WO2007022937A1 - Pyridazine derivatives with antiinflammatory activity - Google Patents

Abstract

The present invention relates to novel pyridazinyl derivatives, pharmaceutical compositions containing these compounds and their use in the treatment of diseases, particularly pain, which diseases are caused directly or indirectly by an increase or decrease in activity of the cannabinoid receptor.

Description

PYRIDAZINE DERIVATIVES WITH ANTIINFLAMMATORY ACTIVITY

The present invention relates to novel pyπdazinyl derivatives, pharmaceutical compositions containing these compounds and their use in the treatment of diseases, particularly pam, which diseases are caused directly or indirectly by an increase or decrease in activity of the cannabinoid receptor.

Cannabinoids are a specific class of psychoactive compounds present in Indian cannabis {Cannabis sativa), including about sixty different molecules, the most representative being cannabinol, cannabidiol and several isomers of tetrahydrocannabinol. Knowledge of the therapeutic activity of cannabis dates back to the ancient dynasties of China, where, 5,000 years ago, cannabis was used for the treatment of asthma, migraine and some gynaecological disorders. These uses later became so established that, around 1850, cannabis extracts were included in the US Pharmacopaeia and remained there until 1947.

Cannabinoids are known to cause different effects on various systems and/or organs, the most important being on the central nervous system and on the cardiovascular system. These effects include alterations in memory and cognition, euphoπa, and sedation. Cannabinoids also increase heart rate and vary systemic arterial pressure. Peripheral effects related to bronchial constriction, immunomodulation, and inflammation have also been observed. The capability of cannabinoids to reduce intraocular pressure and to affect respiratory and endocrine systems is also well documented. See e.g. L.E. Holhster, Health Aspects of Cannabis, Pharmacological Reviews,

Vol. 38, pp. 1-20, (1986). More recently, it was found that cannabinoids suppress the cellular and humoral immune responses and exhibit antiinflammatory properties. Wirth et al.,

Antiinflammatory Properties of Cannabichrome, Life Science, Vol. 26, pp. 1991-1995, (1980).

In spite of the foregoing benefits, the therapeutic use of cannabis is controversial, both due to its relevant psychoactive effects (causing dependence and addiction), and due to manifold side effects that have not yet been completely clarified. Although work in this field has been ongoing since the 1940's, evidence indicating that the peπpheral effects of cannabinoids are directly mediated, and not secondary to a CNS effect, has been limited by the lack of receptor characteπzation, the lack of information concerning an endogenous cannabinoid ligand and, until recently, the lack of receptor subtype selective compounds.

The first cannabinoid receptor was found to be mainly located in the brain, in neural cell lines, and, only to a lesser extent, at the peπpheral level. In view of its location, it was called the central receptor ("CBl"). See Matsuda et al., "Structure of a Cannabinoid Receptor and Functional Expression of the Cloned cDNA," Nature, Vol. 346, pp. 561-564 (1990). The second cannabinoid receptor ("CB2") was identified in the spleen, and was assumed to modulate the non psychoactive effects of the cannabinoids. See Munro et el., "Molecular Characteπzation of a Peripheral Receptor for Cannabinoids," Nature, Vol. 365, pp. 61-65 (1993).

The foregoing indications and the preferential localization of the CB2 receptor m the immune system confirms a specific role of CB2 in modulating the immune and antiinflammatory response to stimuli of different sources.

The total size of the patient population suffeπng from pain is vast (almost 300 million), dominated by those suffering from back pam, osteo-arthπtic pain and post-operative pain. Neuropathic pain (associated with neuronal lesions such as those induced by diabetes, HIV, herpes infection, or stroke) occurs with lower, but still substantial prevalence, as does cancer pain.

The pathogenic mechanisms that give rise to pain symptoms can be grouped into two main categories: - those that are components of inflammatory tissue responses (Inflammatory Pain);

- those that result from a neuronal lesion of some form (Neuropathic Pain).

