WO2003018537A1 - Antiviral protease inhibitors - Google Patents

Abstract

Compounds of the formula (I) wherein: X and Y are hydroxy or H. A' and A'' are terminal amine functions such valinamide or indanolamine. Z' and Z'' along with the adjacent ()n groups are independently alkylaryl have utility as HIV aspartyl protease inhibitors with particularly good activity in the presence of human serum.

Description

ANTIVIRAL PROTEASE INHIBITORS

Technical field

This invention relates to novel protease inhibitors and in particular to inhibitors of the aspartate protease possessed by certain retroviruses, notably HIV. The invention further relates to the use of such protease inhibitors in the treatment of conditions caused by retroviruses and in the preparation of medicaments for this purpose. The invention also relates to novel synthesis methodology for the facile preparation of protease inhibitors and similar chemical structures.

Background of the Invention

Many biological processes are dependent upon the accurate enzymatic abscission of polypeptides at particular amino acid sequences. An example of such an operation is the post-translational processing of the gag and gag-pol gene products of the human immunodeficiency virus HIV to allow for the organisation of core structural proteins and release of viral enzymes. The enzyme responsible for this task, HIV protease, is a virally encoded homodimeric protease belonging to the aspartic protease family of enzymes. The human renin and pepsin enzymes also belong to this family. Inhibition of the HIV protease in cell culture prevents viral maturation and replication and thus this enzyme represents an attractive target for antiviral therapy against HTv^" in humans.

WO 94/13629 describes HIV protease inhibitors based on mannitol esters. Our copending application no. WO98/45330 describes sugar based protease inhibitors based on a mannitol skeleton, but wherein the PI groups are ethers. While the latter compounds are fairly potent and very easy to synthesise, the nature of the HUIV replicative process with its extremely error prone transcription and rapid generation of drug escape mutants means that further more potent compounds with DMPK properties amenable to good patient compliance are required in the HIV armamentarium.

Brief description of the invention

A first aspect of the invention provides novel compounds of the formula I:

wherein:

A' and A" are independently the same or different group of the formula II:

FT

R

1 M

R" II wherein:

R' is H, CH₃, C(CH₃)₂, -OR^a, -N(R^a)₂, -N(R^a)OR^a or -DP

R'" is H or CH₃; R^a is H, C,-C₃ alkyl;

D is a bond, C_l-3 alkylene, -C(=O , -S(O)- or -S(O)₂- ; P is an optionally substituted, mono or bicyclic carbo- or heterocycle;

R" is H, any of the sidechains found in the natural amino acids, Cι-_aalkyl, carboxacetamide, or a group (CH₂)_nDP;

M is a bond or -C(=O)N(R'")-;

Q is absent, a bond, -CH(OH)- or -CH₂-; or R" together with Q , M and R' define an optionally substituted 5 or 6 membered carbo- or heterocyclic ring which is optionally fused with a further 5 or 6 membered carbo- or heterocyclic ring;

X is H, OH, OCH₃;

Y is H, OH, OCH₃, Z' and Z" are independently an optionally substituted mono-or bicyclic carbo-or heterocycle;

() is a methylene group; n and m are independently 0, 1 or 2; and pharmaceutically acceptable salts and prodrugs thereof.

Compounds of the formula I are active inhibitors of aspartyl proteases, such as those from HIV. Further aspects of the invention thus provide: - a pharmaceutical formulation comprising a compound of the formula I in admixture with a pharmaceutical acceptable carrier or diluent;

- the use of a compound of the formula I in the manufacture of a medicament for the prophylaxis or treatment of conditions, such as AIDS caused by retroviruses, such as HIV; and

- a method for treating conditions caused by retroviruses, especially AIDS in humans, comprising administering a compound of formula I to a subject afflicted with said condition.

The compounds have a relatively low molecular weight and should therefore provide good oral absorption properties in mammals.

For ease of synthesis it is generally preferred that the terminal amines A' and A" are identical. However, although the target enzyme is a symmetric dimer, thus implying a tight interaction with symmetric compounds, it can in some circumstances be advantageous for resistance or pharmacokinetic reasons etc to have asymmetric terminal amines. Where is it is desired to have an asymmetric compound, that is where the A' and A" groups differ, it will generally be most convenient to add the respective A' and A" groups sequentially.

This may be achieved by the use of different protecting/leaving groups on the diepoxydicarboxylic acid in conjunction with manipulation of the reagent concentrations, reaction conditions, speed of addition etc to provide a monoprotected, monoamidated diepoxycarboxylic acid followed by displacement of the remaining protecting/leaving group with the second amine.

Alternatively the differential terminal amination can be achieved with a solid phase synthesis where the unprotected or partially protected diepoxydicarboxy acid is secured to a solid phase substrate, such as polymer beads of which many are known in solid phase chemistry, for instance Merrifield resin, in conjunction with an appropriate linker structure. Immobilization of the diepoxydicarboxylic acid in this fashion will only allow amination of one carboxy group. Cleavage from the resin/linker releases the other carboxy terminal which is subsequently amidated with the second terminal amine using conventional peptide chemistry.

Preparation of compounds of Formula I in which X is hydrogen can be carried out by deoxygenation analogously to Examples 2 and 26 of WO 9845330. Preferably X and Y are not both H.

A preferred stereochemistry is the 2R, 3R, 4R, 5R form, the synthesis of which is shown in the examples. A further preferred stereochemistry is

which is readily accessible:

metathesis reaction

cis hydroxylation

Carbocyclic groups for R' as -DP and/or Z'/Z" and/or the optional substituents thereto may be saturated, unsaturated or aromatic and include monocyclic rings such as phenyl, cyclohexenyl, cyclopentenyl, cyclohexanyl, cyclopentanyl, or bicyclic rings such as indanyl, napthyl and the like.

Heterocyclic groups for R' as -DP and/or Z'/Z" and/or the optional substituents thereto may be saturated, unsaturated or aromatic and have 1 to 4 hetero atoms including monocyclic rings such as furyl, thienyl, pyranyl, pyrrolyl, pyrrolinyl, pyrrolidinyl, pyrazolyl, pyrazolinyl, pyrazolidinyl, imidazolyl, imidazolinyl, imidazolidinyl, pyridyl, piperidinyl, pyrazinyl, piperazinyl, pyrimidinyl, pyridazinyl, oxazolyl, oxazolidinyl, isoxazolyl, isoxazolidinyl, morpholinyl, thiazolyl, thiazolidinyl, isothiazolyl, isothiazolidinyl, and the like or bicyclic rings especially of the above fused to a phenyl ring such as indolyl, quinolinyl, isoquinolinyl, benzimidazolyl, benzothiazolyl, benzoxazolyl, benzothienyl etc. The carbo or heterocyclic ring may be bonded via a carbon or via a hetero atom, typically a nitrogen atom, such as N-piperidyl, N-morpholinyl etc.

Preferred embodiments of Formula II for the A7A" groups of the compounds of the invention include those of the formula Ha or lie:

Ila

where n is 1 or 2 and R' is alkyloxy, preferably methyloxy, or those where n is 0 and R' is methyl.

Other preferred groups of formula II include lib below

Mb

An alternative preferred configuration for the A'/A" groups of the compounds of the invention includes groups of the formula lie:

^R" lie

where Q is a bond, methylene or-C(OH)- and R' is -OR^a, -N(R^a) ₂, -NR^a OR^a, where R^a is H or C₁-C₃ alkyl, or a carbo- or heterocyclic group including N-piperidine, N-morpholine, N- piperazine, pyrrolyl, imidazolyl, pyrazolyl, pyridyl, pyrimidyl, pyrazinyl etc .

A favoured subset of compounds within formula lie has the formula lid:

where R^d is hydrogen or methyl (that is a valyl or isoleucyl side chain) and R^e is

where X is methylene, O, S, S=O, S(=O)₂ or NH or R^e is -N(CH₃)₂, -NHOH, -NHOMe, - NHOEt, -NMeOH, -NMeOMe etc.

In each of formulae Ila, lib and He, R" is hydrogen, methyl, ethyl, isopropyl, cycloalkyl such as cyclopropyl, cyclobutyl or cyclohexyl, cycloalkenyl, benzyl, carboxacetamide or 4- imidazolylmethy, any of which may be substituted as defined above. Preferred R" groups include the side chains found in the natural amino acids, especially those of leucine, asparagine, histidine or proline. The most preferred R" groups for formula Ila, lib, lie and lid are the tertiary leucyl, isoleucyl and especially the valyl side chain.

