PL157761B1 - Method of obtaining novel phosphorus containing derivatives of 3-hydroxybutanic acid - Google Patents

Abstract

Compounds which are useful as inhibitors of cholesterol biosynthesis and thus as hypocholesterolemic agents are provided which have the structure <IMAGE> wherein R is OH, or salts thereof or lower alkoxy; R<x> is H or alkyl; X is -CH2-,-CH2-CH2-,-CH=CH-, -C IDENTICAL C-, -CH2CH2CH2- or -CH2O- (where O is linked to Z); Z is a hydrophobic anchor, such as <IMAGE> wherein the dotted lines represent optional double bonds. New intermediates used in preparing the above compounds, pharmaceutical compositions containing such compounds and a method for using such compounds to inhibit cholesterol biosynthesis are also provided.

Description

The present invention relates to a process for the preparation of new phosphorus-containing 3-hydroxybutanoic acid derivatives which inhibit the activity of 3-hydroxy-3-methylglutaryl coenzyme A, useful for inhibiting cholesterol biosynthesis, exhibiting hypocholesterolemic activity. New intermediates are used in the synthesis of these compounds.

F. M. Singer et al., Proc. Soc. Exper. Biol. Med., 102, 570 (1959) and F. H. Hulcher, Arch. Biochem. Biophys., 146,422 (1971) disclosed that certain mevalonic acid derivatives inhibit cholesterol biosynthesis.

In US patents US Am. Nos. 4,049,495, 4,137,322 and 3,983,140 Endo et al. disclose a fermentation product showing inhibitory activity in cholesterol biosynthesis. This product was named compactin and Brown et al., J. Chem. J. Soc. Perkin I 1165 (1976) reported that it has the complex structure of the mevalonic acid lactone.

British Patent No. 1,586,152 discloses a group of synthetic compounds of the general formula 20, wherein E is a direct bond, a C 1-3 alkylene bridge or a vinylene bridge and the various R symbols represent different substituents.

The activity of the compounds described in this British patent is less than 1% of the compactin activity.

In U.S. Patent No. US Am. No. 4,375,475 Willard et al. discloses hypocholesterolemic and hypilipemic compounds of general formula 21 wherein A is hydrogen or methyl, E is a direct bond, -CH2-, -CH2-CH2-, -CH2-CH2-CH2 or -CH = CH-, Ri, R2 and R3 are selected from hydrogen, halogen, C1-4alkyl, C1-4 haloalkyl, phenyl, phenyl substituted with halogen, C1-4alkylyl, C1-4, and C1-4H4 C 1-4 alkyloxy, C 1-4 haloalkyl, and OR 4 groups in which R 4 is hydrogen, C 1-4 H 2 ianoyl group, benzoyl, phenyl, halophenyl, phenyl-C 1-3 alkyl, C 1-9 alkyl, cinnamyl, C1-4 haloalkyl, allyl, cycloalkyl-C1-3alkyl, adamantyl-C1-3alkyl, or substituted phenyl-C1-3alkyl where each substituent is selected from halogen, C1-4alkoxy, C1-4alkyl or C1-4haloalkyl ; and the corresponding dihydroxy acids resulting from the hydrolytic opening of the lactane ring and the pharmacologically acceptable salts of these acids and the C 1-3 alkyl and phenyl-, dimethylamino- or acetylamino-substituted C 1-3al4 esters

157 761 kilograms of dihydroxy acids, all these compounds being enantomers of the 4R configuration in the tetrahydropyran part of the trans racem represented by the above general formula.

In International Publication No. WO 84/02131, PCT / PCT / EP 83/00308 / (based on U.S. Patent Applications No. 443,668, filed November 22, 1982 and No. 548,850, filed November 4, 1983) by Sandoz AG discloses heterocyclic analogs of mevalonolactone and its derivatives of general formula 22, wherein one of the symbols R and Ro is a group of formula 23 and the other is C 1-6 primary or secondary alkyl, C 3-6 cycloalkyl or a phenyl- (CH ² ) group m-, wherein in formula 23 R4 is hydrogen, C1-4alkyl, C1-4alkoxy (except tert-butoxyl), trifluoromethyl, fluoro, chloro, phenoxy or benzyloxy, R5 is hydrogen, C1-3alkyl, Ci_3alkoxy, trifluoromethyl, fluoro, chloro, phenoxy or benzyloxy group, Rsa is hydrogen, Ci_2alkyl, Ci_2alkoxy, fluoro or chloro atom, and m is 1.2 or 3, with the proviso that when R4 is a hydrogen atom then both R5 and Rsa must be an atom hydrogen, no more than one of R4 and Rs is trifluoromethyl, no more than one of R4 and Rs is phenoxy, and no more than one of R4 and Rs is benzyloxy; R2 is hydrogen, C1-4alkyl, C1-4alkyl, C1-4alkoxy (except tert-butoxy), trifluoromethyl, fluoro, chloro, phenoxy or benzyloxy; R3 is hydrogen, C1-3alkyl, C1-3alkoxy, trifluoromethyl, fluoro, chloro, phenoxy or benzyloxy, with the proviso that when R2 is hydrogen then R3 must be hydrogen, not more than one of R2 and R3 is trifluoromethyl, no more than one of R3 and R3 is phenoxy and no more than one of R2 and R3 is benzyloxy; X is - (CH2) n- or -CH = CH (n = 0.1, 2, or 3) and Z is a group of formula 24 wherein Re is hydrogen or C1-3alkyl; in the form of free acid or in the form of a physiologically hydrolysable and physiologically acceptable ester or <lactone, or in the form of a salt.

British Patent Specification No. 2162-179 A discloses naphthyl analogs of mevalolactone useful as inhibitors of chloesterol biosynthesis of the general formula wherein R1 is C1-3aIkyl, Z is a group of formula 26 or formula 27, wherein R7 is hydrogen, a hydrolysable ester group or a cation.

European Patent No. 164-698-A discloses preparation of lactones useful as anti-hipercholesteroloemii by treating the amide orgamczn ^y m hahgenkiem sulfone ^yl uo formula ^SO2X R® ^and usum ^ę five December protective y ^p ^ ^p ^ i ^k summarized in Scheme where X is a halogen atom, Pr is a protecting group karhnolow ^ ^r1 is wo ^d ol hb meth ^y k R ^{3 and R 4} is ^a is hydrogen, CH Alku ht "phenyl-3ćtlkil a phenyl group optionally substituted with Ci- 3alkyl, C 1-3 alkoxy or c ^{h | orowca, R5 g} ru ^PE of formula ²⁸ H ^{B 29 and} Q is ^g ru ^PE of formula ³⁰ h ^b 3 ^1, wherein the wzorac ^h t ^s c ^{h R6} is wo ^d ol hb December ^{EP dihydro k} s ^y hwą; ^R is wo ol h ^{d b} metyk ^a, c ^{and d} represent ewentuahh ^p odwójne wązama; ^R7 ru ^{g u} re fenyhwą or benzyloxy, a ring optionally substituted by a Ci-3alkyl group or halo C ^{hl oΓowca, R8 and R9} is ^and C Alkham hb atom ch ^{| orowca,} and ^R5 is Ci-tflkH, phenyl-H ^b one or di- (Ci-3 alkyl) phenyl.

In the published patent application PS No. 3 525 256 discloses naphthyl Anah ^g mewalonolakton ^yellow wo formula 3 ^2, wherein ^Y m ^{R 1} ALIKA ^{il "Z} is ^g ru ^PE of formula ^26, wherein ^R7 is hydrogen hb he ^g ai ^± c h ^{± y} drohzie December ^EP ester ^and the lu ^b of formula 33 are useful as cholesterol biosynthesis inhibitors and in the treatment of atherosclerosis.

International Publication No. WO 8402-903 of PCT Application (based on US Patent Application No. 460,600, filed Jan. 24, 1983) to Sandoz AG discloses mevalonolactone analogs useful as hypolipoproteinemic agents of general formula 34, wherein two Ro groups together form a radical of formula 35 or a group - (CH ₂ ) 4- in which R 2 represents a hydrogen atom, C 1-4 alkyl, C 1-3 alkoxy (except A1, N, d-butoxy) , trifluoromethyl, fluoro, chloro, phenoxy or benzyloxy, R3 is hydrogen, Ci-3alkyl, Ci-3alkoxy, trifluoromethyl, fluoro, chloro, phenoxy or benzyloxy with the proviso that not more than one of R2 and R3 is

Trifluoromethyl, not more than one of R2 and R3 is phenoxy and not more than one of R2 and R3 is benzyloxy; R 1 is hydrogen, C 1-6 alkyl, fluoro, chloro or benzyloxy; R4 is hydrogen, _Ci-4 alkyl, Ci-4 alkoxy (except Illrzęd. -Butoksylu), trifluoromethyl, fluoro, chloro, phenoxy or benzyloxy; R 5 is hydrogen, C 1-3 alkyl, C 1-3 alkoxy, trifluoromethyl, fluoro, chloro, phenoxy or benzyloxy; Rsa is hydrogen, C1-2alkyl, C1-2alkoxy, fluoro or chloro with the proviso that not more than one of R4 and R5 is trifluoromethyl, not more than one of R4 and Rs is phenoxy and not more than one of R4 and R8 is trifluoromethyl. no more than one of R4 and R8 is phenoxy and no more than one of R4 and R3 is benzyloxy; X represents a group of formula -CH2) n- or a group of formula 36, in which n is the number 0,1,2 or 3 and both q symbols are 0 or one is 0 and the other is 1, Z is the group of Formula 37, wherein Re is hydrogen or C1-3alkyl, provided that the -Χ-Ζ group and the phenyl group with R4 are ortho to each other; in free acid form or in the form of a physiologically hydrolysable and physiologically acceptable ester or a δ lactone thereof, or in the form of a salt.

In U.S. Patent No. US Am. No. 4,613,610 discloses a series of 6-pyrazole-2,4-dihydrohexane-5-carboxylic acid HMG-CoA reductase inhibitors of general formula 38 wherein Ri is Ci-ealkyl containing no asymmetric carbon atom, R2 and Rs each independently is hydrogen, C1-3alkyl, n-butyl, i-butyl, tert-butyl, C1-3alkoxy, n-butoxy, isobutoxy, trifluoromethyl, fluorine, chlorine, phenyl, phenoxy or benzyloxy, each of R3 and Re independently is hydrogen, C1-3alkyl, C1-3alkoxy, trifluoromethyl, fluoro, chloro, phenoxy or benzyloxy; each of R4 and R7 independently is hydrogen, C1-2alkyl, C1-4 alkoxy, fluoro or chloro with the proviso that no more than one of R2 and R5 is trifluoromethyl, no more than one of R2 and R3 is a phenoxy group, not more than one of R2 and R3 is benzyloxy, not more than one of R5 and Re is trifluoromethyl, not more than one of R5 and Re is phenoxy, and not more than one of R5 and Re is benzyloxy; X represents groups of the formulas - (CH2) m-, -CH = CH-, -CH - CH-CH2- or -CH2-CH = CH- where m is 0, 1, 2 or 3 and Z is a group of formula 39 or formula 40, wherein R10 and Ru are hydrogen or C1-3alkyl and R11 is hydrogen, R12 or M, where R12 is a physiologically acceptable salt and a hydrolyzable ester group, and M is a cation with the proviso that (a) the -Χ-Ζ group is on the 4- or 5-position of the pyrazole ring, and (b) the Ri group and the -Χ-Ζ group are ortho to each other.

