 4 hours TLC (acetone: methylene chloride = 1: 1)
2) When it was confirmed that the raw materials had disappeared, a saturated aqueous solution of NaHCO ₃ was added to this reaction solution at 0 ° C.
Extract with (10 ml x 4). The ether layer is washed with saturated aqueous NaCl solution, dried over anhydrous MgSO ₄ , and the solvent is distilled off. The residue was separated and purified by silica gel chromatography (acetone: methyl chloride = 1: 2) to obtain 13 mg (95%) of the target diol 4 . 4 ; Reference example 3 Diol 4 (13 mg, 0.036 mM) was added at room temperature to THF-H ₂ O = 3: 1.
Dissolved in a mixed solvent (1 ml) of 5M-NaOH.
(0.2 ml) is added and stirring is continued overnight. When the temperature was further raised to 40 ° C. and the reaction was carried out for 20 hours under the conditions, disappearance of the raw materials was confirmed by TCL (ether). The reaction solution was returned to room temperature, diluted with ether (20 ml), and then carefully neutralized with 10% HCl water and pH = 4.0 buffer, and finally pH =
It was set to 3-4. This was extracted with ethyl acetate (20 ml x 3), the organic layers were combined and washed with saturated aqueous NaCl (3 ml x 2). This was dried over anhydrous MgSO ₄ , the solvent was distilled off, and the resulting residue was further purified by silica gel column chromatography (ethyl acetate: methanol = 15: 1) to give 12 mg (96% of target 5 ). ) Got. 5 ; Example 1 In Vitro Platelet Aggregation Inhibitory Action The in vitro platelet aggregation inhibitory action of the test drug was assayed using rabbits. That is, blood was collected from the ear vein of a Japanese white male rabbit having a body weight of 2.5 to 3.5 kg at a ratio of 3.8% trisodium citrate solution to 9 blood, and after centrifugation at 1000 rpm for 10 minutes, the upper layer was subjected to PRF (rich). Platelet plasma). The lower layer was further centrifuged at 2800 rpm for 10 minutes and separated into two layers, and the upper layer was separated as PPP (platelet poor plasma). The platelet count was adjusted to 6 × 10 ⁵ / μ with PPP. Adjusted P
25 μ of the test drug was added to 250 μ of RP and pre-incubated at 37 ° C. for 2 minutes, and then 20 μM (final) of ADP was added and the change in permeability was recorded by an acrigometer. The test drug was dissolved in ethanol at 10 mg / ml and then serially diluted with a phosphate buffer (pH 7.4) before use. The aggregation inhibition rate was calculated by the following formula.

T ₀ : Permeability of (phosphate buffer solution-added system) T: Permeability of test drug-added system The lowest concentration of the drug with an inhibition rate of more than 50% was shown as an IC ₅₀ value.

The IC ₅₀ was determined using 9 (0) methano-Δ ^{6 (9)} -prostaglandin I ₁ of Reference Example 3 as the test drug, and was found to be 40 ng / m.
It was l.

Example 2 In Vivo Hypotensive Effect Male wistar rats weighing about 250 g were intraperitoneally administered with a mixed anesthetic solution of urethane and α-chloralose (urethane 500 g / kg, α-chloralose 100 mg / kg). After anesthesia,
The rat was fixed on its back, the blood pressure was measured by releasing the pressure transducer from the common carotid artery, and the heart rate was calculated from the pulse wave. After the test compound of Reference Example 3 was dissolved in ethanol,
Diluted with 0.9% saline, the ethanol concentration was adjusted to be less than 10% even at the highest dose administered. A cannula inserted into the femoral vein was used for administration, and when the test solution of Reference Example 3 was rapidly intravenously injected at a volume of 1 ml / kg, a dose-dependent decrease in blood pressure was observed at 0.1 μg / kg or more. The required dose for lowering blood pressure of 20 mmHg is about 0.5-0.8μ.
It was g / kg. A slight increase in heart rate was observed at 1 μg / kg and above.

Example 3 One tablet having the following composition was produced.