R' will vary depending on the nature of Q and/or M, if present, and may for instance be selected from hydrogen, methyl, ethyl, isopropyl, R^e as defined above, valinol, a heterocycle such as pyridyl, thiazole, oxazole, imidazole, N-piperidine, N-morpholine, N-piperazine, pyrrolyl, imidazolyl, pyrazolyl, pyrimidyl, pyrazinyl, any of which R' groups may be substituted as defined for Z'/Z" below.

Further favoured A'/A" groups include those of formula II where R", Q, M and R' together define an optionally substituted 5 or 6 membered carbo- or heterocylic ring. A preferred group within this definition include groups within formula III:

where

R'" is as defined above,

R¹ is H, NR⁴R⁴, C(=O)R³, CR³R⁴ or a monocyclic, optionally substituted carbo- or heterocycle;

R² s OH, or together with R' is =O, or if R' is NR⁴R\ then R' may be H; R³ s H, halo, C₁-C₃ alkyl, OR⁵, NR⁴R⁴; R⁴ s H, Cι-C₃ alkyl; R⁵ s H or a pharmaceutically acceptable ester; R⁶ s OH, NH₂, carbamoyl or carboxy;

R⁷ s hydrogen, C₁-C₄ straight or branched alkyl or together with the adjacent carbon atoms forms a fused phenyl or heteroaromatic ring;

Preferred groups of formula III include aminoindanol and 1 -amino-azaindan-2-ol, that is moieties of the formulae:

Optional substitutents for the carbo- or heterocyclic moiety of Z'/Z" or A'/A" include one to three substituents such as halo, amino, mercapto, oxo, nitro, NHCι-C₆ alkyl, N(Cι-C₆ alkyl)₂, Cι-C₆ alkyl, C C₆ alkenyl, Cι-C₆ alkynyl, C C₆ alkanoyl, Cι-C₆ alkoxy, thioCι-C₆ alkyl, thioC C₆ alkoxy, hydroxy, hydroxyCι-C₆ alkyl, haloC C₆ alkyl, aminoC C₆ alkyl, C C₆ alkyl, cyano, carboxyl, carbalkoxy, carboxamide, carbamoyl, sulfonylamide, benzyloxy, morpholyl-C_rC₆ alkyloxy, a monocyclic carbo- or heterocycle, as defined above, a carbo- or heterocyclic group spaced by alkyl, such as C_]-3 alkylaryl, etc.

The preferred definitions for -()_mZ' and -{)_m Z" include benzyl and especially phenethyl each unsubstituted or substituted with 1 , 2 or 3 substituents, especially 1 selected from fluoro, chloro, hydroxy, amino, -NH(C_1-6 alkyl), -N(Cι_-6 alkyl)₂, -NPh(C,.₆ alkyl), -NHPh, methoxy, cyano, hydroxymethyl, aminomethyl, alkylsulfonyl, carbamoyl, moφholinethoxy, benzyloxy, benzylamide etc. Other possibilities exhibiting the great freedom in this area are shown in WO 98/45330. It will be apparent that the substituent to Z' and/or Z" may comprise a ring structure (which substituent ring structure is itself substituted as defined herein) such as phenyl or a 5 or 6 membered heterocycle containing one or two hetero atoms such as thiophene, pyridine etc. The preparation of useful heterocyclic substituents for Z' and Z" phenyl are described in Tetrahedron Letters 1997 6359-6359-6367 and J Org Chem 62 (1997) 1264 and 6066, including N-morpholine, N-piperidine, N-piperazine, N'-methyl- N-piperazine, N-pyrrolidone, N-pyrrolidine and the like.

Such substituents may be in the meta but especially the ortho or para positions of Z'/Z", with small groups such as fluoro being favoured for the ortho and meta and with extensive freedom for larger groups in the para such as (optionally substituted) cyclic substituents, including the N-bonded rings in the immediately preceding paragraph. The whole Z' and Z" group or their respective carbo-or heterocyclic moiety may be different but for ease of synthesis it is convenient if they are the same. Appropriate pharmaceutically acceptable salts, both for A'/A" as a free acid or for other charged groups along the compound of formula I include salts of organic carboxylic acids such as acetic, lactic, gluconic, citric, tartaric, maleic, malic, pantothenic, isethionic, oxalic, lactobionic, and succinic acids, organic sulfonic acids such as methanesulfonic acid, ethanesulfonic acid, benzenesulfonic acid, p-chlorobenzenesulfonic acid and p- toluenesulfonic acid; and inorganic acids such as hydrochloric, hydroiodic, sulfuric, phosphoric and sulfamic acids.

Prodrugs of the invention are derivatives that release a compound of formula I in vivo, generally by hydrolysis or other metabolic interaction in the intestine, liver or plasma. Typical prodrugs are esters formed on free hydroxy groups in the compounds. Appropriate pharmaceutically acceptable esters include Cι-C₂₂ fatty acid esters, where the fatty acid is unsaturated, monounsaturated or multiply unsaturated. Saturated fatty acid esters include short chains such as acetyl or butyryl or long chain such as stearoyl.

Unsaturated fatty acid esters are preferably in the ω-9 series, such as palmitoleic or linolenic esters. Other esters include Ci-Cβ alkylaryl esters such as benzyl or methylpyridyl or esters of phosphoric acid, such as monophosphate. Alternative esters include the corresponding fatty acid or alkylaryl carbonate, carbamate or sulphonic esters.

Particularly favoured prodrugs include those described in WO99/41275, especially when A' and A" comprise an hydroxy function such as aminoindanol.

Intermediate compounds of formula III can be prepared by the following reaction scheme:

Aminohydroxylation

l-amino-azaindan-2-ol P-2 filling groups can be prepared analogously to J Med Chem 1991 1228-1230:

In treating conditions caused by retroviruses, the compounds of formula I are preferably administered in an amount to achieve a plasma level of around 10 to 1000 nM and more preferably 100 to 500 nM. This corresponds to a dosage rate, depending on the bioavailability of the formulation of the order 0.001 to 100 mg/kg/day, preferably 10 to 50 mg/kg/day.

In keeping with the usual practice with HIV inhibitors it is advantageous to co-administer one to three additional antivirals, such as AZT, ddl, ddC, D4T, ritonavir, saquinavir, indinavir, nelfinavir, efavirenz, delavirdine, nevirapine, trovirdine, PFA, H2G etc. The molar ratio for such co-administered antivirals will generally be chosen to reflect the respective EC₅₀ performances of the antiviral. Molar ratios of 25:1 to 1 :25, relative to the compound of formula I will often be convenient.

While it is possible for the active agent to be administered alone, it is preferable to present it as part of a pharmaceutical formulation. Such a formulation will comprise the above defined active agent together with one or more acceptable carriers and optionally other therapeutic ingredients. The carrier(s) must be acceptable in the sense of being compatible with the other ingredients of the formulation and not deleterious to the recipient.

The formulations include those suitable for oral, rectal, nasal, topical (including buccal and sublingual), vaginal or parenteral (including subcutaneous, intramuscular, intravenous and intradermal) administration. The formulations may conveniently be presented in unit dosage form, e.g. tablets and sustained release capsules, and may be prepared by any methods well known in the art of pharmacy.

Such methods include the step of bringing into association the above defined active agent with the carrier. In general, the formulations are prepared by uniformly and intimately bringing into association the active agent with liquid carriers or finely divided solid carriers or both, and then if necessary shaping the product.

Formulations for oral administration in the present invention may be presented as discrete units such as capsules, cachets or tablets each containing a predetermined amount of the active agent; as a powder or granules; as a solution or a suspension of the active agent in an aqueous liquid or a non-aqueous liquid; or as an oil-in-water liquid emulsion or a water in oil liquid emulsion and as a bolus etc. With regard to compositions for oral administration (e.g. tablets and capsules), the term suitable carrier includes vehicles such as common excipients e.g. binding agents, for example syrup, acacia, gelatin, sorbitol, tragacanth, polyvinylpyrrolidone (Povidone), methylcellulose, ethylcellulose, sodium carboxymethylcellulose, hydroxypropylmethylcellulose, sucrose and starch; fillers and carriers, for example corn starch, gelatin, lactose, sucrose, microcrystalline cellulose, kaolin, mannitol, dicalcium phosphate, sodium chloride and alginic acid; and lubricants such as magnesium stearate and other metallic stearates, stearic acid, silicone fluid, talc waxes, oils and colloidal silica. Flavouring agents such as peppermint, oil of wintergreen, cherry flavouring or the like can also be used. It may be desirable to add a colouring agent to make the dosage form readily identifiable. Tablets may also be coated by methods well known in the art.