International Publication No. WO 8607-054A of the PCT Application discloses mevalonolactone imidazole analogues useful for the treatment of hyperlipoproteinemia and atherosclerosis, represented by general formula 41, wherein R1 is alkyl, cycloalkyl, adamantyl-1 or R4, Rs Repubstituted phenyl (group A); R2 is alkyl, cycloakyl, adamantyl-1 or R7, Re, R9-substituted phenyl (group B); R3 is hydrogen, alkyl, cycloalkyl, adamantyl-1, styryl or R10, R11, R12 -substituted phenyl (group C); X is a group of the formula - (CH2) _m -, -CH = CH-, -CH - CH-CH2 or -CH2-CH = CH-; m is the number 0-3, Z is the group of the formula -CH (OH) -CH2-C (Ri3) (OHECH2-COOR14 (group a); -Q-CH2- (C / Ri3) (OH) -CH2-COORi4 (group c) or a group of formula 42 (group b), wherein Q is CO or -C (OR-is) 2-; Ri5 is primary or secondary alkyl, where each R15 is the same or R15 + R15 is (CH2) 2 or (CH2) 3 and R13 is hydrogen or C1-3alkyl; R14 is hydrogen, Rw or M; Rie is an ester group and M is a cation, except that Z may be a group c) only when X is CH = CH or CH2-CH-CH and / or R13 is C1-3alkyl; R4, R7 and R10 are C1-3alkyl, η-, iso- or Illzzed.-butyl, C1-4 alkoxy, n- or iso-butoxy, CF3, fluoro, chloro, bromo, phenyl, phenoxly or benzyloxy; R5, Re and R11 are hydrogen, Ci-3alkyl, Ci-3alkoxy, trifluoromethyl, fluoro, chloro, bromo, C ^O OR ^17, N (R19) ^2, Group ^{E f} enoxaparain ^yl head ^{and L} at ^bb enzylo ^{k yl} words ^{s; r17} represents the hydrogens in ^{R l b} of ^{M; Rie} is C aHok η-, theological lu U ^ ^b ^ zed hu ^l ^ u ^b tenzyl; ^r19 is alkfl; ^{R9 and R12} are hydrogen, C1-2alkyl, C1-2alkoxy, fluoro or chloro, with the proviso that (1) no more than one substituent from each of A, B and C is trifluoromethyl, not more than one of

157,761 of each of A, B, and C is trifluoromethyl, no more than one of each of A, B, and C is phenoxy, and no more than one of each of A, B, and C is benzyloxy; (2) when Z is c [Q is C (ORi5) 2], the compound is in the free base form and / or (i) R14 is Rw and each R17 is independently Rie, or (ii) Rn is Each of R17 is independently Rie or M; and (3) when R11 and / or at least one of R17 is M, the compound is in the form of a free base.

Unless otherwise stated, all "alkyl" groups are groups of the formula 1-6 carbon atoms and contain no asymmetric carbon atoms, and "cycloalkyl" is groups of 3-7 carbon atoms.

International Publication No. WO 8603-488-A of the PCT Patent Application discloses identical mevalolactone analogs useful as antihypoliproteinemic and anti-atherosclerotic agents in the form of free acid, ester, δ-lactone or salt, and general formula 43 wherein R is hydrogen. or C1-6 primary or secondary alkyl; R1 is C1-6 primary or secondary alkyl; or R + R1 is (CHz) m or (Z) -CH2-CH = CHCH2; m is the number 2-6; Ro is C1-6alkyl, C3-7cycloalkyl or R4, Rs, R6 - substituted phenyl; R2 and R4 are hydrogen, C1-4alkyl, C1-4alkoxy (except tertiary butoxylate), trifluoromethyl, fluoro, chloro, phenoxy or benzyloxy; R3 and R5 are hydrogen, Ci-3alkyl, Ci-3alkoxy, trifluoromethyl, fluoro, chloro, phenoxy or benzyloxy; R6 is hydrogen, C1-2alkyl, C1-2alkoxy, fluoro or chloro with the proviso that each of the phenyl and indene rings may be only one of the trifluoromethyl, phenoxy or benzyloxy substituents; X is a group of formula (CH2) n or - (CH2) q-CH = CH (CH2) q-; n is 1 to 3; q are both zero, or one of them is zero and the other is 1; Z is a group of formula -Q-CH2-C (Rio) (OH (-CH2COOH, free acid, ester, δ-lactone or salt form; Q is CO-, -C (OR7) 2, or CHOH; R ') 7 are the same primary or secondary alkyls or together form a group of formula (CH2 (2 or (CH2 (3 and R10 is hydrogen or C1-3alkyl, with the proviso that Q may be other than CHOH when X is CH = CH) or CH2-CH = CH and / or R10 is C1-3alkyl.

In U.S. Patent No. US Am. No. 4,647,576 discloses novel C- and N-substituted pyrroles useful as hypolipidaemic and hypocholesterolemic agents of general formulas 44 and 45, wherein X is -CH2-, -CH2CH2- or -CH (CH3) CH2-; R1 is 1- or 2-naphthyl, cyclohexyl, norbornenyl, phenyl optionally substituted with fluoro, chloro, hydroxy, trifluoromethylene, C1-4alkyl, C1-4alkoxy or C2-ealkanoyloxy, 2-, 3- or 4-pyridinyl group, or N-oxides, or a group of formula 46, where R5 is C1-4alkyl and hal is chlorine, bromine, CN, CF3, phenyl, C1-4alkyl, C2-ecarboalkoxy, CH2OR6 or -CH2OCONHR7, where R6 is hydrogen or C1-4alkanoyl and R7 is alkyl or phenyl optionally substituted with chloro, bromo or C1-4alkyl; or R2 and R3 together represent a group of the formula - (C ^ ₂ ) n-, -CH2OCH2-, -CON (Rs) CO- or CON (R9) N (Rio) CO-, in which n is the number 3 or 4; Re is hydrogen, C1-6alkyl, phenyl or benzyl, R9 and R10 are hydrogen, C1-4alkyl or benzyl; and R4 is C1-4alkyl, cyclopropyl, cyclobutyl, or trifluoromethyl.

European Patent Application 0221 025 A1 discloses heterocyclic analogs of mevalonolactone and derivatives thereof, with the general formula 47 in which Ra is -Χ-Ζ, Rb is R2, Rc is R3, Rd is R4, and Y is a group of formula = N-R1 or Ra is R1, Rb is the group -Χ-Ζ, Rc is R2, Rd is R3 and Y is oxygen, sulfur or the group = N (R4); wherein R1, R2, R3 and R4 are independently C1-4 alkyl not containing an asymmetric carbon atom, C3-7cycloalkyl or a ring of general formula 48, or for R3 and R4 additionally hydrogen, or in the case of R3 when Y is oxygen or sulfur, a group of general formula 49 wherein R 17 is hydrogen or C 1-3 alkyl, and R w and R 19 are independently hydrogen, C 1-3 alkyl, or phenyl; each R8 is independently hydrogen, C1-3alkyl, n-butyl, isobutyl, tert-butyl, C1-3alkoxy, n-butoxy, isobutoxy, trifluoromethyl, fluoro, chromium, bromo, phenyl, phenoxy or benzyloxy; each of R4 is independently hydrogen, C1-3alkyl, C1-3alkoxy, trifluoromethyl,

157 761 fluorine, chlorine, bromine, phenoxly or benzyloxy group, each of R? is independently hydrogen, C1-2alkyl, C1-2alkolsyl, fluoro or chloro, with the proviso that there may be only one trifluoromethyl, one phenoxy or benzyloxy group in each ring A; X is the group (Cj or (CH2) qCH = (CHz) q, m is the number 1, 2, 3 or 0, and both q are zero or one of them is zero and the other is 1; Z is the group of formula 50 wherein R9 is hydrogen or C1-3alkyl, in the free acid or ester of its δ lactone, or in the form of a salt, depending on which of these compounds is intended for use as a hypolipoproteinemic and antiatherosclerotic agent.

In Tetrahedron Letters, 29,929,1988, the synthesis of the 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitor Formula 51 wherein R is Na or C2H5 is disclosed.

European Patent Application No. 127 848-A discloses acid derivatives

2-hydroxy-4-thia-o-arylalkanecarboxylic compounds of general formula 52 where Z is a group of formula 53 or formula 54, n is 0, 1 or 2, E is -CH2-, -CH2-CH2 -, -CH2-CH2-CH2-, -CH = CH-CH2- or -CH2-CH = CH-; R1, R2 and R3 are, for example, hydrogen, chlorine, bromine, fluorine, C1-4alkyl, phenyl, substituted phenyl or a group of the formula OR7 where R7 is e.g. hydrogen, C2ealkanoyl, benzoyl, phenyl, substituted phenyl, C 1-4 alkyl, cinnamyl, C 1-4 haloalkyl, allyl, cycloalkyl-C 1-3 alkyl, adamantyl-C 1-3 alkyl, or phenylC 1-3 alkyl; R4, Rs and Re are hydrogen, chlorine, bromine, fluoro or C1-3alkyl; and X is e.g. hydrogen, C1-3alkyl, an alkali metal cation or an ammonium cation.

These compounds have an antihyperocholesterolemic effect due to the ability to inhibit 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMG-Co A) and have an antifungal activity.

Acid derivatives are disclosed in the French patent application mr 2 596 393 A

3-carboxy-2-hydroxy-propane-phosphono and their salts, useful as hypolipaemic agents of general formula 55 wherein R1 and R2 are hydrogen, lower alkyl, or optionally substituted aralkyl and R3 and R4 are hydrogen, lower alkyl or optionally substituted aryl or aralkyl. These compounds were disclosed to provide greater reductions in blood cholesterol, triglycerides, and phospholipids than do meglutol.

European Patent Application No. 142 146-A discloses mevincoline compounds of the general formula 56, in which R1 is e.g. hydrogen or C1-4alkyl, E is -CH2CH2-, CH = CH- or - (CH2) _r - a Z represents: (1) a group of general formula 57 in which X is -O- or a group -NR9 in which R9 is hydrogen or C1-4alkyl; R? is C2-ealkyl; and R ⁸ is hydrogen or methyl; (2) a group of general formula 58 in which R ^w , R and R ¹² are independently, for example, hydrogen, halogen, or C1-3alkyl; (3) a group of the general formula 59, wherein n is 0-2, and R ¹ 4 represents a halogen atom or Cia4alkil; or (4) a group of general formula 60. These compounds are inhibitors of HMG-Co A reductase.

The process of the present invention produces phosphorus-containing compounds which inhibit the enzyme 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMG-Co A reductase) and are therefore useful as hypocholesterolemic agents. These compounds contain a moiety of general formula 6I, where X is -cis -CH = CH and Z is "hydrophobic anchor."

The term "hydrophobic anchor" as used herein refers to a lipophilic group which, when linked to the upper side of a HMG-like chain of a molecule via a -CH2-CH2- binding group, binds to a hydrophobic enzyme pocket not used in the bonding of the HMG-Co A substrate, resulting in increased potency activities compared with compounds where Z is hydrogen.

The compounds of the present invention are compounds of the general formula I wherein R is hydroxy or lower alkoxy, R ^x is hydrogen or lower alkyl and Z is a hydrophobic anchor as defined below, optionally in the form of pharmacologically acceptable salts.

The term "salt or" salts refers to basic salts formed with inorganic and organic bases. Such salts include ammonium salts, alkali metal salts such as lithium, sodium and potassium salts (which are preferred), alkaline earth metal salts such as calcium and magnesium salts, salts with organic bases such as salts with amines, e.g. dicyclohexylamine salt, benzothiazine salt, N-methyl-D-glucamine salt, hydrabamine salts, salts with amino acids such as arginine, lysine and the like. Non-toxic, pharmacologically acceptable salts are preferred, although other salts are also useful, for example, in isolating or purifying the product.

The hydrophobic anchors in the compounds according to the invention are of the general formulas 4,5,6,7, 8,9, 10, 11, 12,13, 14 or 15, where in formula 15 the dashed lines optionally represent double bonds, as, for example, in formulas 62, 63, 64, 65, 66, 67 and 68. In these formulas, R 1, R 2, R 2® and R 2b may be the same or different and are each independently hydrogen, halogen, lower alkyl, haloalkyl, phenyl, substituted phenyl, or OR ^y where R ^y is hydrogen, alkanoyl, benzoyl, phenyl, halophenyl, phenyl lower alkyl, lower alkyl, cinnamyl, haloalkyl, allyl, cycloalkyl lower alkyl, adamantyl lower alkyl or substituted phenyl-lower alkyl.

When Z is a group of general formula 15, then -R ⁵ and R ⁵ , the same or different, are hydrogen, lower alkyl or hydroxy. Re is a group of formula 16, such as a group of formula 17, or an aryl group. CH2-, R ^6a represents lower alkyl, hydroxyl, keto or halogen, q is 0, 1, 2 or 3 and R ⁷ is hydrogen or lower alkyl.

