AP223A

AP223A - "Triazole antifungal agents".

Info

Publication number: AP223A
Application number: APAP/P/1991/000237A
Authority: AP
Inventors: Ray Stephen James Dr; Richardson Kenneth Dr
Original assignee: Pfizer
Priority date: 1990-02-02
Filing date: 1991-01-28
Publication date: 1992-08-27
Also published as: HK219396A; DK0440372T3; FI971238A0; BA97298A; CZ279339B6; MY105494A; JPH09208583A; JP2848811B2; ES2055523T4; IS3671A7; ES2055523T3; OA09480A; NO176796C; CA2285891C; RO109648B1; FI971238A; PL169332B1; LU90960I2; PE31691A1; MA22054A1

Abstract

The invention provides antifungal compounds of the formula

Description

PLC 5,7 (SPC 7704 ) AP 0 0.0 2 2 3

TRIAZOUE ANTIFUNGAL AGENTS

This invention relates to triazole derivatives which haveantifungal activity.

More particularly this invention relates to 2-aryl-3-(3-halopyridin-4-yl or 5-halopyrimidin-4-yl)-l-(lH-l,2,4-triazol-l-yl)alkan-2-ol derivatives which are useful in the treatment offungal infections in animals, including human beings.

Seme of the compounds of the present invention are disclosedin a general sense in our European Patent Application No. 89307920.2 (EP-A-0357241) but none of them are specificallydescribed or exemplified therein.

It has new been discovered that the conpounds of the presentinvention have a surprisingly high level of anti fungal activity,in particular against Aspergillus spp. fungi, which is mainlyattributable to their unexpectedly good pharmacokinetic propertieswhich result in longer half-lives (t'-s values).

The invention provides antifungal agents of the formula:-

(I) PL C 5 17 (SPC 7704 ) 2 and pharmaceutically acceptable salts thereof, wherein R is phenyl substituted by 1 to 3 substituents eachindependently selected from halo, -CF^ and -OCF^; R1 is C -C alkyl; R2 is H or ^"^4 alkyl; X is CH or N; and Y is F or Cl.

In the above definition of compounds of the formula (I) halois F, Cl, Br or I and C3 and C4 alkyl groups may be straight- orbranched-chain. Preferred alkyl groups are methyl and ethyl.

Examples of R include 2-fluorophenyl, 4-fluorophenyl, 2-chlorophenyl, 4-chlorophenyl, 2-branophenyl, 2-iodophenyl,2-trifluorcmethylphenyl, 2,4-dichlorophenyl, 2,4-difluorophenyl,2-chloro-4-fluorcphenyl, 2-fluoro-4-chlorophenyl, 2,5-difluorophenyl, 2,4,6-trifluorophenyl, 4-brano-2,5- difluorophenyl and 2-trifluorcnethoxyphenyl. R is preferably phenyl substituted by 1 to 3 halosubstituents, more preferably by 1 or 2 halo substituents.

Yet more preferably R is phenyl substituted by 1 or 2substituents each independently selected iron fluoro and chloro.

BAD ORIGINAL PL C 517 (SPC 7704 ) AP00O223 3

Preferred individual embodiments of R include 2-f luorophenyl,4-fluorophenyl, 2,4-difluorophenyl, 2-chlorophenyl and 2,4-dichlorophenyl.

Most preferably R is 2-f luorophenyl, 2,4-di fluorophenyl,2-chlorophenyl or 2,4-dichlorophenyl.

Preferably R1 is methyl.

Preferably R2 is H or methyl. 2 .

Most preferably R is H. 1 2 .

Preferably R is methyl and R is H or methyl. l 2 .

Most preferably R is methyl and R is H.

Preferably X is N.

Preferably Y is F.

The pharmaceutically acceptable salts of the ocnpounds of theformula (I) include acid addition salts formed from acids whichform non-toxic salts such as the hydrochloride, hydrobrtmide,hydro iodide, sulphate or bisulphate, phosphate or hydrogenphosphate, acetate, maleate, fumarate, lactate, tartrate, citrate,gluconate, benzoate, methanesulphonate, benzenesulphonate andp-toluenesulphonate salts. For a review on suitablepharmaceutical salts see Berge et al. J. Fharm. Sci., 66, 1-19(1977). 1 . . 2

Where R is identical to R , the compounds of the formula (I)contain one chiral centre and therefore exist as a pair ofenantiomers (a racemate).

Where and R2 are different, the compounds of the formula(I) contain at least two chiral centres (*) and therefore exist asat least two diastereoisomeric pairs of enantiomers, i.e.

BAD ORIGINAL v PIC 517 (SPC 7704)

The invention includes both the individual stereoisomers ofthe compounds of the formula (I) together with mixtures thereof.Separation of diastereoisomers may be achieved by conventionaltechniques, e.g. by fractional crystallisation, chromatography or H.P.L.C. of a diastereoisomeric mixture of a ocnpound of theformula (I) or a suitable salt or derivative thereof. Anindividual enantiomer of a conpound of the formula (I) may also beprepared from a corresponding optically pure intermediate or byresolution, either by H.P.L.C. of the racemate using a suitablechiral support or by fractional crystallisation of thediastereoisomeric salts formed by reaction of the racemate with asuitable optically active acid, e.g. lR-(-)- or IS-(+)-10-camphorsulphonic acid. 2

The preferred compounds of the formula (I) when R is H havethe 2R,3S- configuration, i.e.

BAD ORIGINAL PIC 51? (SPC 7704) AP 0 0 0 2 2 3 5

Particularly preferred individual embodiments of thecompounds of the present invention are 2R, 3S-2- (2,4-di fluorophenyl) —3— (3—f luoropyridin-4-yl) -1- (LH-1,2,4-triazol-l-yl)butan-2-ol, 2R, 3S-2-(2-chlorophenyl) -3- (3-f luoropyridin-4-yl) -1- (LH-1,2,4-triazol-l-yl)butan-2-ol, 2R, 3S-2-(2-fluorophenyl) -3-(3-fluoropyridin-4-yl) -1-(1H-1,2,4-triazol-l-yl)butan-2-ol, 2R, 3S-2- (2,4 -difluorophenyl )-3-( 5-f luoropyrimidin-4 -yl) -1- (1H- 1,2,4-triazol-l-yl) butan-2-ol, and 2R,3S-2-(2,4-dichlorophenyl)- 3- (5-fluoropyrimidin-4-yl )-1-( 1H-1,2,4-triazol-l-yl) butan-2-ol,and pharmaceutically acceptable salts thereof.

Ihe compounds of the formula (I) provided by the inventionmay be prepared by the following methods:- 1) All the compounds of the formula (I) may be prepared as shewnin Scheme 1:-

Scheme 1

1) 3ase, solvent Λ compound οι Cheformula (I)

r a

BAD ORIGINAL PLC 5 17 (SPC 7Z04 ) 6 1 2 wherein R, R , R , X and Y are as defined for a compound of the formula (I) .

In a typical procedure a compound of the formula (II) isdeprotonated by the addition of approximately one equivalent of asuitable base, e.g. lithium diisopropylamide, or sodium orpotassium bis(trimethylsilyl)amide, and the resulting salt(preferably the lithium, sodium or potassium salt) is reacted insitu with a ketone of the formula (III) - The reaction istypically carried out at from -80* to -50*C, preferably at from-70' to -60'C, in a suitable organic solvent, e.g. tetrahydrofuran, toluene or diethyl ether, and under an inertatmosphere, e.g. nitrogen or argon.

The starting materials of the formula (Π) are either kncwnoonpounds (e.g. see D. L. Canins et al, Heterocycles, 22, 339(1984)) or may be prepared by conventional procedures inaccordance with literature precedents. The starting materials ofthe formula (III) are either known compounds (e.g. see EP-A-44605,EP-A-69442 or GB-A-1464224) or may be prepared by similar methods to those described therefor. 2) All the compounds of the formula (I) may also be prepared as shewn in Scheme 2:- BAD ORIGINAL & PLC 5 I Z (SPC ZZ04 ) AP000223 7

Scheme 2

A base sale of acompound of the formula N^NH .y , solvent

H , base,solvent A compound of the formula (I) 1 2 wherein R, R , R , X and Y are as defined for a compound of theformula (I) and Z is a suitable leaving group, e.g. chloro, branoor cj~C4 alkanesulphonyloxy (such as methanesulphonyloxy).Examples of suitable base salts of 1H-1,2,4-triazole are alkalimetal, preferably sodium and potassium, and tetraalkylammonium,preferably tetra-n-butylammonium (see US-A-4259505), salts.

Ihe reaction is preferably carried out using an epoxide ofthe formula (IV) as the starting material. If a ccnpound of theformula (VI) is used in this process, it is probable that thereaction mechanism dictates, at least in part, that thecorresponding epoxide of the formula (IV) is formed in situunder the reaction conditions. Ihe process is therefore, in

BAD ORIGINAL PLC 517 (SPC 7704) 8 this respect, similar to that utilising an epoxide of the formula(IV) as the starting material.

