Field of the Invention

This invention relates to novel compounds which are inhibitors of cytokines and possess anti-inflammatory properties as well as utility in reducing pain in pain conditions where cytokines are involved. Background of the Invention

Immune-driven inflammatory events are a significant cause of many chronic inflammatory diseases where prolonged inflammation causes tissue destruction and results in extensive damage and eventual failure of the effected organ. The cause of these diseases is unknown, so are often called autoimmune, as they appear to originate from an individual's immune system turning on itself. Conditions include those involving multiple organs, such as systemic lupus erythematosus (SLE) and scleroderma. Other types of autoimmune disease can involve specific tissues or organs such as the musculoskeletal tissue (rheumatoid arthritis, ankylosing spondylitis), gastrointestinal tract, (Crohn's disease and ulcerative colitis), the central nervous system (Alzheimer's, multiple sclerosis, motor neurone disease, Parkinson's disease and chronic fatigue syndrome), pancreatic beta cells (insulin dependent diabetes mellitus), the adrenal gland (Addison's disease), the kidney (Goodpasture's syndrome, IgA nephropathy, interstitial nephritis), exocrine glands (Sjogren's syndrome and autoimmune pancreatitis) and skin (psoriasis and atopic dermatitis).

In addition, there are chronic inflammatory diseases whose aetiology is more or less known but whose inflammation is also chronic and unremitting. These also exhibit massive tissue/organ destruction and include conditions such as osteoarthritis. These conditions are a major cause of illness in the developing world and poorly treated by current therapies.

Inflammation of skin structures (dermatitis) is a common set of conditions. These diseases are treated using a wide array of therapies, many of which have very severe side-effects.

Current disease-modifying treatments (if any) for immune-driven conditions include neutralising antibodies, cytotoxics, corticosteriods, immunosuppressants, antihistamines and antimuscarinics. These treatments are often associated with inconvenient routes of administration and severe side- effects, leading to compliance issues. Moreover, certain drug classes are only effective for certain types of inflammatory diseases, e.g. antihistamines for rhinitis. Summary of the Invention

It has been found that compounds of formula (I) are inhibitors of cytokines and possess anti-inflammatory properties as well as utility in reducing pain in pain conditions where cytokines are involved. According to the present invention, an inflammatory condition as previously described is treated by the use of such compounds. According to another aspect of the invention, pain such as acute, chronic or neuropathic pain (including, but not limited to, pain associated with cancer, surgery, arthritis, dental surgery, painful neuropathies, trauma, musculoskeletal injury or disease, and visceral diseases) and migraine headache in mammals, can be treated by the use of compounds of the formula

wherein:

Ri is aryl or heteroaryl optionally substituted with R ₅ , R ₂ is H, alkyl, cycloalkyl or CH ₂ OH (and can form a ring with R ₃ );

R ₃ is H, alkyl, cycloalkyl or CH ₂ OH (and can form a ring with R ₂ ); R ₄ is H or alkyl;

R ₅ is alkyl, CF ₃ , OR ₆ , OCOR ₆ , CONH ₂ , CN, Cl, Br, I, N(R ₆ ) ₂ , NO ₂ , NHCHO, NHCONH ₂ , NHSO ₂ R ₆ , CON(R ₆ ) ₂ , S(O) _n R ₆ , CH ₂ OH or OCON(R ₆ ) ₂ ; R ₆ is H, alkyl or cycloalkyl; and n is 0-2; or a salt thereof. Description of the Invention