Z is a moiety of the formulas 6, 7, 8, 9, 10, 11, one of the symbols R ³ and R ⁴ is a group of general formula 18 and the other is lower alkyl, cycloalkyl or phenyl- (CH 2) p - where p is 0, 1, 2, 3 or 4; wherein in the formula R 18 ⁿ is hydrogen, lower alkyl, lower alkoxy (except t-butoxy), halogen, phenoxy or benzyloxy, R ¹ 4 is hydrogen, lower alkyl, lower alkoxy, halogen, phenoxy or benzyloxy, R ^m ® is hydrogen, lower alkyl, lower alkoxy or halogen, with the proviso that when R1 ³ is hydrogen, then R ™ and R ⁰ must be hydrogen and when R1 is hydrogen then R ^is must be hydrogen, not more than one of R1 ³ and R14 is trifluoromethyl, not more than one of R 1 and R ³ is a group ^of phenoxy, and not more than one of R ¹³ and R ™ is a benzyloxy group; R ⁸ is hydrogen, C 1-4 alkyl, C 3-6 cycloalkyl, C 1-4 alkoxy (except tert-butoxyl), trifluoromethyl, fluoro, chloro, phenoxy or benzyloxy; R ⁹ is hydrogen, Ci-3alkyl, Ci-3alkoxy, trifluoromethyl, fluoro, chloro, phenoxy or benzyloxy, with the proviso that when R8 is hydrogen then R 9 must be hydrogen, not more than one of R 8 and R9 is trifluoromethyl, no more than one of R8 and R9 is trifluoromethyl, no more than one of R8 and R9 is phenoxy, and no more than one of R8 and R9 is benzyloxy; R ¹⁰ and R ¹¹ independently represent hydrogen, alkyl, cycloalkyl, adamantyl-1 or a group of general formula 69 in which R ³ , R ^M and R ^a are as defined above, q is 0, 1, 2, 3 or 4; Y is oxygen, sulfur or NR ^w .

When Z is a group of formula 12, Ra in this formula is hydrogen or a primary or secondary C 1-6 alkyl, R ^b is a primary or secondary C 1-6 alkyl or Ra and R b together form a group of the formula (CH 2), wherein r is the number 2,3,4,5 or 6, or a group of formula (cis) -CH2-CH = CH-CH2; R ² is lower alkyl, cycloalkyl or a group of formula 18, wherein R 8, R 9, R ⁿ , R 14 and R 14® are as defined above.

When Z is a group of formula 13, then R ^{1 and} 6 are both hydrogen, chlorine, bromine, CN, trifluoromethyl, phenyl, C 1-4 alkyl, C 2- ealkoxycarbonyl, -CH 2 OR 17 or -CH ² OCONHR ¹ 8, where R ⁿ is is hydrogen or Ci-6alkanoil and R ¹ 8 is alkyl or phenyl optionally substituted by fluorine, chlorine, bromine or a Ci-4 alkyl; or R 15 and R 6 together represent ^one group of the formula - (CH2) s-, -CH2OCH2-, -CON (R ^W) CO or -CON (R) N (R) -CO-, wherein in these formulas, s is the number of 3 or 4, R is a hydrogen atom, Ci-salkil, phenyl or benzyl, and R20 and R21 are hydrogen, Ci-4 alkyl or benzyl, and ^R3 and R4 are as defined above.

When Z is a group of general formula 14, then R 22 is lower alkyl, cycloalkyl, adamantyl-1 or a group of formula 19 where t is 12.3 or 4, and R2 ³ and R2 ^3a , the same or different, are each independently hydrogen, lower alkyl, lower alkoxy (except tert-butoxy), halogen, phenoxy or benzyloxy, provided that when R2 ³ is hydrogen then R ^{2 3} ® must be hydrogen, no more than one of of R23 and R23a is trifluoromethyl, no more than one of R23 and R23® is phenoxy, and no more than one of R23 and R23® is benzyloxy.

157 761

Thus, the compounds of formula 1 include compounds of the formulas IB ¹ , IB ² , IB ³ and IB ⁴ .

The term "lower alkyl or" alkyl, alone or as part of another group as used herein, includes both straight and branched chain hydrocarbon chains containing up to 12 carbon atoms in the normal chain, preferably 1-7 carbon atoms, such as methyl, ethyl, propyl, isopropyl, butyl. , Tertiary-butyl, isobutyl, pentyl, hexyl, isohexyl, heptyl, 4,4-dimethylpentyl, octyl, 2,2,4-trimethylpentyl, nonyl, decyl, undecyl, dodecyl, their various branched chain isomers and similar hydrocarbon chains as well as the mentioned groups containing a halogen substituent, such as fluoro, bromo, chloro or iodo, trifluoromethyl, alkoxy, aryl, alkylaryl, haloaryl, cycloalkyl, alkylcycloalkyl, hydroxy, alkylamino, alkanoylamino, arylcarbonyl cyano, thiol and alkylthio.

The term "cycloalkyl, alone or as part of another group, as used herein includes saturated cyclic hydrocarbon groups having 3-12 carbon atoms, preferably 3-8 carbon atoms, such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, cyclodecyl and cyclododecyl. each of these groups may be substituted with 1 or 2 halogens, 1 or 2 lower alkyl groups, 1 or 2 lower alkoxy groups, 1 or 2 hydroxyl groups, 1 or 2 alkylamino groups, 1 or 2 arylcarbonylamino groups, 1 or 2 groups amino, 1 or 2 nitro groups, 1 or 2 cyano groups, 1 or 2 thiol groups and / or 1 or 2 alkylthio groups.

The term "aryl or" Ar as used herein refers to monocyclic or bicyclic aromatic groups having 6-10 carbon atoms in the ring portion, such as phenyl, naphthyl, substituted phenyl, or substituted naphthyl, in which groups the substituents of the phenyl or naphthyl group may be 1,2 or 3 lower alkyl groups, halogen atoms (Cl, Br or F), 1,2 or 3 lower alkoxy groups, 1, 2, or 3 hydroxyl 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 adamantylalkyl groups, 1 , 2 or 3 alkylamino groups, 1, 2 or 3 alkanoylamino groups, 1,2 or 3 arylcarbonylamino groups, 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 1, 2 or 3 alkylthio groups, where aryl groups having 3 substituents are preferred ki.

The term "aralkyl," aryl-alkyl, or "aryl-lower alkyl, alone or as part of another group, as used herein, refers to the lower alkyl groups discussed previously having an aryl substituent, such as benzyl.

The term "lower alkoxy," alkoxy, or "aralkoxy, alone or as part of another group, as used herein, means any of the previously described lower alkyl, allyl, aralkyl or aryl groups bonded to an oxygen atom.

The term "lower alkylthio," alkylthio, "arylthio, or" aralkylthio, alone or as part of another group, as used herein, includes any of the previously described lower alkyl, allyl, aralkyl or aryl groups linked to a sulfur atom.

The term "lower alkylamino group," alkylamino group, "arylamino group or" aralkylamino group, alone or as part of another group, as used herein, includes any of the above-described lower alkyl, allyl, aryl or aralkyl groups bonded to the nitrogen atom.

The term "alkanoyl as part of another group as used herein" means lower alkyl attached to a carbonyl group.

The term "halogen as used herein denotes chlorine, bromine, iodine and fluorine, with chlorine and fluorine being preferred. The term also includes trifluoromethyl.

Compounds of formula I of the structure represented by formula 70, in which R is OH, OLi or CH3O, are preferred, R ^x is hydrogen or lithium, X is a group of formula cisCH = CH-, and Z is a group of general formula 4, wherein R1 is phenyl or phenyl substituted with alkyl or halogen (including trifluoromethyl), or R ¹ is cycloalkylalkyl such as cyclohexylmethyl, or R 1 is a benzyloxy group substituted by halogen (including trifluoromethyl), and R 2 and R 2 ^and are the same and are hydrogen, halogen (including trifluoromethyl) or lower alkyl; or Z may preferably represent a group of general formula 71 in which R1 and R2 have the same meaning as previously defined for the group Z of formula 4, or Z may preferably be a group of general formula 72 in which R3 is

157 761 substituted phenyl, lower alkyl, cycloalkyl, or phenylalkyl, and R ⁴ is substituted phenyl, lower alkyl such as isopropyl, cycloalkyl, or phenylalkyl, or Z may also preferably represent a group of general formula 73 in which R ⁵ is hydrogen, methyl or a hydroxyl group, and R ⁶ is a group of general formula 74 wherein R 7 is hydrogen or methyl, or R 6 is substituted phenylmethyl.

Z can also preferably represent groups of formulas 7, 8 or 9 wherein at least one of R3 and R4 is phenyl or substituted phenyl and the remaining R3 or R4 is lower alkyl.

The inventive method for preparing of new phosphorus derivatives of 3-hydroxybutanoic acid of formula 1, wherein R is hydroxy or lower alkoxy, Z is a group of formulas 4, 5, 6, 15, in which R ', R ^2, R ^2a, and R ^2b are the same or different, and each independently is hydrogen, halogen, lower alkyl, haloalkyl, phenyl, substituted phenyl or OR ^y where R ^y is hydrogen, alkanoyl, benzoyl, phenyl, halophenyl, phenyl lower alkyl, lower alkyl, cinamyl, haloalkyl, allyl, cycloalkyl lower alkyl, adamantyl lower alkyl, or substituted phenyl lower alkyl, and when Z is a group of formula 15, R and R are the same or different and represent hydrogen, lower alkyl or OH group, R is a group of formula 17 or aryl-CH2, R6a is lower alkyl, hydroxy, keto or halogen, q is zero, 1,2 or 3, and R ⁷ is hydrogen or lower alkyl, in the case Z is the group of formula 6, j Eden of R ³ and R 4 is a group of the formula 18, and the other is lower alkyl or phenyl- (CHZ) p, wherein p is zero, 1, 2, 3 or 4, R 8 is hydrogen, alkyl of C * , C3-6cycloalkyl, Ci * alkoxy (except tert.-butoxy), trifluoromethyl, fluoro, chloro, phenoxy or benzyloxy, R ⁹ is hydrogen, Ci-3alkyl, Ci-3alkoxy, trifluoromethyl, fluoro, chloro , phenoxy or benzyloxy group, provided that if R ⁸ is hydrogen, R 9 must be hydrogen, no more than one of R ⁰ and R 9 is trifluoromethyl, no more than one of R ⁸ and R 9 is phenoxy and no more than one of R8 and R9 is benzyloxy or Z is the group of the formulas 7,8,9,10,1112,13,14, when Z is the group of the formula 7,8,9,10,11 and 13 one of the substituents R and R, R and R are as defined above, R is hydrogen, lower alkyl, lower alkoxy (except T-butoxy), halogen, phenoxy or benzyloxy R '* is hydrogen, lower alkyl, lower alkoxy, halogen, phenoxy or benzyloxy, R' * is hydrogen, lower alkyl, lower alkoxy or halogen, provided that if R ' ³ is hydrogen both R''* and R' * a must be hydrogen, if R '* is hydrogen, R' * a must be hydrogen, not more than one of R ' ³ and R' * is trifluoromethyl, not more than one of R ¹³ and R '* is phenoxy and not more than one of R' 3 and R '4 represents benzyloxy, R ¹⁰ and R''are each independently hydrogen, alkyl, cycloalkyl, adamantyl-l or a group of the formula 69, where R ' ³ , R' * and R '* a are as defined above, and q is zero, 1, 2, 3 or *, Y is oxygen, sulfur or NR ¹⁰ , in case Z is a group of formula 12, Ra is a hydrogen atom or a primary or secondary C1-6alkyl group, Rb is a primary or secondary C1-6alkyl group or Ra + Rb is a - group (CHzjr or cis-CH2CH = CH-CH 2, r is 2, 3 ', 5, or 6, R' 2 represents lower alkyl, cycloalkyl, or R 18 and R 8, R ^9, R ", R 'and R' are as above and * the meaning given, when Z is a group of formula 13, R ' ⁵ and R ¹ 6 both represent hydrogen, chlorine, bromine, CN, CF3, phenyl, C1- * alkyl, C2- ealkoxycarbonyl, -CH2OR' -CH2OCONHR ⁷ or ¹ 8, R '7 is hydrogen or Ci_6alkyl, R' ⁸ is alkyl or phenyl optionally substituted by fluorine, chlorine, bromine or a Ci * alkyl, or R ^'5 and R 6 together form a ¹ the group - (CH2) s-, -CH2-O-CH2-, -CON (R ' ⁹ ) CO- or -CON (R20) N (R2') CO-, s is 3 or *, R is a hydrogen atom, Ci-6 alkyl, phenyl, or benzyl, R² is hydrogen, C * alkyl or benzyl and R ³ and R are as defined above, where Z is a group of the formula M ² R 2 is lower alkyl, cycloalkyl , Adamtyk-1 or a group of Formula 19 wherein t is 1, 2, 3 or *, R2 ³ and R2 ³ a are the same or different, and each independently represents hydrogen, lower alkyl, n lower alkoxy (except IH ^ i ^^. ^ bl ^ t ^ o ^^^ ylu), halogen, phenoxy or benzyloxy, except that if R2 ³ is hydrogen, R2 ³ a must be hydrogen, not more than one of R23 and R2 ³ i is trifluoromethyl, not more than one of R2 ³ and R23a is phenoxy and not more than one of R2 ³ and R23a is benzyloxy, R ^x

157 761 is hydrogen or a lower alkyl group in the form of the free acid or in the form of a physiologically acceptable and hydrolysable ester in the form of a salt, and the acetylene phosphate salt of formula II is selectively reduced to give the silyl ether of formula 3 (cis), which cleaves the silyl ether group to give the diester of formula 1β1 (cis) and if necessary converted to the monoester of formula IB3 (cis), basic salt of formula IB ² (cis) acid of formula IB (cis), or dibasic a salt of formula IB ⁴ (cis).