When a base salt of lH-l,2,4-tria2ole is used, the reactionis typically carried out at from room temperature to 100‘C,preferably at about 60‘C when using the sodium salt of1H-1,2,4-triazole, and preferably at about room temperature whenusing the corresponding tetra-n-butylanmonium salt, in a suitableorganic solvent, e.g. Ν,Ν-dimethylformamide or tetrahydrofuran.

Alternatively, the reaction may be carried out using1H-1,2,4-triazole in the presence of an additional suitable base,e.g. Ν32°°3 or at fron 50’ to 100’C in a suitable solvent, e.g. Ν,Ν-dimethyl formamide, methanol or aqueous acetone.

Ihe intermediates of the formula (IV) and (VI) may beprepared by conventional techniques as sunroarised by the followingmethods shewn in Schemes 3 and 4:-

Scheme 3 1) Base, solvent (II)

PL C 5 17 < SPC 7 704 ) AP o 0,0 2 2 3 9 1 2 wherein R, R , R , X and Y are as defined for a compound of theformula (I) and Z is a leaving group, preferably Cl or Br.

In a typical procedure, a conpound of the formula (II) isdeprotonated by the addition of approximately one equivalent of asuitable base, e.g. lithium di isopropyl amide, or sodium orpotassium bis (trimethylsilyl) amide, and the resulting organometallic intermediate is reacted in situ with a compound ofthe formula (V). The reaction is typically carried out at from-80* to -50*C, preferably at about -70*C, in a suitable organicsolvent, e.g. tetrahydrofuran, toluene or diethyl ether, and underan inert atmosphere, e.g. nitrogen or argon. The compound of theformula (VI) formed need not be isolated and is generally eye Usedin situ after a period of stirring at a higher temperature, e.g.room temperature, to provide an axirane of the formula (TV). A compound of the formula (VI) when Z is chloro or brans mayalso be prepared by reacting an epoxide of the formula (IV) withthe appropriate hydrogen halide under anhydrous conditions.

BAD ORIGINAL PL C 517 (SPC 7704 ) 1 10

Scheme 4 KCO^CC^-C^ alkyl) (VII)

(where R^ = C^-C^ alkyl) <-

Base, (C^-C^alkyl)Z^

(VIII)

Base, (C^C^ alkyl)Zl

Epoxidation

(where = C “C^ alkyl)

(IVA)

BAD ORIGINAL PLC 517 CSPC 7704) AP00Q223 deprotonation of a compound 11 wherein R, R1, R^, X and Y are as defined for a compound of theformula (I) and Z1 is a suitable leaving group, e.g. Cl, Br, I ormethanesulphonyloxy.

In a typical procedure a compound of the formula (VIII), (IX)or (X) is prepared directly from an ester of the formula (VII) byreaction with an organometallic intermediate derived by of the formula :-

as aDorooriate. wherein R1 2 R , X and Y are as defined for accnpound of the formula (I), with approximately one equivalent ofa suitable base, e.g. lithium diisopropylamide or sodiumbis (trimethylsilyl) amide. The reaction is typically carried outat from -80* to -50*C, preferably at about -70*C, in a suitableorganic solvent, e.g. tetrahydrofuran or diethyl ether, and underan inert atmosphere, e.g. nitrogen or argon.

Alternatively, a compound of the formula (IX) or (X) may beprepared by reacting, respectively, a compound of the formula(VIII) or (IX) with approximately one equivalent of a suitablebase, e.g. sodium hydride, followed by alkylation of the resultantcarbanion in situ with a suitable alkylating agent. The reactionis typically carried out at from O’C to roan temperature in asuitable organic solvent, e.g. N,N-dimethylformamide.

BAD ORIGINAL pW PIC 517 (SPC 7704 ) 12

Preferably, alkylation of a compound of the formula (VTH) or(IX) is performed under phase transfer conditions, e.g. usingNaOH/[ai3(CH2)3]4N® eHSO4/H2O/C«Ciy alkylJZ1 (wherein Z1 is preferably iodo), at from 0‘C to room temperature, and typicallyat rocm temperature.

Epoxidation of a ketone of the formula (IX) or (X) isperformed using conventional methods, e.g. usingdimethyloxosulphonium methylide (e.g. see J.A.C.S. [1965], 87, 1353) or chloranethyllithium (e.g. see Tet. Lett. [1986], 795). . 1 2 3) The compounds of the formula (I) wherein R, R , R and Y areas defined for a oatpound of the formula (I) and X is N may beprepared as shewn in Scheme 5:-

Scheme 5

(XI)

Reduction->

A compound ofthe formula (I)wherein X is N 1 2 wherein R, R , R and Y are as defined for a compound of the2 3

formula (I) and Z and Z are each independently selected from H and a group that may be selectively removed by reduction, with the 2 3 2 proviso that Z and Z cannot both be H. Preferably Z is the

BAD ORIGINAL PLC 5 17 (SPC 7704) AP 0 0-0 2 2 3 13 group that may be selectively removed by reduction and Z3 is H.Preferably the group that may be selectively removed by reductionis halo (defined as F, Cl, Br or I) and most preferably is chloro.

When said group is halo, preferably chloro, the preferredmethod of reduction is by hydrogenolysis. In a typical procedurea compound of the formula (XI) is subjected to hydrogenolysisusing a suitable catalyst, e.g. palladium-on- charcoal, and asuitable solvent, e.g. ethanol, optionally in the presence of anadditional suitable base, e.g. sodium acetate. Ihe reaction maybe carried out at from room tenperature to the reflux temperatureof the solvent and at a pressure of from 1 to 5 atmospheres (100kfd to 500 kPa), but generally proceeds satisfactorily at about room temperature and at about atmospheric pressure. • . . 2

Ihe intermediates of the formula (XI) wherein one of Z andZ3 is H and the other is a group that may be selectively removedby reduction may be conveniently prepared as shewn in Scheme 6:-

Scheme 6

1) Base, solvent Λ compound of ------—--the lonuu la (XI) 2)

(HI) BAD ORIGIN*' PLC 517 (SPC 7704 ) 14 1 2 wherein R, R , R and γ are as defined for a compound of the2 3 · formula (I) and one of Z and Z is H and the other is a group that may be selectively removed by reduction. The reaction may becarried out by a similar procedure to that described in Method (1).

Ihe intermediates of the formula (XI) wherein one of Z2 andZ3 is H and other is a group that may be selectively removed byreduction may also be prepared by an analogous procedure to thatdescribed in Method (2).

Ihe starting materials of the formula (XII) may be preparedby conventional procedures such as are illustrated in the following Preparations section. . . . 2 3

Ihe intermediates of the formula (XI) wherein Z and Z areeach a group that may be selectively removed by reduction may beprepared by an analogous procedure to that described in Method (2)by using an appropriate epoxide starting material which may beprepared as shewn in Scheme 7 using conventional procedures:- bad original £ PLC 517 <SPC 7704) AP 0 04) 2,2 3 15

-,3

1 2 wherein R, R , R and Y are as defined for a compound of the2 3 formula (I), Z and Z are each a group that may be selectively 4 removed by reduction and Z is chloro or C1~C4 alkoxy.

All of the above reactions are conventional and appropriate reagents and reaction conditions for their performance andprocedures for isolating the desired products will be well knownto those skilled in the art, in accordance with literatureprecedents and by reference to the Examples hereto. BAD ORIGINAL £ PLC 5,7 (SPC ,’704, 16 Λ pharmaceutically acceptable acid addition salt is readilyprepared by mixing together solutions containing the free base andthe desired acid. The salt generally precipitates from solutionand is collected by filtration, or is recovered by evaporation of the solvent.

The compounds of the formula (I) and their salts are anti-fungal agents, useful in the curative or prophylactic treatment offungal infections in animals, including humans. For example, theyare useful in treating topical fungal infections in man caused by,among other organisms, species of Candida. Trichophyton.

Microsporum or Epidermophyton. or in mucosal infections caused byCandida albicans (e.g. thrush and vaginal candidiasis). They canalso be used in the treatment of systemic fungal infections causedby, for exanple, species of Candida (e.g. Candida albicans),

Cryptococcus neofortnans. Aspergillus flavus. Aspergillus fumiqatus. Coocidioides. Paracoocidioides. Histoplasma or

Blastomyces.

The compounds of the present invention have been found tohave unexpectedly good activity against the clinically importantAspergillus spp. fungi. This is mainly attributable to theirunexpectedly good pharmacokinetic properties which result inlonger half-lives (t1-, values).

The in vitro evaluation of the antifungal activity of thecompounds can be performed by determining the minimum inhibitoryconcentration (m.i.c.), which is the concentration of the testcompounds, in a suitable medium, at which growth of the particularmicro-organism fails to occur. In practice, a series of agarplates, each having the test oompound incorporated at a particular

bad ORIGINAL PL C 5 17 (SPC 7704) AP0 0 Q2 2 3 17 concentration, is inoculated with a standard culture of, forexample, Candida albicans, and each plate is then incubated for 48hours at 37 *C. The plates are then examined for the presence orabsence of growth of the fungus and the appropriate m.i.c. valueis noted. Other micro-organisms used in such tests can includeAspergillus fumigatus, Trichophyton spp., Microsporum spp.,Epidermoohyton floooosum. Coccidioides immitis and TPrulopsis glabra ta.

