WO2008000777A2 - Phenyl- pyrazole derivatives, process for their preparation and their pharmaceutical use - Google Patents

Phenyl- pyrazole derivatives, process for their preparation and their pharmaceutical use Download PDF

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Publication number
WO2008000777A2
WO2008000777A2 PCT/EP2007/056448 EP2007056448W WO2008000777A2 WO 2008000777 A2 WO2008000777 A2 WO 2008000777A2 EP 2007056448 W EP2007056448 W EP 2007056448W WO 2008000777 A2 WO2008000777 A2 WO 2008000777A2
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WO
WIPO (PCT)
Prior art keywords
amino
fluorophenyl
pyrazole
methyl
trifluoro
Prior art date
Application number
PCT/EP2007/056448
Other languages
French (fr)
Other versions
WO2008000777A3 (en
Inventor
Heather Anne Barnett
Ian Baxter Campbell
Diane Mary Coe
Anthony William James Cooper
Graham George Adam Inglis
Haydn Terence Jones
Steven Philip Keeling
Simon John Fawcett Macdonald
Iain Mcfarlane Mclay
Philip Alan Skone
Gordon Gad Weingarten
James Michael Woolven
Original Assignee
Glaxo Group Limited
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Priority claimed from GB0612942A external-priority patent/GB0612942D0/en
Priority claimed from GB0625458A external-priority patent/GB0625458D0/en
Priority claimed from GB0700077A external-priority patent/GB0700077D0/en
Application filed by Glaxo Group Limited filed Critical Glaxo Group Limited
Publication of WO2008000777A2 publication Critical patent/WO2008000777A2/en
Publication of WO2008000777A3 publication Critical patent/WO2008000777A3/en

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Classifications

    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D231/00—Heterocyclic compounds containing 1,2-diazole or hydrogenated 1,2-diazole rings
    • C07D231/02—Heterocyclic compounds containing 1,2-diazole or hydrogenated 1,2-diazole rings not condensed with other rings
    • C07D231/10—Heterocyclic compounds containing 1,2-diazole or hydrogenated 1,2-diazole rings not condensed with other rings having two or three double bonds between ring members or between ring members and non-ring members
    • C07D231/14—Heterocyclic compounds containing 1,2-diazole or hydrogenated 1,2-diazole rings not condensed with other rings having two or three double bonds between ring members or between ring members and non-ring members with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals, directly attached to ring carbon atoms
    • C07D231/38—Nitrogen atoms
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P19/00—Drugs for skeletal disorders
    • A61P19/02—Drugs for skeletal disorders for joint disorders, e.g. arthritis, arthrosis
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P37/00—Drugs for immunological or allergic disorders
    • A61P37/08—Antiallergic agents

Definitions

  • the present invention relates to non-steroidal compounds and a process for their preparation, to pharmaceutical compositions comprising the compounds and the preparation of said compositions, to intermediates and to use of the compounds for the manufacture of a medicament for therapeutic treatment, particularly for the treatment of inflammation and/or allergic conditions.
  • Nuclear receptors are a class of structurally related proteins involved in the regulation of gene expression.
  • the steroid hormone receptors are a subset of this family whose natural ligands typically comprise endogenous steroids such as estradiol (estrogen receptor), progesterone (progesterone receptor) and Cortisol (glucocorticoid receptor).
  • estradiol estradiol
  • progesterone progesterone receptor
  • Cortisol glucocorticoid receptor
  • glucocorticoids have proved useful in the treatment of inflammation, tissue rejection, auto-immunity, various malignancies, such as leukemias and lymphomas, Cushing's syndrome, rheumatic fever, polyarteritis nodosa, granulomatous polyarteritis, inhibition of myeloid cell lines, immune proliferation/apoptosis, HPA axis suppression and regulation, hypercortisolemia, modulation of the Th1/Th2 cytokine balance, chronic kidney disease, stroke and spinal cord injury, hypercalcemia, hypergylcemia, acute adrenal insufficiency, chronic primary adrenal insufficiency, secondary adrenal insufficiency, congenital adrenal hyperplasia, cerebral edema, thrombocytopenia, Little's syndrome, inflammatory scalp alopecia, panniculitis, psoriasis, discoid lupus erythemnatosus, inflamed cysts, atopic dermatitis, py
  • Glucocorticoids are especially useful in disease states involving systemic inflammation such as inflammatory bowel disease, systemic lupus erythematosus, polyarteritis nodosa, Wegener's granulomatosis, giant cell arteritis, rheumatoid arthritis, osteoarthritis, seasonal rhinitis, allergic rhinitis, vasomotor rhinitis, urticaria, angioneurotic edema, chronic obstructive pulmonary disease, asthma, tendonitis, bursitis, Crohn's disease, ulcerative colitis, autoimmune chronic active hepatitis, organ transplantation, hepatitis and cirrhosis.
  • Glucocorticoids have also been used as immunostimulants and repressors and as wound healing and tissue repair agents.
  • the present invention provides compounds of formula (I):
  • R 1 is selected from hydrogen, -OCHF 2 , fluorine and chlorine;
  • R 2 and R 3 are each independently selected from hydrogen, fluorine, chlorine and bromine; when R 1 is -OCHF 2 , R 2 and R 3 are each hydrogen, or R 2 is chlorine and R 3 is hydrogen; n is an integer selected from 0, 1 and 2, when n is 1 , X is selected from chlorine and fluorine, and when n is 2, each X is fluorine; and salts and solvates thereof (hereinafter "compounds of the invention").
  • R 1 is selected from hydrogen, -OCHF 2 , fluorine and chlorine; R 2 and R 3 are independently selected from hydrogen, fluorine and chlorine; when R 1 is -OCHF 2 , R 2 and R 3 are each hydrogen; and salts and solvates thereof.
  • the compounds of formula (I) each contain a chiral centre and there are two possible stereoisomers (enantiomers) of each compound of formula (I).
  • Enantiomer 1 and Enantiomer 2 are used herein to refer to the enantiomers of a compound of formula (I), based on the order of their elution using the chiral chromatography methodology described herein.
  • Enantiomer 1 refers to the first enantiomer to elute
  • Enantiomer 2 refers to the second enantiomer to elute.
  • a mixture of enantiomers such as a racemic mixture, may be preferred.
  • the compound of formula (I) is the racemic mixture (the racemate).
  • a single enantiomer may be preferred, for example the Enantiomer 1
  • the compound of formula (I) is the Enantiomer 1
  • the compound of formula (I) is the Enantiomer 2
  • stereoisomer and “isomer” as used herein encompass enantiomer, atropisomer and/or rotamer
  • the compounds of the invention are glucocorticoid receptor binders Accordingly, it has been found that at least one of the possible enantiomers of each of the compounds of formula (I) binds to the glucocorticoid receptor
  • At least one of the possible enantiomers of each of the compounds of formula (I) has glucocorticoid receptor binding activity Accordingly, at least one of the possible enantiomers of each compound of formula (I) modulates the glucocorticoid receptor
  • modulator refers to a compound which binds to the glucocorticoid receptor and acts as either an agonist, a partial agonist or an antagonist of the glucocorticoid receptor
  • the compounds of the invention may provide agonism of the glucocorticoid receptor Additionally, it appears that one or more of the possible enantiomers of some of the compounds of formula (I) possess advantageous selectivity in respect of maintaining transrepression activity whilst reducing the transactivation activity. These observations are believed to be indicative that the compounds of the invention provide antiinflammatory properties with fewer or less severe related side effects.
  • R 1 is selected from fluorine and chlorine.
  • R 2 and R 3 are each independently selected from hydrogen, fluorine and chlorine.
  • R 1 is -OCHF 2
  • R 2 and R 3 are each hydrogen.
  • R 2 is selected from fluorine and chlorine.
  • R 3 is hydrogen. In another embodiment, when R 3 is fluorine, R 1 is hydrogen and R 2 is chlorine. In a further embodiment, when R 3 is chlorine, R 1 and R 2 are each hydrogen.
  • R 1 is fluorine
  • R 2 is chlorine and R 3 is hydrogen.
  • R 1 and R 2 are each fluorine and R 3 is hydrogen.
  • R 1 and R 2 are each chlorine and R 3 is hydrogen.
  • n 1
  • X is fluorine.
  • the fluorine is in the para position on the phenyl ring.
  • the compound of formula (I) is:
  • the compound of formula (I) is:
  • the compound of formula (I) is: 5-amino- ⁇ /-(2- ⁇ [( ⁇ 2-[(difluoromethyl)oxy]phenyl ⁇ carbonyl)(2-hydroxyethyl)amino]methyl ⁇ - 3,3,3-trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide; 5-am ⁇ no- ⁇ /-(2- ⁇ [[(3-chlorophenyl)carbonyl](2-hydroxyethyl)ann ⁇ no]nnethyl ⁇ -3,3,3-t ⁇ fluoro-2- hydroxypropyO-i- ⁇ -fluorophenyO-I W-pyrazole ⁇ -carboxamide, 5-am ⁇ no-1-(4-fluorophenyl)- ⁇ y-(3,3,3-tr ⁇ fluoro-2- ⁇ [[(2-fluorophenyl)carbonyl](2- hydroxyethyl)am
  • One embodiment of the invention embraces compounds of formula (I) and salts and solvates thereof. Another embodiment of the invention embraces compounds of formula (I) and salts thereof. Another embodiment of the invention embraces compounds of formula (I) and solvates thereof. A further embodiment of the invention embraces compounds of formula (I) as the free base.
  • Salts and solvates of the compounds of formula (I) which are suitable for use in medicine are those wherein the counter-ion or associated solvent is pharmaceutically acceptable.
  • salts and solvates having non-pharmaceutically acceptable counter-ions or associated solvents are within the scope of the present invention, for example, for use as intermediates in the preparation of other compounds of formula (I) and their pharmaceutically acceptable salts and solvates thereof.
  • One embodiment of the invention embraces compounds of formula (I) and pharmaceutically acceptable salts and solvates thereof.
  • Another embodiment of the invention embraces compounds of formula (I) and pharmaceutically acceptable salts thereof.
  • Another embodiment of the invention embraces compounds of formula (I) and pharmaceutically acceptable solvates thereof.
  • Suitable salts according to the invention include those formed with both organic and inorganic acids or bases.
  • Pharmaceutically acceptable acid addition salts may include those formed from hydrochloric, hydrobromic, sulphuric, phosphoric, trifluoroacetic, sulphamic, sulphanilic, methanesulphonic, ethanesulphonic and arylsulphonic (for example p-toluenesulphonic, benzenesulphonic, naphthalenesulphonic or naphthalenedisulphonic) acids.
  • Pharmaceutically acceptable base salts may include alkali metal salts such as those of sodium and potassium and alkaline earth metal salts such as those of calcium.
  • solvates include hydrates.
  • the compounds of the invention may have the ability to crystallise in more than one form. This is a characteristic known as polymorphism, and it is understood that such polymorphic forms (“polymorphs”) are within the scope of the present invention. Polymorphism generally can occur as a response to changes in temperature or pressure or both and can also result from variations in the crystallisation process. Polymorphs can be distinguished by various physical characteristics known in the art such as x-ray diffraction patterns, solubility, and melting point.
  • the compounds of the invention are expected to have potentially beneficial antiinflammatory and/or anti-allergic effects, particularly upon oral administration, demonstrated by, for example, their ability to bind to the glucocorticoid receptor and to ellicit a response via that receptor.
  • the compounds of the invention may be of use in the treatment of inflammatory and/or allergic disorders.
  • Examples of disease states associated with glucocorticoid receptor activity include skin diseases such as eczema, psoriasis, allergic dermatitis, neurodermatitis, pruritis, exfoliative dermatitis, pemphigus and hypersensitivity reactions; inflammatory conditions of the nose, throat or lungs such as asthma (including allergen-induced asthmatic reactions), rhinitis (including hayfever), nasal polyps, chronic obstructive pulmonary disease (COPD), interstitial lung disease, and fibrosis; inflammatory bowel conditions such as ulcerative colitis and Crohn's disease; auto-immune diseases such as rheumatoid arthritis, systemic lupus erythematosus, termporal arteritis, polyarteritis nodosa, polymyositis, ankylosing spondylitis, sarcoidosis, autoimmune hepatitis; cancers such as acute and lymphatic leukaemia, myelo
  • amphetamine or amphetamine-related drugs e.g. dextroamphetamine, methylamphetamine
  • Compounds having glucocorticoids receptor activity may also have utility in inducing suppression of the immune system during organ transplantation, in acute transplant reject, angioedema of the upper respiratory tract and anaphylactic shock.
  • diseases states in which the compounds of the present invention are expected to have utility include rheumatoid arthritis, asthma, COPD, allergy and rhinitis.
  • compounds of the invention are expected to be of use in human or veterinary medicine, in particular as anti-inflammatory and/or anti-allergic agents.
  • a compound of formula (I) or a pharmaceutically acceptable salt or solvate thereof for use in the treatment of patients with an inflammatory and/or allergic condition.
  • a compound of formula (I) or a pharmaceutically acceptable salt or solvate thereof for use in the treatment of patients with rheumatoid arthritis, asthma, COPD, allergy and/or rhinitis.
  • a compound of formula (I) or a pharmaceutically acceptable salt or solvate thereof for use in the treatment of patients with eczema, psoriasis, allergic dermatitis, neurodermatitis, pruritis and/or hypersensitivity reactions.
  • a compound of formula (I) or a pharmaceutically acceptable salt or solvate thereof for the manufacture of a medicament for the treatment of patients with an inflammatory and/or allergic condition.
  • a compound of formula (I) or a pharmaceutically acceptable salt or solvate thereof for the manufacture of a medicament for the treatment of patients with rheumatoid arthritis, asthma, COPD, allergy and/or rhinitis.
  • a compound of formula (I) or a pharmaceutically acceptable salt or solvate thereof for the manufacture of a medicament for the treatment of patients with eczema, psoriasis, allergic dermatitis, neurodermatitis, pruritis and/or hypersensitivity reactions.
  • a method for the treatment of a human or animal subject with an inflammatory and/or allergic condition comprises administering to said human or animal subject an effective amount of a compound of formula (I) or a pharmaceutically acceptable salt or solvate thereof.
  • a method for the treatment of a human or animal subject with rheumatoid arthritis, asthma, COPD, allergy and/or rhinitis comprises administering to said human or animal subject an effective amount of a compound of formula (I) or a pharmaceutically acceptable salt or solvate thereof.
  • a method for the treatment of a human or animal subject with skin disease comprises administering to said human or animal subject an effective amount of a compound of formula (I) or a pharmaceutically acceptable salt or solvate thereof.
  • a method for the treatment of a human or animal subject with eczema, psoriasis, allergic dermatitis, neurodermatitis, pruritis and/or hypersensitivity reactions comprises administering to said human or animal subject an effective amount of a compound of formula (I) or a pharmaceutically acceptable salt or solvate thereof.
  • the compounds of formula (I) and pharmaceutically acceptable salts and solvates thereof may be formulated for administration in any convenient way, and the invention therefore also includes within its scope pharmaceutical compositions comprising a compound of formula (I) or a pharmaceutically acceptable salt or solvate thereof together, if desirable, in admixture with one or more physiologically acceptable diluents or carriers.
  • the compounds of formula (I) and pharmaceutically acceptable salts and solvates thereof may, for example, be formulated for oral, nasal, inhaled, buccal, sublingual, parenteral, topical rectal administration or other topical administration.
  • the compounds of formula (I) and pharmaceutically acceptable salts and solvates thereof may, for example, be formulated in conventional manner for oral, parenteral or rectal administration.
  • Formulations for oral administration include solutions, syrups, elixirs, powders, granules, tablets and capsules which typically contain conventional excipients such as binding agents, fillers, lubricants, disintegrants, wetting agents, suspending agents, emulsifying agents, preservatives, buffer salts, flavouring, colouring and/or sweetening agents as appropriate.
  • Dosage unit forms may be preferred as described below.
  • the active drug component can be combined with an oral, non-toxic pharmaceutically acceptable inert carrier such as ethanol, glycerol, water and the like.
  • the tablets may also contain excipients such as microcrystalline cellulose, lactose, sodium citrate, calcium carbonate, dibasic calcium phosphate and glycine, disintegrants such as starch (preferably corn, potato or tapioca starch), sodium starch glycollate, croscarmellose sodium and certain complex silicates, and granulation binders such as polyvinylpyrrolidone, hydroxypropylmethylcellulose (HPMC), hydroxypropylcellulose (HPC), sucrose, gelatin and acacia. Additionally, lubricating agents such as magnesium stearate, stearic acid, glyceryl behenate and talc may be included.
  • Solid compositions of a similar type may also be employed as fillers in gelatin capsules.
  • excipients in this regard include lactose, starch, a cellulose, milk sugar or high molecular weight polyethylene glycols.
  • the agent may be combined with various sweetening or flavouring agents, colouring matter or dyes, with emulsifying and/or suspending agents and with diluents such as water, ethanol, propylene glycol and glycerin, and combinations thereof.
  • Powders are prepared by comminuting the compound to a suitable fine size and mixing with a similarly comminuted pharmaceutical carrier such as an edible carbohydrate, as, for example, starch or mannitol.
  • Flavouring, preservative, dispersing and colouring agent can also be present.
  • Capsules can be made by preparing a powder mixture as described above, and filling formed gelatin sheaths.
  • Glidants and lubricants such as colloidal silica, talc, magnesium stearate, calcium stearate or solid polyethylene glycol can be added to the powder mixture before the filling operation.
  • a disintegrating or solubilizing agent such as agar- agar, calcium carbonate or sodium carbonate can also be added to improve the availability of the medicament when the capsule is ingested.
  • suitable binders include starch, gelatin, natural sugars such as glucose or beta-lactose, corn sweeteners, natural and synthetic gums such as acacia, tragacanth or sodium alginate, carboxymethylcellulose, polyethylene glycol, waxes and the like.
  • Lubricants used in these dosage forms include sodium oleate, sodium stearate, magnesium stearate, sodium benzoate, sodium acetate, sodium chloride and the like.
  • Disintegrators include, without limitation, starch, methyl cellulose, agar, bentonite, xanthan gum and the like.
  • Tablets are formulated, for example, by preparing a powder mixture, granulating or slugging, adding a lubricant and disintegrant and pressing into tablets.
  • a powder mixture is prepared by mixing the compound, suitably comminuted, with a diluent or base as described above, and optionally, with a binder such as carboxymethylcellulose, an aliginate, gelatin, or polyvinyl pyrrolidone, a solution retardant such as paraffin, a resorption accelerator such as a quaternary salt and/or an absorption agent such as bentonite, kaolin or dicalcium phosphate.
  • a binder such as carboxymethylcellulose, an aliginate, gelatin, or polyvinyl pyrrolidone
  • a solution retardant such as paraffin
  • a resorption accelerator such as a quaternary salt
  • an absorption agent such as bentonite, kaolin or dicalcium phosphate.
  • the powder mixture can be granulated by wetting with a binder such as syrup, starch paste, acadia mucilage or solutions of cellulosic or polymeric materials and forcing through a screen.
  • a binder such as syrup, starch paste, acadia mucilage or solutions of cellulosic or polymeric materials and forcing through a screen.
  • the powder mixture can be run through the tablet machine and the result is imperfectly formed slugs broken into granules.
  • the granules can be lubricated to prevent sticking to the tablet forming dies by means of the addition of stearic acid, a stearate salt, talc or mineral oil. The lubricated mixture is then compressed into tablets.
  • the compounds of the present invention can also be combined with free flowing inert carrier and compressed into tablets directly without going through the granulating or slugging steps
  • a clear or opaque protective coating consisting of a sealing coat of shellac, a coating of sugar or polymeric material and a polish coating of wax can be provided
  • Dyestuffs can be added to these coatings to distinguish different unit dosages
  • Oral fluids such as solutions, syrups and elixirs can be prepared in dosage unit form so that a given quantity contains a predetermined amount of the compound Syrups can be prepared by dissolving the compound in a suitably flavoured aqueous solution, while elixirs are prepared through the use of a non-toxic alcoholic vehicle Suspensions can be formulated by dispersing the compound in a non-toxic vehicle Solubilizers and emulsifiers such as ethoxylated isostearyl alcohols and polyoxy ethylene sorbitol ethers, preservatives, flavour additives such as peppermint oil or saccharin, and the like can also be added
  • dosage unit formulations for oral administration can be microencapsulated
  • the formulation can also be prepared to prolong or sustain the release as for example by coating or embedding particulate material in polymers, wax or the like
  • the compounds of formula (I) and pharmaceutically acceptable salts and solvates thereof can also be administered in the form of liposome delivery systems, such as small unilamellar vesicles, large unilamellar vesicles and multilamellar vesicles
  • liposome delivery systems such as small unilamellar vesicles, large unilamellar vesicles and multilamellar vesicles
  • Liposomes can be formed from a variety of phospholipids, such as cholesterol, stearylamine or phosphatidylcholines
  • the compounds of formula (I) and pharmaceutically acceptable salts and solvates thereof can also be administered in the form of liposome emulsion delivery systems, such as small unilamellar vesicles, large unilamellar vesicles and multilamellar vesicles
  • liposomes can be formed from a variety of phospholipids, such as cholesterol stearylamine or phosphatidylcholines
  • Topical administration includes administration by insufflation and inhalation
  • preparations for local administration include ointments, lotions, creams, gels, foams, preparations for delivery by transdermal patches, powders, sprays, aerosols, capsules or cartridges for use in an inhaler or insufflator or drops (e g eye or nose drops), solutions/suspensions for nebuhsation, suppositories, pessaries, retention enemas and chewable or suckable tablets or pellets (e g for the treatment of aphthous ulcers) or liposome or microencapsulation preparations
  • Formulations for administration topically to the nose for example, for the treatment of rhinitis include pressurised aerosol formulations and aqueous formulations administered to the nose by pressurised pump
  • Formulations which are non-pressurised and adapted to be administered topically to the nasal cavity are of particular interest Suitable formulations contain water as the diluent or carrier for
  • the compounds of formula (I) or a pharmaceutically acceptable salt or solvate thereof may be formulated for administration topically to the nose as a fluid formulation for delivery from a fluid dispenser, for example a fluid dispenser having a dispensing nozzle or dispensing orifice through which a metered dose of the fluid formulation is dispensed upon the application of a user-applied force to a pump mechanism of the fluid dispenser
  • a fluid dispenser for example a fluid dispenser having a dispensing nozzle or dispensing orifice through which a metered dose of the fluid formulation is dispensed upon the application of a user-applied force to a pump mechanism of the fluid dispenser
  • Such fluid dispensers are generally provided with a reservoir of multiple metered doses of the fluid formulation, the doses being dispensable upon sequential pump actuations
  • the dispensing nozzle or orifice may be configured for insertion into the nostrils of the user for spray dispensing of the fluid formulation into the nasal cavity
  • a fluid dispenser of the aforementioned type is described and illustrated
  • Ointments, creams and gels may, for example, be formulated with an aqueous or oily base with the addition of suitable thickening and/or gelling agent and/or solvents
  • bases may thus, for example, include water and/or an oil such as liquid paraffin or a vegetable oil such as arachis oil or castor oil, or a solvent such as polyethylene glycol Thickening agents and gelling agents which may be used according to the nature of the base include soft paraffin, aluminium stearate, cetostearyl alcohol, polyethylene glycols, woolfat, beeswax, carboxypolymethylene and cellulose derivatives, and/or glyceryl monostearate and/or non-ionic emulsifying agents.
  • Lotions may be formulated with an aqueous or oily base and will in general also contain one or more emulsifying agents, stabilising agents, dispersing agents, suspending agents or thickening agents.
  • Powders for external application may be formed with the aid of any suitable powder base, for example, talc, lactose or starch.
  • Drops may be formulated with an aqueous or non- aqueous base also comprising one or more dispersing agents, solubilising agents, suspending agents or preservatives.
  • the compounds of formula (I) and pharmaceutically acceptable salts and solvates thereof may be formulated as a dry powder for administration by inhalation.
  • a composition suitable for inhaled administration may be incorporated into a plurality of sealed dose containers (e.g. containing the dry powder composition) mounted longitudinally in a strip or ribbon inside a suitable inhalation device.
  • the container is rupturable or peel-openable on demand and the dose of e.g. the dry powder composition may be administered by inhalation via a device such as the DISKUSTM device, marketed by GlaxoSmithKline.
  • the DISKUSTM inhalation device is, for example, described in GB2242134A, and in such a device, at least one container for the composition in powder form (the container or containers preferably being a plurality of sealed dose containers mounted longitudinally in a strip or ribbon) is defined between two members peelably secured to one another; the device comprises: a means of defining an opening station for the said container or containers; a means for peeling the members apart at the opening station to open the container; and an outlet, communicating with the opened container, through which a user can inhale the composition in powder form from the opened container.
  • Spray compositions may for example be formulated as aqueous solutions or suspensions or as aerosols delivered from pressurised packs, such as a metered dose inhaler, with the use of a suitable liquefied propellant.
  • Aerosol compositions suitable for inhalation can be either a suspension or a solution and generally contain a compound of formula (I) and a suitable propellant such as a fluorocarbon or hydrogen-containing chlorofluorocarbon or mixtures thereof, particularly hydrofluoroalkanes, especially 1 ,1 ,1 ,2-tetrafluoroethane, 1,1 ,1 ,2,3,3,3-heptafluoro-n-propane or a mixture thereof.
  • the aerosol composition may optionally contain additional formulation excipients well known in the art such as surfactants e.g. oleic acid, lecithin or an oligolactic acid of derivative e.g. As described in WO94/21229 and WO98/34596 and cosolvents e.g. ethanol.
  • additional formulation excipients well known in the art such as surfactants e.g. oleic acid, lecithin or an oligolactic acid of derivative e.g. As described in WO94/21229 and WO98/34596 and cosolvents e.g. ethanol.
  • a pharmaceutical aerosol formulation comprising a compound of the invention and a fluorocarbon or hydrogen- containing chlorofluorocarbon as propellant, optionally in combination with a surfactant and/or a cosolvent.
  • a pharmaceutical aerosol formulation wherein the propellant is selected from 1 ,1 ,1 ,2-tetrafluoroethane, 1 ,1 ,1 ,2,3,3,3-heptafluoro-n-propane and mixtures thereof.
  • compositions of the invention may be buffered by the addition of suitable buffering agents.
  • Aerosol formulations are preferably arranged so that each metered dose or "puff of aerosol contains from 20 ⁇ g to 10mg, preferably from 20 ⁇ g to 2000 ⁇ g, more preferably from about 20 ⁇ g to 500 ⁇ g of a compound of formula (I). Administration may be once daily or several times daily, for example 2, 3, 4 or 8 times, giving for example 1 , 2 or 3 doses each time.
  • the overall daily dose with an aerosol will be within the range from 100 ⁇ g to 10mg, preferably from 200 ⁇ g to 2000 ⁇ g.
  • the overall daily dose and the metered dose delivered by capsules and cartridges in an inhaler or insufflator will generally be double that delivered with aerosol formulations.
  • the particle size of the particulate (e.g., micronised) drug should be such as to permit inhalation of substantially all the drug into the lungs upon administration of the aerosol formulation and will thus be less than 100 microns, desirably less than 20 microns, and in particular in the range of from 1 to 10 microns, such as from 1 to 5 microns, more preferably from 2 to 3 microns.
  • the formulations of the invention may be prepared by dispersal or dissolution of the medicament and a compound of the invention in the selected propellant in an appropriate container, for example, with the aid of sonication or a high-shear mixer.
  • the process is desirably carried out under controlled humidity conditions.
  • the chemical and physical stability and the pharmaceutical acceptability of the aerosol formulations according to the invention may be determined by techniques well known to those skilled in the art.
  • the chemical stability of the components may be determined by HPLC assay, for example, after prolonged storage of the product.
  • Physical stability data may be gained from other conventional analytical techniques such as, for example, by leak testing, by valve delivery assay (average shot weights per actuation), by dose reproducibility assay (active ingredient per actuation) and spray distribution analysis.
  • the stability of the suspension aerosol formulations according to the invention may be measured by conventional techniques, for example, by measuring flocculation size distribution using a back light scattering instrument or by measuring particle size distribution by cascade impaction or by the "twin impinger” analytical process.
  • twin impinger assay means "Determination of the deposition of the emitted dose in pressurised inhalations using apparatus A” as defined in British Pharmacopaeia 1988, pages A204-207, Appendix XVII C.
  • Such techniques enable the "respirable fraction" of the aerosol formulations to be calculated.
  • MDI canisters generally comprise a container capable of withstanding the vapour pressure of the propellant used such as a plastic or plastic-coated glass bottle or preferably a metal can, for example, aluminium or an alloy thereof which may optionally be anodised, lacquer-coated and/or plastic-coated (for example incorporated herein by reference WO96/32099 wherein part or all of the internal surfaces are coated with one or more fluorocarbon polymers optionally in combination with one or more non-fluorocarbon polymers), which container is closed with a metering valve.
  • the cap may be secured onto the can via ultrasonic welding, screw fitting or crimping.
  • MDIs taught herein may be prepared by methods of the art (e.g.
  • the canister is fitted with a cap assembly, wherein a drug-metering valve is situated in the cap, and said cap is crimped in place.
  • a drug-metering valve is situated in the cap, and said cap is crimped in place.
  • MDI means a unit comprising a can, a secured cap covering the can and a formulation metering valve situated in the cap.
  • MDI system includes a suitable channelling device. Suitable channelling devices comprise for example, a valve actuator and a cylindrical or cone-like passage through which medicament may be delivered from the filled canister via the metering valve to the nose or mouth of a patient such as a mouthpiece actuator.
  • the metallic internal surface of the can is coated with a fluoropolymer, most preferably blended with a non-fluoropolymer.
  • the metallic internal surface of the can is coated with a polymer blend of polytetrafluoroethylene (PTFE) and polyethersulfone (PES).
  • the whole of the metallic internal surface of the can is coated with a polymer blend of polytetrafluoroethylene (PTFE) and polyethersulfone (PES).
  • the metering valves are designed to deliver a metered amount of the formulation per actuation and incorporate a gasket to prevent leakage of propellant through the valve.
  • the gasket may comprise any suitable elastomeric material such as, for example, low density polyethylene, chlorobutyl, bromobutyl, EPDM, black and white butadiene- acrylonitrile rubbers, butyl rubber and neoprene.
  • Suitable valves are commercially available from manufacturers well known in the aerosol industry, for example, from Valois, France (e.g. DF10, DF30, DF60), Bespak pic, UK (e.g. BK300, BK357) and 3M-
  • Neotechnic Ltd UK (e.g. Spraymiser ).
  • the MDIs may also be used in conjunction with other structures such as, without limitation, overwrap packages for storing and containing the MDIs, including those described in U.S. Patent Nos. 6,1 19,853; 6, 179,1 18; 6,315,1 12; 6,352,152; 6,390,291 ; and 6,679,374, as well as dose counter units such as, but not limited to, those described in U.S. Patent Nos. 6,360,739 and 6,431 ,168.
  • overwrap packages for storing and containing the MDIs, including those described in U.S. Patent Nos. 6,1 19,853; 6, 179,1 18; 6,315,1 12; 6,352,152; 6,390,291 ; and 6,679,374, as well as dose counter units such as, but not limited to, those described in U.S. Patent Nos. 6,360,739 and 6,431 ,168.
  • a metering valve is crimped onto an aluminium can to form an empty canister.
  • the particulate medicament is added to a charge vessel and liquefied propellant together with the optional excipients is pressure filled through the charge vessel into a manufacturing vessel.
  • the drug suspension is mixed before recirculation to a filling machine and an aliquot of the drug suspension is then filled through the metering valve into the canister.
  • a metering valve is crimped onto an aluminium can to form an empty canister.
  • the liquefied propellant together with the optional excipients and the dissolved medicament is pressure filled through the charge vessel into a manufacturing vessel.
  • an aliquot of the liquefied formulation is added to an open canister under conditions which are sufficiently cold to ensure the formulation does not vaporise, and then a metering valve crimped onto the canister.
  • each filled canister is check- weighed, coded with a batch number and packed into a tray for storage before release testing.
  • Capsules and cartridges for use in an inhaler or insufflator may be formulated containing a powder mix for inhalation of a compound of the invention and a suitable powder base such as lactose or starch.
  • a powder mix for inhalation of a compound of the invention and a suitable powder base such as lactose or starch.
  • Each capsule or cartridge may generally contain from 20 ⁇ g to 10mg of the compound of formula (I) or a pharmaceutically acceptable salt or solvate thereof.
  • the compound of the invention may be presented without excipients such as lactose.
  • the proportion of the active compound of formula (I) or a pharmaceutically acceptable salt or solvate thereof in the local compositions according to the invention depends on the precise type of formulation to be prepared but will generally be within the range of from 0.001 to 10% by weight. Generally, for most types of preparations, the proportion used will be within the range of from 0.005 to 1%, for example from 0.01 to 0.5%. However, in powders for inhalation or insufflation the proportion used will normally be within the range of from 0.1 to 5%.
  • Topical preparations may be administered by one or more applications per day to the affected area; over skin areas occlusive dressings may advantageously be used. Continuous or prolonged delivery may be achieved by an adhesive reservoir system.
  • a physician will determine the actual dosage which will be most suitable for an individual subject.
  • the specific dose level and frequency of dosage for any particular individual may be varied and will depend upon a variety of factors including the activity of the specific compound employed, the metabolic stability and length of action of that compound, the age, body weight, general health, sex, diet, mode and time of administration, rate of excretion, drug combination, the severity of the particular condition, and the individual undergoing therapy.
  • the daily dosage level of the agent may be in single or divided doses.
  • the daily dose as employed for adult human treatment will range from 0.5-100mg/kg body weight, preferably 0.5-60mg/kg body weight, which may be administered in 1 to 4 daily doses, for example, depending on the route of administration and the condition of the patient.
  • each unit will preferably contain 5mg to 1g of active ingredient.
  • the duration of treatment will be dictated by the rate of response rather than by arbitrary numbers of days.
  • the compounds of the invention may in general be given by internal administration in cases wherein systemic glucocorticoid receptor agonist therapy is indicated.
  • Slow release or enteric coated formulations may be advantageous, particularly for the treatment of inflammatory bowel disorders.
  • the compounds of the invention will be formulated for oral administration. In other embodiments, the compounds of the invention will be formulated for inhaled administration. In further embodiments, the compounds of the invention will be formulated for intranasal administration.
  • the compounds and pharmaceutical formulations according to the invention may be used in combination with or include one or more other therapeutic agents, for example when the compounds of the invention are administered intranasally or by inhalation.
  • Suitable other therapeutic agents may be selected from for example anti-inflammatory agents, anticholinergic agents (particularly an M 1 ZM 2 ZM 3 receptor antagonist), ⁇ 2 -adrenoreceptor agonists, antiinfective agents such as antibiotics or antivirals, or antihistamines.
  • the invention thus provides, in a further aspect, a combination comprising a compound of the invention together with one or more other therapeutically active agents, for example selected from an anti-inflammatory agent such as a corticosteroid or an NSAID, an anticholinergic agent, a ⁇ 2 -adrenoreceptor agonist, an antiinfective agent such as an antibiotic or an antiviral, or an antihistamine.