The compounds of the present invention can be prepared as raceramic mixtures and then separated to isolate the S isomer, which is preferred. However, these compounds can also be prepared directly as S-isomers as described in the description and the examples.

The compounds of formula I are inhibitors of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-Co A) reductase and are therefore useful for inhibiting cholesterol biosynthesis as demonstrated in the following tests.

1. Rat hepatic reductase HMG-Co A.

Rat hepatic HMG-Co A reductase activity was measured using a modified method described by Edwards (Edwards, PA et al., J. Lipid Res. 20:40, 1979). Rat liver microsomes were used as the enzyme source and enzyme activity was determined by measuring the conversion of ™ C-HNC-Co A medium to ^M C-mevalonic acid.

a) Preparation of microsomes. From 2-4 Sprague Dawley rats fed cholestyramine, livers were removed after decapitation and homogenized in phosphate buffer A (potassium phosphate, 0.04 m, pH 7.2; DCI 0.05 m; sucrose, 0.1 m; EDTA 0 0.03 m; aprotinin, 500 KJ units / ml). The homogenization product was centrifuged at 16,000.g. for 15 minutes at 4 ° C. The supernatant was removed and centrifuged again under the same conditions. The supernatant from the second centrifugation was centrifuged for 70 minutes at 4 ° C at 10,000 g. The microsome tablet was resuspended in the minimum volume of buffer A (3-5 ml per liver) and homogenized in a glass / glass homogenizer. Dithiothreitol (10 mmol) was added and the formulation was divided into small aliquots which were quickly frozen in acetone / dry ice and stored at -80 ° C. The specific activity of the first microsomal preparation was 0.68 mmoles of mevalonic acid (mg / protein / minute).

b) Analytical determination of the enzyme. Reductase was determined analytically in 0.25 ml samples, containing the following components at the indicated final concentration: 0.04 m potassium phosphate, pH 7.0, 0.05 m KC1, 0.10 m sucrose, 0.03 m EDTA, 0 , 01 m dithiothreitol, 3.5 mmol NaCl, 1% dimethylsulfoxide, 50-200pg microsomal protein, 100pg ™ C- [DL] HMG-Co A (0.05 µCi, 30-60 mCi (mmol), 2, 7 mmoles of NADPH (nicotinamide adenine dinucleotide phosphate).

Reaction mixtures incubated at 37 ° C. Under the conditions described, the enzyme activity increased linearly to 300 pg of microsomal protein per reaction mixture and remains linear with the incubation time up to 30 minutes. The standard incubation time selected for drug testing is 20 minutes, with 12-15% conversion of the HMG-Co A medium to mevalonic acid. [D, L] HMG-Co A medium with a concentration of 100 [mu] m was used, twice the concentration required to saturate the enzyme under the conditions described. NADPH was used in an excess of 2.7 times the concentration needed to achieve the maximum rate of enzyme activity.

Standard assays for evaluating inhibitors were performed according to the following procedure. The microsomal enzyme was incubated in the presence of NADPH for 15 minutes at 37 ° C. DMSO with or without test compound was added as liquid carrier, and the mixture was incubated further at 37 ° C for 15 minutes. The enzyme sample was activated by adding 1 ⁴ C-HMG-Co A medium. After 20 min incubation at 37 ° C, the reaction was stopped with 28 µl of 33% KOH. ³ H-mevalonic acid (0.05 µCi) was added and the reaction mixture was allowed to stand for 30 minutes at room temperature. 50 µl of 5 N HCl was added to lactonize the mevalonic acid. Bromophenyl blue was added as a pH indicator to track the corresponding decrease in pH. Lactonization was allowed to proceed at room temperature for 30 minutes. The reaction mixture was centrifuged for 15 minutes at

157,761 speeds of 2,800 rpm. Supernatants were layered on 2 g of AG 1-8 anion exchange resin (Biorad, formatted form), poured into glass columns 0.7 cm in diameter and eluted with 2.0 ml H2O. The first 0.5 ml was discarded and the next 1.5 ml were collected and the content of both tritium and ¹⁴ C in 10 ml of Optifluor scintillation liquid was counted. The results were calculated as the amount of mmoles of mevalonic acid produced in 20 minutes and corrected for 100% tritium recovery. Drug efficacy was expressed as value (50) drug concentration inducing 50% inhibition of enzyme activity obtained from given compound dose responses with the indicated 95% confidence interval.

The conversion of the lactone drugs to their sodium salts was achieved by dissolving the lactone in DMSO, adding a 10 fold molar excess of NaOH, and allowing the mixture to stand for 15 minutes at room temperature. The mixture was then neutralized (pH 7.5-8.0) with IN HCl and diluted to obtain an enzyme reaction mixture.

2. Cholesterol synthesis in freshly isolated rat hepatosites.

Compounds showing activity as HMG-Co A reductase inhibitors were evaluated for their ability to inhibit the incorporation of the ^M C-acetate group into cholesterol in a suspension of freshly isolated rat hepatocytes using the method originally described in Capuzzi DM and Margolis, S., Lipids, 6 : 602,1971).

a. Isolation of rat hepatocytes.

Spraue Dawley rats (180-220 g) were anesthetized with Nebutol (50 mg / kg), and their abdominal cavities were opened and portal veins clamped. Heparin (100-200 units) was injected directly into the main vein of the abdominal cavity. A single clamping suture was placed on the distal segment of the portal vein and the portal vein was catheterized between the clamping thread and the first branching vein. The liver was perfused at 20 ml / min, previously warmed to 37 ° C with oxygenated buffer A (HBSS without calcium or magnesium, containing 0.5 mmol EDTA) after disconnecting the vena cava to allow drainage to drain. The liver was additionally perfused with 200 ml of pre-warmed buffer B (HBSS containing 0.05% bacterial colbassenase). After perfusion with buffer B, the liver was excised and degreased in 60 ml of Weymputh medium, allowing free cells to diffuse freely in the medium. Hepatocytes were isolated by slow centrifugation for 3 minutes at room temperature under 50 g overload. The hepatocyte tablet was washed once with Weymouth's medium, scored, their viability was analyzed by counting the trypon blue. Such enriched hepatocyte cell suspensions routinely show 70-90% viability.

b. Incorporation of a group 1 ⁴ C-acetate into chloroestrolu.

Hepatocytes were resuspended in medium (IM) at a concentration of 5 × 10 ⁶ cells per 2 ml of medium [0.02 m Tris-HCl (pH 7.4), 0.1 n KCl, 3.3 mmol sodium citrate, 6 , 7 mmol nicotinamide, 0.23 mmol NADP, 1.7 mmol glycone-6 phosphate].

Test compounds were routinely dissolved in DMSO or in a mixture of DMSO: HiO (1: 3) and added to the IM. Final DMSO concentration in IM was = S11.0% and had no significant effect on cholesterol synthesis.

Incubation was initiated by adding 13 C-acetate (58 m Ci) -mmol, 2 µCi (ml) and placing the cell suspensions (2.0 ml) for 2 hours at 37 ° C in 35 mm diameter tissue culture vessels. After incubation, the cell suspensions were transferred to glass centrifuge tubes and centrifuged at room temperature for 3 minutes at 50 g. Cell pellets were resuspended and lysed in 1.0 ml H 2 O and then placed in an ice bath.

Lipids were extracted essentially as described in Bligh, H. C. and W. J. Dyer. Con. J. Biochem. and Physial., 37: 911, 1959. The lower organic phase was removed and dried under a stream of nitrogen and the residue was resuspended in 100 µl of chloroform: methanol (2: 1). The entire sample was spotted onto silica gel plates (LKGD) for thin layer chromatography and the chromatogram was developed with hexane: ethyl ether: acetic acid (75: 25: 1). Plates were examined and counted using a BioSca automated scanning system. The level of radioactively significant groups in the cholesterol peak (Rf 0.28) was determined and expressed as the total number of counts per peak and as the percentage of groups labeled in the total extract

157 761 lipid. The cholesterol peaks in the control cultures routinely show 800-1000 cpm and contain 9-20% labeled groups present in total lipid extract; the results were obtained by the method

Capuzzi and others show 9% extracted groups significant in cholesterol.

The action of drugs (inhibition of cholesterol synthesis) was determined by comparing the% content of cholesterol labeled groups in control and drug treated cultures. Dose-response curves were plotted from two or more compound dose trials and results were expressed as values (50 with 95% confidence intervals).

3. Cholesterol synthesis in human skin fibroblasts.

The selectivity of a compound favoring greater inhibitory activity in hepatic tissue would be a hallmark of a cholesterol synthesis inhibitor. Therefore, in addition to evaluating inhibitors in hepatocytes, these compounds were also tested for their activity as inhibitors of cholesterol synthesis in cultured fibroblasts.

a. Cultures of human skin fibroblasts.

Human skin fibroblasts (passage 7-27) were cultured in Eagle's Minimal Basic Medium (EM) containing 10% fetal calf serum. Stock cultures for each assay were trypsonized to disperse the cell monolayers, counted, and plated in tissue culture wells 35 mm in diameter (5 105 cells / 2.0 ml). Cultures were incubated for 18 hours at 37 ° C in 5% CO2 air and 95% humidity. Cholesterol biosynthesis enzymes were induced by removing serum containing medium, washing the cell monolayers, and adding 1.0 ml of EM containing 1% fatty acid free bovine serum albumin, and incubating the cultures for an additional 24 hours.

b. Incorporation of ¹⁴ C-acetate groups into cholestreol.

Fibroblast cultures were induced and washed with EMEMwo (Earle's Minimal Basic Medium). Test compounds were dissolved in DMSO or in a DMS: EM (3: 1) mixture (final DMSO concentration in the culture cells> 1.0%), added to the cultures and the cultures pre-incubated at 37 ° C for 30 minutes in room atmosphere. 5% CO2 and 95% humidity. After pre-incubation with the drug, [1-4 CJNa acetate (2.0pCi / ml, 58mCi / mmol) was added and the cultures were re-incubated for 4 hours. After incubation, the culture medium was removed and the cell monolayer (200 µg cell protein per culture) was scraped into 1.0 ml H 2 O. The lipids contained in the lysed cell suspension were extracted with chloroform: methanol as described for hepatocyte suspensions. The organic phase was dried under nitrogen and the residue was resuspended in chloroformrmethanol (2: 1) (100 µl) and the entire sample was spotted onto silica gel (LKOD) plates for thin layer chromatography and then analyzed as described for hepatocytes.

The inhibition of cholesterol synthesis was determined by comparing the content of radioactively significant groups in the cholesterol peak with the control and drug-treated cultures. Results are expressed as values (50, and are derived from a complex dose-response curve for two or more trials. A 95% confidence interval was also calculated from these curves).

The compounds of the invention can be used in the form of pharmacological compositions consisting of at least one compound of formula I in association with a pharmacologically acceptable carrier or diluent. These compositions may be formulated using known solid or liquid pharmacological carriers or additives appropriate to the desired mode of administration. They can be administered orally, e.g. in the form of tablets, capsules, granules or powders, or by parenteral route in the form of injectable preparations, dosage forms containing 1-2000 mg of active ingredient per dose being used for treatment. The administered dose depends on the unit dose, symptoms, age and weight of the patient.

The compounds of formula I can be administered in a similar manner to known compounds suggested for use in inhibiting cholesterol biosynthesis such as levastatin, to mammals such as dogs, cats and the like, and to humans. Thus, compounds of formula 1 may be administered in an amount of from about 4-2000 mg in a single dose or in the form of individual doses 1-4 times daily, preferably 4-200 mg in partial doses of 1-100 mg, suitably in unit doses. , 0.5-50 mg 2-4 times a day, or in the form of sustained release of the active ingredient.

157 761

The following examples illustrate preferred embodiments of the process of the invention. Unless otherwise indicated, all temperatures are expressed in ° C. Flash chromatography was performed on either Merck 60 or Whatmann LPS-1 silica gel. Reverse-phase chromatography was performed on a gel of CMP-20 MCI resin, manufactured by Mitsubisc hi, Ltd.

The abbreviations "Et2O", "EtOAc", "MeOH", and "EtOH" are used in the examples for ethyl ether, ethyl acetate, methanol and ethanol, respectively.

