JPH07504421A - Cyclic ketal derivative - Google Patents

Abstract

(57) [Summary] This bulletin contains application data before electronic filing, so abstract data is not recorded.

Description

[Detailed description of the invention] Cyclic ketal derivative The present invention has blood cholesterol lowering, blood lipid lowering and/or antifungal activity. novel compounds, their production methods, pharmaceutical compositions containing them, medical and special for the treatment and/or prevention of atherosclerosis and related cardiovascular diseases. regarding their uses. The present invention reduces blood cholesterol, blood lipids, and and/or as intermediates for the production of compounds with antifungal activity. It also relates to compounds.

High levels of blood bocholesterol and blood lipids are associated with the development of vascular wall diseases. Ru. Therefore, there is great interest in effective methods for reducing plasma cholesterol levels. Leaning back. Cholesterol concentration is reduced by reducing dietary intake of sterols, e.g. by increasing its metabolism and excretion, or decreasing its rate of biosynthesis. can be reduced. Most effective in lowering physiological cholesterol levels Effective approaches include - as a component, inhibition of cholesterol biosynthesis. it is Cholesterol synthesis undergoes feedback regulation resulting in cholesterol levels This is because the decrease in biosynthesis is compensated for by an increase in biosynthesis.

One rate-limiting step in cholesterol biosynthesis is 3-hydroxy-3-methyl Formation of mevalonic acid from glutaryl coenzyme A (HMGCoA), resulting in high cholesterol The mebic acid group of HMG CoA reductase inhibitors in the treatment of telolemia Clinical success was thus achieved. However, mevalonate is a supplement in animals. Enzyme Q1 is a co-factor of all imprenyl derivatives, including heme A and dolichols. It is a common precursor.

The first biosynthetic step leading exclusively to sterols, i.e. giving squalene2 The condensation of two farnesyl diphosphates is the second regulatory point. farnesirni In the synthesis of squalene from phosphoric acid, there is an isolated intermediate, namely presqualene. phosphoric acid, and the total synthetic pathway involves the membrane-bound enzyme squalene synthase (farne synthase). Cylniphosphate: Fufurnesylniphosphate farnesyltransferase.

EC2,5,1,21>. Therefore, squalene synthase Substances that act to inhibit enzymes are potential drugs for regulating cholesterol formation It is a thing. The use of such substances suggests that non-steroidal pathways are minimally affected. It is attractive because it is not.

The biosynthesis of ergosterol, a major sterol component of fungal cell membranes, is important in humans, including humans. This is similar to the case for cholesterol in mammals, and therefore squalene mediated by the intase enzyme. Therefore, squalene synthase in fungal cells Substances that act to inhibit enzymes are drugs that can be used for antifungal chemotherapy. Ru.

We act as inhibitors of the squalene synthase enzyme and/or A new intermediate for the production of compounds that act as inhibitors of quarene synthase enzymes. We have now discovered a group of novel compounds.

Therefore, in the first aspect of the invention we provide compounds of the general formula (+): [In the above formula, R1 is a hydroxyl group, or -ococH-CHCH(CH3) (CH2)3CH3, -ocE　E OCH-CHC(CH3)-CHCH(CH3)CH2CH3 Also bar OCO -X -CH2CH(CH3)CHCH3 (where X is -CH-CHCH(C H3)-1-CH2CH(OH)CH(CH3)-1-CH-CHC(OR)( CH)-1-CH2CH(OH)CH- or -CHCH2CH(CH3)- represents a group selected from; R2 represents a hydroxyl group; R3 is (Here, R8 is a hydrogen atom or an acetyl group), -C(CH)-CHC) l　(CHR)CH2Ph (where R9 is hydrogen or a hydroxyl group) , -C(CH20H)-ZCHCI((CH3)CH2Ph, -C(-CH)C H(OH)CH(CH20H)CH2Ph, -C(-CH)CH(NHCOCH 3) CH(CH)CHPh, -C(CHNHCOCH3)-CHC)I (C H3) represents a group selected from CH2Ph and; R and R5 each independently represent a hydrogen atom or a methyl group; R6 and R7 each independently represent a hydrogen atom or a C alkyl group; or R6 is the hydrogen atom number, and R7 is the group -C(-Y)NH (where Y is oxygen or is a sulfur atom or NH] and salts thereof.

The compound of formula (1) in which R4 and R5 represent a hydrogen atom is physiologically acceptable. Cations are usually preferred.

R1 is preferably the following group: R3 is preferably the following group: (in the formula, R8 is a hydrogen atom or an acetyl group).

The invention should be understood to encompass any combination of the above selections.

Particularly preferred compounds of formula (+) are (IS-C1α (4R*, 5S), 3α, 4β , 5α, 6α (2E, 4H books.

* * 6R), 7β)) 1-(4-acetyloxy-5-methyl-3-methylene-6- phenylhexyl)-3-(aminomethyl)-4,6,7-drihydroxy-2 ゜8-dioxabicyclo[3,2,11 octane-4,5-dicarboxylic acid, 6- (4,6-dimethyl-2-octenoate) and its salts and protected derivatives .

The compound of the formula in which R1 represents a hydroxyl group has squalene synthase inhibitory activity. It is particularly useful as an intermediate for the production of related structures having

The compounds of the invention may form salts with inorganic and organic acids and bases. Menstruation Scientifically acceptable base salts include alkali metal salts (e.g., disodium and niconica). sodium and potassium salts, including sodium salts), alkaline earth metal salts (e.g. , calcium salts) and inorganic base salts such as ammonium salts. proper organic Base salts include trialkylamines (e.g., triethylamine), dialkylamines, (e.g. dicyclohexylamine), optionally substituted benzylamine (e.g. p-bromobenzylamine), tris(hydroxymethyl)methyl Amine salts and amino acid salts (e.g., lysine and amino acid salts, including di-L-lysine salts) There are amine salts such as luginine salt). Examples of physiologically acceptable acid addition salts include: hydrochloride, hydrobromide, sulfate, alkyl or aryl sulfonate (e.g. , methanesulfonate or p-toluenesulfonate), phosphate, acetate, of citrate, succinate, lactate, tartrate, fumarate and maleate. There are salts derived from organic or inorganic acids such as

Other salts which are not physiologically acceptable may also be useful in the preparation of compounds of formula (1), form another aspect of the invention.

Compounds of the invention inhibit squalene synthase enzyme and cholesterol biosynthesis Therefore, medically, especially in animals (particularly humans), plasma concentration is important. It is useful for a variety of conditions where lowering cholesterol levels would be beneficial. like this Examples of symptoms include those associated with hypercholesterolemia and hyperpoproteinemia. diseases, especially atherosclerosis and cardiovascular diseases (e.g. cardiac ischemic diseases, cerebral ischemic disease and peripheral arterial disease).

Compounds of the present invention that inhibit squalene synthase can be used in animals including humans. It may also be useful in treating fungal infections. For example, they are eg Candid a (for example, Candlda alblcans, candlda glabr ata, Candida parapsilosis and Candjda p. seudotrop), Cryptococcus neor.

rmans, Aspergillus Sp, (for example, Asperg. lus rlavus and Aspergillus fuIIligatus ), Coccidiojdes (e.g. Coccidioides 1Ii tis), Paracoccjdioides (e.g. Paracocc idioides brasiliensjs), Histoplasm direct analogy For example, Hlstoplasma capsulatum) or Blastomy ces (for example, BlastoIl!yces dera+atiNdls) It is useful in treating systemic infections caused by They are Trichol) hytons Old crosporuw or Epidermophyton (e.g. Tri chophyton II lentogral) hytes, Micros poruIIlcants or Epidermophyton rl.

It is also used in the treatment of local infections caused by species of C. ccosum. those Also Torulops glabrata and PItyrosporu ■It is also useful in treating fungal diseases caused by S. ovale.

In vitro evaluation of the antifungal activity of the compounds of the present invention does not result in the growth of specific microorganisms. Determine the minimum inhibitory concentration (MIC), which is the concentration of the test compound in the appropriate medium. This can be done by

Inhibiting squalene synthase in view of their potential in antifungal therapy The compounds of the invention are recommended for the treatment of various fungal infections in humans and animals.

Such infections include candidiasis and chronic mucosal candidiasis (e.g. oral and Fungal infections such as vaginal candidiasis), skin infections caused by fungi, skin and mucosal skin Skin candidiasis, dermatophytosis including albinism and ring infections, athlete's foot, paronychia, and dermatitis. Wind, erythema, intertrigo, fungal fluffy rash, candida vulvitis, candida balanitis and I have otitis externa.

They may also be useful as prophylactic agents to prevent systemic and local fungal infections. prevention Use as a selective intestinal decontamination method, e.g. in the prevention of infection in immunocompromised patients. Suitable as part of. Prevention of fungal overgrowth during antibiotic treatment may also be helpful in some medical conditions. or may be desirable in iatrogenic conditions.

Ability of Compounds of the Invention to Inhibit Squalene Synthase Enzyme in Mammals and Fungi Force is R, M, Talt, Analyt, Biochem, 203, 310-3 16 (1992) as a substrate under assay conditions similar to those described in 2C] farnesyl birophosphate in vitro.

For therapeutic purposes, compounds of the invention that inhibit the enzyme squalene synthase may be administered directly. preferably as an active ingredient in a pharmaceutical formulation. . Therefore, according to the invention, the compounds of the invention that inhibit the squalene synthase enzyme the compound with one or more pharmaceutically acceptable carriers and optionally other treatments and/or Also provided are pharmaceutical formulations comprising a prophylactic or prophylactic ingredient. carrier is prescription ``acceptable'' in the sense of being compatible with the other ingredients of the substance and not harmful to its receptors. Must be.

The compositions of the invention include oral, buccal, parenteral, implantable, rectal, topical, ophthalmic or Forms prescribed for genitourinary administration or suitable for administration by inhalation or insufflation There are compositions that have

Tablets and capsules for oral administration may contain binders such as syrup, acacia, gelatin, etc. of latin, sorbitol, tragacanth, starch or polyvinylpyrrolidone. Slime: fillers such as lactose, sugar, microcrystalline cellulose, maize starch , calcium phosphate or sorbitol; lubricants such as magnesi stearate. disintegrants: umum, stearic acid, talc, polyethylene glycol or silica; For example potato starch or sodium starch glycolate, wetting agents, e.g. It may also contain conventional excipients such as sodium lauryl sulfate. Tablets are in our industry It may be coated according to a well-known method. Oral liquid preparations may be e.g. aqueous or oil-based. It may be in the form of a suspension, solution, emulsion, syrup or elixir; may be served as a dry product that is prepared with water or other suitable vehicle before use. stomach. Such liquid preparations may contain suspending agents such as sorbitol syrup, methyl cell loin, glucose/sugar syrup, gelatin, hydroxyethylcellulose, carbonate ruboxymethylcellulose, aluminum stearate gel or hydrogenated edible fats; emulsifiers such as lecithin, sorbitan monooleate or gum arabic: Non-aqueous vehicles (which may also include edible oils), such as almond oil, fractionated coconut oil , oily esters, propylene glycol or ethyl alcohol; preservatives, e.g. Also contains conventional additives such as methyl or propyl p-hydroxybenzoate You may do so. The composition may include conventional herbal medicines such as cocoa butter or other glycerides. It may also be formulated as a suppository containing a base.

For oral or vaginal administration, the compositions may take the form of conventionally formulated tablets or lozenges. It's okay.

Compositions according to the invention may be formulated for parenteral administration by injection or continuous infusion. stomach. Formulations for injection are presented in unit dosage form in ampoules or multi-use containers and are stored Agents may also be added. The compositions can be prepared as suspensions, solutions or solutions in oily or aqueous vehicles. It may be in the form of a emulsion and may contain suspending, stabilizing and/or dispersing agents. It may also contain prescription agents. On the other hand, the active ingredient must be prepared with a suitable vehicle, e.g. It may also be in powder form prepared in sterile, pyrogen-free water.

For administration by inhalation, the compositions according to the invention may be administered using a suitable propellant, such as dichlorodifluoride. lomethane, trichlorofluoromethane, dichlorotetrafluoroethane, dioxide from a pressurized pack or with a nebulizer under the use of carbon or other suitable gas It is preferably delivered in the form of an aerosol spray dispensing. Place of pressurized aerosol In such cases, the dosage unit may be determined by providing a valve to deliver a metered amount.

