JP2003267868A - Prevention and treatment of malaria infection using hydroxy fatty acid analog as active ingredient - Google Patents

Abstract

(57) [Summary] An agent for preventing and treating malaria infection, which comprises a hydroxy fatty acid analog. PROBLEM TO BE SOLVED: To develop a drug that can exhibit high efficacy against multidrug-resistant malaria and has low toxicity. SOLUTION: An antimalarial drug was developed by using a hydroxy fatty acid analog as an active ingredient.

Description

DETAILED DESCRIPTION OF THE INVENTION [0001] TECHNICAL FIELD The present invention relates to malaria parasites
Of antimalarial agent useful for prevention and treatment of infectious diseases caused by
It is about. [0002] BACKGROUND OF THE INVENTION Caused by malaria parasite infection
Malaria is the most threatening threat to humans
And according to World Health Organization (WHO) statistics,
An estimated 300-500 million people are infected each year by malaria in the world,
The death toll has risen to 2.7 million. 40% of the world's population flows
Living in the region where the global warming
There is a concern that the number of dying people will increase and the area of the endemic area will expand. Further
Malaria is a businessman from a developed country, tourism travel
And immigrants at risk, and
Imports (imported malaria patients) are not available in Europe and North America.
Increasingly in endemic areas. [0003] Plasmodium is a causative pathogen of malaria.
(Plasmodium), for example, tropical fever
Plasmodium (P. falciparum), Plasmodium vivax
(P. vivax), Plasmodium ovale (P. ovale), and the world
Protozoa such as P. malariae (P. malariae)
It infects humans through the cod mosquito. [0004] Conventionally, malaria has been diagnosed and treated properly.
It was said that the disease could be completely cured if it was treated,
In recent years, multi-drug resistant protozoa have been introduced in the Middle East, Indonesia and Malaysia
In the Philippines, the Philippines, and Oceania
Currently available drugs such as chloroquine, prima
Kin, mefloquine, doxycycline, artemisini
And pyrimesamine make treatment more difficult
are doing. [0005] [Problems to be Solved by the Invention]
Only quinine was effective, but the effectiveness was reduced
Has also been reported in some countries in Southeast Asia and Brazil
You. In addition, conventional malaria treatments include renal failure, stomach
Highly likely to cause side effects such as bowel symptoms and dermatosis
No. Therefore, high efficacy against multi-drug resistant malaria
There is a need for a drug that can be demonstrated and has low toxicity. [0006] SUMMARY OF THE INVENTION Hydroxymyristic acid
Leucine (HML) originates from marine Vibrio spp.
New substance found, manufactured based on HML and HML
Heterocapsa circular
Effective against dinoflagellate such as isquama, Prorocentrum micans
It is clear that there is (Japanese Patent Application 2001-83617). By the way, recent molecular phylogenetic analysis (Gajadh
According to ar et al. 1991), dinoflagellates and ciliates,
And apicomplexans (parasitic communities, malari
(Including Plasmodium of Protozoa) is a single line
It has been suggested. These are located just below the cell membrane.
In that they have vesicles called
Albeollata, a group of higher-order classifications
It has been proposed to treat as a group. [0008] In view of such a background, the inventor of the present invention has proposed hydroxy
Anti-Mas against Plasmodium sp.
When the Laria parasite activity was examined, hydroxy fatty acids
Found that the analog has an effective antimalarial action
did. That is, according to the present invention, a hydroxy fatty acid derivative
Body and their hydrates or solvates as active ingredients
An antimalarial agent is provided. Preferred embodiments of the present invention
According to the above, the hydroxy fatty acid analog is represented by the following general formula (1) [0009] Embedded image (In the above general formula (1), n is an integer of 5 to 17, and X is NR
Or O and R is H and is, for example, methyl, ethi
And an alkyl group such as n-propyl. R₁, R _TwoIs replaced
Represents a group. The hydroxy fatty acid analog represented by
It is extremely useful as an antimalarial agent having the characteristics of
And completed the present invention. In the above formula (1), R₁, R_TwoAs various
The substituent is preferably, for example, an alkyl group
(Eg, methyl, ethyl, n-propyl, i-propyl, n-
Butyl, sec-butyl, tert-butyl, pentyl, hex
Phenacyl, pyrimidinylalkyl group, amino
Carbonylamino group, guanodylpropyl group, carboxyl
A silalkyl group and an esterified carboxyalkyl
(For example, carboxymethyl, carboxyethyl,
Methoxycarbonylmethyl, ethoxycarbonylmethyl
, Glucuronide carbonyl ethyl), imidazo
And a pyrimidine group. Also, R₁, R_TwoTomo water
It shall include those of elementary atoms. [0011] The above-mentioned substituent further comprises an ester
And / or amidated, and / or
Group, alkoxyl group, carbonyl group, phenyl group, etc.