  • an anti-inflammatory agent such as a corticosteroid or an NSAID
  • an anticholinergic agent such as a corticosteroid or an NSAID
  • an anticholinergic agent such as a corticosteroid or an NSAID
  • a ⁇ 2 -adrenoreceptor agonist such as an antibiotic or an antiviral
  • an antihistamine such as an antibiotic or an antiviral
  • One embodiment of the invention encompasses combinations comprising a compound of the invention together with a ⁇ 2 - adrenoreceptor agonist, and/or
  • One embodiment of the invention encompasses combinations comprising one or two other therapeutic agents.
  • the other therapeutic ingredient(s) may be used in the form of salts, for example as alkali metal or amine salts or as acid addition salts, or prodrugs, or as esters, for example lower alkyl esters, or as solvates, for example hydrates, to optimise the activity and/or stability and/or physical characteristics, such as solubility, of the therapeutic ingredient. It will be clear also that, where appropriate, the therapeutic ingredients may be used in optically pure form.
  • the invention encompasses a combination comprising a compound of the invention together with a ⁇ 2 -adrenoreceptor agonist.
  • ⁇ 2 -adrenoreceptor agonists examples include salmeterol (e.g. as the racemate or a single enantiomer, such as the f?-enantiomer), salbutamol (e.g. as the racemate or a single enantiomer such as the f?-enantiomer), formoterol (e.g.
  • the ⁇ 2 -adrenoreceptor agonists are long-acting ⁇ 2 -adrenoreceptor agonists, for example compounds which provide effective bronchodilation for about 12 hours or longer.
  • ⁇ 2 -adrenoreceptor agonists may include those described in WO02/066422A, WO02/070490, WO02/076933, WO03/024439, WO03/072539, WO 03/091204, WO04/016578, WO04/022547, WO04/037807, WO04/037773, WO04/037768, WO04/039762, WO04/039766, WO01/42193 and WO03/042160.
  • ⁇ 2 -adrenoreceptor agonists examples include:
  • the ⁇ 2 -adrenoreceptor agonist may be in the form of a salt formed with a pharmaceutically acceptable acid selected from sulphuric, hydrochloric, fumaric, hydroxynaphthoic (for example 1- or 3-hydroxy-2-naphthoic), cinnamic, substituted cinnamic, triphenylacetic, sulphamic, sulphanilic, naphthaleneacrylic, benzoic, 4-methoxybenzoic, 2- or 4-hydroxybenzoic, 4-chlorobenzoic and 4-phenylbenzoic acid.
  • a pharmaceutically acceptable acid selected from sulphuric, hydrochloric, fumaric, hydroxynaphthoic (for example 1- or 3-hydroxy-2-naphthoic), cinnamic, substituted cinnamic, triphenylacetic, sulphamic, sulphanilic, naphthaleneacrylic, benzoic, 4-
  • Suitable anti-inflammatory agents include corticosteroids.
  • corticosteroids which may be used in combination with the compounds of the invention are those oral and inhaled corticosteroids and their pro-drugs which have anti-inflammatory activity. Examples include methyl prednisolone, prednisolone, dexamethasone, fluticasone propionate, 6 ⁇ ,9 ⁇ -difluoro-11 ⁇ -hydroxy-16 ⁇ -methyl-17 ⁇ -[(4-methyl-1 ,3-thiazole-5- carbonyl)oxy]-3-oxo-androsta-1 ,4-diene-17 ⁇ -carbothioic acid S-fluoromethyl ester, 6 ⁇ ,9 ⁇ - difluoro-17 ⁇ -[(2-furanylcarbonyl)oxy]-11 ⁇ -hydroxy-16 ⁇ -methyl-3-oxo-androsta-1 ,4-diene- 17 ⁇ -carbothioic acid S-fluoromethyl ester (fluticasone furoate), 6 ⁇ ,9 ⁇ -difluor
  • corticosteroids include fluticasone propionate, 6 ⁇ ,9 ⁇ -difluoro-11 ⁇ -hydroxy-16 ⁇ -methyl-17 ⁇ -[(4-methyl-1 ,3- thiazole-5-carbonyl)oxy]-3-oxo-androsta-1 ,4-diene-17 ⁇ -carbothioic acid S-fluoromethyl ester, ⁇ . ⁇ -difluoro- ⁇ -f ⁇ -furanylcarbony ⁇ oxyl-i i ⁇ -hydroxy-i ⁇ -methyl-S-oxo- androsta-1 ,4-diene-17 ⁇ -carbothioic acid S-fluoromethyl ester, 6 ⁇ ,9 ⁇ -difluoro-11 ⁇ -hydroxy- 16 ⁇ -methyl-3-oxo-17 ⁇ -(2,2,3,3- tetramethycyclopropylcarbonyOoxy-androsta-i ,4-diene- 17 ⁇ -carbothioic acid S-cyanomethyl ester and 6 ⁇ ,9 ⁇ ,9
  • the corticosteroid is 6 ⁇ ,9 ⁇ -difluoro-17 ⁇ -[(2- furanylcarbonyl)oxy]-11 ⁇ -hydroxy-16 ⁇ -methyl-3-oxo-androsta-1 ,4-diene-17 ⁇ -carbothioic acid S-fluoromethyl ester.
  • corticosteroids may include those described in WO02/088167, WO02/100879, WO02/12265, WO02/12266, WO05/005451 , WO05/005452, WO06/072599 and WO06/072600.
  • Non-steroidal compounds having glucocorticoid agonism that may possess selectivity for transrepression over transactivation and that may be useful in combination therapy include those covered in the following published patent applications and patents: WO2003/082827, WO1998/54159, WO2004/005229, WO2004/009017, WO2004/018429, WO2003/104195, WO2003/082787, WO2003/082280, WO2003/059899,
  • anti-inflammatory agents include non-steroidal anti-inflammatory drugs (NSAID's)
  • NSAID's include sodium cromoglycate, nedocromil sodium, phosphodiesterase (PDE) inhibitors (for example, theophylline, PDE4 inhibitors or mixed PDE3/PDE4 inhibitors), leukotriene antagonists, inhibitors of leukotriene synthesis (for example, montelukast), iNOS inhibitors, tryptase and elastase inhibitors, beta-2 integrin antagonists and adenosine receptor agonists or antagonists (for example, adenosine 2a agonists), cytokine antagonists (for example, chemokine antagonists, such as a CCR3 antagonist) or inhibitors of cytokine synthesis, or 5-l ⁇ poxygenase inhibitors
  • PDE phosphodiesterase
  • iNOS inhibitors inhibitors of leukotriene synthesis
  • tryptase and elastase inhibitors beta-2 integrin antagonists and adenosine
  • the invention provides the use of the compounds of the invention in combination with a phosphodiesterase 4 (PDE4) inhibitor, for example in the case of a formulation adapted for inhalation
  • PDE4-spec ⁇ f ⁇ c inhibitor may be any compound that is known to inhibit the PDE4 enzyme or which is discovered to act as a PDE4 inhibitor, and which are only PDE4 inhibitors, not compounds which inhibit other members of the PDE family, such as PDE3 and PDE5, as well as PDE4
  • Compounds include c/s-4-cyano-4-(3-cyclopentyloxy-4-methoxyphenyl)cyclohexan-1- carboxylic acid, 2-carbomethoxy-4-cyano-4-(3-cyclopropylmethoxy-4- d ⁇ fluoromethoxyphenyl)cyclohexan-1-one and c/s-[4-cyano-4-(3-cyclopropylmethoxy-4- d ⁇ fluoromethoxyphenyl)cyclohexan-1-ol]
  • Another compound is c/s-4-cyano-4-[3- (cyclopentyloxy)-4-methoxyphenyl]cyclohexane-1-carboxyl ⁇ c acid (also known as cilomilast) and its salts, esters, pro-drugs or physical forms, which is described in U S patent 5,552,438 issued 03 September, 1996, this patent and the compounds it discloses are incorporated herein in full by reference
  • AWD-12-281 N-(3,5-d ⁇ chloro-4-py ⁇ d ⁇ nyl)-1-[4- fluorophenyl)methyl]-5-hydroxy- ⁇ -oxo-1 H- ⁇ ndol-3-acetam ⁇ de
  • Elbion Hofgen, N et al 15th EFMC lnt Symp Med Chem (Sept 6-10, Edinburgh) 1998, Abst P 98, CAS reference No 247584020-9
  • INSERM 9-benzyladen ⁇ ne derivative nominated NCS-613
  • D-4418 from Chiroscience and Schering-Plough
  • a benzodioxole derivative disclosed by Kyowa Hakko in WO99/16766
  • K-34 from Kyowa Hakko
  • V- 11294A from Napp (Landells, LJ.
  • anticholinergic agents are those compounds that act as antagonists at the muscarinic receptors, in particular those compounds which are antagonists of the M 1 or M 3 receptors, dual antagonists of the M 1 ZM 3 or M 2 /M 3 , receptors or pan-antagonists of the M 1 ZM 2 ZM 3 receptors.
  • exemplary compounds for administration via inhalation include ipratropium (for example, as the bromide, CAS 22254-24-6, sold under the name Atrovent), oxitropium (for example, as the bromide, CAS 30286-75-0) and tiotropium (for example, as the bromide, CAS 136310-93-5, sold under the name Spiriva).
  • revatropate for example, as the hydrobromide, CAS 262586-79-8) and LAS- 34273 which is disclosed in WO01Z04118.
  • Exemplary compounds for oral administration include pirenzepine (for example, CAS 28797-61-7), darifenacin (for example, CAS 133099-04-4, or CAS 133099-07-7 for the hydrobromide sold under the name Enablex), oxybutynin (for example, CAS 5633-20-5, sold under the name Ditropan), terodiline (for example, CAS 15793-40-5), tolterodine (for example, CAS 124937-51-5, or CAS 124937- 52-6 for the tartrate, sold under the name Detrol), otilonium (for example, as the bromide, CAS 26095-59-0, sold under the name Spasmomen), trospium chloride (for example, CAS 10405-02-4) and solifenacin (for
  • anticholinergic agents include compounds of formula (A) 1 which are disclosed in US patent application 60/487981 :
  • R 31 and R 32 are, independently, selected from the group consisting of straight or branched chain lower alkyl groups having preferably from 1 to 6 carbon atoms, cycloalkyl groups having from 5 to 6 carbon atoms, cycloalkyl-alkyl having from 6 to 10 carbon atoms, 2- thienyl, 2-pyridyl, phenyl, phenyl substituted with an alkyl group having not in excess of 4 carbon atoms and phenyl substituted with an alkoxy group having not in excess of 4 carbon atoms;
  • X " represents an anion associated with the positive charge of the N atom.
  • X " may be but is not limited to chloride, bromide, iodide, sulfate, benzene sulfonate, and toluene sulfonate, including, for example: (3-endo)-3-(2,2-di-2-thienylethenyl)-8,8-dimethyl-8-azoniabicyclo[3.2.1]octane bromide;
  • R 41" represents an anion associated with the positive charge of the N atom;
  • R 41' may be but is not limited to chloride, bromide, iodide, sulfate, benzene sulfonate and toluene sulfonate;
  • R 42 and R 43 are independently selected from the group consisting of straight or branched chain lower alkyl groups (having preferably from 1 to 6 carbon atoms), cycloalkyl groups
  • R 44 is selected from the group consisting of (C r C 6 )alkyl, (C 3 -C 12 )cycloalkyl, (C 3 -
  • C 7 heterocycloalkyl, (C 1 -C 6 )alkyl(C 3 -C 12 )cycloalkyl, (d-CeJalkyKdrdJheterocycloalkyl, aryl, heteroaryl, (d-Ce)alkyl-aryl, (d-C 6 )alkyl-heteroaryl, -OR 45 , -CH 2 OR 45 , -CH 2 OH, -CN,
  • R 45 is selected from the group consisting of (Ci-C ⁇ jalkyl, (C 1 -CeJaIkVl(C 3 -C 12 )CyClOaIkVl, (C 1 -C 6 )alkyl(C 3 -C 7 )heterocycloalkyl, (d-C 6 )alkyl-aryl, (d-C 6 )alkyl-heteroaryl;
  • R 46 is selected from the group consisting of (d-C 6 )alkyl, (C 3 -Ci 2 )cycloalkyl, (C 3 -
  • C 7 heterocycloalkyl, (Ci-C 6 )alkyl(C 3 -C 12 )cycloalkyl, (d-CeJalky ⁇ Cs-CyJheterocycloalkyl, aryl, heteroaryl, (d-C 6 )alkyl-aryl, (d-C 6 )alkyl-heteroaryl;
  • R 47 and R 48 are, independently, selected from the group consisting of H, (d-C 6 )alkyl, (C 3 - Ci 2 )cycloalkyl, (C 3 -C 7 )heterocycloalkyl, (Ci-C 6 )alkyl(C 3 -C 12 )cycloalkyl, (C r C 6 )alkyl(C 3 -
  • C 7 heterocycloalkyl, (C 1 -C 6 )alkyl-aryl, and (d-C 6 )alkyl-heteroaryl, including, for example: (endo)-3-(2-methoxy-2,2-di-thiophen-2-yl-ethyl)-8,8-dimethyl- ⁇ -azonia- bicyclo[3.2.1]octane iodide;
  • antihistamines include any one or more of the numerous antagonists known which inhibit H 1 -receptors, and are safe for human use
  • First generation antagonists include derivatives of ethanolammes, ethylenediamines, and alkylamines, such as diphenylhydramine, py ⁇ lamine, clemastine, chlorpheniramine
  • Second generation antagonists which are non-sedating, include loratidine, desloratidine, terfenadine, astemizole, acrivastine, azelastine, levoceti ⁇ zine fexofenadine and cetirizine
  • antihistamines examples include loratidine, desloratidine, fexofenadine, cetirizine, levocabastine, olopatadine, amlexanox and epinastine
  • H1 antagonists include, without limitation, amelexanox, astemizole, azatadine, azelastine, acrivastine, brompheniramine, cetirizine, levocetirizine, efleti ⁇ zine, chlorpheniramine, clemastine, cyclizine, carebastine, cyproheptadine, carbinoxamine, descarboethoxyloratadine, doxylamine, dimethindene, ebastine, epinastine, efletinzine, fexofenadine, hydroxyzine, ketotifen, loratadine, levocabastine, mizolastine, mequitazine, mianserin, noberastine, meclizine, norastemizole, olopatadine, picumast, pyn
  • the invention thus provides, in a further aspect, a combination comprising a compound of the invention together with a ⁇ 2 -adrenoreceptor agonist.
  • the invention thus provides, in a further aspect, a combination comprising a compound of the invention together with a corticosteroid.
  • the invention thus provides, in a further aspect, a combination comprising a compound of the invention together with another non-steroidal GR agonist.
  • the invention thus provides, in a further aspect, a combination comprising a compound of the invention together with an anticholinergic.
  • the invention thus provides, in a further aspect, a combination comprising a compound of the invention together with an antihistamine.
  • the invention thus provides, in a further aspect, a combination comprising a compound of the invention together with an H1 antagonist.
  • the invention thus provides, in a further aspect, a combination comprising a compound of the invention together with an H1/H3 antagonist.
  • the invention thus provides, in a further aspect, a combination comprising a compound of the invention together with a PDE4 inhibitor and a ⁇ 2 -adrenoreceptor agonist.
  • the invention thus provides, in a further aspect, a combination comprising a compound of the invention together with an anticholinergic and a PDE-4 inhibitor.
  • compositions comprising a combination as defined above together with a pharmaceutically acceptable diluent or carrier represent a further aspect of the invention.
  • the individual compounds of such combinations may be administered either sequentially or simultaneously in separate or combined pharmaceutical formulations In one embodiment, the individual compounds may be administered simultaneously in a combined pharmaceutical formulation
  • Appropriate doses of known therapeutic agents will be readily appreciated by those skilled in the art
  • the invention thus provides, in a further aspect, a pharmaceutical composition
  • a pharmaceutical composition comprising a combination of a compound of the invention together with another therapeutically active agent
  • the invention thus provides, in a further aspect, a pharmaceutical composition
  • a pharmaceutical composition comprising a combination of a compound of the invention together with a PDE4 inhibitor
  • the invention thus provides, in a further aspect, a pharmaceutical composition
  • a pharmaceutical composition comprising a combination of a compound of the invention together with a ⁇ 2 -adrenoreceptor agonist
  • the invention thus provides, in a further aspect, a pharmaceutical composition
  • a pharmaceutical composition comprising a combination of a compound of the invention together with a corticosteroid
  • the invention thus provides, in a further aspect, a pharmaceutical composition
  • a pharmaceutical composition comprising a combination of a compound of the invention together with another non-steroidal GR agonist
  • the invention thus provides, in a further aspect, a pharmaceutical composition
  • a pharmaceutical composition comprising a combination of a compound of the invention together with an anticholinergic
  • the invention thus provides, in a further aspect, a pharmaceutical composition
  • a pharmaceutical composition comprising a combination of a compound of the invention together with an antihistamine
  • the invention thus provides, in a further aspect, a pharmaceutical composition
  • a pharmaceutical composition comprising a combination of a compound of the invention together with an H1 antagonist
  • the invention thus provides, in a further aspect, a pharmaceutical composition comprising a combination of a compound of the invention together with an H1/H3 antagonist
  • a pharmaceutical composition comprising a combination of a compound of the invention together with a PDE4 inhibitor and a ⁇ 2 - adrenoreceptor agonist.
  • the invention thus provides, in a further aspect, a pharmaceutical composition
  • a pharmaceutical composition comprising a combination of a compound of the invention together with an anticholinergic and a PDE- 4 inhibitor.
  • a process according to the invention for the preparation of compounds of formula (I) comprises reaction of an amine of formula (II)
  • X and n are as defined above for compounds of formula (I) and R 4 is hydrogen or a protecting group such as 1 ,1-dimethylethyl or terf-butyldimethylsilyl, with a compound of formula
  • R 1 , R 2 and R 3 are as defined above for compounds of formula (I) and Y is chlorine or hydroxyl.
  • the reaction may be carried out in a conventional organic solvent, for example tetrahydrofuran, in the presence of a base, for example potassium carbonate, triethylamine, pyridine or diisopropylethylamine.
  • a base for example potassium carbonate, triethylamine, pyridine or diisopropylethylamine.
  • the reaction is carried out in the presence of diisopropylethylamine.
  • the reaction may be carried out at a temperature of from -10°C to 100°C, for example at room temperature.
  • the reaction when Y is hydroxyl, the reaction may be carried out in a conventional organic solvent, for example dimethylformamide, in the presence of a coupling agent such as those described in Tetrahedron 2005, 61 , 10827, for example O-(7-azabenzotriazol-1- yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (HATU) 1 and a base, for example triethylamine or diisopropylethylamine.
  • HATU O-(7-azabenzotriazol-1- yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate
  • a base for example triethylamine or diisopropylethylamine.
  • the reaction is carried out in the presence of diisopropylethylamine.
  • the reaction may be carried out at a temperature of from -10°C to 100°C, for
  • R 4 is a protecting group
  • the compound of formula (I) is obtained by deprotection of the protected intermediate.
  • the group may be removed by dissolving the protected intermediate in an organic solvent, for example dichloromethane, and treating with an organic acid, for example trifluoroacetic acid.
  • the reaction may be carried out at a temperature of from - 10°C to 100°C, for example at room temperature.
  • the silyl protecting group may be removed with a fluoride source such as tetrabutylammonium fluoride in tetrahydrofuran.
  • the reaction may be carried out at a temperature of from -10°C to 100 0 C, for example at room temperature.
  • X and n are as defined above for compounds of formula (I), with 2-hydroxyethylamine, or a protected derivative thereof such as 2-t-butoxyethylamine or 2-tert-butyldimethyls ⁇ lyloxyethylamine.
  • the reaction may be carried out in a conventional organic solvent, for example acetonitrile or tetrahydrofuran, and at a temperature of from -10 0 C to 100°C, for example at room temperature.
  • a compound of formula (IV) may be prepared by treating a compound of formula (V)
  • reaction may be carried out in a conventional organic solvent, for example tetrahydrofuran.
  • the reaction may be carried out at a temperature of from -10°C to 100°C, for example at room temperature.
  • a compound of formula (V) may be prepared by treating a compound of formula (Vl)
  • reaction may be carried out in a conventional organic solvent, for example dichloromethane, in the presence of an organic base, for example pyridine.
  • organic base for example pyridine.
  • the reaction may be carried out at a temperature of from - 10°C to 100 0 C, for example at room temperature.
  • a compound of formula (Vl) may be prepared by reacting a compound of formula (VII)
  • the reaction may be carried out in a conventional organic solvent, for example dimethylformamide, in the presence of a coupling agent such as those described in Tetrahedron 2005, 61 , 10827, for example O-(7-azabenzotriazol-1-yl)-N,N,N',N'- tetramethyluronium hexafluorophosphate (HATU), and a base, for example triethylamine or diisopropylethylamine.
  • HATU O-(7-azabenzotriazol-1-yl)-N,N,N',N'- tetramethyluronium hexafluorophosphate
  • a base for example triethylamine or diisopropylethylamine.
  • the reaction is carried out in the presence of diisopropylethylamine.
  • the reaction may be carried out at a temperature of from -10 0 C to 100°C, for example at room temperature.
  • Example of acids of formula (VII) suitable for use in this coupling reaction include:
  • Acids of formula (VII) may be prepared by, for example, reaction of a suitable aryl hydrazine with ethyl 2-cyano-3-ethoxyacrylate followed by conversion of the resulting ethyl ester to the corresponding acid by treatment with, for example, lithium hydroxide in a solvent such as aqueous ethanol.
  • a compound of formula (VIII) may be prepared by treating a compound of formula (IX)
  • reaction with a transition metal catalyst, for example palladium hydroxide on carbon, in the presence of a hydrogen atmosphere.
  • a transition metal catalyst for example palladium hydroxide on carbon
  • the reaction may be carried out in a conventional organic solvent, for example ethanol.
  • the reaction may be carried out at a temperature of from -1O 0 C to 100°C, for example at room temperature.
  • a compound of formula (IX) may be prepared by treating a compound of formula (X)
  • benzylamine followed by treatment with a base, for example sodium hydroxide.
  • a base for example sodium hydroxide.
  • the reaction may be carried out in a conventional organic solvent, for example 1 ,4-dioxan.
  • the treatment with benzylamine may be carried out at a temperature of from -10°C to 100 0 C 1 for example at room temperature, and the treatment with base may be carried out at a temperature of from -10 0 C to 100 0 C, for example at about 9O 0 C.
  • a compound of formula (X) may be prepared by treating a compound of formula (Xl)
  • the reaction may be carried out in a conventional organic solvent, for example dichloromethane. Batch processes or flow processes are suitable equipment for this cyclisation.
  • the reaction may be carried out at a temperature of from -10°C to 100°C, for example at room temperature for a batch process or at about 50 0 C for a flow process.
  • a compound of formula (Xl) may be prepared by treating a compound of formula (XII)
  • a compound of formula (XII) may be prepared by treating a compound of formula (XIII)
  • reaction with a transition metal catalyst, for example 5% palladium on carbon, in the presence of a hydrogen atmosphere.
  • a transition metal catalyst for example 5% palladium on carbon
  • the reaction may be carried out in a conventional organic solvent, for example ethanol.
  • the reaction may be carried out at a temperature of from -10°C to 100 0 C, for example at room temperature for a batch process or at about 80°C for a flow process. Batch processes or flow processes are suitable for this hydrogenation.
  • a compound of formula (XIII) may be prepared by treating a compound of formula (XIV)
  • reaction with trimethyl(trifluoromethyl)silane and tetra-n-butylammonium fluoride.
  • the reaction may be carried out in a conventional organic solvent, for example tetrahydrofuran or dichloromethane.
  • the reaction may be carried out at a temperature of from -10°C to 100 0 C 1 for example at 0 0 C rising to room temperature. Batch processes or flow processes are suitable for this transformation.
  • a compound of formula (XIV) may be prepared by oxidation of 1 ,3-dibenzylglycerol.
  • the oxidation may be carried out using 3A molecular sieves, N- methylmorpholine ⁇ /-oxide and tetrapropylammonium perruthenate in dichloromethane at
  • the oxidation may be carried out using aqueous sodium hypochlorite, saturated sodium bicarbonate solution and 2,2,6,6-tetramethyl-i-piperidinyloxy free radical in toluene at 0 0 C to 50°C, for example at room temperature.
  • the oxidation may be carried out using sulphur trioxide-pyridine complex in the presence of base such as triethylamine in dimethylsulphoxide at 10 0 C to 50°C, for example at room temperature. Batch processes or flow processes are suitable for this oxidation.
  • Certain compounds of formulae (II), (III), (IV), (V), (Vl) 1 (VIII), (IX), (X), (Xl) and (XIII) may be new and form an aspect of the present invention.
  • Compounds of formula (I) may be prepared in the form of mixtures of enantiomers when mixtures of isomers are used as intermediates in the synthesis.
  • a compound of formula (II) as a racemic mixture of enantiomers will lead to a mixture of enantiomers in the final product.
  • These isomers may, if desired, be separated by conventional methods (e.g. HPLC on a chiral column).
  • separation of isomers may be performed earlier in the synthesis, for example individual isomers of compounds of formula (II) or earlier stage intermediates may be employed which may obviate the need to perform a separation of isomers as a final stage in the synthesis.
  • the later process is, in theory, more efficient and is therefore preferred.
  • compositions comprising a compound of the invention also constitute an aspect of the invention.
  • Solvates of compounds of formula (I) or salts thereof, which are not pharmaceutically acceptable may be useful as intermediates in the preparation of other compounds of formula (I) and salts and solvates thereof.
  • Flashmaster 2 is an automated multi user flash chromatography system which utilises disposable SPE cartridges (2g to 10Og) It provides quaternary on-line solvent mixing to enable gradient methods to be run Samples are queued using the multi functional open access software which manages flow rates, gradient profile and collection conditions
  • the system is equipped with a Knauer variable wavelength uv detector and 2 Gilson FC204 fraction collectors enabling automated peak cutting, collection and tracking
  • Aqueous solvent Water + 0 1 % TFA
  • Organic solvent MeCN + 0 1 % TFA
  • Circular dichroism was carried out on an Applied Photophysics Chirascan spectrophotometer at room temperature, using acetonitrile as solvent, over the range 200-
  • the title compound was prepared via a 'flow' process using the following starting materials and solvents.
  • the title compound was prepared via a CPC Cytos Lab System made up of a 47ml reactor block with two Jasco PU - 2080Plus HPLC pumps. Reactor temperature was maintained at 60 0 C via a Huber Unistat 360 unit.
  • Solution A 1 ,3-dibenzyloxy-2-propanol 12Og, 440mmol) in acetonitrile (489ml).
  • Solution B - tetrapropylammonium perruthenate (7.72g, 22mmol, 5mol%) and N-methylmorpholine N-oxide (87.5g, 748mmol) in acetonitrile (611ml).
  • Solutions A and B were pumped through the Cytos Lab system in the ratio of solution A to solution B of 1 : 1.25 with a total flow rate of 7.8ml/min and residence time of 6 minutes. This gave a total reaction time of 2 hours 21 minutes. The total reacted solution was split equally into 2 batches and each was concentrated in vacuo.
  • the reaction mixture was stirred for an additional 5 minutes and then washed with 1M aqueous hydrochloric acid (2 x 15ml), saturated sodium bicarbonate (15ml) and 1 %w/v aqueous sodium chloride solution (2 x 15ml).
  • the organic extract was concentrated in vacuo to give 2.5g of the desired product as dark oil in 99.3% yield.
  • the NMR spectrum of the product was concordant with a reference sample.
  • Tetrabutylammonium fluoride trihydrate (TBAF 3H 2 O) (2.9g, 0.5 equivalent) was dissolved in THF (5ml). This was added cautiously to a stirred and cooled (+15 0 C) solution of 1 ,3- dibenzyloxy-2-propanone in toluene (24.65g, equivalent to 5g of the ketone) and (trifluoromethyl)trimethylsilane (7.5ml). There was an exotherm and lot of gas evolution on addition of the first 1ml of TBAF solution. The temperature rose from 18 to 4O 0 C.
  • the TBAF addition was carried out over 3 minutes and then the mixture was stirred at 15-3O 0 C for a further 2 minutes and then cooled to +10 0 C while carrying out an HPLC analysis.
  • the reaction mixture was sequentially washed with 1 M aqueous hydrochloric acid (50ml), 1% aqueous sodium chloride solution (2 x 25ml) and a mixture of 1 % sodium chloride (25ml) and saturated sodium bicarbonate (5ml) solution.
  • the separated organic extract was concentrated in vacuo to give 6.41 g of the desired product as dark brown oil in 101.8% yield.
  • the NMR spectrum showed the presence of residual toluene (8.8%) and starting material (ca 3%).
  • the title compound was prepared via a 'flow' process using the following starting materials and solvents.
  • the title compound was prepared via a CPC Cytos Lab System made up of a 32ml reactor block with two Jasco PU - 2080Plus HPLC pumps. Reactor temperature was maintained at 22°C via a Huber Unistat 360 unit. The reactor outlet was fitted with a IOOpsi backflow regulator. Two solutions were prepared. Solution A - 1 ,3-bis[(phenylmethyl)oxy]-2-propanone (71.64g, 265mmol) and trimethyl(trifluoromethyl)silane (86.67g, 96ml, 609.5mmol) in tetrahydrofuran(99ml). Solution B - tetrabutylammonium fluoride (1 M in THF, 265ml, 132.5mmol).
  • TMS-CF 3 Trimethyl(trifluoromethyl)silane
  • the title compound was prepared via a 'flow' process using the following starting materials and solvents. Two solutions were prepared.
  • Solution A 2-(trifluoromethyl)-1 ,2 ) 3-propanetriol (4.5gm,27.8mmol), N,N,N ⁇ N'-tetramethyl-1 ,6-hexanediamine (30ml, 139mmol), dichloromethane (550ml).
  • Solution B - p-toluenesulphonyl chloride (21.4g, 111mmol), dichloromethane (550ml).
  • HATU O-(7-azabenzotriazol-1 -yl)-N,N, N', N'-tetramethyluronium hexafluorophosphate
  • Amano PS available from Amano Enzymes (1Og) was suspended in water (30ml) and filtered through a Varian bond elute filter tube, washing with water (20ml) After precipitation with propan-2-ol (200ml), the suspension was allowed to settle and the supernatant decanted to leave a 50ml volume of suspension that was centrifuged at 4000 rpm for 5 minutes and the supernatant decanted to leave the title compound
  • Amano lipase PS (100g) was suspended in 1 M pH 7 potassium phosphate solution
  • Amano PS solution (100ml) was added to sepabeads EC-EP (available from Mitzubishi- Resindion) (4Og) and the mixture shaken at room temperature. After 23 hours as much liquid as possible was removed by pipette and the liquid replaced by an equal volume of a 1 M pH10 potassium phosphate solution and shaking continued. After 92 hours the mixture was filtered and the residue washed with water (3 x 50ml), suspended in a saturated solution of octadecylamine in toluene (50ml) and shaken at room temperature for a further 25 hours. The mixture was filtered, washed with toluene (2 x 100ml) and acetone (25ml) and left under suction to afford a free flowing powder. The immobilised enzyme was obtained as pale brown beads (17.29g).
  • Example 1 was further preparatively separated into its enantiomers, Enantiomer 1 and Enantiomer 2 (Isomers A and B) via method C using a Chiralcel OD column eluting with 40% ethanol in heptane at a flow rate of 75ml/m ⁇ n Elution Time for Enantiomer 1 - 7 5m ⁇ ns Elution Time for Enantiomer 2 - 10 3m ⁇ ns
  • Example 3 5-Amino-1-(4-fluorophenyO- ⁇ /-(3 ⁇ 3-trifluoro-2-(rr(2-fluorophenyl)carbonyll(2- hvdroxvethvl)aminolmethvl ⁇ -2-hvdroxypropvl)-1A7-pyrazole-4-carboxamide
  • 2-Fluorobenzoyl chloride (O.i mmol) was weighed into a vial before a solution of 5-amino- ⁇ /- ⁇ 2-[( ⁇ 2-[(1 ,1-dimethylethyl)oxy]ethyl ⁇ amino)methyl]-3,3,3-trifluoro-2-hydroxypropyl ⁇ -1-(4- fluorophenyl)-1H-pyrazole-4-carboxamide (52mg) and diisopropylethylamine (0.035ml, 0.2mmol) in dichloromethane (0.7ml) was added. It was left to stand at room temperature overnight.
  • Aqueous sodium bicarbonate (0.75ml) and chloroform (1.8ml) were added, the mixture shaken and the separated aqueous phase was further washed with chloroform.
  • the chloroform washings were blown down in a scintillation vial before a mixture of trifluoroacetic acid : water : dichloromethane : anisole (20 : 1.5 : 20 : 1 ) (6ml) was added to the residue. It was left overnight and evaporated in vacuo.
  • the residue was treated with aqueous sodium bicarbonate (0.75ml) and chloroform (1.8ml) and shaken.
  • the organic extract was evaporated and the residue was purified by MDAP.
  • Example 3 was further preparatively separated into its enantiomers, Enantiomer 1 and Enantiomer 2 (Isomers A and B) via method B using a Chiralcel OD column eluting with 40% ethanol in heptane at a flow rate of 15ml/min. Elution Time for Enantiomer 1 - 6mins. Elution Time for Enantiomer 2 - 12mins.
  • 2-Chlorobenzoyl chloride (O.immol) was weighed into a vial before a solution of 5-amino- /V- ⁇ 2-[( ⁇ 2-[(1 ,1-dimethylethyl)oxy]ethyl ⁇ amino)methyl]-3,3,3-trifluoro-2-hydroxypropyl ⁇ -1-(4- fluorophenyl)-1H-pyrazole-4-carboxamide (52mg) and diisopropylethylamine (0.035ml, 0.2mmol) in dichloromethane (0.7ml) was added. It was left to stand at room temperature overnight.
  • Aqueous sodium bicarbonate (0.75ml) and chloroform (1.8ml) were added, the mixture shaken and the separated aqueous phase was further washed with chloroform.
  • the chloroform washings were blown down in a scintillation vial before a mixture of trifluoroacetic acid : water : dichloromethane : anisole ( 20 : 1.5 : 20 : 1 ) (6ml) was added to the residue. It was left overnight and evaporated in vacuo.
  • the residue was treated with aqueous sodium bicarbonate (0.75ml) and chloroform (1.8ml) and shaken.
  • the organic extract was evaporated and the residue was purified by MDAP.
  • Example 4 was further preparatively separated into its enantiomers, Enantiomer 1 and Enantiomer 2 (Isomers A and B) via method B using a Chiralcel OD column eluting with 40% ethanol in heptane at a flow rate of 15ml/min. Elution Time for Enantiomer 1 - 6mins. Elution Time for Enantiomer 2 - 10mins.
  • Example 5 5-Amino-/V-(2-(fr(2-chloro-6-fluorophenyl)carbonyl1(2- hvdroxyethyl)amino1methyl)-3.3,3-trifluoro-2-hvdroxypropyl)-1-(4-fluorophenylV1f/- pyrazole-4-carboxamide
  • 2-Chloro-6-fluorobenzoyl chloride (O.immol) was weighed into a vial before a solution of 5-amino-/V- ⁇ 2-[( ⁇ 2-[(1 ,1-dimethylethyl)oxy]ethyl ⁇ amino)methy
  • Aqueous sodium bicarbonate (0.75ml) and chloroform (1.8ml) were added, the mixture shaken and the separated aqueous phase was further washed with chloroform.