Chronic inflammatory pain consists predominantly of osteoarthritis, chronic low back pain and rheumatoid arthritis. The pain results from acute and on-going injury and/or inflammation. There may be both spontaneous and provoked pain. There is an underlying pathological hypersensitivity as a result of physiological hyperexcitabihty and the release of inflammatory mediators which further potentiate this hyperexcitabihty. CB2 receptors are expressed on inflammatory cells (T cells, B cells, macrophages, mast cells) and mediate immune suppression through inhibition of cellular interaction/ inflammatory mediator release. CB2 receptors may also be expressed on sensory nerve terminals and therefore directly inhibit hyperalgesia.

More recently, data suggests a role for CB2 receptor activation in the CNS. Until recently the CB2 receptor was thought to be restricted to the periphery, however emerging data suggests inflammatory pain-mediated induction of CB2 receptor expression in rat spinal cord which coincides with the appearance of activated microglia (Zhang et. al., 2003). Furthermore CB2 receptor agonists have been shown to reduce mechanically evoked responses and wind-up of wide dynamic range neurones in spinal cord dorsal horn in animal models of inflammatory pain (Zhang et. al., 2003, Eur J. Neurosci. 17: 2750-2754, Nackley et. al., 2004, J. Neurophys. 92: 3562-3574, Elmes et. al., 2004, Eur. J. Neurosci. 20: 2311-2320.)

The role of CB2 in immunomodulation, inflammation, osteoporosis, cardiovascular, renal and other disease conditions is now being examined.

Based on the foregoing, there is a need for compounds which have activity against the CB2 receptor. Thus, CB2 modulators are believed to offer a unique approach toward the pharmacotherapy of immune disorders, inflammation, osteoporosis, renal ischemia and other pathophysiological conditions. WO 04/018433, WO 04/018434, WO04/029027 and WO04/029026 (all m the name of

Glaxo Group Limited) descπbe pyπmidine and pyridine deπvatives useful in the treatment of diseases which are caused directly or indirectly by an increase or decrease in activity of the cannabinoid receptor.

The present invention provides novel pyridazinyl deπvatives of formula (I) and pharmaceutically acceptable denvatives thereof, pharmaceutical compositions containing these compounds or derivatives, and their use as CB2 receptor modulators, which are useful in the treatment of a variety of disorders.

The present invention further comprises a method for treating disease mediated by CB2 receptors in an animal, including humans, which comprises administering to an animal in need thereof an effective, non toxic, amount of a compound of formula (I) or a pharmaceutically acceptable deπvative thereof.

In light of the fact that cannabinoids act on receptors capable of modulating different functional effects, and in view of the low homology between CB2 and CBl, a class of drugs selective for the specific receptor sub-type is desirable. The natural or synthetic cannabmoids currently available do not fulfil this function because they are active on both receptors.

In one embodiment the present invention includes compounds which are capable of selectively modulating the receptors for cannabmoids and therefore the pathologies associated with such receptors.

The invention provides compounds of formula (I):

(I) wherein

Ar represents qumolmyl, lsoquinohnyl, either of which can be unsubstituted or substituted with 1, 2 or 3 substituents selected from halo, Ci_₆alkyl, C_1-6alkoxy, halosubstituted Ci_-6alkyl, halosubstitued C,.₆alkoxy, NH₂, -SO₂Ci.₆alkyl, -NHSO₂Ci_-6alkyl, -C, .₆ alkylNR³R⁴, or Ar represents phenyl substituted with 1, 2 or 3 substituents at the 2, 3, 5 or 6 positions, which substituents are selected from halo, Ci_₆alkyl, C_1-6alkoxy, halosubstituted d.₆alkyl, halosubstitued C,.₆alkoxy, NH₂, -SO₂C,.₆alkyl, -NHSO₂C,.₆alkyl, -C_{1 -6}alkylNR³R⁴;

R¹ and R² together with the nitrogen to which they are attached form a 5- to 8- membered heterocyclyl or a 5-membered heteroaryl ring which can be unsubstituted or substituted with 1, 2 or 3 substituents selected from halo, C_1-6 alkyl, cyano, -C₁ _₆ alkylNR³R⁴;

R³ and R⁴ are independently selected from hydrogen, C_1-6 alkyl or together with the N to which they are attached form a pyrrohdinyl, pφeπdinyl or morpholinyl ring; and pharmaceutically acceptable derivatives thereof.