A tablet may be made by compression or moulding, optionally with one or more accessory ingredients. Compressed tablets may be prepared by compressing in a suitable machine the active agent in a free flowing form such as a powder or granules, optionally mixed with a binder, lubricant, inert diluent, preservative, surface-active or dispersing agent. Moulded tablets may be made by moulding in a suitable machine a mixture of the powdered compound moistened with an inert liquid diluent. The tablets may be optionally be coated or scored and may be formulated so as to provide slow or controlled release of the active agent.

Formulations suitable for topical administration include lozenges comprising the active agent in a flavoured base, usually sucrose and acacia or tragacanth; pastilles comprising the active agent in an inert base such as gelatin and glycerin, or sucrose and acacia; and mouthwashes comprising the active agent in a suitable liquid carrier.

Formulations suitable for topical administration to the skin may be presented as ointments, creams, gels, and pastes comprising the active agent and a pharmaceutically active carrier. An exemplary topical delivery system is a transdermal patch containing the active agent.

Formulations for rectal or vaginal administration may be presented as a suppository or pessary with a suitable base comprising, for example, cocoa butter or a salicylate. Other vaginal preparations can be presented as tampons, creams, gels, pastes, foams or spray formulations containing, in addition to the active agent, such carriers as are known in the art to be appropriate. Formulations suitable for nasal administration wherein the carrier is a solid include a coarse powder having a particle size, for example, in the range 20 to 500 microns which is administered in the manner in which snuff is taken, i.e. by rapid inhalation from a container of the powder held up close to the nose. Suitable formulations wherein the carrier is a liquid for administration, for example, as a nasal spray or as nasal drops, include aqueous or oily solutions of the active agent.

Formulations suitable for parenteral administration include aqueous and non-aqueous sterile injection solutions which may contain antioxidants, buffers, bacteriostats and solutes which render the formulation isotonic with the blood of the intended recipient; and aqueous and non-aqueous sterile suspensions which may include suspending agents and thickening agents. The formulations may be presented in unit-dose or multi-dose containers, for example sealed ampoules and vials, and may be stored in freeze-dried (lyophilized) condition requiring only the addition of the sterile liquid carrier, for example water for injection, immediately prior to use. Extemporaneous injection solutions and suspensions may be prepared from sterile powders, granules, and tablets of the kind previously described.

Detailed Description of Embodiments of the Invention.

The invention will now be further illustrated by reference to the following non-limiting Examples, Figures 1 and 2 which depict synthesis schemes for preparing intermediates, and Figure 3 which is a synthesis scheme for compounds of the invention.

General

Reference numerals refer to the compounds in Figures 1-3.

All glassware was dried over an open flame before use in connection with an inert atmosphere. Concentrations were performed under reduced pressure at <40 °C (bath temperature). Thin layer chromatography was performed using silica gel 60 F-254 plates with detection by UV and charring with 8% sulphuric acid and/or ninhydrin in ethanol. Silica gel (0.040-0.063 mm) was used for column chromatography. Me₄Si (0.0 ppm) was used as an internal standard in Η NMR and Me₄Si or CDC1₃ (77.0 ppm) were used in ¹³C NMR. Melting points are uncorrected. Unless stated otherwise, all materials were obtained from commercial suppliers and used without further purification. Yields are not optimised.

Intermediates

(4R.5S)-Indano[l .2-ttl-oxazolidin-2-one (1).

(lR,25)-(+)-cw-l-Amino-2-indanol (18.4 g, 0.125 mol) was dissolved in CH₂C1₂ (1350 mL) under a nitrogen atmosphere. The temperature was lowered to 0 °C before addition of triphosgene (14.8 g, 49.9 mmol, 0.4 equiv.) and NEt₃ (35.0 mL, 0.251 mol, 2.0 equiv.). Stirring was continued at 0 °C for 1.5 h, when TLC (R 0.39, CHCl₃-MeOH 15:1) showed completion of the reaction. The mixture was concentrated to 675 mL and washed with NH₄C1 (lx) and water (2x) and the combined aqueous layers were extracted with EtOAc (2x). The organic layers were combined with the mixture from above, dried (MgSO₄) and concentrated to give 1 (20.9 g, 0.119 mol, 95%) as white crystals: [D]_D +71 (c 0.65, EtOAc); mp 200-201 °C; Η NMR (300 MHz, CDC1₃ and CD₃OD) D 3.32 (dd, 1 H, J= 1.76, 18.0 Hz), 3.43 (dd, 1 H, J= 6.15, 18.0), 3.74 (s, 1 H), 5.18 (d, 1 H, J= 7.47 Hz) and 5.42 (ddd, 1 H, J= 1.76, 6.15, 7.47); ¹³C NMR (67 MHz, CDC1₃ and CD₃OD) D 38.5, 61.0, 80.5, 124.6, 125.3, 127.6, 129.1, 139.5, 140.1 and 159.9. Anal Calcd for C₁₀H₉NO₂: C, 68.6; H, 5.2; N, 8.0. Found: C, 68.4; H, 5.2; N, 8.0.

N-π-Oxo-4-phenylbutyn-(4R.5S -indano[1.2-tt]-oxazolidin-2-one (^"21

To a stirred solution of 4-phenylbutyric acid (35.0 g, 0.214 mol) and SOCl₂ (98 mL, 1.34 mol, 6.3 equiv.) was added pyridine (0.64 mL, 7.9 mmol, 0.04 equiv.). The solution was stirred at room temperature for 30 min and at 40 °C for an additional 1 h before it was concentrated to give 4-phenylbutanoyl chloride (38.3 g, 0.210 mol, 98%) as a yellow oil, which was used in the next step without further purification: Η ΝMR (300 MHz, CDCI₃) D 2.03 (dt, 2 H, J= 7.6, 7.3 Hz), 2.67 (t, 2 H, J= 7.6 Hz), 2.88 (t, 2 H, J= 7.3 Hz) and 7.15- 7.32 (m, 5 H); ¹³C ΝMR (67 MHz, CDC1₃) D 26.4, 34.0, 46.1, 126.2, 128.3, 128.4, 140.2 and 173.4. ΝaH (1.61 g, 67.1 mmol, 1.1 equiv.) was suspended in DMF (150 mL) under an argon atmosphere and the temperature was lowered to 0 °C. A dropping funnel was charged with the chiral auxiliary 1 (10.7 g, 61.2 mmol) in DMF (90 mL), a second dropping funnel was charged with 4-phenylbutanoyl chloride (13.4 g, 73.4 mmol, 1.2 equiv.) in DMF (90 mL) and the two solutions were then added simultaneously to the stirred NaH suspension during 30 min. Stirring was continued at 0 °C for an additional 3 h (R_/0.56, toluene-EtOAc 10:1), before slow addition of MeOH and H₂0. The layers were separated after dilution with toluene and H₂0. The organic layer was washed with H₂0 (2x), followed by extraction of the combined aqueous layers with toluene (lx). Finally, the combined organic layers were dried (Na₂S0₄) and concentrated. Purification by column chromatography (toluene; toluene- EtOAc 40: 1 ) gave 2 (14.3 g, 44.5 mmol, 73%) as a colourless syrup: [D]²⁰ _D -209 (c 0.948, CHC1₃); Η NMR (300 MHz, CDC1₃) □ 1.98-2.08 (m, 2 H), 2.70 (t, 2 H, J= 7.69), 3.00 (ddd, 2 H, J= 1.76, 7.47, 9.23 Hz), 3.37 (d, 2 H, J= 3.52 Hz), 5.26 (ddd, 1 H, J= 3.08, 4.40, 7.47 Hz), 5.91 (d, 1 H, J = 7.03 Hz), 7.16-7.37 (m, 8 H) and 7.62 (d, 1 H, J= 7.03 Hz); ¹³C NMR (67 MHz, CDC1₃) D 26.0, 34.8, 35.2, 38.0, 63.0, 78.0, 125.2, 126.0, 127.2, 128.2, 128.4, 128.5, 129.8, 139.2, 139.5, 141.6, 153.0 and 173.4. Anal Calcd for C₂₀H₁₉NO₃: C, 74.8; H, 6.0; N, 4.4. Found: C, 74.9; H, 6.0; N, 4.3.