  • the chloroform washings were blown down in a scintillation vial before a mixture of trifluoroacetic acid : water : dichloromethane : anisole (20 : 1.5 : 20 : 1 ) (6ml) was added to the residue. It was left overnight and evaporated in vacuo.
  • the residue was treated with aqueous sodium bicarbonate (0.75ml) and chloroform (1.8ml) and shaken.
  • the organic extract was evaporated and the residue was purified by MDAP.
  • Example 5 was further separated into its Isomers A (probably a mixture of rotamers 1 and 2 and atropisomers 1 and 2) and B (probably a mixture of rotamers 3 and 4 and atropisomers 3 and 4)) using analytical chiral HPLC.
  • Isomers A (probably a mixture of rotamers 1 and 2 and atropisomers 1 and 2)
  • Analytical Chiral HPLC 25 x 0.46cm Chiralcel OD column, 20% ethanol in heptane eluting at 1ml/min) - Retention time 7.55 mins and 8.73 mins.
  • Isomers B (probably a mixture of rotamers 3 and 4 and atropisomers 3 and 4)
  • Analytical Chiral HPLC 25 x 0.46cm Chiralcel OD column, 20% ethanol in heptane eluting at 1 ml/min) - Retention time 20.32 mins and 39.49 mins.
  • Example 6 was further preparatively separated into its enantiomers, Enantiomer 1 and Enantiomer 2 (Isomers A and B) via method C using a Chiralcel OD(5 x 25cm) column eluting with 40% ethanol in heptane at a flow rate of 75ml/min. Elution Time for Enantiomer 1 - 7.6mins. Elution Time for Enantiomer 2 - 12.5mins.
  • Example 7 was further preparatively separated into its enantiomers, Enantiomer 1 and Enantiomer 2 (Isomers A and B) via method C using a Chiralcel OD column (5 x 25cm) eluting with 30% ethanol in heptane at a flow rate of 75ml/min. Elution Time for Enantiomer 1 - 9.0mins. Elution Time for Enantiomer 2 - 15.5mins.
  • Example 8j 5-Amino- ⁇ /-(2-([r(2-chloro-3-fluorophenv ⁇ carbonyll(2- hvdroxyethvhamino1methyl>-3.3.3-trifluoro-2-hvdroxypropy ⁇ -1-(4-fluorophenyl)-1 /-/- pyrazole-4-carboxamide
  • Example 8 was further preparatively separated into its enantiomers (Isomers A and B) using a 2" x 20cm Chiralcel OD column eluti ⁇ g with 40% ethanol in heptane at a flow rate of 75m!/min.
  • the ability of compounds to bind to the glucocorticoid receptor was determined by assessing their ability to compete with an Alexa 555 fluorescently-labelled dexamethasone derivative. Compounds were solvated and diluted in DMSO, and transferred directly into assay plates. Fluorescent dexamethasone and a partially purified full length glucocorticoid receptor were added to the plates, together with buffer components to stabilise the GR protein and incubated at room temperature for 2 hours in the dark. Binding of each compound was assessed by analysing the displacement of fluorescent ligand by measuring the decrease in fluorescence polarisation signal from the mixture. Dose response curves were constructed from which plC 50 values were estimated.
  • Example 1 (racemic), Example 1 Enantiomer 1 , Example 1 Enantiomer 2, Example 2 (racemic), Example 3 (racemic), Example 3 Enantiomer 1 , Example 3 Enantiomer 2, Example 4 (racemic), Example 4 Enantiomer 1, Example 4 Enantiomer 2, Example 5 (racemic), Example 6 (racemic), Example 6 Enantiomer 1 , Example 6 Enantiomer 2, Example 7 (racemic), Example 7 Enantiomer 1 , Example 7 Enantiomer 2, Example 8 (racemic), Example 8 Enantiomer 1 , Example 8 Enantiomer 2, Example 9 (racemic), Example 10 (racemic), Example 11 (racemic), Example 12 (racemic), Example 13 (racemic) and Example 14 (racemic) show glucocorticoid binding with a plC 50 > 6 in this assay.
  • Human Caucasian lung carcinoma A549 cell line (ECACC No. 86012804) has been stably transfected in house with a plasmid containing an ELAM promoter sequence that has a NFKB response element within it. Stimulation of the cell line with TNF ⁇ results in intracellular signal transduction and ultimately translocation of NFKB into the nucleus. This activates the inserted DNA sequence resulting in transcription of the integrated SPAP gene, which is quantified using a colorimetric assay. In this assay, GR agonist compounds inhibit NFKB driven transcription resulting in a decrease in signal.
  • the stably transfected cell line was grown as a monolayer in DMEM supplemented with FCS-HI (10%), Non-essential amino acids (1 %), L-Glutamine (2mM), Pen/Strep (1%) and Geneticin (50mg/ml).
  • NFKB agonist assay A 70% confluent T225 flask of A549 SPAP cells was harvested by centrifugation for 5 min at 20Og, resuspended in assay buffer (DMEM supplemented with 10% FCS 2xHI, 2mM L- Glutamine,1 % Pen/Strep and No ⁇ essential amino acids) and diluted to 0.16 x 10 6 /ml. 60 ⁇ l of cell solution was dispensed to each well of clear Nunc 384-well plates, containing compound at the required concentration. Plates were incubated for 1 h at 37 0 C, 95% humidity, 5% CO 2 before 10 ⁇ l of TNF ⁇ was added at final concentration of 3.2ng/ml and then returned to the cell incubator for 15h.
  • assay buffer DMEM supplemented with 10% FCS 2xHI, 2mM L- Glutamine,1 % Pen/Strep and No ⁇ essential amino acids
  • Example 1 (racemic), Example 1 Enantiomer 2, Example 2 (racemic), Example 3 (racemic), Example 3 Enantiomer 2, Example 4 (racemic), Example 4 Enantiomer 1 , Example 4 Enantiomer 2, Example 5 (racemic), Example 5 Enantiomer 1 , Example 5 Enantiomer 2, Example 6 (racemic), Example 6 Enantiomer 1 , Example 6 Enantiomer 2, Example 7 (racemic), Example 7 Enantiomer 1 , Example 7 Enantiomer 2, Example 8 (racemic), Example 8 Enantiomer 2, Example 9 (racemic), Example 10 (racemic), Example 1 1 (racemic), Example 12 (racemic), Example 13 (racemic), Example 14 (racemic) are > 6.5 for the NFkB assay.
  • Human Caucasian lung carcinoma A549 cell line (ECACC No. 86012804) has been stably transfected in house with a plasmid containing a renilla luciferase reporter with an MMTV promoter. Stimulation of the cell line with GR agonists results in intracellular signal transduction and ultimately translocation of GR into the nucleus. This activates the inserted DNA sequence resulting in transcription of the integrated luciferase gene, which is quantified using a light emission.
  • the stably transfected cell line was grown as a monolayer in DMEM supplemented with FCS-HI (10%), Non-essential amino acids (1 %), L-Glutamine (2mM), Pen/Strep (1%) and Geneticin (50mg/ml).
  • a 90% confluent T175 flask of A549 MMTV cells was harvested by centrifugation for 5 mi ⁇ at 20Og, resuspended in assay buffer (DMEM supplemented with 10% FCS 2xHI, 2mM Glutamax, Non essential amino acids and 25mM HEPES) and diluted to 0.1 x 10 6 /ml.
  • 70 ⁇ l of cell solution was dispensed to each well of white Nunc 384-well plates, containing compound at the required concentration. Plates were incubated for 6h at 37 ° C, 95% humidity, 5% CO 2 . Plates were equilibrated to room temperature for 1 h prior to the addition of 10 ⁇ l of Renilla substrate to each well of assay plates. The plates were covered to protect the reagents from light, and then incubated at room temperature for approximately 15 minutes before reading them on a Viewlux.
  • Example 1 (racemic), Example 1 Enantiomer 2, Example 2 (racemic), Example 3 (racemic), Example 3 Enantiomer 2, Example 4 (racemic), Example 4 Enantiomer 2, Example 5 (racemic), Example 5 Enantiomer 2, Example 6 (racemic), Example 6 Enantiomer 2, Example 7 (racemate), Example 7 Enantiomer 2, Example 8 Enantiomer 2, Example 9 (racemate), Example 10 (racemic), Example 1 1 (racemic), Example 12 (racemic) and Example 13 (racemic).
  • a T225 flask of CV-1 cells at a density of 80% confluency was washed with PBS, detached from the flask using 0.25% trypsin and counted using a Sysmex KX-21 N.
  • Cells were diluted in DMEM containing 10% Hyclone, 2mM L-Glutamate and 1 % Pen/Strep at 140 cells/ ⁇ l and transduced with 10% PRb-BacMam and 10% MMTV-BacMam.
  • 70 ml of suspension cells were dispensed to each well of white Nunc 384-well plates, containing compounds at the required concentration. After 24h 10 ⁇ l of Steadylite were added to each well of the plates. Plates were incubated in the dark for 10 min before reading them on a Viewlux reader. Dose response curves were constructed from which pEC 50 values were estimated.
  • Example 1 (racemic), Example 1 Enantiomer 1 , Example 1 Enantiomer 2, Example 2 (racemic), Example 3 (racemic), Example 3 Enantiomer 1 , Example 3 Enantiomer 2, Example 4 (racemic), Example 4 Enantiomer 1 , Example 4 Enantiomer 2, Example 5 (racemic), Example 5 Enantiomer 1 , Example 5 Enantiomer 2, Example 6 (racemic), Example 6 Enantiomer 1 , Example 6 Enantiomer 2, Example 7 (racemic), Example 7 Enantiomer 1 , Example 7 Enantiomer 2, Example 8 (racemic), Example 8 Enantiomer 1 , Example 8 Enantiomer 2, Example 9 (racemic), Example 10 (racemic), Example 11 (racemic), Example 12 (racemic), Example 13 (racemic) and Example 14 (racemic) show pEC 50 ⁇ 6 in this assay.
  • At least one isomer for example, an enantiomer in a mixture of isomers (such as a racemate) has the described activity.
  • the other enantiomer may have similar activity, less activity, no activity or may have some antagonist activity in the case of a functional assay.

Abstract

The present invention provides compounds of formula (I), a process for their preparation, to pharmaceutical compositions comprising the compounds and the preparation of said compositions, to intermediates and to use of the compounds for the manufacture of a medicament for therapeutic treatment, particularly for the treatment of inflammation and/or allergic conditions.

Description

NOVEL COMPOUNDS
The present invention relates to non-steroidal compounds and a process for their preparation, to pharmaceutical compositions comprising the compounds and the preparation of said compositions, to intermediates and to use of the compounds for the manufacture of a medicament for therapeutic treatment, particularly for the treatment of inflammation and/or allergic conditions.
Nuclear receptors are a class of structurally related proteins involved in the regulation of gene expression. The steroid hormone receptors are a subset of this family whose natural ligands typically comprise endogenous steroids such as estradiol (estrogen receptor), progesterone (progesterone receptor) and Cortisol (glucocorticoid receptor). Man-made ligands to these receptors play an important role in human health, in particular the use of glucocorticoid agonists to treat a wide range of inflammatory conditions.
Current known glucocorticoids have proved useful in the treatment of inflammation, tissue rejection, auto-immunity, various malignancies, such as leukemias and lymphomas, Cushing's syndrome, rheumatic fever, polyarteritis nodosa, granulomatous polyarteritis, inhibition of myeloid cell lines, immune proliferation/apoptosis, HPA axis suppression and regulation, hypercortisolemia, modulation of the Th1/Th2 cytokine balance, chronic kidney disease, stroke and spinal cord injury, hypercalcemia, hypergylcemia, acute adrenal insufficiency, chronic primary adrenal insufficiency, secondary adrenal insufficiency, congenital adrenal hyperplasia, cerebral edema, thrombocytopenia, Little's syndrome, inflammatory scalp alopecia, panniculitis, psoriasis, discoid lupus erythemnatosus, inflamed cysts, atopic dermatitis, pyoderma gangrenosum, pemphigus vulgaris, bullous pemphigoid, systemic lupus erythematosus, dermatomyositis, herpes gestationis, eosinophilic fasciitis, relapsing polychondritis, inflammatory vasculitis, sarcoidosis, Sweet's disease, type 1 reactive leprosy, capillary hemangiomas, contact dermatitis, atopic dermatitis, lichen planus, exfoliative dermatitis, erythema nodosum, acne, hirsutism, toxic epidermal necrolysis, erythema multiform and cutaneous T-cell lymphoma.
Glucocorticoids are especially useful in disease states involving systemic inflammation such as inflammatory bowel disease, systemic lupus erythematosus, polyarteritis nodosa, Wegener's granulomatosis, giant cell arteritis, rheumatoid arthritis, osteoarthritis, seasonal rhinitis, allergic rhinitis, vasomotor rhinitis, urticaria, angioneurotic edema, chronic obstructive pulmonary disease, asthma, tendonitis, bursitis, Crohn's disease, ulcerative colitis, autoimmune chronic active hepatitis, organ transplantation, hepatitis and cirrhosis. Glucocorticoids have also been used as immunostimulants and repressors and as wound healing and tissue repair agents.
There remains a need to find further compounds which bind to the glucocorticoid receptor.
In one embodiment, the present invention provides compounds of formula (I):
Figure imgf000004_0001
wherein
R1 is selected from hydrogen, -OCHF2, fluorine and chlorine;
R2 and R3 are each independently selected from hydrogen, fluorine, chlorine and bromine; when R1 is -OCHF2, R2 and R3 are each hydrogen, or R2 is chlorine and R3 is hydrogen; n is an integer selected from 0, 1 and 2, when n is 1 , X is selected from chlorine and fluorine, and when n is 2, each X is fluorine; and salts and solvates thereof (hereinafter "compounds of the invention").
In a further embodiment, the present invention provides compounds of formula (IA):
Figure imgf000005_0001
wherein
R1 is selected from hydrogen, -OCHF2, fluorine and chlorine; R2 and R3 are independently selected from hydrogen, fluorine and chlorine; when R1 is -OCHF2, R2 and R3 are each hydrogen; and salts and solvates thereof.
The compounds of formula (I) each contain a chiral centre and there are two possible stereoisomers (enantiomers) of each compound of formula (I).
The terms Enantiomer 1 and Enantiomer 2 are used herein to refer to the enantiomers of a compound of formula (I), based on the order of their elution using the chiral chromatography methodology described herein. Enantiomer 1 refers to the first enantiomer to elute, and Enantiomer 2 refers to the second enantiomer to elute.
It will be appreciated by those skilled in the art that although the absolute retention time on chromatography can be variable, the order of elution remains the same when the same column and conditions are employed. However, the use of a different chromatography column and conditions may alter the order of elution.
A mixture of enantiomers, such as a racemic mixture, may be preferred. Thus, in one embodiment of the invention the compound of formula (I) is the racemic mixture (the racemate). Alternatively, a single enantiomer may be preferred, for example the Enantiomer 1 Thus, in one embodiment of the invention the compound of formula (I) is the Enantiomer 1 In a further embodiment of the invention the compound of formula (I) is the Enantiomer 2
It will be appreciated by those skilled in the art that as rotation of the aryl-carbonyl bond becomes less facile due to ortho substitution on the aromatic ring, an axis of symmetry may be observed thus introducing atropisomerism into the compound and thus creating the possibility of four isomers namely Atropisomer 1 , Enantiomer 1 (A1 E1 ), Atropisomer 1 , Enantiomer 2 (A1 E2), Atropisomer 2, Enantiomer 1 (A2E1 ), and Atropisomer 2, Enantiomer 2 (A2E2) Any comment relating to the biological activity of an isomer or stereoisomer should be taken to include these atropisomers It will be appreciated by those skilled in the art that where there is a non equilibrium ratio of atropisomers, this ratio may move towards the equilibrium ratio
It will be further appreciated by those skilled in the art that, for compounds of formula (I) wherein rotation is restricted around the C(O)-NCH2CH2OH bond due to the hydroxyethyl moiety, rotamers may be observed Any comment relating to the biological activity of an isomer or stereoisomer should be taken to include these rotamers It will be appreciated by those skilled in the art that there may not be a 1 1 ratio of rotamers as the ratio can change depending on the half life of interconversion
The terms "stereoisomer" and "isomer" as used herein encompass enantiomer, atropisomer and/or rotamer
The compounds of the invention are glucocorticoid receptor binders Accordingly, it has been found that at least one of the possible enantiomers of each of the compounds of formula (I) binds to the glucocorticoid receptor
Further, it appears that at least one of the possible enantiomers of each of the compounds of formula (I) has glucocorticoid receptor binding activity Accordingly, at least one of the possible enantiomers of each compound of formula (I) modulates the glucocorticoid receptor The term "modulator" as used herein refers to a compound which binds to the glucocorticoid receptor and acts as either an agonist, a partial agonist or an antagonist of the glucocorticoid receptor
The compounds of the invention may provide agonism of the glucocorticoid receptor Additionally, it appears that one or more of the possible enantiomers of some of the compounds of formula (I) possess advantageous selectivity in respect of maintaining transrepression activity whilst reducing the transactivation activity. These observations are believed to be indicative that the compounds of the invention provide antiinflammatory properties with fewer or less severe related side effects.
In one embodiment, R1 is selected from fluorine and chlorine.
In one embodiment, R2 and R3 are each independently selected from hydrogen, fluorine and chlorine.
In one embodiment, when R1 is -OCHF2, R2 and R3 are each hydrogen.
In one embodiment, R2 is selected from fluorine and chlorine.
In one embodiment, R3 is hydrogen. In another embodiment, when R3 is fluorine, R1 is hydrogen and R2 is chlorine. In a further embodiment, when R3 is chlorine, R1 and R2 are each hydrogen.
In one embodiment, R1 is fluorine, R2 is chlorine and R3 is hydrogen. In another embodiment, R1 and R2 are each fluorine and R3 is hydrogen. In a further embodiment, R1 and R2 are each chlorine and R3 is hydrogen.
In one embodiment, n is 1.
In one embodiment, when n is 1 , X is fluorine. In a further embodiment, the fluorine is in the para position on the phenyl ring.
It is to be understood that the present invention covers all combinations of the substituent groups described hereinabove.
In one embodiment, the compound of formula (I) is:
5-amino-Λ/-(2-{[({2-[(difluoromethyl)oxy]phenyl}carbonyl)(2-hydroxyethyl)amino]methyl}- 3,3,3-trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1 H-pyrazole-4-carboxamide; 5-amino-Λ/-(2-{[({2-[(difluoromethyl)oxy]phenyl}carbony])(2-hydroxyethyl)amino]methyl}-
3,3,3-trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide
(Enantiomer 1 );
5-amino-Λ/-(2-{[({2-[(difluoromethyl)oxy]phenyl}carbonyl)(2-hydroxyethyl)amino]methyl}- 3,3,3-trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1/-/-pyrazole-4-carboxamide
(Enantiomer 2);
5-amino-Λ/-(2-{[[(3-chlorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3-trifluoro-2- hydroxypropyl)-1 -(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide;
5-amino-1-(4-fluorophenyl)-Λ/-(3,3,3-trifluoro-2-{[[(2-fluorophenyl)carbonyl](2- hydroxyethyl)amino]methyl}-2-hydroxypropyl)-1 /-/-pyrazole-4-carboxamide;
5-amino-1-(4-fluorophenyl)-Λ/-(3,3,3-trifluoro-2-{[[(2-fluorophenyl)carbonyl](2- hydroxyethyl)amino]methyl}-2-hydroxypropyl)-1 H-pyrazole-4-carboxamide (Enantiomer 1 );
5-amino-1-(4-fluorophenyl)-A/-(3,3,3-trifluoro-2-{[[(2-fluorophenyl)carbony|](2- hydroxyethyl)amino]methyl}-2-hydroxypropyl)-1 /-/-pyrazole-4-carboxamide (Enantiomer 2); 5-amino-Λ/-(2-{[[(2-chlorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3-trifluoro-2- hydroxypropyl)-1 -(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide;
5-amino-Λ/-(2-{[[(2-chlorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3-trifluoro-2- hydroxypropyl)-1-(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide (Enantiomer 1 );
5-amino-Λ/-(2-{[[(2-chlorophenyl)carbony|](2-hydroxyethyl)amino]methyl}-3,3,3-trifluoro-2- hydroxypropyl)-1-(4-fluorophenyl)-1/-/-pyrazole-4-carboxamide (Enantiomer 2);
5-amino-Λ/-(2-{[[(2-chloro-6-fluorophenyl)carbony|](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide;
5-amino-Λ/-(2-{[[(2-chloro-6-fluorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide (Enantiomer 1 ); 5-amino-Λ/-(2-{[[(2-chloro-6-fluorophenyl)carbony|](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1 -(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide (Enantiomer 2);
5-amino-Λ/-(2-{[[(2,6-difluorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3-trifluoro-
2-hydroxypropyl)-1-(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide;
5-amino-Λ/-(2-{[[(2,6-difluorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3-trifluoro- 2-hydroxypropyl)-1-(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide (Enantiomer 1 );
5-amino-Λ/-(2-{[[(2,6-difluorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3-trifluoro-
2-hydroxypropyl)-1 -(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide (Enantiomer 2);
(S)-5-amino-Λ/-(2-{[[(2,6-difluorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide; 5-amino-Λ/-(2-{[[(2,6-dichlorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide; 5-amino-Λ/-(2-{[[(2,6-dichlorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1 -(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide (Enantiomer 1 );
5-amino-Λ/-(2-{[[(2,6-dichlorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1 -(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide (Enantiomer 2); 5-amino-A/-(2-{[[(2-chloro-3-fluorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide;
5-amino-Λ/-(2-{[[(2-chloro-3-fluorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide (Enantiomer 1 );
5-amino-Λ/-(2-{[[(2-chloro-3-fluorophenyl)carbony|](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide (Enantiomer 2);
5-amino-1-(4-fluorophenyl)-Λ/-(3,3,3-trifluoro-2-hydroxy-2-{[(2- hydroxyethyl)(phenylcarbonyl)amino]methyl}propyl)-1 /-/-pyrazole-4-carboxamide;
5-amino-A/-(2-{[[(2,3-difluorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3-trιfluoro-
2-hydroxypropyl)-1-(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide; 5-amino-Λ/-(2-{[[(3-bromo-2-fluorophenyl)carbony|](2-hydroxyethyl)amino]methyl}-3,3,3- trιl:luoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide;
5-amino-Λ/-(2-{[[(3-chloro-2-fluorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1H-pyrazole-4-carboxamide;
5-amino-Λ/-(2-{[({2-chloro-6-[(difluoromethyl)oxy]phenyl}carbonyl)(2- hydroxyethyl)amino]methyl}-3,3,3-trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1 /-/- pyrazole-4-carboxamide;
5-amino-Λ/-(2-{[[(2-bromo-3-fluorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1 -(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide; or a salt or solvate thereof.
In another embodiment, the compound of formula (I) is:
5-amino-Λ/-(2-{[({2-[(difluoromethyl)oxy]phenyl}carbonyl)(2-hydroxyethyl)amino]methyl}-
3,3,3-trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide;
5-amino-A/-(2-{[[(3-chlorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3-trifluoro-2- hydroxypropyl)-1-(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide;
5-amino-1-(4-fluorophenyl)-Λ/-(3,3,3-trifluoro-2-{[[(2-fluorophenyl)carbonyl](2- hydroxyethyl)amino]methyl}-2-hydroxypropyl)-1 /-/-pyrazole-4-carboxamιde;
5-amino-Λ/-(2-{[[(2-chlorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3-trifluoro-2- hydroxypropyl)-1-(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide; 5-amino-Λ/-(2-{[[(2-chlorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3-trifluoro-2- hydroxypropyl)-1-(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide (Enantiomer 1 ); 5-amino-Λ/-(2-{[[(2-chlorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3-trifluoro-2- hydroxypropyl)-1-(4-fluorophenyl)-1 H-pyrazole-4-carboxamide (Enantiomer 2);
5-amino-Λ/-(2-{[[(2-chloro-6-fluorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1/-/-pyrazole-4-carboxamide;5-amino-Λ/-(2- {[[(2,6-difluorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3-trifluoro-2- hydroxypropyl)-1 -(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide;
5-amino-Λ/-(2-{[[(2,6-diNuorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3-trifluoro-
2-hydroxypropyl)-1 -(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide (Enantiomer 1 );
5-amino-/V-(2-{[[(2,6-difluorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3-trifluoro- 2-hydroxypropyl)-1-(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide (Enantiomer 2);
5-amino-Λ/-(2-{[[(2,6-dichlorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1 -(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide;
5-amino-Λ/-(2-{[[(2,6-dichlorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide (Enantiomer 1 ); 5-amino-Λ/-(2-{[[(2,6-dichlorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1 -(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide (Enantiomer 2);
5-amino-Λ/-(2-{[[(2-chloro-3-fluorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide;5-amino-1-(4-
11uorophenyl)-Λ/-(3,3,3-trifluoro-2-hydroxy-2-{[(2- hydroxyethy^phenylcarbonyOaminolmethylJpropyO-I H-pyrazole^-carboxamide;
5-amino-Λ/-(2-{[[(2,3-difluorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3-trifluoro-
2-hydroxypropyl)-1 -(4-fluorophenyl)-1 H-pyrazole-4-carboxamide;
5-amino-/V-(2-{[[(3-bromo-2-fluorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1H-pyrazole-4-carboxamide; 5-amino-Λ/-(2-{[[(3-chloro-2-fluorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide;
5-amino-Λ/-(2-{[({2-chloro-6-[(difluoromethyl)oxy]phenyl}carbonyl)(2- hydroxyethyl)amino]methyl}-3,3,3-trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1H- pyrazole-4-carboxamide; 5-amino-Λ/-(2-{[[(2-bromo-3-fluorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1 -(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide; or a salt or solvate thereof.
In a another embodiment, the compound of formula (I) is: 5-amino-Λ/-(2-{[({2-[(difluoromethyl)oxy]phenyl}carbonyl)(2-hydroxyethyl)amino]methyl}- 3,3,3-trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide; 5-amιno-Λ/-(2-{[[(3-chlorophenyl)carbonyl](2-hydroxyethyl)annιno]nnethyl}-3,3,3-tπfluoro-2- hydroxypropyO-i-^-fluorophenyO-I W-pyrazole^-carboxamide, 5-amιno-1-(4-fluorophenyl)-Λy-(3,3,3-trιfluoro-2-{[[(2-fluorophenyl)carbonyl](2- hydroxyethyl)amιno]methyl}-2-hydroxypropyl)-1W-pyrazole-4-carboxamιde, 5-amιno-Λ/-(2-{[[(2-chlorophenyl)carbonyl](2-hydroxyethyl)annιno]nnethyl}-3,3,3-trιfluoro-2- hydroxypropyl)-1 -(4-fluorophenyl)-1 W-pyrazole-4-carboxamιde,
5-amιno-Λ/-(2-{[[(2-chlorophenyl)carbonyl](2-hydroxyethyl)amιno]methyl}-3,3,3-trιfluoro-2- hydroxypropyl)-1-(4-fluorophenyl)-1/-/-pyrazole-4-carboxamιde (Enantiomer 1 ), 5-amιno-Λy-(2-{[[(2-chlorophenyl)carbonyl](2-hydroxyethyl)annιno]nnethyl}-3,3,3-trιfluoro-2- hydroxypropyl)-1 -(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamιde (Enantiomer 2),
5-amιno-Λ/-(2-{[[(2-chloro-6-fluorophenyl)carbonyl](2-hydroxyethyl)annιno]nnethyl}-3,3,3- trιfluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamιde, 5-amιno-Λ/-(2-{[[(2,6-dιfluorophenyl)carbony|](2-hydroxyethyl)amιno]nnethyl}-3,3,3-tπfluoro- 2-hydroxypropyl)-1-(4-fluorophenyl)-1/-/-pyrazole-4-carboxamιde, 5-amιno-Λ/-(2-{[[(2,6-dιfluorophenyl)carbony|](2-hydroxyethyl)amιno]methyl}-3,3,3-tπfluoro- 2-hydroxypropyl)-1 -(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamιde (Enantiomer 1 ), 5-amιno-Λ/-(2-{[[(2,6-dιfluorophenyl)carbonyl](2-hydroxyethyl)amιno]methyl}-3,3,3-tπfluoro- 2-hydroxypropyl)-1 -(4-fϊuorophenyl)-1 H-pyrazole-4-carboxamιde (Enantiomer 2), 5-amιno-Λ/-(2-{[[(2,6-dιchlorophenyl)carbonyl](2-hydroxyethyl)amιno]methyl}-3,3,3- trιfluoro-2-hydroxypropyl)-1 -(4-fluorophenyl)-1 H-pyrazole-4-carboxamιde,
5-amιno-Λy-(2-{[[(2,6-dιchlorophenyl)carbonyl](2-hydroxyethyl)amιno]methyl}-3,3,3- trιfluoro-2-hydroxypropyl)-1 -(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamιde (Enantiomer 1 ), 5-amιno-Λ/-(2-{[[(2,6-dιchlorophenyl)carbonyl](2-hydroxyethyl)amιno]methyl}-3,3,3- trιfluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1/-/-pyrazole-4-carboxamιde (Enantiomer 2), 5-amιno-Λ/-(2-{[[(2-chloro-3-fluorophenyl)carbonyl](2-hydroxyethyl)amιno]methyl}-3,3,3- trιfluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1 H-pyrazole-4-carboxamιde, 5-amιno-1-(4-fluorophenyl)-Λ/-(3,3,3-trιfluoro-2-hydroxy-2-{[(2- hydroxyethyl)(phenylcarbonyl)amιno]methyl}propyl)-1 /-/-pyrazole-4-carboxamιde, or a salt or solvate thereof
In a further embodiment, the compound of formula (I) is
5-amιno-Λ/-(2-{[({2-[(dιfluoromethyl)oxylphenyl}carbonyl)(2-hydroxyethyl)amιno]methyl}- 3,3,3-trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1 H-pyrazole-4-carboxamide, 5-amιno-1-(4-fluorophenyl)-Λ/-(3,3,3-tπfluoro-2-{[[(2-fluorophenyl)carbony|](2- hydroxyethyl)amιno]methyl}-2-hydroxypropyl)-1 H-pyrazole-4-carboxamιde (Enantiomer 2), 5-amino-Λ/-(2-{[[(2-chlorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3-trifluoro-2- hydroxypropyl)-1-(4-fluorophenyl)-1 H-pyrazole-4-carboxamide (Enantiomer 2); 5-amino-Λ/-(2-{[[(2-chloro-6-fluorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1H-pyrazole-4-carboxamide; 5-amino-Λ/-(2-{[[(2,6-difluorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3-trifluoro- 2-hydroxypropyl)-1-(4-fluorophenyl)-1 H-pyrazole-4-carboxamide;
5-amino-Λ/-(2-{[[(2,6-difluorophenyl)carbony|](2-hydroxyethyl)amino]methyl}-3,3,3-trifluoro- 2-hydroxypropyl)-1 -(4-fluorophenyl)-1 H-pyrazole-4-carboxamide (Enantiomer 2); 5-amino-Λ/-(2-{[[(2,6-dichlorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1 H-pyrazole-4-carboxamide;
5-amino-Λ/-(2-{[[(2>6-dichlorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1 H-pyrazole-4-carboxamide (Enantiomer 2); 5-amino-Λ/-(2-{[[(2-chloro-3-fluorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1 H-pyrazole-4-carboxamide; 5-amino-Λ/-(2-{[[(3-bromo-2-fluorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1 H-pyrazole-4-carboxamide; 5-amino-Λ/-(2-{[[(3-chloro-2-fluorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1 -(4-fluorophenyl)-1H-pyrazole-4-carboxamide; (S)-5-amino-Λ/-(2-{[[(2,6-difluorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1 -(4-fluorophenyl)-1 H-pyrazole-4-carboxamide; or a salt or solvate thereof.
One embodiment of the invention embraces compounds of formula (I) and salts and solvates thereof. Another embodiment of the invention embraces compounds of formula (I) and salts thereof. Another embodiment of the invention embraces compounds of formula (I) and solvates thereof. A further embodiment of the invention embraces compounds of formula (I) as the free base.
Salts and solvates of the compounds of formula (I) which are suitable for use in medicine are those wherein the counter-ion or associated solvent is pharmaceutically acceptable. However, salts and solvates having non-pharmaceutically acceptable counter-ions or associated solvents are within the scope of the present invention, for example, for use as intermediates in the preparation of other compounds of formula (I) and their pharmaceutically acceptable salts and solvates thereof. One embodiment of the invention embraces compounds of formula (I) and pharmaceutically acceptable salts and solvates thereof. Another embodiment of the invention embraces compounds of formula (I) and pharmaceutically acceptable salts thereof. Another embodiment of the invention embraces compounds of formula (I) and pharmaceutically acceptable solvates thereof.
Suitable salts according to the invention include those formed with both organic and inorganic acids or bases. Pharmaceutically acceptable acid addition salts may include those formed from hydrochloric, hydrobromic, sulphuric, phosphoric, trifluoroacetic, sulphamic, sulphanilic, methanesulphonic, ethanesulphonic and arylsulphonic (for example p-toluenesulphonic, benzenesulphonic, naphthalenesulphonic or naphthalenedisulphonic) acids. Pharmaceutically acceptable base salts may include alkali metal salts such as those of sodium and potassium and alkaline earth metal salts such as those of calcium.
Examples of solvates include hydrates.
The compounds of the invention may have the ability to crystallise in more than one form. This is a characteristic known as polymorphism, and it is understood that such polymorphic forms ("polymorphs") are within the scope of the present invention. Polymorphism generally can occur as a response to changes in temperature or pressure or both and can also result from variations in the crystallisation process. Polymorphs can be distinguished by various physical characteristics known in the art such as x-ray diffraction patterns, solubility, and melting point.
The compounds of the invention are expected to have potentially beneficial antiinflammatory and/or anti-allergic effects, particularly upon oral administration, demonstrated by, for example, their ability to bind to the glucocorticoid receptor and to ellicit a response via that receptor. Hence, the compounds of the invention may be of use in the treatment of inflammatory and/or allergic disorders.