It may be further substituted. For example, alkylpyri
The 4- and 6-positions of the midine can be further substituted. Note that R₁And R are combined to form a 5-membered ring or 6
It may form a member ring. Several compounds can be represented by the above formula
Asymmetric carbon, and there are many isomers
In the present invention, these isomers are included in the range.
Shall be. The compound represented by the above formula is in free form
Used in any hydrate or solvate form
You. The fatty acid portion of the compound represented by the above formula is
It is an acid component, and the number of carbon atoms is a general fatty acid n = 5
It is easy to predict that the same activity is observed in ~ 17
You. The compound represented by the above general formula (1) is as follows:
It can be manufactured by the method described above. Embedded image [Where n, X, R₁And R_TwoIs the same as above. X₁Is
NHR or OH, where R is H and
For example, an alkyl group such as methyl, ethyl, n-propyl, etc.
You. Y_Two, Y_FourIs a protecting group for the carboxylic acid, and the protecting group is particularly
Benzyl ester, tertiary butyl ester
, Methyl ester, ethyl ester, phenacyles
Ter, trichloroester, paranitrobenzyl ester
, Diphenyl methyl ester, benzhydryl esthetic
And paramethoxybenzyl ester. Ma
If it is not necessary to protect, it is a hydrogen atom. Y_ThreeIs hydroxyl
T- is a protecting group for the group, and the protecting group is not particularly limited.
Butyl, benzyl, acetyl, methyl, tetra
Hydropyranyl group, cyclohexyl group, etc.
You. If it is not necessary to protect, a hydrogen atom is used. ] Each compound represented by the above formula has several asymmetric carbons.
Having a large number of isomers,
Hydroxy fatty acid analogs and compounds used in the production process
Compounds (2) and (3) include these isomers
You. It is desirable to prepare an optically active substance. In this reaction step, the known compound represented by formula (2)
A condensing agent in an organic solvent having no suitable active hydrogen atom,
Known compound (3) with acidic additives and 20 ° C to 50 ° C, 10
Reaction for up to 24 hours and deprotection by an appropriate method.
To obtain the hydroxy fatty acid analog obtained in (1). As an organic solvent having no active hydrogen atom,
Chloroform, dichloromethane, dichloroethane, tol
Ene, tetrahydrofuran, dimethylformamide, di
Methyl sulfoxide and the like can be mentioned. With condensing agent
As an example, dicyclohexylcarbodiimide, 1-
Ethyl 3-dimethylaminopropyl carbodiimide
Can be Although the acidic additive is not essential, the additive
In order to prevent acyl urea by-product and racemization
Hydroxybenzotriazole,
N-hydroxysuccinimide, 3,4-dihydro-3-hydro
For example, xy-4-oxo-1,2,3-benzotriazine
Can be In the reaction, compound (2)
(3), the condensing agent and the additive are used in substantially equal amounts. Also,
Protection depends on the nature of each protecting group, depending on the other chemical moiety.
Use mild conditions that do not change the position. Got
Compounds can be prepared by conventional separation means, e.g., column chromatography.
Easy isolation and purification from reaction mixture by fee, recrystallization, etc.
can do. [0018] BEST MODE FOR CARRYING OUT THE INVENTION The medicament of the present invention is used for preventing malaria.