When R¹ and R² taken together with the N to which they are attached form an optionally substituted 5- to 8- membered heterocyclyl πng the ring may optionally contain 1, 2, 3 or 4 further hetero atoms. The nng may be saturated or unsaturated. In one embodiment the further hetero atoms are selected from oxygen, nitrogen or sulphur. Examples of a 5-membered heterocyclyl ring are pyrrohdinyl, 2,5,dihydro-lH-pyrrolyl, pyrazolidinyl, lmidazolidinyl. Examples of 6-membered heterocyclyl πngs are morpholmyl, piperazmyl, pipeπdmyl, tetrahydropyridinyl, thiomorpholme- 1,1 -dioxide, thiomorpholinyl and thiomorpholinyl-1 -oxide. Examples of a 7- membered heterocyclyl πng are hexahydro-lH-azepmyl or hexahydro-l,4-oxazepinyl. Examples of 8- membered heterocyclyl rings are 3-oxa-8-azabicyclo[3.2.1]octanyl, azacyclooctanyl, azaoxacyclooctanyl or azathiacyclooctanyl.

When R¹ and R² taken together with the N to which they are attached form an optionally substituted 5-membered heteroaryl πng the πng may optionally contain 1, 2 or 3 further hetero atoms. In one embodiment the further hetero atoms are selected from oxygen, nitrogen or sulphur. Examples of a 5-membered heteroaryl πng are pyrrolyl, pyrazolyl or tπazolylyl.

In one embodiment the substituents for Ar are selected from halo, C_1-6 alkyl, C^alkoxy, halosubstituted C_1-6 alkyl, halosubstituted Ci_-6alkoxy, -SO₂C,.₆alkyl, -NHSO₂Ci_-6alkyl and NH₂ In another embodiment the substituents for Ar are selected from halo, C_1-6 alkyl, halosubstituted Ci- ₆alkyl and NH₂. In one embodiment R¹ and R² together with the nitrogen to which they are attached form a pyrrolidinyl, pyrrolyl, 2,5,dihydro-lH-pyrrolyl, morpholinyl, thiomorpholinyl, piperidinyl, pyrazolyl, hexahydro-lH-azepinyl, hexahydro-l,4-oxazepinyl or 3-oxa-8-azabicyclo[3.2.1]octanyl, any of which can be unsubstituted or substituted with 1, 2 or 3 substituents selected from halo, C).₆ alkyl, ethylpypyrrolidinyl, or cyano. In another embodiment R¹ and R² together with the nitrogen to which they are attached form a pyrrolidinyl, pyrrolyl, 2,5,dihydro-lH-pyrrolyl, morpholinyl, piperidinyl, hexahydro-lH-azepinyl, hexahydro-l,4-oxazepinyl or 3-oxa-8- azabicyclo[3.2.1]octanyl, any of which can be unsubstituted or substituted with 1, 2 or 3 substituents selected from halo, Ci .6 alkyl, ethylpypyrrolidinyl, or cyano.

In one embodiment R³ and R⁴ are independently selected from hydrogen, methyl or together with the N to which they are attached form a pyrolidinyl, piperidinyl or morpholinyl ring. In one embodiment R¹ and R² together with the nitrogen to which they are attached form a pyrrolidinyl, 2,5,dihydro-lH-pyrrolyl, pyrazolidinyl, imidazolidinyl, morpholinyl, piperazinyl, piperidinyl, tetrahydropyridinyl, thiomorpholine- 1 , 1 -dioxide, thiomorpholinyl, thiomorpholinyl- 1 - oxide, hexahydro-lH-azepinyl, hexahydro-l,4-oxazepinyl, 3-oxa-8-azabicyclo[3.2.1]octanyl, azacyclooctanyl, azaoxacyclooctanyl, azathiacyclooctanyl, pyrrolyl or pyrazolyl ring any of which can be unsubstituted or substituted with 1, 2 or 3 substituents selected from halo, Ci_-6 alkyl, methyl, cyano, and ethylpiperidinyl, Ci .₆ alkyl substituted with a NR³R⁴ wherein R³ and R⁴ are independently selected from hydrogen, C].₆ alkyl or together form a pyrolidinyl, piperidinyl or morpholinyl ring.