For the purposes of this specification, aryl is typically phenyl or naphthyl, heteroaryl is typically aryl comprising 1 to 3 heteroatoms selected from O, NH and S, alkyl has 1 to 8 C atoms and cycloalkyl has 3 to 6 C atoms. The compounds of formula (I) can be used to treat inflammatory diseases including, for example, autoimmune diseases involving multiple organs, such as systemic lupus erythematosus (SLE) and scleroderma, specific tissues or organs such as the musculoskeletal tissue (rheumatoid arthritis, ankylosing spondylitis), gastro-intestinal tract' (Crohn's disease and ulcerative colitis), the central nervous system (Alzheimers, multiple sclerosis, motor neurone disease, Parkinson's disease and chronic fatigue syndrome), pancreatic beta cells (insulin dependent diabetes mellitus), the adrenal gland (Addison's disease), the kidney (Goodpasture's syndrome, IgA nephropathy, interstitial nephritis), exocrine glands (Sjogren's syndrome and autoimmune pancreatitis) and skin (psoriasis and atopic dermatitis), chronic inflammatory diseases such as osteoarthritis, periodontal disease, diabetic nephropathy, diabetic ulceration, retinopathy, chronic obstructive pulmonary disease, artherosclerosis, graft versus host disease, chronic pelvic inflammatory disease, endometriosis, chronic hepatitis and tuberculosis, IgE-mediated (Type I) hypersensitivities such as rhinitis, asthma, anaphylaxis, dermatitis and ophthalmic diseases. Dermatitis conditions include: actinic keratosis, acne rosacea, acne vulgaris, allergic contact dermatitis, angioedema, atopic dermatitis, bullous pemiphigoid, cutaneous drug reactions, erythema multiforme, lupus erythematosus, photodermatitis, psoriasis, psoriatic arthritis, scleroderma and urticaria. Ophthalmic diseases include age-related macular degeneration (ARMD), dry eye, uveitis and glaucoma.

Compounds of formula (1) may be used according to the invention when the patient is also administered or in combination with another therapeutic agent selected from corticosteroids (examples including Cortisol, cortisone, hydrocortisone, dihydrocortisone, fludrocortisone, prednisone, prednisolone, deflazacort, flunisolide, beconase, methylprednisolone, triamcinolone, betamethasone, and dexamethasone), disease modifying anti-rheumatic drugs (DMARDs) (examples including azulfidine, aurothiomalate, bucillamine, chlorambucil, cyclophosphamide, leflunomide, methotrexate, mizoribine, penicillamine and sulphasalazine), immunosuppressants (examples including azathioprine, cyclosporin, mycophenolate), COX inhibitors (examples including aceclofenac, acemetacin, alcofenac, alminoprofen, aloxipirin, amfenac, aminophenazone, antraphenine, aspirin, azapropazone, benorilate, benoxaprofen, benzydamine, butibufen, celecoxib, chlorthenoxacine, choline salicylate, chlometacin, dexketoprofen, diclofenac, diflunisal, emorfazone, epirizole, etodolac, feclobuzone, felbinac, fenbufen, fenclofenac, flurbiprofen, glafenine, hydroxylethyl salicylate, ibuprofen, indomethacin, indoprofen, ketoprofen, ketorolac, lactyl phenetidin, loxoprofen, mefenamic acid, metamizole, mofebutazone, mofezolac, nabumetone, naproxen, nifenazone, oxametacin, phenacetin, pipebuzone, pranoprofen, propyphenazone, proquazone, rofecoxib, salicylamide, salsalate, sulindac, suprofen, tiaramide, tinoridine, tolfenamic acid and zomepirac), neutralising antibodies (examples including, etanercept and infliximab), and antibiotics (examples including doxycycline and minocycline). Compounds of formula (I) exhibit analgesic activity in animal models.

The activity of these compounds may be determined by the use of the appropriate in vivo assay.

This invention also relates to a method of treatment for patients (including man and/or mammalian animals raised in the dairy, meat or fur industries or as pets) suffering from chronic, acute or neuropathic pain; and more specifically, a method of treatment involving the administration of the analgesic of formula (I) as the active constituent.

Accordingly, the compounds of formula (I) can be used among other things in the treatment of pain conditions such as acute and chronic pain (as well as, but not limited to, pain associated with cancer, surgery, arthritis, dental surgery, trauma, musculo-skeletal injury or disease, visceral diseases, painful bladder syndrome) and migraine headache. Additionally the painful conditions can be neuropathic (post-herpetic neuralgia, diabetic neuropathy, drug induced neuropathy, HIV mediated neuropathy, sympathetic reflex dystrophy or causalgia, fibromyalgia, myofacial pain, entrapment neuropathy, phantom limb pain, trigeminal neuralgia). Neuropathic conditions include central pain related to stroke, multiple sclerosis, spinal cord injury, arachnoiditis, neoplasms, syringomyelia, Parkinson's and epilepsia.

It will often be advantageous to use compounds of formula (I) in combination with another drug used for pain therapy. Such another drug may be an opiate or a non-opiate such as baclofen. Especially for the treatment of neuropathic pain, coadministration with gabapentin is preferred. Other compounds that may be used include acetaminophen, a non-steroidal antiinflammatory drug, a narcotic analgesic, a local anaesthetic, an NMDA antagonist, a neuroleptic agent, an anticonvulsant, an anti-spasmodic, an antidepressant or a muscle relaxant.

