JP2680347B2 - Enzyme inhibitor compound - Google Patents

Description

TECHNICAL FIELD OF THE INVENTION The present invention relates to enzyme-inhibiting compounds, more particularly 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-C
oA) novel phosphorus-containing compounds useful in inhibiting the activity of reductase and thus in inhibiting cholesterol biosynthesis, hypocholesterolemic agents containing such compounds, novel intermediates formed in the manufacture of such compounds, and It relates to the use of such compounds for such purposes.

Prior art FM Singer et al .: Proc.Soc.Exper.Biol.Me
d.102, 370 (1959) and FH Hulche
r): Arch. Biochem. Biophys. 146, 422 (1971) discloses that certain mevalonic acid derivatives inhibit cholesterol biosynthesis.

Endo et al .: US Pat. Nos. 4,049,495, 4,137,322 and 3,983,140 disclose fermentation products that are active in inhibiting cholesterol biosynthesis. This product is called compactin and is called brown (B
rown) et al. (J. Chem. Soc. Perkin I., 1165 (1976)) have been reported to have a complex mevalonolactone structure.

GB No. 1,586,152 is [Wherein E represents a direct bond, a C _1-3 alkylene bridge or a vinylene bridge, and R having a suffix of each number represents various substituents].

The activity reported in this UK patent is less than 1% of that of compactin.

US Patent No. 4,375,475 to Willard et al. Has the formula: [In the formula, A is H or methyl; E is a direct bond, -CH _2- ,
_{-CH 2 -CH 2 -, - CH} 2 -CH 2 -CH 2 - or -CH = CH-;
R ₁ , R ₂ and R ₃ are each halogen, C _1-4 alkyl, C _1-4 haloalkyl, phenyl, substituted phenyl (substituents are halogen, C _1-4 alkoxy, C _2-8 alkanoyloxy, C _{1 to 4} alkyl or _{C 1 to 4} haloalkyl) and oR ₄ (R ₄ is _{H, C 2 to 8} alkanoyl, benzoyl, phenyl, Harofueniru, phenyl -
C _1-3 -alkyl, C _1-9 alkyl, cinnamyl,
C _1-4 haloalkyl, allyl, cycloalkyl-C
_1-3 -alkyl, adamantyl-C _1-3 -alkyl or substituted phenyl-C _1-3 alkyl (the substituents in each group are halogen, C _1-4 alkoxy, C
_{1 to 4} alkyl or C ₁ to ₄ haloalkyl))) a hypocholesterolemic and hypolipidemic compound represented by the following) and the corresponding dihydroxy formed from the hydrolytic ring opening of the lactone ring thereof. Of acids and pharmaceutically acceptable salts of the acids, and C _1-3 alkyl and phenyl, dimethylamino or acetylamino substituted C _1-3 -alkyl esters of the dihydroxy acid and the trans racemate of the above formula In the tetrahydropyran moiety
All compounds in the 4R configuration are disclosed.

WO 84/02131 (PCT / EP83 / 00308) (filing date November 1982
US Application No. 443,668 on 22nd and filing date November 4, 1983
(US application No. 548,850) (application in the name of Sandoz AG) has the formula: [Wherein one of R and R ₀ is The other is primary or secondary C _1-6 alkyl, C _3-6 cycloalkyl or phenyl- (CH ₂ ).
_m −, (in the group, R ₄ is hydrogen, C _1-4 alkyl, C _1-4 alkoxy (excluding t-butoxy), trifluoromethyl, fluoro, chloro, phenoxy or benzyloxy), R ₅ is hydrogen, C _1-3 alkyl, C _1-3 alkoxy,
Trifluoromethyl, fluoro, chloro, phenoxy or benzyloxy, R _5a is hydrogen, C _1-2 alkyl, C _1-2 alkyl,
C _1-2 alkoxy, fluoro or chloro, m is 1,
2 or 3, (wherein R ₅ and R _5a when R ₄ is hydrogen is hydrogen, when R ₅ is hydrogen R _5a is hydrogen, further at most one is trifluoromethyl R ₄ and R _5, R ₄ And at least one of R ₅ is phenoxy, and at least one of R ₄ and R ₅ is benzyloxy), R ₂ is hydrogen, C _1-4 alkyl, C _3-6 cycloalkyl, C _1-4 alkoxy (t -Excluding butoxy), trifluoromethyl, fluoro, chloro, phenoxy or benzyloxy, R ₃ is hydrogen, C _1-3 alkyl, C _1-3 alkoxy,
Trifluoromethyl, fluoro, chloro, R ₃ when phenoxy or benzyloxy (wherein R ₂ is hydrogen is hydrogen, further at most one is trifluoromethyl R ₂ and R _3, at most one of R ₂ and R ₃ Phenoxy, and R ₂ and R ₃
At most one of which is benzyloxy) of, X is - (CH _{2) n} - or -CH = CH- (n is 0, 1,
2 or 3), Z is (Wherein R ₆ is hydrogen or C _1-3 alkyl)] The heterocyclic analogue (free acid form) of mevalonolactone and its derivatives represented by: or physiologically hydrolyzed and physiologically acceptable Ester or its δ-
Lactones or salt-type compounds are disclosed.

GB No. 2162-179-A has the formula: [Wherein R ₁ is 1-3C alkyl; (Wherein R ₇ is any of H, a hydrolyzable ester group or a cation)], and a naphthyl analogue of mevalonolactone useful as a cholesterol biosynthesis inhibitor having a structure shown in .

European Patent No. 164-698-A discloses that an antihypercholesterolemic agent is prepared by treating an amide compound with an organic sulfonyl halide R ⁵ SO ₂ X as shown in the following formula, and then removing the protecting group Pr. Discloses a method for producing lactones useful as: [Wherein X is halo; Pr is carbinol-protecting group; R ¹ is H or CH ₃ ; R ³ and R ⁴ are H, 1-3 C alkyl or phenyl- (1
~3C alkyl) (1-3C alkyl if the phenyl is optionally may be substituted by 1-3C alkoxy or halo); R ² is the formula: A group represented by (wherein Q is R ⁶ R ⁶ is H or OH, R═H or CH ₃ , a, b, c or d is a double bond which may be optionally present, R ⁷ is phenyl or benzyloxy (the ring in each case is optional) 1 to 3C alkyl or halo), R ⁸ and R ⁹ are 1 to 3C alkyl or halo); R ⁵ is 1 to 3C alkyl, phenyl or mono- or di- (1 to 3C) alkyl. ) Represents phenyl].

Anderson, Paul Leroy
eroy) (German published DE 3,525,256) has the formula: [Wherein R ¹ is alkyl, Z is Q or Q ¹ , and Q is R ⁷ is a hydrolyzable ester group useful as an inhibitor of H or cholesterol biosynthesis and in the treatment of atherosclerosis].] And discloses naphthyl analogs of the mevalonolactones.

WO No. 8402 filed in the name of Sandoz AG
-903 (US Application No. 460,600, filed Jan. 24, 1983) has the formula: [In the formula, two R ₀ groups are formed by combining these. (In the group, R ₂ is hydrogen, C _1-4 alkyl, C _1-4 alkoxy (excluding t-butoxy), trifluoromethyl, fluoro, chloro, phenoxy or benzyloxy, R ₃ is hydrogen, C _1-3 Alkyl, C _1-3 alkoxy,
Trifluoromethyl, fluoro, chloro, phenoxy or benzyloxy (provided that at least one of R ₂ and R ₃ is trifluoromethyl, at least one of R ₂ and R ₃ is phenoxy,
And at least one of R ₂ and R ₃ is benzyloxy), R ₁ is hydrogen, C _1-6 alkyl, fluoro, chloro or benzyloxy, R ₄ is hydrogen, C _1-4 alkyl, C _1-4 alkoxy. (Excluding t-butoxy), trifluoromethyl, fluoro, chloro, phenoxy or benzyloxy, R ₅ is hydrogen, C _1-3 alkyl, C _1-3 alkoxy,
Trifluoromethyl, fluoro, chloro, phenoxy or benzyloxy, R _5a is hydrogen, C _1-2 alkyl, C _1-2 alkoxy, fluoro or chloro (provided that at least one of R ₄ and R ₅ is trifluoromethyl, R 5 _Four
And at least one of R ₅ is phenoxy, and at least one of R ₄ and R ₅ is benzyloxy), X is — (CH ₂ ) _n —, (Wherein n is 0, 1, 2 or 3, both q are 0 or one of them is 0 and the other is 1), (Wherein R ₆ is hydrogen or C _1-3 alkyl), (provided that the phenyl groups having —XZ and R ₄ substituents are in the ortho position relative to each other). Disclosed are mevalono-lactone analogs (free acid form) or physiologically hydrolyzed and physiologically acceptable esters or δ-lactones or salts thereof useful as proteinemia agents.

European Patent Application No. 127,848-A (Merck C
o., Inc.)) is the formula: n is 0, 1 or 2; E is _{-CH 2 -, - CH 2 -CH} 2, -CH 2 -CH 2 -CH 2 -, - CH =
CH-CH ₂ - or _{-CH 2 -CH = CH-; R 1} , R 2 and R ₃ each for example hydrogen, chloro, bromo, fluoro, C ₁ - alkyl, phenyl, substituted phenyl or OR ₇ (R ₇ is For example, hydrogen, C _2-8 alkanoyl, benzoyl, phenyl, substituted phenyl, C _1-9 alkyl, cinnamyl, C _1-4 haloalkyl, allyl,
Cycloalkyl-C _1-3 -alkyl, adamantyl-
C _1-3 -alkyl or phenyl-C _1-3 -alkyl; R ⁴ , R ⁵ and R ⁶ are each hydrogen, chloro, bromo, fluoro or C _1-3 alkyl; X is, for example, hydrogen, C _1-3 alkyl Is a cation derived from an alkali metal or ammonium] 3-hydroxy-5-thia-ω
-Allyl-alkanoic acid derivatives are disclosed.

These compounds have antihypercholesterolemic activity based on the inhibitory activity of 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase and antibacterial activity.

French patent application No. 2,596,393A (filing date 1986 April
On the 1st of January, Sanofi SA) has the formula: Wherein R ₁ and R ₂ are each H, lower alkyl or optionally substituted aralkyl; R ₃ and R ₄ are each H, lower alkyl or optionally substituted aryl or Which is useful as a hypolipidemic agent represented by
-Hydroxy-propane-phosphonic acid derivatives, including salts thereof, are disclosed.

These compounds have the effect of reducing cholesterol, triglycerides and phospholipids, meglitol.
utol) is disclosed.

European Patent Application No. 142,146-A (Merck Co., Inc.)
formula: [In the formula, R ¹ is, for example, hydrogen or C _1-4 alkyl; E is —CH ₂ CH ₂ —, —CH═CH— or — (CH ₂ ) _r —; and Z is the formula (1): [In the formula, X is -O- or -NR ⁹ (R ⁹ is hydrogen or C
_1-3 alkyl), R ⁷ is C _2-8 alkyl, R ⁸ is hydrogen or CH ₃ , [Wherein R ¹⁰ , R ¹¹ and R ¹² are each independently hydrogen, halogen or C _1-4 alkyl], And a mevinolin-like compound represented by the formula

According to the present invention, the formula: [In the Formula, X is -O- or -NH-, n is 1 or 2, Z
Represents a lipophilic group], and the enzyme 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMG-C
The present invention provides a phosphorus-containing compound which is useful as a hypocholesterolemic agent because it inhibits oA reductase).

In the present invention, the lipophilic group represented by Z in the present invention is used for binding the substrate HMG-CoA by binding to the enzyme pocket when binding to the upper side chain of the HMG-like molecule by its appropriate binding moiety. , And as a result, it is a lipophilic group whose activity is enhanced compared to compounds where Z = H.

The compound of the present invention [Wherein R represents OH, lower alkoxy or lower alkyl; R ^X represents H or lower alkyl; X represents -O- or -NH-; n represents 1 or 2; Z represents a lipophilic group; It includes compounds (including salts thereof) and pharmaceutically acceptable salts thereof.

Salts and salts refer to basic salts formed with inorganic and organic bases. Such salts include alkali metal salts such as ammonium salts, lithium salts, sodium salts and potassium salts (which are preferred), calcium salts,
Alkaline earth metal salts such as magnesium salts, and amine salts (for example, dicyclohexylamine salt, benzathine salt, N-methyl-D-glucamine salt, hydrabamine salt), salts with amino acids (eg arginine salt, lysine salt, etc.) Such salts with organic bases are included. Non-toxic and pharmaceutically acceptable salts are preferred, but other salts are also useful, eg for isolating or purifying the product.