In certain embodiments compounds of formula (I) show selectivity for CB2 over CB 1. In one embodiment compounds of formula (I) have a pEC50 value at the cloned human cannabinoid CB2 receptor of at least about 2 units higher than the pEC50 values at the cloned human cannabinoid CBl receptor and/or have less than 30% efficacy at the CBl receptor.

The invention is described using the following definitions unless otherwise indicated. The term "pharmaceutically acceptable derivative" means any pharmaceutically acceptable salt, ester, salt of such ester or solvate (including solvates of salts, esters, or salts of esters) of the compounds of formula (I), or any other compound which upon administration to the recipient is capable of providing (directly or indirectly) a compound of formula (I) or an active metabolite or residue thereof. In one embodiment the pharmaceutically acceptable derivative is a salt or solvate of a compound of formula (I).

It will be appreciated by those skilled in the art that compounds of formula (I) may be modified to provide pharmaceutically acceptable derivatives thereof at any of the functional groups in the compounds, and that the compounds of formula (I) may be derivatised at more than one position.

It will be appreciated that, for pharmaceutical use, the salts, esters, salts of esters and solvates referred to above will be physiologically acceptable salts, esters, salts of esters and solvates but other salts, esters, salts of esters and solvates may find use, for example in the preparation of compounds of formula (I) and the physiological acceptable salts, esters, salts of esters and solvates thereof. Pharmaceutically acceptable salts include those described by Berge, Bighley and Monkhouse , J. Pharm. Sci., 1977, 66, 1-19. The term "pharmaceutically acceptable salts" includes salts prepared from pharmaceutically acceptable non-toxic bases including inorganic bases and organic bases. Salts derived from inorganic bases include aluminum, ammonium, calcium, copper, ferric, ferrous, lithium, magnesium, manganic salts, manganous, potassium, sodium, zinc, and the like. Salts derived from pharmaceutically acceptable organic non-toxic bases include salts of primary, secondary, and tertiary amines, substituted amines including naturally occurring substituted amines, cyclic amines, and basic ion exchange resins, such as arginine, betaine, caffeine, choline, N,N'-dibenzylethylenediamine, diethylamine, 2-diethylammoethanol, 2-dimethylaminoethanol, ethanolamine, ethylenediamine, N-ethyl-morpholine, N- ethylpiperidme, glucamine, glucosamine, histidine, hydrabamine, lsopropylamine, lysine, methylglucamine, morpholine, piperazine, pipeπdine, polyamine resins, procaine, purines, theobromine, tπethylamine, tπmethylamine, tπshydroxylmethyl amino methane, tnpropyl amine, tromethamine, and the like. When a the compound of the present invention is basic, salts may be prepared from pharmaceutically acceptable non-toxic acids, including inorganic and organic acids. Such acids include acetic, benzenesulfonic, benzoic, camphorsulfonic, citric, ethanesulfonic, fumaπc, gluconic, glutamic, hydrobromic, hydrochloric, lsethionic, lactic, maleic, malic, mandelic, methanesulfonic, mucic, mtnc, pamoic, pantothenic, phosphoπc, succinic, sulfuric, tartaric, p-toluenesulfonic acid, and the like.