Examples of disease states associated with glucocorticoid receptor activity include skin diseases such as eczema, psoriasis, allergic dermatitis, neurodermatitis, pruritis, exfoliative dermatitis, pemphigus and hypersensitivity reactions; inflammatory conditions of the nose, throat or lungs such as asthma (including allergen-induced asthmatic reactions), rhinitis (including hayfever), nasal polyps, chronic obstructive pulmonary disease (COPD), interstitial lung disease, and fibrosis; inflammatory bowel conditions such as ulcerative colitis and Crohn's disease; auto-immune diseases such as rheumatoid arthritis, systemic lupus erythematosus, termporal arteritis, polyarteritis nodosa, polymyositis, ankylosing spondylitis, sarcoidosis, autoimmune hepatitis; cancers such as acute and lymphatic leukaemia, myeloma, lymphoma; nephritic syndrome; septic shock; adrenal insufficiency; ophthalmic inflammation and allergic conjunctivitis; obesity; diabetes; chronic inflammatory pain including musculoskeletal pain; lower back and neck pain; sprains and strains; neuropathic pain; sympathetically maintained pain; myositis; pain associated with cancer and fibromyalgia; pain associated with migraine; pain associated with influenza or other viral infections, such as the common cold; rheumatic fever; pain associated with functional bowel disorders such as non-ulcer dyspepsia, non- cardiac chest pain and irritable bowel syndrome; pain associated with myocardial ischemia; post operative pain; headache; toothache; and dysmenorrhea; psychiatric disease for example schizophrenia, depression (which term is used herein to include bipolar depression, unipolar depression, single or recurrent major depressive episodes with or without psychotic features, catatonic features, melancholic features, atypical features or postpartum onset, seasonal affective disorder, dysthymic disorders with early or late onset and with or without atypical features, neurotic depression and social phobia, depression accompanying dementia for example of the Alzheimer's type, schizoaffective disorder or the depressed type, and depressive disorders resulting from general medical conditions including, but not limited to, myocardial infarction, diabetes, miscarriage or abortion, etc), anxiety disorders (including generalised anxiety disorder and social anxiety disorder), panic disorder, agoraphobia, social phobia, obsessive compulsive disorder and post-traumatic stress disorder, memory disorders, including dementia, amnesic disorders and age-associated memory impairment, disorders of eating behaviours, including anorexia nervosa and bulimia nervosa, sleep disorders (including disturbances of circadian rhythm, dyssomnia, insomnia, sleep apnea and narcolepsy), withdrawal from abuse of drugs such as of ***e, ethanol, nicotine, benzodiazepines, alcohol, caffeine, phencyclidine (phencyclidine-like compounds), opiates (e.g. cannabis, heroin, morphine), amphetamine or amphetamine-related drugs (e.g. dextroamphetamine, methylamphetamine) or a combination thereof. Compounds having glucocorticoids receptor activity may also have utility in inducing suppression of the immune system during organ transplantation, in acute transplant reject, angioedema of the upper respiratory tract and anaphylactic shock. Examples of disease states in which the compounds of the present invention are expected to have utility include rheumatoid arthritis, asthma, COPD, allergy and rhinitis.
It will be appreciated by those skilled in the art that reference herein to treatment extends to prophylaxis as well as the treatment of established conditions.
As mentioned above, compounds of the invention are expected to be of use in human or veterinary medicine, in particular as anti-inflammatory and/or anti-allergic agents.
There is thus provided as a further aspect of the invention a compound of formula (I) or a pharmaceutically acceptable salt or solvate thereof for use in therapy.
In another aspect of the invention there is provided a compound of formula (I) or a pharmaceutically acceptable salt or solvate thereof for use in the treatment of patients with an inflammatory and/or allergic condition.
In another aspect of the invention there is provided a compound of formula (I) or a pharmaceutically acceptable salt or solvate thereof for use in the treatment of patients with rheumatoid arthritis, asthma, COPD, allergy and/or rhinitis.
In another aspect of the invention there is provided a compound of formula (I) or a pharmaceutically acceptable salt or solvate thereof for use in the treatment of patients with skin disease.
In another aspect of the invention there is provided a compound of formula (I) or a pharmaceutically acceptable salt or solvate thereof for use in the treatment of patients with eczema, psoriasis, allergic dermatitis, neurodermatitis, pruritis and/or hypersensitivity reactions.
According to another aspect of the invention, there is provided the use of a compound of formula (I) or a pharmaceutically acceptable salt or solvate thereof for the manufacture of a medicament for the treatment of patients with an inflammatory and/or allergic condition.
According to another aspect of the invention, there is provided the use of a compound of formula (I) or a pharmaceutically acceptable salt or solvate thereof for the manufacture of a medicament for the treatment of patients with rheumatoid arthritis, asthma, COPD, allergy and/or rhinitis.
According to another aspect of the invention, there is provided the use of a compound of formula (I) or a pharmaceutically acceptable salt or solvate thereof for the manufacture of a medicament for the treatment of patients with skin disease.
According to another aspect of the invention, there is provided the use of a compound of formula (I) or a pharmaceutically acceptable salt or solvate thereof for the manufacture of a medicament for the treatment of patients with eczema, psoriasis, allergic dermatitis, neurodermatitis, pruritis and/or hypersensitivity reactions.
In another aspect, there is provided a method for the treatment of a human or animal subject with an inflammatory and/or allergic condition which method comprises administering to said human or animal subject an effective amount of a compound of formula (I) or a pharmaceutically acceptable salt or solvate thereof.
In another aspect, there is provided a method for the treatment of a human or animal subject with rheumatoid arthritis, asthma, COPD, allergy and/or rhinitis which method comprises administering to said human or animal subject an effective amount of a compound of formula (I) or a pharmaceutically acceptable salt or solvate thereof.
In another aspect, there is provided a method for the treatment of a human or animal subject with skin disease which method comprises administering to said human or animal subject an effective amount of a compound of formula (I) or a pharmaceutically acceptable salt or solvate thereof.
In a further aspect, there is provided a method for the treatment of a human or animal subject with eczema, psoriasis, allergic dermatitis, neurodermatitis, pruritis and/or hypersensitivity reactions, which method comprises administering to said human or animal subject an effective amount of a compound of formula (I) or a pharmaceutically acceptable salt or solvate thereof.
The compounds of formula (I) and pharmaceutically acceptable salts and solvates thereof may be formulated for administration in any convenient way, and the invention therefore also includes within its scope pharmaceutical compositions comprising a compound of formula (I) or a pharmaceutically acceptable salt or solvate thereof together, if desirable, in admixture with one or more physiologically acceptable diluents or carriers.
Further, there is provided a process for the preparation of such pharmaceutical compositions which comprises mixing the ingredients.
The compounds of formula (I) and pharmaceutically acceptable salts and solvates thereof may, for example, be formulated for oral, nasal, inhaled, buccal, sublingual, parenteral, topical rectal administration or other topical administration.
For internal administration the compounds of formula (I) and pharmaceutically acceptable salts and solvates thereof may, for example, be formulated in conventional manner for oral, parenteral or rectal administration. Formulations for oral administration include solutions, syrups, elixirs, powders, granules, tablets and capsules which typically contain conventional excipients such as binding agents, fillers, lubricants, disintegrants, wetting agents, suspending agents, emulsifying agents, preservatives, buffer salts, flavouring, colouring and/or sweetening agents as appropriate. Dosage unit forms may be preferred as described below.
For instance, for oral administration in the form of a tablet or capsule, the active drug component can be combined with an oral, non-toxic pharmaceutically acceptable inert carrier such as ethanol, glycerol, water and the like. The tablets may also contain excipients such as microcrystalline cellulose, lactose, sodium citrate, calcium carbonate, dibasic calcium phosphate and glycine, disintegrants such as starch (preferably corn, potato or tapioca starch), sodium starch glycollate, croscarmellose sodium and certain complex silicates, and granulation binders such as polyvinylpyrrolidone, hydroxypropylmethylcellulose (HPMC), hydroxypropylcellulose (HPC), sucrose, gelatin and acacia. Additionally, lubricating agents such as magnesium stearate, stearic acid, glyceryl behenate and talc may be included.
Solid compositions of a similar type may also be employed as fillers in gelatin capsules. Examples of excipients in this regard include lactose, starch, a cellulose, milk sugar or high molecular weight polyethylene glycols. For aqueous suspensions and/or elixirs, the agent may be combined with various sweetening or flavouring agents, colouring matter or dyes, with emulsifying and/or suspending agents and with diluents such as water, ethanol, propylene glycol and glycerin, and combinations thereof. Powders are prepared by comminuting the compound to a suitable fine size and mixing with a similarly comminuted pharmaceutical carrier such as an edible carbohydrate, as, for example, starch or mannitol. Flavouring, preservative, dispersing and colouring agent can also be present.
Capsules can be made by preparing a powder mixture as described above, and filling formed gelatin sheaths. Glidants and lubricants such as colloidal silica, talc, magnesium stearate, calcium stearate or solid polyethylene glycol can be added to the powder mixture before the filling operation. A disintegrating or solubilizing agent such as agar- agar, calcium carbonate or sodium carbonate can also be added to improve the availability of the medicament when the capsule is ingested.
Moreover, when desired or necessary, suitable binders, lubricants, disintegrating agents and coloring agents can also be incorporated into the mixture. Suitable binders include starch, gelatin, natural sugars such as glucose or beta-lactose, corn sweeteners, natural and synthetic gums such as acacia, tragacanth or sodium alginate, carboxymethylcellulose, polyethylene glycol, waxes and the like. Lubricants used in these dosage forms include sodium oleate, sodium stearate, magnesium stearate, sodium benzoate, sodium acetate, sodium chloride and the like. Disintegrators include, without limitation, starch, methyl cellulose, agar, bentonite, xanthan gum and the like.
Tablets are formulated, for example, by preparing a powder mixture, granulating or slugging, adding a lubricant and disintegrant and pressing into tablets. A powder mixture is prepared by mixing the compound, suitably comminuted, with a diluent or base as described above, and optionally, with a binder such as carboxymethylcellulose, an aliginate, gelatin, or polyvinyl pyrrolidone, a solution retardant such as paraffin, a resorption accelerator such as a quaternary salt and/or an absorption agent such as bentonite, kaolin or dicalcium phosphate. The powder mixture can be granulated by wetting with a binder such as syrup, starch paste, acadia mucilage or solutions of cellulosic or polymeric materials and forcing through a screen. As an alternative to granulating, the powder mixture can be run through the tablet machine and the result is imperfectly formed slugs broken into granules. The granules can be lubricated to prevent sticking to the tablet forming dies by means of the addition of stearic acid, a stearate salt, talc or mineral oil. The lubricated mixture is then compressed into tablets. The compounds of the present invention can also be combined with free flowing inert carrier and compressed into tablets directly without going through the granulating or slugging steps A clear or opaque protective coating consisting of a sealing coat of shellac, a coating of sugar or polymeric material and a polish coating of wax can be provided Dyestuffs can be added to these coatings to distinguish different unit dosages
Oral fluids such as solutions, syrups and elixirs can be prepared in dosage unit form so that a given quantity contains a predetermined amount of the compound Syrups can be prepared by dissolving the compound in a suitably flavoured aqueous solution, while elixirs are prepared through the use of a non-toxic alcoholic vehicle Suspensions can be formulated by dispersing the compound in a non-toxic vehicle Solubilizers and emulsifiers such as ethoxylated isostearyl alcohols and polyoxy ethylene sorbitol ethers, preservatives, flavour additives such as peppermint oil or saccharin, and the like can also be added Where appropriate, dosage unit formulations for oral administration can be microencapsulated The formulation can also be prepared to prolong or sustain the release as for example by coating or embedding particulate material in polymers, wax or the like
The compounds of formula (I) and pharmaceutically acceptable salts and solvates thereof can also be administered in the form of liposome delivery systems, such as small unilamellar vesicles, large unilamellar vesicles and multilamellar vesicles Liposomes can be formed from a variety of phospholipids, such as cholesterol, stearylamine or phosphatidylcholines
The compounds of formula (I) and pharmaceutically acceptable salts and solvates thereof can also be administered in the form of liposome emulsion delivery systems, such as small unilamellar vesicles, large unilamellar vesicles and multilamellar vesicles Liposomes can be formed from a variety of phospholipids, such as cholesterol stearylamine or phosphatidylcholines
Topical administration as used herein, includes administration by insufflation and inhalation Examples of various types of preparation for local administration include ointments, lotions, creams, gels, foams, preparations for delivery by transdermal patches, powders, sprays, aerosols, capsules or cartridges for use in an inhaler or insufflator or drops (e g eye or nose drops), solutions/suspensions for nebuhsation, suppositories, pessaries, retention enemas and chewable or suckable tablets or pellets (e g for the treatment of aphthous ulcers) or liposome or microencapsulation preparations Formulations for administration topically to the nose for example, for the treatment of rhinitis, include pressurised aerosol formulations and aqueous formulations administered to the nose by pressurised pump Formulations which are non-pressurised and adapted to be administered topically to the nasal cavity are of particular interest Suitable formulations contain water as the diluent or carrier for this purpose Aqueous formulations for administration to the lung or nose may be provided with conventional excipients such as buffering agents, tonicity modifying agents and the like Aqueous formulations may also be administered to the nose by nebulisation
In one embodiment the compounds of formula (I) or a pharmaceutically acceptable salt or solvate thereof may be formulated for administration topically to the nose as a fluid formulation for delivery from a fluid dispenser, for example a fluid dispenser having a dispensing nozzle or dispensing orifice through which a metered dose of the fluid formulation is dispensed upon the application of a user-applied force to a pump mechanism of the fluid dispenser Such fluid dispensers are generally provided with a reservoir of multiple metered doses of the fluid formulation, the doses being dispensable upon sequential pump actuations The dispensing nozzle or orifice may be configured for insertion into the nostrils of the user for spray dispensing of the fluid formulation into the nasal cavity A fluid dispenser of the aforementioned type is described and illustrated in WO05/044354, the entire content of which is hereby incorporated herein by reference The dispenser has a housing which houses a fluid discharge device having a compression pump mounted on a container for containing a fluid formulation The housing has at least one finger-operable side lever which is movable inwardly with respect to the housing to cam the container upwardly in the housing to cause the pump to compress and pump a metered dose of the formulation out of a pump stem through a nasal nozzle of the housing In one embodiment, the fluid dispenser is of the general type illustrated in Figures 30-40 of WO05/044354
Ointments, creams and gels may, for example, be formulated with an aqueous or oily base with the addition of suitable thickening and/or gelling agent and/or solvents Such bases may thus, for example, include water and/or an oil such as liquid paraffin or a vegetable oil such as arachis oil or castor oil, or a solvent such as polyethylene glycol Thickening agents and gelling agents which may be used according to the nature of the base include soft paraffin, aluminium stearate, cetostearyl alcohol, polyethylene glycols, woolfat, beeswax, carboxypolymethylene and cellulose derivatives, and/or glyceryl monostearate and/or non-ionic emulsifying agents.
Lotions may be formulated with an aqueous or oily base and will in general also contain one or more emulsifying agents, stabilising agents, dispersing agents, suspending agents or thickening agents.
Powders for external application may be formed with the aid of any suitable powder base, for example, talc, lactose or starch. Drops may be formulated with an aqueous or non- aqueous base also comprising one or more dispersing agents, solubilising agents, suspending agents or preservatives.
In one embodiment the compounds of formula (I) and pharmaceutically acceptable salts and solvates thereof may be formulated as a dry powder for administration by inhalation.
Optionally, in particular for dry powder inhalable compositions, a composition suitable for inhaled administration may be incorporated into a plurality of sealed dose containers (e.g. containing the dry powder composition) mounted longitudinally in a strip or ribbon inside a suitable inhalation device. The container is rupturable or peel-openable on demand and the dose of e.g. the dry powder composition may be administered by inhalation via a device such as the DISKUS™ device, marketed by GlaxoSmithKline. The DISKUS™ inhalation device is, for example, described in GB2242134A, and in such a device, at least one container for the composition in powder form (the container or containers preferably being a plurality of sealed dose containers mounted longitudinally in a strip or ribbon) is defined between two members peelably secured to one another; the device comprises: a means of defining an opening station for the said container or containers; a means for peeling the members apart at the opening station to open the container; and an outlet, communicating with the opened container, through which a user can inhale the composition in powder form from the opened container.
Spray compositions may for example be formulated as aqueous solutions or suspensions or as aerosols delivered from pressurised packs, such as a metered dose inhaler, with the use of a suitable liquefied propellant. Aerosol compositions suitable for inhalation can be either a suspension or a solution and generally contain a compound of formula (I) and a suitable propellant such as a fluorocarbon or hydrogen-containing chlorofluorocarbon or mixtures thereof, particularly hydrofluoroalkanes, especially 1 ,1 ,1 ,2-tetrafluoroethane, 1,1 ,1 ,2,3,3,3-heptafluoro-n-propane or a mixture thereof. The aerosol composition may optionally contain additional formulation excipients well known in the art such as surfactants e.g. oleic acid, lecithin or an oligolactic acid of derivative e.g. As described in WO94/21229 and WO98/34596 and cosolvents e.g. ethanol.
There is thus provided as a further aspect of the invention a pharmaceutical aerosol formulation comprising a compound of the invention and a fluorocarbon or hydrogen- containing chlorofluorocarbon as propellant, optionally in combination with a surfactant and/or a cosolvent.
According to another aspect of the invention, there is provided a pharmaceutical aerosol formulation wherein the propellant is selected from 1 ,1 ,1 ,2-tetrafluoroethane, 1 ,1 ,1 ,2,3,3,3-heptafluoro-n-propane and mixtures thereof.
The formulations of the invention may be buffered by the addition of suitable buffering agents.
Aerosol formulations are preferably arranged so that each metered dose or "puff of aerosol contains from 20μg to 10mg, preferably from 20μg to 2000μg, more preferably from about 20μg to 500μg of a compound of formula (I). Administration may be once daily or several times daily, for example 2, 3, 4 or 8 times, giving for example 1 , 2 or 3 doses each time. The overall daily dose with an aerosol will be within the range from 100μg to 10mg, preferably from 200μg to 2000μg. The overall daily dose and the metered dose delivered by capsules and cartridges in an inhaler or insufflator will generally be double that delivered with aerosol formulations.
In the case of suspension aerosol formulations, the particle size of the particulate (e.g., micronised) drug should be such as to permit inhalation of substantially all the drug into the lungs upon administration of the aerosol formulation and will thus be less than 100 microns, desirably less than 20 microns, and in particular in the range of from 1 to 10 microns, such as from 1 to 5 microns, more preferably from 2 to 3 microns.
The formulations of the invention may be prepared by dispersal or dissolution of the medicament and a compound of the invention in the selected propellant in an appropriate container, for example, with the aid of sonication or a high-shear mixer. The process is desirably carried out under controlled humidity conditions. The chemical and physical stability and the pharmaceutical acceptability of the aerosol formulations according to the invention may be determined by techniques well known to those skilled in the art. Thus, for example, the chemical stability of the components may be determined by HPLC assay, for example, after prolonged storage of the product. Physical stability data may be gained from other conventional analytical techniques such as, for example, by leak testing, by valve delivery assay (average shot weights per actuation), by dose reproducibility assay (active ingredient per actuation) and spray distribution analysis.
The stability of the suspension aerosol formulations according to the invention may be measured by conventional techniques, for example, by measuring flocculation size distribution using a back light scattering instrument or by measuring particle size distribution by cascade impaction or by the "twin impinger" analytical process. As used herein reference to the "twin impinger" assay means "Determination of the deposition of the emitted dose in pressurised inhalations using apparatus A" as defined in British Pharmacopaeia 1988, pages A204-207, Appendix XVII C. Such techniques enable the "respirable fraction" of the aerosol formulations to be calculated. One method used to calculate the "respirable fraction" is by reference to "fine particle fraction" which is the amount of active ingredient collected in the lower impingement chamber per actuation expressed as a percentage of the total amount of active ingredient delivered per actuation using the twin impinger method described above.
MDI canisters generally comprise a container capable of withstanding the vapour pressure of the propellant used such as a plastic or plastic-coated glass bottle or preferably a metal can, for example, aluminium or an alloy thereof which may optionally be anodised, lacquer-coated and/or plastic-coated (for example incorporated herein by reference WO96/32099 wherein part or all of the internal surfaces are coated with one or more fluorocarbon polymers optionally in combination with one or more non-fluorocarbon polymers), which container is closed with a metering valve. The cap may be secured onto the can via ultrasonic welding, screw fitting or crimping. MDIs taught herein may be prepared by methods of the art (e.g. see Byron, above and WO96/32099). Preferably the canister is fitted with a cap assembly, wherein a drug-metering valve is situated in the cap, and said cap is crimped in place. The term "metered dose inhaler" or MDI means a unit comprising a can, a secured cap covering the can and a formulation metering valve situated in the cap. MDI system includes a suitable channelling device. Suitable channelling devices comprise for example, a valve actuator and a cylindrical or cone-like passage through which medicament may be delivered from the filled canister via the metering valve to the nose or mouth of a patient such as a mouthpiece actuator.
In one embodiment of the invention the metallic internal surface of the can is coated with a fluoropolymer, most preferably blended with a non-fluoropolymer. In another embodiment of the invention the metallic internal surface of the can is coated with a polymer blend of polytetrafluoroethylene (PTFE) and polyethersulfone (PES). In a further embodiment of the invention the whole of the metallic internal surface of the can is coated with a polymer blend of polytetrafluoroethylene (PTFE) and polyethersulfone (PES).
The metering valves are designed to deliver a metered amount of the formulation per actuation and incorporate a gasket to prevent leakage of propellant through the valve. The gasket may comprise any suitable elastomeric material such as, for example, low density polyethylene, chlorobutyl, bromobutyl, EPDM, black and white butadiene- acrylonitrile rubbers, butyl rubber and neoprene. Suitable valves are commercially available from manufacturers well known in the aerosol industry, for example, from Valois, France (e.g. DF10, DF30, DF60), Bespak pic, UK (e.g. BK300, BK357) and 3M-
TM
Neotechnic Ltd, UK (e.g. Spraymiser ).
In various embodiments, the MDIs may also be used in conjunction with other structures such as, without limitation, overwrap packages for storing and containing the MDIs, including those described in U.S. Patent Nos. 6,1 19,853; 6, 179,1 18; 6,315,1 12; 6,352,152; 6,390,291 ; and 6,679,374, as well as dose counter units such as, but not limited to, those described in U.S. Patent Nos. 6,360,739 and 6,431 ,168.
Conventional bulk manufacturing methods and machinery well known to those skilled in the art of pharmaceutical aerosol manufacture may be employed for the preparation of large-scale batches for the commercial production of filled canisters. Thus, for example, in one bulk manufacturing method for preparing suspension aerosol formulations a metering valve is crimped onto an aluminium can to form an empty canister. The particulate medicament is added to a charge vessel and liquefied propellant together with the optional excipients is pressure filled through the charge vessel into a manufacturing vessel. The drug suspension is mixed before recirculation to a filling machine and an aliquot of the drug suspension is then filled through the metering valve into the canister. In one example bulk manufacturing method for preparing solution aerosol formulations a metering valve is crimped onto an aluminium can to form an empty canister. The liquefied propellant together with the optional excipients and the dissolved medicament is pressure filled through the charge vessel into a manufacturing vessel.
In an alternative process, an aliquot of the liquefied formulation is added to an open canister under conditions which are sufficiently cold to ensure the formulation does not vaporise, and then a metering valve crimped onto the canister.
Typically, in batches prepared for pharmaceutical use, each filled canister is check- weighed, coded with a batch number and packed into a tray for storage before release testing.
Capsules and cartridges for use in an inhaler or insufflator, of for example gelatine, may be formulated containing a powder mix for inhalation of a compound of the invention and a suitable powder base such as lactose or starch. Each capsule or cartridge may generally contain from 20μg to 10mg of the compound of formula (I) or a pharmaceutically acceptable salt or solvate thereof. Alternatively, the compound of the invention may be presented without excipients such as lactose.
The proportion of the active compound of formula (I) or a pharmaceutically acceptable salt or solvate thereof in the local compositions according to the invention depends on the precise type of formulation to be prepared but will generally be within the range of from 0.001 to 10% by weight. Generally, for most types of preparations, the proportion used will be within the range of from 0.005 to 1%, for example from 0.01 to 0.5%. However, in powders for inhalation or insufflation the proportion used will normally be within the range of from 0.1 to 5%.
Topical preparations may be administered by one or more applications per day to the affected area; over skin areas occlusive dressings may advantageously be used. Continuous or prolonged delivery may be achieved by an adhesive reservoir system.
Typically, a physician will determine the actual dosage which will be most suitable for an individual subject. The specific dose level and frequency of dosage for any particular individual may be varied and will depend upon a variety of factors including the activity of the specific compound employed, the metabolic stability and length of action of that compound, the age, body weight, general health, sex, diet, mode and time of administration, rate of excretion, drug combination, the severity of the particular condition, and the individual undergoing therapy.
For oral administration to humans, the daily dosage level of the agent may be in single or divided doses.
For systemic administration the daily dose as employed for adult human treatment will range from 0.5-100mg/kg body weight, preferably 0.5-60mg/kg body weight, which may be administered in 1 to 4 daily doses, for example, depending on the route of administration and the condition of the patient. When the composition comprises dosage units, each unit will preferably contain 5mg to 1g of active ingredient. The duration of treatment will be dictated by the rate of response rather than by arbitrary numbers of days.
The compounds of the invention may in general be given by internal administration in cases wherein systemic glucocorticoid receptor agonist therapy is indicated.
Slow release or enteric coated formulations may be advantageous, particularly for the treatment of inflammatory bowel disorders.
In some embodiments, the compounds of the invention will be formulated for oral administration. In other embodiments, the compounds of the invention will be formulated for inhaled administration. In further embodiments, the compounds of the invention will be formulated for intranasal administration.
The compounds and pharmaceutical formulations according to the invention may be used in combination with or include one or more other therapeutic agents, for example when the compounds of the invention are administered intranasally or by inhalation. Suitable other therapeutic agents may be selected from for example anti-inflammatory agents, anticholinergic agents (particularly an M1ZM2ZM3 receptor antagonist), β2-adrenoreceptor agonists, antiinfective agents such as antibiotics or antivirals, or antihistamines. The invention thus provides, in a further aspect, a combination comprising a compound of the invention together with one or more other therapeutically active agents, for example selected from an anti-inflammatory agent such as a corticosteroid or an NSAID, an anticholinergic agent, a β2-adrenoreceptor agonist, an antiinfective agent such as an antibiotic or an antiviral, or an antihistamine. One embodiment of the invention encompasses combinations comprising a compound of the invention together with a β2- adrenoreceptor agonist, and/or an anticholinergic, and/or a PDE-4 inhibitor, and/or an antihistamine.
One embodiment of the invention encompasses combinations comprising one or two other therapeutic agents.
It will be clear to a person skilled in the art that, where appropriate, the other therapeutic ingredient(s) may be used in the form of salts, for example as alkali metal or amine salts or as acid addition salts, or prodrugs, or as esters, for example lower alkyl esters, or as solvates, for example hydrates, to optimise the activity and/or stability and/or physical characteristics, such as solubility, of the therapeutic ingredient. It will be clear also that, where appropriate, the therapeutic ingredients may be used in optically pure form.
In one embodiment, the invention encompasses a combination comprising a compound of the invention together with a β2-adrenoreceptor agonist.
Examples of β2-adrenoreceptor agonists include salmeterol (e.g. as the racemate or a single enantiomer, such as the f?-enantiomer), salbutamol (e.g. as the racemate or a single enantiomer such as the f?-enantiomer), formoterol (e.g. as the racemate or a single diastereomer such as the f?,f?-diastereomer), salmefamol, fenoterol, carmoterol, etanterol, naminterol, clenbuterol, pirbuterol, flerobuterol, reproterol, bambuterol, indacaterol or terbutaline and salts thereof, for example the xinafoate (1-hydroxy-2- naphthalenecarboxylate) salt of salmeterol, the sulphate salt or free base of salbutamol or the fumarate salt of formoterol. In one embodiment, the β2-adrenoreceptor agonists are long-acting β2-adrenoreceptor agonists, for example compounds which provide effective bronchodilation for about 12 hours or longer. Examples of β2-adrenoreceptor agonists may include those described in WO02/066422A, WO02/070490, WO02/076933, WO03/024439, WO03/072539, WO 03/091204, WO04/016578, WO04/022547, WO04/037807, WO04/037773, WO04/037768, WO04/039762, WO04/039766, WO01/42193 and WO03/042160.
Examples of β2-adrenoreceptor agonists include:
3-(4-{[6-({(2/?)-2-hydroxy-2-[4-hydroxy-3-(hydroxymethyl)pheny|]ethyl}amino) hexyl]oxy}butyl)benzenesulfonamide; 3-(3-{[7-({(2/?)-2-hydroxy-2-[4-hydroxy-3-hydroxymethyl)phenyl]ethyl}- amino)heptyl]oxy}propyl)benzenesulfonamide;
4-{(1f?)-2-[(6-{2-[(2,6-dichlorobenzyl)oxy]ethoxy}hexyl)amino]-1-hydroxyethyl}-2-
(hydroxymethyl)phenol;
4-{(1f?)-2-[(6-{4-[3-(cyclopentylsulfonyl)phenyl]butoxy}hexyl)amino]-1-hydroxyethyl}-2- (hydroxymethyl)phenol;
N-[2-hydroxyl-5-[( 1 R)-1 -hydroxy-2-[[2-4-[[(2/?)-2-hydroxy-2- phenylethyl]amino]pheny|]ethy|]amino]ethy|]pheny|]foramide,
N-2{2-[4-(3-phenyl-4-methoxyphenyl)aminophenyl]ethyl}-2-hydroxy-2-(8-hydroxy-2(1 W)- quinolinon-5-yl)ethylamine, and 5-[(/?)-2-(2-{4-[4-(2-amino-2-methyl-propoxy)-phenylamino]-phenyl}-ethylamino)-1- hydroxy-ethyl]-8-hydroxy-1 H-quinolin-2-one, and salts thereof.
The β2-adrenoreceptor agonist may be in the form of a salt formed with a pharmaceutically acceptable acid selected from sulphuric, hydrochloric, fumaric, hydroxynaphthoic (for example 1- or 3-hydroxy-2-naphthoic), cinnamic, substituted cinnamic, triphenylacetic, sulphamic, sulphanilic, naphthaleneacrylic, benzoic, 4-methoxybenzoic, 2- or 4-hydroxybenzoic, 4-chlorobenzoic and 4-phenylbenzoic acid.
Suitable anti-inflammatory agents include corticosteroids. Examples of corticosteroids which may be used in combination with the compounds of the invention are those oral and inhaled corticosteroids and their pro-drugs which have anti-inflammatory activity. Examples include methyl prednisolone, prednisolone, dexamethasone, fluticasone propionate, 6α,9α-difluoro-11β-hydroxy-16α-methyl-17α-[(4-methyl-1 ,3-thiazole-5- carbonyl)oxy]-3-oxo-androsta-1 ,4-diene-17β-carbothioic acid S-fluoromethyl ester, 6α,9α- difluoro-17α-[(2-furanylcarbonyl)oxy]-11 β-hydroxy-16α-methyl-3-oxo-androsta-1 ,4-diene- 17β-carbothioic acid S-fluoromethyl ester (fluticasone furoate), 6α,9α-difluoro-11β- hydroxy-iθα-methyl-S-oxo-ITα-propionyloxy- androsta-1 ,4-diene-17β-carbothioic acid S- (2-oxo-tetrahydro-furan-3S-yl) ester, 6α,9α-difluoro-11 β-hydroxy-16α-methyl-3-oxo-17α- (2,2,3,3- tetramethycyclopropylcarbonyl)oxy-androsta-1 ,4-diene-17β-carbothioic acid S- cyanomethyl ester and 6α,9α-difluoro-11β-hydroxy-16α-methyl-17α-(1- methycyclopropylcarbonyl)oxy-3-oxo-androsta-1 ,4-diene-17β-carbothioic acid S- fluoromethyl ester, beclomethasone esters (for example the 17-propionate ester or the 17,21-dipropionate ester), budesonide, flunisolide, mometasone esters (for example mometasone furoate), triamcinolone acetonide, rofleponide, ciclesonide (16α,17-[[(R)- cyclohexylmethylene]bis(oxy)]-11 β,21-dihydroxy-pregna-1 ,4-diene-3,20-dione), butixocort propionate, RPR-106541 , and ST-126. In one embodiment corticosteroids include fluticasone propionate, 6α,9α-difluoro-11 β-hydroxy-16α-methyl-17α-[(4-methyl-1 ,3- thiazole-5-carbonyl)oxy]-3-oxo-androsta-1 ,4-diene-17β-carbothioic acid S-fluoromethyl ester, δα.θα-difluoro-^α-f^-furanylcarbony^oxyl-i i β-hydroxy-iδα-methyl-S-oxo- androsta-1 ,4-diene-17β-carbothioic acid S-fluoromethyl ester, 6α,9α-difluoro-11 β-hydroxy- 16α-methyl-3-oxo-17α-(2,2,3,3- tetramethycyclopropylcarbonyOoxy-androsta-i ,4-diene- 17β-carbothioic acid S-cyanomethyl ester and 6α,9α-difluoro-1 1 β-hydroxy-16α-methyl- 17α-(1-methycyclopropylcarbonyl)oxy-3-oxo-androsta-1 ,4-diene-17β-carbothioic acid S- fluoromethyl ester. In one embodiment the corticosteroid is 6α,9α-difluoro-17α-[(2- furanylcarbonyl)oxy]-11 β-hydroxy-16α-methyl-3-oxo-androsta-1 ,4-diene-17β-carbothioic acid S-fluoromethyl ester.
Examples of corticosteroids may include those described in WO02/088167, WO02/100879, WO02/12265, WO02/12266, WO05/005451 , WO05/005452, WO06/072599 and WO06/072600.