Examples of the lipophilic group (Z) in the present invention include the following groups. In addition, "alkyl", "lower alkyl" or "alkali metal" in the groups or formulas shown below is respectively translated into alkyl, lower alkyl or alkali metal.

[In the formula, a dotted line represents a double bond which may be present if necessary] For example, [Wherein R ¹ , R ² , R ^2a and R ^2b are the same or different and each independently represents H, halogen, lower alkyl,
Haloalkyl, phenyl, substituted phenyl or OR ^y (R ^y
Represents H, alkanoyl, benzoyl, phenyl, halophenyl, phenyl (lower) alkyl, lower alkyl, cinnamyl, haloalkyl, allyl, cycloalkyl (lower) alkyl, adamantyl (lower) alkyl or substituted phenyl (lower) alkyl].

Z is R ⁵ and R ⁵ ′ are the same or different and are H, lower alkyl or OH; R ⁶ is lower alkyl Or aryl -CH ₂ -; R ^6a is lower alkyl, hydroxy, oxo or halogen, q is 0, 1, 2 or 3; R ⁷ represents H or lower alkyl.

Therefore, the compound [I] is Embedded image

As used herein, the term lower alkyl or alkyl, whether alone or as a part of another group, has 1 to 12 carbon atoms (preferably 1 to 7) in a straight chain or Branched hydrocarbon groups such as methyl, ethyl, propyl, isopropyl, butyl, t-butyl, isobutyl, pentyl, hexyl, isohexyl, heptyl, 4,
4-dimethylpentyl, octyl, 2,2,4-trimethylpentyl, nonyl, decyl, undecyl, dodecyl, various branched chain isomers thereof, and the like, and substituents such as F, Br, Cl or I or CF ₃
Halo-substituents, alkoxy substituents, aryl substituents, alkyl-aryl substituents, haloaryl substituents, cycloalkyl substituents, alkyl-cycloalkyl substituents, hydroxy and alkylamino substituents, alkanoylamino substituents, Included are alkyl containing aryl-carbonylamino, nitro, cyano, thiol or alkylthio substituents.

Cycloalkyl in the present specification includes a saturated cyclic hydrocarbon group having 3 to 12 carbon atoms (preferably 3 to 8 carbon atoms), whether alone or as a part of another group, and these are For example, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, cyclodecyl and cyclododecyl, and any of these substituents such as 1-2 halogens, 1-2 lower alkyl groups, 1- 2 lower alkoxy groups, 1 to 2 hydroxy groups, 1 to 2 alkylamino groups, 1 to 2 alkanoylamino groups, 1 to 2
Arylcarbonylamino groups, 1-2 amino groups, 1-2 nitro groups, 1-2 cyano groups, 1-2
Thiol groups and / or alkyl having substituents such as 1-2 alkylthio groups.

As used herein, aryl or Ar is a monocyclic or bicyclic aromatic group containing 6 to 10 carbon atoms in the ring portion, such as phenyl, naphthyl, substituted phenyl or substituted naphthyl (on the phenyl group or naphthyl group). Any of the above substituents may be 1, 2 or 3 lower alkyl groups, halogen (Cl, Br or F), 1, 2 or 3 lower alkoxy groups, 1, 2 or 3 hydroxy groups, 1 2 or 3 phenyl groups, 1, 2 or 3 alkanoyloxy groups, 1, 2 or 3 benzoyloxy groups, 1,
2 or 3 haloalkyl groups, 1, 2 or 3 halophenyl groups, 1, 2 or 3 allyl groups, 1, 2 or 3 cycloalkylalkyl groups, 1, 2 or 3
Adamantyl alkyl groups, 1, 2 or 3 alkylamino groups, 1, 2 or 3 alkanoylamino groups, 1, 2 or 3 arylcarbonylamino groups,
1, 2 or 3 with 1, 2 or 3 amino groups, 1, 2 or 3 nitro groups, 1, 2 or 3 cyano groups, 1, 2 or 3 thiol groups and / or aryl groups It means 3 (preferably containing 3 substituents) alkylthio groups.

As used herein, aralkyl, aryl-alkyl or aryl (lower) alkyl means a lower alkyl group (eg benzyl) as defined above having an aryl substituent, whether alone or as part of another group. To do.

As used herein, lower alkoxy, alkoxy or aryl or aralkoxy means a lower alkyl, alkyl, aralkyl or aryl group as defined above attached to an oxygen atom, whether alone or as part of another group. .

In the present specification, lower alkylthio, alkylthio, arylthio or aralkylthio is any of the above lower alkyl, alkyl, aralkyl or aryl groups bonded to a sulfur atom, whether alone or as a part of another group. Means

Lower alkylamino, alkylamino in the present specification,
Arylamino, arylalkylamino, either alone or as part of another group, refers to any of the above lower alkyl, alkyl, aryl or arylalkyl group attached to a nitrogen atom.

Alkanoyl, as used herein, refers to a lower alkyl, as defined above, attached to the carbonyl group, whether alone or as part of another group.

Halogen or halo in the present specification means base, bromine, fluorine, iodine and CF ₃ (chlorine and fluorine are preferable).

Compound [I], preferably of formula: [Where R is OH and OLi; R ^X is Li or H; X is O or NH; (Wherein R ¹ is phenyl including alkyl and / or halo substituents, or R ¹ is benzyloxy including halo substituents, R ² and R ^2a are the same and are halogen or lower alkyl), Z is preferably (Wherein R ¹ and R ² have the same meanings as described above for the compound [II]), or Z is (Here, R ⁵ is H, CH ₃ or OH, R ⁶ is Or (substituted) phenylmethyl (wherein R ⁷ is H or CH
₃ ))]] is preferred.

The compound [I] of the present invention can be produced by the reaction steps shown below and the subsequent description.

As is apparent from the above reaction step, compound [I] can be produced as described in detail below.

And a phosphonite / phosphite represented by the formula: R _a -P (O-alkyl) ₂ [III] [wherein R ^a represents lower alkyl or lower alkoxy] under standard Arbuzov reaction conditions. The iodide [A] is subjected to an Arbuzov reaction by heating to the formula: The phosphinate / phosphonate represented by

This phosphinate / phosphonate [IV] is a novel compound and forms a part of the present invention.

Then a solution of this compound [IV] in an inert organic solvent (eg methylene chloride) is subsequently treated with bis (trimethylsilyl) trifluoroacetamide (BSTFA) and trimethylsilyl bromide (under an inert atmosphere (eg argon)). The phosphinate / phosphonate [IV] is cleaved with a phosphoric ester to give a phosphinic acid [VA] (a product when R ^{a of} formula [IV] is lower alkyl), that is, the formula: Or a phosphonic acid [VB] (formula [IV]
Where R ^a is lower alkoxy) ie the formula: Is obtained.

The compounds [VA] and [VB] are novel intermediates and form part of the present invention.

When phosphonic acid [VB] was obtained, this compound [VB]
Is treated in dry pyridine with an alcohol of the formula: R ^b OH [VC] where R ^b represents lower alkyl and dicyclohexylcarbodiimide and the mixture is stirred under an inert atmosphere (eg argon). Esterified by the formula: To form a monoalkyl phosphonate represented by

The ester [VI] or phosphinic acid [VA] is then dissolved in an inert organic solvent (e.g. methylene chloride, benzene or tetrahydrofuran (THF)), treated with trimethylsilyldiethylamine and stirred under an inert atmosphere (e.g. argon), The mixture is evaporated and dissolved in methylene chloride (or other suitable inert organic solvent). The solution is cooled to about 0-25 ° C, treated with oxalyl chloride, and evaporated to give crude phosphonochloridate. This phosphonochloridate is dissolved in an inert organic solvent such as methylene chloride, benzene, pyridine or THF, the solution is cooled to about -20 to 0 ° C. and the compound [VI] or [VA] and the formula: Z- ( CH ₂ ) _n —XH Molar ratio of the compound represented by [B] ([VI] or [VA]:
[B]) (about 0.5: 1) to (about 3: 1), preferably (about 1: 1).
1)-(about 2: 1) compound [VI] or [VA] is treated with compound [B], followed by triethylamine and catalytic amount of 4-dimethylaminopyridine (DMAP),
formula: [Wherein R ^c represents lower alkyl or lower alkoxy] to form an addition product.

The obtained compound [VII] is treated with glacial acetic acid and tetrabutylammonium fluoride in an inert organic solvent such as tetrahydrofuran to give the compound [VI].
I] is cleaved with a silyl ether to give the formula: To obtain an ester form (wherein R ^X is alkyl).

Then, the ester form [VIII] is hydrolyzed to produce a corresponding alkali metal salt or acid (that is, a compound in which R ^X is an alkali metal or H, respectively). That is, the ester form [VIII] is added in the presence of dioxane, tetrahydrofuran or another inert organic solvent in an inert atmosphere such as argon at a molar ratio of strong base: ester [VIII] of (about 1: 1) to ( About 1.1: 1) and treated with the strong hydrochloric acid (eg lithium hydroxide) at 25 ° C., formula: To form the corresponding alkali metal salt, wherein R is lower alkyl or lower alkoxy.

By treating the above compound [VIII A] with a strong acid such as hydrochloric acid, the compound of formula: A corresponding acid represented by the formula (Compound [I] of the present invention wherein R ^X is H) can be obtained.

Next, R is a lower alkoxy ester compound [VIII]
Is a strong base: ester [VIII] molar ratio (about 2: 1) to
(About 4: 1) by treatment with the strong group at 50-60 ° C. to give the formula: Can be converted to the corresponding dialkali metal salt represented by.

By treating this dialkali metal salt [VIII C] with a strong acid such as hydrochloric acid, the formula: The compound (I) of the present invention wherein R is OH and R ^X is H can be converted to the corresponding acid represented by

The iodide [A] starting material can be produced using bromide [C] as follows. formula: (Produced by the method described in Tetrahedron Lett. Vol. 26, page 2951 (1985)) was dissolved in dimethylformamide (DMF) together with imidazole and 4-dimethylaminopyridine, and the solution was diluted with argon. Treated with t-butyldiphenylsilyl chloride in an inert atmosphere and of the formula: To form a silyl ether form. By treating a solution of this silyl ether compound [D] in an inert organic solvent (for example, methyl ethyl ketone or DMF) with sodium iodide under an inert atmosphere such as argon,
Iodide [A] can be produced.

Then the formula: Z- (CH ₂₎ starting materials represented by _n -XH (B) can be produced by the method described accordance connexion defined below Z and X.

Z is The compound [B] in which X is O, that is, the formula: The compound represented by the formula: It can be produced by treating the aldehyde derivative represented by with a reducing agent such as lithium aluminum hydride or sodium borohydride.

Z is The compound [B] in which X is NH, that is, the formula: The compound having the structure represented by can be produced as follows. The aldehyde form [E] is oxidized in solution with acetone, for example by the use of the Djones reagent to oxidize the aldehyde form to give the formula: To form the corresponding acid chloride in suspension with methylene chloride, which is dissolved in an inert organic solvent such as tetrahydrofuran, Treated with a mixture of ammonium and tetrahydrofuran, The corresponding amino compound [B ² ] can be obtained by producing an amide compound represented by and reducing this with a reducing agent such as lithium aluminum hydride.

The compound [B] in which X is O or —NH—, that is, the formula: Among the compounds represented by the formula, compounds in which X is O are those described by CHHeathcock et al. (J. Org. Chem. Vol.
Page (1985)). The compound [H ′] in which X is NH has the formula: A compound of formula (made as disclosed by Heskotsk et al., Supra) by reductive amination with ammonium acetate and sodium cyanoborohydride in the presence of an alcohol solvent such as methanol. You can

A starting material [B] in which X is O, ie of the formula: The compounds represented by are disclosed in both WO 8402-903-A and GB 2,162,179A filed in the name of Sandoz.

The starting material [B] where X is NH, ie the formula: The compound represented by the formula: The aldehyde derivative represented by can be produced by treatment with ammonium acetate and sodium cyanoborohydride in the presence of an alcohol solvent such as methanol for reductive amination.