Example IA (S) - · 4-f [2- [4'-nuoro-3,3 ', 5-trimethyl] o (1,1'-biphenyl-2-yl] ethenyl] -methoxyphosphinyl] -3 acid methyl ester -tert-butyldiphenylsiiioxybutane.

To a solution cooled to -78 ° C (in a dry ice and acetone bath) solution of 2.678 g (11.2 mmol) of 2-ethynyl-4'-fluoro-3,3 ', 5-trimethyl'] ol, 1-biphenyl 7ml (11.2mmol, 1.0eq) of a 1.6M solution of n-butyllithium in hexane was added dropwise. The purple mixture was stirred at -78 ° C under argon for 1 hour, quickly warmed to 0 ° C, re-cooled to -78 ° C, transferred via a catheter to a dropping funnel and added dropwise to cooled dry ice bath and acetone to -78 ° C of a solution of 8.27 g (18.4 mmol, 1.6 eq) phosphonochloridate in 20 mL of dry THF. After 1 hour at -78 ° C, the reaction was quenched with saturated NEUCi solution, then the mixture was allowed to warm to room temperature and extracted with diethyl ether. The ether layer was washed with saturated NaHCOe and brine, dried over anhydrous MgSO4 and evaporated to give 11.70 g of a brown oil. The crude oil was purified by flash chromatography on silica gel, eluting with 7: 3 hexane-ethyl acetate. Product containing fractions were combined and evaporated to yield 4.246 g (-6%) of the desired title acetylenic phosphinate as a light brown oil. An additional 4-7 mg of bufenylacetylene was isolated from part E such that the corrected yield was 68%. TLC (7: 3) hexane-ethyl acetate, Rf = 0.20, UV and PMA.

B. (S) - · 4- [2- [4'-PyroΓO-3,3'-5-trimethyl] o (l, Γ-biphenyl-2-yl-thie] yIo] methoxyphosphinyl] -3- acid methyl ester tflt-butyIodiphene IosiliIoxybutane.

To an argon purged solution of 333 mg of acetylene phosphinate from Part A in - ml of CH3OH was added 121 mg (36 wt%) of a 10% Pd / C and the mixture was shaken on a Parr apparatus under 0.28MPa hydrogen pressure for 30 hours. The catalyst was removed by filtration through a celite packet and the filtrate was evaporated to give a pale yellow oil. The crude oil was purified by flash chromatography on silica gel, eluting with 1: 1 ethyl acetate-hexane. Product containing fractions were evaporated to yield 2-0 mg (7-%) of the title saturated phosphinate as a clear oil. TLC (4: 1) ethyl acetate-hexane, Rf = 4 O 3 4 3, UV and PMA.

C. (S) - · 4 [[2- [4'-yluol-3,3 ', - trimphthylol 1,1-biffnyl) -2-yl] phthyl] -methoxyphosphinyl] -3-hydroxybutylic acid methyl ester .

To a solution of 330 mg (0.489 mmol) of Part B silylester in 6 mL of dry THF was added 11-μϊ (1.96 mmol, 4.0 eq.) Of glacial acetic acid followed by 1.47 mL (1.47 mmol, 3 0 eq.) Of a 1.0 M tetrabutylammonium fluoride solution in THF, then the resulting mixture was stirred overnight at room temperature under argon. The mixture was diluted with 10 mL of ice water, then extracted twice with ethyl acetate. The organic phase was washed with saturated NaHCOe and brine, dried over anhydrous Na2SO4 and evaporated to give 364 mg of a pale yellow oil. The crude oil was purified by flash chromatography on silica gel, eluting with -: 4 acetone-hexane. Product containing fractions were evaporated to afford 20 mg (96%) of the desired title free alcohol as a clear oil which slowly crystallized on standing. TLC (7: 3) acetone-hexane, Rf = 0.28, UV and PMA.

Example II A. 2) - 4 - [[2- [3- (4-Ωυο-nyl) -1- (1-methyl-ethyl) -1Hmdol-2-yl] phthimyl] -methoxyphosphinyl] -3 acid methyl ester -tfrt-butyIodiphenylsillloxy) butane.

For a solution cooled in a mixture of dry ice and acetone to -78 ° C, a solution of 678 mg (244 mmol, 1.0 eq.) 3- (4-pyrophenes] o)) 1- (1-mphthylethyl) -2-ethine] o- h lu ^{indole in 6 mL of} su ^c h g ^{e of THF} wa ^ ar ^g osferze one wlcro it ^{en 1} -3 ml ^(2, 44 ^{mmol, 1,} 0 ^y yellow ^equivalence disappears) of a 1.6 molar solution of n-butyllithium in hexanes. After 30 minutes at -78 ° C, the mixture was transferred via a catheter to a cooled -78 ° C solution.

157 761 about 4.3 mmoles (1.75 eq.) Of phosphonochloridate of methyl ester of (S3 -3 - [[(1,1-dimethyl / diphenylsilyloxy] -4- (chloromethoxyphosphinyl / b) tanoic acid in 5 mL of dry THF). color was stirred at -78 ° C for -9 minutes, then quenched by dropwise addition of 5 ml of saturated NH4Cl solution and the mixture was allowed to warm to room temperature.The mixture was extracted twice with diethyl ether, washed with saturated NH4Cl solution and brine, dried over anhydrous MgSO4 and evaporated in vacuo to give 2.567 g of a reddish brown oil The crude product was purified by flash chromatography on silica gel eluting with 3: 2 hexane-ethyl acetate to give 756 mg (44%) of the desired title acetylene phosphinate as a dark oil. yellow solution. TLC (7: 3) hexane - acetone, Rf = 0.27, UV and PMA ¹ H NMR (CDCl): δ (ppm) 1.0 (9H, s), 1.64 (6H, d) , 2.10 - 2.90 (4H, m), 3.56 (3H, s), 4.6 (IB, bm), 4.90 (1H, m), 7.05-7.55 (18H, m).

13C NMR (CDCU): δ (ppm) 14.2, 19.1, 21.0, 26.7, 27.8, 37.5, 39.2,42,2,45,1,49.2, 51.4, 51.9, 60.3.65.5 (Jcc-p = 15.1Hz), 88.1,91.2, 98.3, 111.3, 115.3, 115.6, 120.8 (J = 5.7Hz), 122.3, 124.9, 125.9, 126.4, 127.6, 129.2, 130.7, 133.0, 135.7, 136.1 , 170.9.

B. Methyl (S) -4 [[2--3 - (^ _Or flu rfenylo0l - l [methylethyl) -lH [indol-2-yl] [ethyI] metoksyfosflnyIo] [3- (tert-butylodifenylosillloksy ) butane gas.

To an argon purged solution of 422 mg of Part A acetylene phosphinate in 9 mL of methanol was added 420 mg of a 10% solution of Pt / C and the resulting mixture was shaken in a Parr apparatus for 2 hours under 0.28 MPa hydrogen pressure. The catalyst was removed by filtering the mixture through celite and the filtrate was evaporated to yield 380 mg (90%) of the title indolophosphinate as a yellow colored foam. TLC (4: 1) ethyl acetate-hexane, Rf = 0.27, UV and PMA

1 H NMR (CDCU): δ (ppm) 1.00 (9H, s), 1.63 (6H, d), 1.5-2.0 (2H, m), 2.20 (1H, m), 2.58-3.00 (5H, m), 3.44 (3H, d, Jh-p = 10.6Hz), 3.61 (3H, s), 4.52 (2H, m), 7 . 07-7.66 (18H, m).

ⁿ C NMR (CDCU): δ (ppm) 12.6, 16.8, 17.2, 19.1, 21.5, 26.7, 36.0.42.1.47.2, 50.9 , 51.4.65.8, 111.8, 115.3, 119.1, 121.1, 121.7, 128.3, 129.9, 131.2, 132.8, 133.4, 134 , 3, 134.8, 135.7, 171.3.

Example 3 A. N - 4 - (2 - (phosphinyl] [3- (tert-butyldiphenylsilyloxy] butanoic acid) methyl ester.

0.332 g (1.86 mmol) of 2-ethylen (-1,1'-biphenyl) was stirred at -78 ° C in 10 ml of THF under argon. 0.75 mL of a 2.5 M n-butyl lithium solution in hexane was added to the solution of the acetylene compound over 5 minutes. The reaction mixture was stirred at -78 ° C for 1 hour, warmed to room temperature, stirred for 1 hour, warmed to room temperature, stirred for 10 minutes, and re-cooled to -78 ° C. The solution of the acetylene anion was then added dropwise over 8 minutes to a solution of 2.98 mmol of phosphonochloridate as in Example 2A, in 10 ml of THF, cooled to -78 ° C under argon. After completion of the dropwise addition, the reaction mixture was stirred at -78 ° C for 1 hour, then quenched with saturated aqueous NH4Cl solution. The mixture was warmed to room temperature, diluted with half saturated aq. NaCl and extracted three times with diethyl ether, the combined ethereal extracts were washed with saturated aq. NaHCO3 and saturated aq. NaCl. The ether layer was dried over MgSO4 and evaporated to give 1.5 g of a yellow oil. Purification by flash chromatography eluting with 5: 1: 4 hexane-toluene-ethyl acetate provided 0.543 g (48%) of the title acetylene phosphinate. TLC (5: 1: 4 hexane - toluene - ethyl acetate, silica gel) Rf = RHF2O.

IR (CHC13) 3070, 3053, 3035, 3000, 2952, 2934, 2896, 2859, 2178, 1735, 1474, 1448, 1436, ¹⁴²⁹ cm ¹

1 H NMR (270 MHz, CDCU): δ 7.65 (m, 3H), 7.65-7.28 (m, 16H), 4.55 (m, 1H), 3.55 (d, 3H), 3.40 (dd, 3H), 2.80 (m, 1H), 2.55 (m, 1H), 2.35 (m, 1H), 2.08 (m, 1H), 1.00 (s , 9H).

13C NMR (67.8 MHz, CDCU) δ 170.83,145.29,145.19,139.22.135.95,135.59,133.86,133.75, 133.16,132.86,130.57,129.56,129.34,128.81 127.92,127.75,127,44,127, 39,126.94,117.90,100.91, 100.38, 100.18, 84.51, 81.60, 65.53, 65.53, 65.42, 60.06, 51.61, 51.50, 51, 11, 42.07, 41.90, 38.86, 37.16, 26.56, 20.75, 18.97, 13.97.

Mass spectrum, m / e 611 (M + + H).

B. (S) -4 - [[2 - ((1, 1'-b-phenyl] -yl-ylmethyl-methoxyphosphinyl-tert-butyldiphenylsilyloxy) butanoic acid methyl ester.

Argon was bubbled through a solution of the acetylene phosphinate from Part A, 0.515 g (0.85 mmol) in 8 ml of methanol for 10 minutes. After adding 0.190 g of 10% Pd / c to the acetylene compound solution was hydrogenated in a Parr apparatus at a pressure of 0.30 MPa. After shaking for 25 minutes at 0.30 MPa, the methanol solution was filtered through celite and the filtrate was evaporated to give 0.510 g (98%) of the title phosphinate as a colorless oil. TLC (4: 1 ethyl acetate-hexane) Rf = 0.21. TLC (4: 1 ethyl acetate-hexane) Rf = 0.21.

IR (CHC13) 3071,3054,2998,2954,2934,2902,2859,1734,1477,1462,1448,1438,1428 cm-1 ¹ H NMR (270 MHz, CDC1 _3): δ 7.65 (m, 3H), 7.55-7.00 (m, 16H), 4.45 (m, 1H), 3.58 (s, 3H), 3.30-3.20 (dd, 3H, J = 11Hz ), 2.88 (m, 1H), 2.60 (m, 3H), 2.17-1. 80 (m, 1H), 1.8O-1.30 (m, 1H), 1.00 ( s, 3H).

1 ³ C NMR (67.8 MHz, CDCl, identifying peaks) δ 171.33, 65.78, 51.36, 42.24, 26.75.

Mass spectrum, m / e 615 (M + + H).

c. (S) -4- [2 - [[1,1'bbifei ^; yI] - ^ - illo] etjllo] n ^ et (^ l ^^ iff (^: ^ fii ^) lk) acid methyl ester ] - ^: ^ - ^ ł ^> ^ c ^ r (^ lss; -butane.

A solution of 0.500 g (0.82 mmol) of part B phosphinate in 10 ml of THF was stirred under nitrogen with 0.19 ml (3.3 mmol) of acetic acid. 2.45 ml of a fO-molar solution of tetra (n-butyl) ammoniumquegk fluorine in THF was added dropwise at room temperature. The reaction mixture was stirred at room temperature for 23 hours, then quenched with 15 ml of ice water. The aqueous phase was extracted three times with ethyl acetate. The combined organic solutions were washed twice with saturated aqueous NaHCO3 solution and once with saturated aqueous NaCl solution. The organic phase was dried over Na2SO4 and evaporated to yield 0.437 g of a colorless oil. Purification by flash chromatography eluting with 7: 3 acetone-hexane provided 0.247 g (81%) of the title alcohol as a colorless oil. TLC (7: 3 acetone-hexane, silica gel) Rf = 0.22.