The in vivo evaluation of the compounds can be carried out ata series of dose levels by intraperitoneal or intravenousinjection, or by oral administration, to mice which are inoculatedwith, e.g., a strain of Candida albicans or Aspergillus fumigatus.Activity is based on the survival of a treated group of mice afterthe death of an untreated group of mice. The dose level at whichthe compound provides 50% protection against the lethal effect ofthe infection (Ρ°5θ) is noted. For Aspergillus spp. infectionmodels, the number of mice cured of the infection after a set doseallows further assessment of activity.

For human use, the antifungal compounds of the formula (I)and their salts can be administered alone, but will generally beadministered in admixture with a pharmaceutical carrier selectedwith regard to the intended route of administration and standardpharmaceutical practice- For example, they can be administeredorally in the form of tablets containing such excipients as starchor lactose, or in capsules or ovules either alone or in admixturewith excipients, or in the form of elixirs, solutions orsuspensions containing flavouring or colouring agents. They can

BAD ORIGINAL PIC 517 (SPC 7 70 4 ) 13 be injected parenterally, for example, intravenously, intramuscularly or subcutaneously. For parenteral administration,they are best used in the form of a sterile aqueous solution whichmay contain other substances, for example, enough salts or glucose to make the solution isotonic with blood.

The solubility of a oonpound of the formula (I) in an aqueousmedium may be improved by conplexation with a hydroxyalkylderivative of a cyclodextrin in the preparation of an appropriatepharmaceutical composition. Preferably the cyclodextrin used isalpha-, beta-, or gamma-cyclodextrin and most preferably isbeta-cydodextrin. Preferably the hydroxyalkyl derivative is ahydroxypropyl derivative.

For oral and parenteral administration to human patients, thedaily dosage level of the antifungal compounds of the formula (I)and their salts will be from 0.01 to 20 mg/kg (in single ordivided doses) when administered by either the oral or parenteralroute. Thus tablets or capsules of the compounds will containfrcro 5 mg to 0.5 g of active ccnpound for administration singly ortwo or more at a time, as appropriate. The physician in any eventwill determine the actual dosage which will be most suitable foran individual patient and it will vary with the age, weight andresponse of the particular patient. The above dosages areexemplary of the average case; there can, of course, be individualinstances where higher or lower dosage ranges are merited, andsuch are within the scope of this invention.

Alternatively, the antifungal compounds of formula (I) can beadministered in the form of a suppository or pessary, or they maybe applied topically in the form of a lotion, solution, cream,ointment or dusting powder. For example, they can be incorporated BAD ORIGINAL £ PIC 517 (SPC 7704) AP O 0 0 2 2 3 19 into a cream consisting of an aqueous emulsion of polyethyleneglycols or liquid paraffin; or they can be incorporated, at aconcentration between 1 and 10%, into an ointment consisting of awhite wax or white soft paraffin base together with suchstabilizers and preservatives as may be required.

Thus the invention further provides a pharmaceuticalcomposition comprising a compound of the formula (I), or apharmaceutically acceptable salt thereof, together with apharmaceutically acceptable diluent or carrier.

Ihe invention yet further provides a compound of the formula(I), or a pharmaceutically acceptable salt or oonposition thereof,for use as a medicament, in particular as an antifungal agent.

Ihe invention also provides the use of a compound of theformula (I), or of a pharmaceutically acceptable salt orcomposition thereof, for the manufacture of an antifungal agent.

Ihe invention yet further provides a method of treating ananimal (including a human being) to cure or prevent a fungalinfection, which comprises treating said animal with an effectiveamount of a compound of the formula (I), or with, as appropriate,a pharmaceutically acceptable salt or composition thereof.

Ihe invention also provides novel intermediates of theformulae (IV), (VI) and (XI), 4-ethyl-5-fluoropyridimine and 4-chloro-6-ethyl-5-fluoropyrimidine.

Ihe following Examples illustrate the preparation of thecompounds of the formula (I). It is believed that enantiomericpair B, when referred to in any following Example or Preparation,and the products of Examples 1, 3, 4 and 5 (in each of which onlyone of the two possible enantiomeric pairs was obtained) are aracemic mixture of the 2R,3S- and 2S,3R- enantiomers. BAD ORIGINAL & PLC 5 17 ( SPC 7704 )

JO E?<AMPLE 1 3-(3-Chioropyridin-4-yl·) -2-(2,4-di fluorophenyl) -1-( 1H-1,2.4- trianol-l-yl)butan-2-ol

F

To a solution of diiscpropylamine (1.01 g, 10 nmol) in dry1HF (60ml) at -60 "C and under a nitrogen atmosphere was addeddropwise a 1.6 M solution of n-butyllithium in hexane (6.25ml, 10nmol). The mixture was allowed to warm to -20*C then recooled to-70*C and to the resulting solution of lithium diisopropylamide(LEA) (10 mmol) at -70’C was added dropwise 3-chloro-4-ethylpyridine (see D. L. Comins et al, Heterocycles, 22, 339(1984)) (1.41 g, 10 mmol). The resulting mixture was stirred atthis temperature for 15 minutes after which time a solution of1-(2,4-difluorophenyl)-2-(1H-1, 2,4-triazol-l-yl)ethancne (2.23 g,10 mmol) in THF (15ml) was added. This mixture was allowed towarm to room temperature over a 30 minute period and the reactionwas quenched by the addition of water (30ml) and extracted with bad ORIGINAL g PLC 517 (SPC 7704) AP00D223 2: ethyl acetate (3 x 60ml) . The combined organic extracts were dried over magnesium sulphate, filtered, concentrated under reduced pressure and the title compound isolated by "flash" chromatography on silica eluting with ethyl acetate. The product was recrystallised from ethyl acetate (yield = 0.46 g), m.p. 182-184’C. Found: C,55.76; H,4.15; N,15.23; C._H.CC1F.N.O1I lb 24 requires: C,55.98; H,4.14; N,15.36%. EXAMPLE 2 2-(2,4-Difluorophenvl) -3-(3-fluoropyridin-4-yl) -1- ( 1H-1.2.4- triazol-l-vl)butan-2-ol

The reaction was carried out by a similar method to thatdescribed for Example 1 using 4-ethyl-3-fluoropyridine (seePreparation 1) instead of 3-chloro-4-ethylpyridine as the startingmaterial. Column chromatography of the crude reaction product onsilica using ethyl acetate as the eluant first gave, aftercombination and evaporation of the appropriate fractions, thetitle compound, enantiomeric pair A, m.p. 178-181’C, which wascharacterised by Hi-NMR spectroscopy.

BAD ORIGINAL P-L C 5 1 7 ( SPC 7704 ) 22 ^Ή-NMR (CDC1 ): 6 - 1.6 (d, 3H) , 3.95 (q, 1H) , 4.7 and 5.15 (AB q,2H), 5.1 (s, IB (OH)), 6.5 (m, IB), 6.7 (m, IB), 6.95 (m, IB), 7.45 (t, IB), 7.8 (s, IB), 7.95 (s, IB), 3.15 (s, IB), 8.25 (d, IB) ppm.

Further elution with 95:5 ethyl acetate/methanol provided,after combination and evaporation of the appropriate fractions,the impure title compound, enantiomeric pair B. Ibis was furtherpurified by column chrcsnatography on silica using 93:7:1dichlorcsnethane/methanol/0.880 aqueous ammonia as the eluant. Theappropriate fractions were combined and evaporated to provide,after trituration with diethyl ether, the title compound,enantiomeric pair B, m.p. 188-9’C. Found: C,57.63; H,4.32; N,15.71; C17H15F3N40.0.25 1^0 requires: C,57.87; H,4.43; N,15.88%.

Enantiomeric pair B was resolved by H.P.L.C. using a chiral support (CHIRACEL ' 0G) and eluting with 1:1 isopropanol/hexane.

The appropriate fractions were combined and evaporated to providethe resolved individual enantiomers, each contaminated with thechiral support.

Each impure enantiomer was further purified by columnchromatography on silica using dichloromethane/methanol (95:5) asthe eluant. The appropriate fractions were combined andevaporated to give, after trituration with hexane/diethyl ether, apurified individual enantiomer. 25

One enantiomer of m.p. 57-59*C and (ce]D -59’ (c = lmg/ml in25 methanol) and another of m.p. 56-57’C and [α]θ + 57’ (c = lmg/mlin methanol) were obtained. BAD ORIGINAL Ά PIC 517 (SPC 7704) AP 0 0 6 2 2 3 23 EXAMPLES 3 to 6

The following tabulated compounds of the general formula:-

were prepared by a similar method to that described for Exanple 1using the appropriate 4-ethyl-3-halcpyridine and l-(halophenyl)-2-(lH-l,2,4-triazol-l-yl)ethanone as the starting materials.