Examples of pharmaceutically acceptable salts include the ammonium, calcium, magnesium, potassium, and sodium salts, and those formed from maleic, fumaπc, benzoic, ascorbic, pamoic, succinic, hydrochloπc, sulfuric, bismethylenesahcylic, methanesulfonic, ethanedisulfonic, propionic, tartaric, salicylic, citric, gluconic, aspartic, steaπc, palmitic, ltaconic, glycolic, p-aminobenzoic, glutamic, benzenesulfonic, cyclohexylsulfamic, phosphoπc and nitπc acids.

The terms 'halogen or halo' are used to represent fluoπne, chloπne, bromine or iodine.

The term 'alkyF as a group or part of a group means a straight or branched chain alkyl group or combinations thereof, for example a methyl, ethyl, n-propyl, i-propyl, n-butyl, s-butyl, t-butyl, i- butyl, pentyl, hexyl, 1,1-dimethylethyl, heptyl, octyl, nonyl, decyl or combinations thereof.

The term 'alkoxy' as a group or as part of a group means a straight, branched or cyclic chain alkyl group having an oxygen atom attached to the chain, for example a methoxy, ethoxy, n-propoxy, l-propoxy, n-butoxy, s-butoxy, t-butoxy group, i-butoxy, pentoxy, hexyloxy group, cyclopentoxy or cyclohexyloxy group.

Compounds of formula (I) can be prepared as set out in scheme 1 to 3:

Scheme 1:

In Scheme 1, LG¹ and LG² represent leaving groups. In compounds of formula (V), either LG¹ and LG² both represent bromine or LG¹ represents iodine and LG² represents chloπne. Compounds of formula (I) are conveniently prepared by treating a compound of formula (III) wherein LG² represents bromo or chloro with an amine of formula (II). This is conveniently earned out in the presence of a base, such as a tertiary amine (e.g. tπethylamine) and in a solvent, such as a nitπle (e.g. methylnitπle) at elevated temperature with or without microwave irradiation. An excess of the amine may be used in place of the base. The reaction can be earned out in a sealed system.

Alternatively, the treatment of compounds of formula (III) with an amine of formula (II) is carried out in a solvent, such as an amine (e.g. pyridine) and at elevated temperature (e.g. reflux.) Amines of formula (II) are either known compounds or may be prepared by standard literature methods, such as those described in 'Comprehensive Organic Transformations: a guide to functional group preparations' by Richard Larock (VCH, 1989), incorporated herein by reference. 3-oxa-8-azabicyclo[3.2.1]octane may be prepared according to the method descπbed by J. v. Braun and W. Leistner in Chem Ber 1926, 59, 2327.

Compounds of formula (III) may be available commercially or may be obtained from compounds of formula (V), e.g. 3,6-dibromopyπdazine or 3-chloro-6-iodopyridazme, by treatment with a compound of formula (FV) wherein Ar is as defined for compounds of formula (I) and M is a metal for example boron, via a Suzuki coupling reaction employing a palladium source, such as palladium tetrakis(trφhenylphosphine) Pd(PPh₃)₄ or tris(dibenzyhdeneacetone)dipalladium Pd₂(dba)₃, a hgand, such as tπphenylphosphine or tri(tert-butyl)phosphine, and a base, such as sodium carbonate, potassium phosphate or potassium fluoride, in a solvent such as a water/toluene mix, a water/1, 2-dimethoxyethane mix or 1,4-dioxane. Boronate esters or boronic acids of formula (IV) are either known compounds or may be prepared by standard literature methods, such as those described by T Ishiyama, M Murata, and N Miyaura in J Org Chem 1995, 60, 7508, or A. V. Ivachtchenko, D. V.

Kravchenko, V. I. Zheludeva and D. G. Pershin in J Heterocyclic Chem 2004, 41, 931, or R. L. Letsinger and S, H, Dandegaonker in J Am. Chem Soc. 1959, 81, 498.

The compound of formula (V) wherein LG¹ is iodo and LG² is chloro, e.g. 3-Chloro-6- lodopyridazine is readily obtained from 2,6-dichloropyridazine by treatment with sodium iodide and hydπodic acid, according to the method described by AJ. Goodman, S. P. Stanforth and B. Tarbit in Tetrahedron 1999, 55, 15067.