IR (CHC13) 3600-3171 (wide), 3064, 3009, 2954, 1731, 1479, 1439, 1237, 1180, 1042, 999 cm ^-1 1H NMR (270 MHz, CDCb): δ 7.50-7, 10 (m, 9H), 4.50-5.15 (m, 1H), 3.70 (s, 3H), 3.53 and 3.50 (dd, 3H, J = 11Hz), 2.88 (m, 2H), 2.50 (m, 2H), 2.00-1.60 (m, 4H).

1 ³ C NMR (67.8 MHz, CDCb): δ 171.49,141.39,141.00,138.10,137.88,129.95,128.81, 128.06, 127.53, 126.83, 126.22, 63.08 , 63.08, 63.02, 62.85, 51.39, 50.58, 50.47, 42.35, 42.15, 42.07, 41.87, 34.31, 33.06, 33 .00, 30.77, 30.52, 29.49, 29.21, 25.41.

Band spectrum, m / e 377 (M + + H).

Example IV A. (S) -4 -] [2-] 1-] 4-phylophenite -] -] 1-methylethyl] -lH-indol] 2] yl] ethynylI-methoxyphosphinyl] -3 -] [(1,1, ] dimethylsilyl) diphenylsilyl] kxy] butane.

Phosphonochloridate was prepared from the dicyclic salt of methyl ester of (S) -4 (hydroxymethoxyphosphinyl) -3 - [[(1,1-dimethylketyl) diphenylxylkxyIbutanoic acid according to the following method: Free acid was isolated from 4.33 mmol, 75 equivalents) by dissolving 1.0 N HCl and ethyl acetate in a mixture, washing the organic phase twice with 1.0 N HCl and brine, then drying over anhydrous Na2SO4 and evaporating under reduced pressure. 6.8 mmol of free acid are obtained in the form of a clear, viscous oil. 1.72 ml (13.7 mmol, 2 eq.) Of distilled trimethylsilyl] diethylamine were added to 6.8 mmol of monophosphonic acid methyl ester in 10 ml of dry CH2Cl2 and the clear solution was stirred under argon at room temperature for 1 hour. The mixture was evaporated under reduced pressure, azeotroped twice with 20 ml of dry benzene and dried for 15 minutes using a vacuum pump. The crude silylated acid in 10 mL of dry CH 2 Cl 2 and 1 drop of dry DMF was cooled in an ice bath to 0 ° C and 655 µl (7.5 mmol, 1.1 eq) distilled COCl 3 was added dropwise thereto over 5 minutes. The yellow mixture was stirred at 0 ° C for 15 minutes and then for 45 minutes at room temperature under argon. The mixture was evaporated under reduced pressure, azeotroped twice with 20 ml of benzene and dried for 15 minutes with a vacuum pump, yielding crude phosphonochloridate as a viscous yellow oil.

To a solution of 1.084 g (3.90 mmol, 1 eq.) Of 2-ethynyl-1- (4-fluorphenyl) -3-1-methylethyl) -1H-indole cooled in a mixture of acetone with dry ice to -78 ° C 10 mL of dry THF was added dropwise over 10 minutes 2.44 mL (3.9 mmol, 1 eq.) 1.6 molar

A solution of n-butyllithium in hexanes, the purple suspension was stirred under argon at -78 ° C for 30 minutes. The anion solution was added dropwise via a catheter over 30 minutes at -78 ° C to a cooled to -78 ° C solution of phosphonochloridate in dry THF. The dark brown mixture was stirred at -78 ° C for 30 minutes, then quenched by dropwise addition of 10 mL of saturated NH 4 Cl. The mixture was warmed to room temperature and dissolved in a mixture of ethyl acetate and saturated NH 4 Cl, washed with brine, dried over anhydrous Na 2 SO 4 and evaporated to give 1.968 g (71.7%) of the title acetylene phosphinate as a pale yellow oil. TLC (7: 3 hexane-acetone) Rf = 0.25, UV and PMA.

Mass Spectrum, M + H + = 710+ observed

B. (S) -3 - [[1,1-dimethylethyl) diphenylsilyl] oxy] -4- [2- [1- (4-fluorophenyl) -3-1-methylethyl) -1H-indole- acid methyl ester 2-yl] ethyl] methoxyphosphinyl] butane.

To an argon purged solution of 950 mg of acetylene from Part A and 10 ml of methanol was added 238 mg (25% by weight) of 10%. Pt / C and the resulting black suspension were stirred overnight under a hydrogen atmosphere at 0.1 MPa. The catalyst was removed by filtering the suspension through a 0.4 µm Millipore polycarbonate filter and a prefilter, and the filtrate was evaporated in vacuo to yield a yellow oil. The crude oil was purified by flash chromatography on Merck silica gel eluting with 8: 2 hexane-ethyl acetate. Product fractions were evaporated to yield 915 mg (86.7%) of pure title saturated phosphinate as a white foam. TLC (4: 1 ethyl acetate-hexane) Rf = 0.39, UV and PMA.

Mass Spectrum: M = H + = 714+ observed.

C. 1S) -4 - [[2- [1- (4-Fuoro-phenyl) -3- (1-methylethyl) -1H-incol-2-yl] ethyl] methoxyphosphinyl] -3-hydroxybutanoic acid methyl ester.

To a solution of 915 g (1.22 mmol) of Part B silyl ether in 10 ml of THF was added successively 280 µl (4.88 mmol, 4 eq.) Of glacial acetic acid and 3.3 ml (3.66 mmol, 3 eq.) A 1.1 molar solution of n-C4HgNF in THF, then the mixture was stirred under argon at room temperature overnight. 8 ml of ice-cold water were added, the mixture was extracted with ethyl acetate and the organic phase was washed twice with 5%. KHSO4, saturated NaHCO3, and brine, dried over anhydrous NazSO4, and evaporated under reduced pressure to give 955 mg of a yellow oil. The crude oil was purified by flash chromatography on Merck silica gel eluting with 1: 1 hexane-acetone. Product fractions were evaporated to afford 521 mg (85.5%) of the desired title alcohol as a pale yellow oil. TLC (3: 2 acetone-hexane) Rf = 0.21, UV and PMA.

EXAMPLE 5 (S) -4 - [[[[2,4- ^ dimethyl-6- [i ^ ^ ^ ^ and ^ orophenyl) methoxy] phenyl] ethyl] hydroxyphosphinyl] -3-hydroxybutanoic acid salt.

A. (S) -3 - [[(1,1-dimethylethyl) diphenylsilyl] oxy] -4 - [[2 - [(4-fluorophenyl) methoxy] -4,6-dimethylphenyl] ethyl] methoxyphosphinyl] acid methyl ester butane gas.

1.34 g (1.95 mmol) of the dipithium salt of (S) -4 - [[[[^, '4 ^ and ^ ^ methyl-6 - [[(4 ^ ^ ^ ^ uorophenyl) methoxy] phenyl] -ethynyl] hydroxyphosphinyl acid ] -3- (tert-butyl-diphenylsilyloxy) butane was stirred in 12 ml of methanol, then 0.040 g of PtO2 was added. Hydrogen was bubbled through the methanol solution for 10 minutes, then maintaining a hydrogen atmosphere with a balloon. After 5 hours and 15 minutes at room temperature, the reaction was complete and argon was bubbled through the reaction solution. The mixture was filtered through a pad of celite on a fine sintered funnel, and the catalyst was washed with methanol. The solvent was removed from the filtrate to give 1.4 g of the title saturated phosphinate as a clear gum. The title saturated phosphinate was purified by flash chromatography eluting with 60%. ethyl acetate in hexane followed by rechromatography to remove contaminating fractions eluting with 6: 2.5: 1.5 hexane / acetone / toluene. 1.17 g (86% yield) of the title saturated phosphinate were obtained.

TLC: Rf = 0.045 (80% ethyl acetate in hexane, silica gel). Mass spectrum: m / e 691 (M + H) +, 659 (M-OCH 3) +, 635 (M-C 8 HigOSi /.

B. 1S) -4 - [[2,4-dimethyl-6 - [(4-fluoro-phenyl) methoxy] phenyl] ethyl] methoxyphosphinyl] -3-hydroxybutanoic acid, methyl ester.

1.16 g (1.68 mmol) of part A phosphinate was mixed with 25 ml of THF under argon at room temperature. 0.40 ml of glacial acetic acid was added dropwise to the phosphinate solution, then 4.6 ml of a 1.1 molar solution of n-C4HgNF in THF was added dropwise over 5 minutes. The reaction mixture was stirred at room temperature overnight (18 hours), then quenched with 50 mL of ice water. After stirring for several minutes, saturated NaCl solution was added and the layers were separated. The organic layer was evaporated on a rotary evaporator in ethyl acetate. The aqueous layer was extracted twice with ethyl acetate, then all ethyl acetate solutions were combined, washed twice with saturated aqueous NaHCO3 solution and once with saturated HCl solution, then dried over NazSCb. After filtration and removal of the solvent, 1.13 g of the title hydroxyphosphinate was obtained as a clear oil. The title hydroxyphosphinate was purified by flash chromatography eluting with 100%. ethyl acetate to afford the title hydroxyphosphinate compound in 83% yield. TLC: Rf = 0.27 (6: 4 acetone / hexane, silica gel). Mass spectrum: m / e 453 (M + H) +, 343 (M-CtH6F) ⁺ .

Example VI A. (S) -3 - [[1,1-dimethylethyl) diphenylsilyl] oxy] -4 [[2 - [[3,5-dimethyl [1,1'-3-phenyl] -2-yl) acid methyl ester ] methoxyphosphinyl] 3u tane.

A solution of 0.950 g (4.61 mmol) of 3,5-dimethyl-2 -1-propynyl) (Ι, Γ-biphenyl) in 27.3 ml of dry THF was stirred under argon and cooled to -78 ° C. 1.84 mL (4.61 mmol) of a 2.5 M n-butyllithium solution in hexanes was added dropwise over 20 minutes to give a dark purple-brown solution. The reaction mixture was stirred at -78 ° C for 1 hour, then became a slurry, then warmed to 0 ° C and stirred for 15 minutes, returning the mixture to a homogeneous dark purple solution again, and then re-cooled to -40 ° C except that the mixture remained homogeneous. This acetylene anion solution was added dropwise at -40 ° C over 25 minutes to a solution of 8.12 mmoles of phosphinylchlorodane as in Example 2 in 27.3 ml of dry THF, cooled to -78 ° C and stirred under argon. After completion of dropwise addition of the acetyl anion solution to the phosphinylchloridate solution, the dark orange mixture was stirred at -78 ° C for 1 hour, then quenched at -78 ° C with 50 ml of saturated NH4Cl, warmed to room temperature and diluted with water. The aqueous layer was extracted with ether. The organic extracts were combined, washed once with saturated aqueous NaHCO 3 and once with brine, dried over MgSOb, filtered and evaporated under reduced pressure. The product was isolated by flash chromatography on a 6 "Merck silica gel column 50% eluting with 50% ethyl acetate in hexane at a flow rate of 2ml / min. The yield was 0.609 g (0.945 mmol, 21%) of the titular phosphinate in the form of a golden orange oil.

TLC: silica gel, Rf = 0.32, 50%. ethyl acetate in hexane. Mass spectrum: (Cl): m / e 639 (M + H) +.

B. (S) -4 - [[2- [3,5-Dimethyl- [1,1 ' -bufen-l] -2-yl] ethyl] hydroxyphosphinyl] 3- (tert-butyldiphenylsilyloxy) butanoic acid dithium salt.

Argon was passed through a solution of 0.867 g (1.37 mmol) of acetylated phosphinate from Part A in 13 ml of methanol for 10 minutes. 0.315 g of 10% was added. Pd / C and the reaction mixture was hydrogenated in a Parr apparatus at 0.28MPa. After shaking for 24 hours, the reaction mixture was filtered through a celite pad on a sintered glass funnel. The celite was washed with methanol and the filtrate was evaporated in vacuo to give 0.896 g of a yellow oil which was purified by flash chromatography on a 50 mm diameter column of 6 "Merck silica gel, eluting with 40% of a yellow oil. ethyl acetate in hexane gradually going to 50% solution. 0.680 g (1.06 mmol, 77% yield) of the title saturated phosphinate was obtained as a pale yellow foam. Purification of the flash chromatography column by eluting with methanol gave an additional 0.087 g of slightly impure product. TLC: Rf = 0.27, silica gel, 50%. acetone in hexane. Mass spectrum (CI): m / e 643 (M + H) +.