BAD ORIGINAL Λ 2*

BAD ORIGINAL & AP o 0 0 2 2 PLC 5 17 (SPC 7 704 )

bad original s PIC 517 (SPC 7704) » ___ 26 (1) Column chromatography was carried out on silica with agradient elution using 2:1 ethyl acetate/dichloromethanefollowed by ethyl acetate as the eluant. The resulting solidobtained was triturated with diethyl ether to provide thedesired product. (2) See Example 1 for reference to starting material. (3) See Preparation 1 for starting material. (4) Column chromatography was carried out on silica with agradient elution using 2:1 ethyl acetate/dichloromethanefollowed by ethyl acetate as the eluant. The appropriatefractions were combined and evaporated and the materialobtained was further purified by column chromatography onsilica using 93:7:1 dichloromethane/methanol/0.880 aqueousammonia as the eluant. The appropriate fractions werecombined and evaporated and the residue triturated withdiethyl ether to provide the desired product. (5) The enantiomeric pair obtained was resolved by H.P.L.C. usinga similar method to that described in Example 2. Thisprovided the individual enantiomers, one of m.p. 83-84*C and 25 [α]θ -80* (c = lmg/ml in methanol) and the other of m.p.78-79*C and [α]θ5 +82* (c = lmg/ml in methanol). (6) Column chromatography was carried out on silica using80:20:1.5 hexane/isopropanol/ 0.880 aqueous ammonia as theeluant. The appropriate fractions were combined andevaporated and the material obtained was further purified bycolumn chromatography on silica using 97:3 ethyl acetate/ethanol as the eluant. The appropriate fractions werecombined and evaporated to provide the separated enantiomericpairs. Each enantiomeric pair was triturated with diethylether to provide the desired product. (7) The enantiomeric pair obtained was resolved by H.P.L.C. usinga similar method to that described in Example 2. BAD ORIGINAL ά PL C 517 (SPC 7 704) AP 0 0,0 2 2 3 EXAMPLE 7 2-(2.4-Dif luorophenyl) -3-( 5-fluoropyrrimidin-4-yl) -1-( 1H-1,2,4-tri- azol-l-yl) butan-2-ol

A solution of 3-(4-chloro-5-fluoropyriinidin-6-yl)-2-(2,4- difluorophenyl) -1- (1H-1,2,4-triazol-l-yl) butan-2-ol, enantiomeric pair B (see Preparation 2(iii)) (0.307 g, 0.8 mmol) in ethanol (20ml) was hydrogenated at atmospheric pressure and at room temperature in the presence of 10% palladium-on-charcoal (30 mg) and sodium acetate (0.082 g, 1 mmol). After 5 hours a further 10 mg of 10% palladium-on-charcoal was added and hydrogenation was continued for an additional 1 hour period. Ihe catalyst was removed by filtration and the filtrate v/as concentrated in vacuo. "Flash" chromatography of the residue on silica using 97:3 ethyl acetate/methanol as the eluant provided, after combination and evaporation of appropriate fractions and trituration with diethyl ether, the title compound, enantiomeric pair B, (0.249 g, 89%), m.p. 127-C. Found: C,55.08; H,4.00; N,19.96; C, H FNOlo 14 3 5 requires: C,55.01; H,4.01; N,20.05%.

BAD ORIGINAL PL C 517 (SPC 7704) 28 Λ saitple of the title compound, enantiomeric pair B, (0.105g,0.3 mmol) and lR-(-)-10-camphorsulphonic acid (0.07g, 0.3 iranol)were dissolved in methanol (4ml) then cooled to 0*C for 2 hours.Ihe resulting crystalline solid was collected by filtration togive 2R, 3S-2- (2,4-difluorophenyl) -3-(5-fluoropyrimidin-4-yl) -1-(1H-1,2,4-triazol-l-yl) butan-2-ol lR-(-)-10-canphorsulphonate 0.5methanol (0.06g), m.p. 176’C, [α]θ5-49.5* (c = 2 mg/ml inmethanol). Found: C,53.09; H,5.36; N,11.43; C^gH^F.jNgOgS.O.SCH^OH requires: C,53.27; H,5.36; N,11.73%. Ihe absoluteconfiguration of the compound was confirmed by single crystalX-ray analysis.

Ihe filtrate from the crystallisation was evaporated in vacuo and partitioned between dichloromethane (10ml) and saturated aqueous sodium bicarbonate solution (5ml). Ihe organic layer was dried over magnesium sulphate, filtered and concentrated under reduced pressure. The residue and IS-(+)-10-camphorsulphonic acid (0.46g, 0.2 mmol) were dissolved in methanol (3ml) then cooled to 0*C for 2 hours. Ihe crystalline solid was collected by filtration to give 2S,3R-2-(2,4-difluorophenyl)-3- (5-f luoropyrimidin-4-yl) -1- (1H-1,2,4-triazol-l-yl) butan-2-ol IS-(+)-10-camphorsulphonate 0.5 methanol (0.052g), m.p. 176*C, [a]25+ 54.5· (c = 2mg/ml in methanol). Found: C,53.27; H,5.31; N,11.64; C_.HFNOS. 0.5 CH_,OH requires: C,53.27; H,5.36;

ZD JU J □ □ J N,11.73%. bad original PLC 5 1 Z ( S P C Z Z 0 4 ) AP 0 0Ό 2 2 3 29 A sample of the 1R- (-) -10-camphorsulphonate salt (1.22g, 2.1mmol) prepared according to the above method was partitionedbetween dichloromethane (20ml) and saturated aqueous sodiumbicarbonate (3ml). Ihe organic layer was washed with water (5ml)then dried over magnesium sulphate, filtered and evaporated invacuo to give 2R,3S-2-(2,4-difluorophenyl)-3-(5- f luoropyrimidin-4-yl) -1- (1H-1,2,4-triazol-l-yl) butan-2-ol(0.64g), m.p. 127*C, [α]θ5 -62' (c = lmq/ml in methanol). A sample of the IS-(+)-10-camphorsulphonate salt (1.17g, 2.0mmol) prepared according to the above method was treated by asimilar method to that described above for the 1R-(-)-10-camphorsulphonate salt to give 2S,3R-2-(2,4-difluorcphenyl)-3-(5-fluorcpyrimidin-4-yl) -1- (1H-1,2,4-triazol-l-yl) butan-2-ol(0.63g),-m.p. 127*C, [α]θ5 +59.5* (c = 2mg/ml in methanol). EXAMPLE 8 2-(2,4-Difluorophenvl) -3- (5-fluoropvrimidin-4-vl) -1- (1H-1.2.4-

triazol-l-yl)butan-2-ol, enantiomeric pair B

BAD ORIGINAL £ PLC 5 1 7 t S PC /7 0 i ) 30

To THF (200ml) was added sodium bis (trimethylsilyl) amide(79ml of a 1.0M solution in THF) and the solution cooled to -65"Cunder nitrogen. A solution of 4-ethyl-5-fluoropyrimidine (lOg)(see Preparation 8) in THF (100ml) was added over 30 minutes.After stirring for 3 hours at -65 ’C the thin slurry was treatedwith a solution of l-(2,4-difluorophenyl)-2-(lH-l,2,4-triazol-l-yl)ethanone (17.7g) in THF (100ml), dropwise over 30 minutes.

The solution was stirred for a further 1 hour at -65’C and then treated with acetic acid (20ml). After warming to -20 *C thesolution was washed with water (200ml) and the organic layerseparated and combined with an ethyl acetate (200ml) back extractof the aqueous phase. The combined organic layers wereconcentrated under reduced pressure to provide a solid that wastriturated with diethyl ether (230ml) and filtered. The filtratewas concentrated under reduced pressure and chromatographed onsilica with 1:1 diethyl ether/ethyl acetate as the eluent. Thefractions containing the title compound were combined,concentrated under reduced pressure and the residuechromatographed on silica with 1:1 ethyl acetate/hexane as theeluent. The appropriate fractions were combined and evaporatedunder reduced pressure to provide the purified title compound(0.82g), m.p. 125-127’C. Found: C,54.89; H,4.06; N,19.66;C16H14F3N5° ret3uires: C,55.01; H,4.01; 14,20.05%.

BAD ORIGINAL PLC 517 (SPC 7704) AP 0 0 0_2 2 3 31 EXAMPLE 9 2-(2,4-Difluorophenyl) -3-(5-fluoropyrimidin-4-yl)-1-( 1H-1,2,4-

triazol-l-yl)butan-2-ol, enantiomeric pair A

Ihe title compound was prepared by a similar method to thatused in Example 7 using 3-(4-chloro-5-fluoropyrimidin-6-yl) - 2- (2,4-di fluorophenyl) -1- (1H-1,2,4-triazol-l-yl) butan-2-ol, enantiomeric pair A (see Preparation 2(iii)) as the startingmaterial. This gave the product, m.p. 137’C. Found: C,54.39;H,4.06; N,19.82; F^O requires: C,55.01; H,4.01; N,20.05%. EXAMPLE 10 3- (5-Chloropvrimidin-4-yl) -2- (2,4-difluorophenyl) -1- (1H-1,2,4-tri-