Active ingredient 200 μg Lactose 280 mg Potato starch 80 mg Polyvinylpyrrolidone 11 mg Magnesium stearate 5 mg 576 mg Active ingredient, lactose and potato starch are mixed and evenly moistened with 20% ethanol solution of polyvinylpyrrolidone, and passed through a 20 mm mesh sieve, 45 It was dried at ° C and passed through a 15 mm mesh screen again. The granules thus obtained are mixed with magnesium stearate,
Compressed into tablets.

The compound of Reference Example 3 was typically used as the active ingredient.

Example 4 A hard gelatin capsule was prepared in which one capsule contained the following composition:

Active ingredient 200 μg Microcrystalline cellulose 195 mg Amorphous silicic acid 5 mg 400 mg Finely powdered active ingredient, microcrystalline cellulose and unpressed amorphous silicic acid were thoroughly mixed and packed into a hard gelatin capsule.

The compound of Reference Example 3 was typically used as the active ingredient.

Example 5 (Preparation of Ampoule) An ampoule containing the following composition in one ampoule (5 ml volume) was produced.

Active ingredient 200 μg Polyethylene glycol 600 200 mg Distilled water Total 50 ml Polyethylene glycol and active ingredient were dissolved in water under nitrogen, boiled, cooled under nitrogen and distilled. Pre-treated water was added to this solution and the mixture was passed under aseptic conditions at a given volume. This manufacturing is performed in the diffused light.

The filling was performed in a nitrogen stream, and the sterilization was performed at 121 ° C. for 20 minutes.

The compound of Reference Example 3 was used as the active ingredient.

Example 6 10 mg of the compound of Reference Example 3 is dissolved in 5 ml of ethanol, sterilized through a bacteria-retaining filter, 0.1 ml is added per 1 ml ampoule, and the ampoule is sealed. The contents of the ampoule are diluted to an appropriate volume. For example, it is diluted to 1 ml with Tris-HCl buffer of pH 8.6 for injection administration.

Example 7 In Vitro Platelet Aggregation Inhibitory Action Platelets used were prepared by the following method.

Rabbit Platelets Under ether anesthesia, blood was collected from the carotid artery of rabbits (JW, ♂) as citrated blood, and the blood was collected at 20 ° C and 1000 rpm for 10 min.
The platelet-rich plasma (PRP) was collected by centrifugation. The remaining blood was further centrifuged at 20 ° C. at 3000 rpm for 10 minutes to collect platelet poor plasma (PPP). The number of platelets in PRP was measured with a hemocytometer (Nippon Kohden, NEK-4500), and PRP was diluted with PPP to obtain a platelet number of 30-40 × 10 ⁴ cells / μl.
Was used as a PRP for measuring platelet aggregation.

Human platelets Blood collected from healthy adult male volunteers as citrated blood was centrifuged at 20 ° C at 800 rpm for 10 minutes to obtain platelet rich plasma (PRP). Remaining blood at 20 ℃ at 3000rpm for 10min
The platelet-poor plasma (PPP) was obtained by centrifugation. The number of platelets in PRP is measured and P is adjusted to 30-40x10 ⁴ cells / μl.
It was diluted with PP to obtain PRP for measurement.

The test drug to be further used was prepared by the following method.

That is, the compound 4 obtained in Reference Example 2 described above was treated with EtO
After dissolving and diluting with H, just before adding to PRP, 50 mM P
It was diluted 20-fold with B-Saline (pH 7.4) before use. In addition, Con
As trol, EtOH similarly diluted was used.

In addition, ADP (5'-adenos as a platelet aggregation inducer
ine diphosphate Na, Seikagaku Corporation) is 0.1 M Tris-H
It was used after being dissolved and diluted with Cl Buffer (pH 7.4).

After making the above preparations, 15 μl of the test drug containing the compound 4 of Reference Example 2 was added to 270 μl of PRP and heated at 37 ° C. for 20 minutes in the case of rabbit platelets and 30 minutes in the case of human platelets, and then in an aggregometer. ADP 15 μl (final concentration; rabbit 20 μM, human 6 μM) was added, and the maximum aggregation value was measured. The control was prepared by adding a solvent instead of the test drug, and the inhibition rate (Inhib
ition%), a dose-response curve was prepared, and the IC ₅₀ value was determined by the method of least squares.