Non-steroidal compounds having glucocorticoid agonism that may possess selectivity for transrepression over transactivation and that may be useful in combination therapy include those covered in the following published patent applications and patents: WO2003/082827, WO1998/54159, WO2004/005229, WO2004/009017, WO2004/018429, WO2003/104195, WO2003/082787, WO2003/082280, WO2003/059899,
WO2003/101932, WO2002/02565, WO2001/16128, WO2000/66590, WO2003/086294, WO2004/026248, WO2003/061651 , WO2003/08277, WO2006/000401 , WO2006/000398 and WO2006/015870. Examples of anti-inflammatory agents include non-steroidal anti-inflammatory drugs (NSAID's)
Examples of NSAID's include sodium cromoglycate, nedocromil sodium, phosphodiesterase (PDE) inhibitors (for example, theophylline, PDE4 inhibitors or mixed PDE3/PDE4 inhibitors), leukotriene antagonists, inhibitors of leukotriene synthesis (for example, montelukast), iNOS inhibitors, tryptase and elastase inhibitors, beta-2 integrin antagonists and adenosine receptor agonists or antagonists (for example, adenosine 2a agonists), cytokine antagonists (for example, chemokine antagonists, such as a CCR3 antagonist) or inhibitors of cytokine synthesis, or 5-lιpoxygenase inhibitors An iNOS (inducible nitric oxide synthase inhibitor) is preferably for oral administration Suitable iNOS inhibitors include those disclosed in WO93/13055, WO98/30537, WO02/50021 , WO95/34534 and WO99/62875 Suitable CCR3 inhibitors include those disclosed in WO02/26722 Adenosine 2a agonists include those discussed in WO05/116037
In one embodiment, the invention provides the use of the compounds of the invention in combination with a phosphodiesterase 4 (PDE4) inhibitor, for example in the case of a formulation adapted for inhalation The PDE4-specιfιc inhibitor may be any compound that is known to inhibit the PDE4 enzyme or which is discovered to act as a PDE4 inhibitor, and which are only PDE4 inhibitors, not compounds which inhibit other members of the PDE family, such as PDE3 and PDE5, as well as PDE4
Compounds include c/s-4-cyano-4-(3-cyclopentyloxy-4-methoxyphenyl)cyclohexan-1- carboxylic acid, 2-carbomethoxy-4-cyano-4-(3-cyclopropylmethoxy-4- dιfluoromethoxyphenyl)cyclohexan-1-one and c/s-[4-cyano-4-(3-cyclopropylmethoxy-4- dιfluoromethoxyphenyl)cyclohexan-1-ol] Another compound is c/s-4-cyano-4-[3- (cyclopentyloxy)-4-methoxyphenyl]cyclohexane-1-carboxylιc acid (also known as cilomilast) and its salts, esters, pro-drugs or physical forms, which is described in U S patent 5,552,438 issued 03 September, 1996, this patent and the compounds it discloses are incorporated herein in full by reference
Other compounds include AWD-12-281 (N-(3,5-dιchloro-4-pyπdιnyl)-1-[4- fluorophenyl)methyl]-5-hydroxy-α-oxo-1 H-ιndol-3-acetamιde) from Elbion (Hofgen, N et al 15th EFMC lnt Symp Med Chem (Sept 6-10, Edinburgh) 1998, Abst P 98, CAS reference No 247584020-9), a 9-benzyladenιne derivative nominated NCS-613 (INSERM), D-4418 from Chiroscience and Schering-Plough, a benzodiazepine PDE4 inhibitor identified as CI-1018 (PD-168787) and attributed to Pfizer; a benzodioxole derivative disclosed by Kyowa Hakko in WO99/16766; K-34 from Kyowa Hakko; V- 11294A from Napp (Landells, LJ. et al. Eur Resp J [Annu Cong Eur Resp Soc (Sept 19- 23, Geneva) 1998] 1998, 12 (Suppl. 28): Abst P2393); roflumilast (CAS reference No 162401-32-3) and a pthalazinone (WO99/47505, the disclosure of which is hereby incorporated by reference) from Byk-Gulden; Pumafentrine, (-)-p-[(4aR*,10ύS*)-9-ethoxy- 1 ,2,3,4,4a, 10b-hexahyd ro-8-methoxy-2-methylbenzo[c][1 ,6]naphthyridin-6-yl]-N, N- diisopropylbenzamide which is a mixed PDE3/PDE4 inhibitor which has been prepared and published on by Byk-Gulden, now Altana; arofylline under development by Almirall- Prodesfarma; VM554/UM565 from Vernalis; or T-440 (Tanabe Seiyaku; Fuji, K. et al. J Pharmacol Exp Ther,1998, 284(1 ): 162), and T2585.
Further compounds are disclosed in the published international patent application WO04/024728 (Glaxo Group Ltd), WO04/056823 (Glaxo Group Ltd) and WO04/103998 (Glaxo Group Ltd).
Examples of anticholinergic agents are those compounds that act as antagonists at the muscarinic receptors, in particular those compounds which are antagonists of the M1 or M3 receptors, dual antagonists of the M1ZM3 or M2/M3, receptors or pan-antagonists of the M1ZM2ZM3 receptors. Exemplary compounds for administration via inhalation include ipratropium (for example, as the bromide, CAS 22254-24-6, sold under the name Atrovent), oxitropium (for example, as the bromide, CAS 30286-75-0) and tiotropium (for example, as the bromide, CAS 136310-93-5, sold under the name Spiriva). Also of interest are revatropate (for example, as the hydrobromide, CAS 262586-79-8) and LAS- 34273 which is disclosed in WO01Z04118. Exemplary compounds for oral administration include pirenzepine (for example, CAS 28797-61-7), darifenacin (for example, CAS 133099-04-4, or CAS 133099-07-7 for the hydrobromide sold under the name Enablex), oxybutynin (for example, CAS 5633-20-5, sold under the name Ditropan), terodiline (for example, CAS 15793-40-5), tolterodine (for example, CAS 124937-51-5, or CAS 124937- 52-6 for the tartrate, sold under the name Detrol), otilonium (for example, as the bromide, CAS 26095-59-0, sold under the name Spasmomen), trospium chloride (for example, CAS 10405-02-4) and solifenacin (for example, CAS 242478-37-1 , or CAS 242478-38-2, or the succinate also known as YM-905 and sold under the name Vesicare). Additional compounds are disclosed in WO 2005/037280, WO 2005/046586 and WO 2005/104745, incorporated herein by reference. The present combinations include, but are not limited to:
(3-eπdo)-3-(2,2-di-2-thienylethenyl)-8,8-dimethyl-8-azoniabicyclo[3.2.1 Joctane iodide; (3-eπdo)-3-(2-cyano-2,2-diphenylethyl)-8,8-dimethyl-8-azoniabicyclo[3.2.'l]octane bromide;
4-[hydroxy(diphenyl)methy|]-1-{2-[(phenylmethyl)oxy]ethyl}-1-azoniabicyclo[2.2.2]octane bromide; and
(IR.SSJ-S^-cyano^^-diphenylethylJ-S-methyl-S^-^phenylmethyOoxylethylJ-S- azoniabicyclo[3.2.1]octane bromide.
Other anticholinergic agents include compounds of formula (A)1 which are disclosed in US patent application 60/487981 :
Figure imgf000032_0001
in which the preferred orientation of the alkyl chain attached to the tropane ring is endo;
R31 and R32 are, independently, selected from the group consisting of straight or branched chain lower alkyl groups having preferably from 1 to 6 carbon atoms, cycloalkyl groups having from 5 to 6 carbon atoms, cycloalkyl-alkyl having from 6 to 10 carbon atoms, 2- thienyl, 2-pyridyl, phenyl, phenyl substituted with an alkyl group having not in excess of 4 carbon atoms and phenyl substituted with an alkoxy group having not in excess of 4 carbon atoms;
X" represents an anion associated with the positive charge of the N atom. X" may be but is not limited to chloride, bromide, iodide, sulfate, benzene sulfonate, and toluene sulfonate, including, for example: (3-endo)-3-(2,2-di-2-thienylethenyl)-8,8-dimethyl-8-azoniabicyclo[3.2.1]octane bromide;
(3-endo)-3-(2,2-diphenylethenyl)-8,8-dimethyl-8-azoniabicyclo[3.2.1]octane bromide;
(3-endo)-3-(2,2-diphenylethenyl)-8,8-dimethyl-8-azoniabicyclo[3.2.1 ]octane 4- methylbenzenesulfonate;
(3-eπdo)-8,8-dimethyl-3-[2-phenyl-2-(2-thienyl)ethenyl]-8-azoniabicyclo[3.2.1]octane bromide; and/or
(3-endo)-8,8-dimethyl-3-[2-phenyl-2-(2-pyridinyl)ethenyl]-8-azoniabicyclo[3.2.1]octane bromide. Further anticholinergic agents include compounds of formula (B) or (C), which are disclosed in US patent application 60/511009:
Figure imgf000033_0001
wherein: the H atom indicated is in the exo position;
R41" represents an anion associated with the positive charge of the N atom; R41' may be but is not limited to chloride, bromide, iodide, sulfate, benzene sulfonate and toluene sulfonate;
R42 and R43 are independently selected from the group consisting of straight or branched chain lower alkyl groups (having preferably from 1 to 6 carbon atoms), cycloalkyl groups
(having from 5 to 6 carbon atoms), cycloalkyl-alkyl (having from 6 to 10 carbon atoms), heterocycloalkyl (having from 5 to 6 carbon atoms) and N or O as the heteroatom, heterocycloalkyl-alkyl (having from 6 to 10 carbon atoms) and N or O as the heteroatom, aryl, optionally substituted aryl, heteroaryl, and optionally substituted heteroaryl;
R44 is selected from the group consisting of (CrC6)alkyl, (C3-C12)cycloalkyl, (C3-
C7)heterocycloalkyl, (C1-C6)alkyl(C3-C12)cycloalkyl, (d-CeJalkyKdrdJheterocycloalkyl, aryl, heteroaryl, (d-Ce)alkyl-aryl, (d-C6)alkyl-heteroaryl, -OR45, -CH2OR45, -CH2OH, -CN,
-CF3, -CH2O(CO)R46, -CO2R47, -CH2NH2, -CH2N(R47)SO2R45, -SO2N(R47J(R48), -
CON(R47XR48), -CH2N(R48)CO(R46), -CH2N(R48JSO2(R46), -CH2N(R48JCO2(R45), -
CH2N(R48JCONH(R47);
R45 is selected from the group consisting of (Ci-Cβjalkyl, (C1-CeJaIkVl(C3-C12)CyClOaIkVl, (C1-C6)alkyl(C3-C7)heterocycloalkyl, (d-C6)alkyl-aryl, (d-C6)alkyl-heteroaryl;
R46 is selected from the group consisting of (d-C6)alkyl, (C3-Ci2)cycloalkyl, (C3-
C7)heterocycloalkyl, (Ci-C6)alkyl(C3-C12)cycloalkyl, (d-CeJalky^Cs-CyJheterocycloalkyl, aryl, heteroaryl, (d-C6)alkyl-aryl, (d-C6)alkyl-heteroaryl;
R47 and R48 are, independently, selected from the group consisting of H, (d-C6)alkyl, (C3- Ci2)cycloalkyl, (C3-C7)heterocycloalkyl, (Ci-C6)alkyl(C3-C12)cycloalkyl, (CrC6)alkyl(C3-
C7)heterocycloalkyl, (C1-C6)alkyl-aryl, and (d-C6)alkyl-heteroaryl, including, for example: (endo)-3-(2-methoxy-2,2-di-thiophen-2-yl-ethyl)-8,8-dimethyl-δ-azonia- bicyclo[3.2.1]octane iodide;
3-((endo)-8-methyl-8-aza-bicyclo[3.2.1]oct-3-yl)-2,2-diphenyl-propionitrile;
(endoJ-δ-methyl-S^^^-triphenyl-ethylVδ-aza-bicyclop^.iJoctane; 3-((endo)-δ-methyl-δ-aza-bicydo[3.2.1]oct-3-yl)-2,2-diphenylφropionamide;
3-((endo)-8-methyl-δ-aza-bicyclo[3.2.1]oct-3-yl)-2,2-diphenyl-propionic acid;
(endo)-3-(2-cyano-2,2-diphenyl-ethyl)-δ,δ-dimethyl-δ-azonia-bicyclo[3.2.1]octane iodide;
(endo)-3-(2-cyano-2,2-diphenyl-ethyl)-δ,δ-dimethyl-δ-azonia-bicyclo[3.2.1]octane bromide;
3-((endo)-δ-methyl-δ-aza-bicyclo[3.2.1]oct-3-yl)-2,2-diphenyl-propan-1-ol; Λ/-benzyl-3-((endo)-δ-methyl-δ-aza-bicyclo[3.2.1]oct-3-yl)-2,2-diphenyl-propionamide;
(endo)-3-(2-carbamoyl-2,2-diphenyl-ethyl)-δ,δ-dimethyl-δ-azonia-bicyclo[3.2.1]octane iodide;
1-benzyl-3-[3-((endo)-δ-methyl-δ-aza-bicyclo[3.2.1]oct-3-yl)-2,2-diphenyl-propyl]-urea;
1-ethyl-3-[3-((endo)-δ-methyl-δ-aza-bicyclo[3.2.1]oct-3-yl)-2,2-diphenyl-propy|]-urea; Λ/-[3-((endo)-δ-methyl-δ-aza-bicyclo[3.2.1]oct-3-yl)-2,2-diphenyl-propyl]-acetamide;
/V-β-^endoJ-δ-methyl-δ-aza-bicycloβ^.iloct-S-yO^-diphenyl-propylJ-benzamide;
3-((endo)-δ-methyl-δ-aza-bicyclo[3.2.1]oct-3-yl)-2,2-di-thiophen-2-yl-propionitrile;
(endo)-3-(2-cyano-2,2-di-thiophen-2-yl-ethyl)-δ,δ-dimethyl-δ-azonia-bicyclo[3.2.1]octane iodide; Λ/-[3-((endo)-δ-methyl-8-aza-bicyclo[3.2.1]oct-3-yl)-2,2-diphenyl-propyl]- benzenesulfonamide;
^-((endo^δ-methyl-δ-aza-bicyclo^^.^oct-S-yO^^-diphenyl-propyll-urea;
/^-((endoJ-δ-methyl-δ-aza-bicycloβ^.ijoct-S-yO^-diphenyl-propyl]- methanesulfonamide; and/or (endo)-3-{2,2-diphenyl-3-[(1-phenyl-methanoyl)-amino]-propyl}-δ,δ-dimethyl-δ-azonia- bicyclo[3.2.1]octane bromide.
Further compounds include:
(endo)-3-(2-methoxy-2,2-di-thiophen-2-yl-ethyl)-8,δ-dimethyl-δ-azonia- bicyclo[3.2.1]octane iodide;
(endo)-3-(2-cyano-2,2-diphenyl-ethyl)-δ,8-dimethyl-8-azonia-bicyclo[3.2.1]octane iodide;
(endo)-3-(2-cyano-2,2-diphenyl-ethyl)-8,8-dimethyl-8-azonia-bicyclo[3.2.1]octane bromide;
(endo)-3-(2-carbamoyl-2,2-diphenyl-ethyl)-8,8-dimethyl-8-azonia-bicyclo[3.2.1]octane iodide; (endo)-3-(2-cyano-2,2-di-thiophen-2-yl-ethyl)-8,δ-dimethyl-δ-azonia-bicyclo[3.2.1]octane iodide; and/or (endoJ-S^^-ciiphenyl-S-^i-phenyl-methanoyO-aminol-propylJ-δ.δ-dimethyl-δ-azonia- bιcyclo[3 2 1]octane bromide
Examples of antihistamines (also referred to as H 1 -receptor antagonists) include any one or more of the numerous antagonists known which inhibit H 1 -receptors, and are safe for human use First generation antagonists, include derivatives of ethanolammes, ethylenediamines, and alkylamines, such as diphenylhydramine, pyπlamine, clemastine, chlorpheniramine Second generation antagonists, which are non-sedating, include loratidine, desloratidine, terfenadine, astemizole, acrivastine, azelastine, levocetiπzine fexofenadine and cetirizine
Examples of antihistamines include loratidine, desloratidine, fexofenadine, cetirizine, levocabastine, olopatadine, amlexanox and epinastine
Of particular interest is a combination comprising a compound of the invention together with an H1 antagonist Suitable H1 antagonists include, without limitation, amelexanox, astemizole, azatadine, azelastine, acrivastine, brompheniramine, cetirizine, levocetirizine, efletiπzine, chlorpheniramine, clemastine, cyclizine, carebastine, cyproheptadine, carbinoxamine, descarboethoxyloratadine, doxylamine, dimethindene, ebastine, epinastine, efletinzine, fexofenadine, hydroxyzine, ketotifen, loratadine, levocabastine, mizolastine, mequitazine, mianserin, noberastine, meclizine, norastemizole, olopatadine, picumast, pynlamine, promethazine, terfenadine, tripelennamine, temelastme, trimeprazine and tπprolidine, particularly cetirizine, levocetirizine, efletinzine and fexofenadine Another combination of interest is a combination comprising a compound of formula (I), or a pharmaceutically acceptable salt thereof together with an H3 antagonist (and/or inverse agonist) Suitable H3 antagonists include, for example, those compounds disclosed in WO2004/035556 and in WO2006/045416 Other histamine receptor antagonists which may be used in combination with the compounds of the present invention include antagonists (and/or inverse agonists) of the H4 receptor, for example, the compounds disclosed in Jablonowski et al , J Med Chem 46 3957-3960 (2003) In a further embodiment, the invention provides a combination comprising a compound of formula (I) or a pharmaceutically acceptable salt thereof together with a dual H1/H3 antagonist Examples of dual H1/H3 antagonists include 4-[(4-chlorophenyl)methyl]-2- ({(2R)-1 -[4-(4-{[3-(hexahydro-1 /-/-azepιn-1 -yl)propyl]oxy}phenyl)butyl]-2- pyrrolιdιnyl}methyl)-1(2/-/)-phthalazιnone or a pharmaceutically acceptable salt thereof as described in priority application GB0607839 8 The invention thus provides, in a further aspect, a combination comprising a compound of the invention together with a PDE4 inhibitor.
The invention thus provides, in a further aspect, a combination comprising a compound of the invention together with a β2-adrenoreceptor agonist.
The invention thus provides, in a further aspect, a combination comprising a compound of the invention together with a corticosteroid.
The invention thus provides, in a further aspect, a combination comprising a compound of the invention together with another non-steroidal GR agonist.
The invention thus provides, in a further aspect, a combination comprising a compound of the invention together with an anticholinergic.
The invention thus provides, in a further aspect, a combination comprising a compound of the invention together with an antihistamine.
The invention thus provides, in a further aspect, a combination comprising a compound of the invention together with an H1 antagonist.
The invention thus provides, in a further aspect, a combination comprising a compound of the invention together with an H1/H3 antagonist.
The invention thus provides, in a further aspect, a combination comprising a compound of the invention together with a PDE4 inhibitor and a β2-adrenoreceptor agonist.
The invention thus provides, in a further aspect, a combination comprising a compound of the invention together with an anticholinergic and a PDE-4 inhibitor.
The combinations referred to above may conveniently be presented for use in the form of a pharmaceutical formulation and thus pharmaceutical formulations comprising a combination as defined above together with a pharmaceutically acceptable diluent or carrier represent a further aspect of the invention. The individual compounds of such combinations may be administered either sequentially or simultaneously in separate or combined pharmaceutical formulations In one embodiment, the individual compounds may be administered simultaneously in a combined pharmaceutical formulation Appropriate doses of known therapeutic agents will be readily appreciated by those skilled in the art
The invention thus provides, in a further aspect, a pharmaceutical composition comprising a combination of a compound of the invention together with another therapeutically active agent
The invention thus provides, in a further aspect, a pharmaceutical composition comprising a combination of a compound of the invention together with a PDE4 inhibitor
The invention thus provides, in a further aspect, a pharmaceutical composition comprising a combination of a compound of the invention together with a β2-adrenoreceptor agonist
The invention thus provides, in a further aspect, a pharmaceutical composition comprising a combination of a compound of the invention together with a corticosteroid
The invention thus provides, in a further aspect, a pharmaceutical composition comprising a combination of a compound of the invention together with another non-steroidal GR agonist
The invention thus provides, in a further aspect, a pharmaceutical composition comprising a combination of a compound of the invention together with an anticholinergic
The invention thus provides, in a further aspect, a pharmaceutical composition comprising a combination of a compound of the invention together with an antihistamine
The invention thus provides, in a further aspect, a pharmaceutical composition comprising a combination of a compound of the invention together with an H1 antagonist
The invention thus provides, in a further aspect, a pharmaceutical composition comprising a combination of a compound of the invention together with an H1/H3 antagonist The invention thus provides, in a further aspect, a pharmaceutical composition comprising a combination of a compound of the invention together with a PDE4 inhibitor and a β2- adrenoreceptor agonist.
The invention thus provides, in a further aspect, a pharmaceutical composition comprising a combination of a compound of the invention together with an anticholinergic and a PDE- 4 inhibitor.
A process according to the invention for the preparation of compounds of formula (I) comprises reaction of an amine of formula (II)
Figure imgf000038_0001
wherein X and n are as defined above for compounds of formula (I) and R4 is hydrogen or a protecting group such as 1 ,1-dimethylethyl or terf-butyldimethylsilyl, with a compound of formula
Figure imgf000038_0002
wherein R1, R2 and R3 are as defined above for compounds of formula (I) and Y is chlorine or hydroxyl.
When Y is chlorine, the reaction may be carried out in a conventional organic solvent, for example tetrahydrofuran, in the presence of a base, for example potassium carbonate, triethylamine, pyridine or diisopropylethylamine. In one embodiment, the reaction is carried out in the presence of diisopropylethylamine. The reaction may be carried out at a temperature of from -10°C to 100°C, for example at room temperature.
Alternatively, when Y is hydroxyl, the reaction may be carried out in a conventional organic solvent, for example dimethylformamide, in the presence of a coupling agent such as those described in Tetrahedron 2005, 61 , 10827, for example O-(7-azabenzotriazol-1- yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (HATU)1 and a base, for example triethylamine or diisopropylethylamine. In one embodiment, the reaction is carried out in the presence of diisopropylethylamine. The reaction may be carried out at a temperature of from -10°C to 100°C, for example at room temperature.
When R4 is a protecting group, the compound of formula (I) is obtained by deprotection of the protected intermediate. For example, when R4 is 1 ,1-dimethylethyl or tert- butyldimethylsilyl, the group may be removed by dissolving the protected intermediate in an organic solvent, for example dichloromethane, and treating with an organic acid, for example trifluoroacetic acid. The reaction may be carried out at a temperature of from - 10°C to 100°C, for example at room temperature. Alternatively, the silyl protecting group may be removed with a fluoride source such as tetrabutylammonium fluoride in tetrahydrofuran. The reaction may be carried out at a temperature of from -10°C to 1000C, for example at room temperature.
Compounds of formula (II) may be prepared by reaction of a compound of formula (IV)
Figure imgf000039_0001
wherein X and n are as defined above for compounds of formula (I), with 2-hydroxyethylamine, or a protected derivative thereof such as 2-t-butoxyethylamine or 2-tert-butyldimethylsιlyloxyethylamine. The reaction may be carried out in a conventional organic solvent, for example acetonitrile or tetrahydrofuran, and at a temperature of from -100C to 100°C, for example at room temperature. A compound of formula (IV) may be prepared by treating a compound of formula (V)
Figure imgf000040_0001
(V) wherein X and n are as defined above for compounds of formula (I), with a polymer supported carbonate resin. The reaction may be carried out in a conventional organic solvent, for example tetrahydrofuran. The reaction may be carried out at a temperature of from -10°C to 100°C, for example at room temperature.
A compound of formula (V) may be prepared by treating a compound of formula (Vl)
Figure imgf000040_0002
(Vl) wherein X and n are as defined above for compounds of formula (I), with 4-methylbenzenesulphonyl chloride. The reaction may be carried out in a conventional organic solvent, for example dichloromethane, in the presence of an organic base, for example pyridine. The reaction may be carried out at a temperature of from - 10°C to 1000C, for example at room temperature.
A compound of formula (Vl) may be prepared by reacting a compound of formula (VII)
Figure imgf000040_0003
(VII) wherein X and n are as defined above for compounds of formula (I), with a compound of formula (VIII)
Figure imgf000041_0001
(VIII)
The reaction may be carried out in a conventional organic solvent, for example dimethylformamide, in the presence of a coupling agent such as those described in Tetrahedron 2005, 61 , 10827, for example O-(7-azabenzotriazol-1-yl)-N,N,N',N'- tetramethyluronium hexafluorophosphate (HATU), and a base, for example triethylamine or diisopropylethylamine. In one embodiment, the reaction is carried out in the presence of diisopropylethylamine. The reaction may be carried out at a temperature of from -100C to 100°C, for example at room temperature.
Example of acids of formula (VII) suitable for use in this coupling reaction include:
5-amino-1-(2-fluorophenyl)-1AV-pyrazole-4-carboxylic acid;
5-amino-1-(3-fluorophenyl)-1/-/-pyrazole-4-carboxylic acid;
5-amino-1-(4-fluorophenyl)-1/-/-pyrazole-4-carboxylic acid; 5-amino-1-(4-chlorophenyl)-1/7-pyrazole-4-carboxylic acid;
5-amino-1-(3-chlorophenyl)-1/-/-pyrazole-4-carboxylic acid;
5-amino-1 -(2-chlorophenyl)-1 H-pyrazole-4-carboxylic acid;
5-amino-1-(2,3-difluorophenyl)-1/-/-pyrazole-4-carboxylic acid;
5-amino-1-(2,4-difluorophenyl)-1/-/-pyrazole-4-carboxylic acid; 5-amino-1 -(2,5-difluorophenyl)-1 /-/-pyrazole-4-carboxylic acid;
5-amino-1-(2,6-difluorophenyl)-1H-pyrazole-4-carboxylic acid;
5-amino-1 -(3,4-difluorophenyl)-1 /-/-pyrazole-4-carboxylic acid;
5-amino-1 -(3, 5-difluorophenyl)-1 /-/-pyrazole-4-carboxylic acid; and
5-amino-1 -phenyl-1 /-/-pyrazole-4-carboxylic acid.
Acids of formula (VII) may be prepared by, for example, reaction of a suitable aryl hydrazine with ethyl 2-cyano-3-ethoxyacrylate followed by conversion of the resulting ethyl ester to the corresponding acid by treatment with, for example, lithium hydroxide in a solvent such as aqueous ethanol.
A compound of formula (VIII) may be prepared by treating a compound of formula (IX)
Figure imgf000042_0001
(IX) with a transition metal catalyst, for example palladium hydroxide on carbon, in the presence of a hydrogen atmosphere. The reaction may be carried out in a conventional organic solvent, for example ethanol. The reaction may be carried out at a temperature of from -1O0C to 100°C, for example at room temperature.
A compound of formula (IX) may be prepared by treating a compound of formula (X)
Figure imgf000042_0002
with benzylamine followed by treatment with a base, for example sodium hydroxide. The reaction may be carried out in a conventional organic solvent, for example 1 ,4-dioxan. The treatment with benzylamine may be carried out at a temperature of from -10°C to 1000C1 for example at room temperature, and the treatment with base may be carried out at a temperature of from -100C to 1000C, for example at about 9O0C.
A compound of formula (X) may be prepared by treating a compound of formula (Xl)
Figure imgf000042_0003
(Xl) with a polymer supported carbonate resin. The reaction may be carried out in a conventional organic solvent, for example dichloromethane. Batch processes or flow processes are suitable equipment for this cyclisation. The reaction may be carried out at a temperature of from -10°C to 100°C, for example at room temperature for a batch process or at about 500C for a flow process.
A compound of formula (Xl) may be prepared by treating a compound of formula (XII)
HO OH
OH
F3C
(XII) with 4-methylbenzenesulphonyl chloride in the presence of an organic base, for example pyridine. The reaction may be carried out at a temperature of from -1O0C to 100°C, for example at room temperature. Alternatively, when a flow process is used, the compound of formula (XII) may be treated with 4-methylsulphonyl chloride in the presence of an organic base, for example N,N,N',N'-tetramethyl-1 ,6-hexanediamine, in dichloromethane at room temperature.
A compound of formula (XII) may be prepared by treating a compound of formula (XIII)
Figure imgf000043_0001
(XIII) with a transition metal catalyst, for example 5% palladium on carbon, in the presence of a hydrogen atmosphere. The reaction may be carried out in a conventional organic solvent, for example ethanol. The reaction may be carried out at a temperature of from -10°C to 1000C, for example at room temperature for a batch process or at about 80°C for a flow process. Batch processes or flow processes are suitable for this hydrogenation.
A compound of formula (XIII) may be prepared by treating a compound of formula (XIV)
Figure imgf000043_0002
(XIV) with trimethyl(trifluoromethyl)silane and tetra-n-butylammonium fluoride. The reaction may be carried out in a conventional organic solvent, for example tetrahydrofuran or dichloromethane. The reaction may be carried out at a temperature of from -10°C to 1000C1 for example at 00C rising to room temperature. Batch processes or flow processes are suitable for this transformation.
A compound of formula (XIV) may be prepared by oxidation of 1 ,3-dibenzylglycerol. In one embodiment, the oxidation may be carried out using 3A molecular sieves, N- methylmorpholine Λ/-oxide and tetrapropylammonium perruthenate in dichloromethane at
O0C to reflux, for example at room temperature. In another embodiment, the oxidation may be carried out using aqueous sodium hypochlorite, saturated sodium bicarbonate solution and 2,2,6,6-tetramethyl-i-piperidinyloxy free radical in toluene at 00C to 50°C, for example at room temperature. In a further embodiment, the oxidation may be carried out using sulphur trioxide-pyridine complex in the presence of base such as triethylamine in dimethylsulphoxide at 100C to 50°C, for example at room temperature. Batch processes or flow processes are suitable for this oxidation.
Certain compounds of formulae (II), (III), (IV), (V), (Vl)1 (VIII), (IX), (X), (Xl) and (XIII) may be new and form an aspect of the present invention.
Compounds of formula (I) may be prepared in the form of mixtures of enantiomers when mixtures of isomers are used as intermediates in the synthesis. For example, the use of a compound of formula (II) as a racemic mixture of enantiomers will lead to a mixture of enantiomers in the final product. These isomers may, if desired, be separated by conventional methods (e.g. HPLC on a chiral column).
Alternatively, separation of isomers may be performed earlier in the synthesis, for example individual isomers of compounds of formula (II) or earlier stage intermediates may be employed which may obviate the need to perform a separation of isomers as a final stage in the synthesis. The later process is, in theory, more efficient and is therefore preferred.
A chiral method of preparing a compound of formula (I) is shown in Scheme 1:
OH
1,3-dιbθnzylglycθral
Figure imgf000045_0001
OB 6n
Pd-C
Figure imgf000045_0002
ι) PS-CO,. THF il) PhCHjNH2
Figure imgf000045_0003
(V)
PS-CO, THF
Figure imgf000045_0004
Scheme 1
Abbreviations
Figure imgf000045_0005
Figure imgf000046_0001
In addition, processes for preparing formulations including one or more compounds of formula (I) form an aspect of this invention.
Compositions comprising a compound of the invention also constitute an aspect of the invention.
Solvates of compounds of formula (I) or salts thereof, which are not pharmaceutically acceptable may be useful as intermediates in the preparation of other compounds of formula (I) and salts and solvates thereof.
The invention will now be illustrated by way of the following non-limiting examples.
EXAMPLES
The following non-limiting Examples illustrate the invention:
GENERAL Abbreviations
Figure imgf000047_0001
Chromatography
Chromatographic purification was performed using pre-packed Bond Elut silica gel cartridges available commercially from Varian. The Flashmaster 2 is an automated multi user flash chromatography system which utilises disposable SPE cartridges (2g to 10Og) It provides quaternary on-line solvent mixing to enable gradient methods to be run Samples are queued using the multi functional open access software which manages flow rates, gradient profile and collection conditions The system is equipped with a Knauer variable wavelength uv detector and 2 Gilson FC204 fraction collectors enabling automated peak cutting, collection and tracking
Mass Directed Autopreparative HPLC (MDAP)
Agilent 1100 series LC/MSD hardware, using electrospray positive mode (ES +ve) running chemstation 32 purification software
Column Zorbax Eclipse XDB-C18 prep HT (dimensions 212 x 100mm, 5μm packing), 20ml/mιn solvent speed
Aqueous solvent = Water + 0 1 % TFA Organic solvent = MeCN + 0 1 % TFA
Specific gradients used
Gradient 1 (collects on uv / mass ion trigger)
1 mm 70% Water (0 1% TFA) 30% MeCN (0 1%TFA) increasing over 9 mins to 5% Water (0 1%TFA) 95% MeCN (0 1 %TFA) to elute compounds
Gradient 2 (collects on uv only)
1 mm 70% Water (0 1% TFA) 30% MeCN (0 1%TFA) increasing over 9 mins to 5% Water (0 1 %TFA) 95% MeCN (0 1 %TFA) to elute compounds
LCMS System The LCMS system used was as follows
• Column 3 3cm x 4 6mm ID, 3μm ABZ+PLUS from Supelco
• Flow Rate 3ml/mιn
• Injection Volume 5μl
• Temp RT • UV Detection Range 215 to 330nm Solvents: A: 0.1 % Formic Acid + 10mMolar Ammonium Acetate. B: 95% Acetonitrile + 0.05% Formic Acid
Gradient: Time A% B%
0.00 100 0
0.70 100 0
4.20 0 100
5.30 0 100
5.50 100 0
Chiral HPLC
(1) Analytical Conditions - Method A - ChiralCel OD Column Dimensions 4.6 x 250mm Particle Size 10 μm prepacked by Chiral Technologies (Europe) a subsidiary of Diacel
Chemical Industries Ltd.
(2) Semi Preparative Conditions - Method B - ChiralCel OD Column Dimensions - 20 x 250mm Particle Size - 10 μm prepacked by Chiral Technologies (Europe) a subsidiary of Diacel Chemical Industries Ltd.
(3) Preparative Conditions - Method C - ChiralCel OD Column Dimensions 5 x 25cm packed with silica supplied by Diacel Chemical Industries Ltd.
NMR
1H NMR spectra were recorded in DMSO-d6 or chloroform-d or MeOD on a Bruker DPX 400, a Bruker AV 400 working at 400MHz. The internal standard used was either tetramethylsilane or the residual protonated solvent at 2.50 ppm for DMSO-d6 or at 7.27 ppm for chloroform-d or at 3.35 ppm for MeOD.
Circular Dichroism
Circular dichroism was carried out on an Applied Photophysics Chirascan spectrophotometer at room temperature, using acetonitrile as solvent, over the range 200-
350nm.
Intermediate 1 : 1 ,3-Bisr(benzv0oxyl-2-propanone
Figure imgf000049_0001
3A Molecular sieve powder (5Og) was dried at 1000C in a vacuum oven The sieves and N-methylmorpholine N-oxide (35 1g, 300mmol) were suspended in dry dichloromethane (700ml) before 1 ,3-dιbenzyloxy-2-propanol (41 ml, 165mmol) in dichloromethane (100ml) was added to the stirred suspension The mixture was stirred under an atmosphere of nitrogen for 90 minutes before tetrapropylammonium perruthenate (3g, 8 53mmol) was added (The reaction was sufficiently exothermic to cause the dichloromethane to boil and therefore a reflux condenser was fitted ) The reaction was stirred at 210C for 23 hours before being filtered through celite It was then washed with 2M hydrochloric acid (400ml) and saturated brine (500ml) The combined aqueous washings were filtered through celite and re-extracted with dichloromethane (500ml) and then this was washed with saturated brine (200ml) The organic extracts were combined, dried over magnesium sulphate and concentrated under reduced pressure to give a dark oil (43 6g) Diethyl ether (ca 200ml) was added and the resultant black solid was filtered off The filtrate was concentrated under reduced pressure to give the title compound (42g) as a grey white solid 1H NMR (400 MHz, CHLOROFORM-d) δ ppm 7 31 - 7 40 (m, 10H) 4 59 (s, 4H) 4 26 (s, 4H) LC-MS Retention Time 3 27mιns, MNH4 + 288
Alternative Preparation A of Intermediate 1
A mixture of sodium hypochlorite (100ml, 13%w/v) and saturated sodium bicarbonate (25ml) was added in one charge to a stirred solution of 1 ,3-dιbenzyloxy-2-propanol (10g), 2,2,6,6-tetramethyl-i-pιperιdιnyloxy, free radical, (TEMPO) (0 3g) in toluene (40ml) The biphasic mixture was stirred at 20-25°C for 15 minutes when HPLC analysis showed reaction to be complete The reaction mixture was stirred for a total of 25 minutes at 23°C The reaction mixture was separated and the organic extract was washed with 5% w/v sodium thiosulfate solution (40ml) and separated The organic extract was washed with 1 % w/v sodium chloride solution (2 x 25ml) The organic extract was then concentrated in vacuo to give an oil which crystallised on standing to give 8 8g of 1 ,3-dιbenzyloxy-2- propanone in 88 7% yield The NMR spectrum of the product was concordant with a reference sample
Alternative Preparation B of Intermediate 1
A mixture of sulfur trioxide/pyridine complex (2 33g, 4 equivalents) and triethylamine (2 05ml, 4 equivalents) in DMSO (3ml) was stirred to give a pale yellow solution To this was added a solution of 1 ,3-dibenzyloxy-2-propanol (1g) in DMSO (1 ml) over 2 minutes. (The reaction mixture was kept in a water bath). The temperature of the reaction mixture reached 30°C. After 10 minutes the water bath was removed and the reaction mixture was stirred at room temperature (ca 20 - 25°C) for 3 hours. The reaction mixture was diluted with ethyl acetate (15ml) and water (15ml), stirred and the organic extract was separated. The organic extract was washed with 5% w/v sodium chloride (2 x 10ml) and water (10ml). The separated organic extract was concentrated in vacuo to give an oil which solidified to provide 0.75g of 1 ,3-dibenzyloxy-2-propanone in 75.8% yield. An NMR spectrum of product was concordant with a reference sample.