azol-l-yl)butan-2-ol, enantiomeric pair B

BAD ORIGINAL PLC 5 17 (SPC 7 704 ) 32 A solution of 3-(4,5-dichloropyrimidin-6-yl)-2-(2,4-d ifluorophenyl)-1-( IB-1,2,4-triazol-l-yl)butan-2-ol, enantiomericpair B (see Preparation 6(iii)) (0.58 g, 1.46 mmol) in ethanol (20ml) was hydrogenated at atmospheric pressure and at roomtemperature in the presence of 10% palladium-on-charcoal (45 mg)and sodium acetate (122 mg, 1.5 nmol) for 7 hours. The catalystwas then removed by filtration and the filtrate was concentratedunder reduced pressure. "Flash" chromatography of the residue onsilica using ethyl acetate as the eluant provided, aftercombination and evaporation of the appropriate fractions, thetitle compound (0.35 g, 72%), m.p. 128’C. Found: C,51.68; H,3.89; N,18.58; 1^0 requires: C,51.76; H,3.94; N,18.87%. EXAMPLE 11 3- (5-Chloropvrimidin-4-vl) -2-(2,4-difluorophenvl) -1- (1H-1.2.4-

triazol-l-vl)butan-2-ol, enantiomeric pair A

The title compound was obtained by a similar method to that used in Exanple 10 using 3-(4,5-dichloropyrimidin-6-yl)-2-(2,4-difluorophenyl) -1-(1H-1,2,4-triazol-l-yl)butan-2-ol, enantiomericpair A (see Preparation 6(iii)) as the starting material. Thisgave the product as a gum that was characterised by Si-NMRspectroscopy. ^H-NMR (CDC13) 5 = 1.50 (d, 3H), 4.4 (q, IB), 4.67 and 4.82 (AB q,2H), 6.35 (s, 1H (OH)), 6.45 (m, 1H) , 6.62 (m, IB), 7.07 (m, 1H),7.6 (s, 1H) , 8.05 (s, IB), 8.5 (s, IB), 8.8 (s, IB) ppm. bad original PLC 517 (SPC 7704 ) AP 0 0 0 2 2 3 33 EXAMPLES 12 to 16

The following tabulated compounds of the general formula :-

were prepared by a similar method to that described for Example 10using the appropriate 2-aryl-3-(4-chloro-5-fluorcpyrimidin-6-yl)-l-(lH-l,2,4-triazol-l-yl)butan-2-ol as the starting material. bad original V.

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Cl cn in vo vo BAD ORIGINAL d PL C 5 17 (SPC 7704 ) AP 0 0>0 2 2 3 35 (1) Column chromatography was carried out on silica using 96:4ethyl ace tate/methanol as the eluant. (2) Column chromatography was carried out on silica usingisobutylmethylketone as the eluant. (3) See Preparation 3 for starting material. (4) See Preparation 4 for starting material. (5) See Preparation 5 for starting material. (6) The enantiomeric pair obtained was resolved by H.P.L.C. usinga similar method to that described in Example 2.

BAD ORIGINAL P L C 5 1? (SPC '704) 36 EXAMPLE 17

An aqueous saline soLution of 2R,3S-2-(2.4-difi.uorophenyl)- 3-(5-fluoropvrimidin-4-yl) -1-(LH-l,2,4-triazol-l-yl) butan-2-ol and hydroxypropyl-^-cyclodextrin

Hydroxypropyl-/?-cyclodextrin (Molar Substitution = 0.41, lg)was placed in a 10ml volumetric flask and dissolved in distilledwater (ca. 7ml). Sodium chloride (90mg) was added and dissolvedin the solution and the volume made up to 10ml with distilledwater. The resulting solution was added to 2R,3S-2-(2,4-difluorophenyl) -3- (5-f luoropyrimidin-4-yl) -1- (1H-1,2,4-triazol-l-yl)butan-2-ol (lOOmg) (see Exairple 7) in a vial and the mixturewas sonicated for 15 minutes and then further mixed by mechanicalrotation of the vial for 2 days. A further quantity ofhydroxyprcpyl-/3- cyclodextrin (200mg) was then added and themixture mixed by mechanical rotation of the vial for 1 hour toprovide the title solution. bad original $ PLC 517 (SPC 7 704) 0 0 0 22 3 37

The follcwing Preparations illustrate the preparation ofcertain novel starting materials used in the Examples.

Preparation 1 4-Ethvl-3-fluoropyridine CH.

1) LDA, THF

2) c2h5!

To a stirred solution of IDA (200 nmol) in dry 1HF (400 ml)(prepared by a similar method to that used in Example 1) at -70*Cand under a nitrogen atmosphere was added dropwise 3-fluoro-pyridine (20 g, 200 mmol). After 30 minutes at this temperatureethyl iodide (60 g, 370 nmol) was added dropwise to the reactionand the mixture was allowed to warm slowly to between -10* and-5*C whereupon an exotherm occurred and the temperature rose to 15* to 20 ‘C. Ihe mixture was stirred for a further 30 minutesafter which time the reaction was quenched by the addition ofwater (50 ml) and the organic phase separated. Ihe aqueous phasewas extracted with ether (3 x 50 ml) and the combined organiclayers were dried over magnesium sulphate and concentrated underreduced pressure. Ihe resulting liquid was distilled atatmospheric pressure to yield the title compound (13 g), b.p.154-158*C, which was characterised by Si-NMR spectroscopy.

BAD ORIGINAL PLC 517 (SPC 7 70t) )8 ^H-NMR (CDC1 ): ό - 1.25 (t, 3H, J = 10 Hz), 2.65 (q, 2H, J = 10Hz), 7.1 {t, IB, J - 8Hz), 3.3 (d, 1H, J = 8Hz), 8.33 (s, IB) ppm.

Preparation 2 3-(4-Οΐ1θΓθ-5-ΠυθΓθρντϊιηϊάίη-6-ν1) -2-(2,4-difluorophenyl)-1- (1H-1,2,4-triazol-l-yl)butan-2-ol

BAD ORIGINAL ft PLC 517 (SPC 7704) AP0O0223 39 (i) 6-Ethyl-5-f luoroovrimidίη-4 (3H) -one

To a solution of sodium methoxide (8.64 g, 160 mmol) inmethanol (50 ml) at O’C was added a solution of ethyla-fluoropropionylacetate (see E. D. Bergmann et al, J.

Chem. Soc., 1959, 3278 and D. J. Burton et al, Tet. Lett.,30, 6113 (1989)) (12.96 g, 80 mmol) and fonnamidine acetate(8.32 g, 80 mmol) in methanol (50 ml) and the resultingmixture was stirred at 0*C for 1 hour, overnight at roomtemperature and finally for 30 minutes under reflux. Themixture was cooled and the excess sodium methoxide wasneutralised by the addition of glacial acetic acid (10 g).The reaction was concentrated under reduced pressure andthe residue was dissolved in hot ethyl acetate, theinsoluble sodium acetate was removed by filtration and thefiltrate was concentrated under reduced pressure. "Flash"chromatography of the residue using ethyl acetate as theeluant provided, after combination and evaporation ofappropriate fractions and trituration with diethyl ether,the title oompound (5.5 g, 48%), m.p. 105-106'C. Found: C,50.38; H,4.85; N,19.63; CJ^F^O requires: C,50.70;H,4.93; N,19.72%.

The title compound may also be prepared as described inPreparation 7. (ii) 4-Chloro-6-ethvl-5-fluoropyrinu.dine A mixture of the product of part (i) (6.4 g, 45 irnnol) andphosphoryl chloride (30 ml) was heated under reflux for 3hours. The excess phosphoryl chloride was removed by

BAD ORIGINAL PL C 5 17 (SPC 7 704) 40 distillation under reduced pressure and the residue waspoured into ice-water. The resulting .mixture was extractedwith methylene chloride (3 x 50 ml) and the combined organicextracts were washed with water and dried over magnesiumsulphate. The solvent was removed under reduced pressureand the resulting oil was distilled under reduced pressureto provide the title compound (4.81 g, 66%), b.p. 74*C at22 ran Hg, which was characterised by Si-NMR spectroscopy. ^H-NMR (CDC13) : & = 1.3 (t, 3H, J = 10Hz), 2.9 (q, 2H, J =10Hz), 8.68 (s, 1H) ppm. (iii) 3- (4-Odoro-5-fluorcovrimidin-6-vl) -2- f 2.4-difluorophenvl) -1- (1H-1,2,4-triazol-l-yl) butan-2-ol

To a solution of IDA (20 nmol) in 1HF1 (50 ml) (prepared bya similar method to that used in Example 1) under anitrogen atmosphere and at -70’C was added dropwise asolution of the product of part (ii) (3.2 g, 20 nmol) inTHF1 (30 ml) over 15 minutes. The resulting mixture wasstirred at this temperature for 3 hours. To the resultingsolution was added a solution of l-(2,4-difluorophenyl)-2-(1H-1,2,4-triazol-l-yl)ethanone (4.46 g, 20 nmol) in THF(50 ml) and the mixture was maintained at -70*C for 1 hour and then at -50*C for a further 1 hour. The reaction was quenched by the addition of a solution of glacial aceticacid (1.2 g) in water (10 ml) and the mixture was allowedto warm to rocm temperature. The organic phase wasseparated, the aqueous phase extracted with ethyl acetate

BAD ORIGINAL PLC S 1 7 (SPC 7 704) AP ο 0·0 2 2 3 41 (20 ml) and the combined organic layers were dried overmagnesium sulphate and concentrated under reduced pressure.Column chromatography of the residue on silica using 3:2ethyl acetate/diethyl ether as the eluant first gave, aftercombination and evaporation of appropriate fractions andtrituration with diethyl ether, the title compound,enantiomeric pair B (0.94 g, 12%), m.p. 92*C. Found:C,49.93; H,3.57; N,18.17; C requires: C,50.06; H,3.39; N,18.25%.