The compound of the present invention can be produced as a racemic mixture and then resolved to obtain a preferred S-isomer.
However, the compounds of the present invention can be prepared directly in their S-isomeric form, as described above and as shown in the Examples below.

The compounds of the present invention are inhibitors of 3-hydroxy-3-methyl-glutaryl-coenzyme A (HMG-CoA) reductase and are therefore useful for inhibiting cholesterol biosynthesis as demonstrated in the tests described below. Is.

1. Rat liver HMG-CoA reductase The method described by Edwards (Edwards, PA et al.,
J. Lipid Res. 20:40 (1979)) is used to measure rat liver HMG-CoA reductase activity. Using rat liver microsomes as the enzyme source, the enzyme activity is measured by measuring the conversion amount of ¹⁴ C-HMG-CoA substrate into ¹⁴ C-mevalonate.

Preparation of microsomes Liver was separated from 2 to 4 Sprague Dawley rats decapitated by administration of cholestyramine, and phosphate buffer A (potassium phosphate 0.04M, pH 7.2; potassium chloride 0.05M; sucrose 0.1M; EDTA 0.03M; aprotinin 500 KI units / ml). The homogenate is spun at 16,000 xg for 15 minutes at 4 ° C. Separate the supernatant and centrifuge again under the same conditions. The second supernatant, which was centrifuged at 16,000xg, was stored at 4 ℃ for 10
Spin at 0,000xg for 70 minutes. The pelleted microsomes are resuspended in a minimum amount (3-5 ml / liver) of buffer and homogenized in a glass / glass homogenizer. Dithiothreitol was added (10 mM) to divide the product into unit fractions,
Quick-freeze in acetone / dry ice and store at -80 ° C. The specific activity of the first microsome preparation
ctivity) was 0.68 nmole mevalonate / mg protein / min.

b. Enzyme test Test the reductase in 0.25 ml of a solution containing the following final concentrations: 0.04M potassium phosphate pH 7.0 0.05M potassium chloride 0.10M sucrose 0.03M EDTA 0.01M dithiothreitol 3.5. mM Sodium chloride 1% Dimethyl sulfoxide 50-200 μg Microsome protein 100 μM ¹⁴ C- [DL] HMG-CoA (0.05 μCi, 30-60 mCi /
2.7 mM NADPH (nicotinamide adenine dinucleotide phosphate) The reaction mixture is aged at 37 ° C. Under the conditions described, the enzymatic activity increases linearly up to 300 μg microsomal protein per reaction mixture and linearly up to 30 minutes with respect to aging time. The standard aging time selected for drug research is
After 20 minutes, this results in a 12-15% conversion of the HMG-CoA substrate to the mevalonate product. [DL at 100 μM twice the concentration required to saturate the enzyme under the conditions described
-] HMG-CoA substrate is used. NADPH is used in a 2.7-fold excess over the concentration required to achieve maximum enzyme rate.

A standard test for inhibitor evaluation is performed according to the following procedure. 37 in the presence of NADPH
Aged for 15 minutes at ℃. DMSO vehicle is added with or without the test compound and the mixture is aged for a further 15 minutes at 37 ° C. ^The enzyme test is started by adding ¹⁴ C-HMG-CoA substrate. After aging for 20 minutes at 37 ℃, add 25μ of 33% potassium hydroxide to stop the reaction. ³ H-mevalonic acid (0.05 μCi) is added and the reaction mixture is left at room temperature for 30 minutes. Mevalonic acid is lactonized by adding 50 μ of 5N hydrochloric acid. Bromophenol blue is added as a pH indicator to monitor appropriate pH drop. Allow the lactonization to proceed for 30 minutes at room temperature. The reaction mixture is centrifuged at 2800 rpm for 15 minutes. The supernatant is poured into 2 g of AG1-X8 anion exchange resin (Biorad formic acid form) on a 0.7 cm (id) glass column and separated by elution with 2.0 ml of water. Discard the first 0.5 ml,
The next 1.5 ml was collected and added to tritium and carbon-14 in 10.0 ml of Opti-fluor scintillation fluid.
Is counted. Results are calculated as nanomoles mevalonate produced per 20 minutes and corrected to 100% recovery of tritium.
Drug effects are expressed as I ₅₀ values (drug concentration that inhibits enzyme activity by 50%) derived from pooled dose response data with specified 95% confidence intervals.

The process of converting the lactone type drug into its sodium salt is performed as follows. The lactone form was dissolved in DMSO, a 10-fold molar excess of sodium hydroxide was added, and the mixture was stirred at room temperature for 15 minutes.
Leave for a minute. The mixture is then partially neutralized (pH 7.5-8.0) with 1N hydrochloric acid and diluted in the enzyme reaction mixture.

2. To demonstrate the inhibitory activity of the compounds against cholesterol-synthesizing HMG-CoA reductase in freshly isolated rat hepatocytes, the method initially described by Capuzzi et al. (Capuzzi, DM) and Margolis, S. .): Lipids 6: 602 (1971)) is used to evaluate the compound activity of inhibiting ¹⁴ C-acetate complex cholesterol in freshly isolated rat hepatocyte suspensions.

Isolation of rat hepatocytes Sprague-Dawley rat (180-220 g) is anesthetized with Nembutol 50 mg / kg. An incision is made in the abdomen and the first branch of the portal vein is ligated. Heparin (100-200
Unit) directly into the abdominal vena cava. A single closed suture is applied to the terminal portion of the portal vein, and the portal vein is inserted between the suture line and the first branch vein. After cutting the vena cava to allow the drainage of effluent, buffer solution A (HBS containing calcium or magnesium deficient 0.5 mM EDTA and preheated (37 ° C) and oxygenated)
S) is sprinkled on the liver at a rate of 20 ml / min. Furthermore, heated buffer B (HBSS containing 0.05% bacterial collagenase) 200 m
Disperse l on the liver. After spraying buffer solution B, remove the liver,
Free cells are dispersed in 60 ml of Waymouth medium by stripping the medium. Hepatocytes are isolated by low speed centrifugation at 50xg for 3 minutes at room temperature. The pelleted hepatocytes are washed once in the weight-mouse medium and the viability is calculated and tested by trypan exclusion. These hepatocyte-rich cell suspensions usually show 70-90% viability.

b. Conjugation of ¹⁴ C-acetate to cholesterol Hepatocytes were aged at 5 x 10 ⁶ cells / 2.0 ml in a medium (IM) [0.02M].
Tris-hydrochloric acid (pH7.4), 0.1 M potassium chloride, 3.3 mM sodium citrate, 6.7 mM nicotinamide, 0.23 mM NAD
P, 1.7 mM glucose-6-phosphate]. The test compound is routinely treated with DMSO or DMSO: water (1:
Dissolve in 3) and add to IM above. The final DMSO concentration in IM is
At 1.0%, it has no significant effect on cholesterol synthesis.

¹⁴ C-acetate (58 mCi / mmol, 2 μCi / ml) was added to disclose aging, and 2.0 ml of this cell suspension was placed in a 35 mm tissue culture pot at 37 ° C. for 2.0 hours. After aging, the cell suspension is transferred to a glass centrifuge tube and spun at 50 xg for 3 minutes at room temperature. Resuspend and lyse the cell pellet in 1.0 ml of water,
Place on ice bath.

Substantially Bligh, EG and WJDy
er) [Can.J.Biochem.and Physiol.Vol.37: 911
(1959)] to extract lipids. The lower organic layer is separated, dried under a stream of nitrogen and the residue is resuspended in 100 μl chloroform: methanol (2: 1). Drop all of this sample onto a silica gel (LK6D) thin layer plate,
Expand to hexane: ethyl ether: acetic acid (75: 25: 1). Plates are scanned and counted using the BioScan automated scanning system. Cholesterol peak (RF0.
The radiolabel of 28) was measured and expressed as% of label of total lipid extract in total counts per peak. Cholesterol peaks in control cultures usually contained 800-1000 cpm, 9-20% of the label present in the total lipid extract, and the above results showed 9% of the extracted cholesterol labels, the values of Kaputui et al. Does not conflict with.

The drug effect (% inhibition of cholesterol synthesis) is measured by comparing the% cholesterol labeled in control and drug-treated cultures. Plot a dose-response curve from aggregated data from two or more studies,
Results are expressed as I ₅₀ values with 95% confidence intervals.

3. Cholesterol synthesis in human skin fibroblasts The compound selectivity favoring the greater inhibitory activity of liver tissue is a contributing factor for inhibitors of cholesterol synthesis. Therefore, in addition to evaluating inhibitors of cholesterol synthesis in hepatocytes, it is necessary to test the compounds of the present invention for activity as inhibitors of cholesterol synthesis in cultured fibroblasts.

Human skin fibroblast culture Human skin fibroblasts (passage cells 7 to 27) were cultured in 10% fetal stage calf serum-containing Eagle (Eagle) minimum essential medium (E).
M) grow in. In each test, the stock cultures were trypsonized and dispersed into cell monolayers,
Counted to be the plate in 35mm tissue culture vessel (5 × 10 ⁵ cells /2.0ml). The culture is incubated at 37 ° C. for 18 hours in 5% carbon dioxide / 95% hygroscopic chamber air. Remove serum-containing medium, wash cell monolayer, fatty acid-free bovine serum albumin 1.0%
Cholesterol biosynthesis enzyme is induced by adding 1.0 ml of EM containing broth and culturing the culture for another 24 hours.

b. Wash ^14C- acetate complex-induced cholesterol fibroblasts with EMEM ₁₀₀ (Eagle's minimal essential medium). The test compound was dissolved in DMSO or DMSO: EM (1: 3) (final DMSO concentration in cell culture was 1.0%), this was added to the culture, and this was added in 5% carbon dioxide / 95% humidified chamber air, 3 at 37 ° C
Pre-incubate for 0 minutes. After pre-culture, ^[1-14 C] sodium acetate (2.0μCi / ml, 58mCi / mmol) was added and 4 hours recultivation cultures. After culturing, the culture medium is removed and the cell monolayer (200 μg cellular protein per culture) is scraped into 1.0 ml of water. Lipids in the lysed cell suspension are extracted into chloroform / methanol as described above for hepatocyte suspensions. The organic layer was dried under a nitrogen atmosphere, the residue was resuspended in 100 μm of chloroform: methanol (2: 1), and the whole sample was dropped on a silica gel (LK6D) thin layer plate and analyzed as described above for hepatocytes. To do.

The inhibitory effect on cholesterol synthesis is tested by comparing the% cholesterol peak labeling from control and drug treated cultures. Results are expressed as I ₅₀ values and are derived from the cumulative dose response curves obtained from 2 or more trials. The 95% confidence interval for the I ₅₀ value is also calculated from the cumulative dose response curve.

Yet another embodiment of the present invention is compound [I] at least 1
A pharmacological composition containing a seed in combination with a pharmaceutical carrier or diluent. Such pharmacological compositions may be prepared using a conventional diluent, a solid or liquid carrier, and a pharmacological adduct of a type suitable for a desired administration method. The active compound is administered orally in the form of tablets, capsules, granules or powders, or the active compound is administered parenterally in the form of injections, for example in dosage units for use in medical treatment. It can be administered in a dosage form containing 1 to 2000 mg of active compound per dose. The dose depends on the dose, symptoms, age and weight of the patient.

The compound [I] is a known compound suggested for use in suppressing cholesterol biosynthesis in humans, dogs, cats and other mammals, such as lovastati.
It can be administered by the same method as the administration method of n).
Therefore, the compound of the present invention is a single-dose type or a single-dose type of about 4-2000 mg per day, preferably 4-200 mg per day or as a sustained release type (divided type 1-100 mg),
Suitably, 0.5 to 50 mg can be administered as a dosage form divided into 2 to 4 times.

The following examples represent preferred embodiments of the invention. Unless otherwise specified, temperatures are expressed in ° C. Flash chromatography was performed on either Merck 60 or Whatmann LPS-1 silica gel. Reversed-phase chromatography is Mitsubishi (Mitsubish
i, Ltd.) on a CHP-20 MCI gel.