On the other hand, for inhalation administration, the composition according to the invention may be a dry powder composition, e.g. and in the form of a powder mix with a suitable powder base such as lactose or starch. You can. Powder compositions can be packaged in capsules or cartridges, such as gelatin. , or as a blister pack in which the powder is administered with the help of an inhaler or ventilator Can be provided in unit dosage form.

The composition may be, for example, a suppository containing a conventional herbal base, or, for example, a conventional PET. It may also take the form of a pessary containing a sari base.

Compositions include ointments, creams, gels, lotions, shampoos, powders (spray powders) pessaries, tampons, sprays, dips, aerosols, preparations (e.g. , eye, ear or nose preparations) or for topical administration in the form of a pour-on. May be prescribed. Ointments and creams may contain suitable thickening and/or gelling agents. In addition, they may be formulated with an aqueous or oily base, for example. Ointments for ocular administration are sterile Manufactured aseptically using ingredients. Boa-on may optionally contain prescription drugs, e.g. For example, they are formulated for veterinary use in oils containing organic solvents along with stabilizing and solubilizing agents.

Pessaries and tampons for vaginal insertion are prescribed using conventional techniques and are If so, it contains an effervescent vehicle. Such compositions may optionally contain fluticosteroids. It may also contain other active ingredients such as antibiotics, antibiotics, or anthelmintics.

Liquid formulations for intranasal delivery take the form of solutions or suspensions and may include tonicity adjusting agents, e.g. Sodium, dextrose or mannitol; preservatives such as benzal chloride Conium, thiomersal, phenylethyl alcohol; other prescription agents, e.g. It may also contain conventional excipients such as clouding, buffering, stabilizing and/or dispersing agents.

Transdermal administration facilitates absorption of the active compound through the skin and typically includes a backing film. Adhesive stick-on containing adhesive film, membrane and release liner ) This can be done by designing an appropriate system consisting of a base formulation enclosed within a batch.

Compositions according to the invention may be formulated as depot preparations.

Such long-acting formulations may be implanted (e.g., subcutaneously or intramuscularly) or intramuscularly. Administered by injection. Thus, for example, the compounds of the present invention may be combined with suitable polymers. or hydrophobic substances (e.g. as emulsions in acceptable oils) or ions. It is formulated with exchange resins or as poorly soluble derivatives, such as poorly soluble salts.

When the composition consists of dosage units, each The units are preferably from 0.001 to 1000B, advantageously from 0.01 to 400mg of activity. Contains sexual components. The daily dose as used for adult treatment depends, for example, on the route of administration and on the patient. Depending on the person's condition and the disease being treated, preferably from 0.001 to 50% of the active ingredient 001g, most preferably in the range of 0.01-2000mg, 1-2000mg per day. Administered 4 times.

The compound can be administered by intravenous infusion using, for example, 50 mg/kg of active ingredient within 7 days. more administered. Treatment duration is determined by response rate rather than an arbitrary number of days. It will be done.

Compounds of the invention that inhibit squalene synthase may be used in combination with other therapeutic agents. Therefore, the present invention utilizes compounds of the present invention that inhibit squalene synthase in other treatments. Activators, such as 3-hydroxy-3-methylglutaryl coenzyme A (HMG Co A) Inhibitors of reductase or drugs that reduce serum cholesterol and/or or other drugs that inhibit cholesterol biosynthesis, such as bile acid sequestrants, or Antihyperpoproteinemic or antihyperlipidemic agents, such as probucol, gemfi Brozil, clofibrate, dextrothyroxine or its sodium salt, Cholestyramine or its hydrochloride, cholestyramine, nicotinic acid, neomycin , p-aminosalicylic acid, aspirin, DEAE-Sephadex, poly(dia (lylmethylamine) derivatives, ionene or poly(diallyldimethylammonium) In another aspect, the combination is provided with chloride.

The above combinations are preferably put to use in the form of pharmaceutical formulations, so that Pharmaceutical formulations containing the combination together with a pharmaceutically acceptable carrier therefor may be forming another aspect of the invention. The individual components of such combinations may be used separately or in combination. Administered sequentially or simultaneously as a separate pharmaceutical formulation.

When a compound of the present invention is used in combination with a second therapeutic agent for a similar condition, each compound The dosage of the compound may be different than when the compound is used alone. appropriate use Amounts will be readily apparent to those skilled in the art.

According to another aspect of the invention we provide a compound of formula (+) or its physiologically acceptable salts that can be tolerated or for therapeutic use, especially plasma cholesterol in animals (especially humans) Treatment of conditions where lowering of drug levels is beneficial or true in animals (particularly humans) Compounds of formula (1) or physiologically acceptable compounds thereof as described above for the treatment of fungal infections The present invention provides a pharmaceutical composition containing a salt of the present invention.

In a specific aspect of the invention, we provide a compound of formula (1) or its physiologically acceptable or hypercholesterolemia and/or hyperpoproteinemia diseases related to heart disease, especially atherosclerosis and cardiovascular disease (e.g. heart failure). Formula (1) as above for the treatment of blood diseases, cerebral ischemic diseases and peripheral arterial diseases) or a physiologically acceptable salt thereof.

According to another aspect of the invention, we provide a method for treating hypercholesterolemia and/or high blood pressure. Diseases associated with proteinemia, especially atherosclerosis and cardiovascular diseases ( For example, 1. Manufacture of drugs for the treatment of cardiac ischemic disease, cerebral ischemic disease, and peripheral arterial disease. Provided is the use of a compound of formula (1) or a physiologically acceptable salt thereof for.

According to another aspect of the invention, we provide a method for treating fungal infections in human or non-human animal patients. Compounds of formula (1) or physiologically acceptable compounds thereof for the manufacture of medicaments for the treatment of infections Provides the use of salt.

According to another aspect of the invention, we provide a method for treating hypercholesterolemia and/or high blood pressure. Diseases associated with proteinemia, especially atherosclerosis and cardiovascular diseases ( For example, to treat cardiac ischemic disease, cerebral ischemic disease and peripheral arterial disease) or to treat fungal Provided is a method of treating a human or non-human animal individual for treating a disease; The method includes administering an effective amount of the compound of formula (1) or a physiologically acceptable salt thereof to the above individual. It consists of administering.

When we refer to treatment here, we refer to prevention as well as treatment of established symptoms or infections. It will be clear to those skilled in the art that this also applies.

The compounds of the present invention are produced by the following process.

A process for producing a metal compound of formula (1) in which R and R7 both represent a hydrogen atom (A) is a compound of the following formula (11) using conventional conditions: (In the above formula, R-R has the same meaning as above, and R4a and R5a are protecting groups) and then removing the protective groups present. The reduction is preferably , a water-containing ether solvent (e.g., tetrahydrocarbon) at a temperature in the range of 20-50 A compound of formula (11) is added to a trisubstituted phosphine such as triphenylphosphine in This is done by processing with a sphin.

The compound of formula (I+) is a compound of the following formula (II+): (in the above formula, R, R, R and and R5a have the same meanings as above 1 3 4a , R2a is a protected hydroxyl group, and L is an alkyl or arylsulfur group. A suitable deactivation group such as honyloxy group, e.g. trifluoromethanesulfonyloxy, is a leaving group) preferably in the presence of a solvent such as dimethylformamide. treatment with an azide source such as sodium azide at about room temperature followed by hydroxyl retention. Produced by removing the protecting group.

The compound of formula (II+) can be converted to the following formula (1v) under conventional conditions: a Manufactured from (in the above formula, R, R, R, Rt5, Yohi, R5a, H'J) be done. For this reason, for example, alkaline compounds such as trifluoromethanesulfonyloxy The reaction to introduce a kyl or arylsulfonyloxy group is preferably carried out using an organic base. (e.g. 2,4,6-collidine). of formula (1v) in a solvent such as chloromethane) at a temperature of about -10 to +20°C. treating the compound with a suitable anhydride such as trifluoromethanesulfonic anhydride. This is done by

A compound of formula (II+) in which R1 represents a hydroxyl group is produced from a compound of formula (1■) When it is desired to produce a compound of formula (IV), the 6-hydroxyl It may be necessary to protect the base. Appropriate protecting groups and deprotection methods are described below. do.

The compound of formula (!■) is preferably a compound of the following formula (■): (in the above formula, R%R , R, R and R5a are the same as above 1 2a 3 4a (synonymous with).

The reaction preferably involves activation of the 3-carboxyl group followed by dimethylformamide optionally in a solvent such as hydrogen or an ether (e.g. tetrahydrofuran). in the presence of water and at a suitable temperature, e.g. in the range of 0 to 50°C (e.g. around room temperature). with a suitable reducing agent such as borohydride (e.g. sodium borohydride). This is done through reduction.

Activation of the 3-carboxyl group can be performed, for example, by using an ether (e.g., tetrahydrofuran). carbodiimide in a suitable solvent such as C e.g. 1-cyclohexyl-3-(2-morpholinoethyl)carbodiimide Metho-p-)luenesulfonate or N,N''-dicyclohexyl carbodi by reaction with reagents such as N-hydroxysuccinimide in the presence of or a suitable solvent such as a halogenated hydrocarbon (e.g. dichloromethane) at temperatures ranging from 0 to 20 °C in the presence of a non-nucleophilic organic base such as triethylamine. Reaction with 2-chloro-3-ethylbenzoxacilium tetrafluoroborate conversion to active esters by reaction. On the other hand, activation is in dimethylform amide, preferably in the presence of a suitable solvent such as dichloromethane. ethers (e.g. tetrahydrofuran), nitriles (e.g. acetonitrile) or a mixture thereof, preferably at a temperature of about 0°C. It is carried out by reaction with oxalyl.

The compound of formula (■) is preferably prepared using a suitable base such as sodium hydroxide. By saponifying the 3-position carboxylic acid methyl ester, a compound of the following formula (■1): f /ISa/kl! To thing + +1. /, 711−→=kV/7’1) Afrfu+ 1-4=kK1 2a 3 4a (In the above formula, R, R, RSR Oyohi R"a has the same meaning as above) .

The compound of formula (■1) can be converted to the following formula (VI +) by standard protection/deprotection methods. Compound: (In the above formula, R-R3 has the same meaning as the above formula (+)).

Another process (B) for the production of the compound of formula (1) is the production of the compound of formula (1) It consists of converting a compound into a different compound of formula (1).

Therefore, in the embodiment of process (B), R6 and 7.

The compound of formula (1) in which at least one of the groups represents a C alkyl group is preferably In an organic solvent such as acetonitrile, when R and R both represent a hydrogen atom, I■, -C(-N!()NH-(L- is S(')-1(like eight(1) Reno 1 (, c + 1 sword I and 2 Riichi Lum I Lua mouth r Reno 5) l 5 I Non-T This C and borohydride (e.g., sodium borohydride, Sodium cyanoborohydride or triacetate hydride)・, -J-^Senichi 1- L　II　fy人) Manufactured by raising or processing in an appropriate amount of 1,000 people. .

In another embodiment of process (B), R6 represents a hydrogen atom and R is a group - Compounds of formula (1) representing C(-Y)NH2 introduce a group -C(-Y)NH2 Under conditions suitable as above, compounds of formula (1) in which R6 and R7 both represent a hydrogen atom Manufactured by processing things. Therefore, when Y represents an oxygen atom, the group - C(-Y) NH2 is dissolved in an ether solvent ( (e.g., tetrahydrofuran), preferably at elevated temperature (e.g., reflux) aminomethyl compound with a carbamate ester such as phenyl carbamate. It is introduced by reacting. When Y represents a sulfur atom, the group -C(-S ) NH2 preferably converts the aminomethyl compound of formula (1) into a thiocarbonyldiyl compound. Like midazole, thiophosgene or phenylcarbonyl isothiocyanate "thione" introduced by reacting with a donor and ammonia gas . When Y represents NH, the group -C(-N)()NH2 is in the presence of a base, in an alcoholic solvent (e.g. methanol), preferably at about room temperature Then, by reacting the aminomethyl compound of formula (1) with a suitable leaving group) will be introduced.