N-[f2S)-2-(Hvdroxymethvπ-l-oxo-4-phenylbutyl1-(4R.5S)-indanori.2-^]-oxazolidin-2-one

TiCl (3.6 mL, 32.8 mmol, 1.05 equiv.) was added to a stirred solution of compound 2 (10.0 g, 31.1 mmol) in CH₂C1₂ (525 mL), under an argon atmosphere, with the temperature kept at 0 °C. A yellow precipitate was formed within 30 min, DIEA (5.5 mL, 31.9 mmol, 1.0 equiv.) was added (the colour changed from yellow to red), and stirring was continued at 0 °C for 1 h. Trioxane (2.08 g, 23.1 mmol, 0.74 equiv.) and TiCl₄ (4.5 mL, 40.9 mmol, 1.3 equiv.) were added, and stirring was continued until TLC (R_/0.15, toluene- EtOAc 10:1) showed complete reaction (3 h). During this time the colour of the reaction changed from dark red to light brown. Neutralisation (pH=7) of the cold reaction was performed by adding sat. aqueous NaHC0₃ (500 L). The layers were separated and the aqueous layer was extracted with CH₂C1₂ (4x). The combined organic layers were dried (Na₂S0₄) and concentrated. The crude product was immediately purified using column chromatography (toluene; toluene-EtOAc 10:1) to give alcohol 3 (8.1 g, 23.0 mmol, 74%) as a colourless syrup, which solidified upon standing: [π]²⁰ _D -174 (c 1.19, CHC1₃); Η NMR (300 MHz, CDC1₃) D 1.58 (s, 1 H), 1.90- 2.02 (m, 1 H), 2.15-2.24 ( , 1 H), 2.63-2.84 (m, 2 H), 3.36 (d, 2 H, J= 3.52 Hz), 3.84 (m, 2 H), 3.94-4.02 (m, 1 H), 5.14-5.19 (m, 1 H), 5.72 (d, 1 H, J= 7.03 Hz), 7.18-7.36 (m, 8 H) and 7.58 (d, 1 H, J= 7.47 Hz); ¹³C NMR (67 MHz, CDC1₃) Q29.9, 33.8, 37.7, 45.2, 63.2, 63.8, 78.2, 125.1, 126.0, 127.1, 128.2, 128.3, 128.6, 129.8, 138.9, 139.3, 141.2, 152.9 and 175.9. Anal Calcd for C₂ιH₂ιN0₄: C, 71.8; H, 6.0; N, 4.0. Found: C, 71.6; H, 6.1; N, 4.0.

N- (2S)-2-(^fBenzyloxymethyl)-l-oxo-4-phenylbutyl1-t4R.5S)-indano[1.2-ιsπ-oxazolidin-2-one

To a stirred solution of alcohol 3 (18.2 g, 51.7 mmol) and benzyl 2,2,2-trichloroacetimidate (11.6 mL, 62.4 mmol, 1.2 equiv.) in 1,4-dioxane (900 mL) was added TMS-OTf (0.9 mL, 4.96 mmol, 0.1 equiv.) dropwise under argon. Within 1 h the colour changed to brown- yellow and TLC (R 0.47, toluene-EtOAc 10: 1) showed completion of the reaction. The reaction mixture was filtered through a pad of SiO₂/ΝaHCO₃/SiO₂ in a glass filter-funnel and concentrated. Purification by column chromatography (toluene-EtOAc 40:1) gave the benzylated compound 4 (20.2 g, 45.8 mmol, 89%) as a colourless syrup: [D]²⁰ _D -117 (c 1.16, CHCI3); ¹H NMR (300 MHz, CDC1₃) D 1.87-1.98 (m, IH), 2.15-2.27 (m, 1 H), 2.61-2.78 (m, 2 H), 3.35 (d, 2 H, J= 3.08 Hz), 3.64 (dd, 1 H, J= 5.27, 9.22 Hz), 3.78 (dd, 1 H, J= 7.03, 9.22 Hz) 4.23 (m, 1 H), 4.41 (s, 2 H), 5.13-5.17 (m, 1 H), 5.78 (d, 1 H, J= 7.03 Hz), 7.1 1-7.47 (m, 14 H) and 7.55 (d, 1 H, J= 7.91 Hz); ¹³C NMR (67 MHz, CDC1₃) D 30.3, 33.7, 37.8, 43.2, 63.0, 71.2, 72.9, 77.8, 125.0, 126.0, 127.2, 127.3, 128.0, 128.2, 128.3, 128.6, 129.7, 138.1, 139.1, 139.3, 141.4, 152.6 and 174.8. Anal Calcd for C₂₈H₂7N0₄: C, 76.2; H, 6.2; N, 3.2. Found: C, 76.2; H, 6.2; N, 3.1.

( RV2-(Benzyloxymethyl)-4-phenyl- 1 -butanol (5).

The benzylated compound 4 (3.85 g, 8.72 mmol) was dissolved in THF (90 mL) under an argon atmosphere, the temperature was lowered to -60 °C and LAH (3.38 g, 89.0 mmol, 10 equiv.) was added. Stirring was continued at -60 °C for 30 min and at 0 °C for 1 h (R_/0.2, toluene-EtOAc 9:1). The reaction mixture was acidified by the addition of cold 0.6 M HC1. The suspension was extracted with Et₂0 (5x) and the combined organic layers were dried (MgS0₄) and concentrated. Purification by column chromatography (toluene-EtOAc 3: 1) gave alcohol 5 (1.96 g, 7.25 mmol, 83%) as a colourless oil: [π]²⁰ _D +14 (c 0.90, CHC1₃); Η NMR (400 MHz, CDC1₃) D 1.57-1.71 (m, 2 H), 1.86-1.93 (m, 1 H), 2.49 (s, 1 H), 2.57-2.67 (m, 2 H), 3.50-3.53 (m, 1 H), 3.62-3.69 (m, 2 H), 3.74-3.78 (m, 1 H), 4.51 (q, 2 H, J= 12.1 and 17.6 Hz) and 7.15-7.36 (m, 10 H); ¹³C NMR (67 MHz, CDC1₃) Q 29.8, 33.3, 40.1, 65.5, 73.4, 125.8, 127.6, 127.7, 128.3, 128.4, 138.0 and 142.1. Anal Calcd for C₁₈H₂₂0₂: C, 80.0; H, 8.2. Found: C, 80.1 ; H, 8.2.

(2S)-2-[(Benzyloxymethyn-4-phenylbutyll (^'2RV3.3.3-Trifluoro-2-methoxy-2-phenyl- propanoate.

Moshers ester of alcohol 5 was prepared according to the method described by Wipf and Fritch, using 26 mg (0.096 mmol) of 5 in 1.4 ml of CH₂C1₂ together with 580 GL of pyridine and 210 DL of (S)-(+)-D -methoxy- D-(trifluoromethyl)phenylacetic acid chloride with stirring during 40 min. Purification by column chromatography (toluene-EtOAc 40:1) gave the ester (38 mg, 0.078 mmol, 81%) as an oil: ¹³C NMR (75 MHz, CDC1₃) Q 29.9, 33.1, 37.8, 55.4, 66.4, 69.6, 73.2, 121.3, 125.2, 125.8, 127.2, 127.4, 127.5, 128.2, 128.3, 129.5, 132.2, 138.1, 141.5 and 166.4.

2-[fBenzyloxymethyO-4-phenylbutyl] (^"2RV3.3.3-Trifluoro-2-methoxy-2-phenyl-propanoate.

Moshers ester of racemic alcohol 5 (ref) was prepared as described above, using 20 mg (0.074 mmol) of racemic 5, which gave the racemic ester (28 mg, 0.058 mmol, 78%) as an oil: ¹³C NMR (75 MHz, CDC1₃) Q 29.8, 29.9, 33.0, 37.8 (2 C), 55.4, 66.2, 66.4, 69.6, 69.7, 73.2, 121.4, 125.2, 125.8, 127.2, 127.4 (2 C), 127.5, 128.2, 128.3, 129.5, 138.1 and 141.5.

(2SV2-(BenzyloxymethyO-4-phenylbutanaU6^').