Example 1 (S) -4-[[[4'-fluoro-3,3 ', 5-trimethyl [1,1'-biphenyl] -2-yl] methoxy]
Methoxyphosphoryl] -3-hydroxy-butanoic acid monolithium salt AN- (2,4-dimethylbenzylidene) benzenamine (see Merck U.S. Pat. No. 4,375,475 page 39) Freshly distilled 2,4-dimethylbenzaldehyde ( Ald
rich) 6.97 ml (50 mmol) and distilled aniline (Aldr
ich) 4.56 ml (50 ml) of 80.0 ml of dry toluene is refluxed in a flask equipped with a Dean-Stark apparatus for 3.0 hours under an argon atmosphere. The mixture is cooled and then evaporated under reduced pressure to give a yellow oil. This crude oil was purified by Kugelrohr distillation (0.5 mmHg, 160-180 ° C) and the light yellow oily product was left to crystallize to give 8.172 g of the desired title benzenimine as a low melting solid ( 78.1
%). TLC: hexane-acetone (4: 1), RF = 0.6
7 and 0.77 (geometric isomers), UV and I ₂ .

(See Merck U.S. Pat. No. 4,375,475, page 39) In 144 ml of glacial acetic acid, 6.0 g of the benzeneimine derivative of the above-mentioned item A (2
8.7 mmol) mixture of palladium (II) acetate 6.44 g
(28.7 mmol) and the clear, homogeneous red solution is refluxed under argon for 1 hour. This cloudy mixture is warmed and passed through a Celite-filled (1/2 ″) bed with 900 ml of water.
Have inside. The orange solid that precipitated was collected and dried under vacuum over phosphorus pentoxide at 65 ° C for 16.0 hours to give a melting point of 194-196.
Desired title palladium complex 1 as an orange solid at ℃ 1
0.627 g (85.5%) is obtained (melting point of recrystallized analytical sample described in literature is 203-205 ° C.).

C. 4'-Fluoro-3,3 ', 5-trimethyl [1,1'-biphenyl] -2-acetaldehyde 1.Bromo [4-fluoro-3-methylphenyl] magnesium (Merck U.S. Pat. No. 4,375,475 37- (See page 38) The Grignard reagent of the section C1. Is prepared as follows. Magnesium slice 1.35 in dried ethyl ether 70.0 ml
To g (55.4 mmol, 80 equivalents), 2-bromo-2-fluorotoluene (Fairfield Chemical Co.) 22.5 g (60.9
Mmol) is added dropwise with stirring at a rate sufficient to keep the mixture at reflux. Start the reaction in an ultrasonic device. After completion of the bromide addition, the mixture is stirred at room temperature for 1 hour under an argon atmosphere, refluxed for 15 minutes, and then cooled to room temperature.

2.4'-Fluoro-3,3 ', 5-trimethyl [1,1'-biphenyl] -2-acetaldehyde In 100 ml of dry benzene, the dipalladium complex of the above item B is used.
3.0 g (6.92 mmol) and triphenylphosphine 1
A mixture of 4.25 g (55.4 mmol, 8.0 eq) is stirred for 30 minutes at room temperature under an atmosphere of argon. To this solution is added freshly made (glass wool-stuffed) Section C1 Grignard reagent all at once with a cannula and the mixture is stirred at room temperature under an argon atmosphere for 1.5 hours. 6.0N hydrochloric acid 35ml
And the mixture is stirred at room temperature for another hour and then passed through a packed Celite (1/2 ″) bed. The solution is extracted with 250 ml of diethyl ether and the extract is extracted with sodium chloride (100 ml × 2).
Wash once, dry over anhydrous magnesium sulphate and evaporate under reduced pressure 13.35 g of a viscous orange oil which is left to crystallize. The crude orange solid is purified by flash chromatography on 700 g of silica gel, eluting with hexane then hexane-diethyl ether (95: 5). The product fractions were evaporated to give 1.507 g (89.9 g) of the desired title aldehyde as a light yellow solid.
%). Melting point 72-75 ° C (melting point reported in literature = 73
~ 74 ℃). TLC: hexane-diethyl ether (95: 5),
R _f = 0.40, UV and PMA.

D. 4'-Fluoro-3,3 ', 5-trimethyl [1,1'-biphenyl] -2-methanol A cold (on an ice bath, 0 ° C) solution of 15.0 ml of dry diethyl ether was added with 259 mg of lithium aluminum hydride ( 6.82 mmol, 0.55 equiv.) And added to the gray suspension to the C
A solution of 3.0 g (12.4 mmol) of the aldehyde of the above in 15 ml of dry diethyl ether is added dropwise over 15 minutes. After stirring the mixture at room temperature for 30 minutes under an argon atmosphere, it is cooled to 0 ° C., and the reaction is stopped by adding dropwise 260 μ of water, 260 μ of 15% sodium hydroxide and 780 μ of water successively. The suspension is diluted with ethyl acetate and passed through anhydrous sodium sulfate over a packed Celite (1/4 ") bed and the colorless liquid is evaporated under reduced pressure to give 2.99g (98.8%) of a white solid. .
The crude solid was triturated with cold hexane and dried under reduced pressure to give 2.467 g (81.6%) of the desired title alcohol as a white solid.
Get. 102-103 ° C. TLC: hexane-ethyl acetate (9: 1), _Rf = 0.24, UV and PMA.

E. (S) -3-[[(1,1-Dimethylethyl) diphenylsilyl] oxy] -4- (hydroxymethoxyphosphoryl) butanoic acid methyl ester 1. (S) -4-Bromo-3-hydroxybutanoic acid methyl ester Ester 1a. [R- (R ^* , R ^* )]-2,3,4-trihydroxybutanoic acid calcium salt hydrate (Carbohydrate Research Vol. 72, pages 301-304 (1979))
Reference) D-Isoascorbic acid 44.0 g (250 mmol) in water (625 ml) was added with calcium carbonate 50 g, the suspension was cooled to 0 ° C (ice bath), and 30% hydrogen peroxide 100 ml was added little by little. To process. The mixture is stirred at 30-40 ° C (oil bath) for 30 minutes. 10 g of Darco are added and the black suspension is heated on a steam bath until oxygen evolution ceases. The suspension is passed through Celite and evaporated under reduced pressure (40 ° C. bath). The residue is dissolved in 50 ml of water, warmed on a steam bath and methyl alcohol is added until the solution becomes cloudy. The gummy solid is collected and air dried to give 30.836 g (75.2%) of the desired calcium salt as a white powdery solid. TLC: isopropyl alcohol-ammonium hydroxide-water (7: 2: 1), R _f
= 0.19, PMA.

1b. [S- (R ^* , R ^* )]-2,4-dibromo-3-hydroxybutanoic acid methyl ester (Bock, K et al .: Acta Scandinavica (B) Vol. 37
Pp.341-344 (1983)) Dissolve 30 g of the calcium salt of paragraph 1a above in 30-32% hydrogen bromide in 210 ml acetic acid and stir at room temperature for 24 hours. Add 990 ml of methanol to this brown solution and stir overnight. The mixture is evaporated, the orange oil obtained is dissolved in 75 ml of methyl alcohol, refluxed for 2.0 hours and evaporated. The residue was partitioned between 100 ml of ethyl acetate and water and the organic layer was washed with water (2
After washing with water) and saline, dried over anhydrous sodium sulfate,
Evaporate to give crude dibromide as a light orange oil.
22.83 g (90.5%) are obtained. TLC: ethyl acetate-hexane (1: 1), _Rf = 0.69, UV and PMA.

1c. (S) -4-Bromo-3-hydroxybutanoic acid methyl ester (similar to the production method in section 1b (see the same method)) The dibromide derivative 2 was added to 370 ml of ethyl acetate and 37 ml of glacial acetic acid.
A solution of 0.80 g (75.4 mmol) and anhydrous sodium acetate 21.0 g (under argon atmosphere) was added to 5% palladium / carbon 1.30.
Treat with g and stir the black suspension under hydrogen (1 atm), monitoring the uptake of hydrogen. After the hydrogen absorption had ended after 2 hours, the mixture was passed through Celite,
After washing the liquid with saturated sodium hydrogen carbonate and brine,
Dry over anhydrous magnesium sulfate and evaporate to give the crude dibromo ester as a brown oil. This crude oil is combined with another batch (starting from 36.77 g of dibromide) and distilled under reduced pressure to give 25.77 g (61.3%) of the desired title bromoester as a clear oil. Boiling point 79-80 ℃ (1.0mmH
g). TLC: ethyl acetate-hexane (1: 1), R _f = 0.44, PM
A.

Elemental analysis, calculated as C ₅ H ₉ O ₃ Br, C H Br calculated: 30.48; 4.60; 40.56 found: 29.76; 4.50; 39.86 2. (S) -4-bromo-3-[[(1,1- Dimethylethyl) diphenylsilyl] oxy] butanoic acid methyl ester 4.0 g (20.4 mmol) of bromohydrin of the above item E1, 6.94 g (5.0 equivalents) of imidazole and 12 mg (0.005 equivalents) of 4-dimethylaminopyridine in dry DMF (40 ml).
The solution was added with 5.84 ml of t-butyldiphenylsilyl chloride.
Treat (1.1 eq.) And stir the homogeneous mixture at room temperature under an atmosphere of argon overnight. The mixture was partitioned between 5% potassium hydrogen sulfate and ethyl acetate, the organic layer was washed with water and brine, dried over anhydrous sodium sulfate and evaporated to give 9.32 g of crude silyl ether as a clear viscous oil ( 100
%). TLC: hexane-ethyl acetate (3: 1), _Rf silyl ether = 0.75, UV and PMA.

3. (S) -4-iodo-3-[[(1,1-dimethylethyl) diphenylsilyl] oxy] butanoic acid methyl ester 9.32 g (201 mmol) of methyl ethyl ketone (dried with 4Å sieves) of the crude bromide derivative of E2 above. ) A 60 ml solution is treated with 15.06 g (100.5 mmol, 5.0 eq) of sodium iodide and the yellow suspension is refluxed for 5.0 hours under an atmosphere of argon. The mixture was cooled, diluted with ethyl acetate, washed with dilute sodium bisulfite (until colorless) and brine, dried over anhydrous sodium sulfate and evaporated under reduced pressure to give 10.17 g of a yellow oil. obtain. The crude oil is purified by flash chromatography on 600 g of silica gel, eluting with hexane-methylene chloride (3: 1). The product fractions are combined and evaporated to give 7.691 g (74.2%, total yield of both steps) of the desired title iodide as a clear colorless viscous oil.

TLC: hexane-ethyl acetate (3: 1) product, _Rf = 0.75, U
V and PMA (Note-Iodide formed is common spot with starting bromide).

4. (S) -4- (diethoxyphosphoryl) -3-
[[(1,1-Dimethylethyl) diphenylsilyl] oxy] butanoic acid methyl ester A solution of 7.691 g of the above iodide in 20 ml of triethyl phosphite is heated at 155 ° C (oil bath) for 3.5 hours under an argon atmosphere.
Cool the mixture and remove excess phosphite under reduced pressure (0.5 mmH
g, 75 ° C.) to give a yellow oil (approximately 8.0 g). The crude oil is purified by flash chromatography on 400 g of silica gel, eluting with hexane-acetone (4: 1). Evaporate the product fractions to give 3.222 g (41.1%) of the desired title phosphonate as a clear colorless viscous oil. TLC: hexane-acetone (1: 1), R _f = 0.51, UV
And PMA. An additional 2.519 g of product (corrected yield 61.1%) is recovered using the starting iodide of paragraph 3 above.

5. (S) -3-[[(1,1-Dimethylethyl) diphenylsilyl] oxy] -4-phosphonobutanoic acid methyl ester A solution of 9.85 g (20.0 mmol) of the phosphonate of the above item 4 in 60 ml of dry methylene chloride was added to bistrimethylsilyl. 5.31 ml (32.0 mmol, 1.6 eq) of trifluoroacetamide (BSTFA) and trimethylsilyl bromite (TMSBr) 6.
Sequentially treat with 60 ml (50.0 mmol, 2.5 eq) and stir the clear mixture overnight at room temperature under argon atmosphere, 5%
80 ml potassium hydrogen sulphate are added and the mixture is extracted with ethyl acetate. The aqueous layer is saturated with sodium chloride and reextracted with ethyl acetate. The combined organic layers are washed with brine, dried over anhydrous sodium sulfate and evaporated under reduced pressure to give the crude title phosphonic acid compound as a viscous oil. TLC: isopropanol-ammonium hydroxide-water (7: 2: 1), R _f = 0.3
0, UV and PMA.