Alternative Preparation C of Intermediate 1
The title compound was prepared via a 'flow' process using the following starting materials and solvents.
The title compound was prepared via a CPC Cytos Lab System made up of a 47ml reactor block with two Jasco PU - 2080Plus HPLC pumps. Reactor temperature was maintained at 600C via a Huber Unistat 360 unit.
Two solutions were prepared. Solution A - 1 ,3-dibenzyloxy-2-propanol 12Og, 440mmol) in acetonitrile (489ml). Solution B - tetrapropylammonium perruthenate (7.72g, 22mmol, 5mol%) and N-methylmorpholine N-oxide (87.5g, 748mmol) in acetonitrile (611ml). Solutions A and B were pumped through the Cytos Lab system in the ratio of solution A to solution B of 1 : 1.25 with a total flow rate of 7.8ml/min and residence time of 6 minutes. This gave a total reaction time of 2 hours 21 minutes. The total reacted solution was split equally into 2 batches and each was concentrated in vacuo. Diethyl ether (250ml) was added before being washed with sodium sulphite, brine, cupric sulphate then filtered through celite, dried and evaporated. The batches were recombined to give upon evaporation in vacuo the title compound (71.64g).
Alternative Preparation D of Intermediate 1
A solution of 1 , 3-dibenzyloxy-2-propanol (50Og, 1.84mol, 1.0eq.), TEMPO (5.5g, 0.034mol) in dichloromethane (1.25L) was placed in a 10L flange flask fitted with overhead stirrer. A solution of potassium bromide (48g, 0.40mol) in water (185ml) was added and the reaction stirred and cooled to -10°C. A 14% aqueous NaOCI solution was diluted to 1 M (2145g diluted to 4050ml). The pH of this solution was then adjusted to 9.5 by dissolving NaHCO3 (8Og) immediately before use. This NaOCI solution was added over 1 hour, keeping the temperature of the reaction mixture between 10-150C. The mixture was the stirred for 60 minutes. The orange coloured organic phase was separated and the aqueous layer was extracted with dichloromethane (5.0L, 2.0L). The combined organics were washed with 10% aq. HCI (10.75L) containing potassium iodide (143g), 10% aq. Na2S2O5 (5.0L) and water (5.0L). The organics were dried over MgSO4 and concentrated under reduced pressure to give the crude title compound (893g, 90%). This compound was taken through to the next step without further purification.
Intermediate 2: 1.1 ■1-Trifluoro-3-r(benzvltoxvl-2-(r(benzvl)oxvlmethyl}-2-propanol
Figure imgf000052_0001
To a solution of 1 ,3-bis[(benzyl)oxy]-2-propanone (42g, 155mmol) in anhydrous tetrahydrofuran (600ml) was added trimethyl(trifluoromethyl)silane (35ml, 236mmol) under nitrogen. The mixture was then cooled in an ice/ethanol bath to -30C before tetrabutylammonium fluoride (1 M in THF, 180ml, 180mmol) was added dropwise (initial 10ml of addition resulted in a slight exotherm with the temperature rising to 90C before being allowed to cool to 60C and then the addition was resumed, the temperature dropping to the range of -10C to +30C). The addition was completed after 30 minutes. The mixture was stirred for a further 4 hours during which, gas was evolved all the time and then 2M hydrochloric acid (750ml) was added with stirring. Diethyl ether (600ml) was added and the separated aqueous phase was reextracted with diethyl ether (1 x 600ml, 1 x 300ml) and the combined organic extracts were washed with saturated brine (1 x 300ml), dried over sodium sulphate and concentrated under reduced pressure to give an oil (52.9g). This oil was purified via flash chromatography (Silica, 80Og) using cyclohexane:ethyl acetate (9:1) as eluent. This gave the title compound as a yellow oil (39.5g).
1H NMR (400 MHz, CHLOROFORM-d) δ ppm 7.29 - 7.39 (m, 10H) 4.60 (s, 4H) 3.72 (s, 4H) 3.38 (s, 1H). LC-MS Retention Time 3.69 min, MNH4 + 358.
Alternative Preparation A of Intermediate 2
A mixture of 1,3-dibenzyloxy-2-propanone (2g) and (trifluoromethyl)trimethylsilane
(2.56ml, 2.3 equivalents) in dichloromethane (20ml) was stirred and cooled to O0C. A solution of 1 M tetrabutylammonium fluoride in THF (4ml) was added drop wise over 3 minutes. Initial addition of a few drops gave an exotherm of 1O0C. Throughout the addition the batch temperature was maintained below 10°C. After completing the addition the dark brown mixture was stirred at +50C for 5 minutes when HPLC analysis indicated the reaction to be complete. The reaction mixture was stirred for an additional 5 minutes and then washed with 1M aqueous hydrochloric acid (2 x 15ml), saturated sodium bicarbonate (15ml) and 1 %w/v aqueous sodium chloride solution (2 x 15ml). The organic extract was concentrated in vacuo to give 2.5g of the desired product as dark oil in 99.3% yield. The NMR spectrum of the product was concordant with a reference sample.
Alternative Preparation B of Intermediate 2
Tetrabutylammonium fluoride trihydrate (TBAF 3H2O) (2.9g, 0.5 equivalent) was dissolved in THF (5ml). This was added cautiously to a stirred and cooled (+150C) solution of 1 ,3- dibenzyloxy-2-propanone in toluene (24.65g, equivalent to 5g of the ketone) and (trifluoromethyl)trimethylsilane (7.5ml). There was an exotherm and lot of gas evolution on addition of the first 1ml of TBAF solution. The temperature rose from 18 to 4O0C. The TBAF addition was carried out over 3 minutes and then the mixture was stirred at 15-3O0C for a further 2 minutes and then cooled to +100C while carrying out an HPLC analysis. The reaction mixture was sequentially washed with 1 M aqueous hydrochloric acid (50ml), 1% aqueous sodium chloride solution (2 x 25ml) and a mixture of 1 % sodium chloride (25ml) and saturated sodium bicarbonate (5ml) solution. The separated organic extract was concentrated in vacuo to give 6.41 g of the desired product as dark brown oil in 101.8% yield. The NMR spectrum showed the presence of residual toluene (8.8%) and starting material (ca 3%).
Alternative Preparation C of Intermediate 2
The title compound was prepared via a 'flow' process using the following starting materials and solvents.
The title compound was prepared via a CPC Cytos Lab System made up of a 32ml reactor block with two Jasco PU - 2080Plus HPLC pumps. Reactor temperature was maintained at 22°C via a Huber Unistat 360 unit. The reactor outlet was fitted with a IOOpsi backflow regulator. Two solutions were prepared. Solution A - 1 ,3-bis[(phenylmethyl)oxy]-2-propanone (71.64g, 265mmol) and trimethyl(trifluoromethyl)silane (86.67g, 96ml, 609.5mmol) in tetrahydrofuran(99ml). Solution B - tetrabutylammonium fluoride (1 M in THF, 265ml, 132.5mmol).
Solutions A and B were pumped through the Cytos Lab System with a flow rate of 6.4ml/min and a 5 minute residence time giving a total reaction time of 82 minutes. The reaction mixture was quenched with 2M hydrochloric acid (560ml) and then divided into 2 equal batches (2 x 280ml). Diethyl ether (100ml) was added to each batch, extracted and then washed with brine (2 x 100ml), dried (MgSO4) and evaporated to give a residue (82.99g). Part of the residue was taken up in dichloromethane and applied to SPE silica cartridges. Using 10% hexane in dichloromethane as eluent and concentration of the relevant 15ml fractions, the title compound was obtained. The bulk of the crude sample was purified on the Combiflash Companion XL. 8g of material was run on a 120gmcolumn with a solvent gradient of 10%-70% dichloromethane in hexane as eluent. Any mixed fractions from each run were combined and repurified in an identical manner. All pure fractions were combined and evaporated to give the title compound (68.68g).
Alternative Preparation D of Intermediate 2
A solution of 1 ,3-dibenzyloxy-2-propanone (31 Og, 1.15mol, 1.0eq.) was placed in a 10L flange flask, equipped with magnetic stirrer, condenser under argon, followed by THF (3.5L). This was stirred at 15°C. Trimethyl(trifluoromethyl)silane (TMS-CF3) (Matrix, 231 g, 1.62mol, 1.41eq.) was added dropwise over 1 hour. The solution was then cooled to 0°C using an ice-water bath and tetrabutylammonium fluoride (TBAF) (337g, 1.29L, 1 M in THF, 1.29mol, 1.12eq.) was added dropwise keeping the temperature in the range 0-80C (initial sharp delayed exotherms). After the addition was complete, the temperature was raised cautiously to 20°C and the reaction stirred at 27°C overnight (oil bath). (TLC Rf starting material=0.3, product=0.4, 80:20 petroleum ether/ethyl acetate showed reaction was essentially complete). Reaction mixture was then cooled to 15°C using an ice-water bath, then quenched by slow addition of 1 M HCI (10.5L) (transfer to a 2OL separator after 1.0L added). The mixture was extracted with diethyl ether (3x 5.0L). The combined organics were washed with water (2.5L) and brine (2.5L). The organics were dried over MgSO4 and concentrated under reduced pressure to yield a brown oil (388g with THF). Intermediate 3: 2-(Trifluoromethyl)-1.2.3-propanetriol
F3C OH
A solution of 1 , 1 ,1-trifluoro-3-[(benzyl)oxy]-2-{[(benzyl)oxy]methyl}-2-propanol (98.9g, 290.9mmol) in ethanol (1750ml) was added to 5% palladium on carbon (9.73g, wet, Degussa, E101 No/W) under nitrogen. The mixture was then stirred under an atmosphere of hydrogen using a Wright valve in a 5 litre hydrogenation vessel. After approximately 3 hours most of the theoretical volume of hydrogen had been taken up. After stirring under hydrogen overnight (approximately a further 1 litre of hydrogen had been taken up overnight), the catalyst was filtered off through a pad of celite and the pad washed with ethanol. The filtrate and washings were then concentrated under reduced pressure and the residue azeotroped (x2) with dichloromethane whereupon the residue became solid. This material was left on the vacuum pump overnight to give the title compound (48.56g) as an off white solid. 1H NMR (400 MHz, DMSO-Cf6) δ ppm 5.65 (s, 1 H) 4.89 (t, 2H) 3.54 (d, J=5.8 Hz, 4H). LC-MS Retention Time 0.42mins, ES" MH" 159.
Alternative Preparation A of Intermediate 3
The title compound was prepared employing the Thales H-Cube hydrogenator and milligat pump in full hydrogen mode. A solution of 1 ,1 ,1-trifluoro-3-[(phenylmethyl)oxy]-2-
{[(phenylmethyl)oxy] methyl}-2-propanol (58g) in ethanol (580ml) was prepared. The flow rate was 1.3ml/min, the temperature was set to 80°C and the cartridge employed was a
10%Pd/C Cat Cart 70 which was replaced every 2 hours. Any fractions which still contained starting material and the mono benzyl intermediate were reprocessed. All pure fractions were combined and evaporated to give the title compound (26.48g).
Alternative Preparation B of Intermediate 3
1 ,1 ,1-Trifluoro-3-[(benzyl)oxy]-2-{[(benzyl)oxy]methyl}-2-propanol (1.3kg, 3.82mol, 1.0eq.) was placed in a 1OL flange flask equipped with a overhead stirrer, followed by ethanol
(4.5L). 10% palladium on carbon (27g) was added under Argon atmosphere. The reaction was then subjected to hydrogenolysis at atmospheric pressure (6 balloons) and stirred overnight at 50°C (the balloons were topped up repeatably during the day). 1H
NMR showed completion of reaction had been reached after 1 week, required additional 10% palladium on carbon (4g). The reaction mixture was filtered through a pad of celite and washed with ethanol (2.5L). The filtrate was concentrated under reduced pressure to give an oil. This was placed under high vacuum overnight to obtain a solid material. Toluene (1.5L) was added and the mixture heated until the solid dissolved (-6O0C), 2 layers were observed. The mixture was stirred using a magnetic stirrer and cooled using an ice-water bath, where a solid precipitated. The solid was broken up and stirred for a further 30 minutes, then isolated by filtration. The solid was washed with toluene (250ml) and petroleum ether (250ml). The solid was dried under high vacuum overnight to yield desired product (465g).
Intermediate 4j 3,3,3-Trifluoro-2-hvdroxy-2-αr(4-methylphenvn sulfonvπoxylmethyltoropyl 4-methylbenzenesulfonate
Figure imgf000056_0001
To a stirred solution of 2-(trifluoromethyl)-1 ,2,3-propanetriol (18.9g, 118mmo!) in pyridine (200ml) which had been cooled in an ice bath was added p-toluenesulphonyl chloride
(67g, 351mmol) to give an orange solution. The ice bath was removed after 45 minutes and stirring was continued for 21 hours during which time a solid formed. Most of the pyridine was removed under reduced pressure and the residue was partitioned between ethyl acetate (500ml) and water (300ml). The separated aqueous phase was further extracted with ethyl acetate (1 x 250ml) and the combined organic extracts were washed with 2M hydrochloric acid (1 x 200ml), water (1 x 200ml), saturated sodium bicarbonate (1 x 200ml), water (1 x 200ml) and saturated brine (1 x 200ml) before being dried over sodium sulphate and concentrated under reduced pressure to give an oil (72.8g). This oil was purified on a Flash silica column (80Og) eluting with cyclohexane.ethyl acetate (5:1 ) to give the title product (49g, 95%) as an oil which crystallised on standing.
1H NMR (400 MHz, CHLOROFORM-d) δ ppm 7.78 (d, J=8.3 Hz, 4H) 7.38 (d, J=8.3 Hz, 4H) 4.18 (s, 4H) 3.66 (s, 1 H) 2.48 (s, 6H). LC-MS Retention Time, 3.62mins, MNH4 + 486.
Alternative Preparation A of Intermediate 4
The title compound was prepared via a 'flow' process using the following starting materials and solvents. Two solutions were prepared. Solution A - 2-(trifluoromethyl)-1 ,2)3-propanetriol (4.5gm,27.8mmol), N,N,N\N'-tetramethyl-1 ,6-hexanediamine (30ml, 139mmol), dichloromethane (550ml). Solution B - p-toluenesulphonyl chloride (21.4g, 111mmol), dichloromethane (550ml).
Solutions A and B were pumped through a CPC Cytos reactor (reactor volume 47ml) at a flow rate each of 2.35ml/min. It was noted that the pressure for the pump containing solution B was fluctuating. After 110 minutes, the reaction was abandoned as it was evident that the pumps were not operating 1 : 1. The collected material was extracted with dichloromethane (x3) before being washed with brine, dried (MgSO4), filtered and concentrated to give a residue which was discarded. The pump was replaced and the remainder of the reagents were reacted. The collected material was extracted with dichloromethane (x3) before being washed with brine, dried (MgSO4), filtered and concentrated to give a residue. It was adsorbed onto silica and eluted over a silica column (12gm) with dichloromethane : hexane (1 : 1 ). Four fractions were eluted and fraction 4 gave the title compound (2.31g).
Alternative Preparation B of Intermediate 4
2-(Trifluoromethyl)-1 ,2,3-propanetriol ( 30Og, 1.86mol, 1.0eq.) was dissolved in pyridine (2.0L) in a 10L flange flask fitted with an overhead stirrer, thermoprobe under argon. This was cooled to 00C with a solid CO2-acetone bath. Tosyl chloride (809g, 4.1 mol, 2.2eq) was added portionwise, maintaining temperature <10°C. On completion of addition, the cold bath was removed and reaction mixture allowed to stir at room temperature overnight. The reaction mixture was filtered and concentrated under reduced pressure. The resulting brown oil was partitioned between EtOAc (4.0L) and 2M HCI (4.0L), stirred for 5 minutes and separated. The aqueous layer was further extracted with EtOAc (2 x 2.5L). The combined organics were washed with saturated NaHCO3 (3.5L, 5mins stir period), brine (2.5L) and dried over MgSO4. This was concentrated under reduced pressure to give a thick brown oil. The crude material was taken to the next step without further purification (95Og, overweight, assume 100%).
Intermediate 5: f2-(Trifluoromethyl)-2-oxiranyl1methyl 4-methylbenzenesulfonate
Figure imgf000057_0001
A solution of the bis tosylate, 3,3,3-trifluoro-2-hydroxy-2-({[(4-methylphenyl) sulfony|]oxy}methy|)propyl 4-methylbenzenesulfonate (186.5g, 398.5mmol) in dichloromethane (2500ml) was stirred under nitrogen whilst polymer supported carbonate resin (ex Fluka, ca. 3.5mmoles carbonate/g resin) (232g) was added. The mixture was stirred at room temperature overnight. The resin was filtered off and the resin was washed with dichloromethane. The combined filtrate and washings were concentrated under reduced pressure to give the title compound (116.2g) as a brown oil. 1H NMR (400 MHz, CHLOROFORM-c/) δ ppm 7.80 (d, J=8.3 Hz1 2H) 7.38 (d, J=8.0 Hz, 2H) 4.41 (d, J=11.9 Hz, 1 H) 4.29 (d, J=11.9 Hz, 1 H) 3.14 (d, J=4.8 Hz, 1 H) 3.01 (dd, J=4.5, 1.5 Hz, 1 H) 2.47 (s, 3H).
LC-MS Retention Time 3.2mins, MNH4 + 314.
Alternative Preparation A of Intermediate 5
3,3,3-Trifluoro-2-hydroxy-2-({[(4-methylphenyl) sulfonyl]oxy}methyl)propyl 4- methylbenzenesulfonate (14.29g, 29mmol) in dichloromethane (75ml) was pumped through a cartridge containing PS-carbonate resin (not pre-swelled) (3mmol/g, 25g, 75mmol) at 675 microlitres/min. The temperature was set to approx. 500C by wrapping a Whatman thin film heater around the cartridge. The pressure was regulated at 40psi. After all the reagent had been aspirated, the column was washed through with dichloromethane - at this point the column started to leak slightly and the back pressure regulator had to be removed to reduce the pressure. The collected solution was concentrated in vacuo to afford the title compound (7.25g).
Alternative Preparation B of Intermediate 5
Bis tosylate, 3,3,3-trifluoro-2-hydroxy-2-({[(4-methylphenyl) sulfonyl]oxy}methyl)propyl 4-methylbenzenesulfonate (1.047kg, 2.24mol, 1.0eq.) was dissolved in dichloromethane (9.0L) in a 2OL flange flask, fitted with overhead stirrer under argon. Potassium carbonate (1.24kg, 8.95mol, 4.0eq.) was added portionwise over 8 hours and stirred overnight at room temperature. 1H NMR showed -41 % completion of reaction. Additional potassium carbonate (60Og, 2.0eq.) was added over 8 hours and stirred overnight. 1H NMR showed 89% completion of reaction. Additional potassium carbonate (30Og, 1.0eq.) was added over 8 hours and stirred overnight. 1H NMR showed completion of reaction reached. The reaction mixture was filtered on two glass sinters with a pad of celite and washed with dichloromethane. The filtrate was washed with NaHCO3 (3.5L)1 brine (2.5L) and dried over MgSO4. The filtrate was then concentrated under reduced pressure to give a dark red oil. The crude product was purified by suction chromatography (~13cm silica on 4L sinter, collected -1.5L fractions, starting with 95:5 petroleum ether/EtOAc as eluent until all the tosyl chloride was removed, the polarity of the eluent was then increased slowly to 90:10, 80:20 and 70:30. TLC was run with 70:30 Petroleum ether/EtOAc giving Rf tosyl chloride=0.55, Product=0.5, impurity=0.3. Some mixed fractions were obtained, they were re-columned using same conditions as above. This gave clean product (417g, 63%) as an orange oil, which solidified with high vacuum overnight.
Intermediate 6: 3.3.3-Trifluoro-2-fflphenylmethyl)aminolmethyl)-1 ,2-propanediol
Figure imgf000059_0001
To a stirred solution of the [2-(trifluoromethyl)-2-oxiranyl]methyl 4-methylbenzenesulfonate (10.07g, 34mmol) in anhydrous 1 ,4-dioxan (70ml) cooled in an ice bath under nitrogen was added benzylamine (4.1 ml, 37.4 mmol) in small portions over 10 minutes. The mixture was stirred at ice bath temperature for a further hour before being allowed to warm to 210C and then stirred for 18 hours. 2M Sodium hydroxide (50ml) and 1 ,4-dioxan (50ml) were added and stirred for 2 hours at room temperature before being heated at 900C for 22 hours. The mixture was allowed to cool before being concentrated to low volume and partitioned between ethyl acetate (250ml) and water (100ml). The separated aqueous layer was further extracted with ethyl acetate (1 x 250ml) and the combined organic extracts were washed with water (1 x 100ml), saturated brine (1 x 100ml), dried over sodium sulphate and concentrated under reduced pressure to give an oil (9.4g). This was purified on 3 x 100g SPE cartridges using a 0-100% cyclohexane-ethyl acetate gradient over 60 minutes. This gave upon concentration of the relevant fractions under reduced pressure, the title compound (5.09g) as an oil.
1H NMR (400 MHz, CHLOROFORM-d) δ ppm 7.30 - 7.35 (m, 2H) 7.23 - 7.29 (m, 3H) 3.75 - 3.86 (m, 4H) 3.57 (d, J=11.6 Hz, 1 H) 3.06 (d, J=13.1 Hz, 1 H) 2.86 (d, J=13.1 Hz, 1 H), OH & NH are very broad 2.5-3.0ppm. LC-MS Retention Time 1.45mins, MH+ 250.
Intermediate 7: 2-(Aminomethyl)-3,3,3-trifluoro-1 ,2-propanediol
Figure imgf000060_0001
A solution of S.S.S-trifluoro^-t^phenylmethyOaminoJmethylJ-i ^-propanediol (8 33g, 33 4mmol) in ethanol (550ml) containing palladium hydroxide on carbon (20%, 800mg) was stirred under an atmosphere of hydrogen for 24 hours The catalyst was filtered off via a pad of celite and the filtrate was concentrated under reduced pressure, toluene was added and the solution evaporated again under reduced pressure to give the title compound (5 06g) as an oil
1H NMR (400 MHz1 MeOD) δ ppm 3 69 - 3 69 (m, 2H) 2 96 (d, J=13 5 Hz, 1 H) 2 87 (d, J=13 5 Hz, 2H) LC-MS Retention Time 0 32mιns, MH+ 160
Intermediate 8 Ethyl 5-amιno-1-(4-fluorophenyl)-1/-/-pyrazole-4-carboxylate
Figure imgf000060_0002
To a stirred suspension of 4-fluorophenylhydrazιne hydrochloride (9 76g, 60mmol) in ethanol (250ml) was added triethylamine (9 2ml, 62mmol) and to the resulting amber solution was added ethyl 2-cyano-3-ethoxyacrylate (10 15g, 60mmol) The solution was heated at reflux temperature for 3 5 hours The solution was allowed to cool to room temperature and after standing overnight the resultant solid was filtered off, washed with small amount of ethanol and then ether before being dried under vacuum to give the title compound (12 1g) as an off white solid
1H NMR (400 MHz, DMSO-d6) δ ppm 7 70 (s, 1 H) 7 55 (s, J=5 0 Hz, 2H) 7 34 - 7 41 (m, 2 H) 6 34 (br s , 2H) 4 21 (q, J=I 0, 7 0 Hz, 2H) 1 26 (t, J=I 0 Hz, 3H)
Intermediate 9 5-Amιno-1-(4-fluorophenyl)-1 /-/-pyrazole-4-carboxylιc acid
Figure imgf000060_0003
To a suspension of ethyl 5-amιno-1-(4-fluorophenyl)-1 /-/-pyrazole-4-carboxylate (12 1g, 48 5mmol) in ethanol (250ml) was added a solution of lithium hydroxide (5 8g, 242mmol) in water (100ml) The mixture was stirred at reflux for 2 5 hours It was allowed to cool and concentrated to 50% of its volume before 5M hydrochloric acid (47ml) was added. After stirring for 15 minutes, the resulting white solid was filtered off and further 5M hydrochloric acid (3ml) was added to the filtrate and the resulting solid was filtered and the combined solids were washed with water and diethyl ether and then dried under vacuum to give the title compound (10.27g).
1H NMR (400 MHz, DMSOd6) δ ppm 12.09 (br. s., 1 H) 7.67 (s, 1 H) 7.54 - 7.60 (m, 2H) 7.34 - 7.41 (m, 2H) 6.29 (br. s., 2H). LC-MS Retention Time 2.20mins, MH+ 222.
Intermediate 10: 5-Amino-1-(4-fluorophenyl)-Λ/-r3.3.3-trifluoro-2-hvdroxy-2- (hvdroxymethvπproDyll-1 /-/-pyrazole-4-carboxamide
Figure imgf000061_0001
A solution of 5-amino-1-(4-fluorophenyl)-1/-/-pyrazole-4-carboxylic acid (5.86g, 26.5mmol) in anhydrous dimethylformamide (60ml) and diisopropylethylamine (17.5ml, lOOmmol) was cooled in an ice bath for 5 mins before O-(7-azabenzotπazol-1-yl)-N,N,N',N'- tetramethyluronium hexafluorophosphate (HATU) (11.1g, 29.2mmol) was added. After 5 minutes, the solution was removed from the ice bath and stirred under nitrogen for 20 minutes. The stirred mixture was recooled in ice for 5 minutes before a solution of the amine, 2-(aminomethyl)-3,3,3-trifluoro-1 ,2-propanediol (5g, 31.4mmol) in anhydrous dimethylformamide (20ml) was added. The ice bath was again removed and stirring continued for 2.5 hours. The mixture was then partitioned between ethyl acetate (500ml) and water (500ml) and the separated aqueous phase was reextracted with ethyl acetate (300ml). The combined organic extracts were washed with water (1x 500ml, 1 x 300ml), 1 M hydrochloric acid (1 x 400ml), concentrated lithium chloride (2 x 200ml), saturated sodium hydrogen carbonate (1 x 200ml), water (200ml) and saturated brine (2 x 200ml) before being dried over sodium sulphate and concentrated under vacuum to give a foam (9.5g). Ethyl acetate (5ml) was added followed by dichloromethane (50ml) and the mixture was swirled to initiate crystallisation. It was left to stand in the fridge for 15 hours and the resultant solid was filtered off and washed with small amounts of dichloromethane and heptane before being dried under vacuum to give the title compound (7.Og, 73%) as a white solid. 1H NMR (400 MHz, DMSOd6) δ ppm 8.17 (t, ./=6.1 Hz, 1 H) 7.99 (s, I H) 7.54 - 7.60 (m, 2H) 7.33 - 7.40 (m, 2H) 6.37 (br. s., 2H) 6.30 (s, 1 H) 5.19 (t, J=6.4 Hz, 1 H) 3.51 - 3.69 (m, 2H) 3.38 - 3.50 (m, 2H). LC-MS Retention Time 2.20mins, MH+363.
Intermediate 11 : 2-IY([5-Amino-1-(4-fluorophenyl)-1 H-pyrazol-4- yllcarbonyl)amino)methyl1-3,3.3-trifluoro-2-hvdroxypropyl 4-methylbenzenesulfonate
Figure imgf000062_0001
To a stirred solution of 5-amino-1-(4-fluorophenyl)-Λ/-[3,3,3-trifluoro-2-hydroxy-2- (hydroxymethyl)propyl]-1H-pyrazole-4-carboxamide (2.47g, 6.82mmol) in anhydrous dichloromethane (20ml) and anhydrous pyridine (20ml) cooled in an ice bath under a nitrogen atmosphere was added p-toluenesulphonyl chloride (1.7g, 8.9mmol). The mixture was stirred for 6 hours at ice bath temperature before being allowed to warm to room temperature and stirred overnight. The solution was evaporated under vacuum and the residue was partitioned between ethyl acetate (100ml) and water (30ml). The separated organic phase was washed with 2M hydrochloric acid (2 x 30ml), water (30ml), saturated sodium hydrogen carbonate (30ml), water (30ml) and saturated brine (50ml) before being dried over sodium sulphate and evaporated under reduced pressure to give a foam (3.45g). This foam was purified on a Flashmaster column of Silica (100g) using a 0-100% ethyl acetate in cyclohexane gradient over 1 hour. This afforded the title compound (2.8g, 79%) as a foam.
1H NMR (400 MHz, DMSO-d6) δ ppm 8.04 (t, J=6.3 Hz, 1 H) 7.90 (s, 1 H) 7.78 (d, J=8.1 Hz, 2H) 7.55 - 7.60 (m, 2H) 7.45 (d, J=8.1 Hz, 2H) 7.34 - 7.41 (m, 2H) 3.97 - 4.06 (m, 2H) 3.63 (dd, J=UJ, 6.6 Hz, 1 H) 3.45 (dd, 1 H) 2.38 (s, 3H). LC-MS Retention Time 3.41 mins, MH+ 517.
Intermediate 12: 5-Amino-1-(4-fluorophenylVΛ/-(r2-(trifluoromethyl)-2-oxiranyl1methyl)-1 H- pyrazole-4-carboxamide
Figure imgf000063_0001
A solution 2-[({[5-amino-1 -(4-f luorophenyl)-1 H-pyrazol-4-yl]carbonyl}amino)methyl]-3,3,3- trifluoro-2-hydroxypropyl 4-methylbenzenesulfonate (2.8g, 5.4mmol) in anhydrous tetrahydrofuran (150ml) was shaken with polymer supported carbonate resin (ex Fluka, 5g, 3.5mmol/g, 17.5mmol) which had been pre washed with tetrahydrofuran (5x). After shaking for 15 hours, the resin was filtered off and the filtrate was evaporated under reduced pressure to give a semi-solid (1.986g). Diethyl ether (ca. 10ml) was added and after standing for 3 hours, the resultant crystallised solid (1g) was filtered off and washed with heptane. The filtrate and precipitate were evaporated under reduced pressure and the residue, dissolved in dichloromethane, was purified on a Flashmaster silica column (100g) eluting with 0-100% ethyl acetate in cyclohexane over 60 minutes to afford a cream solid (0.46g). This was combined with the crystallised solid to give the title compound (1.46g). 1H NMR (400 MHz, DMSO-Cf6) δ ppm 8.15 (t, J=6.1 Hz, 1 H) 7.94 (s, 1 H) 7.53 - 7.60 (m, 2H) 7.36 (t, J=8.8 Hz, 2H) 6.38 (s, 2H) 3.88 (dd, J=14.9, 6.1 Hz, 1 H) 3.69 (dd, .7=14.8, 6.0 Hz, 1 H) 3.17 (d, J=4.3 Hz, 1 H) 2.90 - 2.98 (m, J=4.0 Hz, 1 H). LC-MS Retention Time 2.83mins, MH+ 345.
Intermediate 13: 5-Amino-Λ/-(2-r((2-r(1 ,1-dimethylethvnoxy1ethyllamino)methyl1-3,3,3- trifluoro-2-hvdroxypropyl}-1-(4-fluorophenyl)-1H-pyrazole-4-carboxamide
Figure imgf000063_0002
To a solution of 5-amino-1-(4-fluorophenyl)-Λ/-{[2-(trifluoromethyl)-2-oxiranyl]methyl}-1H- pyrazole-4-carboxamide (2.065g, 6mmol) in anhydrous acetonitrile (20ml) was added t- butoxyethylamine (2.06ml, 15mmol). The mixture was stirred under nitrogen for 18.5 hours. The solvent was evaporated off and the resulting oil was triturated with water (ca. 25 ml) to give a white solid. The suspension was stirred for 20 minutes, the solid was filtered off, washed with water (ca. 10ml) and 40-60° petroleum ether (2 x 20 ml) and dried in vacuo at 400C over phosphorous pentoxide to give the title compound (2.547g). The solid melted in the drying pistol but resolidified as a glass.
1H NMR (400 MHz1 DMSO-Cf6) δ ppm 7.86 (s, 1 H) 7.45 - 7.53 (m, 2H) 7.28 - 7.36 (m, 2H)
3.43 - 3.65 (m, J=37.9 Hz1 2H) 3.30 - 3.37 (m, 2H) 2.75 (s, 2H) 2.57 - 2.68 (m, 2H) 1.08
(S1 9H).
LC-MS Retention Time 2.47mins, MH+ 462.
Intermediate 14: 5-Amino-A/-(2-rfrf2.6-diriuoroDhenvhcarbonvlU2-rM .1- dimethylethvπoxylethyl>amino)methyl1-3.3.3-trifluoro-2-hvdroxypropyl)-1 -(4-fluoroDhenvπ- 1 /-/-pyrazole-4-carboxamide
Figure imgf000064_0001
To a solution of 5-amino-Λ/-{2-[({2-[(1 ,1-dimethylethyl)oxy]ethyl}amino)methyl]-3,3,3- trifluoro-2-hydroxypropyl}-1-(4-fluorophenyl)-1/-/-pyrazole-4-carboxamide (1.84g, 4mmol) in anhydrous tetrahydrofuran (20 ml) was added diisopropylethylamine (1.44ml, δmmol). The solution was divided into two equal portions and cooled in ice. To one solution was added 2,6-difluorobenzoyl chloride (0.27ml, 2.14mmol). After 30 minutes, the reaction was removed from the ice bath and stirring was continued under nitrogen for 3 days. The mixture was added to ethyl acetate (50 ml) and the solution was washed with 2M hydrochloric acid (10ml), water (10ml), saturated sodium hydrogen carbonate (15ml), water (10ml) and saturated brine (10ml), dried over magnesium sulphate and evaporated to give the title compound (1.221g) as a foam.
1H NMR (400 MHz, DMSO-c/6) δ ppm 8.07 (t, 1 H) 7.88 (s, 1 H) 7.52 - 7.61 (m, 3H) 7.31 - 7.41 (m, 2H) 7.18 - 7.26 (m, 2H) 7.01 (s, 1 H) 6.38 (br. s., 2H) 3.36 - 4.18 (m, 8H) 1.03 (s, 9H).
LC-MS Retention Time 3.45 and 3.59mins, MH+ 602.