In another embodiment of process (B), a compound of formula (1) in which R1 represents a hydroxyl group The compounds can be prepared using the general deacylation conditions described below, where R1 is By deacylation of the corresponding compound of formula (1) representing an acyloxy group as noted Manufactured.

Carboxylic acid and hydroxyl protecting groups suitable for use herein include, for example, P protective Groups in Organic Cheajstr y', Ed, J, FJ, McOmle (Plenum Press, 1973) or °protect Groups in Organic 5yn thesls’by Theodora V, Greene (John Vil ey and 5ons, 1991). There is a protector. Examples of suitable carboxylic acid protecting groups include methyl or t-butyl Alkyl groups such as 2-methoxyethoxymethyl, or diphenylmethyl or an aralkyl group such as p-nitrobenzyl. Adequate hydroxyl protection Examples of groups include 2-methoxyethoxymethyl and silyl groups (e.g. t-butyl methyldimethylsilyl). Amino groups are, for example, aralkoxylic carbonyl (e.g. benzyloxycarbonyl) or alkoxycarbonyl (e.g. protected by a group selected from, for example, a t-butoxycarbonyl group.

Protecting groups are removed using conventional techniques. For this reason, for example, t-butyl The alkyl group (e.g. ether, hydrogen chloride in a solvent such as dioxane) ) under anhydrous acid conditions. The p-nitrobenzyl group is preferably such as esters (e.g. tetrahydrofuran or aqueous tetrahydrofuran). Removed using zinc metal and hydrochloric acid in a solvent. diphenylmethyl group or 2 - the methoxyethoxymethyl group is preferably aqueous formic acid or trifluoroacetic acid; removed using

Silyl groups such as t-butyldimethylsilyl are preferably treated with fluoride ions. removed. Removal of the amino protecting group as described above is amenable to hydrolysis under standard conditions. It is done more. The conditions described above for the removal of the t-butyl group are preferably t-butyl. It may also be used to remove butoxycarbonyl groups. Compatible methyl ester group Esterification of the carboxylic acid group of a suitable intermediate compound to an amide (e.g. for example, dimethylacetamide or preferably dimethylformamide). The presence of a base such as a bicarbonate (e.g., sodium bicarbonate) in a clean organic solvent in the presence of methyl halides (e.g. methyl iodide) or dimethyl sulfate. This is done by treatment with a methylating agent. The reaction is preferably from 0 to 100"C1 The process is preferably carried out at a temperature in the range of 20-30°C. On the other hand, esterification is similar to methanol. Treatment of diazomethane in a suitable solvent such as a mineral acid (e.g., hydrochloric acid) at about room temperature This can also be done by treatment with methanol in the presence of a suitable acid.

Conversion of one t-butyl ester to a different t-butyl ester can be accomplished by appropriate deeupholization. This is done by a stealth step. Deesterification is carried out under acidic conditions, e.g. This is carried out by hydrolysis using anhydrous hydrogen chloride in a suitable solvent such as chloride.

2 ml) of the compound can be prepared according to the fermentation process described below or by adding the appropriate acyl by acylation or deacylation at the 6-position as appropriate according to the It may also be produced from the product of a fermentation process. Suitable acylation methods are described below. There is. Deacylation is preferably carried out in a solvent such as an alcohol (e.g. methanol). carried out by base-catalyzed hydrolysis using a base such as aqueous sodium hydroxide in a medium It will be done. On the other hand, deacylation of α,β-unsaturated esters can be A suitable base (e.g. triethylamine) in a solvent such as hydroxylamine (e.g., N-methylhydroxy amine hydrochloride). The fermentation process involves the use of one or more formulas (ml). Cultivate microorganisms that can produce the desired compounds.

The desired compound is then isolated from the culture and acylated or de-acylated if desired. Acylated and/or esterified to the corresponding methyl ester.

Suitable microorganisms can be used to test samples obtained from microbial fermentation using small scale tests. is easily confirmed by analysis using standard methods.

In particular, the commonly used microorganism is CAB International Mycol.

gical In5tituto, Ferry Road, Kew, 5urr ey, England. The stock was released in May 1989. It was accepted by the institution on the 25th, and subsequently the accession numbers 1 old 3329B2 and 19 A mortar dated June 27, 1989 (confirmation of survival) was attached. Deposited stocks are held in trust It will be referred to in this specification as number 1 old 332962. IMI　33296 The characteristics so far identified for 2 are shown in Example 10 below.

It is clear that the desired intermediates may also be produced from the mutant strain of IMl 332962. Will.

Variants of I old 332962 occur naturally or from Radiatjon an d Radioisotopes f’or Industrial Micr oorganisas’, Proceedings of the Sympos iuIIl, Vienna, 1973. p241゜International H, I in Atomic Energy Authority.

Techniques ror the Developem by Adler Various people, including those described in ent of Hieroorganisms. produced by the method. Such methods include ionizing radiation, chemical methods such as N- Treatment with methyl-N-nitro-N-nitrosoguanidine (NTG), heating, Genetic technologies, such as recombination and transformation and selective techniques for natural variants, are Ru.

Fermentation is carried out by conventional means, i.e. in the presence of assimilable sources of carbon, nitrogen and inorganic salts. This is done by culturing.

Assimilated sources of carbon, nitrogen and minerals are provided by simple or complex nutrients. charcoal Sources usually include glucose, maltose, starch, glycerol, molasses, and dextrose. Thorin, lactose, sucrose, fructose, galactose, myo-inocyto alcohol, D-mannitol, soybean oil, carboxylic acid, amino acid, glyceride, alcohol oils, alkanes and vegetable oils. The carbon source is usually 0.5-10% by weight of the fermentation medium. %. Fructose, glucose and sucrose are the preferred carbon sources .

Nitrogen sources typically include soybean meal, corn steeping liquid, distiller's soluble ingredients, yeast extract, Cottonseed meal, peptone, ground nut meal, malt extract, molasses, casein, aminos acid mixtures, ammonia (gas or solution), ammonium salts or nitrates. Ru. Urea and other amides may also be used. The nitrogen source is usually 0.1 to 1 in the fermentation medium. o% by weight.

Nutrient inorganic salts added to the culture medium include sodium, potassium, ammonium, Iron, magnesium, zinc, nickel, cobalt, manganese, vanadium, chromium , calcium, copper, molybdenum, boron, phosphoric acid, sulfuric acid, chloride and carbonate ions There are commonly used salts that can produce ions.

Cultivation of organisms is usually carried out at 20-40°C, preferably at 20-35°C, particularly at about 25-28°C. temperature and may be carried out by aeration and agitation, e.g. shaking or stirring. desirable. The medium is first inoculated with a small amount of mycelia and/or spores. obtained The nutrient inoculum is either a fermentation medium or is subjected to further growth before being transferred to the main fermentation medium. transferred to one or more seed stages. Fermentation is usually 3.5-9.5, preferably 4.5-7 ．． It is carried out in a pH range of 5. To keep the pH within the desired range, the fermentation medium must be It is necessary to add a base or acid. Suitable bases to be added include aqueous There are alkali metal hydroxides such as sodium hydroxide or potassium hydroxide. suitable Correct acids include mineral acids such as hydrochloric acid, sulfuric acid, or phosphoric acid.

Fermentation is carried out over a period of 4 to 30 days, preferably about 7 to 18 days. Defoaming Agents may also be present to control excessive foaming and should be added at the required intervals. It will be done.

Carbon and/or nitrogen sources may also be provided into the fermentation medium if necessary.

The products of the fermentation process are present both in the fermentation liquor and in the mycelial fraction, preferably filtered. Or separated by centrifugation.

Optionally, the solution then has a pH of pI (below 6 (e.g., about pH 3)). until treated with an acid such as sulfuric acid in the presence of an organic solvent.

Products of the fermentation process are separated from the fermentation broth by conventional isolation and separation techniques. You may. Obviously, the choice of isolation technique is wide-ranging.

The products of the fermentation process are subjected to various fractionation techniques, such as adsorption-elution, sedimentation, fractional crystallization, Isolated and purified by solvent extraction and liquid-liquid partitioning, which can be You can combine them either way.

Adsorption to a solid support followed by elution facilitates the isolation and purification of the compounds of the present invention. It was found to be suitable for manufacturing.

The products of the fermentation process are ketones (e.g. acetone, methyl ethyl ketone or methyl isobutyl ketone), halogenated hydrocarbons, alcohols, diols (e.g. propane-1,2-diol or butane-1,3-diol) or Cells are washed with a suitable organic solvent such as methyl acetate or ethyl acetate (e.g. methyl acetate or ethyl acetate). and extracted from the aqueous phase. Typically, two or more extractions will help achieve the best recovery. desirable. The water-immiscible solvent extract itself is extracted with a basic aqueous solution, and these basic After acidification of the solution, the desired compounds are re-extracted into the water-immiscible organic phase.

Removal of the solvent (e.g., by evaporation) from the organic extract containing the desired compounds Obtain the substance.

Chromatography (including high performance liquid chromatography) Suitable supports; non-functional reticulated adsorption resins, such as Amberllte XAD- 2. XAD-4, XAD-16 or XAD-1180 resin (Rohm & H aas Ltd) or Kastel I S112 (Montedison) Cross-linked styrene-divinylbenzene polymer resins such as; substituted styrene-divinyl Benzene polymers, such as halogens such as Dlaion 5P207 (Mitsubishi) (e.g., brominated) styrene divinylbenzene polymer; anion exchange agent ( For example, IRA-35 or IRA-18); 5ephadex LH20 ( Organic solvent compatible cross-linked dextran such as Pharmacia UK Ltd) carried out on a reverse phase support such as hydrocarbon-bonded silica, e.g. C18-bonded silica . Another means of chromatography for the purification/separation of products of fermentation processes is Countercurrent chromatography using a coil extractor, such as a bed coil extractor.

The products of the fermentation process can be isolated or isolated by the use of liquid anion exchangers such as LA2. It may also be purified.

11? A-68 or especially I]? When A-35 is used as a solid adsorbent, Cell extracts can be added directly without removal of solvent.

The extract may be added directly at about pH 3 or about pH 8 after filtration of solid impurities.

2 ml) of suitable solvents for chromatographic purification/separation of suitable compounds/ The eluent will of course depend on the column type and the nature of the support. Countercurrent chromatograph When using fission, we use ethyl acetate, hexane, methanol and aqueous acid ( We have found that solvent systems containing (eg, aqueous sulfuric acid) are particularly suitable. IRA-35 When using anion exchange agents such as and then eluted with sulfuric acid in aqueous acetone.

The presence of products of the fermentation process during the extraction/isolation operation is determined by h, p, 1. c, ma or by conventional techniques such as UV spectroscopy, or by taking advantage of the properties of the compound. It can be re-monitored by using

The products of the fermentation process are dissolved in organic solvents after e.g. chromatographic purification If the solvent is obtained in the form of a removed by exposure. If desired, compounds can be further chromatographed as described above. Refined.

giving a compound of formula ml> in which R represents an acyloxy group as defined above in formula (1); Acylation of the formula (Vl+) in which R1 is a hydroxyl group or a protected derivative thereof by treating the corresponding compound with a suitable acylating agent under conventional esterification conditions. This is carried out by removing the protective groups present after . For this, for example, the formula ml ) in R1c When R represents a compound of the formula (Vl+) in which R1 is a hydroxy group, the R group is represented by the following formula ( vllll) acid; or by treatment with an activated derivative thereof. For this reason, the formula (vl The acid acylation of ll) is preferably performed with an acid such as dicyclohexylcarbodiimide. in the presence of a suitable carbodiimide such as 4-dimethylaminopyridine. Solvents such as halogenated hydrocarbons (e.g. dichloromethane) in the presence of a suitable base. It is done in medium. On the other hand, the acid of the formula ml+) can be prepared using, for example, thionyl chloride to convert the corresponding acid into lorides, and then the acylation reaction is carried out with halogenated hydrocarbons (e.g. dichlorohydrocarbons). 2°4,6-drimethylpyridine or N,N-dimethylpyridine in a solvent such as in the presence of a base such as tylaniline or an alkali metal carbonate or an alkali carbonate. It is carried out using potash earth metals (eg calcium carbonate).

Acylation or deacylation and esterification processes may be carried out sequentially or simultaneously, as appropriate. It is understood that the steps may be combined as appropriate.