And / or can be used for therapy. Malaria Causes
The pathogen is not particularly limited, and Plasmodium falciparum,
Malaria parasite, oval malaria parasite, vivax malaria parasite
Malaria caused by protozoa such as
Is applicable. As the medicine of the present invention, the above formula
Compounds represented by (I), hydrates thereof, and solvates thereof
Substances selected from the group consisting of
No. As the HMLs used in the present invention, commercially available HMLs
Extracted and purified bacteria and chemistry
Any of the synthetic ones can be used,
Usually contains the above-mentioned substance as an active ingredient and a pharmaceutical additive
Desirably, it is prepared as a pharmaceutical composition. Of the present invention
The route of administration of the drug is not particularly limited, and is orally or parenterally.
Can be administered. The form of the medicament of the present invention is not particularly limited.
It is possible to select an appropriate form depending on the route given,
Manufacture according to the general method in the industry according to its form
be able to. For example, tablets, capsules, soft capsules
Preparations, granules, powders, fine granules, syrups, solutions, suspensions
Or a pharmaceutical composition for oral administration, such as subcutaneous, intramuscular,
Or intravenous injections, drops, suppositories, etc.
Can be prepared as a pharmaceutical composition. The preparations used in the production of these pharmaceutical compositions
The type of additive for the agent is not particularly limited and can be used in the industry.
One or two of these depending on the form of the pharmaceutical composition
More than one species can be selected as appropriate. For example, injection
The preparation of the diluent is carried out using diluents (for example,
Brine, glucose injection, lactose injection, mannitol injection
Etc.), dissolve the above substances as active ingredients, sterilize by filtration, etc.
Appropriate sterilization treatment and filling in sealed containers such as ampules
do it. Also, freeze-dried form based on Japanese Pharmacopoeia
Injection and powdered injection mixed with sodium chloride.
It may be made. In addition, as a pharmaceutical additive, for example, po
Polyethylene glycol, HCO-60 (surfactant; Nikko Chemi
(Manufactured by Cal Corporation), ethanol and / or lipo
Even if it contains carriers such as chromosomes and cyclodextrins
Good. Pharmaceutical compositions suitable for oral administration or rectal administration
Preparation of a suitable pharmaceutical composition comprises excipients, disintegrants,
Agents, lubricants, suspending agents, isotonic agents, emulsifiers, etc.
Mix and mold the excipients for pharmaceuticals and the above substances by the usual method.
And can be manufactured. As excipient used
A cellulose derivative (crystalline cellulose, methylcell
Loin, hydroxypropylcellulose, hydroxyp
Propyl methylcellulose), polyvinylpyrrolidone, de
Kisstrin, starch, lactose, mannitol, sorbitol
Oil, vegetable oils (eg, corn oil, cottonseed oil, cocoa
Oil, almond oil, olive oil, peanut oil, etc.),
Oil esters such as medium-chain fatty acid glyceride oil, mineral oil,
Glycerin esters such as licaprylin and triacetin
, Alcohol such as ethanol, physiological saline,
Ren glycol, polyethylene glycol, animal fats and oils,
Vaseline and the like can be mentioned. Each of these formulations
They can be produced by conventional production methods known to those skilled in the art. The dose and frequency of administration of the medicament of the present invention are particularly
Not limited, therapeutic or prophylactic purpose, route of administration, patient year
Selectable according to conditions such as age, weight, and severity of disease
It is noh, but generally, 0.1 to 1,
About 000 mg (weight of active ingredient), preferably 1 to 60
0 mg, administered once or twice a day
Is preferred. The medicament of the present invention is used for malaria.
May be used in combination with other drugs used for treatment or prevention,
Compositions (such as "iwawa") in combination with the active ingredients of such medicaments
May be used. [0023] EXAMPLES Hereinafter, the present invention will be described more specifically with reference to examples.
However, the scope of the present invention is not limited to the following examples.
And not. <Culture and assay of Plasmodium falciparum> Evaluation method and
The method of Murakami et al. Was used.¹⁾Tropical as malaria parasite
P. falciparu FCR-3 strain (cancer
Biaitol cycloguanyl-resistant strain) and treated with sorbitol
After synchronization, the hematocrit value (red
Ratio of red blood cell volume in blood cell suspension) 2%, infection
Transfer 50 μL of culture solution with a ratio of 0.55% to a 96-well microplate
Inoculated. The test sample is dissolved in DMSO and cultured to an appropriate concentration.