DMSO (4.03 mL, 56.7 mmol, 2.2 equiv.) in CH₂C1₂ (11.3 mL) was added to a solution of oxalyl chloride (2.43 mL, 28.3 mmol, 1.1 equiv.) in CH₂C1₂ (64 mL) at -70 °C under argon. After stirring for 5 min, alcohol 5 (6.97 g, 25.8 mmol) dissolved in CH₂C1₂ (26 mL) was added dropwise (20 min). Stirring was continued for an additional 20 min. NEt₃ (7.18 mL, 54.0 mmol, 2.1 equiv.) was added, and the reaction mixture was stirred during 5 min at -70 °C, and subsequently during 1 h while the reaction mixture slowly was allowed to attain room temperature (R 0.35, pentane-EtOAc 15: 1 and R 0.57, toluene-EtOAc 9: 1). Dilution with H₂0, extraction of the aqueous layer with CH₂C1 (3x), drying (MgS0₄) and concentration gave the crude aldehyde 6. Purification by column chromatography (pentane- EtOAc 25: 1; 15: 1 and 10:1) gave 6 (5.64 g, 21.0 mmol, 82%) as a transparent oil, which was used immediately in the next step: [D]²⁰ _D +17 (c 1.1 , CHC1₃); Η NMR (300 MHz, CDC1₃) D 1.72-1.86 (m, 1 H), 1.97-2.12 (m, 1 H), 2.48-2.58 (m, 1 H), 2.62 (t, 2 H, J= 7.97 Hz), 3.67 (d, 2 H, J= 5.50 Hz), 4.47 (d, 2 H, J= 1.65 Hz), 7.12-7.40 (m, 10 H) and 9.68 (s, 1 H); ¹³C NMR (75 MHz, CDC1₃) D 27.3, 32.9, 51.3, 68.3, 73.1, 125.8, 127.4, 127.5, 128.1, 128.2 (2 C), 137.6, 140.9 and 203.1. Anal Calcd for Cι₈H₂₀O₂ • ^lΛ H₂0: C, 79.2; H, 7.6. Found: C, 79.4; H, 7.6.

Reduction of aldehyde 6 with borane-dimethylsulfide complex (BMS).

Aldehyde 6 was reduced with BMS according to the method described by Brown et al. , using 64 mg (0.238 mmol) of 6 in 2.0 ml of diethyl ether together with 60 DL of BMS with stirring at 0 °C during 30 min, followed by stirring at room temperature during 2.5 h. Purification by column chromatography (toluene-EtOAc 3: 1) gave alcohol 5 (50 mg, 0.185 mmol, 78%>) as an oil. Alcohol 5 (20 mg, 0.074 mmol) was converted to Moshers ester (28 mg, 0.058 mmol, 78%) according to the method described above. ¹³C NMR spectral data were in agreement with those reported above for (2S)-2-[(benzyloxymethyl)-4-phenylbutyl] (2R)-3,3,3-trifluoro-2-methoxy-2-phenyl-propanoate.

(2S)-2- {(4R,5S)-5-[( IS)- 1 -(Hydroxymethyl)-3-phenylpropyl]-2,2-dimethyl- 1 ,3-dioxolan-4- y 1 } -4-phenyl-l-butanol (9 a). ^SI^-I^R-SRVS-rd^-l-ChydroxymethylVS-phenylpropyll^^-dimethyl-lJ-dioxolan^- yl} -4-phenyl-l-butanol (9 b) and

(2^-2-(f4R.5RV5-[OS)-l-(^,hvdroxymethvn-3-phenylpropyll-2.2-dimethyl-1.3-dioxolan-4- y -4-phenyl-l-butanol (9 c .

Zinc dust (pre-washed with 1M HC1, EtOH, acetone and CH₂C1₂) (2.72 g, 41.6 mg-atom, 2.0 equiv.) was added to a solution of VC1₃ (THF)₃ in CH₂C1₂ (50 mL, 0.5 M, 25 mmol, 1.2 equiv.), under an argon atmosphere, changing the colour of the solution from deep-red to violet. The mixture was stirred at room temperature during 80 min, while the colour slowly changed from violet to black-green. l,3-Dimethyl-2-imidazolidinone (16.0 mL, 146 mmol, 7.0 equiv.) was added, and stirring was continued during 15 min, before dropwise addition (40 min) of aldehyde 6 (5.59 g, 20.8 mmol) dissolved in CH₂C1₂ (56 mL) which gave a brown reaction mixture. The reaction was quenched after 20 h (R/7 a 0.32 and R/7 b, 7 c 0.21, toluene-EtOAc 9: 1), by the addition of 1 M HC1 (127 mL). Stirring of the two-phase mixture during 30 min gave a turquoise aqueous layer and a brown organic layer, which were separated. Additional extraction of the aqueous layer with CH₂CI₂ (3x), drying (MgS0₄), concentration and subsequent separation by column chromatography (toluene; toluene-EtOAc 25: 1 and 15: 1) gave 7 a (2.29 g) and a mixture of 7 b and 7 c (2.16 g), all of which were used in the next step without further purification.

(7 a): ¹³C NMR (75 MHz, CDC1₃) 026.6, 31.3, 33.8, 34.0, 38.4, 39.8, 70.2, 72.9, 73.5, 73.6 (2 C), 75.9, 125.6, 127.5, 127.7, 127.8, 128.1 , 128.2 (2 C), 128.3 (2 C), 137.4, 137.7, 142.2 and 142.4. (7 b and 7 c): ¹³C NMR (75 MHz, CDC1₃) 028.5, 31.1, 33.5, 33.8, 40.7 (2 C), 70.3, 70.4, 72.6, 73.4, 73.7, 73.9, 125.7 (2 C), 127.5, 127.6 (2 C), 127.7, 128.2 (2 C), 128.3 (2 C), 137.7, 142.2 and 142.1.

Step II. Isopropylidene protection.

2,2-Dimethoxypropane (2.5 equiv.) and (±)-lO-camphorsulfonic acid (0.8 equiv.) were added to a solution of 7 a-c (1.0 equiv.) in acetone (5.3 mL/mmol) under nitrogen. After stirring for 30 min, when TLC (R_f 8 a 0.63 and R_f 8 b, 8 c 0.72, toluene-EtOAc 9: 1) showed completion of the reaction, the reaction mixture was poured into sat. aqueous NaHC0₃ and extracted with EtOAc (3x). Activated charcoal and NaS0₄ were added to the organic layer, which was stirred for 10 min before filtration through a pad of Celite and NaS0₄, and concentration. Column chromatography (toluene; toluene-EtOAc 20:1 and 9:1) gave the crude product, which was used in the next step without further purification.

(8 a). The isopropylidene protected compound was prepared according to step II, using 2.29 g of 7 a, and isolated as a white solid (2.29 g): ¹ C NMR (75 MHz, CDC1₃) D 25.1, 26.2, 29.6, 31.0, 33.0, 33.3, 38.0, 38.2, 69.6, 71.1, 73.0, 73.1, 77.1, 77.5, 106.9, 125.6 (2 C), 127.3 (2 C), 127.4, 128.1, 128.2 (3 C), 128.3, 128.4, 138.5, 138.8, 142.3 and 142.4.

(8 b and 8 c). The isopropylidene protected compounds was prepared according to step II, using 2.16 g of the mixture 7 b and 7 c, and isolated a transparent syrup (1.95 g). After column chromatography a small amount of alcohol 5 (265 mg, 0.980 mmol) was isolated as well. ¹³C NMR (75 MHz, CDC1₃) 0 27.4, 28.6, 31.5, 33.4, 33.8, 40.2, 40.7, 69.4, 70.7, 72.9, 73.0, 79.1, 79.3, 107.5, 107.8, 125.6, 125.7, 127.3 (2 C), 127.5, 128.1, 128.2 (2 C), 128.3, 138.4 (2 C), 142.2 and 142.3.

Step III. Hydrogenolysis. Compounds 8 a-c (1.0 equiv.) was dissolved in EtOAc (23 mL/mmol), and NaHCO₃ (3.0 equiv.), a minor amount of H O and Pd/C (0.8 g) was added. The suspension was stirred at room temperature under a hydrogen atmosphere. After 19 h, 39 h and 47 h, the suspension was filtered through a pad of Celite and Na₂SO₄, fresh reagents were added and stirring under hydrogen was continued. When TLC showed completion of the reaction (R 9 a 0.33, Rf 9 b, 9 c 0.31, toluene-EtOAc 1 : 1) after 63 h, filtration and concentration followed by column chromatography (toluene-EtOAc 2: 1) gave compounds 9 a-c.