As a result, the compound 4 of Reference Example 2 dose-dependently affected the ADP.
It inhibited the platelet aggregation induced by γ. The IC ₅₀ value was 11.02 ng / ml when rabbit platelets were used and 2.06 ng / ml when human platelets were used.

Reference example 4 Exoenone 6 was synthesized from cyclooctadiene,
It was used as a mixture of α and β. 6 (435 mg, 0.72 mM) was dissolved in methanol (36 ml), and RhCl ₃ .3H ₂ O (0.1
(3 mol equivalent) and anhydrous K ₂ CO ₃ (0.72 mol equivalent) are added, and the mixture is stirred with heating under reflux for 2 hours. After confirming the disappearance of the raw materials by TLC (ether.n-hexane = 1: 2), the solvent is distilled off under reduced pressure. Rhodium was removed from the residue by alumina column chromatography (ether), and then silica gel column chromatography (ether: n-hexane =).
Endoenone 7 by separating and purifying in 1: 2)
Was obtained in a yield of 335 mg (77%). 7 ; Endoenone 7 (303 mg, 0.50 mM) was dissolved in THF (3 ml), and tetrabutylammonium fluoride was added to this at room temperature.
(2.0 eq.) Was added and stirred for 12 hours, the starting material disappeared on TLC (ether: methanol = 95: 5), and the 15α, 15β-isomer of the diol was observed as two spots.
Saturated NaCl water is added to this and extraction is performed with ether. The ether layer is dried over anhydrous MgSO ₄ , and the solvent is distilled off. When the obtained residue is separated and purified by silica gel column chromatography (ether: methanol = 95: 5),
180m total of 15α- and 15β-isomers of diol 8
g (95%) was obtained. 8 ; Reference example 5 15α-diol 8 (180mg, 0.46mM) methylene chloride (3m
l) Add dihydropyran to the solution at room temperature. When a catalytic amount of p-toluenesulfonic acid was further added, the reaction instantaneously proceeded, and disappearance of the raw material was observed on TLC (ether: n-hexane = 2: 1). Saturated NH ₄ Cl water is added to the reaction solution, and extraction is performed with ether. The ether layer is washed with saturated aqueous NaCl and then dried over anhydrous MgSO ₄ . After the solvent was distilled off, the residue was purified by silica gel column chromatography (ether: n-hexane = 1: 1) to obtain 241 mg (96%) of enone 9 .

Reference example 6 To a solution of enone 9 (229 mg, 0.42 mM) in HMPA (2 ml) was added hexamethyldisilazane (1.5 eq) at room temperature, and tetrabutylammonium fluoride was added in 0.1 molar equivalents each.
0.3 molar equivalent of TLC (ether: n-hexane = 2:
Add while observing the progress of the reaction in 1). When the disappearance of the raw materials was confirmed (after 1 hour), the reaction solution was added with ether (5
ml) and cool in an ice bath. Water (1 ml) is added little by little to this to terminate the reaction. After removing the water layer, the organic layer is washed several times with a small amount of water to remove HMPA. This was dried over anhydrous MgSO ₄ , the solvent was distilled off, and the resulting residue was separated and purified by silica gel column chromatography (ether green n-hexane = 1: 1) to give 214 mg (82%) of ketone 10. Was obtained. 10 ;

Claims (3)

[Claims] 1. The following formula [I]: With prostacyclin represented by as an active ingredient,
A pharmaceutical composition for lowering blood pressure or suppressing platelet aggregation, which is contained together with a pharmaceutically acceptable carrier. 2. A pharmaceutical composition according to claim 1 for vasodilation, hypotension or antithrombosis. 3. For anti-atherosclerosis, anti-angina, anti-myocardial infarction, anti-pulmonary arterial hypertension, anti-stroke, anti-transient Ischemic Attack, anti-deep vein thrombosis or anti-peripheral vascular disease. The pharmaceutical composition according to claim 1 or 2.