Further elution gave, after combination and evaporation ofappropriate fractions, the title compound, enantiomericpair A contaminated with ketone starting material. Thiswas purified by several recrystallisations from diethylether to provide the product, m.p. 132*C. Found: C,49.93;H,3.58; N, 18.23; C dEjNgO requires: C,50.06; H,3.39;N,18.25%. (1) IHF may be replaced by toluene. BAD ORIGINAL ./ PLC 517 CSPC 7704) 42

Preparations 3 to 5

The following tabulated compounds of the general formula :-

were prepared by a similar method to that described in Preparation2(iii) using 4-chloro-6-ethyl-5-fluoropyrimidine and theappropriate l-aryl-2-(lH-l,2,4-triazol-l-yl)ethanone as thestarting materials. BAD ORIGINAL ΌΪ co o oM> in 04 O' w in o X»«z d. g o

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Preparation 6 3-(4.5-Dichloropyrimiriin-6-yI) -2-(2,4-difluoroptieny].)-I.-(lH- 1,2, 4-^Ϊ3"οΙ-1-γ1) butan-2-ol

och3

CH^ONa, CS^OH^NH HC\

NH2.CH3COOH aq. H2°2’ c.HCl (ii)

BAD ORIGINAL PL C 5 17 (SPC 7 704) AP000223 ( i) 6-Ethylpyrimidin-4 (3H) -one

To a solution of sodium methoxide (4.19 kg, 77.6 mol) and forrumidine acetate (3.0 kg, 28.8 mol) in methanol (45 L)at 5-10’C was added slowly a solution of methylpropionylacetate (2.5 kg, 19.2 mol) in methanol (10 L)maintaining the temperature belcw 20*C throughout theaddition. The resulting mixture was stirred at roomtemperature overnight after which time the pH was adjustedto 7 by the addition of concentrated hydrochloric acid. Thereaction mixture was concentrated under reduced pressure toca.10 L in volume, diluted with water (10 L) and was extractedwith 2-butanone (2 x 30 L). The combined organic extracts wereconcentrated under reduced pressure to ca. 2 L in volume anddiluted with ethyl acetate (4 L). The desired productcrystallised from the solution (2.4 kg, 70%) and wasrecrystallised from isopropanol to yield a product of m.p. 132-134’C. Found: C,58.45; H,6.37; N,22.41; CgHg^O requires: C,58.05; H,6.50; N,22.57%. (ii) 4,5-Dichloro-6-ethylpyrimidine

To a solution of 6-ethylpyrimidin-4 (3H) -one (the product ofpart (i)) (18.6 g, 150 mmol) in concentrated hydrochloricacid (120 ml) at 30-40’C was added dropwise a 30 wt. %solution of hydrogen peroxide in water (18 ml) over aperiod of 30 minutes (slight exotherm resulted) and theresulting mixture was stirred overnight at 40'C. Themixture was concentrated under reduced pressure and theresidue was suspended/dissolved in toluene and the tolueneremoved under reduced pressure.

BAD ORIGINAL PL C S ! 7 ( SPC Z Λ0 £. } 46

The residue was dissolved in phosphorus oxychloride (150ml) and heated under reflux for 3 hours after which timethe excess phosphorus oxychloride was removed under reducedpressure. The residue was poured into ice/water, extractedwith methylene chloride (3 x 50 ml) and the combinedorganic extracts were washed with water (30 ml) and driedover magnesium sulphate. The solvent was removed underreduced pressure and the resulting oil was distilled underreduced pressure to yield the title compound (5.4 g, 20%),b.p. 104*C at 22 mm Hg, which was characterised by Si-NMRspectroscopy. V-NMR (CDC13): i = 1.3 (t, 3H, J = 10Hz), 3.04 (q, 2H, J =10Hz), 8.75 (s, 1H) ppm. (i ii) 3-(4,5-Dichloropyrimidin-6-yl) -2- (2,4-difluorophenyl) -1- (1H-1,2,4-triazol-l-vl) butan-2-ol

To a solution of LEA (13.6 mmol) in THF (50 ml) (preparedby a similar method to that used in Example 1) at -70 *C wasadded dropwise 4,5-dichloro-6-ethylpyrimidine (the productof part (ii)) (2.37 g, 13.3 mmol) and the resultingsolution was stirred at this temperature for 10 minutes. Asolution of 1-(2,4-difluorophenyl)- 2-(lH-l,2,4-triazol-l-yl)ethanone (2.97 g, 13.3 mmol) in THF (50ml) was added to the reaction mixture at such a rate so as tomaintain the reaction temperature belcw -50’C. After stirring at -70*C for 1 hour and at -50*C for a further 1 hour the reactionwas quenched by the addition of 10% aqueous acetic acid (11 ml).

BAD ORIGINAL ?LC 517 (SPC 7704) AP00D223 47

The organic phase was separated, the aqueous phaseextracted with ethyl acetate (2 x 20 ml) and the combinedorganic layers were dried over magnesium sulphate. Afterremoval of the solvent under reduced pressure, the residuewas triturated with diethyl ether (25 ml) and the unreactedketone starting material (1.7 g) was removed by filtration.The filtrate was concentrated under reduced pressure and"flash" chromatography of the residue on silica using 65:35ethyl acetate/diethyl ether cis the eluant first provided,after combination and evaporation of appropriate fractionsand trituration with diethyl ether, the title compound,enantiomeric pair B as a solid (670 mg, 13%), m.p. 124’C.Found: C,47.78; H,3.33; N,17.13; C^^C^F^O requires:C,48.00; H,3.25; N,17.50%.

Further elution gave, after combination and evaporation ofappropriate fractions and trituration with diethyl ether,the title compound, enantiomeric pair A as a solid (527 mg,10%), m.p. 137*C. Found: C,48.02; H,3.30; N,17.39;C16H13Cl2F2N5° C,48.00; H,3.25; N,17.50%. bad original PL C 5 17 ( SPC 7 704 )

Preparation 7 6-Ethyl-5-fIucropvri-midin-4 (3H) -one "Δ (i)

(ii) i) CH^CH^MgBr,

DME.THF

ii) AcOH

Pd/C. C2H5OH BAD ORIGINAL ft PL.: 5 1 1 (SPC //0-) AP000223 ( i) 2.4-DichIoro-5-fluoropyrimidine

To phosphorus oxychloride (141.4g) at 25"C was addedpowdered 5-fluorouracil (20g). The resulting slurry washeated to 90 "C and N,N-dimethyloniline (37.3g) was added over 1 hour. The reaction was then heated at reflux for 5 hours and 70g of the phosphorus oxychloride was removed bydistillation. The mixture was then cooled to 25’C and quenched into 3N HC1 (200ml) at O'C, portionwise over 1hour. The title compound was then extracted from themixture using dichloranethane (2 x 70ml). The combineddichloromethane layers were washed with water (50ml) andconcentrated under vacuum to give an oil (24g), which wascharacterised by ^H-NMR and mass spectrometry. H-NMR (CDC13): <S = 8.5 (s, 1H) ppm.

Mass Spec.: m/e = 166. (ii) 2,4-Dichloro-l, 6-dihvdro-6-ethvl-5-f luoropyrimidine

To magnesium turnings (4.27g) in tetrahydro furan (56ml) wasadded a solution of bromoethane (19g) in THF (19ml) over 5hours. To this slurry at 0’C was added a solution of theproduct of part (i) (24g) in 1,2-dimethoxyethane (70ml)over 1 hour. The reaction was quenched at 10*C usingglacial acetic acid (lOg) to give a solution of the titleconpound which was used directly in the next step.

BAD ORIGINAL PLC 5 1 7 (SPC 7 7 0 -) 50 (l i i) 2,4-Dichloro-6^thyl-5-f luoropyrimidine

To the solution obtained as the product of part (ii) wasadded a solution of potassium permanganate (23g) in water(260ml) over 2 hours, keeping the temperature of thereaction be lew 20 °C. 5N hydrochloric acid (30ml) was thenadded followed by a solution of sodium metabisulphite (14g)in water (42ml). After decolourisation of the mixture theproduct was extracted into ethyl acetate (250ml). Theorganic layer was then concentrated to give an oil. Theoil was partitioned between dichloromethane (50ml) and 2Nsodium hydroxide (105ml) and the organic layer was washedwith 5% brine (100ml). The organic layer was concentratedto give a solution of the title ccnpound which was useddirectly in the next step. (iv) 2-Chloro-6-ethvl-5-fluoropvrimidin-4 (3H) -one

To the solution obtained as the product of part (iii) wasadded water (6ml). The mixture was stirred at 80°C and4N sodium hydroxide (45ml) was added slowly over 2 hours.