Compounds of the general formula (1) may be prepared by any suitable method known in the art and/or by the processes described below. It will be appreciated that where a particular stereoisomer of formula (1) is required, the synthetic processes described herein may be used with the appropriate homochiral starting material and/or isomers maybe resolved from mixtures using conventional separation techniques (e.g. HPLC).

Compounds (1) may be prepared by any suitable method known in the art and/or by the following process. It will be appreciated that functional groups, such as amino, hydroxyl or carboxyl groups, present in the various compounds described, and which it is desired to retain, may need to be in protected form before any reaction is initiated. In such instances, removal of the protecting group may be the final step in a particular reaction. Suitable protecting groups for such functionality will be apparent to those skilled in the art. For specific details see "Protective Groups in Organic Synthesis", Wiley Interscience, T W Greene, PGM Wuts.

A process for preparing compounds of general formula (1) comprises reacting a compound of formula (2) where X is a leaving group with the corresponding aminoalcohol, followed by cyclisation The aminoalcohols and the carbonyl derivatives are either commercially available or readily obtained from commercially available materials by people who are skilled in the art of synthetic organic chemistry. This procedure is shown in the following Scheme.

Any mixtures of final products or intermediates obtained can be separated on the basis of the physico-chemical differences of the constituents, in known manner, into the pure final products or intermediates, for example by chromatography, distillation, fractional crystallization, or by formation of a salt if appropriate or possible under the circumstances.

Any suitable route of administration can be used. For example, any of oral, topical, parenteral, ocular, rectal, vaginal, inhalation, buccal, sublingual and intranasal delivery routes may be suitable. The pharmaceutical composition containing the active ingredient may be in a form suitable for oral use, for example, as tablets, troches, lozenges, aqueous or oily suspensions, dispersible powders or granules, emulsions, hard or soft capsules, or syrups or elixirs. The composition may be in immediate or controlled release form. Compositions intended for oral use may be prepared according to any method known to the art for the manufacture of pharmaceutical compositions and such compositions may contain one or more agents selected from the group consisting of sweetening agents, flavouring agents, colouring agents and preserving agents in order to provide pharmaceutically elegant and palatable preparations. Tablets contain the active ingredient in admixture with non-toxic pharmaceutically acceptable excipients which are suitable for the manufacture of tablets. These excipients may be for example, inert diluents, such as calcium carbonate, sodium carbonate, lactose, calcium phosphate or sodium phosphate; granulating and disintegrating agents, for example corn starch, or alginic acid; binding agents, for example starch, gelatin or acacia, and lubricating agents, for example magnesium stearate, stearic acid or talc. The tablets may be uncoated or they may be coated by known techniques to delay disintegration and absorption in the gastointestinal tract and thereby provide a sustained action over a longer period. For example, a time delay material such as glyceryl monostearate or glyeryl distearate may be employed. They may also be coated by the techniques described in the US4256108, US4166452 and US4265874, to form osmotic therapeutic tablets for control release.

Formulations for oral use may also be presented as hard gelatin capsules where in the active ingredient is mixed with an inert solid diluent, for example calcium carbonate, calcium phosphate or kaolin, or as soft gelatin capsules wherein the active ingredient is mixed with water or an oil medium, for example peanut oil, liquid paraffin or olive oil.

Aqueous suspensions contain the active materials in admixture with excipients suitable for the manufacture of aqueous suspensions. Such excipients are suspending agents, for example sodium carboxymethylcellulose, methylcellulose, hydroxypropylmethylcellulose, sodium alginate polyvinyl- pyrrolidone, gum tragacanth and gum acacia; dispersing or wetting agents may be a naturally occuring phosphatide, for example lecithin, or condensation products of an alkylene oxide with fatty acids, for example polyoxyethylene stearate, or condensation products of ethylene oxide with long chain aliphatic alcohols, for example heptadecaethyleneoxycetanol, or condensation products of ethylene oxide with partial esters dervied from fatty acids and a hexitol such a polyoxyethylene with partial esters derived from fatty acids and hexitol anhydrides, for example polyoxyethylene sorbitan monooleate. The aqueous suspensions may also contain one or more preservatives, for example ethyl or n-propyl p-hydroxybenzoate, one or more colouring agents, one or more flavouring agents, and one or more sweetening agents, such as sucrose or saccharin.