The compound of formula (vllll) is preferably a compound of formula (vllll) such as By hydrolysis of a compound of formula (Vl+) in the presence of a co-solvent, e.g. of aqueous sodium hydroxide in a solvent such as an alcohol (e.g. methanol). prepared by base-catalyzed hydrolysis using bases such as

The base salt of the compound of formula (1) is preferably a base salt of the compound of formula (1). formed by treatment with a group. For this reason, for example, salt is preferably with sodium or potassium hydroxide, bicarbonate, carbonate or acetate (e.g. , potassium hydroxide, potassium bicarbonate, sodium bicarbonate or potassium acetate ), ammonium acetate, calcium acetate and L-lysine; is prepared by treating a compound of formula (1) with a base. For example, salt is a suitable solvent and/or alcohol (e.g. methanol) or nitric acid. A solution of a compound of formula (+) in a co-solvent such as lyl (e.g. acetonitrile) or is added to the suspension at a temperature of e.g. 0 to 80°C, preferably at about room temperature. or by adding a base (if necessary as an aqueous solution).

Acid addition salts can be prepared in a suitable solvent such as water and/or alcohol (e.g. methane). water) or nitrile (e.g. distilled water, total volume up to 1 liter) Prepared by treating a compound of formula (+) with a suitable acid.

Physiologically acceptable salts can be prepared using conventional methods using other physiologically acceptable salts of the compound of formula (1). It may also be made from other salts, including those salts.

Compounds of formula (11) are novel intermediates and form another aspect of the invention.

The following examples are given to illustrate the invention and are not intended to limit it in any way. isn't it.

Ruhexyl'I-4.6.7-1-lihydroxy-2,8-dioxabicyclo [ 3,2,1]octane-3,4,5-tricarboxylic acid, 6-(4,6-cymethylate) 2-octenoate) (a) I) It 3329fi2 was grown on agar plates with the following composition: Malt extract (Oxoid R39) 30g Fungal peptone (Oxoid R40) ) 5g yeast extract (Oxoid R21) 0.5g agar (Oxoid No. , 3) The pH of the medium before 20g autoclaving is in the range of 5.3 to 5.5. It was surrounded. The inoculated plates were incubated at 28°C for 14 days. fungus Cut several 6-diameter plugs of mycelium-covered agar from the growing end of the culture. and plug two plugs into each of several cryotubes containing 1.6 liters of sterile distilled water. I put it in. The tube was capped and stored at room temperature until needed.

contained in a 250+el Erlenmeyer flask using two agar plugs. Seed culture medium (A) 5C1+l was inoculated into 8 cells each: Seed medium (A): Peptone (Oxoid R34) 10g malt extract (Oxoid d R39) 21g glycerol 40g Junlon 110 (l(oneyville & 5teln) Ltd,, W alltngton, 5urrey) 1g distilled water, total amount up to 1 liter The pH of the medium was adjusted to 6.3-6.5 with aqueous sodium hydroxide before autoclaving. It was adjusted.

The flask of the inoculated seed medium was moved at 25 orpm while moving in a 50a+m diameter orbit. The cells were incubated for 5 days at 25° C. on a shaker plate rotating at 50°C.

The contents of the flasks were pooled and homogenized. homogenized seed culture was used at 3% (V) to prepare a fermentation medium (B ) 120 were inoculated with 50 wl each: Fermentation medium (B): Glycerol 50g Soybean oil 30g Cottonseed flour (Sigma) 10g Distilled water up to 1 liter The pH of the medium before autoclaving was in the range of 6.1 to 6.3. centre The flasks were incubated as above for 8 days with shaking.

Fermentation broth (approximately 6 L) from flasks incubated for 8 days was filtered to remove bacteria. The strands were removed and the filtrate was adjusted to pI (2.8) with sulfuric acid (20% v/v) and diluted to double the volume. The mixture was extracted three times with ethyl acetate. Combine the ethyl acetate extracts and dilute with aqueous sodium bicarbonate. Back-extraction was carried out twice with 400 ml of um solution (1% v/v). Combine the aqueous back extract , adjusted to pH 2.8 as above and re-extracted twice into ethyl acetate 8001.

These extracts were combined and evaporated to dryness to give a brown oil. This oil is It o Multi-1ayer Co11 Extractor (P, C, Inc. Countercurrent chromatography using a Potomac, Maryland, USA) It was further processed in The coil used was about 70 meters, indicating the total capacity piece 3801. Standard prevarication consisting of Tor's 2.6m+I++ inner diameter PTFE tubing It was a coil. The solvent system used was ethyl acetate, hexane, methanol and It was a mixture of N/100 sulfuric acid (5:5:5:6 by volume). The lower phase remains unchanged. Ta. G11son Model 303 pump and Model H2 on the coil OManosetricModule (Gilson, Vlliers Le Be1. France) and filled with the lower phase. Then oil (upper phase 41+ The lower phase 497 a+g in 41) was injected at the "tail" end of the column. Then centrifuge was operated at 800 revolutions/sin, and the mobile (upper) phase was transferred from the end of the column tail 1 to 4. Pumping was carried out at ml/1 Iin. The 201 fractions were collected and the squalene synthase This was monitored by measuring inhibition.

Consecutive fractions showing activity against squalene synthase were combined. Steaming the early fractions The mixture was evaporated to dryness to give the title compound (90 ig) as a pale yellow oil.

(b) Mycelium isolated from 6L broth for 8 days according to the procedure in part (a) above. Extract the incubated flask with methanol (2X 3 L) and filter. did. The filtrate was concentrated to about 5001 by evaporation, adjusted to pH 3.0 with formic acid, and diluted with vinegar. Extraction was performed three times with 500 ml of ethyl acetate. Combine the ethyl acetate extracts and dilute with sodium bicarbonate. Back extraction was carried out twice with Lithium solution (1% v/v) 2001. Combine the aqueous back extract , adjusted to pH 3.0 and re-extracted twice into 500 ml of ethyl acetate. all have The machine fractions were combined and reduced to dryness using a rotary evaporator to form a brown oil. I got it. The oil (578 mg) was collected using three G11son solvent delivery pumps (model 303), 811 Dynamic Mixer and 802CManometer G11son autopreparative system consisting of jcModule Further processing was performed using high performance liquid chromatography (HPLC). chromatog Raffy is Dynamax Microsorb C1g (5um) semi-pre It was carried out using a Varativ column (250×10 aa+). Mobile phase runs for 65 minutes Acetonitrile pumped at 2.8 to 5,6 ml/l inch over hours tolyl and 0.1% v/v formic acid and ammonium acetate (1:3 →4:1→1:3). This method was repeated 16 times. . 13X4°95m1n fractions were collected and measured for inhibition of squalene synthase. It was monitored by Combine fraction N015 from each run and pH 3 with formic acid. , 0 and extracted twice with ethyl acetate 1001. Remove organic phase and evaporate It was dried to give the title compound (172+eg) as a pale yellow oil.

(c) (1) A 5-day fermentation carried out as in part (a) above.　5IIl Seed culture contained in 250 ml Erlenmeyer flasks using 8 l each The soil (A) 50a+1 was inoculated into 8 pieces each. The flask was transported in a 50 tnm diameter orbit. Incubate for 4 days at 25°C on a shaker plate rotating at 25 rpm while moving. Cubated. The contents of the flasks were pooled and homogenized.

Using a homogenized seed culture at 3% (V/V), 120 pieces of fermentation medium (B) were inoculated in a jar flask. flask 1o Incubate for 1 day with shaking as above.

(Q) (if) Homogenized seed culture prepared as in part (e) (i) above Fermentation of 5 liters and 3 liters each using nutrients at 3% (V/V) Two fermentation vessels containing medium (B) were inoculated. The inoculation medium was maintained at 25°C; Stirring with two six-sided turbine rotor blades (70°C, 1m diameter) rotating at 500 rpm did. The culture was aerated by sparging with sterile air at 3 Lpm. Siri Excess of culture by addition of Corning antifoam (Day Corning 1520) Prepared to control foaming. After 11 days of growth, the contents of two culture vessels were further processed by countercurrent chromatography according to the procedure in part (a) above. , the title compound (13.7 mg) was obtained; b) N, I1. r, Teha approximately δ0.84-0.90 (m, 9H), 1.03 (d, 7.38), 1.09-1.19 (m, 2) 1), 2.10 (s , 3H), 2.24 (IIl, IH), 2.34 (w, IH) , 2D68 (dd, 13.6.1) 1), 4.04 (d, 2. IH), 4.97 (s, LH), 5.02 (s, LH).

5.08 (d, 5, LH), 5.27 (s, LH), 5.80 ( d, 1B, IH), 6.31 (d, 2.Lg) , 8.85 (dd, IB, 8.IH), 7.14 (t, 7,1) (), 7.19(d, 7.2H), 7.2[i(t, 7,2H) indicates signal; Combined pulse decoupling 125.75MHz charcoal in deuterium-methanol Element-13N.

m, ", then about 6172.5 (0), 172.1 (0), 170.1 (0), 1 68.5 (0).

LH, 5 (0), 157.6 (1), 147.7 (0), 141 ．． 6 (0), 130.2 (1), 129.382, 5 (1), 8L, O (1), 80.1 (1), 78.6 (1), 75.6 (0), 44.4 (2).

40.9 (2), 37.7 (1), 35.8 (1), 34.9 (2), 33.1(1), 30.8(2).

26.5 (2), 20.9 (3), 20.5 (3), 19.2 (3), Peaks are shown at 14.1 (3) and 11.4 (3).

(d) (i) Freeze stock of inoculum for 5 days following the procedure in part (a) above. Prepared from fermentation. Centrifuge the culture sample for 10 minutes and remove the mycelium from 15% glycerinase. Roll and resuspended in 0.01% Tveen 80 to original volume. mycelium Spin down and resuspend again, then add 1.81 volumes into a plastic tube. and stored at -20°C. Using 8 x 0.51 frozen inoculum, 250 1. Patch 50 ml of seed medium (A) contained in 1 Erlenmeyer flask to 8 pieces. I planted a seed. Rotate the flask at 25 Orpm with a 50+aa+radial orbital motion. The cells were incubated for 4 days at 25°C on a shaker plate. Seed flask contents The samples were pooled and used at 3% (V/V) in a 250 ml Erlenmeyer flask. 120 cells were inoculated with 50 ml each of medium fermentation medium (B). Keep the flask as described above for 9 days. The cells were incubated with constant shaking.

(d) (i) Final stage flora grown as in Ha part (d) (i) above. After 7 days of incubation, the contents of 4 Scos were pooled and mixed into fermentation medium (B) 50+ Homogenize to obtain seeds for 120 pieces of nl each, and add this to the above part ( c) (i) and (d) Incubated as in (i) for 8 days. 8 days in Filter the fermentation broth (approximately 6 L) from the incubated flask to remove mycelium. I left. The filtrate was adjusted to pH 2.8 with sulfuric acid (20% v/v) and extracted with ethyl acetate. and back-extracted into sodium bicarbonate and purified as described in part (a) above. Re-extracted into ethyl acetate with H2.8. Concentrate the ethyl acetate extract under reduced pressure to give a yellow A colored oil was obtained which was dissolved in methanol (10ml). Add this solution to 3-1 OD S-3 (Vhatman Partlsil Biopr ep40.75 people, slurry filled with acetonitrile-water 20:80) column (32×2.5 cm).

The column has acetonitrile ratio of 1=4.3, 2:3.1:1.3:2. eluted with a stepwise gradient of a mixture of acetonitrile and water, increasing to . Fractions were monitored by HPLC and fractions containing the title compound were evaporated and aliquoted. Setonitrile was removed. The resulting aqueous suspensions were pooled, lyophilized overnight, and the ash The title compound (59 mg) was obtained as a white solid.

(e) Followed the procedure in part (d)(i) except that the pooled seed flasks were % (v/v) to inoculate 4 L of seed medium (A) in a 7 L fermentor. culture 500 rpm under agitation as described above while aerating the culture at 4 L/ll1 in. ll for 2 days. Remove 1.2 L of culture and add 40 ml of seed medium (A). It was used to inoculate a 70L fermentor filled with L. Culture at 40L/m Cultures were incubated as above for 2 days at 500 rpII without aeration. I did it. Remove 15L of culture and fill 780L with 500L of fermentation medium (C) Added to fermenter.