After dilution with a nutrient solution, 50 μL thereof was added. (DMSO medium
Concentration is 1%. ) After culturing at 37 ° C for 48 hours,
P. The growth of falciparum is thin-layer smear by Giemsa staining
It was determined by measuring 10,000 red blood cells per book. Anti-ma
Quinine was used as an indicator as a laria drug. Table is individualized
It represents the compound IC50. 1) N. Murakami, H. M. Mostaqul, S. Tamura, S. Ita
gaki, T. Horii and M. Kobayashi, Bioorganic & Medic
inal Chemistry Letters 11 (2001), 2445-2447. Of the compounds represented by the general formula (I),
As examples, five types represented by the following chemical structural formulas are used.
Laria parasite growth inhibition screening test results
Was. The method for producing these five compounds is described in Japanese Patent Application No. 2001-8361.
No. 7 and the application as the original application dated March 15, 2002
Application for priority claim filed in Japan). [Compound 1] 3 (S) -hydroxymyris
(L) formic acid Embedded image [Compound 2] 3 (S) -hydroxylauri
Luic acid (L) leucine Embedded image [Compound 3] 3 (S) -hydroxypalmi
(L) formic acid Embedded image [Compound 4] 3 (S) -hydroxymyris
Formic acid (L) glutamic acid Embedded image[Compound 5] 3 (S) -hydroxymyris
Arginine formic acid (L) Embedded image <Production Example 1> 3 (S) -hydroxymyris
Synthesis of formic acid (L) -leucine (compound 1) Embedded image 1) Production of L-leucine-benzyl ester-tosylate L-leucine (3.3g), tosylic acid (5.7g), benzyl alcohol
(15 ml) and toluene (30 ml) into a round bottom flask.
Oil bath (1) using a Dean-Stark device equipped with a reflux condenser.
(10-130 ° C) for 6 hours. Cool the round bottom flask
And add ether (50 ml) and petroleum ether (50 ml)
Crystallize, filter, hot ethanol (30ml) -ether (30ml)
Further recrystallization gave the desired product in a yield of 8.4 g (85%). 2) Synthesis of 3-hydroxymyristic acid a) Production of 3-oxomyristic acid ethyl ester 2,2-dimethyl-1,3-dioxane-4,6-dione (2.26 g)
Dissolve in methylene chloride (30 ml), cool to 0 ° C and stir.
You. Add anhydrous pyridine (2.7 ml) and n-dodecanoyl chloride
(3.6 ml) and continue at 0 ° C for 1 hour,
While stirring. The reaction mixture was poured into ice water and 2N-HCl (30 ml) was added.
And then extract twice with 30 ml of dichloromethane.
You. The dichloromethane extraction part further includes 2N-HCl (3
0 ml), washed with saturated saline and dried over sodium sulfate.
Thereafter, the solvent is distilled off under reduced pressure. Ethanol obtained
Dissolve in 30 ml of ethanol and heat to reflux for 3 hours. Reaction volume reduced
After evaporating the solvent under pressure, n-hexane-ethyl acetate (5: 1)
To give 2.8 g of product (80% yield). B) 3- (S) -hydroxymyristic acid
Production of ethyl ester (S) -BINAP (8mg) and bis (methylallyl) cycloocta-
Put 1,5 diene-ruthenium in a 50 ml shrink tube and depressurize
Replace with an argon stream and fill. 2 ml of acetone and
Add aqueous hydrogen bromide (0.11 ml of 0.29M) and stir at room temperature for 30 minutes.
Mix. After distilling off the solvent under reduced pressure, the residue
Add ethyl ristate (1 g) and put on an oil bath at 87 ° C.
Heat and stir for 72 hours. The solvent is distilled off from the reaction solution under reduced pressure.
To dryness, then purified with n-hexane-ethyl acetate (10: 1)
This gives 0.73 g (yield 73%) of the product. C) 3- (S) -hydroxymyristic acid
Manufacturing of 3- (S) -hydroxymyristic acid ethyl ester (0.5
g), add 1.1N NaOH (2 ml) and THF (5.0 ml) and stir for 20 minutes
I do. THF was distilled off under reduced pressure from the reaction solution, and 10%
After adjusting the pH to 3 by adding an acid, extract twice with ether (20 ml).