Oily suspensions may be formulated by suspending the active ingredient in a vegetable oil, for example arachis oil, olive oil, sesame oil or coconut oil, or in a mineral oil such as liquid paraffin. The oily suspensions may contain a thickening agent, for example beeswax, hard paraffin or cetyl alcohol. Sweetening agents such as those set forth above, and flavouring agents may be added to provide a palatable oral preparation. These compositions may be preserved by the addition of an anti-oxidant such as ascorbic acid. Dispersible powders and granules suitable for preparation of an aqueous suspension by the addition of water provide the active ingredient in admixture with a dispersing or wetting agent, suspending agent and one or more preservatives. Suitable dispersing or wetting agents and suspending agents are exemplified, for example sweetening, flavouring and colouring agents, may also be present.

The pharmaceutical compositions of the invention may also be in the form of oil-in-water emulsions. The oily phase may be a vegetable oil, for example olive oil or arachis oil, or a mineral oil, for example liquid paraffin or mixtures of these. Suitable emulsifying agents may be naturally-occuring gums, for example gum acacia or gum tragacanth, naturally-occuring phosphatides, for example soya bean, lecithin, and esters or partial esters derived from fatty acids and hexitol anhydrides, for example sorbitan monooleate and condensation products of the said partial esters with ethylene oxide, for example polyoxyethylene sorbitan monooleate. The emulsions may also contain sweetening and flavouring agents.

Syrups and elixirs may be formulated with sweetening agents, for example gycerol, propylene glycol, sorbitol or sucrose. Such formulations may also contain a demulcent, a preservative and flavouring and colouring agents. The pharmaceutical compositions may be in the form of a sterile injectable aqueous or oleagenous suspension. This suspension may be formulated according to the known art using those suitable dispersing or wetting agents and suspending agents which have been mentioned above. The sterile injectable preparation may also be in a sterile injectable solution or suspension in a nontoxic parenterally-acceptable diluent or solvent, for example as a solution in 1 ,3- butanediol. Among the acceptable vehicles and solvents that may be employed are water, Ringer's solution and isotonic sodium chloride solution. In addition, sterile, fixed oils are conventionally employed as a solvent or suspending medium. For this purpose any bland fixed oil may be employed including synthetic mono- or diglyce rides. In addition, fatty acids such as oleic acid find use in the preparation of injectables.

The compounds of formula (1) may also be administered in the form of suppositories for rectal administration of the drug. These compositions can be prepared by mixing the drug with a suitable non-irritating excipient which is solid at ordinary temperatures but liquid at the rectal temperature and will therefore melt in the rectum to release the drug. Such materials are cocoa butter and polyethylene glycols.

For topical use, creams, ointments, jellies, solutions or suspensions, etc containing the compounds of Formula (1) are employed. For the purposes of this application, topical application includes mouth washes and gargles.

The amount of active ingredient that may be combined with the carrier materials to produce a single dosage form will vary depending upon the host treated and the particular mode of administration. For example, a formulation intended for the oral administration of humans may vary from about 5 to about 95 percent of the total composition. Dosage unit forms will generally contain between from about 1 mg to about 500 mg of an active ingredient.

It will be understood, however, that the specific dose level for any particular patient will depend upon a variety of factors including the activity of the specific compound employed, the age, body weight, general health, sex, diet time of administration, route of administration, rate of excretion, drug combination and the severity of the particular disease undergoing therapy.

The following Examples illustrate the preparation of compounds of the invention, following the scheme shown above, wherein Ri is 4-amino-3,5- dichlorophenyl, X is Br, R ₂ , R ₃ and R ₇ are each CH ₃ and R ₄ is H. 4-Amino-3,5-dichloro-bromoacetophenone (2)

Bromine (5 ml, 98 mmol) was added to a mixture of 4-amino-3,5- dichloroacetophenone (1) (20 g, 98 mmol) in CHCI ₃ (240 ml) at room temperature. The mixture was stirred for 1 h then EtOH (40 ml) was added. The mixture was cooled to 0 ⁰ C and stirred for 1 h. The precipitate was filtered and air-dried (20.4 g, 72%). ¹ H NMR (400 MHz, DMSO): 4.77 (2H, s), 6.61 (2H, bs), 7.86 (2H, s); ¹³ C NMR (100 MHz, DMSO): 63.39, 117.89, 128.57, 129.75, 146.17, 195.99. N-methyl-2-amino-2-methyl-propan-1 -ol (4)