Fermentation medium (C): Fructose 50g Soybean oil 30g Cottonseed flour (Sigma) 20g neutral pH Aerate the culture at 500 L/win for 450 hours at 200 rpm. Incubate under shaking as above and add 50% (V/V) solution of fructose. The solution was increased to 5 g/L/day at 120 hours and 7.5 g/L/day at 162 hours. and supplied it. Analysis of the broth at 450 hours yielded 1056 mg/L. The yield of the compound is shown.

The above operation was repeated on a small scale, but fructose was replaced with glucose, galactose, Sucrose, maltose, lactose, myo-inositol, D-mannitol and other carbon sources selected from soybean oil. Each experiment at the 450th hour? Analysis of their broth showed substantial potency of the title compound.

The title compound prepared according to the above procedure was consistent with a product having the following characteristics.

Approximate molecule ff1690;-FAB mass spectrum GJ constant CM-H]-689, 2789; + F A B mass spectrometry [M + Na co-deuterium-chlorophore 500 MHz proton N, m, r, spectrum [multiplicity and cutoff are in parentheses] Pulling constant (H) and δ value with integral value): 0.79-0.85 ( m, 9H), 1.00 (d, 7.3H), 1.04-1.15 (m, 2 1(), 2.09(s, 3H), 2.4[i(a+,LH).

2.69 (dd, 13.5. LH), 4.05 (s, IH), 4.9 4 (s, LH), 4.96 (s, 1) 1).

5.06 (d, 4. LH), 5.30 (s, IH), 5.78 (d, 16. LH), 5.92 (s, IH).

6.88 (dd, 16,111.1) 1), 7.11 (d, 7.2) 1 ), 7.14 (t, 7. LH), 7.24 (t, 7.2H).

Combined pulse decoupling 125.75MHz in deuterium-chloroform Carbon-13 N, m, r, spectrum [δ value showing the number of bonded protons in parentheses) : 171.5(0), 171.0(0).

169, 1 (0), 167.0 (0), LH, 7 (0), 157 ．． 9 (1), 145.4 (0), 140.1 (0), 12g. 9 (1 ), 128.1(1), 125.8(1), 117.8(1), 111.4(2 ).

105.8 (0), H, 5 (0), 81.6 (1), 80.7 (1 ), 79.3 (1), 75.1 (1).

74, 2 (0), 42.9 (2), 39.7 (2), 36.7 (1) , 34.2(1), 33.8(2).

31.6 (1), 29.4 (2), 25.4 (2), 20.9 (3) , 19.8 (3), 18.8 (3).

13.5(3), 10.9(3).

A suspension of the lyophilized product (1 g) of intermediate 1 in water (100 ml) was added to water (10 ml). It was treated with a solution of potassium hydrogen hydroxide (430 g). Lyophilize the resulting solution The title compound (1.08 g) was obtained as a beige solid; (NLIjOI) 3491-3187 (br OH), 1731 (Ester c-o>.

16140m-' (carboxylate c-o and c-c); Actual analysis value: C48,85; H5,70; K14.1; H20B, 2; CHKo ・3 H20 calculation value 2 C4g, 93; H5, 75; K 13.65; H20B, 29% methanol ( A stirred suspension of Intermediate 2 (15.6 g) in IL) was added dropwise with concentrated hydrochloric acid (13 ml). I understood. The resulting clear solution was stirred at room temperature for 24 hours. Then convert it into solid carbonic acid It was treated with sodium hydrogen (13.2 g) and most of the solvent was evaporated under reduced pressure.

The residue was treated with aqueous hydrochloric acid (2M 1500 ml) and extracted with ethyl acetate (ILX3). Output table Hei 7-504421 (14) Ta. The organic extract was washed with water (IL), dried over magnesium sulfate, filtered, and evaporated. I let it emanate. The residue was dissolved in dry toluene (130ml) and heated to 80°C under nitrogen. and then N,N-dimethylformamide di-t-butyl acetal (38 ml) The solution was added dropwise for 30 minutes. The reaction mixture was stirred at 80°C for 3114 hours and then It was then cooled down. It was diluted with ether (700i1) and brine (600ml). Washed. The organic layer was dried over magnesium sulphate and the solvent was evaporated under reduced pressure. Residue The residue was subjected to silica gel (Merck 9385.900g) flash column chromatography. Graph and eluted with 5:1 to 1:1 cyclohexane:ethyl acetate.

The appropriate fractions were combined and the solvent was evaporated to give the title compound (5,93 g) obtained; protons N, m.

r, (CDCI) value is 60.8-0.9 (G9H13Cf13), 1.05 (d, 3H1-ClClCl, J"6.2Hz), 1.6 and 1.48 (2 S, 18H, 2CO2C (Cl.

3) 3), 2.1 (s, 3H, Cl, co□), 2.71 (dd, LH , Pb0 and one of J-13,7 and 5Hz), 2.98 (d, LH, cHO H, J-2Hz>, 3.73 (s, 311° CO2Cl,), 4.1 (s, IH , OH), 4.05 (t, IH, cHOH, J-2Hz), 4.97 (s, 2H , C-CH2), 5.1 (d, IH, Ac0c:, J-5H2), 5.26 (S , lH, ClCo CH>, 5.77 (d, IH, CH-C, CO02, J-1 6,2H2), 5.97 (d, ratio CHOCOCH-CI, J-2Hz), 6.9 1 (dd, LH, Cl-CHCo 2.J-16, 2 and 8.7Hz), 7 ．． 1-7.3 (m, 5H, Ph); t, 1. c, (Si02) ethyl acetate /cyclohexane (1:1) Rfo, 53 Intermediate 4 1 and 2 2 Q2 hi "Nino" tips 2-" 21 Sho 3 begging Σ- Decoy Gai 222" "Shomu" Standing on the ground 2L" 3m begging 2" 2 above the ground - L Nim L Usumi "New Shadow Yu Mu1!慇ヱ正"nee 2-1) vyy",=纺"-A-ΣTwo sleep sosashi 1-to ni tu soil - rainbow - ';if) to' 2! Mumumu sevenyoyu 4,5 two-story construction soil-Lyoyol] three l) ester, 3-methyl ester Intermediate 3 (10,38 g) in dry dimethylformamide (26 ml), t-bu Tyldimethylsilylchloride (19,6 g) and imidazole (17,7 g ) solution was stirred at 65°C under nitrogen for 16.5 h, then ethyl acetate (200 ml ) and 2M hydrochloric acid (200+al).

The aqueous phase was extracted with ethyl acetate (200ml). The combined organic extracts were diluted with 2M hydrochloric acid ( 1001111), washed with water and brine (2X100ml each), (MgSO 4) Dry and evaporate to an orange oil. Add this to silica gel (@erak 7734.500 g) and 4:1 cyclohexane. : Eluted with ethyl acetate. Combine the required fractions and evaporate to a colorless gum. The title compound (9.01 g) was obtained; protons N, m, r, (CDCIs) (60.05 (s, (C) I) Sl), 0.8-0.9 (m, c crow and and (Cl,)3C9i), 1.02 (d, J-6H2, -CHCHCH,), 1.4 and 1.65 (2s.

Co2C(CH,)3), 2.1(s, CHaCo), 3.73(s, Co2Cl,).

4.02 (s, 4-0H), 4.12 (d, J-2) 1z, 74), 4.98 and 5.0 (2s.

C-C and 2), 5.12 (d, J-5Hz, CHOAc), 5.28 (s, 3- 4), 5.8 (d, J-16Hz, 0COCH-CI), 6.38 (d , J-2Hz, 8-)j), 6.93 (dd, J-9 and 1 6Hz, 0COCR"Cji), 7.1-7.3 (ToCsdan5).

Intermediate 5 [IS-(la (4R', 53 pieces), 3a, 4β, 5α.

* 6α(2E, 4R, 6R*), 7β)) 1-(4-acetyloxy-5-methyl -3-methylene-6-phenylhexyl)-7-((dimethyl(1,1-dimethyl ethyl)]silyloxy)-4,6-cyhydroxy-2゜8-dioxabisic b[3,2,1)octane-3,4゜5-tricarboxylic acid, 6-(4,6-syl methyl-2-octenoate), 4,5-bis(1,1-dimethylethyl)S Tell A solution of intermediate 4 (9, OIg) in tetrahydrofuran (450 ml) at room temperature It was treated with 0.1M sodium hydroxide (116ml) under stirring. Dissolved after 0.5 hours The liquid was evaporated to a small volume and then mixed with ethyl acetate (250ml) and 2M hydrochloric acid (500ml). l). The aqueous phase was further extracted with ethyl acetate (2x250ml). Matching The extracted extract was washed with water and brine (2 x 2501 each) and dried (MgSO4). , evaporated to give the title compound (8.7 g) as a white foam; protons N, m , r, (CDCIa) is 60.04 (s, (C anal 3) 2 Si), 0.8 -0.9 (Otori, CH and (Cl3)3C3i), 1.02 (d, J-8Hz , -CHCHCdan, ), 1.4 and 1.65 (2S, CO2C(Cl, > 3).

2.1 (s7CHa Co), 4.13 (di-2Hz, 74) 65.02 and and 5.05 (2s.

C-CH2), 5.11 (d, J-5Hz, C) (OAc), 5.22 (s, 3 - and), 5.78 (d, J-16Hz, 0COCR-C) I), 6.34 (d, J-2) 1z, 6-H), 6.95 (dd, J-9 and 16Hz, 0COCH-C)l), 7.1-7.3(m, C6dan5). vinegar.

Intermediate 5 (5,89 g) in tetrahydrofuran (60 ml), N-hydroxy Succinimide (0,8 g) and N,N-dicyclohexylcarbodiimide A solution of (1.46 g) was stirred at room temperature for 17 hours. The resulting suspension was filtered; The liquid was evaporated to a white foam.

This was dissolved in dimethylformamide (60 ml), stirred at room temperature, and dissolved in dimethylformamide (60 ml). treated with sodium hydroxide (24211 g). After 55 minutes the suspension was filtered. filter The liquid was partitioned between ethyl acetate (500*I) and 2M hydrochloric acid (500*I). organic phase Water (500*I), saturated aqueous sodium bicarbonate and brine (2X500a each +l), dried (MgSO4) and evaporated. The residue was treated with silica (Merc 7734.300 g) and 3:1 cyclohexane. : Eluted with ethyl acetate. Combine the required fractions and evaporate as a white foam. The title compound (2.55 g) was obtained; protons N, m, r, (CDCI) 60.04 (s, (C, Qa　)2　Si), 0.8-0.95 (II, Cl3 and (Cl3)3CSi), 1.02 (d.

J-6Hz, = CHCHCl3), 1.4 and 1.61 (2S, Co2C (CH,) 3), 2.1 (s, CHCO), 3.58 and 3.78 (m, CH20H), 3.86 (s, 4 10th), 4.11 (d, J-2Hz, 74) , 4.85 (dd, J-4 and 6Hz, 3-1).

4.98 and 5.0 (2s, C-C top 2), 5.12 (d, J-5H2, C HOAc), 5.8 (d, J=16.Hz, 0COCH-CH), 6. 32 (d, J-2Hz, 6-H), 8.92 (dd, J-9 and and 16Hz, 0COCH-CH), 7.1-7.3 (IIl, C6H5) show.

Tan-4,5-tricarboxylic acid, 6-(4,6-dimethydry dichloromethane ( A solution of intermediate 6 (1,19 g) in 20 ml) was stirred under nitrogen in a water bath and 2,4. 6-collidine (225 μL, 208 mg) followed by anhydrous trifluoromethanesulfate. Treated with fonic acid (280 μL, 476 mg). After 45 minutes, remove the yellow solution by dichloromethane. diluted with lomethane (1501), 2M hydrochloric acid (2X50011), water (1001) , saturated aqueous sodium bicarbonate (2X 100+ ol) and brine (2X 10 0 ml), then dried (MgSO4) and evaporated to give the title as a foam. Compound (1,296 g) was obtained: with protons N, rA, r, (CDCla) is δ0.05 and 0.0B (2s, (CH) St), 0.8-0.9 (m , CH and (CH3)a C3+), 1.02 (d, J-6) (z, -CH CHCfi3), L-41 and 1.65 (2s.