At the end of this period the reaction was cooled andwashed with dichloromethane (15ml). The aqueous layer wasthen added to dichloromethane (60ml) and the pH adjusted to1 with concentrated hydrochloric acid. The organic layer BAD ORIGINAL & PLC 5 17 (S P C 7704) AP 0 0 0 2-2 3 51 was separated and the pH adjusted to 3 using concentratedaqueous ammonia solution. The precipitate of ammoniumchloride was removed by filtration and the filtrate was then concentrated to a volume of 15ml and diluted with ethyl acetate (150ml). This solution was concentrated to avolume of 30ml and the crystals of the title compound thatformed were collected by filtration and dried (8g), thencharacterised by Si-NMR and mass spectrometry. *H-NMR (dmso-d.): 6 = 7.3 (exchangeable), 2.4 (m, 2H), 1.1o (t, 3H) ppm.

Mass Snec.: ιη/e ~ 176. (v) 6-Ethyl-5-fluoropyrimidin-4 f 3H) -one

To the product of part (iv) (6g) in ethanol (60ml) wasadded sodium acetate (5.5g) and 5% palladium-on-carton(0.6g). The mixture was hydrogenated at 3 atmospherespressure for 8 hours. Ihe catalyst was removed by filtration and the filtrate was concentrated to a volume of 10ml then mixed with water (2ml) and dichlorcmethane(80ml). Toluene (32ml) was added and the solution was concentrated to a volume of 5-6ml and then mixed withfurther toluene (8ml). The crystals of the title compoundthat separated were isolated by filtration and characterised by Ηί-NMR and mass spectrometry (Yield = 3.9g).

BAD ORIGINAL p l c 5 ; z (s p c z 7 ο ς > * _ 52

Sl-NMR (dmso-d J : <S = 8.0 (s, 1H), 2.5 (m, 2H), 1.15 (t,6 3H) ppm.

Mass spec.: m/e = 142.

Precarat ion 8 4-Ethyl-5-fluoropyrimidine

A mixture of 2,4-dichloro-6-ethyl-5-fluoropyrimidine (lOg)(see Preparation 7(iii)), sodium acetate (8.83g), 5%palladium-on-charcoal (50% "wet", 2g) and methanol (30ml) washydrogenated at 50“C and 3 atmospheres pressure for 5 hours. Theresulting slurry was filtered carefully through a cellulose-basedfilter-aid, the pad was washed with further methanol (5ml) and theresulting orange filtrate was distilled at 64 *C and atmosphericpressure to provide a colourless distillate. This was partitionedbetween water (300ml) and ether (40ml) and the two BAD ORIGINAL ft PIC S 1 7 ( S P C 7 0) APOO’0^2 3 53 phases separated. The organic phase was washed with water (4 x50ml), dried over MgSO^ and the solvent was removed at roomtemperature under reduced pressure to provide the title compoundas a pale yellow liquid (2.2g).

Preparation 9 2-Chloro-4-ethy 1-5- fluoropyrimidine

CH3CH(CO2CH2CH3)2,NaH.THF -> (i)

(ii)

aq.HCl,CH3CO2H

(i) 2-Methyl-2- (2-chloro-5-fluoropyrimidin-4-yl) -1,3- propanedioc acid, diethyl ester

Sodium hydride (60% oil dispersion, 2.8g) and diethylmethylmalonate (6g) were reacted at -10’C in THF (200ml).After 30 minutes a solution of 2,4-dichloro-5- fluoropyrimidine (5g) (see Preparation 7) in THF (25ml) wasadded over 30 minutes at -10*C. The reaction was partitioned between dichlorcmethane (200ml) and water (200ml), acidified with

BAD ORIGINAL PIC 5 1’ ( 5 ?: ’ 0 <· ) » 54 acetic acid and the layers separated, ’lhe organic layerwas concentrated under reduced pressure to an oil andchromo cographed on silica gel using dichloromethane as theeluent. Ihis gave, after the combination and evaporationof appropriate fractions, the title compound (9g) which wascharacterised using ^H-NMR and mass spectrometry. ^l-NMR (CDCip: & = 8.5 (d, 1H) , 4.6 (m, 4H) , 1.9 (s, 3H) , 1.3 (t, 3H) ppm.

Mass spec.: m/e = 304. (ii) 2-OTioro-4-ethyl-5-fluorwvrimidine

The product of part (i) (3.2g) was dissolved in acetic acid(25ml) and diluted with 5N HCl (10ml). After heating themixture at 100 "C for 16 hours the mixture was cooled andpartitioned between water (30ml) and dichloromethane(45ml). The dichloromethane layer was separated, dried andconcentrated under reduced pressure to give an oil. Thetitle compound was isolated by chromatography on silica gelusing dichloromethane as the eluent. The product wascharacterised by ''"H-NMR and mass spectrometry (yield =350mg).

Si-NMR (CDC13): & = 8.4 (s, 1H), 2.9 (m, 2H), 1.3 (t, 3H)ppm.

Mass spec.: m/e = 160. BAD ORIGINAL ft

PlC 51Z (SPC 7704) ΑΡ0 0Ό22 3 55

Assessment of in vivo activity against Aspergillus fumigatus in mice

Using the general test procedure outlined on page 17 of thedescription, a group of mice was inoculated with a strain ofAspergillus fumigatus. Each mouse was then treated with the testcompound at a standard dose of 20 mg/kg b.i.d. for 5 days. Themice were then assessed on the tenth day.

Activity is based on the survival of a treated group of miceafter the death of an untreated group of mice, and also on thenumber of mice cured of the infect ion.

The results obtained in a comparative study using twocompounds described in the specific Examples of the presentapplication and two ccmpounds described in the specific Examplesof European Patent Application No. 89307920.2 (EP-A-0357241) areshewn in the following table:-

BAD ORIGINAL

Reference to Structure Survivors Cured test compound (expressed as number /’expressed as number from a test group from a test itroup of five mice) of five mice) iT\

UA ιΛ

VA

i/A

I u Ή i j 1 * -J □ c O X. · 0 0 z c 0 CM CM <υ 1 CM <y U-. « ε i u *-» o 0) ο o X <Z Cv »-H H Oj F-'» XX ex i-» » if. •H t'- e Z ε '*4 o 00 Π3 τΗ Φ •W a.<n X C <Q 1 x “C Q CL Ch ω <U O- i ω C. < 00

I

BAD ORIGINAL A

X CL 5 c x—·>. c lu ^U <£ x— 'Λ Φ -a as kJ u φ cn u “3 Φ £ Φ w tft Φ W <C > Φ -w u £ Mu CL O X lu Mu Φ Mu O ΑΡ Ο 0,0 2 2 3 5/

$u φ JO a. £ 3 □ O c ju (0 00 iu (A Φ o (0 4-» a m > CO Ή T-t •Ό οι ε m > Φ kJ lu CO Φ 3 0) <0 > cn 41 τΚ lu e «- o. o X Li Mu 41 <M O m <η >> <0 *Χ3

C

V ω Φ

X

kJ φ u □

4U Ο □

U

XJ

LO

υ φ

Mu

C

C φ φ lu

l”U φ φ r-i α. u c. · U 0 lu o c u Φ c kj □ e o c o * L* • c lu o CM φ CL p*. Φ <*> φ MU «Η · CJ ε C lu L» C C o Φ O Φ a x co cm Φ u «"U •tU SQ ·—< kJ X U 9» L- o. kJ ο. λ •κ r*» Φ kJ E C lu £ a lu r-f O Mu <0 co eg -H 41 u •Η Ο.ΓΊ Φ Φ κ c as X Ό CO CL Os oc U UJ φ a. ω ; C. < 00 'Λ “3 Ο

C φ Ό tn -σ φ l- □ (2) Although in these cases the mice were not "cured" as defined above, progression of the infection wassignificantly reduced.

BAD ORIGINAL

Claims

P L C 5 ! 7 ( S p C 7 7 3 4 ) CI ATMS Λ compound ot' the formula:-

(I) or a pharmaceutically acceptable salt thereof, wherein R isphenyl substituted by 1 to 3 substituents each independentlyselected from halo, -CF^ and -OCF^; R1 is C1~C4 alkyl; R2 is H or C^-C alkyl; 2. 2. 3. 3. X is CH or N; and Y is F or Cl. A compound as claimed in claim 1 wherein R is phenylsubstituted by 1 or 2 halo substituents. A compound as claimed in claim 2 wherein R is phenylsubstituted by 1 or 2 substituents 'each independently selected from fluoro and chloro. A compound as claimed in claim 3 wherein R is 2-fluorophenyl,4-fluorophenyl, 2,4-difluorophenyl, 2-chlorophenyl or 2,4-dichlorophenyl. bad original

PL C 5 17 (SPC 7 7 0 4 ) APο Οΰ2 2 3 59
5. A compound as claimed in claim 4 wherein R is 2-fluorophenyl, 2,4-difluorophenyl, 2-chlorophenyl or 2,4-dichlorophenyl.
6. A compound as claimed in any preceding claim wherein R1 ismethyl.
7. A compound as claimed in any preceding claim wherein R is Hor methyl. . . 2
8. A compound as claimed in claim 7 wherein R is H.
9. A compound as claimed in any preceding claim wherein X is N.
10. A ccmpcund as claimed in any preceding claim wherein Y is F.
11. A compound as claimed in any preceding claim wherein R is H and which has the 2R,3S- configuration, that is