6. (S) -3-[[(1,1-Dimethylethyl) diphenylsilyl] oxy] -4- (hydroxymethoxyphosphoryl) butanoic acid methyl ester In 25 ml of dry pyridine, the crude phosphonic acid compound of the above item 5 (approximately 20.0 1.62 ml (40.0 mmol, 2.0 equivalents) of dry methanol (on 3Å sieve) and 4.54 g (22.0 mmol, dicyclohexylcarbodiimide (DCC)) of dicyclohexylcarbodiimide (DCC).
1.10 eq.) And stir the white suspension overnight at room temperature under argon atmosphere. Remove pyridine under reduced pressure and azeotrope with benzene (15 ml x 2). The residual oil was dissolved in ethyl acetate, washed with 1.0N hydrochloric acid and brine,
After drying over anhydrous sodium acetate and evaporating under reduced pressure, 8.272 g of the title crude ester compound is obtained as an oily substance containing a small amount of dicyclohexylurea (DCU) precipitated. TLC: isopropanol-ammonium hydroxide-water (7: 2: 1), _Rf = 0.60, U
V and PMA.

F. (S) -4-[[[4'-fluoro-3,3 ', 5-trimethyl [1,1'-biphenyl] -2-yl] [methoxy] methoxyphosphoryl] -3-t-butyldiph Ethylsilyloxybutanoic acid methyl ester 6.595 g (approximately 14.7 mmol) of the crude phosphonic acid monomethyl ester of the previous item E was dissolved in 30 ml of dry methylene chloride, treated with distilled trimethylsilyldiethylamine 5.60 ml (29.4 mmol, 2.0 equivalents), and charged with argon. Stir at room temperature under an atmosphere for 1 hour. The mixture is evaporated under reduced pressure, chase with benzene (1 x 30 ml) and dried under reduced pressure. The light yellow viscous oil obtained was dried with methylene chloride.
1.4 ml of oxalyl chloride (16.2 mmol, 1.1 eq) dissolved in 2 ml of DMF (4Å sieve) and cooled to -10 ° C (salt / ice bath) and distilled using a syringe.
Is added dropwise. A strong outgassing is revealed and the solution color is deeper yellow. The mixture is stirred under an argon atmosphere at -10 ° C for 15 minutes and then at room temperature for 1 hour. The mixture is evaporated under reduced pressure, washed with benzene (1 x 30 ml) and dried under reduced pressure to give crude phosphonochloridate as a yellow oil.

A solution of crude phosphonochloridate (approximately 14.7 mmol) in 10 ml of dry methylene chloride was added dropwise with a solution of 2.06 g (8.43 mmol) of the biphenyl alcohol derivative of the above item D in 15 ml of dry pyridine, and the mixture was stirred at room temperature under an argon atmosphere. Stir for 16 hours. The mixture is evaporated to dryness and the residue is partitioned between 5% potassium hydrogen sulfate and ethyl acetate. The organic layer was washed with saturated sodium hydrogen carbonate and brine, dried over anhydrous sodium sulfate and evaporated under reduced pressure to give a brown oil, 8.290 g.
Get. The crude product was subjected to flash chromatography on 370 g of silica gel and hexane-acetone (70:30).
Purify by eluting with The product fractions were combined and evaporated to give 3.681 g (66%) of the desired title phosphonate as a pale yellow oil.
%). TLC: hexane-acetone (3: 2), R _f = 0.5
9, UV and PMA.

G. (S) -4-[[[4'-fluoro 3,3 ', 5-trimethyl [1,1'-biphenyl] -2-yl] [methoxy]
Methoxyphosphoryl] -3-hydroxybutanoic acid methyl ester In 20.0 ml of dry THF, 1.103 of the silyl ether form of the above item F
A mixture of g (1.66 mmol) was treated with 380 μ (6.64 mmol, 4.0 eq) glacial acetic acid and 4.98 ml (4.98 mmol, 3.0 eq) 1.0 M tetrabutylammonium fluoride solution and the clear yellow solution was placed under an argon atmosphere. , Stir overnight at room temperature. The mixture was partitioned between cold water and ethyl acetate, the organic layer was washed with saturated sodium hydrogen carbonate and brine, dried over anhydrous sodium sulfate and evaporated to give 1.174g of a viscous yellow oil. Crude oil on silica gel 47 g,
Attach to flash chromatography, methylene chloride-
Purify by eluting with acetone (85:15). The product fractions are evaporated to give 679 mg (93.1%) of the desired title alcohol as a clear viscous oil. TLC: hexane-acetone (1: 1), Rf = 0.41, UV and PMA.

H. (S) -4-[[[4'-fluoro-3,3 ', 5-trimethyl [1,1'-biphenyl] -2-yl] methoxy]
Methoxyphosphoryl] -3-hydroxy-butanoic acid monolithium salt 184 mg (0.420 mmol) of the methyl ester derivative of the above-mentioned section G
Dioxane 5.0 ml solution 1.0 N lithium hydroxide 0.50 ml
Treat (1.2 eq) and stir the mixture at room temperature under an atmosphere of argon for 3 hours. The mixture is diluted with water, passed through a 0.4 μm polycarbonate film and evaporated under reduced pressure. The residue is dissolved in 75 ml of water, frozen and lyophilized.
Dissolve the crude acid product in the minimum amount of water and mix with CHP-20 resin bed 1
Chromatograph on 00 ml and elute with a water / acetonitrile linear gradient eluent. Evaporate the product fractions,
Dissolved in 50 ml of water, passed through a 0.4 μm polycarbonate film and lyophilized to give 174 mg (89.1% based on hydrate weight) of the desired title monolithium salt as a white solid. TLC: isopropanol-ammonium hydroxide-water (7: 2: 1), _Rf = 0.58, UV and PMA.

Elemental analysis, C ₂₁ H ₂₅ O ₆ PFLi + 1.95 mol H ₂ O (MW465.46)
As C H F P calculated value: 54.19; 6.26; 4.08; 6.65 actual value: 54.19; 6.21; 4.29; 6.43 H ¹・ NMR (400M Hz): δ 1.74 to 2.08 ppm (2H, m, -PO (OCH ₃ ) CH ₂ −) 2.30 (3H, s, aromatic methyl) 2.32 (3H, d, aromatic methyl α-F, J _HF 2.2Hz) 2.35 to 2.62 (2H, m, −CH ₂ CO ₂ Li) 2.46 (3H, s, aromatic methyl) 3.57 and 3.63 (3H, 2 double lines, -OP (OCH ₃ )-, 2 diastereomers, J _HP = 10.3Hz) 4.28 (1H, m, -CH ₂ CH (OH) CH ₂ CO ₂ Li) 6.87 to 7.25 (5H, m, aromatic H) Example 2 (S) -4-[[[4'-fluoro-3,3 ', 5-trimethyl [1,1'-biphenyl] -2-yl]] Methoxy] hydroxyphosphoryl] -3-hydroxybutanoic acid dilithium salt 374 mg (0.853 mmol) of the diester derivative of Example 1
A solution of 8.0 ml of dioxane in 2.6 ml of 1.0 N lithium hydroxide (3.
(0 eq) and under argon atmosphere at 50 ° C (oil bath)
Heat for 5.0 hours. A clear white precipitate forms. Dilute the mixture with water and pass. The aqueous solution is extracted once with diethyl ether, passed through a 0.4 μm polycarbonate membrane and concentrated under reduced pressure. The crude product was converted into CHP-20 resin (1
Chromatography on 0 ml bed) and elute with a water / acetonitrile linear gradient elution system. The product fractions were evaporated under reduced pressure, dissolved in 50 ml of water and lyophilized by passing through a 0.4 μm polycarbonate film to give 260 mg of the desired title dilithium salt as a white solid (6% based on hydrate weight).
7.1%). TLC: propanol-ammonium hydroxide-water (7: 2: 1), _Rf = 0.47, UV and PMA.

Elemental analysis, calculated as C ₂₀ H ₂₂ O ₆ PFLi ₂ +1.77 mol H ₂ O, C H F P calculated value: 52.88; 5.67; 4.18; 6.82 found value: 52.88; 5.26; 4.24; 6.43 H ¹・ NMR (400M Hz, CD ₃ OD): 2.26 ~ 2.42 (2H, m, CH ₂ CO ₂ Li) 2.30 (3H, s, aromatic methyl) 2.31 (3H, d, aromatic methyl α-F, J _HF = 1.9Hz) 2.38 (3H, s, aromatic family methyl) 4.22 (1H, m, -CH (OH) CH 2 -) 6.86-7.23 (5H, m, aromatic proton) Example 3 (3S) -4-[[[4'-fluoro-3,3 ', 5-trimethyl [1,1'-biphenyl] -2-yl]] Meoxy] methylphosphoryl] -3-hydroxybutanoic acid monolithium salt A. (S) -4-[(chloro) methylphosphoryl] -3-
[[[(1,1-Dimethylethyl) diphenylsilyl] oxy] butanoic acid methyl ester The title phosphonochloridate compound is prepared as described in Example 6B, paragraphs 1 to 3.

B. (3S) -4-[[[4'-fluoro-3,3 ', 5-trimethyl [1,1'-biphenyl] -2-yl] methoxy]
Methylphosphoryl] -3-t-butyldiphenylsilyloxybutanoic acid methyl ester phosphonochloridate (approximately 2.2 mmol) of the above item A and 429 mg (2.2 mg of the biphenyl alcohol compound of the example 1D item)
10 mL of a dry solution of 10 mL of dry methylene chloride (0 ° C. ice bath) was treated with 425 μ (3.04 mmol, 1.4 eq) of triethylamine and 27 mg (0.22 mmol) of 4-DMAP, and the orange solution was treated with argon atmosphere under argon atmosphere. Stir overnight at room temperature. The mixture was partitioned between 5% potassium hydrogen sulfate and ethyl acetate, the organic layer was washed with brine, dried over anhydrous sodium sulfate and evaporated to give 1.1 g of an orange oil. This crude oil is purified by flash chromatography on 44 g of LPS-1 silica gel, eluting with ethyl acetate: hexane (1: 1). The product fractions are combined and evaporated to give 298 mg (21%) of the desired title product coupled as a pale yellow oil. In addition, 460 mg of the biphenyl alcohol starting material (corrected recovery rate 67%) of Example 1D is recovered. TLC: Ethyl acetate: Hexane (1:
1), R _f = 0.18, UV and PMA.

C. (3S) -4-[[[4'-fluoro-3,3 ', 5-trimethyl [1,1'-biphenyl] -2-yl] methoxy]
Methylphosphoryl] -3-hydroxybutanoic acid methyl ester 298 mg (0.46 mmol) of the silyl ether of the above-mentioned section B
A solution of 6.0 ml of dry THF in 110 μl of glacial acetic acid (1.84 mmol, 4.0 eq) and tetrabutylammonium fluoride
Treat with 1.43 ml (3.1 eq) of a THF solution (1.0 M) and stir the solution at room temperature under an atmosphere of argon overnight. The mixture was partitioned between cold water and ethyl acetate, the organic layer was washed with saturated sodium hydrogen carbonate and brine, dried over anhydrous sodium sulfate and evaporated to give a yellow oil 273mg. The crude oil is purified by flash chromatography on 11 g of LPS-1 silica gel, eluting with hexane-acetone.
The product fractions are combined and evaporated to give 150 mg (80%) of the desired title alcohol as a viscous oil. TLC: hexane-acetone (1: 1), _Rf = 0.23, UV and PMA.

D. (3S) -4-[[[4'-fluoro-3,3 ', 5-trimethyl [1,1'-biphenyl] -2-yl] methoxy]
Methylphosphoryl] -3-hydroxybutanoic acid monolithium salt 150 mg (0.367 mmol) of the methyl ester derivative of the above item C
Dioxane 3.0 ml solution, 1.0N lithium hydroxide 0.44 ml
Treat with (1.2 eq) and stir the resulting white suspension for 2 hours at room temperature under an atmosphere of argon. The mixture is diluted with water, passed through a 0.4 μm polycarbonate film and evaporated under reduced pressure to give a colorless glass-like material.

Dissolve this crude product in the minimum amount of water and add HP-20 (100 ml
Bed), chromatograph and elute with a water / acetonitrile linear gradient eluent. The product fractions were evaporated, dissolved in 50 ml of water, passed through a 0.4 μm polycarbonate film and lyophilized to give 130 mg (79% (based on hydrate weight) of the desired title lithium salt as a white solid. Obtained. TLC: methylene chloride-methanol-acetic acid (8: 1: 1), R
_f = 0.52, UV and PMA.