Compounds wherein R¹ and R² together with the N to which they are attached form a 5- membered heteroaryl πng, can be prepared according to scheme 2.

Scheme 2

Compounds of formula (I) wherein R¹ and R² together with the N to which they are attached form a 5-membered heteroaryl πng are conveniently prepared by treatment of a compound of formula (VI) with a compound of formula (II) in the presence of a catalytic quantity of a copper (I) salt, such as copper (I) iodide, a diamine hgand, such as trans-N,N -dimethyl- 1,2- cyclohexanediamine, and a base, such as potassium carbonate. The reaction is conveniently earned out in a solvent, such as toluene and at elevated temperature according, for example, to the method described by J. C. Antilla, A. Klapars and S. L. Buchwald in J Am Chem Soc 2002, 124, 1 1684.

Compounds of formula (VI) may be obtained by treating a compound of formula (III) wherein LG² is bromo, with sodium iodide in the presence of a catalytic quantity of a copper (I) salt, such as copper (I) iodide and a diamine ligand, such as £rø«.y-N,N-dimethyl-l,2- cyclohexanediamine. The reaction is conveniently carried out in a solvent, such as 1,4-dioxane and at elevated temperature according, for example, to the method described by A. Klapars and S. L. Buchwald in J. Am. Chem. Soc. 2002, 124, 14844.

Scheme 3:

In Scheme 3, X represents a leaving group selected from chloro or bromo. Compounds of formula (VII) may be converted to compounds of formula (I) by treatment with a compound of formula (IV) wherein Ar is as defined for compounds of formula (I) and M is a metal for example boron, via a Suzuki coupling reaction employing a palladium source, such as palladium tetrakis(triphenylphosphine) Pd(PPh₃)₄ or tris(dibenzylideneacetone)dipalladium Pd₂(dba)₃, a ligand, such as triphenylphosphine or tri(tert-butyl)phosphine, and a base, such as sodium carbonate, potassium phosphate or potassium fluoride, in a solvent such as a water/toluene mix, a water/ 1 ,2-dimethoxyethane mix or 1 ,4-dioxane.

Compounds of formula (VII) are conveniently prepared by treating a compound of formula (VIII) with an amine of formula (II). This is conveniently carried out in the presence of a base, such as a tertiary amine (e.g. triethylamine) and in a solvent, such as a nitrile (e.g. methylnitrile) in a sealed system at elevated temperature (e.g. from approx 5O⁰C to 16O⁰C) with microwave irradiation. An excess of the amine may be used in place of the base.

It is to be understood that the present invention encompasses all isomers of compounds of formula (I) and their pharmaceutically acceptable derivatives, including all geometric, tautomeric and optical forms, and mixtures thereof (e.g. racemic mixtures). Where additional chiral centres are present in compounds of formula (I), the present invention includes within its scope all possible diastereoismers, including mixtures thereof. The different isomeric forms may be separated or resolved one from the other by conventional methods, or any given isomer may be obtained by conventional synthetic methods or by stereospecific or asymmetric syntheses.

The subject invention also includes isotopically-labeled compounds, which are identical to those recited in formula (I) and following, but for the fact that one or more atoms are replaced by an atom having an atomic mass or mass number different from the atomic mass or mass number usually found in nature. Examples of isotopes that can be incorporated into compounds of the invention include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorous, fluorine, iodine, and chlorine, such as ³H, ¹¹C, ¹⁴C, ¹⁸F, ¹²³I and ¹²⁵I.