(9 a). The title compound was prepared according to step III, using 2.47 g of 8 a, and isolated as a transparent syrup (1.27 g, 3.19 mmol): [D]²⁰ _D +91 (c 1.4, CHC1₃); Η NMR (300 MHz, CDC1₃) □ 1.26 (s, 3 H), 1.37 (s, 3 H), 1.59-1.71 (m, 4 H), 1.90 (s, 1 H), 1.97 (s, IH), 2.12-2.28 (m, 3 H), 2.49-2.67 (m, 3 H), 3.57-3.77 (m, 4 H), 4.03-4.08 (m, 1 H), 4.29- 4.32 (dd, 1 H, J= 2.64, 6.59 Hz) and 7.03-7.25 (m, 10 H); ¹³C NMR (75 MHz, CDC1₃) D 25.0, 26.1, 27.3, 30.4, 32.8, 33.1, 38.6, 39.5, 64.0, 64.2, 79.2, 81.0, 108.1, 125.8, 126.0, 128.3 (2 C), 128.4, 128.5, 141.4 and 141.9. Anal Calcd for C₂3H3₄O₄ • ^lλ H₂0: C, 74.5; H, 8.6. Found: C, 74.5; H, 8.6.

(9 b and 9 c). The mixture of the title compounds was prepared according to step III, using 1.77 g of the mixture of 8 b and 8 c, and isolated as a transparent syrup (1.03 g, 2.58 mmol): ^l3C NMR (75 MHz, CDC1₃) □ 26.9, 27.1, 27.3, 30.6, 33.3, 33.5, 40.6, 41.1, 62.2, 63.4, 79.9, 81.9, 108.8, 108.3, 125.8 (2 C), 128.2 (2 C), 128.3, 141.5 and 141.8. Anal Calcd for C₂₅H₃₄θ₄ ' 1/3 H₂0: C, 74.2; H, 8.6. Found: C, 74.3; H, 8.7.

Diol 9 a and the mixture of diols 9 b and 9 c were isolated with a total yield of 64% over three steps (2.30 g 5.77 mmol). A small amount of alcohol 5 (265 mg, 0.980 mmol) was also isolated.

General method for the preparation of compounds 10 a-c. Method I. The diol 9 (1.0 equiv.) was dissolved in a stirred mixture of CH₃CN (4 mL/mmol), CH₂C1₂ (4 mL/mmol) and H₂0 (6 mL/mmol). RuCl₃ • xH₂0 (0.05 equiv.) and NaI0 (9.0 equiv.) were added. TLC (toluene-EtOAc 1 :3) showed completion of the reaction after vigorous stirring for 105 min. The reaction mixture was diluted with CH₂C1₂ (lx volume) and stirred for an additional 10 min. Water was added (lx volume) and the aqueous layer was extracted with EtOAc (4x). The combined organic layers were dried (MgS0₄), and concentrated to provide the corresponding carboxylic acid, which was immediately activated without further purification.

A mixture of the carboxylic acid, NN-disuccinimidyl carbonate (5.0 equiv.) and pyridine (6.9 equiv.) in CH₃CΝ (16 mL/mmol) was stirred under an argon atmosphere. When TLC showed completion of the reaction after 23 h (R/10 a 0.63, R 10 b 0.47 and R_/10 c 0.58, toluene-EtOAc 1 : 1), toluene (V2 x volume) was added and the reaction mixture was concentrated. The residue was dissolved in EtOAc, extracted with water (5x) and the combined aqueous layers were washed with EtOAc (3x). The combined organic layers were dried (Na₂S0₄), concentrated and purificated by column chromatography (toluene-EtOAc 4:1) to provide the disuccinimidyl esters 10 a-c.

Succinimidyl (2RV2-r(4R.5Sy5-(3-phenylpropyl-(lR -succinimidyloxycarbonyO-2.2- dimethyl-1.3-dioxolan-4-yll-4-phenylbutanoat (10 a). The title compound was prepared according to method I, using 1.14 g (2.87 mmol) of diol 9 a, and isolated as a white foam in 22% yield (393 mg, 0.633 mmol): [O]²⁰ _D +19 (c 0.35, CDCI₃); ^l3C NMR (75 MHz, CDC1₃) O 25.1, 25.7, 26.8, 31.8, 32.0, 32.5, 32.6, 42.2, 43.5, 76.0, 77.8, 108.9, 125.2, 125.9 (2 C), 128.1, 128.2, 128.3, 128.5, 128.7, 128.9, 140.9, 141.1, 168.2, 168.5, 168.6 and 168.8. Anal Calcd for C₃₃H₃₆N₂O₁₀: C, 63.9; H, 5.9; N, 4.5. Found: C, 63.8; H, 5.9; N, 4.4.

Succinimidyl (2RV2- (4R.5RV5-(^'3-phenylpropyl-(lR^')-l-succinimidyloxycarbonyπ-2.2- dimethyl-1.3-dioxolan-4-vn-4-phenylbutanoat (10 b) and succinimidyl (2R -2- (4S.5S)-5-(3-phenylpropyl-(lRVl-succinimidyloxycarbonvπ-2.2- dimethyl- 1.3-dioxolan-4-yl]-4-phenylbutanoat (10 cY

The title compounds, separated by column chromatography, were prepared according to method I, using 965 mg (2.42 mmol) of the mixture of diols 9 b and 9 c, and isolated as white foams in 43% total yield (10 b: 126 mg, 0.203 mmol; 10 c: 527 mg, 0.849 mmol).

(10 b): [O] ⁰ _D +86 (c 0.40, CDC1₃); ¹³C NMR (75 MHz, CDC1₃) 0 25.7, 27.4, 29.8, 32.7, 45.6, 78.8, 110.4, 126.0, 128.1, 128.3, 128.6, 128.9, 140.8, 168.3 and 168.5. Anal Calcd for C₃3H36N₂O₁₀ • V* H₂0: C, 63.4; H, 5.9; N, 4.5. Found: C, 63.4; H, 5.1; N, 4.3.

(10 c): [D]²⁰ _D -5.3 (c 0.75, CDC1₃); ¹³C NMR (75 MHz, CDC1₃) 0 25.7, 27.6, 27.9, 31.4, 31.7, 32.6, 33.0, 45.0, 78.5, 80.2, 110.3, 125.2, 125.9, 126.1, 128.1, 128.3, 128.4, 128.6, 128.9, 140.4, 167.5 and 168.6. Anal Calcd for C^N o: C, 63.9; H, 5.9; N, 4.5. Found: C, 63.8; H, 5.9; N, 4.5.

General method for the preparation of compounds 12 a and 12 b.

Method II. The disuccinimidyl esters 10 a-c (1.0 equiv.) and (lS,2R)-(-)-cw-l-amino-2- indanol (6.3 equiv.) were dissolved in 1 ,2-dichloroethane (30 mL/mmol) under a nitrogen atmosphere. The reaction was stirred at 60 °C until TLC (R_/ll a 0.31, R/ll b 0.43 and R_f 11 c 0.19, toluene-EtOAc 1 : 1) showed completion of the reaction (16-20 h). Concentration followed by column chromatography (toluene-EtOAc 3:1 and 1:1) gave the amides 11 a-c. A mixture of FeCl₃ • 6H₂O (3.5 equiv.) and the amides 11 a and 11 b (1.0 equiv.) were dissolved in warm CH₂CI₂ (15 mL/mmol) and stirred at room temperature for 1 h and 8 h (R_/ 12 a 0.18, R_/12 b 0.21, CHCL-MeOH 20:1), respectively. Sat. aqueous NaHC0₃ (lx volume) was added and the aqueous layer was extracted with CH₂C1₂ (3x) and EtOAc (3x). Drying (MgSU₄), concentration, and purification by column chromatography (CHC1₃; CHCl₃-MeOH 40: 1 and 30: 1), followed by trituration from EtOAc, gave the target compounds 12 a and 12 b. (11 a): ¹³C NMR (75 MHz, CDC1₃) 023.3, 25.4, 32.7, 32.9, 33.4, 34.0, 39.2, 39.7, 46.1, 46.7, 58.0, 58.2, 73.4, 73.6, 77.4, 77.7, 107.5, 123.6, 125.0, 125.3, 125.4, 125.9, 126.0,

126.8, 127.1, 128.1, 128.3 (2 C), 128.4, 128.6, 139.8, 140.1, 140.4, 141.1, 141.5, 173.2 and

174.7.