Co　C(CM　)　), 2.1(s, C4a　Co)　, 3-88　(s, 4-01), 4.13 (bs.7-11), 4.32 (dd, J-2,5 and 1IHz, IH, C) I OSO2), 4.57 (dd, J-7, 5 and 1 1Hz, 3-H), 4.95 and 5.05 (m, C-Cpz and printing 20S 02-1), 5.11 (d, J-5Hz, CHOAc), 5.8 (d.

J=IGHz, 0COCH-C1(), 6.3(bs, 6-H), 6.95(d d, J=9 and 16H2,0COCH-CH), 7.1-7.3 (l11. C6...5) is shown.

6α (2E, 4R, 6H books), 7β) El-(4-ane Cetyloxy-5-methyl-3-methylene-6-pheny(1,1-dimethylethyl) )]]silyloxy]-4.6=dihydroxy2,8-dioxabicyclo(3 ,2゜1] Octane-4,5-dicarboxylic acid, 6-(4,6-dimethyl-2- octenoate)), 4,5-bis(1゜1-dimethylethyl)ester A solution of intermediate 7 (584 mg) in dry dimethylformamide (61) was stirred at room temperature. Stir and treat with sodium azide (112 mg). After 4 hours, the mixture was diluted with acetic acid. Partitioned between chill (10C1al) and 2M hydrochloric acid (50ml). The organic phase was dissolved in 2M hydrochloric acid ( 501), drained and washed with brine (2 x 50 ml each), (MgS04) Drying and evaporation gave the title compound (4751g) as a colorless gum; Ton N, II1. r, (CDC13) has δ0,04 and 0.06 (2s, (CHa)2Si), 0.8-1.0(m, CH and (Cl3)a C3i), 1.02 (d, J-6Hz, -CHCHC)13), 1.42 and 1.64(2S,C02C(CH,3)3>.

2.09 (S, CH3Co), 2-91 and 3.52 (2dd, J-2,5 , 13 and 7.5, 13H2, cl (2N3), 3.81 (s, 4-011 ), 4.12 (bs, 7-ri, 4.78 (dd, J-2,5 and 7.5Hz, 3-H), 4.97 and 5.0 (2s, C-Cl2), 5.11 (d, J-5Hz, CHOAc), 5.8 (di-16Hz, 0COC day-〇〇), 6 ．． 32 (bs, 6-H), 6.94 (dd, J-9 and 1 [iHz], 0 COCH-CI(), 7, 1-7, 35(m, C6dan,) is shown.

Intermediate 9 cetyloxy-5-methyl-3-methylene-6-phenylhexyl)-3-azi Domethyl-4,6,7-)lihydroxy-2,8-dioxabicyclo(3,2, 1) Octacy-4,5-dicarboxylic acid, 6-(4,6-cymethyl-2-octacyl) 4,5-bis(1,1-dimethylethyl)ester A solution of intermediate 8 (453 IIg) in tetrahydrofuran (10 ml) was stirred at room temperature. Stir and treat with tetrabutylammonium fluoride (0.5 ml of a 1M solution). Ta. After 70 minutes the solution was evaporated to dryness. The residue was mixed with ethyl acetate (100ml) and water. (50ml). The organic phase was dissolved in water (50if) and brine (2X50111 Wash with 1), dry (MgS O4), and evaporate until it becomes a colorless rubbery substance. Ta. This was chromatographed on silica (Merck 7734.43g). and eluted with cyclohexane:ethyl acetate 5:1. Combine the required fractions and evaporate. The title compound (363 mg) was obtained as a colorless gum; protons N, m , r, (CDCl2) has δ0.8-0.9 (m, CH), 1.05 (d, J -6Hz, -CHCHCH3), 1.5 and 1.58 (2s, C02C(CH ,) 3), 2.1 (s, Cdan, Co), 2.98 (d, J-2H2, 7-0 )1), 3.12 and 3.55 (2dd, J-5,12,5 and 7.5.1 2.5) 1z, CH2N3), 3, 85 (s, 4-Ofi), 4 -06 (i, 7-H), 4, 65 (dd, J-5 and 7.5Hz , 3-H), 4.95 and 4.98 (2s, CCdan 2), 5.09 (d Te Intermediate 9 (283B) and triphenylphosph in trihydrofuran (3 ml) A solution of fin (100+eg) was stirred and treated with water (0,61). Stir 44 C for 23 hours, after which the solution was evaporated. The residue was dissolved in ethyl acetate (10C1+l ) and the solution was dissolved in saturated aqueous sodium bicarbonate (2X2511) and brine ( 2 x 25 ml) and then dried (MgS O4) to form a foam. This was chromatographed on silica (Merck 7734.40g). The chloroform was increased from 95% methanol to 10% methanol. eluted. The required fractions were combined and evaporated to give the title compound ( 180 mg) was obtained; proton N, m, r, (CDCl2) was 60.8-0 ．． 9 (m, cH), 1.04 (d, J-6Hz, =CHCHCCH3C0J5 and 1.56 (28, Co2C (CH3) 3), 2.1 (S, CH, CO), 2.9・2 (1, C, j42NH2), 4.02 (d, J-2Hz, 74), 4.4 (t, J-4Hz, 3-1), 4.98 and 4°97 (2s, C-C) 12), 5.09 (d, J-5Hz, CHOAc), 5.78 (d, J-18H z, QC6α (2E, 4R, 6R*), 7β)) 1-(4-A* cetyloxy-5-methyl-3-methylene-6-)dinylhexyl')-3-( aminomethyl)-4,6,7-dolihydroxy-2,8-dioxabicyclo(3 ,2,1) octane-4,5-dicarboxylic acid, 6-(4,6-cymethyl-2- octenoate), 5-(1,1-dimethylethyl) ester 6.5 M chloride in dioxane (115 ml) 0 in hydrogen chloride (2,85 g) The solution was kept at room temperature for 8 hours and then evaporated. Dissolve the residue in ether and make the solution Evaporate to a yellow foam and add 5pherisorb 0DS-2 (2x 2 Trifluoroacetic acid ( 1ml/L) Containing water 70% then 98% acetonitrile at a flow rate of 15a+l Purification was performed by elution with /ll1n. from the later running peak. The fractions were combined, concentrated by rotary evaporation, and then lyophilized to form an off-white solid. The title compound (374-g) was obtained as a body; protons N, a, r, (dB-D MSO), 60.75-0.88(1, CH3), 0.9[1(d, JJHz , -CHCHCH), 1.31(s, (CH) C), 2.1(s, CH3C 0J 3 3 ), 3.85 (bd, J=5Hz, 7-4), 4.82 (bd , J-7Hz, 34), 4.91 (bs, C| CH2), 4.98 (d, J=5Hz, Cdan0Ac), 5.82 (d, J-1 6Hz, 0COCH-○○).

6.18 (d, J=5Hz, 7-0H), 6.28 (bs, 8-H) , 6.79 (dd, J-9 and 16Hz, 0cOcH-CH), 7.1 2-7.35 (w, C61j 5).

Analysis actual values: C55.8, H8.8, N 1.9. F　8.7CHNo　-C HF 0/2H20 calculated value: C55,8; H7,1,N 1.6. F B, 5% intermediate 12 (Is-[1α(4R,5S*), 3α, 4β, 5α.

* 6α(2E, 4R, 6R*), 7β)) 1-(4-A* cetyloxy-5-methyl-3-methylene-6-)dinylhexyl)-3-(( ((1,1-dimethylethoxy)carbonyl]amino]methyl)-4,6,7- Dolihydroxy-2,8-dioxabicyclo(3,2,1)octane-4,5- Intermediate 11 in dicarboxylic acid, 6-(4,6-dimethyldichloromethane (21) (110 mg), di-t-butyl dicarbonate (35 mg) and triethyl acetate A solution of min (36 μL) was stirred at room temperature for 24 hours and then evaporated to dryness. residue was partitioned between ethyl acetate (50ml) and 2M hydrochloric acid (5C1+I). 2M organic phase Washed with hydrochloric acid (2X50ml), water and brine (3x50+al each), (M g S O4) Dry and evaporate to give the title compound (104B) as a white foam. obtained; proton N, a+, r, (CDCIa) is 60.8-0.9 (m, Cdan>, 1.02 (d, J-fliHz, -CHClICdan, ), 1゜4 5 and 1.52 (2s, (CH) C), 2.09 (s, 0 animals, Co), 3.13 (bS, 7-0H), 3.27 (1, NC)! 2), 3.98 (bs. , 7 once), 4.7 & (i, 3 once), 4.94 and 4.96 (2s, C-C dan2), 5.09 (d, J-5Hz, CHOAc), 5.75 (bs, 6-)1 ), 5.78 (d, J-16Hz, 0COCH-CH), 6.92 (2d, J- 9 and 16Hz, 0COCH-CH), 7.1-7.3 (m, C6dan5). vinegar.

Analysis actual value: CB2.1, H8, lN 1.9 CHNo - HO calculated value: CB2.2, H7,9, N 1.7% Ruhexyl)-3-((1:(1,1 -dimethylethoxy)carbonyl]amino]methyl)-4,6,7-dolihydro xy-2,8-dioxabicycloC3,2,1)octane-4,5-dicarvone acid, 6-(4,6-dimethyl-2-octenoate), 5-(1,1-dimethyl A solution of intermediate 12 (96 mg) in thylethidimethylformamide (51) was diluted with carbonate. Treated with potassium hydrogen (15 mg) and methyl iodide (11 μL) and incubated at room temperature for 8 Stir for hours. The mixture was then diluted with ethyl acetate (501) and 2M hydrochloric acid (50ml). distributed. The organic phase was dissolved in 2M hydrochloric acid (50ml), water and brine (2X50ml each). Washed with water, dried (MgSO4) and evaporated to a gum. Siri this Chromatography was performed using Merck 7734.6 g. Elution was carried out with San:ethyl acetate 2:1. Combine the required fractions and evaporate to a white foam. The title compound (74 mg) was obtained as a product; proton N, m, ry (CDCl2 ) is 60.8-0.9(a, cI(3), 1.02(d, J-6Hz, CH 'CHCH3), 1.42(bs, ((43)3C), 1.52(bs, (C O) C), 2.09 (s, CHa Co), 3.02 (d, J-3Hz, 7- October), 3.22, NCH-2), 3. [18 (BS, 4-0 Dan), 3.89 (s, Co. 0 animals 3).

4.0 (brt, J-3Hz, 7 once), 4. f39 (m, 3-H), 4.9 5 and 4.98 (2s.

C-Cdan, ), 5.09 (d, J-5) 1z, cHOAc), 5.76 (d, J -16Hz, 0COCi-C8), 5.82 (d, J=2Hz, GH), 6.91 (dd, J-9 and 16Hz, 0COCH-CH), 7.1-7.3 (m9C6H5).

Analysis actual value + CB3.4; H7, 8, N 1.6CH No. 0.25H 20 calculated value: C[i3.6; H8,0:N L, S26 cetyloxy-5-methyl-3 -methylene-6-)2dioctane-4,5-dicarboxylic acid, 6-(4,6-dicarboxylic acid) methyl-2-octenoate) Dissolution of intermediate 10 (170 mg) in 6.5 M hydrogen chloride in dioxane (10 ml) The liquid was kept at room temperature for 8625 hours and then evaporated to a pale yellow solid. this was redissolved in 6,5M hydrogen chloride in dioxane (5 ml) and the solution was kept at room temperature for 4 hours. Then it was evaporated. Dissolve the residue in ether until the solution becomes a pale yellow solid. 5pher+5orb 0DS-2 (2X 25’cm) column. Trifluoroacetic acid (1 ml/L) containing 7 purified by elution with 70% acetonitrile at a flow rate of 15 ml/min. Manufactured. The required fractions were combined and concentrated by rotor evaporation. white solid substance A precipitate was collected by filtration, washed with water and dried under vacuum to give the title compound (61 mg). Proton N, Lr, (dB-DMSO) is 60.75-0.88 (m, c I3), 0.98 (d, J-6Hz, -CHCHCH3), 2.1 (s, C a Co), 2-85 (Ol.