Elemental analysis, C ₂₁ H ₂₅ O ₅ FLiP + 1.73 mol H ₂ O (MW445.49)
As calculated, C H F P calculated value: 56.61; 6.44; 4.26; 6.95 actual value: 56.67; 6.36; 4.31; 7.43 H ¹・ NMR (400 MHz): 2.27 ~ 2.40 (2H, m, CH ₂ CO ₂ Li) 2.30 (6H, s, 2 aromatic methyls) 2.44 (3H, s, aromatic methyl) 4.26 (1H, m, -CH ₂ CH (OH) CH ₂ CO ₂ Li) 6.90 to 7.20 (5H, m, aromatic H) Example 4 (S) -4-[[[2,4-dichloro-6-[(4-fluorophenyl) methoxy] phenyl] methoxy] methoxyphosphoryl]- 3-Hydroxybutanoic acid monolithium salt A.2,4-dichloro-6- (4-fluorophenylmethoxy) benzaldehyde (see J. Med. Chem. (1986) Vol. 29, p. 167) 4,6-dichloro- 2-hydroxybenzaldehyde 13.
A solution of 77 g (72.5 mmol) in 100 ml DMF is stirred and 12.02 g (87 mmol) potassium carbonate are added. After heating the mixture to approximately 70 ° C for 60 minutes, 4-fluorobenzyl bromide 11.7
Add ml. The solution obtained is stirred at 70 ° C. for 3.5 hours, then poured into ice-water 1.5, filtered, washed with water and recrystallized from diethyl ether / petroleum ether. Yield of off-white crystals: 17.88 g (83%). Melting point 107-108 ° C.

B.2,4-Dichloro-6-[(4-fluorophenyl) methoxy] benzenemethanol Cold (0 ° C. ice bath) dry diethyl ether 10.0 ml was treated with lithium aluminum hydrogen 158 mg (4.16 mmol, 0.6 eq), 2.06 g of the aldehyde compound of the above section A in a gray suspension
(6.93 mmol of dry THF 10 ml solution is added dropwise. The mixture is warmed to room temperature and stirred under an atmosphere of argon for 1 hour.
The mixture was cooled again to 0 ° C (ice bath), water 160μ, 15%
The reaction is stopped by the sequential addition of 160 µ of sodium hydroxide and 475 µ of water. Fill the precipitated salt with Celite (1 /
It is passed over anhydrous sodium sulfate over 4 ″ bed) and evaporated. The clear liquid is evaporated to obtain 2.052 g (98.9%) of crude alcohol as white crystals. Milled once with cold hexane to give white crystalline Labeled as solid Alcohol Pure 1.89
2 g (91.2%) are obtained. 72-73 ° C.

TLC: hexane-acetone (4: 1), R _f = 0.31, UV and P
MA.

Elemental analysis, as C ₁₄ H ₁₁ O ₂ Cl ₂ F (MW301.142), C H Cl F calculated: 55.84; 3.68; 23.55; 6.31 found: 55.97; 3.71; 23.42; 6.30 C. (S) -4 -[[[2,4-Dichloro-6-[(4-fluorophenyl) methoxy] phenyl] methoxyphosphoryl 4] -3-t-butyldiphenylsilyloxybutanoic acid methyl ester The methyl ester of Example 1E6 A solution of (approximately 3.84 mmol) in 10 ml of dry methylene chloride was treated with 1.46 ml of distilled trimethylsilyldiethylamine (7.68 mmol, 2.0 eq) and the solution was 1.
Stir for 0 hours. The mixture is evaporated under reduced pressure, washed with benzene (1 x 20 ml) and dried under reduced pressure to give crude silylated phosphonic acid monomethyl ester as a colorless oil.

A solution of the above crude ester (approximately 3.84 mmol) in 10 ml of dry methylene chloride and dry DMF (1 drop) was cooled to -10 ° C (salt / ice bath) and distilled oxalyl chloride 368μ.
(4.22 mmol, 1.1 eq) is added dropwise. Gas is clearly evolved from this clear yellow mixture. The mixture was stirred under argon atmosphere at room temperature for 1 hour, evaporated under reduced pressure, washed with benzene (20 ml x 2 times), and crude phosphonochloridat as a viscous yellow oil.
e) get.

The crude phosphonochloridate (approximately 3.84 mmol) (0 ° C. ice bath) in 10 ml of dry methylene chloride was mixed with 1.15 g (3.84 mmol, 1.0 equivalent) of the alcohol compound of the section B, and then 805 μ (5.76 mmol, 1.5 equivalent) of triethylamine. ) And 4-DMAP 47 mg (0.384 mmol, 0.1 eq),
The dry mixture is stirred overnight at room temperature under an atmosphere of argon.
The mixture was partitioned between 5% potassium hydrogen sulfate and ethyl acetate, the organic layer was washed with brine, dried over anhydrous sodium sulfate and evaporated to give 3.197g of a dark brown oil. The crude product is purified by flash chromatography on 160 g silica gel, eluting with hexane-ethyl acetate (7: 3). The product fractions are combined and evaporated to give 594 mg (21.1%) of the desired title phosphonate as a yellow oil. Further, 688 mg of the alcohol starting material of Section B (corrected recovery rate 52.4%) is recovered. TLC: hexane-acetone (1:
1), R _f = 0.29, UV and PMA.

D. (S) -4-[[[2,4-dichloro-6-[(4-fluorophenyl) methoxy] phenyl] methoxy] methoxyphosphoryl] -3-hydroxytanic acid methyl ester Methyl silyl ester Body 578 mg (0.788 mmol)
A solution of 8 mL of dry THF in 180 μL of glacial acetic acid (3.2 mmol, 4.
0 eq), then THF 2.36 ml (2.36 mmol, 3.0 eq)
Medium, tetra-n-butylammonium fluoride (n-Bu
_4- NF) solution (1.0 M) and the resulting pale yellow solution is stirred overnight at room temperature under an argon atmosphere. The mixture is poured into cold water and extracted twice with ethyl acetate. The organic layer is washed with saturated sodium hydrogen carbonate and brine, dried over anhydrous sodium sulfate and evaporated to give 625 mg of a yellow oil. The crude product is purified by flash chromatography on 31 g silica gel, eluting with hexane-acetone (7: 3). The product fractions were combined and evaporated to give the desired title alcohol as a colorless transparent viscous oil 339m
Get g (86.9%). TLC: hexane-acetone (1: 1),
R _f = 0.25, UV and PMA.

E. (S)-[[[2,4-Dichloro-6-[(4-fluorophenyl) methoxy] phenyl] methoxy] methoxyphosphoryl] -3-hydroxybutanoic acid monolithium salt 132 mg of the phosphonate of the above section D ( 0.267 mmol) in 2.5 ml of dioxane was added to 0.32 ml of 1.0 N lithium hydroxide (1.2
Equivalent) and the mixture is at room temperature under an argon atmosphere 4.
Stir for 0 hours. A white precipitate forms. The mixture is diluted with water and the liquid is evaporated to dryness under reduced pressure. Residue
Chromatograph on HP-20 resin (100 ml bed) and elute with a water / acetonitrile linear gradient elution system.
The product fractions were combined, evaporated, dissolved in water, passed through a 0.4 μm polycarbonate film and lyophilized to give 108 mg (79% based on hydrate weight) of the desired title lithium salt as a white solid. Obtained. TLC: Methylene chloride-methanol-
Acetic acid (20: 1: 1), R _f = 0.41, UV and PMA.

Elemental analysis, C ₁₉ H ₁₈ O ₇ Cl ₂ FLi ₂ P + 1.42 mol H ₂ O (MW511.7
2) as calculated value: C, 44.59; H, 4.10; Cl, 13.86; F, 3.71; P, 6.05 Actual value: C, 44.22; H, 4.09; Cl, 13.91; F, 3.72; P, 6.11 H ¹・ NMR (400M Hz): 2.26~2.45 (2H, m, -CH ( OH) CH 2 CO 2 Li) 3.63 and 3.62 (3H, 2 pieces doublets, diastereomer 2
Pieces, 4.23 (1H, m, −CH ₂ CH (OH) CH ₂ CO ₂ Li) 5.16 (2H, s, F−PhCH ₂ O) 5.24 (2H, d, ArCH ₂ OP, J _HP = 6.2Hz) 7.13 to 7.53 (6H, m, aromatic H) Example 5 (3S) -4-[[[[2,4-dichloro-6-[(4-fluorophenyl) methoxy] phenyl] methoxy] hydroxyphosphonyl] -3- Hydroxybutanoic acid dilithium salt In 4.0 ml of dioxane, diester 2 of the above-mentioned Example 4D is used.
10 mg (0.424 mmol) of a mixture was added to 1.0 N lithium hydroxide.
Treat with 1.30 ml (3.0 eq) and warm the colorless solution at 50 ° C. (oil bath) for 3.5 hours under an atmosphere of argon. A clear white precipitate forms after 15 minutes. The mixture is diluted with water and the liquid is evaporated under reduced pressure. The residue is dissolved in a minimum amount of water, chromatographed on HP-20 resin (100 ml bed) and eluted with a water / acetonitrile linear gradient eluent. The product fractions are combined and evaporated. Residue in water 50
Treated with ml, passed through a 0.4 μm polycarbonate film and lyophilized to give 175 mg (81% based on hydrate weight) of the desired title dilithium salt as a white solid. TLC:
Methylene chloride-methanol-acetic acid (8: 1: 1), _Rf = 0.0
7, UV and PMA.

Elemental analysis, C ₁₈ H ₁₆ O ₇ Cl ₂ FLi ₂ P + 1.70 mol H ₂ O (MW509.6
2) as calculated value: C, 42.42; H, 3.84; F, 3.73; Cl, 13.91; P, 6.08, actual value: C, 42.46; H, 3.09; F, 3.90; Cl, 13.42; P, 5.66.

H ¹ · NMR (400 MHz): 2.27 (1H, dd, −CH (OH) CH ₂ CO ₂ Li, J _HH = 8.8Hz) 2.39 (1H, dd, −CH (OH) CH ₂ CO ₂ Li, J _HH = 4.4Hz) 4.26 (1H, m, CH ₂ CH (OH) CH ₂ CO ₂ Li) 5.08 (2H, s, F-Ph-CH ₂ OAr) 7.03 to 7.53 (6H, m, aromatic H) Example 6 (3S) -4- [ [2,4-Dichloro-6-[(4-fluorophenyl) methoxy] phenyl] methoxy] methylphosphoryl] -3-hydroxybutanoic acid methyl ester A. (S) 3-[[(1,1-dimethylethyl ) Diphenylsilyl] oxy] -4- (ethoxymethylphosphoryl)
Butanoic acid methyl ester 5.0 g (36.7 mmol, Strem Chemicals) of methyldiethoxyphosphine, Example 1E3
A mixture of 4.68 g (9.18 mmol) of iodide in paragraph is heated at 100 ° C. (oil bath) for 2.5 hours under an argon atmosphere and then at 150 ° C. for a further 3 hours. A dull white precipitate forms in this yellow solution. Excess phosphine is removed under reduced pressure (0.5 mmH
g) Evaporate and subject the crude product to silica gel flash chromatography on hexane-acetone (65:35).
Purify by eluting with. The product fractions were combined and distilled to give 1.590 g (38%) of the desired title phosphinic acid ester (a mixture of diastereomers) as a clear viscous oil.
Get. TLC: hexane-acetone (3: 2), _Rf (2 diastereomers) = 0.19 and 0.22, UV and PMA.

B. (3S) -4-[[2,4-dichloro-6-[(4-fluorophenyl) methoxy] phenyl] methoxy] methylphosphoryl] -3-t-butyldiphenylsilylbutanoic acid methyl ester A A solution of 605 mg (1.3 mmol) of the phosphinic acid ester in section 6.0 ml of dry methylene chloride was added to 280 μ of bis (trimethylsilyl) trifluoroacetamide (BSTFA).
(1.05 mmol, 0.8 eq) and trimethylsilyl bromide (TMS Br) 210μ (1.57 mmol, 1.2 eq), this solution was stirred at room temperature overnight under argon atmosphere, 15 ml of 5% potassium hydrogen sulfate was added and the mixture was Extract with ethyl acetate. The organic layer is washed with brine, dried over anhydrous sodium sulfate and evaporated under reduced pressure to give crude phosphinic acid as a colorless oil.