Take out, dry with magnesium sulfate, 0.41 g of the desired product
(93% yield). 3) Leucine and hydroxymyristic acid
Condensation and purification L-leucine-benzyl ester-tosylate and (S) -3-H
Droxymyristate and 1.1 equivalents of them
Dissolve hydroxybenzotriazole in DMF and bring to -20 ° C
While cooling, 1.1 equivalents of 1-ethyl 3-dimethylaminopropyl
Lecarbodiimide is added dropwise. Remove the coolant tank after dropping
The mixture is further stirred overnight to complete the reaction. Vinegar reaction
Dissolved in ethyl acetate, and the solution was diluted with 1N HCl, water, 5% aqueous sodium bicarbonate,
After washing with water, it is dried over anhydrous magnesium sulfate. solvent
, And dissolved in methanol, and after adding carbon black, water
Stir overnight in a stream of air. The reaction solution was filtered to remove carbon black, and the solvent was removed.
Is distilled off. Crude product is 80% acetonitrile-0.1% ODS
Reversed-phase high-performance liquid chromatography using
Ram: Shiseido Capsule Pack C18SG120, 15mm x 250mm)
The title compound 1 was obtained. Chemical Properties of Compound 1 of the Invention Material color: colorless Molecular weight: 357 Molecular formula: C₂₀H₃₉NO_Four <Production Example 2> 3- (S) -hydroxylauric acid (L) -roy
Synthesis of syn (compound 2) Embedded image 1) L-leucine-benzyl ester-toshi
Synthesis of Lurate This compound was synthesized by the method described in Production Example 1. 2) Synthesis of 3- (S) -hydroxylauric acid
Success a) Production of 3-oxolauric acid ethyl ester 2,2-dimethyl-1,3-dioxane-4,6-dione (2.26 g)
Dissolve in methylene chloride (30 ml), cool to 0 ° C and stir.
You. To this was added anhydrous pyridine (2.7 ml) and n-decanoyl chloride (3.
(0 ml) and stirred at 0 ° C for 1 hour, followed by stirring at room temperature for 1 hour.
Mix. The reaction solution was poured into ice water and 2N-HCl (30 ml) was added.
After acidification, extract twice with 30 ml of dichloromethane. The
The chloromethane extraction section further contains 2N-HCl (30 ml),
After washing with brine and drying over sodium sulfate, dissolve under reduced pressure.
The medium is distilled off. Dissolve the obtained oil in 30 ml of ethanol.
Dissolve and heat to reflux for 3 hours. The reaction solution is distilled under reduced pressure.
After purification, purified by n-hexane-ethyl acetate (5: 1)
2.7 g (80% yield) of the product are obtained. B) 3- (S) -hydroxylauric acid ethyl
Production of luster (S) -BINAP (8mg) and bis (methylallyl) cycloocta-
Put 1,5 diene-ruthenium in a 50 ml shrink tube and depressurize
Replace with an argon stream and fill. 2 ml of acetone and
Add aqueous hydrogen bromide (0.11 ml of 0.29M) and stir at room temperature for 30 minutes.
Mix. After distilling off the solvent under reduced pressure, 3-oxola
Add uric acid ethyl ester (1 g) and put on an oil bath at 87 ° C.
Heat and stir for hours. The solvent is distilled off from the reaction solution under reduced pressure.
After drying, the residue was purified with n-hexane-ethyl acetate (10: 1).
This gives 0.68 g (68% yield) of the product. C) Production of 3- (S) -hydroxylauric acid
Construction 3- (S) -hydroxymyristic acid ethyl ester (0.5
g), add 1.1N NaOH (2 ml) and THF (5.0 ml) and stir for 20 minutes
I do. THF was distilled off under reduced pressure from the reaction solution, and 10%
After adjusting the pH to 3 by adding an acid, extract twice with ether (20 ml).