CHNl (), 3.82 (dd, J-2 and 5Hz, 7-H), 4.58 (d d, J-4 and [iHz, 3-H), 4.9 (bs, C-fjj2), 4.9 8 (d, J-5Hz, CHOAc), 5°75 (d, J-16Hz, 0CO CII-CH), 6.12 (d, J-2Hz, G4), 6.72 (dd, J-9 and 1fiHz, 0COCH-CH), 12-7.3 (m, C6dan5) is shown.

Analysis actual value: C59,8, H7,1:N 1.9CHN O, O, LC2HF 302.1.2H20 calculated value: C59.7, H7.3; N 2.0% tetra Hydrofuran (5ml) Medium 1 (93+g), phenyl carbamate (123m A solution of g) and triethylamine (86 μL, 60 mg) was refluxed for 27 hours, It was then evaporated to dryness. The residue was mixed with ethyl acetate (25a+I) and 2M hydrochloric acid (25ml). ). The organic phase was further treated with acid (10 ml), drained and brine (2 x 2 each). 5ml), dried (MgSO4) and evaporated to give a white solid, 5ph Preparation using erisorb 0DS-2 (2X 25 cm) column Fluid with 60% acetonitrile in water containing sulfuric acid (0,15w1/L) by HPLC. Purification was carried out by elution at a speed of 15 ml/1n. Rotate the required fractions Concentrate by evaporation then treat with solid sodium chloride and with ethyl acetate (3x) Extracted. The combined extracts were washed with brine, dried (MgSO4) and evaporated. The title compound (38I1g) was obtained as a white solid; protons N, m, r , (dB-DMSO) is 60.75-0.9 (s, C day, ), 0.98 ( d, No 7Hz, -C)lcHcpa), 2.

09 (s, CH3C0), 2.95 (s, C, fi2NH), 3.85 (d d, J-2 Oyohi 5Hz, 7-)ri, 4.311 (dd, J-5 and 7Hz, 3-mouse), 4-91 (bs, C-%).

Analysis actual value: C59,4; H7,1,N 3.9C3,H5,N 2013 ．． 0.5) I20 calculated value: C59.4, H7.1, N C8% Bonic acid, 6-(4,6-dimethyl-2-octenoate), 4,5-dicassium mu salt Add the solution of side 2 (50B) in dioxane (5 ml) to water (11) and potassium hydrogen mesothelate. (14 mg).

The resulting solution was lyophilized to give the title compound (59 mg) as a white powder; For roton N, m, r, (D 20), δ0.82 (m, CH,), 0.9 1 (d, J-7Hz, CH3), 1. (11(d, J-7)1z, -CHCH CHa), 2.08 (s, 0COCHa), 3.19 (m, Cdan2N)l) , 3.94 (dd, J-2Hz, 7-)j), 4.49 (m, 3-f i), 4.9 (d, J”4Hz, CjjOAc), 4.97 and and 5゜02 (2s, = CC20), 5, 91 (d, J-18Hz, 0 COC and -C)l), 6.09 (bs, 84), 6.94 (dd, J-9 and 16Hz, 0COCR-CH), 7.2-7.4(m, C6H* (Is-[1α (4R, 55 lines), 3α, 4β, 5α.

6α, 7β)) 1-(4-acetyloxy-5-methyldimethylformamide ( 11) Middle side 2 (25 mg), N=methylhydroxylamine hydrochloride (7 mg ) and triethylamine (28 μL, 19 mg) were stirred at room temperature for 5 hours. Ta. It was then evaporated to dryness and the residue was dissolved in ethyl acetate (50 ml) and 2M hydrochloric acid (2 0 ml). Wash the organic phase with acid (20 ml) and brine (2x20 ml). Cleaned, dried (MgS04) and evaporated to a white solid. this Dissolve in dimethylformamide (11) and convert the solution to N-methylhydroxylamine. treated with hydrochloride (4 sg) and triethylamine (15 μL, 10 mg) and incubated at room temperature. The mixture was stirred at room temperature for 20 hours. It was then evaporated to dryness.

Treat the residue with trifluoroacetic acid (IIIll) and evaporate the solution to a gum. eluted using a 5pherisorb 0DS-2 (2X 25cm) column. 3 in water containing trifluoroacetic acid (11111/L) by preparative HPLC. Purification was achieved by elution with 5% acetonitrile at a flow rate of 151/win.

The required fractions were combined and evaporated to dryness. Treat the residue with methanol, leaving a cloudy solution. was filtered. The filtrate was evaporated to give the title compound (2IIlg) as a white solid. : Proton N, m, r, (CD300) has δ0.85 (d, J-6Hz, CHC Dans), 2.1 (s.

CH, CO), 2.7 (m1% NH), 4.02 (d, J-2) 1z, 7-H ), 4.98 (2s, C-CH2), 5.07 (d, J-5Hz, C)! 0Ac ), 5.1 (bs, 8-H), 7.1-7.3 (+a, C6 and 5); mass Svector (MW 588.8) LSIMS-585* (Is-(1α(4R,5S*), 3α, 4β, 5α.

6α (2E, 4R, 6R*), 7β]: 1l-(4-A book cetyloxy-5-methyl-3-methylene-6-phenylhexyl)-3-[( (aminoiminomethyl)aminocomethyl)-4,6,7-drihydroxyl2. 8-dioxabicycloC3,2,1)octane-4,5-dicarboxylic acid, 6- (4,6-dimethyl-2-octenoate), hydrochloride A solution of 1 (61.3g 1g) in methanol (1if) was mixed with triethylamine ( 0,051ml), then (aminoiminomethane)sulfonic acid (17mg) The mixture was stirred at 20° C. for 18 hours. The solvent was removed under reduced pressure and the residue was dissolved in vinegar. The mixture was partitioned between ethyl chloride and 2M hydrochloric acid. The organic phase was washed with 2M hydrochloric acid, brine, dried, Evaporation to dryness gave the title compound (62 mg) as a glassy substance; proton N , m, r, (DMSO-d 6) 60.75-0.9 (m, Cdan a), 0.99 (d, J-7Hz, = CIIC) IC) 13).

2.10 (s, Aco), 3.18 (m, ″) JCH2), 3.89 (br) d, J-6Hz, 7-1f), 4.52 (br t, J-4Hz, 3-H), 4 ．． 90 (s, = cH,,), 4.96 (d, J-5Hz, C, QO^C), 5. 79 (d, No. 15) 12. CI-C [CO2) 4.04 (d, J-8) 1z , O and), 8.08 (br　s, 6-H), 6.75 (dd, J=15 and 8 H2, CH-CHCO2), 7.15-7.13 ．． 35 (m, CH), CN, w, r, (CD30D) δ170.4, 1137 ゜0,165,0,157,5,155,9,lt5.9.139.8.128 ．． 4.127.8.125.2.

118, l, 109.7.105. L, 89.2.80.9, 79.4. 7g, 0, 73.1.42.7.40.9.39.2.36.1.33.8. 33.3.3L4.29.0.25.2.19.2.1g, g, 17.5. 12.4, 9.7° Analysis actual value: C57.05, H7.05, N5.34; C12,5C36H 5, N 30.2.0.5HC1・H20 (754,1) Calculated value: C57,34 ;H7,15,N5.57. CI 2.35% (4,6-cymethyl-2-oc A suspension of 1 (20 mg) in acetonitrile (11) was heated to room temperature under nitrogen. Stir with aqueous formaldehyde (2 μL of 40% solution) and hydrogenated triacetate. Treatment with sodium oxyboronate (19 mg) gave a clear solution. 3 hours 10 Add formaldehyde solution (4 μL) after 6 h and evaporate the reaction solution. I set it. The residue was stirred with ethyl acetate and filtered. Filter the filtrate until it becomes a white solid. Evaporate and apply a 5pherisorb 0DS-2 (2x 25 Cot) column. 7 in water containing trifluoroacetic acid (11/L) by preparative HPLC using Purification was carried out by elution with 0% acetonitrile at a flow rate of 15 ml/min.

The required fractions were concentrated by rotary evaporation and then lyophilized to form an off-white solid. The title compound (1 mg) was obtained. 5pherisorb 0DS-2 (4,6 in water containing trifluoroacetic acid (1 ml/L) using a mIIX 25 cm) column. Analytical HPLC eluting with 70% acetonitrile at a flow rate of 1°5 a+I/min retention time 6°4 l1lin; mass spectrum CMV 889.8) 10F ’toB [i90 (Mll”), -T’AB1:l5-(1α (4R, 5 5*), 3α, 4β, 5α.

* (hexyl)-3-[(dimethylamino)methyl]-4゜6.7-dolihydro xy2,8-dioxabicycloC3,2,1)octane-4,5-dicarvone acid, 6-(4,6-simethyl-2-octenoate)acetonitrile (12ml ) The suspension of middle side 1 (250B) was stirred at room temperature under nitrogen, and added with glacial acetic acid (21 μL) and water. formaldehyde (280 μL of 40% solution) and triacetoxyborohydride Treatment with sodium hydroxide (235 mg) gave a clear solution. After 2 hours remove the solution Evaporated to dryness. The residue was stirred with ethyl acetate (~101) and filtered. filtrate Evaporate to a yellow foam, 5pheris.

Preparative HPLC using rb0DS-2 (2X 25 cm) column 70% acetonitrile in water containing trifluoroacetic acid (1 ml/L) at a flow rate of 1. Purification was achieved by elution at 5 ml/+in. Combine the required fractions and roll Concentration by tally evaporation followed by lyophilization afforded the title compound (1 89IIlg) was obtained; proton N, m, r, (dB-DMso) was 60. 75-0.9(II, CH), 0.99(d, J-GHz, -Ct(CHCH3 ), 2.1 (s, C) I CO), 2.81 (s, (CHa) 2 N), 3 ．． 99 (bd, J-3Hz, 7-H), 4.7 (d, J=9Hz, 3-)1), 4.92 (bs, C-(Ij2), 4.97 (d, J-5Hz, CHOAc ), 5.8 (d, J-16Hz, 0COCH-CH), 6.2 ( bs, 6-H), [i, 77 (dd, J-9 and 16Hz, 0CO CR-CO) + 7, 1-7, 3 (a, C6H5); Analysis actual values: C52,6; H6,2; N 1.9; F 9. ICH No. 1.5CHF 0 ・2H20 calculated value: 3753 1J 2 3 2 C52,7,HG,5. N 1.5; F 9.4%-3-methylene-6-phene nylhexyl)-3-(amidimethyl)-4,6,7-drihydroxy-2,8 -dioxabicycloC32,1]octane-4,5-dicarboxylic acid A solution of side 1 (100 rAg) in dimethylformamide (0.5 ml) was diluted with N-methyl Hydroxylamine hydrochloride (28a+g) and triethylamine (110μ L). The mixture was stirred at room temperature for 7 hours, then the solvent was removed by rotary evaporation. removed. The obtained white solid was treated with 5pherisorb 0DS-2 (2X 25 am) trifluoroacetic acid by Brenoglatative HPLC using a column. Dissolved in 65% acetonitrile in water containing acid (1 ml/L) at a flow rate of 15 ml/min. The necessary fractions were combined and subjected to rotary evaporation. It was concentrated and then lyophilized to a white solid. This can be done using the blurring method shown above. Nokulative HPL (contains trifluoroacetic acid (l ml/L)) Repurification using 35% acetonitrile in water gave the title compound (5%) as a white solid. 7 mg) was obtained; proton N, rn, r, (d 6-DMSO) 60.78 (d, J-6Hz, CHOH), 2.1 (s, ct13Co), 3 ．． 9 (bd, J-4Hz, 7-H), 4.48 (bd, J-6Hz, 3- H), 4.85 and 4, 91 (2s, C”CH2 and 6-H), 4.99 (d, J-5Hz, CHOAc), 7, 1-7, 3 (ToC Mass spectrum (MY 523.5) + FAB 524 (M H”).