C. (S) -4 - [[2- [3,5-Dimethyl] - [, 1'-biphenyl] -2-yl] ethyl] hydroxyphosphinyl] 3-hydroxybutanoic acid dithium salt.

0.66 g (1.03 mmol) of Part B phosphinate was mixed under argon at room temperature with 12.65 ml of dry THF. To this solution was added dropwise 0.247 g (0.236 ml, 4.12 mmol) of glacial acetic acid, followed by 2.81 ml (3.09 mmol) of a 1.1 M solution of nC-HgNF in THF. After stirring at room temperature for 16 hours, the reaction was quenched by adding 25 ml of ice water. The aqueous layer was extracted with ethyl acetate. The organic extracts were combined, washed twice with saturated aqueous NaHCO 3 and once with brine, dried over MgSO 4, filtered and evaporated under reduced pressure. The product was purified by flash chromatography on a 6 "Merck silica gel 40 mm column, eluting with 50% of the product. acetone in hexane. 0.363 g (0.898 mmol, 87% yield) of the title alcohol was obtained in the form of a white solid. TLC: silica gel, R = ^0, 3 ^{0, 50} -proc. acetone ^h EK sleigh. Mass Depth ( ^F AB): m / e ⁴⁰⁵ ( ^M + ^H ) ⁺ .

Example VII A. Acid methyl ester (S) - ^ - ^ - [[: ^ - [^ '- ffl ^ oi ^ (^ -; ^, 5-ddT ^ er / k3 [1,1- ^ bffenyl] -2-yl] ethyl] hydroxyphosphinyl] -3- (tert-butyldiphenyl] osliyloxy) butane.

- Argon was bubbled through a solution of 0.685 g (1.04 mmol) of (S) -4 - [[2 [4'-fluoro-3,5-dimethyl] 1,1'-biphenyl-2-yl] phtnyl] acid methyl ester] hydrocyphosphinyl] -3- (tfrt-butyldiphenylsilyloxy) butane in 9.9 ml of methanol for 10 minutes. 0.239 g of 10% was added. Pd / C and the reaction mixture was hydrogenated under Parr pressure at 0.28 MPa. After shaking for 24 hours, the reaction mixture was filtered through a pad of celite on a sintered glass funnel, the celite was washed with methanol and the filtrate was evaporated to give 0.638 g of a green oil. The product was purified by flash chromatography on a 6 "Merck silica gel column 40 mm in diameter, eluting with 45%. ethyl acetate in hexane at a flow rate of 2 ml / min. 0.530 g (0.802 mmol, 77% yield) of the title saturated phosphinate was obtained in the form of a pale yellow foam. In addition, 0.09 g (0.136 mmol, 13%) of a slightly impure product was obtained. TLC: silica gel, R "= 0.30 ⁽⁵ 0- ^p roc. Etyto in ethyl hexamethylene). Mass Depth ( ^CI ): m / e ⁶⁶¹ ( ^M + ^H ) +.

B. (S) -4 - [[2- [4'-Fluoro-O-3,5-dimefyl [1,1-biphenyl] -2-yl] ethyl] methoxyphosphinyl] -3-hydroxybutanoic acid methyl ester.

0.525 g (0.794 mmol) of Part A phosphinate was stirred under argon at room temperature with 9.74 ml of anhydrous THF. To this solution, 0.191 g (0.182 ml, 3.18 mmol) of glacial acetic acid was added dropwise, followed by dropwise addition of 2.17 ml (2.38 mmol) of a 1.1 molar solution of n-C4H9NF THF. After stirring for 16 hours at room temperature, the reaction was quenched by adding 15 ml of ice water. The aqueous layer was extracted 3 times with ethyl acetate. The organic extracts were combined, washed 2 times with saturated aqueous NaHCO 3 and once with brine, dried over MgSO 4, filtered and evaporated. The product was purified by flash chromatography on a 6 "Merck silica gel column 40 mm in diameter with 50% elution. acetone in hexane at a flow rate of 2 ml / mmut. The product fractions were concentrated and azeotroped to dryness with toluene to give 0.281 g (0.665 mmol, 84% yield) of the title alcohol as a white solid. Impurity was observed in ¹ 1 H ^NMR at ^{270 MH} not ^d k> t ^o about ^dd ziehć ^{and was} wtóziatae ^Getting rctenych systems ^{TL TL} C. ^C: silica gel ^R <= ^{0.31 (50} - ^p roc. Acetone in ebony). ^{1 H} NM ^r : ^{270 MH} z ^, CDCl 2. Mass spectrum (CI): m / e 423 (M + H) +.

Example VIII. (S) -4 - [[2- [1- (4-Fluoroffnyl) -4- (1-methylethyl) -2-phenyl-1H-imidazo-5-yl-phenyl] -me [oxyphosphinyl-3-hyd [ oxybutane.

A. (S) -3 - [(1,1-dimethylethyl) diffnylsil] okky] -4 - [[2- [1- (4-fluorophenyl) -1-methylethyl-4-phenyl] acid methyl ester -1H-imidazol-5-yl] ethyl] methoxyphoxinyl] butane.

To a solution of 839 g (1.14 mmol) of (S) -3 - [(1,1-dimethyl) diffnylsiiyloxy] -4 - [[1- (4-fluorophenyl) -4- (1) -phthylethyl) -2-phenyl-1H-imidazole-yl] ethynyl] methoxyphosphinylbutane in 86 ml of dry methanol was added 213 mg of 10%. Pd / C and hydrogenated at room temperature in a Parr apparatus overnight at ca 0.28MPa. The suspension was filtered through celite and the clear filtrate was evaporated to dryness and dried in vacuo to give 853 g (100% yield) of the title compound as a thick syrup. TLC: Rf = 0.17 (silica gel, hexane - acetone 7: 3).

B. (S) -4 - [2- [1- (4-Fluorophenyl) -4- (1-methyloethyl) -2-phenyl] -so-1-himidazole --- yl] ethyl] methoxyphosphinyl] -3- ^e hydroksybutanow it.

To a solution of 853 mg (approximately 1.14 mmol) of Part A compound in 11.0 ml of dry tetrahydrofuran was added successively 270 μ / 1 (4.60 mmol) of glacial acetic acid and 3.62 ml (3.62 mmol) of 1.0 -mol (C4H9NF in hexane then stirred overnight at room temperature at rt

157 761 argon atmosphere. The reaction mixture was diluted with 25 ml of ice-water then extracted with three portions of 100 ml of ethyl acetate. The combined organic extracts were washed with 15 ml of saturated NaHCO 3 and 25 ml of brine, dried over anhydrous MgSO 4, filtered and evaporated to dryness.

The crude product, 958 mg, was chromatographed on a silica gel column, eluting the column with 1: 1 and 4: 1 acetone-hexane. The desired fractions were combined, evaporated to dryness and dried in vacuo to give 443 mg (77.0% of the title compound as a solid). TLC: Rt = 0.13 (silica gel, acetone - hexane 1: 1).

Example IX. (S) -4 - [[2- [1- (4-fluoro-phenyl) -3-methyl-2-naphthalenyl] ethyl] hydroxyphosphinyl] -3-hydroxybutanoic acid dipithium salt. .

A. (S) -3 - [[1,1-dimethylethyl / dieenylsil] oxy] -4 - [[2- [1- (4-fluorophenyl} -3-methyl-2-naphthalenyl] ethynyl] methoxyphosphinyl acid methyl ester ] butane.

0.675 g (0.974 mmol) of (S) -3 - [[1,1-dimethylethyl) diphenylsilyl] oxy] -4- [[[1-4-fluoro-phenyl) -3-methyl- [beta] -2-naphthalenyl] methoxyphosphate The 'i ^ y ^^] ^ ^ tan tane was dissolved in 14.3 ml of methanol. Argon was bubbled through the solution for 10 minutes before adding 0.270 g, 10%. Pd / C and the reaction mixture was shaken on a Parr hydrogenator under 0.28MPa hydrogen for 24 hours. The reaction mixture was filtered through a pad of celite that was washed thoroughly with methanol, then the filtrate was evaporated under reduced pressure to give a white foam. The product was purified by flash chromatography on a 50 mm diameter column containing a Merck 113 mm silica gel bed, using 70% of the eluent. ethyl acetate in hexane with a linear flow rate of 5 cm / min. 0.556 g (0.798 mmol, 82% yield) of the title saturated phosphonate was obtained in the form of a white foam. Eluting the column with methanol gave an additional 0.101 g (0.145 mmol, 15%) of product. TLC: Rf = 0.24 (60% ethyl acetate / hexane, silica gel, PMA).

¹ H NMR (270 MHz, CDCl): δ 7,78-7,14 (Ar, 19H) 4.44 (m, 1 H), 3.61 (s, 3H), 3.35 + 3.23 (2 X d, 3H, J = 10.6Hz), 2.92 - 2.83 (m, 1H), 2.63-2.54 (m, 3H), 2.21-1.27 (m, 4H ), 2.45-2.42 (2xs, 3H), 1.00 (s, 9H).

Mass spectrum: CI m / e 697 (M + H) ⁺

IR (CHC3 solution): 3028, 3019, 3007, 2997, 2953, 2859, 1735, 1510, 1497, 1472, 1463, 1439, 1428, 1378, 1364, 1314, 1236, 1197, 1157, 1142, 1112, 1091, 1073, 1065, 1043, 823 αηΛ

B. (S) -4 - [[2- [1- (4-fluorophenyl) -3-methyl-2-naphthalenyl] ethyl] methoxyphosphinyl] -3-hydroxybutanoic acid methyl ester.

To 0.540 g (0.775 mmol) of Part A silyl ether, stirred under argon in 9.45 ml of dry THF, 0.186 g (0.177 ml, 3.10 mmol) of glacial acetic acid was added, followed by dropwise addition of 2.1 ml (2.33 mmol). ) 1.1 M tetrabutylammonium fluoride solution in THF. The reaction mixture was stirred at room temperature for 16 hours, then quenched with 30 ml of ice water and diluted with ethyl acetate. The aqueous layer was extracted 3 times with ethyl acetate. The organic extracts were combined, washed once with saturated NaHCO3 in water and once with brine, dried under MgSO4, filtered and evaporated under reduced pressure. Preliminary purification by flash chromatography on a 40 mm diameter column containing a bed of 15 cm Merck silica gel, eluting with

4% methanol in CH2Cl212 at a linear flow rate of 5 cm / min gave 0.40 g of a white solid. This material was triturated in hexane, filtered and dried under high vacuum for 8 hours to give 0.317 g (0.691 mmol, 89% yield) of the title hydroxyphosphinate as a white solid, mp 120-122 ° C. TLC: Rf = 0.12 (2% methanol / CH-Cl-, silica gel)

1 H NMR (270 MHz, CDCl 3): δ 7.76 (d, 1H, J = 7.9 Hz), 7.69 (s, 1H), 7.42 - 7.16 (m, 7H), 4, 42 + 4.26 (2 X m, 1H), 3.92 + 3.84 (2 X d, 1H, J = 3.16 Hz), 3.72 (s, 3H), 3.58 + 3, 5 (2 X d, 3H, J = 3.69 Hz), 2.89 - 2.76 (m, 2H), 2.56 (s, 3H), 2.63 - 2.41 (m, 2H) , 1.92 - 1.61 (m, 4H).

Mass spectrum: CI m / e 459 (M + H) +.

IR (KBr): 3428 (wide, 3287 (wide), 3064,3050,3017,2989,2952,2921,1737,1603,1510,1497,1458,1438,1234,1221, 1191, 1175, 1042, 826 cm !

Example XA Acid methyl ester [1S- [1 ^a / R ^x / 2 ^a , 4 ^b , 8 ^b , 83 N -4- ^ 8 - N (1,1-dimethylethyl) dimethylsilyl] -oxy]. 4a, 5,6,7,8,8a-octahydro-2-methyl-1-naphthalenyl] -ethynyl] methoxy [phosphinyl] -3 - [[[-1 [-dimethylethyldieenylsil] oxy] butane.