(11 b): ¹³C NMR (75 MHz, CDC1₃) 027.3, 29.6, 33.4, 39.5, 49.8, 57.7, 73.5, 79.6, 108.9,

124.7, 125.2, 126.0, 127.2, 128.3, 128.5, 140.0, 140.2, 141.0 and 172.9. (11 c): 29% yield (73 mg, 0.106 mmol); [D]²⁰ _D +15 (c 0.48, CHCl₃-MeOH 1 : 1); 'H NMR (400 MHz, CDCI₃ and CD₃OD) O 1.40 (s, 6H), 1.77-1.84 (m, 2 H), 2.00-2.10 (m, 2 H), 2.60-2.68 (m, 4 H), 2.74-2.82 (m, 2 H), 2.96 (d, 2 H, J= 16.41 Hz), 3.17 (dd, 2 H, J= 4.69, 16.02 Hz), 3.67 (d, 2 H, J= 3.12 Hz), 4.17 (d, 2 H, J= 5.86 Hz), 4.61-4.64 (m, 2 H), 5.42 (d, 2 H, J= 5.08 Hz) and 7.15-7.35 (m, 18 H);¹³C NMR (75 MHz, CDCI₃ and CD₃OD) 026.9, 31.5, 33.1, 39.6, 50.6, 57.2, 72.5, 79.7, 108.9, 124.0, 124.9, 125.7, 126.5, 127.7, 128.0,

128.1, 139.9, 140.4, 140.7 and 173.2. Anal Calcd for GuH^Oe * 1 H₂O: C, 73.1 ; H, 7.1 ; N, 4.0. Found: C, 72.9; H, 6.8; N, 4.0. Nl.N6-Dir(2RVhvdroxy-(lS -indanyll-(2R.3R.4S.5RV2.5-di(2-phenylethylV3.4- dihydroxyhexanediamide (12 a)

The title compound was prepared according to method II, using 195 mg (0.314 mmol) of the disuccinimidyl ester 10 a, and isolated as a white solid in 47% yield (96 mg, 0.148 mmol): [O]²⁰ _D +20 (c 0.94, CHCl₃-MeOH 1 : 1); Η ΝMR (400 MHz, CDC1₃ and CD₃OD) 01.97- 2.23 (m, 4 H), 2.61-2.85 (m, 6 H), 2.92-2.99 (m, 2 H), 3.12-3.19 (m, 2 H), 3.69 (bs, 1 H), 3.91-3.94 (m, 1 H), 4.48 (s, 6 H), 4.54-4.61 (m, 2 H), 5.38-5.41 (m, 2 H) and 7.13-7.30 (m, 18 H); ¹³C ΝMR (75 MHz, CDC1₃ and CD₃OD) 0 33.3 (2 C), 39.3, 47.0, 56.8, 56.9, 72.3, 72.4, 72.7, 73.4, 123.6, 123.8, 124.7, 125.3, 125.4, 126.5, 127.5, 127.8 (3 C), 139.7, 139.8, 140.2, 141.1, 141.5 and 175.9. Anal Calcd for C₄oH₄₄Ν₂0₆ • 1 V2 H₂0: C, 71.1; H, 7.0; N, 4.2. Found: C, 71.3; H, 6.7; N, 4.1.

Nl.N6-Dir(2RVhvdroxy-(lS)-indanyll-(2R.3R.4R.5RV2.5-di(2-phenylethvn-3.4- dihydroxyhexanediamide (12 b).

The title compound was prepared according to method II, using 67 mg (0.108 mmol) of the disuccinimidyl ester 10 c, and isolated as a white solid in 58% yield (41 mg, 0.063 mmol): [O]²⁰ _D +30 (c 0.77, CHCl₃-MeOH 1 :1); 'H ΝMR: (400 MHz, CDC1₃ and CD₃OD) 01 -99- 2.03 (m, 2 H), 2.05-2.15 (m, 2 H), 2.60-2.69 (m, 4 H), 2.76-2.84 (m, 2 H), 2.95 (dd, 2 H, J= 2.20, 16.48 Hz), 3.15 (dd, 2 H, J= 5.13, 16.48 Hz), 3.81 (d, 2 H, J= 8.79 Hz), 4.24 (s, 6 H), 4.60-4.63 (m, 2 H), 5.41 (d, 2 H, J= 5.13 Hz), and 7.16-7.33 (m, 18 H); ¹³C ΝMR (100 MHz, CDCI3 and CD₃OD) D 30.8, 33.4, 39.4, 50.8, 57.1, 71.6, 72.6, 124.0, 124.7, 125.5, 126.6, 127.6, 127.9, 139.8, 140.0, 141.2 and 174.6. Anal Calcd for C₄oH₄₄Ν₂0₆ • 1 H₂0: C, 72.1 ; H, 7.0; N, 4.2. Found: C, 72.4; H, 6.6; N, 4.1.

General method for the preparation of compounds 14 a and 14 b. Method III. The disuccinimidyl esters 10 a-c (1.0 equiv.) and H-Val-NHMe (6.3 equiv.) were dissolved in 1 ,2-dichloroethane (30 mL/mmol) under a nitrogen atmosphere. The reaction was stirred at 50 °C until TLC (R_/13 a 0.34, R/13 b 0.41, CHCh-MeOH 9: 1) showed completion of the reaction (22 h). Concentration followed by column chromatography (CHCl -MeOH 20:1) gave the amides 13 a and 13 b. A mixture of FeCl₃ • 6H₂0 (3.5 equiv.) and the amides 13 a and 13 b (1.0 equiv.) were dissolved in warm CH₂C1₂ (17.5 mL/mmol) and stirred at room temperature. The reaction was followed with TLC (R/14 a 0.23, R/14 b 0.40, CHCl₃-MeOH 9: 1), and every other hour the reaction was concentrated and new CH₂C1₂ was added. When TLC showed completion of the reaction (14 a: 4 h and 14 b: 8 h) sat. aqueous NaHCO₃ (lx volume) was added and the aqueous layer was extracted with CH₂C1₂ (3x) and EtOAc (3x). Drying (MgSO₄), concentration, and purification by column chromatography (CHC1₃; CHCl₃-MeOH 30: 1 and 20:1), followed by trituration from EtOAc, gave the target compounds 14 a and 14 b. (13 a): ¹³C NMR (75 MHz, CDC1₃ and CD₃OD) 019.0, 24.3, 25.6, 25.7, 25.8, 30.8, 31.0, 31.6, 32.0, 32.3, 33.0, 46.1, 46.5, 58.5, 59.0, 76.4, 77.9, 107.7, 125.8, 128.0, 128.2, 140.9, 141.6, 172.1, 172.2, 173.0 and 174.0.

(13 b): ¹³C NMR (75 MHz, CDC1₃) 018.6, 19.5, 26.2, 27.2, 29.7, 30.6, 33.6, 49.1, 58.8, 79.1, 109.1, 125.9, 128.3, 141.2, 171.6 and 172.6.

Nl.N6-Di-r(lS -2-methyl-l-(methylcarbamovnpropyl1-(2R.3R.4S.5RV2.5-di(2-phenylethvn- 3.4-dihydroxyhexanediamide (14 a)

The title compound was prepared according to method III, using 153 mg (0.247 mmol) of the disuccinimidyl ester 10 b, and isolated as a white solid in 7% yield (10 mg, 0.017 mmol) together with recovered 13 a (26 mg, 0.040 mmol). [O]²⁰ _D -27 (c 0.32, CHCl₃-MeOH 1 : 1); Η ΝMR: (400 MHz, CDC1₃ and CD₃OD) 00.92 (d, 3 H, J= 6.64 Hz), 0.97 (d, 9 H, J= 6.64 Hz), 1.85-2.19 (m, 6 H), 2.54-2.72 (m, 6 H), 2.76 (s, 3 H), 2.77 (s, 3 H), 3.60-3.62 (m, 1 H), 3.70-3.73 (m, 1 H), 4.12-4.18 (m, 2 H), 4.56 (s, 6 H) and 7.16-7.55 (m, 10 H); ¹³C ΝMR (100 MHz, CDC1₃ and CD₃OD) 0 17.8, 18.7, 18.8, 25.2, 28.4, 30.1, 30.3, 32.5, 33.2, 46.5, 58.3, 58.5, 72.5, 73.5, 125.2, 125.3, 127.6, 127.7, 127.8, 141.0, 141.3, 171.8, 171.9, 175.2 and 175.4. HRMS Calcd for C₃₄H₅o0₆Ν₄ [M+Na] 633.3628. Found [M+Na] 633.3604.