BAD ORIGINAL PL C 5 17 (SPC 7704 ) 60 12. 2R, 3S-2-(2,4-di f luorophenyl) -3-( 3-f luoropyridin-4-yl) -1-( 1H- 1,2,4-triazol-l-yl)butan-2-ol, 2R, 3S-2-(2-chlorophcnyl) -3-(3-fluoropyridin-4-yl) -1-(1H- 1,2,4-triazol-l-yl)butan-2-ol, 2R, 3S-2- (2-fluorophenyl) -3- (3-f luoropyridin-4-yl) -1- (1H- 1,2,4-triazol-l-yl)butan-2-ol, 2R, 3S-2- (2,4-di fluorophenyl) -3- (5-fluoropyrimidin-4-yl) -1-(1H-1,2,4-triazol-l-yl)butan-2-ol, or 2R, 3S-2- (2,4-dichlorophenyl) -3-(5-fluoropyrimidin-4-yl) -1- (1H-1,2,4-triazol-l-yl) butan-2-ol: or a pharmaceutically acceptable salt thereof.
13. A pharmaceutical composition comprising a compound of theformula (I) or a pharmaceutically acceptable salt thereof, asclaimed in any preceding claim, together with a pharmaceutically acceptable diluent or carrier.
14. A pharmaceutical composition as claimed in claim 13 in whichthe compound of the formula (I) is ccmplexed with ahydroxyalkyl derivative of a cyclodextrin.
15. A pharmaceutical composition as claimed in claim 14 whereinsaid hydroxyalkyl derivative is a hydroxypropyl derivativeand said cyclodextrin is alpha- or beta-cyclodextrin. r BAD ORIGIN*1- P L C Sir (SPC 7 7 0 4 ) AP000223 61
16. A compound of the formula (I) , or a pharmaceuticallyacceptable salt or composition thereof, as claimed in any oneof claims 1 to 12 and any one of claims 13 to 15 respectively, for use as a medicament.
17. The use of a compound of the formula (I), or of apharmaceutically acceptable salt or composition thereof, asclaimed in any one of claims 1 to 12 and any one of claims 13to 15 respectively, for the manufacture of an antifungalagent.
18. A compound of the formula:-

1 2 wherein R, R , R , X and Y are as defined in claim 1 and Z isa leaving group.
19. A compound of the formula (VI) as claimed in claim 18 whereinZ is chloro, bromo or alkanesulphony loxy. BAD ORIGINAL Sir ( S P C 7 7 0 4 ) 62
20. A compound of the formula:- 1

(XI) 12 wherein R, R , R and Y are as defined in claim 1 and2 3 Z and Z are each independently selected from H and a group that may be selectively removed by reduction, with the2 3 proviso that Z and Z cannot both be H.
21. A compound as claimed in claim 20 wherein the group that maybe selectively removed by reduction is halo. 2 3
22. A compound as claimed in claim 21 wherein Z is chloro and Z is H. 23. 4-Ethyl-5-f luoropyr imidine or 4-chloro-6-ethyl-5- f luoropyr imidine.
24. A method of treatment of an animal (including a human being)to cure or prevent a fungal infection comprising treatingsaid animal with an effective amount of a compound of theformula (I), or with a pharmaceutically acceptable salt orcomposition thereof, as claimed in any one of claims 1 to 12and any one of claims 13 to 15 respectively.

BAD ORIGINAL PIC 5,/ AP Ο 0-0 2 2 3 ¢,3 tTOCESS CLAIMS '-Γ'- A process Cor the preparation of a compound of the formula: -

or a pharmaceutically acceptable salt thereof, wherein R isphenyl substituted by 1 to 3 substituents each independentlyselected from halo, -CF3 and -OCF^; R1 is Cj-C4 alkyl; R2 is H or ^-<4 alkyl; X is CH or N; and Y is F or Cl, comprising reacting a 1’-deprotonated form of a compound of the formula :- .,1

( N ) <&Sf« BAD original 44» i n wherein R , R , X and Y are as 'let mod above, with a corrrourxi of the formula:- (III) wherein R is as defined above: said process being optionallyfollowed by conversion of the compound of the formula (I) toa pharmaceutically acceptable salt thereof. J20 A process as claimed in claim 1 in which said deprotonated form is a lithium, sodium or potassium salt of the compoundof the formula (II). 7 A process for the preparation of a compound of the formula:-

(i) BAD ORIGINAL £ AP 0 0 0 2 2 3 or a pharmaceutically acceptable salt tlaereof, therein R, r\ Ί R“, >; and Y are as defined in claim 1, comprising re.icfirxj a coir;x^::>! of the formula:-

το#. 12 . ...wherein R, R , R , X and Y are as previously defined in thisclaim and Z is a leaving group, either with a base salt oflH-l,2,4-triazole or with lH-l,2,4-triazole in the presenceof an additional base: said process being optionally followedby conversion of the compound of the formula (I) into apharmaceutically acceptable salt thereof. A process as claimed in claim 3 in which Z is chloro, bromoor alkanesulphonylcxy. A process as claimed in claim 3 in which a compound of theformula (IV) is used. A process as claimed in any one of claims 3 to 5 in whichsaid base salt of 111-1,2,4-triazole is either a sodium,potassium or a tetra-n-butylammonium salt. bad original I ' ( S ? C 7 z’ 0 A ) 31 >f. A process as claimed in any one of claims 3 said additional base is sodium or potassium carbonate. to 5 in which 32X- Λ process for the preparation of a compound of the formula:-

or a pharmaceutically acceptable salt thereof, 2 . . . . R and Y are as defined in claim 1, comprisingcompound of the formula:- wherein R, R1,reduction of a

Zb XT. Stir. 37 JrfT. 33 x. βΡ0Ο0223 47 1 ? .wherein R, R , R and Y are an previously defined in this -> i claim and Z and Z arc each indopondentiy selected from H and a tjreiip tint may be selectively removed by reduction, 2 3 with the proviso that Z ard Z cannot both be H: saidprocess being optionally followed by conversion of thecompound of the formula (IA) into a pharmaceuticallyacceptable salt thereof. 2 . A process as claimed in claim 8 in which Z is a group thatmay be selectively removed by reduction and Z^ is H. A process as claimed in claim 8 or 9 in which said group thatmay be selectively removed by reduction is halo. A process as claimed in claim 10 in which said halo group is chloro. A process as claimed in any one of claims 8 to 11 in whichsaid reduction is carried out by hydrogenolysis using apalladium-on-charcoal catalyst. A process as claimed in claim 12 in which sodium acetate isalso present. Λ process as claimed in any preceding claim whereinR is phenyl substituted by 1 or 2 halo substituents. BAD ORIGINAL & .- . ; ’ ’ η . : * 3Ί >5. Λ process as cLaLined in claim 14 wherein R is phenylsubstituted by I or 2 substituents each independently selected from fluoro and chloro. l+° -hi. A process as claimed in claim 15 wherein R is 2-fluorophenyl·,4-fluorophenyl, 2,4-difluorophenyl, 2-chlorophenyl or 2.4- dichlorophenyl; R1 is methyl; andR2 is H. If r -i7*. A process as claimed in claim 16 wherein R is 2-fluorophenyl, 2.4- difluorophenyl, 2-chlorophenyl or 2,4-dichlorophenyl; R1 is methyl; 2 . R is H; and Y is F. iff. A process as claimed in any preceding claim which is used to2 prepare a compound of the formula (I) wherein R is H andwhich has the 2R,3S- configuration, that is

bad origin*1- ΑΡ ο Ο 0 2 2 3 Λ prvM.x-k; us cluim»·,! in .liin 13 which is iuuil to prep.tro2k, :: :-2- (2, 3 -d i! Itiorot h· ny 1) - 5- (3-t' luoroj >y r i-1 :n - ; -y L) -1-( 1H-1,3, ;-tr ias.i .1-1 -y 1) Inhirtl’.-o I , 2R, :::--2-(2-chlorophony I )-3-( 3-L luoropyr i.din-4-y L) -1-(11-h, 2,4-triazol-L-yl)butan-2-oL, 2R,3S-2-(2-fluoropheryl) -3-(3-fluoropyridin-4-yl) -1-(1H- 1,2,4-triazol-l-yl)butan-2-ol, 2R,3S-2-(2,4-difluorophenyI)-3-(5-fluoropyrimidin-4-yl) -1-(1H-1,2,4-triazol-l-yl)butan-2-ol, or 2R, 3S-2- (2,4-dichlorophenyl) -3- (5-fluoropyrimidin-4-yl) - 1-(1H-1,2,4-triazol-l-yl)butan-2-ol: or a pharmaceutically acceptable salt thereof. 2+1* 2Π. A process for the preparation of a pharmaceutical compositioncomprising combining a compound of the formula (I), or apharmaceutically acceptable salt thereof, which has beenprepared by a process as claimed in any preceding claim,with a pharmaceutically acceptable diluent or carrier. . A process as claimed in claim 20 in which the compound of theformula (I) is complexed with a hydroxyalkyl derivative of acyclodextrin. *+<=> &r. A process as claimed in claim derivative is a hydroxypropyl derivative and said I cyclodcxtrin is alpha- or beta-cyclodextrrincv DATED this 28th day of January 1991

1——PATENT AGENT FOR THE APPLICANTS

BAD ORIGINAL