Intermediate 15: 5-Amino-Λ/-(2-f(rf2.6-dichlorophenyl)carbonyll(2-r(1.1- dimethylethvnoxylethyl)amino)methyll-3.3,3-trifluoro-2-hvdroxypropyl)-1-(4-fluorophenyl)- 1 /-/-Dvrazole-4-carboxamide
Figure imgf000065_0001
To a solution of 5-amino-Λ/-{2-[({2-[(1 ,1-dimethylethyl)oxy]ethyl}amino)methyl]-3,3,3- trifluoro-2-hydroxypropyl}-1-(4-fluorophenyl)-1W-pyrazole-4-carboxamide (1.84g, 4mmol) in anhydrous tetrahydrofuran (20ml) was added diisopropylethylamine (1.44ml, δmmol). The solution was divided into two equal portions and cooled in ice. To one solution was added 2,6 dichlorobenzoyl chloride (0.30ml, 2.14mmol). After 30 minutes, the reaction was removed from the ice bath and stirring continued under nitrogen for 3 days. It was added to ethyl acetate (50ml) and the solution was washed with 2M hydrochloric acid (1 x 10ml), water (10ml), saturated sodium hydrogen carbonate (15ml), water (10ml) and saturated brine (10ml) before being dried over magnesium sulphate and evaporated to give the title compound (1.32g) as a foam.
1H NMR (400 MHz, DMSO-cfe) δ ppm 8.10 (t, 1H) 7.85 (s, 1H) 7.51 - 7.63 (m, 5H) 7.34 - 7.40 (m, 2H) 7.04 (s, 1 H) 6.39 (br. s., 2H) 3.21 - 3.99 (m, 8H) 1.03 (s, 9H). LC-MS Retention Time 3.74mins, MH+ 634.
Intermediate 16; 5-Amino-Λ/-(2-r(r(3-chlorophenyl)carbonvn(2-r(1 ,1- dimethylethvπoxy1ethyl)amino)methyl1-3.3.3-trifluoro-2-hvdroxypropyl)-1-(4-fluorophenyl)- 1 H-pyrazole-4-carboxamide
Figure imgf000065_0002
A solution of 3-chlorobenzoic acid (13.8mg, 0.088mmol), 0-(7-azabenzotriazol-1-yl)- N,N,N',N'-tetramethyluronium hexafluorophosphate (HATU) (33mg, O.Oδδmmol) in anhydrous dimethylformamide (0.2ml) and diisopropylethylamine (0.03ml) was left at 21 °C for ca. 5 minutes. A solution of 5-amino-Λ/-{2-[({2-[(1 ,1-dimethylethyl) oxy]ethyl}amino)methyl]-3,3,3-trifluoro-2-hydroxypropyl}-1-(4-fluorophenyl)-1 H-pyrazole-4- carboxamide (0.07mmol) in dimethylformamide (0.1ml) was added. The solution was left at 210C for 19 hours before being diluted with methanol (0.3ml) and dimethylsulphoxide (0.1ml) and purified by MDAP. The appropriate fractions were eluted through bicarbonate cartridges (500mg) and evaporated. Toluene was added and evaporated to give the title compound (28.9mg). LC-MS Retention Time 3.67mins, MH+ 600.
Intermediate 17: 5-Amino-Λ/-(2-r(((2-r(difluoromethvπoxylphenyl|carbonyl)(2-r(1.1- dimethylethvπoxy1ethyl}amino)methyl1-3,3,3-trifluoro-2-hvdroxypropyl>-1-(4-fluorophenvπ- 1/-/-pyrazole-4-carboxamide
Figure imgf000066_0001
solution of O-(7-azabenzotriazol-1 -yl)-N,N, N', N'-tetramethyluronium hexafluorophosphate (HATU) (498.1 mg, 1.31 mmol) in dimethylformamide (2.5ml) was added to 2-difluoromethoxybenzoic acid (246.3mg, 1.31 mmol ). After 10 minutes, a solution of 5-amino-Λ/-{2-[({2-[(1 ,1-dimethylethyl)oxy]ethyl}amino)methyl]-3,3,3-trifluoro-2- hydroxypropyl}-1-(4-fluorophenyl)-1/-/-pyrazole-4-carboxamide (550mg, 1.19mmol) and diisopropylethylamine (461.6mg, 3.57mmol) in dimethylformamide (2.5ml) was added and the solution was stirred under nitrogen at room temperature. After 3.5 hours, the reaction mixture was concentrated in vacuo and the residue was purified by the Flashmaster with 0-100% ethyl acetate in cyclohexane gradient over 60 minutes as eluent. This gave the title product (423mg).
1H NMR (400 MHz, CHLOROFORM-cQ δ ppm 7.74 (s, 1 H) 7.47 (s, 2H) 7.42 (s, 3H) 7.25 - 7.31 (m, 1 H) 7.13 - 7.24 (m, 2H) 5.52 (br. s., 2H) 3.15 - 4.24 (m, 9H) 1.13 (s, 9H). LC-MS Retention Time 3.37mins, MH+ 631.
Intermediate 18: 5-Amino-1-(4-fluorophenvh-Λ/-(3.3,3-trifluoro-2-hvdroxy-2-(r(2- hydroxyethv0aminolmethyl}propyl)-1H-pyrazole-4-carboxamide
Figure imgf000066_0002
A solution of 5-amino-1-(4-fluorophenyl)-Λ/-{[2-(trifluoromethyl)-2-oxiranyl]methyl}-1H- pyrazolθ-4-carboxamidθ (413mg,1.2mmol) and athanolamine (217μlitres, 3.6mmol) in anhydrous tetrahydrofuran (13ml) was stirred at 210C under nitrogen for 20 hours. It was diluted with ethyl acetate (60ml) and washed with water (40ml) and saturated brine (2 x 30ml), dried over sodium sulphate and evaporated to give a solid foam (496mg) This was purified on a 1Og Silica SPE cartridge eluting with dichloromethane and then dichloromethane : ethanol (99-1 , 98.2, 96.4, 94:6, 92:8 (4 times), 90 10 (2 times) The product was eluted in the 94:6 and 92-8 fractions. These were combined and evaporated to give the title compound (405mg) as a solid.
1H NMR (400 MHz, DMSOd6) δ ppm 8 08 (t, I H) 7.95 (s, 1 H) 7.54 - 7.60 (m, 2H) 7 33 - 7 40 (in, 2H) 6.37 (br s., 3H) 4.59 (t, 1 H) 3.62 - 3 71 (m, 1 H) 3 42 - 3 53 (m, 3H) 2 72 - 2 82 (m, 2H) 2 57 - 2 68 (m, 2H) LC-MS Retention Time 1.96mιns, ES+ 406.
Intermediate 19- (S)-3.3,3-Trιfluoro-2-hvdroxy-2-(hvdroxymethyl)propyl butanoate
Figure imgf000067_0001
Preparation of Lipase Amano PS Precipitate
Amano PS (available from Amano Enzymes) (1Og) was suspended in water (30ml) and filtered through a Varian bond elute filter tube, washing with water (20ml) After precipitation with propan-2-ol (200ml), the suspension was allowed to settle and the supernatant decanted to leave a 50ml volume of suspension that was centrifuged at 4000 rpm for 5 minutes and the supernatant decanted to leave the title compound
Lipase Catalysed Desymmetrisation using Amano PS Precipitate
Vinyl butyrate (1ml) was added to a stirred mixture of 4A molecular sieves (2g), lipase Amano PS precipitate (100mg) and 2-(trιfluoromethyl)-1 ,2,3-propanetrιol (1g) suspended in 1 ,1-dιmethylethyl methyl ether (10ml) and the mixture stirred at room temperature The mixture was filtered through celite after 21.5 hours, washing with dichloromethane, concentrated, diluted with toluene (20ml) and washed with saturated aqueous sodium chloride (2 x 20ml), dried (Na2SO4) and evaporated to give a light yellow oil 19F NMR spectroscopy displayed a 20 1 mixture of monoester/diester with no starting tπol The yellow oil was purified by chromatography over silica using a 1 1 mixture of dichloromethane/ cyclohexane as eluent to remove the side products followed by elution with dichloromethaπe/ethyl acetate to afford the title compound as a pale yellow oil (0.64g).
1H NMR (400 MHz, CHLOROFORM-d) δ ppm 4.45 (d, 1 H) 4.35 (d, 1 H) 3.85 (dd, 1H) 3.73 (dd, 1H) 2.37 (t, 2H) 1.64 - 1.74 (m, 2H) 0.98 (t, 3H).
Enantiomeric excess Determination
(R)-(-)-a-Methoxy-a-trifluoromethylphenylacetyl chloride (45μl) and (S)-(+)-α-methoxy-α- trifluoromethylphenylacetyl chloride (45μl) were added to two parallel reactions containing mixtures of the title compound (20mg) in pyridine (22μl) and dichloromethane (1ml) and the resultant mixtures stirred at room temperature. After 1 hour, the mixtures were washed with a saturated aqueous solution of sodium bicarbonate (1 ml) then a 2M aqueous solution of HCI (1 ml) and finally water (1 ml) and the organic portions concentrated and the crude mixtures analysed by 19F NMR from which the title compound was found to have an enantiomeric excess of 88 % and 86 % respectively.
Alternative Preparation A of Intermediate 19
Preparation of Amano PS Solution
Amano lipase PS (100g) was suspended in 1 M pH 7 potassium phosphate solution
(400ml) and filtered, washing the cake with further buffer solution (100ml) to give a yellow/brown solution as the filtrate.
Immobilisation of Amano PS onto Sepabeads
Amano PS solution (100ml) was added to sepabeads EC-EP (available from Mitzubishi- Resindion) (4Og) and the mixture shaken at room temperature. After 23 hours as much liquid as possible was removed by pipette and the liquid replaced by an equal volume of a 1 M pH10 potassium phosphate solution and shaking continued. After 92 hours the mixture was filtered and the residue washed with water (3 x 50ml), suspended in a saturated solution of octadecylamine in toluene (50ml) and shaken at room temperature for a further 25 hours. The mixture was filtered, washed with toluene (2 x 100ml) and acetone (25ml) and left under suction to afford a free flowing powder. The immobilised enzyme was obtained as pale brown beads (17.29g).
Lipase Catalysed Desymmetrisation using Amano PS on Sepabeads
In parallel reactions, a mixture of 2-(trifluoromethyl)-1 ,2,3-propanetriol (100mg), Amano PS onto Sepabeads (100mg), vinyl butyrate (0.3ml) and solvent (as shown in table 1 below) (1 ml) was shaken at room temperature. The reactions were analysed by 19F NMR after 42 hours to give 2-(trifluoromethyl)-1 ,2,3-propanetriol (triol) / title compound /2- [(butanoyloxy)methyl]-3,3,3-trifluoro-2-hydroxypropyl butanoate (diester) peak ratios shown in table 1 below.
Figure imgf000069_0001
10 beads of 4 molecular sieves were also added at the start of the reaction.
Table 1 Enantiomeric Excess Determination
To parallel reactions, (R)-(-)-α-methoxy-α-trifluoromethylphenylacetyl chloride (45μl) was added to mixtures of crude title compound (20mg) in pyridine (22μl) and dichloromethane
(1 ml) and the resultant mixtures were stirred at room temperature. After 1 hour, the mixtures were washed with 2M HCI (1 ml), separated and the organic portions washed with a saturated aqueous solution of sodium bicarbonate (1 ml), the organic portions were then concentrated under a stream of nitrogen and analysed by 19F NMR from which the title compound enantiomeric excesses were obtained.
Figure imgf000069_0002
Table 2
Alternative Preparation B of Intermediate 19 A 10 g/L solution of 2-(trifluoromethyl)-1 ,2,3-propanetriol in vinyl butyrate/TBME (1 : 9) was prepared and passed through an Omnifit column (10cm x 0.6cm) for Entries 1 - 3 and 2 parallel Omnifit columns (10cm x 1.2cm) for Entries 4 and 5, all packed to full capacity with lipase Amano PS on sepabeads at room temperature using an HPLC pump and a pulse damper fitted after the column. The column was allowed to equilibrate for 1 hour on changing the conditions for each Entry before sampling (about 5ml samples taken) from the column outlet. Each sample taken was evaporated and analysed by 19F NMR.
Figure imgf000070_0001
Table 3
Enantiomeric Excess Determination
(R)-(-)-α-Methoxy-α-trifluoromethylphenylacetyl chloride (40μl) or (S)-(+)-α-methoxy-α- trifluoromethylphenylacetyl chloride was added to parallel reactions containing mixtures of monoesters (20mg) shown in Table 4 below in pyridine (20μ!) and dichloromethane (1 ml) and the resultant mixtures stirred at room temperature. After 1 hour the mixtures were washed with a 2M aqueous solution of HCI (1 ml) then a saturated aqueous solution of sodium bicarbonate (1 ml), the organic portions were concentrated and the crude mixtures analysed by 19F NMR from which the title compound enantiomeric excesses were obtained.
Figure imgf000070_0002
Figure imgf000071_0002
Table 4
*Samples 1 A, 5 and 5A were reacted with (S)-(+)-α-methoxy-α-trifluoromethylphenylacetyl chloride.
Intermediate 20: (ff)-3,3.3-Trifluoro-2-hvdroxy-2-((r(4- methylphenyl)sulfonyl1oxy)methylbropyl butanoate
Figure imgf000071_0001
(S)-3,3,3-trifluoro-2-hydroxy-2-(hydroxymethyl)propyl butanoate (372mg) was dissolved in dry dichloromethane (4ml) and dry pyridine (3ml) and cooled in an ice bath under nitrogen. P-Toluenesulphonyl chloride (0.4g) was added and the reaction was stirred at ice bath temperature for 1 hour and then at room temperature overnight. The reaction was left stirring at room temperature under nitrogen for a further 2 days. The solvent was evaporated and the residue was partitioned between ethyl acetate and water. The organic layer was washed with 1 M hydrochloric acid, water, sodium bicarbonate, brine, dried using a hydrophobic frit and evaporated to give an orange oil (0.68g). This was dissolved in dichloromethane and applied to a 5Og SPE cartridge and eluted with dichloromethane followed by 10% methanol in dichloromethane. The relevant fractions were combined and evaporated to give the title compound as a pale yellow oil (0.36g).
1H NMR (400 MHz, CHLOROFORM-d) d ppm 7.82 (d, 2H) 7.39 (d, 2H) 4.32 - 4.41 (m, 2H) 4.16 - 4.22 (m, 2H) 3.68 - 3.69 (m, 1 H) 2.48 (s, 3H) 2.34 (t, 2H) 1.60 - 1.69 (m, 2H) 0.95 (t, 3H). LC-MS Retention Time 3.33mins, MNH4 + 402.
Four further fractions which contained product by TLC (eluted with 10%MeOH/DCM) were combined and evaporated to give an orange oil (0.39g) which was dissolved in DCM and applied to a 2Og SPE column and purified on the Flashmaster eluting with a 0-50% ethyl acetate/cyclohexane 20 minute gradient. The fractions containing the product were evaporated to give a colourless oil (0.3g). Total yield (0.66g). LC-MS Retention Time 3.48mins, MNH4 + 402.
Intermediate 21 : { SV3.3.3-Trifluoro-2-hvdroxy-2-(r(phenylmethyl)aminolmethyl)propyl butanoate
Figure imgf000072_0001
Polymer Supported Carbonate Resin (ex Fluka, 3.5mmol/g, 0.96g) was washed thoroughly with dry tetrahydrofuran (5 times). (ft)-3,3,3-trifluoro-2-hydroxy-2-({[(4- methylphenyl) sulfonyl]oxy}methyl)propyl butanoate (0.43g) was dissolved in dry tetrahydrofuran (5ml) and shaken with the polymer supported carbonate overnight. The resin was filtered off and the reaction solution was treated with benzylamine (122μl). The reaction was stirred at room temperature under nitrogen, overnight. A further portion of benzylamine (24μl) was added and the reaction was stirred at room temperature for 2 hours. The solvent was evaporated to give the title compound as a colourless liquid (0.38g). 1H NMR (400 MHz, CHLOROFORM-d) 3 ppm 7.27 - 7.38 (m, 5H) 4.28 - 4.32 (m, 1 H) 4.16 - 4.20 (m, 1 H) 3.82 - 3.92 (m, 4H) 3.01 - 3.06 (m, 1 H) 2.71 - 2.77 (m, 1 H) 2.28 - 2.32 (m, 2H) 1.60 - 1.69 (m, 2H) 0.91 - 0.99 (m, 3H). LC-MS Retention Time 2.72mins, MH+ 320.
Intermediate 22: (S)-3,3,3-Trifluoro-2-(r(phenylmethyl^amino1methyl)-1.2-propanediol
Figure imgf000072_0002
(S)-3,3,3-Trifluoro-2-hydroxy-2-{[(phenylmethyl)amino]methyl}propyl butanoate (0.32g) was dissolved in 5M hydrochloric acid (5ml) and ethanol (5ml) and heated at 1000C for 3 hours. After cooling, the ethanol was evaporated and the remaining aqueous was basified with 2M sodium hydroxide and extracted with ethyl acetate (2 times). The combined organics were dried using a hydrophobic frit and evaporated to give the title compound as a pale yellow oil (0.26g).
1H NMR (400 MHz, CHLOROFORM-d) 3 ppm 7.28 - 7.40 (m, 5H) 3.82 - 3.91 (m, 3H) 3.61 (d, 1 H) 3.52 (s, 1 H) 3.12 (d, 1 H) 2.91 (d, 1 H). LC-MS Retention Time 1.65mins, MH+ 250.
Intermediate 23: (S)-2-( Aminomethyl)-3,3.3-trifluoro-1 ,2-propanediol
Figure imgf000073_0001
(S)-3,3,3-Trifluoro-2-{[(phenylmethyl)amino]methyl}-1 ,2-propanediol (160mg) was dissolved in methanol (1.5ml) and hydrogenated using 10% palladium on carbon as the catalyst and the H-cube (Thales) as the hydrogen source. The solvent was carefully evaporated to give a pinkish volatile oil (112mg) which by 1NMR still contained 13% starting material. Therefore this was re-dissolved in methanol (2ml) and re-hydrogenated using the H-cube. The solvent was evaporated to give the title compound as a pink/brown oil (93mg).
1H NMR (400 MHz, CHLOROFORM-d) 3 ppm 3.85 (d, 1 H) 3.57 (d, 1 H) 3.50 (s, 4H) 3.15
(s, 1 H) 2.98 (d, 1H).
LC-MS Retention Time 0.41 mins, MH+ 160.
Intermediate 24: (S)-5-Amino-1-(4-fluorophenv0-ftH3.3,3-trifluoro-2-hvdroxy-2-
(hvdroxymethyl)propyl1-1 /-/-pyrazole-4-carboxamide
Figure imgf000073_0002
5-Amino-1-(4-fluorophenyl)-1H-pyrazole-4-carboxylic acid (104rηg) was dissolved in dry dimethylformamide (3ml) and diisopropylethylamine (0.33ml) was added followed by the addition of O-(7-Azabenzotriazol-1-yl)-N,N,N',N'-tetramethyl uronium hexafluorophosphate (196mg). The reaction was stirred at room temperature, under nitrogen, for 20 minutes. (S)-2-(Aminomethyl)-3,3,3-trifluoro-1 ,2-propanediol (90mg) in dry dimethylformamide (2ml) was added and the reaction was stirred at room temperature for 3 hours. The reaction was partitioned between ethyl acetate (50ml) and water (50ml) and the separated aqueous phase was re-extracted with ethyl acetate (50ml). The combined organics were washed with water, 1 M hydrochloric acid, 10% lithium chloride solution (2 times), bicarbonate and brine. The organics were dried using a hydrophobic frit and evaporated to give a pale yellow oil (200mg). This was dissolved in dichloromethane (ca.
5ml) and left to stand at room temperature for 4 hours wherein a precipitate formed. The precipitate was filtered and washed with a small amount of dichloromethane to give the title compound as a white solid (93mg).
1H NMR (400 MHz, DMSO-cfe) δ ppm 8.15 - 8.22 (m, 1 H) 8.01 (s, 1 H) 7.55 - 7.60 (m, 2H)
7.34 - 7.40 (m, 2H) 5.76 (s, 2H) 5.17 - 5.21 (m, 1 H) 3.40 - 3.68 (m. 4H).
LC-MS Retention Time 2.47mins, MH+ 363.
The filtrate from above was evaporated and the residue was dissolved in DMSO/MeOH
(1 ml 1 :1 ) and purified on the MDAP. Fractions containing product were combined and evaporated to give the title compound as a colourless oil (28mg) (total yield 121 mg)
1H NMR showed pure product.
LC/MS Retention Time 2.51 mins, MH+ 363.
Intermediate 25: (S)-2-r(ir5-Amino-1 -(4-fluorophenvπ-1 H-pyrazol-4- yl1carbonyl}amino)methyl1-3.3,3-trifluoro-2-hydroxypropyl 4-methylbenzenesulfonate
Figure imgf000074_0001
(S)-5-Amino-1-(4-fluorophenyl)-Λ/-[3,3,3-trifluoro-2-hydroxy-2-(hydroxymethyl)propyl]-1 /-/- pyrazole-4-carboxamide (90mg) was dissolved in dry dichloromethane (3ml) and dry pyridine (3ml) and cooled in an ice bath under nitrogen. p-Toluenesulphonyl chloride (62mg) was added and the reaction was stirred at ice bath temperature for 1 hour and then at room temperature for 24 hours. p-Toluenesulphonyl chloride (25mg) was added and the reaction was stirred at room temperature, under nitrogen over the weekend, p- Toluenesulphonyl chloride (25mg) was added and the reaction was stirred at room temperature under nitrogen for 2 hours and heated at 5O0C for 2 hours and left stirring at room temperature overnight. The solvent was evaporated and the residue was partitioned between dichloromethane and water. The organic layer was washed with 1 M hydrochloric acid, water, sodium bicarbonate and brine, dried using a hydrophobic frit and evaporated to give a beige foam (94mg). This was dissolved in dichloromethane and applied to a 10g SPE cartridge and purified on the Flashmaster eluting with a 0-50% ethyl acetate in cyclohexane over a 20 minute gradient. The appropriate fraction was evaporated to give the title compound as a colourless oil (83mg).
1H NMR (400 MHz, CHLOROFORM-d) d ppm 7.79 - 7.83 (m, 2H) 7.71 (s, 1 H) 7.50 - 7.55 (m, 2H) 7.37 - 7.41 (m, 2H) 7.20 - 7.26 (m, 2H) 6.60 - 6.68 (m, 1 H) 6.56 (s, 1 H) 5.46 (s, 2H) 4.17 - 4.24 (m, I H) 3.98 - 4.07 (m, 1 H) 3.60 - 3.70 (m, 1 H) 3.45 - 3.55 (m, 1 H) 2.47 (S, 3H). LC-MS Retention Time 3.33mins, MH+ 517.
Intermediate 26: (S)-5-Amino-1 -(4-fluorophenyl)-Λ/-(f2-(trifluoromethyl)-2-oxiranyllmethyl)- 1 AV-pyrazole-4-carboxamide.
Figure imgf000075_0001
Polymer Supported carbonate resin (3.5mmol/g, 137mg) was washed thoroughly with dry tetrahydrofuran (5 times). (S)-2-[({[5-Amino-1-(4-fluorophenyl)-1 H-pyrazol-4- y|]carbonyl}amino)methy|]-3,3,3-trifluoro-2-hydroxypropyl 4-methyl benzene sulfonate
(83mg) was dissolved in dry tetrahydrofuran (5ml) and shaken with the polymer supported resin overnight. The resin was filtered off and the solution was evaporated to give the title compound as a beige solid (60mg).
1H NMR (400 MHz, CHLOROFORM-d) d ppm 7.60 (s, 1 H) 7.50 - 7.55 (m, 2H) 7.19 - 7.25
(m, 2H) 5.73 (br. s., 1 H) 5.47 (br. s., 2H) 4.21 - 4.31 (m, 1 H) 3.73 - 3.78 (m, 1 H) 3.13 (d,
1 H) 2.95 - 2.98 (m, 1 H).
LC-MS Retention Time 2.84mins, MH+ 345.
Intermediate 27: (RV5-Amino-/V-(2-f K2-W1.1 - dimethylethvπ(dimethvπsilylloxy)ethyl)aminolmethylV3,3.3-trifluoro-2-hvdroxypropyl)-1-(4- fluorophenvπ-1/-/-pyrazole-4-carboxamide
Figure imgf000075_0002
(S];5-Amino-1-(4-fluorophenyl)-Λ/-{[2-(trifluoromethyl)-2-oxiranyl]methyl}-1A7-pyrazole-4- carboxamide (50mg) was dissolved in acetonitrile (5ml) and (2-{[(1 ,1- dimethylethyl)(dimethyl)silyl]oxy}ethyl)amine (28mg) was added. The reaction was heated at reflux overnight. After cooling, the solvent was evaporated to give the title compound as a colourless oil (76mg).
1H NMR (400 MHz, CHLOROFORM-d) d ppm 7.62 (s, 1 H) 7.50 - 7.55 (m, 2H) 7.19 - 7.25 (m, 2H) 6.13 - 6.21 (m, 1 H) 5.47 (s, 2H) 3.80 - 3.92 (m, 1 H) 3.63 - 3.73 (m, 2H) 3.51 - 3.61 (m, 1 H) 3.10 (d, 1 H) 2.69 - 2.84 (m, 2H) 2.65 (d, 1 H) 0.90 (s, 9H) 0.06 (s, 6H). LC-MS Retention Time 3.51 mins, MH+ 520.
Intermediate 28: (S)-5-Amino-Λ/-(2-(rr(2.6-difluorophenvncarbonyll(2-(r(1.1- dimethylethyl)(dimethyl)sιlylloxy>ethvπamino1methyl)-3.3.3-trifluoro-2-hvdroxypropyπ-1-(4- fluorophenyl)-1/-/-pyrazole-4-carboxamide
Figure imgf000076_0001
(R)-5-Amino-Λ/-(2-{[(2-{[(1 ,1-dimethylethyl)(dimethyl)silyl]oxy}ethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1 -(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide (38mg) was dissolved in dichloromethane (5ml) and 2,6-difluorobenzoyl chloride (1 Oμl) and diisopropylethylamine (32μl) were added. The mixture was stirred at room temperature overnight.The reaction was then partitioned between dichloromethane and water. The organic layer was dried using a hydrophobic frit and the solvent was evaporated to give a pale yellow solid (54mg).
1H NMR (400 MHz, CHLOROFORM-cO d ppm 7.74 (s, 1H) 7.50 - 7.58 (m, 2H) 7.36 - 7.45 (m, 1 H) 7.16 - 7.25 (m, 2H) 6.96 - 7.06 (m, 2 H) 5.95 (br. S., 1 H) 5.40 - 5.51 (m, 1H) 4.13 -
4.28 (m, 1 H) 4.04 - 4.13 (m, 1 H) 3.80 - 3.91 (m, 1 H) 3.59 - 3.68 (m, 2H) 3.47 - 3.59 (m,
2H) 3.35 - 3.46 (m, 1 H) 0.85 (s, 9H) -0.02 - 0.01 (m, 6H).
LC-MS Retention Time 4.03 mins, MH+ 660. Example 1 5-Amιno-Λ/-(2-(r({2-r(dιfluoromethvπoxyiphenyl)carbonyl)(2- hvdroxyethvπamιnolmethyl)-3,3.3-trιfluoro-2-hvdroxypropyπ-1-(4-fluorophenyl)-1 AV- pyrazole-4-carboxamιde
Figure imgf000077_0001
5-Amιno-Λ/-{2-[(({2-[(dιfluoromethyl)oxy]phenyl}carbonyl){2-[(1 ,1- dιmethylethyl)oxy]ethyl}amιno)methy|]-3,3,3-tπfluoro-2-hydroxypropyl}-1-(4-fluorophenyl)- IH-pyrazole-4-carboxamιde (50mg, 0.079mmol) was dissolved in dichloromethane (0 5ml) before trifluoroacetic acid (0 5ml) was added The reaction mixture was then stirred at room temperature for 3 5 hours The reaction mixture was concentrated via the blow down unit The residue was purified via MDAP and the relevant fraction was passed down a bicarbonate SPE cartridge and the cartridge was flushed with acetonitπle The filtrate was blown down to give the title product (17mg) 1H NMR (400 MHz, CHLOROFORM-cf) δ ppm 7 72 (s, 1H) 7 50 - 7 56 (m, 2H) 7 43 (s, 1H) 7 25 - 7 32 (m, 3H) 7.19 - 7.25 (m, 2H) 5 45 (s, 2H) 3 29 - 4 27 (m, 9H) LC-MS Retention Time 2 91 mιns, MH+ 576
Example 1 was further preparatively separated into its enantiomers, Enantiomer 1 and Enantiomer 2 (Isomers A and B) via method C using a Chiralcel OD column eluting with 40% ethanol in heptane at a flow rate of 75ml/mιn Elution Time for Enantiomer 1 - 7 5mιns Elution Time for Enantiomer 2 - 10 3mιns
Enantiomer 1 (Isomer A)
Analytical Chiral HPLC - Method A (25 x 0 46cm ChiralCel OD Column, 40% ethanol in heptane eluting at 1 ml/mιn) - Retention Time 4 2mιns
Circular Dichroism (MeCN, RT, 0 00013OM, v = 350-200nm, cell length 0 2cm)
227 2nm (de = 2 38)
267 0nm (de = 2 15)
Enantiomer 2 (Isomer B) Analytical Chiral HPLC - Method A (25 x 0.46cm ChiralCel OD Column, 40% ethanol in heptane eluting at 1 ml/min) - Retention Time 6.9mins.
Circular Dichroism (MeCN, RT, 0.000131 M1 v = 350-200nm, cell length 0.2cm) 223.2nm (de = -2.76) 267.0nm (de = -2.39)
Example 2: 5-Amino-Λ/-(2-(r[(3-chlorophenyl)carbonyll(2-hvdroxyethvπamino1methyl>- 3,3,3-trifluoro-2-hvdroxypropyl)-1-(4-fluorophenyl)-1/7-pyrazole-4-carboxamide
Figure imgf000078_0001
solution of 5-amino-Λ/-{2-[([(3-chlorophenyl)carbonyl]{2-[(1 ,1- dimethylethyl)oxy]ethyl}amino)methyl]-3,3,3-trifluoro-2-hydroxypropyl}-1-(4-fluorophenyl)- 1/-/-pyrazole-4-carboxamide (28mg) in trifluoroacetic acid : dichloromethane (1 : 1 ) (0.5ml) was left at 21 °C for 2 hours. It was then blown down and then purified by MDAP. This gave the title product (9.4mg).
1H NMR (400 MHz, DMSO-cfe) δ ppm 8.07 (s, 1 H) 7.89 (s, 1H) 7.55 - 7.60 (m, 2H) 7.46 - 7.54 (m, 3H) 7.33 - 7.42 (m, 3H) 6.92 (s, 1 H) 6.38 (s, 2H) 4.82 - 4.93 (m, 1 H) 4.00 - 4.12 (m, 1 H) 3.84 (dd, 1 H) 3.71 - 3.79 (m, 1H) 3.40 - 3.52 (m, 5H). LC-MS Retention Time 3.08mins, MH+ 544.
Example 3: 5-Amino-1-(4-fluorophenyO-Λ/-(3Λ3-trifluoro-2-(rr(2-fluorophenyl)carbonyll(2- hvdroxvethvl)aminolmethvl}-2-hvdroxypropvl)-1A7-pyrazole-4-carboxamide
Figure imgf000078_0002
2-Fluorobenzoyl chloride (O.i mmol) was weighed into a vial before a solution of 5-amino- Λ/-{2-[({2-[(1 ,1-dimethylethyl)oxy]ethyl}amino)methyl]-3,3,3-trifluoro-2-hydroxypropyl}-1-(4- fluorophenyl)-1H-pyrazole-4-carboxamide (52mg) and diisopropylethylamine (0.035ml, 0.2mmol) in dichloromethane (0.7ml) was added. It was left to stand at room temperature overnight. Aqueous sodium bicarbonate (0.75ml) and chloroform (1.8ml) were added, the mixture shaken and the separated aqueous phase was further washed with chloroform. The chloroform washings were blown down in a scintillation vial before a mixture of trifluoroacetic acid : water : dichloromethane : anisole (20 : 1.5 : 20 : 1 ) (6ml) was added to the residue. It was left overnight and evaporated in vacuo. The residue was treated with aqueous sodium bicarbonate (0.75ml) and chloroform (1.8ml) and shaken. The organic extract was evaporated and the residue was purified by MDAP. The appropriate fractions from the MDAP were made basic with aqueous sodium bicarbonate, evaporated to remove the acetonitrile and then extracted with chloroform. The extract was filtered through a phase separator and evaporated to give the crude title compound. This was triturated twice with petroleum ether (40° - 60°) to give title compound (41.3mg) as a white solid.
1H NMR (400 MHz, DMSO-c/6) δ ppm 8.06 (t, 1H) 7.85 (s, 1H) 7.47 - 7.58 (m, 2H) 7.28 - 7.38 (m, 6H) 3.27 - 3.94 (m, 10H). LC-MS Retention Time 2.96mins, ES+ 528.
Example 3 was further preparatively separated into its enantiomers, Enantiomer 1 and Enantiomer 2 (Isomers A and B) via method B using a Chiralcel OD column eluting with 40% ethanol in heptane at a flow rate of 15ml/min. Elution Time for Enantiomer 1 - 6mins. Elution Time for Enantiomer 2 - 12mins.
Enantiomer 1 (Isomer A)
Analytical Chiral HPLC Method A (25 x 0.46cm ChiralCel OD Column, 40% ethanol in heptane eluting at 1ml/min) - Retention Time 4.9mins. Circular Dichroism (MeCN, RT, 0.000148M, v = 350-200nm, cell length 0.2cm) 202.2nm (de = 8.00) 226.0nm (de = 1.87) 264.6nm (de = 2.54).
Enantiomer 2 (Isomer B)
Analytical Chiral HPLC Method A (25 x 0.46cm ChiralCel OD Column, 40% ethanol in heptane eluting at 1 ml/min) - Retention Time 9.22mins.
Circular Dichroism (MeCN1 RT, 0.000114M, v = 350-200nm, cell length 0.2cm)
202.0nm (de = -9.00) 226.0nm (de = -2.47)
266.4nm (de = -3.31 ). Example 4: 5-Amino-Λ/-(2-{[r(2-chlorophenyl)carbonyll(2-hvdroxyethyl)aminolmethyl)- 3,3,3-trifluoro-2-hvdroxypropylV1-(4-fluorophenyl)-1H-pyrazole-4-carboxamide
Figure imgf000080_0001
2-Chlorobenzoyl chloride (O.immol) was weighed into a vial before a solution of 5-amino- /V-{2-[({2-[(1 ,1-dimethylethyl)oxy]ethyl}amino)methyl]-3,3,3-trifluoro-2-hydroxypropyl}-1-(4- fluorophenyl)-1H-pyrazole-4-carboxamide (52mg) and diisopropylethylamine (0.035ml, 0.2mmol) in dichloromethane (0.7ml) was added. It was left to stand at room temperature overnight. Aqueous sodium bicarbonate (0.75ml) and chloroform (1.8ml) were added, the mixture shaken and the separated aqueous phase was further washed with chloroform. The chloroform washings were blown down in a scintillation vial before a mixture of trifluoroacetic acid : water : dichloromethane : anisole ( 20 : 1.5 : 20 : 1 ) (6ml) was added to the residue. It was left overnight and evaporated in vacuo. The residue was treated with aqueous sodium bicarbonate (0.75ml) and chloroform (1.8ml) and shaken. The organic extract was evaporated and the residue was purified by MDAP. The appropriate fractions from the MDAP were made basic with aqueous sodium bicarbonate, evaporated to remove the acetonitrile and then extracted with chloroform. The extract was filtered through a phase separator and evaporated to give the crude title compound. This was triturated twice with petroleum ether (40° - 60°) to give title compound (49mg) as a white solid.