A solution of this crude phosphinic acid (approximately 1.3 mmol) in 6.0 ml of dry methylene chloride was treated with 270 μm of distilled trimethylsilyldiethylamine (1.44 mmol, 1.1 eq) and the clear mixture was stirred at room temperature under an atmosphere of argon at 1.0.
Stir for hours. The mixture is evaporated under reduced pressure, washed with benzene (1 x 15 ml) and dried under reduced pressure.

To a cold (0 ° C. ice bath) solution of the above crude silylated phosphinic acid (approximately 1.3 mmol) in 6.0 ml of dry methylene chloride and 1 drop of DMF, 130 μ (1.44 mmol, 1.1 eq) of distilled oxalyl chloride was added dropwise with a syringe. To do. Clearly gas is released. The mixture was stirred under argon atmosphere at room temperature for 1 hour, then evaporated under reduced pressure and treated with benzene (15 ml x 2).
Wash twice and dry under reduced pressure to give crude phosphonochloridate as a yellow oil.

Phosphonochloridate (approximately 1.3 mmol) and alcohol 392 m of Example 1E6 in 6.0 ml of dry methylene chloride.
A solution of g (1.3 mmol) in cold (0 ° C. ice bath) was added with triethylamine 275 μ (1.97 mmol, 1.5 eq) and 4-DM.
Treat with AP (16 mg, 0.13 mmol, 0.1 eq) and stir the yellow mixture at room temperature overnight under an atmosphere of argon. The mixture was partitioned between 5% potassium hydrogen sulfate and ethyl acetate, the organic layer was washed with brine and dried over anhydrous sodium sulfate,
Evaporate to give 908 mg of crude product as a dark yellow oil.
The crude product is purified by flash chromatography on 45 g of silica gel, eluting with hexane-ethyl acetate (3: 2). The product fractions were combined and evaporated,
266 mg (28.3 mg) of the desired title product as a clear colorless oil.
%). Also recover 197 mg of alcohol starting material (corrected recovery rate 57%).

C. (3S) -4-[[2,4-dichloro-6-[(4-fluorophenyl) methoxy] phenyl] methoxy] methylphosphoryl] -3-hydroxybutanoic acid methyl ester The silyl ester form of the above section B 275 mg (0.38 mmol)
A solution of 6.0 ml of dry THF in 90 μl of glacial acetic acid (1.53 mmol,
4.0 equivalents) and tetrabutylammonium fluoride
Treat with 1.2 ml (3.1 equiv.) THF solution (1.0 M). The solution is stirred overnight at room temperature under an argon atmosphere. The mixture was partitioned between cold water and ethyl acetate, the organic layer was washed with saturated sodium hydrogen carbonate and brine, dried over anhydrous sodium sulfate and evaporated to give a yellow oil 258mg. The crude product is purified by flash chromatography on 8 g LPS-1 silica gel, eluting with hexane-acetone (1: 1). The product fractions were combined and evaporated to give 142 mg (77%) of the desired title alcohol as a clear colorless oil.
%). TLC: hexane-acetone (1: 1), R _f = 0.2
0, UV and PMA.

D. (3S) -4-[[2,4-Dichloro-6-[(4-fluorophenyl) methoxy] phenyl] methoxy] methylphosphoryl] -3-hydroxybutanoic acid monolithium salt The methyl ester of the above item C 142 mg (0.296 mmol)
Dioxane 3.0 ml solution, 1.0N lithium hydroxide 0.36 ml
Treat with (1.2 eq) and stir the white suspension for 2.0 h at room temperature under argon atmosphere. Dilute the mixture with water,
Pass through 0.4 μm polycarbonate film and allow liquid to evaporate under reduced pressure.

The crude product is dissolved in a minimum amount of water, chromatographed on a 100 ml bed of HP-20 resin and eluted with a water / acetonitrile linear gradient eluent. The product fractions are combined and evaporated. The residue is treated with water, passed through a polycarbonate film and lyophilized to give 93 mg (63% based on hydrate weight) of the desired title lithium salt as a white solid.
TLC: methylene chloride-methanol-acetic acid (8: 1: 1), UV and PMA.

Elemental analysis, C ₁₉ H ₁₉ O ₇ Cl ₂ FLiP + 1.38 mol H ₂ O (MW495.9
4), calculated value: C, 46.01; H, 4.42; F, 3.83; Cl, 14.30; P, 6.24, measured value: C, 46.10; H, 4.49; F, 3.82; Cl, 14.32; P, 6.43.

H ¹ · NMR (400 MHz): 2.27 (1H, dd, −CH (OH) CH ₂ CO ₂ Li, J _HH ＝ 8.4Hz, J _HP ＝
1.1Hz) 2.38 (1H, dd, −CH (OH) CH ₂ CO ₂ Li, J _HH ＝ 4.7Hz, J _HP ＝
1.1Hz) 4.29 (1H, m, −CH ₂ CH (OH) CH ₂ CO ₂ Li) 5.16 and 5.18 (4H, m, ArCH ₂ OP and F−PhCH ₂ O) −) 7.11 to 7.52 (6H, m, Aromatic) Example 7 (S) -4-[[[4'-fluoro-3,3 ', 5-trimethyl [1,1'-biphenyl-2-yl] methyl] amino] methoxyphosphoryl] -3- Hydroxybutanoic acid monolithic acid A. 4'-fluoro-3,3 ', 5-trimethyl [1,1'-biphenyl] -2-carboxylic acid 1.0 g (4.13 mmol) of acetone of the aldehyde derivative of Example 1C2 above To 10.0 ml of the liquid (0 ° C. ice bath), 4.1 ml (excess amount) of 8.0N Jyon's reagent was added dropwise, and the obtained greenish brown suspension was stirred overnight at room temperature under an argon atmosphere. Separate the chromium salt precipitate by adding 10.0 ml of isopropanol to decompose excess oxidant and passing through a celite (1/4 ″) pad. Evaporate the solution and dissolve in ethyl acetate, 2 times with 1.0N hydrochloric acid and 2 times with saturated ammonium chloride
Wash with brine and brine, dry over anhydrous sodium sulfate and evaporate to give a green solid 1.011 g. Melting point 153-154 ° C. The acid product is purified as follows via the dicyclohexylamine salt of this crude acid product. That is, to a 5.0 ml solution of the crude acid product in ethyl acetate is added 823 μ (1.0 eq) of dicyclohexylamine (DCHA). This solution was diluted with hexane and the precipitated crystalline salt was collected, giving off-white crystalline
997 mg (55% yield from aldehyde) of the desired product are obtained as a DCHA salt (mp 181-183 [deg.] C).

The DCHA salt is treated as follows to regenerate the title free acid. That is, the salt is partitioned between 5% potassium hydrogen sulfate and ethyl acetate. The organic layer was washed with brine, dried over anhydrous sodium sulfate and evaporated under reduced pressure to give
554 mg of the desired title acid (52% yield from the aldehyde form) are obtained.

TLC: Methylene chloride-methanol (9: 1), R _f = 0.37, UV
And PMA.

B. 4'-Fluoro-3,3 ', 5-trimethyl [1,1'-biphenyl] -2-carboxamide In 6.0 ml of dry methylene chloride and 1 drop of dry DMF, the acid of 5
To a suspension of 54 mg (2.14 mmol) (0 ° C. ice bath), 205 μ of distilled oxalyl chloride (2.35 mmol, 1.1 eq)
Is added dropwise with a syringe and the clear yellow solution is stirred for 1 hour at room temperature under an atmosphere of argon. The mixture is evaporated under reduced pressure, washed twice with benzene and dried under reduced pressure to give the crude acid chloride as a yellow oil.

To a mixture of 3.0 ml of THF and 2.0 ml of concentrated ammonium hydroxide (excess amount of cold (0 ° C. ice bath) mixture was added a solution of the above crude acid chloride in THF.
3.0 ml are added dropwise and the clear orange solution is stirred for 1 hour at room temperature under an atmosphere of argon. The mixture was partitioned between water and ethyl acetate, the organic layer was washed with saturated sodium hydrogen carbonate, water and brine, dried over anhydrous sodium sulfate and evaporated to give 528 mg of crude amide as a light orange solid.
(96.1%) is obtained. Recrystallized once from ethyl acetate-hexane to give 435 mg (79.
1%). 197-198 ° C. TLC: diethyl ether-acetone (1: 1), _Rf = 0.83, UV and PMA.

C. 4'-Fluoro-3,3 ', 5-trimethyl [1,1'-biphenyl] -2-methanamine A solution of 5.0 ml of dry THF in cold (0 ° C ice bath) was added to solid lithium ammonium hydride (125 mg, 3.3 mmol). ), And to this gray suspension is added dropwise a solution of 424 mg (1.65 mmol) of the amide of section B in 5.0 ml of THF over 5 minutes. The obtained suspension was stirred at room temperature for 2.5 hours under an argon atmosphere, and then 4
Reflux for 5 minutes. Cool the mixture to 0 ° C (ice bath) and water 125
The reaction is stopped by successively adding μ, 125 μ of 15% sodium hydroxide and 375 μ of water. This is passed through anhydrous sodium sulfate over packed Celite to remove the aluminum salt precipitate. The clear liquid is evaporated under reduced pressure to give the crude amine form as a clear oil. TLC: diethyl ether-acetone (7: 3), _Rf = 0.60, UV and PMA. The amine form is purified as its hydrochloride salt.

A solution of this crude amine form (approximately 1.65 mmol) in 8.0 ml absolute ethanol is treated with concentrated hydrochloric acid 152 μ (1.82 mmol) and the mixture is stirred for 15 minutes at room temperature under an argon atmosphere. The mixture is evaporated under reduced pressure to give a white crystalline solid. The solid is triturated with cold diethyl ether, collected and dried under reduced pressure to give 426 mg (92.4%) of the hydrochloride salt of the title amine as white crystallites.

D. (S) -4-[[[[4'-fluoro-3,3 ', 5-trimethyl [1,1'-biphenyl] -2-yl] methyl]
Amino] methoxyphosphoryl] -3-t-butyldiphenylsilyloxybutanoic acid methyl ester A solution of the methyl ester of Example 1E6 (approximately 2.0 mmol) in 5.0 ml of dry methylene chloride was distilled to 758 μl (4.0 mmol, 2.0 mmol). Equivalent)
And the clear mixture was treated with argon at room temperature for 1
Stir for hours. The mixture is evaporated under reduced pressure, washed with benzene (1 x 15 ml) and dried under reduced pressure.

To a cold (0 ° C.) solution of the above crude silylphosphonate in 7.0 ml dry methylene chloride and DMF (1 drop) is added dropwise 192 μ (2.2 mmol, 1.1 eq) distilled oxalyl chloride. Clear gasses evolve from the clear yellow mixture. The solution is stirred at room temperature for 1 hour, evaporated under reduced pressure, washed with benzene (15 ml x 2) and dried under reduced pressure to give crude phosphonochloridate as a yellow viscous oil.

This phosphonochloridate and 416 mg (1.49 mmol) of dry methylene chloride of the above item C, biphenylamine hydrochloride 1
0 ml cold (0 ° C.) solution was added to triethylamine 641 μ (4.6
Mmol, 2.3 eq) and 4-DMAP 24 mg (0.2 mmol, 0.1
1 eq.) And stir the clear yellow mixture at room temperature under argon atmosphere overnight. The mixture was partitioned between 5% potassium hydrogen sulfate and ethyl acetate, the organic layer was washed with brine, dried over anhydrous sodium sulfate and evaporated under reduced pressure to give 1.19 g of a yellow oil. The crude product on silica gel 60 g,
Purify by flash chromatography and elute with hexane-acetone (7: 3). The product fractions are evaporated to give 588 mg (59.5%) of the desired title phosphonamide as a pale yellow viscous oil. TLC: Hexane
Acetone (7: 3), R _f = 0.20, UV and PMA.