Take out, dry with magnesium sulfate, 0.40 g of the desired product
(91% yield). 3) Condensation reaction and purification L-leucine-benzyl ester-tosylate and 3- (s) -H
Droxylauric acid and 1.1 equivalents of
Dissolve droxybenzotriazole in DMF and cool to -20 ° C
1.1 equivalents of 1-ethyl 3-dimethylaminopropyl
Carbodiimide is added dropwise. Remove the coolant tank after dropping
The mixture is further stirred overnight to complete the reaction. Vinegar reaction
Dissolved in ethyl acetate, and the solution was diluted with 1N HCl, water, 5% aqueous sodium bicarbonate,
After washing with water, it is dried over anhydrous magnesium sulfate. solvent
, And dissolved in methanol, and after adding carbon black, water
Stir overnight in a stream of air. The reaction solution is filtered to remove carbon
The solvent is distilled off except for black. 80% acetonitrile
Reversed-phase high-performance liquid chromatography using 0.1% ODS as eluent
Raffy (Column: Shiseido Capsule Pack C18SG120,15mm
× 250 mm) to give the title compound 2. Chemical Properties of Compound 2 of the Invention (1) Color of substance: colorless (2) Molecular weight: 329 (3) Molecular formula: C₁₈H₃₅NO_Four <Production Example 3> 3- (S) -hydroxymyristin
Synthesis of acid (L) -arginine (compound 5) Embedded image 1) N^g-NO_TwoOf L-arginine L-arginine hydrochloride (1.0 g) in concentrated sulfuric acid solution (10 ml)
Add ammonium acid (2.0 mg) and stir. ether
(50 ml) and petroleum ether (50 ml) for crystallization,
Filtration and recrystallization from hot ethanol (30 ml) -ether (30 ml)
The target product was obtained in a yield of 0.83 g (65%). 2) 3- (S) -hydroxymyristic acid
Synthesis It was synthesized by the method described in Production Example 1. 3) Condensation reaction and purification N^g-NO_Two-L-arginine and 3- (S) -hydroxymyristin
Acids and 1.1 equivalents of hydroxybenzoto
Lyazole is dissolved in DMF and cooled to -20 ° C while 1.1 equivalents
1-ethyl 3-dimethylaminopropyl carbodiimide is dropped
Down. After dropping, remove the refrigerant tank and stir overnight.
To complete the reaction. Dissolve the reaction in ethyl acetate and add
The solution was washed with 1N HCl, water, 5% aqueous sodium bicarbonate, water, and then sulfuric anhydride.
Dry with magnesium. The solvent is distilled off and methanol
And add carbon black and stir overnight under a stream of hydrogen.
You. The reaction solution was filtered to remove the carbon black and remove the solvent.
Is distilled off. Crude product is 80% acetonitrile-0.1% ODS
Reversed-phase high-performance liquid chromatography using
Ram: Shiseido Capsule Pack C18SG120, 15mm x 250mm)
Scrutiny provided the title compound 5. Chemical properties of compound 5 of the present invention (1) Color of substance: colorless (2) Molecular weight: 400 (3) Molecular formula: C₂₀H₄₀N_FourO_Four Compound 3-4 can be produced in the same manner.
You. [0050] [Table 1][0051] EFFECT OF THE INVENTION Formula (I) which is an active ingredient of the medicament of the present invention.
Is a compound that exhibits antimalarial activity at low concentrations,
Antimalarials containing it can prevent malaria infection or
Is useful as a therapeutic.

   ────────────────────────────────────────────────── ─── Continuation of front page    (72) Inventor Hirohisa Murakami             Osaka University Pharmacy 1-6 Yamadaoka, Suita-shi, Osaka             Department of Natural Product Chemistry (72) Inventor Kazuma Kobayashi             Osaka University Pharmacy 1-6 Yamadaoka, Suita-shi, Osaka             Department of Natural Product Chemistry F term (reference) 4C206 AA01 AA02 GA04 GA25 HA31                       MA01 MA04 NA14 ZB38

Claims (1)

Claims: 1. An agent for preventing and treating malaria infection, comprising a hydroxy fatty acid analog represented by the following general formula (1). General formula (1) (In the above general formula (1), n is an integer of 5 to 17, and X is NR
Or 0, wherein R is H and an alkyl group such as, for example, methyl, ethyl, n-propyl, and the like. R ₁ , R ₂
Represents a substituent. )