Nl.N6-Di-[(lS)-2-methyl-l-(methylcarbamoyl)propyll-(2R.3R.4R.5R)-2.5-di(2-phenylethyl)- 3.4-dihvdroxyhexanediamide (14 b).

The title compound was prepared according to method III, using 52 mg (0.084 mmol) of the disuccinimidyl ester 10 b, and isolated as a white solid in 22% yield (1 1 mg, 0.018 mmol) together with recovered 13 b (16 mg, 0.025 mmol). [O]²⁰ _D -23 (c 0.75, CHCl₃-MeOH 1 : 1); 'H ΝMR: (400 MHz, CDC1₃ and CD₃OD) D 0.60-0.64 (m, 12 H), 1.61-1.74 (m, 6 H), 2.18- 2.26 (m, 6 H), 2.42 (s, 6 H), 3.34 (d, 2 H, J= 8.59 Hz), 3.76-3.78 (m, 2 H), 4.32 (s, 6 H) and 6.81-7.27 (m, 10 H); ¹³C ΝMR (100 MHz, CDC1₃ and CD₃OD) 0 17.8, 18.5, 25.1, 29.9, 30.8, 33.0, 50.4, 58.7, 71.1, 125.1, 127.5, 141.0, 171.9 and 174.1. HRMS Calcd for C₃₄H₅o0₆N₄ [M+Na] 633.3628. Found [M+Na] 633.3638.

Biological Example 1

Compounds arre tested for HTV protease activity in a spectrophotometric enzyme assay using the chromogenic substrate His-Lys-Ala-Arg-Val-Leu-p-nitro-Phe-Glu-Ala-Nle-Ser- amide and purified HIV proteinase. The rate of cleavage is followed by continuously registering the change in absorbance at 300 nm. The IC₅o representing the compound concentration which inhibits enzyme performance by 50%) is calculated from the dose response curve. Extremely active compounds are best compared with affinity constants (K,). Compounds are also tested for HTV protease activity in cell culture. MT4 cells grown in RPMI 1640 cell culture medium including 10 % fetal calf serum are infected with 10 TCID HIV-1 per 2 x 10⁵ cells and cultured for 6 days. XTT is added and the amount of XTT formazan produced in the following 6 hours represents the number of surviving cells. Results are expressed as the ED₅o, that is the concentration in μg/ml of the compound of the invention which suppresses viral replication by 50%. Additionally, activity measurements are run in the corresponding system further comprising 40% human serum to mimic protein binding effects in vivo.

1 Example 1 of WO 98/45330:

2 Example 10 of WO98/45330 The compounds of the invention have a particularly good performance in the presence of human serum relative to the closest compounds of the prior art .

Claims

1. A compound of the formula I:

wherein:

A' and A" are independently the same or different group of the formula II:

wherein:

R' is H, CH₃, C(CH₃)₂, -OR^a, -N(R^a)₂, -N(R^a)OR^a or -DP

R'" is H or CH₃; R^a is H, C_rC₃ alkyl;

D is a bond, C,_-3 alkylene, -C(=0)-, -S(O)- or -S(0)₂- ;

P is an optionally substituted, mono or bicyclic carbo- or heterocycle; R" is H, any of the sidechains found in the natural amino acids, Cι-_aalkyl, carboxacetamide, or a group (CH₂)„DP;

M is a bond or -C(=0)N(R" ;

Q is absent, a bond, -CH(OH)- or -CH₂-; or R" together with Q , M and R' define an optionally substituted 5 or 6 membered carbo- or heterocyclic ring which is optionally fused with a further 5 or 6 membered carbo- or heterocyclic ring;

X is H, OH, OCH₃;

Y is H, OH, OCH₃,

Z' and Z" are independently an optionally substituted mono-or bicyclic carbo-or heterocycle;

() is a methylene group; n and m are independently 0, 1 or 2; and pharmaceutically acceptable salts and prodrugs thereof.

2. A compound according to claim 1, wherein A' and/or A" are a group of the formula

Ila:

Ila wherein

n is 0, 1 or 2; R' is methyl or methyloxy; and

R" is hydrogen, alkyl, including methyl, ethyl, isopropyl, isobutyl, cycloalkyl such as cyclopropyl, cyclobutyl or cyclohexyl, cycloalkenyl, benzyl, carboxacetamide or 4- imidazolylmethyl or a side chain found in the natural amino acids.

3. A compound according to claim 2 wherein R" is the side chain of L-valine.

4. A compound according to claim 2 or 3 wherein n is 0 and R' is methyl.

5. A compound according to claim 1, wherein A' and/or A" are a group of the formula

where

R'" is H or CH₃,

R¹ is H, NR⁴R⁴, C(=0)R³, CR³R⁴ or a monocyclic, optionally substituted, carbo- or heterocycle; R² is OH, or together with R¹ is =0, or if R¹ is NR⁴R⁴, then R² may be H; R³ is H, halo, C,-C₃ alkyl, OR⁵, NR⁴R⁴; R⁴ is H, C,-C₃ alkyl; R⁵ is H or a pharmaceutically acceptable ester; R⁶ is OH, NH₂, carbamoyl or carboxy;

R⁷ is hydrogen, C]-C₄ straight or branched alkyl or together with the adjacent carbon atoms forms a fused phenyl or pyrimidine;

6. A compound according to claim 5, wherein the moiety of formula III has the structure:

7. A compound according to claim 1, wherein both A' and A" are identical.

8 A compound as defined in claim 1 wherein A' is as defined in claim 2 and A" is as defined in claim 6.

9. A compound according to claim 1 , wherein each m is 2 and Z' and /or Z" is phenyl unsubstituted or substituted with one to three substituents selected from halo, methoxy, hydroxy, amino, cyano, hydroxymethyl, aminomethyl, morpholinethoxy, alkylsulfonyl, carbamoyl, benzyloxy, phenyl or a 5 or 6 membered heterocycle containing one or two hetero atoms such as thiophene, pyrimidine, N-morpholine, N-piperidine, N-piperazine, N'- methyl-N-piperazine, N-pyrrolidone, N-pyrrolidine and the like, any of which cyclic optional substituents may optionally be substituted as defined herein.

10. A compound according to claim 9 wherein Z' and/or Z" comprise phenyl, 2- fluorophenyl, 2-methylphenyl, 2,4-difluorophenyl, 4-fluorophenyl, 4-bromophenyl, 4- phenylphenylyl, 4-thiophenylphenyl, 4-(4'-nitrophenyl)phenyl, 4-thienylphenylyl, 4- thiazolylphenylyl or 4-(pyridyl)phenyl.

1 1. A compound according to claim 9 wherein both Z' and Z" are halophenyl, especially fluoro or difluorophenyl.

12 A compound according to any one of claims 1-11 wherein each m is 1 and Z' and/or Z" is phenyl, unsubstituted or substituted with one to three substituents selected from halo, methoxy, hydroxy, amino, cyano, hydroxymethyl, aminomethyl, morpholinethoxy, alkylsulfonyl, carbamoyl, benzyloxy, phenyl or a 5 or 6 membered heterocycle containing one or two hetero atoms such as thiophene, pyrimidine, N-morpholine, N-piperidine, N- piperazine, N'-methyl-N-piperazine, N-pyrrolidone, N-pyrrolidine and the like, any of which cyclic optional substituents may optionally be substituted as defined herein.

13 A compound according to claim 12 wherein __' and/or Z" comprise phenyl, 2- fluorophenyl, 2-methylphenyl, 2,4-difluorophenyl, 4-fluorophenyl, or 4-bromophenyl.

14. A compound according to claim 1, wherein X and Y are both OH or one is H and the other is OH.

15. A compound according to claim 1, with the stereochemistry 2R,3R,4R,5R.

16 A compound according to claim 1 wherein X and Y are OH, n and m are 2, Zi and Zii are phenyl, halophenyl or dihalophenyl and A' and A" are indanolamine.

17. A pharmaceutical composition comprising a compound as defined in any preceding claim and a pharmaceutically acceptable carrier or diluent therefor.

18. A method for inhibiting the replication of HIV comprising administering an effective amount of a compound as defined in any one of claims 1 to 17 to a subject afflicted with said condition.