E. (S) -4-[[[[4'-fluoro-3,3 ', 5-trimethyl (1,1'-biphenyl) -2-yl] methyl]
Amino] methoxyphosphoryl] -3-hydroxybutanoic acid methyl ester 588 mg (0.888 mmol) of the silyl ester form of the above item D
A solution of 10.0 ml of dry THF in 203 μl (3.55 mmol, 4.0 eq) of glacial acetic acid and 2.66 ml (2.66 mmol, 3.0 eq) of tetrabutylammonium fluoride in THF (1.0
M) and stir the resulting solution at room temperature under an argon atmosphere overnight. The mixture is poured into cold water and extracted with ethyl acetate. The organic layer is washed with saturated sodium hydrogen carbonate and brine, dried over anhydrous sodium sulfate and evaporated under reduced pressure to give 605 mg of an orange oil. The crude product was subjected to flash chromatography on 36 g of silica gel,
Purify by eluting with hexane-acetone (1: 1). The product fractions are combined and evaporated to give 196 mg (50.4%) of the desired title diester as a light orange oil. TLC: hexane-acetone (1: 1), _Rf = 0.16, UV
And PMA.

F. (S) -4-[[[[4'-fluoro-3,3 ', 5-trimethyl [1,1'-biphenyl] -2-yl] methyl]
Amino] methoxyphosphoryl] -3-hydroxybutanoic acid monolithium salt A solution of 105 mg (0.240 mmol) of the diester of the above-mentioned E in 2.0 ml of dioxane was added to 1.0N lithium hydroxide 288 μ (1.2 μm).
Eq.) And stir the white suspension at room temperature under argon for 4.0 h. The mixture is diluted with water and the liquid is evaporated under reduced pressure. The residue is chromatographed on HP-20 (100 ml bed) and eluted with a water / acetonitrile linear gradient eluent. The product fractions are combined and evaporated. The residue is taken up in 50 ml of water, passed through a 0.4 μm polycarbonate film and lyophilized to give 70 mg of the desired title lithium salt as a white solid (62.7% based on hydrate weight). TLC: methylene chloride-methanol acetic acid (20: 1: 1), _Rf = 0.19, UV and PMA.

Elemental analysis, as C ₂₁ H ₂₆ NO ₅ PFLi + 2.41 mol H ₂ O (MW472.75), calculated value: C, 53.35; H, 6.57; N, 2.96; F, 4.02; P, 6.55, measured value: C , 53.35; H, 6.52; N, 2,98; F, 4.05; P, 6.59.

H ¹ · NMR (400 MHz): 2.26 ~ 2.44 (2H, m, -CH ₂ CO ₂ Li) 2.29 (3H, s, aromatic methyl) 2.31 (3H, d, aromatic methyl α-fluorine, J _HF = 1.4Hz) 2.47 (3H, aromatic Group methyl) 3.46 and 3.50 (3H, 2 double bonds, 2 diastereomers,
J _HP = 10.5Hz) _{4.17 (1H, m, -CH 2} CH (OH) CH 2 CO 2 Li) 6.84~7.21 (5H, m, aromatic protons) and described in Examples according to the method described in configuration and effect of the embodiment 8-20 INVENTION As described above, a compound of the present invention represented by the following formula (substituents thereof are shown below) is produced.

Example 21 (S) -4- (diisopropyloxyphosphoryl) -3
-[[(1,1-Dimethylethyl) diphenylsilyl] oxy] butanoic acid methyl ester 21.70 g (45.1 mmol) of iodide of Example 1E3 is stirred for 30 minutes under high vacuum. 93.92 g (113.37 ml, 0.451 mol) of freshly distilled triisopropyl phosphonate are added in one portion and the mixture is stirred and heated under an argon atmosphere on an oil bath at 155 ° C. for 16.5 hours. Then the mixture is cooled to room temperature. Excess triisopropyl phosphonate and volatile reaction products are distilled off by short path distillation (10 mmHg) followed by Kugelrohr distillation (0.50 mmHg, 100 ° C., 8 hours). The product was further purified by flash chromatography (95 mm diameter column, 6 ″ / Merck silica gel, hexane / acetone / toluene (6/3/1) eluent, 2 ″ / min flow rate, 50 ml fraction), 17.68 g (33.9 g) of the title isopropyl phosphonate as a transparent viscous oil.
6 mmol, 75% yield) was obtained. TLC: silica gel, R _f =
0.32 (hexane / acetone / toluene (6: 3: 1)). ¹ H ・ NMR: (270 MHz, CDCl ₃ ) δ 7.70 to 7.65 (m, 4H) 7.45 to 7.35 (m, 6H) 4.57 to 4.44 (m, 3H) 3.59 (s, 3H) 2.94 and 2.88 (2xd, 1H, J = 3.7Hz) 2.65 and 2.60 (2xd, 1H, J = 7.4Hz) 2.24 to 1.87 (m continuous, 2H) 1.19 and 1.12 (2xd, 12H, J = 6.3Hz) 1.01 (s, 9H) Example 22 (S) -4- (hydroxymethoxyphosphoryl) -3-
[[(1,1-Dimethylethyl) diphenylsilyl] oxy] butanoic acid methyl ester dicyclohexylamine (1: 1) salt 10.66 g (3
0.5 mmol) is stirred in 80 ml of dry methylene chloride under an atmosphere of argon at room temperature. To this solution, 8.44 g (8.71 g) of bistrimethylsilyltrifluoroacetamide (BSTFA)
ml, 32.8 mmol) over 5 minutes and then 7.84 g (6.75 ml, 51.3 ml) of trimethylsilyl bromide (TMSBr).
Mmol) is added dropwise over 10 minutes. After stirring at room temperature for 20 hours, 200 ml of 5% potassium hydrogen sulfate aqueous solution is added to the mixture to stop the reaction, and the mixture is stirred vigorously for 15 minutes. The aqueous layer is extracted 3 times with ethyl acetate. Combine the organic extracts with brine 1
Wash twice, dry over sodium sulfate and concentrate under reduced pressure.
The residue is azeotroped twice with 50 ml of toluene. The precipitate formed is suspended in toluene. Concentrate the liquid and repeat the azeotropic / over-operation. The liquid obtained is evaporated under reduced pressure and then kept under high vacuum for 5 hours with suction. The viscous, clear oil obtained is stirred at room temperature in 50 ml of dry pyridine under an atmosphere of argon. To this solution was added 4.65 g (22.6 mmol) of dicyclohexylcarbodiimide (DCC), and then 1.31 g of methanol (1.67 ml, 4.10 mmol).
Add. After stirring for 20 hours at room temperature, the reaction mixture is passed through a Celite pad in a sintered glass funnel. Wash celite with ethyl acetate, combine with the solution and evaporate under reduced pressure. The residue is redissolved in ethyl acetate and washed twice with 5% aqueous potassium hydrogensulfate solution and once with brine. The organic extracts are dried over sodium sulphate, filtered, concentrated, azeotroped twice with toluene and suspended in toluene. The solution was concentrated, azeotroped and evaporated again under reduced pressure and left under high vacuum for 6 hours to obtain 10.2 g of phosphonate monoester as a transparent viscous oil (yield> 100%).
Get. TLC: silica gel, _Rf = 0.50 (n-propanol / ammonium hydroxide / water (7: 2: 1)). This phosphonate monoester [1.21 g was aspirated and kept under high vacuum for 4 hours to give 1.16 g (2.57 mmol)] was dissolved in 10 ml dry ethyl ether to give 0.48 dicyclohexylamine.
1 g (0.528 ml, 2.65 mmol) is added dropwise. The resulting homogeneous solution is left at room temperature for 7 hours to allow significant crystals to form. The mixture is kept at -20 ° C for 16 hours, then warmed to room temperature and passed. The crystals are washed with cold dry ethyl ether, then suctioned over phosphorus pentoxide and kept under high vacuum for 18 hours. Then, the crystals were sucked at 45 ° C. and kept under high vacuum for 4 hours to obtain the title dicyclohexylamine as a white powdery solid.
1.25 g (1.98 mmol, 77% yield) was obtained. Melting point 155-15
6 ° C. TLC: silica gel, _Rf = 0.57 (20% methanol / methylene chloride). ¹ H ・ NMR (270 MHz, CDCl ₃ ): δ7.71 to 7.65 (m, 4H) 7.40 to 7.32 (m, 6H) 4.02 (m, 1H) 3.52 (s, 3H) 3.28 and 3.22 (m, 1H) 3.11 (d, 3H, J = 11Hz) 2.77 to 2.64 (m, 2H) 2.62 to 2.56 (m, 1H) 1.92 to 1.08 (continuous m, 22H) 1.00 (S, 9H) Mass spectrum: (FAB) 632 ( M + H) ⁺ IR: (KBr) 3466 to 3457 (wide) 3046, 3016, 2997, 2937, 2858, 2836, 2798, 2721, 27
04, 2633, 2533, 2447, 1736, 1449, 1435, 1426, 137
9, 1243, 1231, 1191, 1107, 1074, 1061, 1051, 820cm
^-1 .

Elemental analysis as _{C 22 H 31 O 6 PSi ·} C 12 H 23 N, Calculated: C, 64.63; H, 8.61 ; N, 2.22, Found: C, 64.51; H, 8.4
9; N, 2.18.

 ─────────────────────────────────────────────────── ─── Continued Front Page (72) Inventor Scott Adams Biller, Nancy Lane 136, Ewing, New Jersey, USA (72) Inventor Eric Michael Gordon 126th Raining Avenue, Pennington, New Jersey, USA

Claims (6)

(57) [Claims] 1. The formula: [Wherein R is OH, lower alkoxy or lower alkyl,
^Rx is H or lower alkyl; X is -O- or -NH-; n is 1 or 2; Z is A lipophilic group represented by (in the group, a double bond may be present if necessary; R ¹ , R ² , R ^2a and R ^2b may be the same or different; H, halogen, lower Alkyl, haloalkyl, phenyl, substituted phenyl or OR ^y (R ^y is H, alkanoyl, benzoyl, phenyl, halophenyl, phenyl (lower) alkyl, lower alkyl, cinnamyl, haloalkyl, allyl, cycloalkyl (lower) alkyl, adamantyl (lower ) Alkyl or substituted phenyl (lower) alkyl); R ⁵ and R ⁵ ′ are the same or different and are H, lower alkyl or OH (one of R ⁵ and R ⁵ ′ is present when the carbon to which they are bound to a double bond) , R ⁶ is lower alkyl Or aryl -CH ₂ -, R ^6a is lower alkyl, hydroxy, oxo or halogen, and q is 0, 1, 2 or 3); represent. When R ^x is hydrogen or lower alkyl, it forms a physiologically hydrolyzed and physiologically acceptable ester form in the free acid form or salt form, respectively.], The enzyme 3-hydroxy- A compound useful for suppressing 3-methylglutaryl-coenzyme A reductase or a salt thereof. 2. X is --O--, R is lower alkoxy, and Z is The compound according to claim 1, which is 3. X is --NH--, R is lower alkoxy, (C
H ₂ ) _n is CH ₂ , and Z is The compound according to claim 1, which is 4. (S) -4-[[[4'-fluoro-3,
3 ', 5-trimethyl [1,1'-biphenyl] -2-yl]
Methoxy] methoxyphosphoryl] -3-hydroxybutanoic acid methyl ester or its monolithium salt, (S) -4-[[[4'-fluoro-3,3 ', 5-trimethyl [1,1'-biphenyl]- 2-yl] methoxy] hydroxyphosphoryl] -3-hydroxybutanoic acid dilithium salt, (3S) -4-[[[4'-fluoro-3,3 ', 5-trimethyl [1,1'-biphenyl] -2 -Yl] methoxy] methylphosphoryl] -3-hydroxybutanoic acid monolithium salt, (S) -4-[[[2,4-dichloro-6-[(4-fluorophenyl) methoxy] phenyl] methoxy] methoxyphosphoryl ] -3-Hydroxybutanoic acid monolithium salt, (3S) -4-[[[2,4-dichloro-6-[(4-fluorophenyl) methoxy] phenyl] methoxy] hydroxyphosphoryl] 3-Hydroxybutanoic acid dilithium salt, (3S) -4-[[[2,4-dichloro-6-[(4-fluorophenyl) methoxy] phenyl] methoxy] methylphosphoryl] -3-hydroxybutanoic acid or its methyl Ester, or (S) -4-[[[4'-fluoro-3,3 ', 5-trimethyl [1,1'-biphenyl-2-yl] methyl] amino] methoxyphosphoryl] -3-hydroxybutanoic acid The compound according to claim 2, which has a compound name of a monolithium salt. 5. The formula: [Wherein, R ^a represents lower alkyl or lower alkoxy] or a stereoisomer thereof. 6. The formula: [In the formula, R ^a ′ represents lower alkyl or OH] or a stereoisomer thereof.