Dihydroxy-2,8-dioxabicyclo (in 3,2,11 dioxane (21) A solution of 3 (56ig) in 7.5M aqueous chloride and hydrogen chloride was kept at room temperature for 8 hours, then Evaporated to dryness. Dissolve the residue in a small amount of ether and stir the solution until it becomes a white solid. Evaporated. Add this to 5pherisorb ODS-2 (2X25cm) column Containing trifluoroacetic acid (1 ml/L) was analyzed by pre-variative HPLC using 70% in water and then eluted with 95% acetonitrile at a flow rate of 15 sl/+in. It was purified by Concentrate the required fractions by rotary evaporation and then freeze Drying gave the title compound (22 mg) as a white solid; protons N, a+ , r, (dB-DMSO) is 60.7-0.9 (i, CH), 1.0 (d , J-GHz, -C)ICHC)Ia), 2.09(s, CHCO), 3 ．． 7 (s, c02C) 13), 3.97 (bd, J-5Hz, 7-H), 4.6 1(d, J-7)]z, :34), 4.91 (bs, C-C top 2), 4.99 (d, J-5) 1z, CHOAcd, J-9 and 16Hz, 0cOcH-CI ), 7.1-7.3 (s, C6H5).

Analysis actual value: C52, 1, H8, 1, N 2.1; F g, aCHNo, 1 ．． 3CHF O・3H20 calculated value: C52.0, H6.8, N 1.6:F 8 ．． After 2-3 weeks of growth at 25°C on 3% oatmeal agar, colonies The color was smoke gray-mouse gray on both the front and back sides (Ra yner's Mycol.

gical Co1our Chart, 1970; Co+mmonvealt h　Published by Agricultural Bureau).

Morphological observations of the strains grown on oatmeal agar at 25°C were performed under a light microscope.

Although the fungus did not have a sexual reproductive stage, it did form powder sporophores and thus Coe. It belongs to loBcetes. The fungus produces beak-like spore organs, and the hyphae are loose. The conidia were bright and had both septate and 1 septate conidia. Conidia are about 5-9 in size X1.5-3 μM (usually 7-9X1.50-2.5 μM). chlamydospore Numerous multiseptate/multicellular spherical structures resembling powder or sporophyte progenitor cells were also observed. Akira There are no definite reticulospores.

The isolate was identified as a species of the genus Phoma, and the identity was confirmed by CAB International. Confirmed by onal Mycologjcal In5 titute.

In vitro results The ability of the compounds of the present invention to inhibit the squalene synthase enzyme is determined by R, M, Ta1 t, Analyt, Biochem, 203, 310-316 (1992). (2-140]falnesyl diphosphate as substrate under the assay conditions listed. It was demonstrated using Inhibition of sphalene synthase is [2C]farnesyl diphosphate Incubate rat liver homogenate with various concentrations of test compound in the presence of It was quantified by

Concentration of compound showing 50% inhibition of squalene production (14c) in 30 minute assay was taken as the IC5o value.

In this test, the title compounds of Examples 1.2.5 and 7 have an I' of up to 1100n It had a Cso value.

Formulation example In the examples below, the term ``active ingredient'' refers to a compound of the invention, such as those mentioned in the examples above. Regarding the listed compounds.

Example 1 - Tablet a) Active ingredient 5. OB Lactose 95.0 g Microcrystalline cellulose 90. OB Crosslinked polyvinylpyrrolidone 8.0+gg magnesium stearate 2. hg Compressed weight: 200.0mg Active ingredients: microcrystalline cellulose, lactose and cross-linked polyvinylpyrrolidone. 00 micron sieve and blend in a suitable mixer. Magnesium stearate sieve through a 250 micron sieve and blend with the active blend. Apply brand/blind Compress into tablets or tablets using a sharp punch.

b) Active ingredient 5.0mg Lactose 165. hg Pre-gelatinized starch 20. hg Crosslinked polyvinylpyrrolidone 8.0+ng Magnesium stearate 2.0m g Compressed weight 200. hg The active ingredients, lactose and pre-gelatinized starch are blended together and prepared with water. Grainy. Dry the wet mass and grind. Magnesium stearate and cross-linked poly Vinylpyrrolidone is passed through a 250 micron sieve and blended with the granules. The obtained block Compress the blend using a suitable tablet punch.

Example 2 - Capsule a) Active ingredient 5.0mg Pre-gelatinized starch 193.0+ng Magnesium stearate 2.0μ g Filling weight 200.0+g The active ingredient and pre-gelatinized starch were passed through a 500 micron mesh sieve. blended with magnesium stearate (250 micron sieved) to become Fill the blend into appropriately sized hard gelatin capsules.

b) °Active ingredient 5.0mg Lactose 177.0mg Polyvinylpyrrolidone 8.0mg Crosslinked polyvinylpyrrolidone 8.0mg magnesium stearate 2. OB Filling weight 200.0mg The active ingredient and lactose are blended together and in a solution of polyvinylpyrrolidone. Granulate. Dry the wet mass and grind. Magnesium stearate and crosslinking The ribinylpyrrolidone is passed through a 250 micron sieve and blended with the granules. obtained Fill the blend into appropriately sized hard gelatin capsules.

Example 3 - Syrup a) Active ingredient 5.0mg Hydroxypropyl methylcellulose 45.0mg Propyl hydroxybenzoate 1.5B Butyl hydroxybenzoate 0.75mg Sodium saccharin 5. hg Sorbitol solution 1.01 Appropriate buffer qs Appropriate flavor qs Purified water total amount 10. up to hl Hydroxypropyl methylcellulose with hydroxybenzoate in thermally purified water and cool the solution to room temperature. Sodium saccharin, flavor and sorbitol solution to the bulk solution. Dissolve the active ingredient in some of the residual water and add to the bulk solution. Appropriate buffers control pH in the region of maximum stability. You can add it to make a difference. Adjust the solution to the desired volume, filter, and fill into a suitable container. Ru.

Active ingredient 0.1 Dextrose (anhydrous) 5.0 Benzalkonium chloride 0.02 Appropriate buffer 9s Purified water up to 100% Dissolve the active ingredient and dextrose in a portion of the bulk solution. A suitable buffer is May be added to control pH in the region of maximum stability. Adjust the solution to the specified volume. Strain, filter and fill into suitable containers.

On the other hand, solutions may be provided as sterile unit doses such that preservatives are omitted from the formulation. good.

b) Sterile solution without preservatives %v/v Active ingredient 0.1 Dextrose (anhydrous) 5.0 Appropriate buffer qs Purified water up to 100% , , PCT/EP 93100486 Front page continuation (51) Int, C1,' Identification symbol, internal office reference number A61K 31/3 4 AED 31/70 ADZ 9454-4C CO7H91028615-4C 15/10 8615-4C (81) Specified times EP (AT, BE, CH, DE.

DK, ES, FR, GB, GR, IE, IT, LU, MC, NL, PT, SE) , 0A (BF, BJ, CF, CG, CI, CM, GA, GN, ML, MR, SN , T.D.

TG), AT, AU, BB, BG, BR, CA, CH.

CZ, DE, DK, ES, FI, GB, HU, JP, KP, KR, LK, LU, MG, MN, MW, NL, N.

, NZ, PL, PT, RO, RU, SD, SE, SK.

UA, US FI (72) Inventor: Watson, Nizel Stephen Middlesepras, UK Greenford, Berkeley, Avenue (Street address) ), GRAKTS, GROUP, RESEARCH, LIMITED. (72) Inventors Procobio, Panayiotis Alexandrou Avenue, Middleseplace, Greenford, Berkeley, United Kingdom (street address) ), GRAKTS, GROUP, RESEARCH, LIMITED.

Claims (1)

[Claims] 1. A compound having the following formula (1) and a salt thereof. ▲There are mathematical formulas, chemical formulas, tables, etc.▼(1) [In the above formula, R1 is a hydroxyl group, Or [there is an array] or -OCOC H-ECHCH(CH3)(CH2)3CH3, -OCOCH-ECHC(CH 3) -ECHCH(CH3)CH2CH3 or -OCO-X-CH2CH(C H3) CH2CH3 (where X is -CH-ECHCH(CH3)-, -CH2 CH(OH)CH(CH3)-, -CH-ECHC(OH)(CH3)-, -C H2CH(OH)CH2- or -CH2CH2CH(CH3)) represents a selected group; R2 represents a hydroxyl group; R3 is ▲Contains mathematical formulas, chemical formulas, tables, etc.▼ (In the above formula, R8 is a hydrogen atom or an acetyl group), -C(CH3)=EC HCH(CH2R9)CH2Ph (where R9 is hydrogen or a hydroxyl group) ), -C(CH2H)-ZCHCH(CH3)CH2Ph, -C(-CH 2) CH(OH)CH(CH2OH)CH2Ph, -C(-CH2)CH(NH COCH3)CH(CH3)CH2Ph, -C(CH2NHCOCH3)-EC HCH (CH3) CH2Ph and ▲Mathematical formulas, chemical formulas, tables, etc.▼ represents a group selected from; R4 and R5 each independently represent a hydrogen atom or a methyl group; R6 and R7 each independently represent a hydrogen atom or a C1-4 alkyl group; or R6 represents a hydrogen atom, and R7 is a group -C(=Y)NH2 (here, Y is represents an oxygen or sulfur atom or NH]2. R4 and R5 are hydrogen atoms 2. A compound according to claim 1, which represents a child. 3. R1 is the following group: ▲Contains mathematical formulas, chemical formulas, tables, etc.▼ represents, 4. A compound according to claim 1 or 2. ▲Contains mathematical formulas, chemical formulas, tables, etc.▼ (wherein R8 is a hydrogen atom or an acetyl group) The compound described in any one of the above. 5. [1S-[1α (4R*, 5S*), 3α, 4β, 5a, 6a (2E, 4R *,6R*),7β]]1-(4-acetyloxy-5-methyl-3-methylene -6-phenylhexyl)-3-(aminomethyl)-4,6,7-trihydroxy C-2,8-dioxabicyclo[3.2.1]octane-4,5-dicarboxylic acid , 6-(4,6-dimethyl-2-octenoate) and its salts and protected derivatives. 6. 6. A compound according to any one of claims 1 to 5 for use in therapy. 7. Diseases in which lowering plasma cholesterol levels is beneficial in animals, including humans 6. A compound according to any one of claims 1 to 5 for use in the treatment of. 8. Claim 1: Used for the treatment of fungal infections in human or non-human animal patients. 5. The compound according to any one of . 9. Diseases associated with hypercholesterolemia and/or hyperlipoproteinemia treatment of a human or non-human animal individual to treat a fungal disease or to treat a fungal disease. A therapeutic method, A compound according to any one of claims 1 to 5 that inhibits squalene synthase. A method comprising administering to said individual an effective amount of a substance. 10. Any of claims 1 to 5 together with one or more carriers and/or excipients. A pharmaceutical composition comprising a compound according to item 1. 11. Reducing plasma cholesterol levels in animals, including humans, is beneficial The activity of a compound according to any one of claims 1 to 5 for use in the treatment of symptoms. 1. A pharmaceutical composition comprising: 12. The claim is for use in the treatment of fungal infections in human or non-human animal patients. 6. A pharmaceutical composition comprising an active amount of a compound as described in any one of 1 to 5. 13. Suitable for oral, buccal, topical, parenteral, implantable, rectal, ocular or genitourinary administration. or in a form suitable for administration by inhalation or insufflation. 2. The pharmaceutical composition according to any one of 2. 14. 14. A pharmaceutical composition according to any one of claims 10 to 13 in unit dosage form. 15. Hypercholesterolemia and/or in human or non-human animal patients Any one of claims 1 to 5 for the manufacture of a medicament for treating hyperriboproteinemia. USE OF THE COMPOUNDS DESCRIBED IN SECTION. 16. For the manufacture of medicaments for the treatment of fungal infections in human or non-human animal patients. , use of a compound according to any one of claims 1 to 5. 17. A method for producing the compound according to claim 1, comprising: (A) (for the production of a compound of formula (I) in which R6 and R7 both represent a hydrogen atom) (2) Compound of the following formula (ll): ▲ There are mathematical formulas, chemical formulas, tables, etc. ▼ (II) ( In the above formula, R1 to R3 have the same meanings as in claim 1, and R4a and R5a are protecting groups. or (B) converting a compound of formula (I) into a different compound of formula (I); Method. 18. A compound of formula (II). 19. A compound according to any one of claims 1 to 5 substantially as herein described. thing. 20. 15. A method according to any one of claims 10 to 14 substantially as herein described. Composition.