157 761

For a solution of 356 mg (1.17 mmol) [1S- / 1 - <a, 2 <a, 4a <b, 8 <b, 8a <a] -8 - [[1,1-dimethylethyl] dimethylsilyl] -oxy ] -1- [ethynyl-1,2,4a, 5,6,7,8a-oktahvdro-2-methylnaphthalene in 5 ml of dry THF at -78 ° C 740μΙ (1.17 mmol) 1.1 (6 molar) was added dropwise n-butyllithium solution in hexanes and the resulting clear mixture was stirred under argon at -78 ° C for 30 minutes The anion of the acetylene compound was then added dropwise via a catheter over 15 minutes to a cooled to -78 ° C acid methyl ester solution (S> - 4- (chlorome [oxyphosphyl) -3 - [- (1,1-dimethylethyl) diphenylsilyl] oxy] butane in 6 ml dry THF The yellow mixture was stirred at -78 ° C for 30 minutes, then quenched by dropwise addition of saturated 5 ml of NH4Cl and the mixture was warmed to room temperature Water and ethyl acetate were added to the mixture, the organic phase was washed with brine, dried over anhydrous Na2SO4 and evaporated under reduced pressure to give 1.282 g of a pale yellow oil. was performed by flash chromatography on silica gel eluting with hexane-ethyl acetate 7: 3. Product fractions were evaporated to yield 624 mg (72.4%) of the title acetylenic phosphinate as a colorless gum. TLC: Ri = 0.49 (hexane-acetone, 7: 3, UV + PMA).

B. EisUf metytówy k ^wa s u [^ - [/ 1 ⁴ R ^χ / 2 ^4, 4 ^4, 8 ^- 84 ^4]] - 4 - [[2- [8 - [/ 1 J-dimethyl ^ lo (dimethylsilyl] -oxy] dec4-hydro-2-methyl-1-naphthalenyl-ethyl-ethyl-methoxyphosphinyl] -4 - [[(1,1-dimethylethyl) diphenylsilyl] oxy] butane.

To a solution of 498 mg of acetylenic phosphinate from part A in 6 ml of methanol was added 200 mg of 10%. The Pd / C and the black slurry were shaken in a Parr vapor under 0.28 MPa hydrogen pressure for 48 hours. The catalyst was removed by filtering the suspension through celite, a new 150 mg of catalyst was added to the reaction mixture and shaking in a Parr apparatus under a hydrogen pressure of 0.28 MPa was continued for 24 hours. The catalyst was removed by filtering the suspension through celite and the filtrate was evaporated under reduced pressure to give 448 mg of a colorless glassy substance. The crude product was purified by flash chromatography on silica gel eluting with hexane-ethyl acetate 8: 2. Product fractions were evaporated under reduced pressure to yield 334 mg (66.5%) of the title compound as a colorless glass. TLC (7: 3 ethyl acetate-hexane UV + PMA): Rf of the three diastereoisomers as one spot - 0.42, Rf of the fourth diastereoisomer - 0.49.

C. ^E ster methyl ester ^[s 1 - [1 ⁴ / ^RX / ²⁴ '' La ^ 8 ^ ⁴ and ^{8 /} l]] - 4- [L2 / dekahyd] ^{o-8-hydroxy-2-methyl l} ol -n4phthal [nyl] -e [yto] me [oxyfo] flnyto] --- [[[l,] - dimethyl] oeth] o (diphenylsilyl] ckt-butane.

To a solution of 248 mg (0.334 mmol) of Part B compound in 4 ml of CH3CN was added 36 µM (1 mmol) of 48%. Hr in water and the mixture was stirred for 6.5 hours under argon at room temperature. The mixture was partitioned between ethyl acetate and saturated NaHCO3, the organic phase was washed with brine, dried over anhydrous Na2SO4 and evaporated under reduced pressure to give 227 mg of a colorless glass. The crude product was purified by flash silica gel chromatography eluting with hexane-ethyl acetate 4: 1 followed by ethyl acetate alone. Product fractions were evaporated in vacuo to afford 159 mg (75.8%) of pure title mono alcohol as a colorless oil. TLC: Rf = 0.5 (acetone-hexane, 7: 3, UV (weak) + PMA).

Pattern 74

R ⁵

R ⁶ i

-CUT

Formula 68 (R ^la l _t

RO CH ₂ H CH ₂ PC CO, R ^X

OH

Pattern 70

R6

AND

R6

ABOUT

-p-ch ₂ -ch-ch, -go i I ł

X OH i

WITH

Pattern 61

R ⁶ i

Pattern 58

Pattern 59

COOH

Pattern 46 hal \ / ^R ie cc / \ ^ 17 ^R e

Pattern 49

-ch-ch ₂ -c - ch ₂ -cooh

OH

OH

Pattern 50

H \

Z =

-Cch ₂ ) <1

Formula 36 (CH ₂ ) q

Re * 3 | 2 1 ch ch ₂ -c ch ₂ cooh

OH OH

Pattern 37

Pattern 32

Pattern 33

Pattern 34 Pattern 35

Pattern 21

R *

II nitrile alkyl - C.

CH3CH2C cFormula 16

_R 14Q

Formula 1

Formula 17

157 761

alkyl - O - P - CH ₂ - CH - CH ₂ - CO ₂ alkyl CH (cis) CH

Z Formula alkyl - O- P-CH ₂ -CH- CH ₂ - O) ₂ R ^xa

CH AT OH (cis) CH WITH Pattern

O alkyl-O-P- (H2 CH (cis) CH z

CH -CH2 'CO2H ÓH

IB pattern by o

R ^xa aP - CH-CH - CH2 - (^ 2R ^x a CH ÓH (cis) CH

Z Formula 1B ⁴

Ο ll w

R- Ρ - OH- CH - (Hj-COjR *

CH ÓH u

CH (cis) r

WITH

O alkyl-OP-CH ₂ -CH - CHy-CO ₂ alkyl OC Ó i J ° c (ch ₃ ) ₃ ^C 6 ^H 5 ^C 6 ^H 5

Pattern 2 o

alkyl-O-P- (H.

Z-ch (cis)

CH

CH-CH ₂ CO ₂ alkyl &

J _x i - (OHjIj

Ch c, h ₅

Formula 3

Department of Publishing of the UP RP. Circulation of 90 copies. Price PLN 5,000.

Claims (2)

Patent claims A method for the preparation of new phosphorus-containing derivatives of 3-hydroxybutanoic acid, of the formula I, in which R is a hydroxyl group or a lower alkoxy group, Z is a group of the formulas 4,5,6,15, wherein R ¹ , R ² , R ^2a and ^R2b are the same or different, and each independently is hydrogen, halogen, lower alkyl, haloalkyl, phenyl, substituted phenyl, or the group OR ^y where R ^y is hydrogen, alkanoyl, benzoyl, phenyl , halogen-lower alkyl, phenyl-lower alkyl, lower alkyl, cinamyl, halo-alkyl, allyl, cycloalkyl-lower alkyl, adamantyl-lower alkyl or substituted phenyl-lower alkyl, and when Z is the group of formula 15, R ⁵ and R ⁵ are the same or different and represent hydrogen, lower alkyl or OH group, R6 is a group of formula 16 such as formula 17 or aryl-CH2, R6a is lower alkyl, hydroxy, keto or halogen, q is zero, 1, 2 or 3 and R7 is hydrogen or lower alkyl when Z is the group of formula 6 One of R ³ and R ⁴ is a group of formula 18 and the other is lower alkyl, cycloalkyl or phenyl- (CH2) p-, wherein p is zero, 12.3, or 4, R8 is hydrogen, -4alkyl, Cca-cycloalkyl, C 1-4 alkoxy (except T-butoxyl), trifluoromethyl, fluoro, chloro, phenoxy or benzyloxy, R ⁹ is hydrogen, Ci-3alkyl, Ci-3alkoxy, trifluoromethyl, fluoro, chloro , phenoxy or benzyloxy group, provided that if R8 is hydrogen, R9 must be hydrogen, not more than one of R8 and R9 is trifluoromethyl, not more than one of R8 and R9 is phenoxy and not more than one of R8 and R9 is benzyloxy, R ^x is hydrogen or a lower alkyl group as the free acid or in the form of a physiologically acceptable and hydrolysable salt ester, characterized in that the acetylenephosphinate of formula II, wherein Z is as defined above, is subjected to the reduction to form the sil ether III (cis), in which the silyl ether group is cleaved to give the diester of formula 1B1, if necessary, it is converted to the monoester of formula 1B ³ (cis), basic salt of formula 1B2 (cis), acid of formula IB (cis), or a dibasic salt of formula IB4 (cis). 2. Process for the preparation of new phosphorus-containing derivatives of 3-hydroxybutanoic acid of formula 1, in which R is a hydroxyl group or a lower alkoxy group, Z is a group of formulas 7,8,9,10,11, 12,13,14, when Z is a group of formula 7,8, 9, 10 and 11, one of the substituents R ³ and R 4 is a group of the formula 18, and the other is lower alkyl, cycloalkyl or phenyl- (CH2) p-, wherein p is zero 1, 2, 3 or 4, R ¹³ is hydrogen, lower alkyl, lower alkoxy (except t-butoxy), halogen, phenoxy or benzyloxy, R ¹ 4 is hydrogen, lower alkyl, lower alkoxy, halogen , phenoxy or benzyloxy group, R 5 is hydrogen, lower alkyl, lower alkoxy or halogen, provided that when R ³ is hydrogen, both R "and R ¹ * must be hydrogen if R ^u is hydrogen R 5 must be a hydrogen atom, not more than one of R ⁿ and R14 is trifluoromethyl, not more than one of R1 ³ and R "is and phenoxy, and not more depressions one of R ⁵ and R ^M is a benzyloxy group, R ¹⁰ and R "are each independently hydrogen, alkyl, cycloalkyl, adamantyl-1 or a group of formula 69 wherein R1 ^3, R * and R5a are as defined above, and q is zero, 1, 2, 3 or 4, Y is oxygen, sulfur or N-R10, in case Z is a group of formula 12, Ra is hydrogen or primary or a secondary C1-6alkyl group, Rb is a primary or secondary C1-6alkyl group, or Ra + Rb is a group - (Ccyr- or cis-CH2-CH = CH-CH2-, r is 2, 3, 4, 5 or 6, ^R8 is hydrogen, Ci-4alkyl, 6C ^^ ^ c ^^ * ^ ^{<^^} lkil, Ci-4 alkoxy (except t-butoxy), trifluoromethyl, fluoro, chloro, phenoxy or benzyloxy , R9 is hydrogen, Ci-3alkyl, Ci-3alkoxy, trifluoromethyl, fluoro, chloro, phenoxy or benzyloxy with the proviso that when R ⁸ is hydrogen, R 9 must be hydrogen, not more than one of the R ^e and R 9 is trifluoromethyl, not more than one of R ^e and R 9 is phenoxy, and not more than one of ^R8 and R9 is benzyloxy, R ^tt is 157 761 3 lower alkyl, cycloalkyl or the group of formula 18, and R ¹³ , R ¹⁴ and R ^14a are as defined above, when Z is a group of formula 13, R ¹⁵ and R ¹⁶ are both hydrogen, chlorine, bromine , CN, CF 3, phenyl, Ci-C4alkyl, C2-ealkoksykarbonylową, CH 2 oR ¹⁷ or CH2OCONHR ^18, R 1 ⁷ is hydrogen or Ci_6alkyl, R 18 is alkyl or phenyl optionally substituted by fluorine, chlorine, bromine or a Ci-4 alkyl, or R ^w and R ^w together form a group - (CH2) ₃ -, -CH2-O-CH2-, -CON (R1 ⁹ ) CO- or -CON (R2 ⁰ ) N (R21) CO-, s is 3 or 4 , R ¹⁹ is hydrogen, C 1-6 alkyl, phenyl or benzyl, R 20 and R ²¹ are hydrogen, C 1-4 alkyl or benzyl, and R ³ and R 4 are as defined above when Z is a group of formula 14, R 22 is lower alkyl, cycloalkyl, adamantyl-1 or a group of formula 19 where t is 1, 2, ³ or 4, R2 ³ and R23a are the same or different and each independently is hydrogen, lower alkyl, lower alkoxy (with except I-butoxyl ), halogen, phenoxy or benzyloxy group, provided that when R is hydrogen, R must be hydrogen, not more than one of R and R23a is trifluoromethyl, not more than one of R23 and R2 ^3a is phenoxy and no more than one of R23 and ^R2 R3a represents a benzyloxy group, R ^x is hydrogen or lower alkyl in free acid form or in the form of a physiologically acceptable and hydrolysable to acid in salt form, characterized in that the acetylenofosfinian of formula 2, wherein Z is as defined above is subjected to reduction to give the silyl ether of formula 3 (cis ^ which cleaved the silyl ether group to form a diester of the formula 1 B ¹ (cis), and if necessary it is converted into the monoester of formula IB (cis ), a basic salt of formula IB (cis) acid of formula IB (cis), or a dibasic salt of formula IB4 (cis).