1H NMR (400 MHz, DMSOd6) δ ppm 8.27 (s, I H) 8.07 (t, I H) 7.51 - 7.60 (m, 3H) 7.41 - 7.49 (m, 3H) 7.31 - 7.41 (m, 2H) 3.11 - 4.37 (m, 10H). LC-MS Retention Time 3.05mins, ES+ 544.
Example 4 was further preparatively separated into its enantiomers, Enantiomer 1 and Enantiomer 2 (Isomers A and B) via method B using a Chiralcel OD column eluting with 40% ethanol in heptane at a flow rate of 15ml/min. Elution Time for Enantiomer 1 - 6mins. Elution Time for Enantiomer 2 - 10mins.
Enantiomer 1 (Isomer A) Analytical Chiral HPLC - Method A (25 x 0.46cm ChiralCel OD Column, 40% ethanol in heptane eluting at 1 ml/min) - Retention Time 4.7mins.
Circular Dichroism (MeCN, RT, 0.000125M1 v = 350-200nm, cell length 0.2cm)
201.6nm (de = 1 1.60)
225.4nm (de = 5.13)
262.2nm (de = 2.86)
Enantiomer 2 (Isomer B)
Analytical Chiral HPLC - Method A (25 x 0.46cm ChiralCel OD Column, 40% ethanol in heptane eluting at 1 ml/min) - Retention Time 8.3mins.
Circular Dichroism (MeCN, RT, 0.000122M, v = 350-200nm, cell length 0.2cm)
202.2nm (de = -1 1.20)
225.4nm (de = -4.80)
263.2nm (de = -3.01 )
Example 5: 5-Amino-/V-(2-(fr(2-chloro-6-fluorophenyl)carbonyl1(2- hvdroxyethyl)amino1methyl)-3.3,3-trifluoro-2-hvdroxypropyl)-1-(4-fluorophenylV1f/- pyrazole-4-carboxamide
Figure imgf000081_0001
2-Chloro-6-fluorobenzoyl chloride (O.immol) was weighed into a vial before a solution of 5-amino-/V-{2-[({2-[(1 ,1-dimethylethyl)oxy]ethyl}amino)methy|]-3,3,3-trifluoro-2- hydroxypropyl}-1 -(4-fluorophenyl)-1 H-pyrazole-4-carboxamide (46mg) and diisopropylethylamine (0.035ml, 0.2mmol) in dichloromethane (0.7ml) was added. It was left to stand at room temperature overnight. Aqueous sodium bicarbonate (0.75ml) and chloroform (1.8ml) were added, the mixture shaken and the separated aqueous phase was further washed with chloroform. The chloroform washings were blown down in a scintillation vial before a mixture of trifluoroacetic acid : water : dichloromethane : anisole (20 : 1.5 : 20 : 1 ) (6ml) was added to the residue. It was left overnight and evaporated in vacuo. The residue was treated with aqueous sodium bicarbonate (0.75ml) and chloroform (1.8ml) and shaken. The organic extract was evaporated and the residue was purified by MDAP. The appropriate fractions from the MDAP were made basic with aqueous sodium bicarbonate, evaporated to remove the acetonitrile and then extracted with chloroform. The extract was filtered through a phase separator and evaporated to give the crude title compound. This was triturated twice with petroleum ether (40° - 60°) to give the title compound (34mg) as a white solid. 1H NMR (400 MHz, DMSO-Cf6) δ ppm 8.25 (s, 1 H) 8.06 (t, 1 H) 7.83 (d, 1 H) 7.50 - 7.60 (m, 3H) 7.30 - 7.48 (m, 3H) 3.24 - 4.26 (m, 10H). LC-MS Retention Time 3.09mins, ES+ 562.
Example 5 was further separated into its Isomers A (probably a mixture of rotamers 1 and 2 and atropisomers 1 and 2) and B (probably a mixture of rotamers 3 and 4 and atropisomers 3 and 4)) using analytical chiral HPLC.
Isomers A (probably a mixture of rotamers 1 and 2 and atropisomers 1 and 2) Analytical Chiral HPLC (25 x 0.46cm Chiralcel OD column, 20% ethanol in heptane eluting at 1ml/min) - Retention time 7.55 mins and 8.73 mins.
Isomers B (probably a mixture of rotamers 3 and 4 and atropisomers 3 and 4) Analytical Chiral HPLC (25 x 0.46cm Chiralcel OD column, 20% ethanol in heptane eluting at 1 ml/min) - Retention time 20.32 mins and 39.49 mins.
Example 6: 5-Amino-Λ/-(2-ffr(2.6-difluorophenvπcarbonylK2-hvdroxyethyl)amino1methyl)- 3.3.3-trifluoro-2-hvdroxypropylV1-(4-fluorophenyl)-1H-pyrazole-4-carboxamide
Figure imgf000082_0001
A solution of 5-amino-Λ/-{2-[([(2,6-difluorophenyl)carbonyl]{2-[(1 ,1- dimethylethyl)oxy]ethyl}amino)methyl]-3,3,3-trifluoro-2-hydroxypropyl}-1-(4-fluorophenyl)- 1H-pyrazole-4-carboxamide (1.22g, 2.02mmol) was dissolved in dichloromethane (6ml) and trifluoroacetic acid (6ml) and water (0.05ml) was added. It was stirred at 21 °C for 5.5 hours before being added slowly to concentrated aqueous sodium bicarbonate (100ml) and extracted with dichloromethane (50ml). The organic layer was separated and evaporated. The residue (1.209g) was dissolved in methanol (20ml) and stirred with polymer supported carbonate resin (ca. 1.5g, 3.5mmol/g) for 4 hours. The resin was filtered off and the filtrate was evaporated. The residue was purified on the Flashmaster 2 (100g column) using 0-100% ethyl acetate in cyclohexane gradient over 60 minutes to give the title compound (843mg) as a foam.
1H NMR (400 MHz, DMSO-cfe) δ ppm 8.06 (br. s., 1 H) 7.87 (s, 1 H) 7.53 - 7.62 (m, 3H) 7.33 - 7.40 (m, 2H) 7.19 - 7.28 (m, 2H) 7.03 (s, 1 H) 6.38 (s, 2H) 3.36 - 3.89 (m, 9H). LC-MS Retention Time 2.94mins, MH+ 546.
Example 6 was further preparatively separated into its enantiomers, Enantiomer 1 and Enantiomer 2 (Isomers A and B) via method C using a Chiralcel OD(5 x 25cm) column eluting with 40% ethanol in heptane at a flow rate of 75ml/min. Elution Time for Enantiomer 1 - 7.6mins. Elution Time for Enantiomer 2 - 12.5mins.
Enantiomer 1 (Isomer A)
Analytical Chiral HPLC - Method A (25 x 0.46cm ChiralCel OD Column, 50% ethanol in heptane eluting at 1 ml/min) - Retention Time 3.76mins.
Circular Dichroism (MeCN, RT, 0.000123M, v = 350-200nm, cell length 0.2cm)
203.4nm (de = 8.25) 228.2nm (de = 3.90)
265.0nm (de = 3.58).
Enantiomer 2 (Isomer B)
Analytical Chiral HPLC - Method A (25 x 0.46cm ChiralCel OD Column, 50% ethanol in heptane eluting at 1 ml/min) - Retention Time 7.22mins.
Circular Dichroism (MeCN, RT, 0.000118M1 v = 350-200nm, cell length 0.2cm)
204.6nm (de = -7.82)
228.4nm (de = -3.69)
264.0nm (de = -3.72).
Example 6 Enantiomer 2: (S)-5-Amino-Λ/-(2-(rr(2,6-difluorophenyl)carbonvn(2- hvdroxyethyl)amino1methyl)-3.3.3-trifluoro-2-hvdroxypropyπ-1-(4-fluorophenyl)-1/-/- pyrazole-4-carboxamide
Figure imgf000084_0001
(S)-5-Amiπo-Λ/-(2-{tt(2,6-difluorophenyl)carbony|](2-{[(1,1-dimethylethyl)
(dimethyl)silyl]oxy}ethyl)amino]methyl}-3,3,3-trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)- IH-pyrazole-4-carboxamide (54mg) was dissolved in tetrahydrofuran (5ml) and tert- butylammonium fluoride (1 M in tetrahydrofuran, 0.22ml) was added. The reaction was stirred at room temperature for 1 hour. The solvent was evaporated and the residue was partitioned between dichloromethane and water. The organic layer was washed with water, dried using a hydrophobic frit and evaporated to give a pale yellow solid (45mg). This solid was dissolved in methanokdimethylsulphoxide (1 : 1 ) (1 ml) and purified via MDAP to give the title compound as a colourless oil (19mg).
1H NMR (400 MHz, CHLOROFORM-d) d ppm 7.73 (s, 1H) 7.49 - 7.55 (m, 2H) 7.37 - 7.45 (m, 1 H) 7.19 - 7.25 (m, 2H) 7.04 - 7.11 (m, 1 H) 6.96 - 7.03 (m, 2H) 5.46 (s, 2H) 3.95 - 4.10 (m, 2H) 3.60 - 3.73 (m, 4H) 3.48 - 3.52 (m, 2H) LC-MS Retention Time 3.03 mins, MH+ 546.
Chiral HPLC showed an enantiomeric excess of 83% (cf. starting enantiomeric excess 84%).
Example 7: 5-Amino-Λ/-(2-f[[(2.6-dichlorophenyl)carbonyll(2-hvdroxyethyl)aminolmethyl)- 3,3,3-trifluoro-2-hvdroxypropyl)-1-(4-fluorophenyl)-1 H-pyrazole-4-carboxamide
Figure imgf000084_0002
To a solution of 5-amino-Λ/-{2-[([(2,6-dichlorophenyl)carbonyl]{2-[(1 ,1- dimethylethyl)oxy]ethyl}amino)methy|]-3,3,3-trifluoro-2-hydroxypropyl}-1-(4-fluorophenyl)- 1H-pyrazole-4-carboxamide (1.32g, 2.08mmol) in dichloromethane (6ml) was added trifluoroacetic acid (6ml) and water (0.05ml). It was stirred at 21 °C for 5 hours before being added slowly to concentrated aqueous sodium bicarbonate (100ml) and extracted with dichloromethane (50ml). The organic layer was separated and evaporated. The residue (1.209g) was dissolved in methanol (20ml) and stirred with polymer supported carbonate resin (ca. 1.5g, 3.5mmol/g) for 4 hours. The resin was filtered off and the filtrate was evaporated to give a residue (1.17g). The residue was purified on the Flashmaster 2 (100g column) using a 0-100% ethyl acetate in cyclohexane gradient over 60 minutes to give the title compound (923mg) as a foam.
1H NMR (400 MHz1 DMSO-d6) δ ppm 8.09 (t, 1 H) 7.85 (s, 1 H) 7.45 - 7.67 (m, 5H) 7.33 - 7.41 ( m, 2H) 7.09 (s, 1 H) 6.37 (br. s., 2H) 4.88 (br. s., 1 H) 3.16 - 4.08 (m, 8H). LC-MS Retention Time 3.08mins, MH+ 578, 582.
Example 7 was further preparatively separated into its enantiomers, Enantiomer 1 and Enantiomer 2 (Isomers A and B) via method C using a Chiralcel OD column (5 x 25cm) eluting with 30% ethanol in heptane at a flow rate of 75ml/min. Elution Time for Enantiomer 1 - 9.0mins. Elution Time for Enantiomer 2 - 15.5mins.
Enantiomer 1 (Isomer A)
Analytical Chiral HPLC - Method A (25 x 0.46cm ChiralCel OD Column, 40% ethanol in heptane eluting at 1ml/min) - Retention Time 4.48mins.
Circular Dichroism (MeCN, RT, 0.000111M, v = 350-200nm, cell length 0.2cm) 206.6nm (de =13.21 ) 229.8nm (de = 6.18) 263.4nm (de = 3.08).
Enantiomer 2 (Isomer B) Analytical Chiral HPLC - Method A (25 x 0.46cm ChiralCel OD Column, 40% ethanol in heptane eluting at 1 ml/min) - Retention Time 8.98mins.
Circular Dichroism (MeCN, RT, 0.000129M, v = 350-200nm, cell length 0.2cm)
207.0nm (de = -13.64)
231.0nm (de = -6.33) 264.0nm (de = -3.25).
Example 8j 5-Amino-Λ/-(2-([r(2-chloro-3-fluorophenvπcarbonyll(2- hvdroxyethvhamino1methyl>-3.3.3-trifluoro-2-hvdroxypropyπ-1-(4-fluorophenyl)-1 /-/- pyrazole-4-carboxamide
Figure imgf000086_0001
A solution of 2-chloro-3-fluorobenzoic acid (0.088mmol) and 0-( 7-azabenzotriazol-1-yl ) - N,N,N',N'-tetramethyluronium hexafluorophosphate (HATU) (33mg, O.Oδδmmol) in dimethylformamide (0.2ml) was treated with diisopropylethylamine (0.03ml, O.iδmol). The mixture was shaken for 1 minute before a solution of 5-amino-Λ/-{2-[({2-[(1 ,1- dimethylethyl)oxy]ethyl}amino)methyl]-3,3,3-trifluoro-2-hydroxypropyl}-1-(4-fluorophenyl)- 1H-pyrazole-4-carboxamide (32mg, 0.07mmol) in dimethylformamide (0.1 ml) was added. The mixture was shaken for a further minute and then allowed to stand at room temperature for 1δ hours. Dimethylsulphoxide (0.3ml) was added and the mixture purified by MDAP. This gave the intermediate fert-butoxy derivative (14.3mg). This was dissolved in trifluoroacetic acid : dichloromethane(1 : 1 ) (0.5ml) and left to stand at room temperature for 2 hours. The residue was purified by MDAP to give the title compound (6.1 mg). 1H NMR (600 MHz, DMSO-cfe) δ ppm 8.06 (t, 1 H) 7.65 (s, 1 H) 7.55 - 7.61 (m, 2H) 7.48 - 7.53 (m, 2H) 7.32 - 7.39 (m, 3H) 6.99 (s, 1 H) 6.35 (br. s., 2H) 3.19 - 4.12 (m, 8H). LC-MS Retention Time 3.04mins, ES+ 562.
Example 8 was further preparatively separated into its enantiomers (Isomers A and B) using a 2" x 20cm Chiralcel OD column elutiπg with 40% ethanol in heptane at a flow rate of 75m!/min.
Enantiomer 1 (Isomer A)
Analytical Chiral HPLC (25 x 4.6cm Chiralcel OD column, 40% ethanol in heptane eluting at 1 ml/min) - Retention time 4.26mins.
Enantiomer 2 (Isomer B)
Analytical Chiral HPLC (25 x 0.46cm Chiralcel OD column, 40% ethanol in heptane eluting at 1 ml/min) - Retention time 8.37mins. Example 9: 5-Amino-1-(4-fluoroDhenvl)-Λ/-(3,3,3-trifluoro-2-hvdroxv-2-(rι'2- hvdroxvethvDfphenvlcarbonvπaminolmethvDpropvn-I H-pvrazole-^carboxamide
Figure imgf000087_0001
A solution of benzoyl chloride (8.8μlitre, 0.05mmol), diisopropylethylamine (7.4μlitre, O.immol) and 5-amino-1-(4-fluorophenyl)-Λ/-(3,3,3-trifluoro-2-hydroxy-2-{[(2- hydroxyethyl)amino]methyl}propyl)-1H-pyrazole-4-carboxamide (0.05mmol) in dichloromethane (200μlitre) was stirred at room temperature in a greenhouse and the reactions monitored. Further benzoyl chloride (8.8μlitre, 0.05mmol) was added to push the reaction to completion. The reaction mixture was concentrated in vacuo and the crude product was purified by MDAP.
1H NMR (400 MHz, CHLOROFORM-d) δ ppm 7.67 (s, 1 H) 7.52 (s, 2H) 7.43 (s, 5H) 7.19 - 7.25 (m, 2H) 7.09 (s, 1 H) 5.46 (s, 2H) 3.34 - 4.12 (m, 8H). LC-MS Retention Time 2.78mins, ES+ 510.
The following Examples were similarly prepared:
Figure imgf000087_0002
Figure imgf000088_0001
BIOLOGICAL EXAMPLES
Glucocorticoid receptor binding assay
The ability of compounds to bind to the glucocorticoid receptor was determined by assessing their ability to compete with an Alexa 555 fluorescently-labelled dexamethasone derivative. Compounds were solvated and diluted in DMSO, and transferred directly into assay plates. Fluorescent dexamethasone and a partially purified full length glucocorticoid receptor were added to the plates, together with buffer components to stabilise the GR protein and incubated at room temperature for 2 hours in the dark. Binding of each compound was assessed by analysing the displacement of fluorescent ligand by measuring the decrease in fluorescence polarisation signal from the mixture. Dose response curves were constructed from which plC50 values were estimated. Example 1 (racemic), Example 1 Enantiomer 1 , Example 1 Enantiomer 2, Example 2 (racemic), Example 3 (racemic), Example 3 Enantiomer 1 , Example 3 Enantiomer 2, Example 4 (racemic), Example 4 Enantiomer 1, Example 4 Enantiomer 2, Example 5 (racemic), Example 6 (racemic), Example 6 Enantiomer 1 , Example 6 Enantiomer 2, Example 7 (racemic), Example 7 Enantiomer 1 , Example 7 Enantiomer 2, Example 8 (racemic), Example 8 Enantiomer 1 , Example 8 Enantiomer 2, Example 9 (racemic), Example 10 (racemic), Example 11 (racemic), Example 12 (racemic), Example 13 (racemic) and Example 14 (racemic) show glucocorticoid binding with a plC50 > 6 in this assay.
NFKB assay
A549 SPAP cells
Human Caucasian lung carcinoma A549 cell line (ECACC No. 86012804) has been stably transfected in house with a plasmid containing an ELAM promoter sequence that has a NFKB response element within it. Stimulation of the cell line with TNFα results in intracellular signal transduction and ultimately translocation of NFKB into the nucleus. This activates the inserted DNA sequence resulting in transcription of the integrated SPAP gene, which is quantified using a colorimetric assay. In this assay, GR agonist compounds inhibit NFKB driven transcription resulting in a decrease in signal. The stably transfected cell line was grown as a monolayer in DMEM supplemented with FCS-HI (10%), Non-essential amino acids (1 %), L-Glutamine (2mM), Pen/Strep (1%) and Geneticin (50mg/ml).
NFKB agonist assay A 70% confluent T225 flask of A549 SPAP cells was harvested by centrifugation for 5 min at 20Og, resuspended in assay buffer (DMEM supplemented with 10% FCS 2xHI, 2mM L- Glutamine,1 % Pen/Strep and Noπ essential amino acids) and diluted to 0.16 x 106/ml. 60 μl of cell solution was dispensed to each well of clear Nunc 384-well plates, containing compound at the required concentration. Plates were incubated for 1 h at 370C, 95% humidity, 5% CO2 before 10 μl of TNFα was added at final concentration of 3.2ng/ml and then returned to the cell incubator for 15h. Plates were equilibrated to room temperature for 1 h prior to the addition of 25μl of pNPP buffer (1 M Diethanolamine pH 9.8, 0.5mM MgCI2, 0.28M NaCI, 2mg/ml pNPP) to each well of assay plates. The plates were covered to protect the reagents from light, and then incubated at room temperature for approximately 1 hour before reading them on an Ascent using a 405 nm single filter. Dose response curves were constructed from which plC50 values were estimated. The plC50 values for Example 1 (racemic), Example 1 Enantiomer 2, Example 2 (racemic), Example 3 (racemic), Example 3 Enantiomer 2, Example 4 (racemic), Example 4 Enantiomer 1 , Example 4 Enantiomer 2, Example 5 (racemic), Example 5 Enantiomer 1 , Example 5 Enantiomer 2, Example 6 (racemic), Example 6 Enantiomer 1 , Example 6 Enantiomer 2, Example 7 (racemic), Example 7 Enantiomer 1 , Example 7 Enantiomer 2, Example 8 (racemic), Example 8 Enantiomer 2, Example 9 (racemic), Example 10 (racemic), Example 1 1 (racemic), Example 12 (racemic), Example 13 (racemic), Example 14 (racemic) are > 6.5 for the NFkB assay.
MMTV Assay
A549 MMTV cells
Human Caucasian lung carcinoma A549 cell line (ECACC No. 86012804) has been stably transfected in house with a plasmid containing a renilla luciferase reporter with an MMTV promoter. Stimulation of the cell line with GR agonists results in intracellular signal transduction and ultimately translocation of GR into the nucleus. This activates the inserted DNA sequence resulting in transcription of the integrated luciferase gene, which is quantified using a light emission. The stably transfected cell line was grown as a monolayer in DMEM supplemented with FCS-HI (10%), Non-essential amino acids (1 %), L-Glutamine (2mM), Pen/Strep (1%) and Geneticin (50mg/ml).
MMTV Agonist assay
A 90% confluent T175 flask of A549 MMTV cells was harvested by centrifugation for 5 miπ at 20Og, resuspended in assay buffer (DMEM supplemented with 10% FCS 2xHI, 2mM Glutamax, Non essential amino acids and 25mM HEPES) and diluted to 0.1 x 106/ml. 70 μl of cell solution was dispensed to each well of white Nunc 384-well plates, containing compound at the required concentration. Plates were incubated for 6h at 37°C, 95% humidity, 5% CO2. Plates were equilibrated to room temperature for 1 h prior to the addition of 10μl of Renilla substrate to each well of assay plates. The plates were covered to protect the reagents from light, and then incubated at room temperature for approximately 15 minutes before reading them on a Viewlux.
The following examples are full agonists (i.e. have an average maximum asymptote of ≥ 85%) in the NFkB and MMTV agonist assays and have potency in the NFkB assay of > 6.5: Example 1 (racemic), Example 1 Enantiomer 2, Example 2 (racemic), Example 3 (racemic), Example 3 Enantiomer 2, Example 4 (racemic), Example 4 Enantiomer 2, Example 5 (racemic), Example 5 Enantiomer 2, Example 6 (racemic), Example 6 Enantiomer 2, Example 7 (racemate), Example 7 Enantiomer 2, Example 8 Enantiomer 2, Example 9 (racemate), Example 10 (racemic), Example 1 1 (racemic), Example 12 (racemic) and Example 13 (racemic).
Assay for Progesterone Receptor Agonist Activity
A T225 flask of CV-1 cells at a density of 80% confluency was washed with PBS, detached from the flask using 0.25% trypsin and counted using a Sysmex KX-21 N. Cells were diluted in DMEM containing 10% Hyclone, 2mM L-Glutamate and 1 % Pen/Strep at 140 cells/ μl and transduced with 10% PRb-BacMam and 10% MMTV-BacMam. 70 ml of suspension cells were dispensed to each well of white Nunc 384-well plates, containing compounds at the required concentration. After 24h 10 μl of Steadylite were added to each well of the plates. Plates were incubated in the dark for 10 min before reading them on a Viewlux reader. Dose response curves were constructed from which pEC50 values were estimated.
Example 1 (racemic), Example 1 Enantiomer 1 , Example 1 Enantiomer 2, Example 2 (racemic), Example 3 (racemic), Example 3 Enantiomer 1 , Example 3 Enantiomer 2, Example 4 (racemic), Example 4 Enantiomer 1 , Example 4 Enantiomer 2, Example 5 (racemic), Example 5 Enantiomer 1 , Example 5 Enantiomer 2, Example 6 (racemic), Example 6 Enantiomer 1 , Example 6 Enantiomer 2, Example 7 (racemic), Example 7 Enantiomer 1 , Example 7 Enantiomer 2, Example 8 (racemic), Example 8 Enantiomer 1 , Example 8 Enantiomer 2, Example 9 (racemic), Example 10 (racemic), Example 11 (racemic), Example 12 (racemic), Example 13 (racemic) and Example 14 (racemic) show pEC50 <6 in this assay.
In describing examples according to their activity in the assays above, it will be appreciated that at least one isomer, for example, an enantiomer in a mixture of isomers (such as a racemate) has the described activity. The other enantiomer may have similar activity, less activity, no activity or may have some antagonist activity in the case of a functional assay.
Throughout the specification and the claims which follow, unless the context requires otherwise, the word 'comprise1, and variations such as 'comprises' and 'comprising', will be understood to imply the inclusion of a stated integer or step or group of integers but not to the exclusion of any other integer or step or group of integers or steps. The application of which this description and claims forms part may be used as a basis for priority in respect of any subsequent application. The claims of such subsequent application may be directed to any feature or combination of features described herein. They may take the form of product, composition, process, or use claims and may include, by way of example and without limitation, the following claims.
The patents, patent applications and other references described in this application are herein incorporated by reference.

Claims

1. A compound of formula (I):
Figure imgf000093_0001
wherein
R1 is selected from hydrogen, -OCHF2, fluorine and chlorine; R2 and R3 are each independently selected from hydrogen, fluorine, chlorine and bromine; when R1 is -OCHF2, R2 and R3 are each hydrogen, or R2 is chlorine and R3 is hydrogen; n is an integer selected from O1 1 and 2, when n is 1 , X is selected from chlorine and fluorine, and when n is 2, each X is fluorine; or a salt or solvate thereof.
2. A compound according to claim 1 wherein R1 is selected from fluorine and chlorine.
3. A compound according to claim 1 or claim 2 wherein R2 is selected from fluorine and chlorine.
4. A compound according to any one of the preceding claims wherein R3 is hydrogen.
5. A compound according to any one of the preceding claims wherein R1 is fluorine, R2 is chlorine and R3 is hydrogen.
6. A compound according to any one of claims 1 to 4 wherein R1 and R2 are each fluorine and R3 is hydrogen.
7. A compound according to any one of claims 1 to 4 wherein R1 and R2 are each chlorine and R3 is hydrogen.
8. A compound according to any one of the preceding claims wherein n is 1.
9. A compound according to claim 8 wherein X is fluorine.
10 A compound substantially as described in any one of Examples 1 to 14, or a salt or solvate thereof.
11. A compound which is:
5-amino-Λ/-(2-{[({2-[(difluoromethyl)oxy]phenyl}carbonyl)(2-hydroxyethyl)amino]methyl}-
3,3,3-trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1/-/-pyrazole-4-carboxamide;
5-amino-Λ/-(2-{[({2-[(difluoromethyl)oxy]phenyl}carbonyl)(2-hydroxyethyl)amino]methyl}-
3,3,3-trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1/-/-pyrazole-4-carboxamide (Enantiomer 1 );
5-amino-Λ/-(2-{[({2-[(difluoromethyl)oxy]phenyl}carbonyl)(2-hydroxyethyl)amino]methyl}-
3,3,3-trifluoro-2-hydroxypropyl)-1 -(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide
(Enantiomer 2);
5-amino-Λ/-(2-{[[(3-chlorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3-trifluoro-2- hydroxypropyl)-1 -(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide;
5-amino-1-(4-fluorophenyl)-Λ/-(3,3,3-trifluoro-2-{[[(2-fluorophenyl)carbonyl](2- hydroxyethyl)amino]methyl}-2-hydroxypropyl)-1 /-/-pyrazole-4-carboxamide;
5-amino-1-(4-fluorophenyl)-Λ/-(3,3,3-trifluoro-2-{[[(2-fluorophenyl)carbonyl](2- hydroxyethyl)amino]methyl}-2-hydroxypropyl)-1 /-/-pyrazole-4-carboxamide (Enantiomer 1 ); 5-amino-1-(4-fluorophenyl)-Λ/-(3,3,3-trifluoro-2-{[[(2-fluorophenyl)carbonyl](2- hydroxyethyl)amino]methyl}-2-hydroxypropyl)-1 /-/-pyrazole-4-carboxamide (Enantiomer 2);
5-amino-Λ/-(2-{[[(2-chlorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3-trifluoro-2- hydroxypropyl)-1 -(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide;
5-amino-Λ/-(2-{[[(2-chlorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3-trifluoro-2- hydroxypropyl)-1-(4-fluorophenyl)-1H-pyrazole-4-carboxamide (Enantiomer 1 ); 5-amino-Λ/-(2-{[[(2-chlorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3-trifluoro-2- hydroxypropyl)-1 -(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide (Enantiomer 2); 5-amino-Λ/-(2-{[[(2-chloro-6-fluorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,313- trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1W-pyrazole-4-carboxamide; 5-amino-Λ/-(2-{[[(2-chloro-6-fluorophenyl)carbony|](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1/-/-pyrazo!e-4-carboxamide (Enantiomer 1 ); 5-amino-Λ/-(2-{[[(2-chloro-6-fluorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1 -(4-fluorophenyl)-1 H-pyrazole-4-carboxamide (Enantiomer 2); 5-amino-Λ/-(2-{[[(2,6-difluorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3-trifluoro- 2-hydroxypropyl)-1 -(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide;
5-amino-Λ/-(2-{[[(2,6-difluorophenyl)carbony|](2-hydroxyethyl)amino]methyl}-3,3,3-trifluoro- 2-hydroxypropyl)-1 -(4-fluorophenyl)-1 H-pyrazole-4-carboxamide (Enantiomer 1 ); 5-amino-Λ/-(2-{[[(2,6-difluorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3-trifluoro- 2-hydroxypropyl)-1-(4-fluorophenyl)-1H-pyrazole-4-carboxamide (Enantiomer 2); (S)-5-amino-Λ/-(2-{[[(2,6-difluorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1A7-pyrazole-4-carboxamide; 5-amino-Λ/-(2-{[[(2,6-dichlorophenyl)carbony|](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1 H-pyrazole-4-carboxamide; 5-amino-Λ/-(2-{[[(2,6-dichlorophenyl)carbony|](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide (Enantiomer 1 ); 5-amino-Λ/-(2-{[[(2>6-dichlorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1 -(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide (Enantiomer 2); 5-amino-Λ/-(2-{[[(2-chloro-3-fluorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1H-pyrazole-4-carboxamide; 5-amino-Λ/-(2-{[[(2-chloro-3-fluorophenyl)carbony|](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide (Enantiomer 1 ); 5-amino-Λ/-(2-{[[(2-chloro-3-fluorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1 -(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide (Enantiomer 2); 5-amino-1-(4-fluorophenyl)-Λ/-(3,3,3-trifluoro-2-hydroxy-2-{[(2- hydroxyethyl)(phenylcarbonyl)amino]methyl}propyl)-1 /-/-pyrazole-4-carboxamide;
5-amino-Λ/-(2-{[[(2,3-difluorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3-trifluoro- 2-hydroxypropyl)-1-(4-fluorophenyl)-1 /-/-pyrazole-4-carboxamide; 5-amino-Λ/-(2-{[[(3-bromo-2-fluorophenyl)carbonyl](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1H-pyrazole-4-carboxamide; 5-amino-Λ/-(2-{[[(3-chloro-2-fluorophenyl)carbony|](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1H-pyrazole-4-carboxamide; 5-amino-Λ/-(2-{[({2-chloro-6-[(difluoromethyl)oxy]phenyl}carbonyl)(2- hydroxyethyl)amino]methyl}-3,3,3-trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1 H- pyrazole-4-carboxamide;
5-amino-Λ/-(2-{[[(2-bromo-3-fluorophenyl)carbony|](2-hydroxyethyl)amino]methyl}-3,3,3- trifluoro-2-hydroxypropyl)-1-(4-fluorophenyl)-1H-pyrazole-4-carboxamide; or a salt or solvate thereof.
12. A compound as claimed in any one of claims 1 to 1 1 , or a pharmaceutically acceptable salt or solvate thereof, for use in therapy.
13. A compound as claimed in any one of claims 1 to 11 , or a pharmaceutically acceptable salt or solvate thereof, for use in the treatment of patients with an inflammatory and/or allergic condition.
14. A compound as claimed in any one of claims 1 to 11 , or a pharmaceutically acceptable salt or solvate thereof, for use in the treatment of patients with rheumatoid arthritis, asthma, COPD, allergy and/or rhinitis.
15. A compound as claimed in any one of claims 1 to 11 , or a pharmaceutically acceptable salt or solvate thereof, for use in the treatment of patients with skin disease.
16. A compound as claimed in any one of claims 1 to 11 , or a pharmaceutically acceptable salt or solvate thereof, for use in the treatment of patients with eczema, psoriasis, allergic dermatitis, neurodermatitis, pruritis and/or hypersensitivity reactions.
17. Use of a compound as claimed in any one of claims 1 to 11 , or a pharmaceutically acceptable salt or solvate thereof, for the manufacture of a medicament for the treatment of patients with an inflammatory and/or allergic condition.
18. Use of a compound as claimed in any one of claims 1 to 11 , or a pharmaceutically acceptable salt or solvate thereof, for the manufacture of a medicament for the treatment of patients with rheumatoid arthritis, asthma, COPD, allergy and/or rhinitis.
19. Use of a compound as claimed in any one of claims 1 to 11 , or a pharmaceutically acceptable salt or solvate thereof, for the manufacture of a medicament for the treatment of patients with skin disease.
20. Use of a compound as claimed in any one of claims 1 to 1 1 , or a pharmaceutically acceptable salt or solvate thereof, for the manufacture of a medicament for the treatment of patients with eczema, psoriasis, allergic dermatitis, neurodermatitis, pruritis and/or hypersensitivity reactions.
21. A method for the treatment of a human or animal subject with an inflammatory and/or allergic condition which method comprises administering to said human or animal subject an effective amount of a compound of formula (I) or a pharmaceutically acceptable salt or solvate thereof.
22. A method for the treatment of a human or animal subject with rheumatoid arthritis, asthma, COPD, allergy and/or rhinitis which method comprises administering to said human or animal subject an effective amount of a compound of formula (I) or a pharmaceutically acceptable salt or solvate thereof.
23. A method for the treatment of a human or animal subject with skin disease which method comprises administering to said human or animal subject an effective amount of a compound of formula (I) or a pharmaceutically acceptable salt or solvate thereof.
24. A method for the treatment of a human or animal subject with eczema, psoriasis, allergic dermatitis, neurodermatitis, pruritis and/or hypersensitivity reactions, which method comprises administering to said human or animal subject an effective amount of a compound of formula (I) or a pharmaceutically acceptable salt or solvate thereof.
25. A pharmaceutical composition comprising a compound of formula (I) as claimed in any one of claims 1 to 11 , or a pharmaceutically acceptable salt or solvate thereof, in admixture with one or more physiologically acceptable diluents or carriers.
26. A process for the preparation of a compound of formula (I) as claimed in any one of claims 1 to 11 , or a salt or solvate thereof, comprising reaction of an amine of formula
(II)
Figure imgf000098_0001
wherein X and n are as defined in claim 1 and R4 is hydrogen or a protecting group, with a compound of formula (III)
Figure imgf000098_0002
wherein R , R and R are as defined in claim 1 and Y is chlorine or hydroxy.
27. A compound of formula (II)
Figure imgf000098_0003
(H) wherein X and n are as defined in claim 1 and R4 is hydrogen or a protecting group.
28. A compound of formula (III)
Figure imgf000099_0001
wherein R , R and R are as defined in claim 1 and Y is chlorine or hydroxy.
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