2-Amino-2-methylpropanol (3) (10 g, 0.11 mol), Na ₂ CO ₃ (0.41 g), NaHCO ₃ (0.41 g) and 3:1 dioxane/water (41 ml) were stirred together at 0 ⁰ C. Boc anhydride (29.4 g, 0.13 mol) was added slowly and the reaction was stirred at room temperature for 6 h. The salts formed were filtered and washed with DCM. The filtrate was diluted with water (50ml) and extracted with DCM (3 x 50 ml), dried (Na ₂ SO ₄ ) and evaporated to dryness, giving a white solid. This was dissolved in THF (100 ml) and added dropwise to a mixture of lithium aluminium hydride (12.7 g, 0.33 mol) in THF (200 ml). The mixture was heated to reflux overnight then cooled to room temperature. Water (15 ml) was added slowly with ice-cooling followed by 4 M NaOH (15 ml), and then more water (45 ml). The white solids were filtered, washed with THF, and the filtrate was evaporated to dryness, giving a pink solid (8.9 g, 77%). ¹ H NMR (400 MHz, CDCI ₃ ): 0.97 (6H, s), 2.24 (3H, s), 3.26 (2H, s); ¹³ C NMR (100 MHz, CDCI ₃ ): 23.27, 28.41 , 53.62, 68.26.

2-(4-Amino-3,5-dichlorophenyl)-4,5,5-trimethylmorpholin-2 -ol (5)

Λ/-Methyl-2-amino-2-methyl-propan-1-ol (4) (4.6 g, 44.6 mmol) was added to a mixture of compound 2 (4.2 g, 14.8 mmol) in chloroform (17 ml). The mixture was stirred at room temperature for 2 h, and then water (9 ml) was added. The mixture was stirred for 1 h, and then the solid was filtered. The solid was triturated with water (1 L) to give the desired compound (1.2 g, 91 %). ¹ H NMR (400 MHz, DMSO): 0.94 (6H, s), 3.18 (2H, d J=4.4 Hz), 3,93 (2H, s), 4.55 (1 H, m), 6.40 (2H, s), 7.84 (2H, s); ¹³ C NMR (100 MHz ₁ DMSO): 24.21 , 48.87, 53.73, 68.52, 117.92, 124.57, 125.79, 128.62, 146.07, 195.30; LC-MS: 291 , 292, 293 (M + H ⁺ ).

The following illustrates the utility of compounds of the invention, by evaluation of the above compound (the "test compound") in a mouse LPS cytokine assay. Group size: n=8 Protocol and Study Design:

At the end of the acclimatization period, non-fasted mice are weighed, individually identified on the tail with a permanent marker and administered by oral route with either vehicle or test compound in a volume of 10 ml/kg adapted to the body weight. Mice receive an intraperitoneal (i.p.) injection of 1 mg/kg LPS in a volume of 5 ml/kg of body weight.

2 hours after LPS challenge, blood samples are collected into tubes with anticoagulant by cardiac puncture, under gaseous anesthesia (isoflurane).

Samples are put on ice until they are prepared by centrifugation (6000 x g for 5 min at 4°C), aliquoted and stored at -20 ⁰ C until use.

TNFα levels are measured in duplicate by ELISA technique in plasma samples. Data are reported as IL-10 and TNFα (pg/mL), mean ± S. E. M.

(standard error of the mean).

% effect are calculated according to the following formula: (Mean test or reference substance - Mean vehicle)/Mean vehicle x 100.

Inter-group deviations are statistically analyzed by a one-way analysis of variance (ANOVA). In the case of significant difference in the mean values among the different levels of treatment, comparisons versus the vehicle group are carried out using Dunnett's test. In case the equal variance test fails, a Kruskal-Wallis one-way analysis of variance on ranks followed by a Dunn's test is proposed. p< 0.05 is considered statistically significant. Drawings

Results of the preceding study are shown in the accompanying drawings. In these drawings:

Figure 1 is a diagram showing the effect of vehicle and the test compound on LPS-induced TNFα plasma levels in mice (pg/ml; x axis); and Figure 2 is a diagram showing the effect of vehicle and the test compound on LPS-induced IL-10 plasma levels in mice (pg/ml; x axis).