Compounds of the present invention and pharmaceutically acceptable salts of said compounds that contain the aforementioned isotopes and/or other isotopes of other atoms are within the scope of the present invention. Isotopically-labeled compounds of the present invention, for example those into which radioactive isotopes such as ³H, ¹⁴C are incorporated, are useful in drug and/or substrate tissue distribution assays. Tritiated, i.e., ³H, and carbon-14, i.e., ¹⁴C, isotopes are particularly preferred for their ease of preparation and detectability. ¹¹C and ⁸F isotopes are particularly useful in PET (positron emission tomography), and ¹²⁵I isotopes are particularly useful in SPECT (single photon emission computerized tomography), all useful in brain imaging. Further, substitution with heavier isotopes such as deuterium, i.e., ²H, can afford certain therapeutic advantages resulting from greater metabolic stability, for example increased in vivo half-life or reduced dosage requirements and, hence, may be preferred in some circumstances. Isotopically labeled compounds of formula (I) and following of this invention can generally be prepared by carrying out the procedures disclosed in the Schemes and/or in the Examples below, by substituting a readily available isotopically labeled reagent for a non-isotopically labeled reagent.

Compounds of formula (I) and their pharmaceutically acceptable derivatives may be prepared in crystalline or non-crystalline form, and, if crystalline, may optionally be solvated. References to solvates herein include hydrates. This invention includes within its scope stoichiometric solvates (including hydrates) as well as compounds containing variable amounts of water and/or solvent.

In view of their ability to bind to the CB2 receptor, it is believed that compounds of the invention will be useful in the treatment of the disorders that follow. Thus, compounds of formula (I) and pharmaceutically acceptable derivatives thereof may be useful as analgesics. For example they may be useful in the treatment of chronic inflammatory pain (e.g. pain associated with rheumatoid arthritis, osteoarthritis, rheumatoid spondylitis, gouty arthritis and juvenile arthritis) including the property of disease modification and joint structure preservation; 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.

Compounds of the invention may also have disease modification or joint structure preservation properties in multiple sclerosis, rheumatoid arthritis, osteo-arthritis, rheumatoid spondylitis, gouty arthritis and juvenile arthritis.

Compounds of the invention may be particularly useful in the treatment of neuropathic pain. Neuropathic pain syndromes can develop following neuronal injury and the resulting pain may persist for months or years, even after the original injury has healed. Neuronal injury may occur in the peripheral nerves, dorsal roots, spinal cord or certain regions in the brain. Neuropathic pain syndromes are traditionally classified according to the disease or event that precipitated them. Neuropathic pain syndromes include: diabetic neuropathy; sciatica; non-specific lower back pain; multiple sclerosis pain; fibromyalgia; HIV-related neuropathy; post-herpetic neuralgia; trigeminal neuralgia; and pain resulting from physical trauma, amputation, cancer, toxins or chronic inflammatory conditions. These conditions are difficult to treat and although several drugs are known to have limited efficacy, complete pain control is rarely achieved. The symptoms of neuropathic pain are incredibly heterogeneous and are often described as spontaneous shooting and lancinating pain, or ongoing, burning pain. In addition, there is pain associated with normally non- painful sensations such as "pins and needles" (paraesthesias and dysesthesias), increased sensitivity to touch (hyperesthesia), painful sensation following innocuous stimulation (dynamic, static or thermal allodynia), increased sensitivity to noxious stimuli (thermal, cold, mechanical hyperalgesia), continuing pain sensation after removal of the stimulation (hyperpathia) or an absence of or deficit in selective sensory pathways (hypoalgesia).

Compounds of formula (I) and their pharmaceutically acceptable derivatives may also be useful in the treatment of fever. Compounds of formula (I) and their pharmaceutically acceptable derivatives may also be useful in the treatment of inflammation, for example in the treatment of skin conditions (e.g. sunburn, burns, eczema, dermatitis, psoriasis); ophthalmic diseases such as glaucoma, retinitis, retinopathies, uveitis and of acute injury to the eye tissue (e.g. conjunctivitis); lung disorders (e.g. asthma, bronchitis, emphysema, allergic rhinitis, respiratory distress syndrome, pigeon fancier's disease, farmer's lung, chronic obstructive pulmonary disease, (COPD); gastrointestinal tract disorders (e.g. aphthous ulcer, Crohn's disease, atopic gastritis, gastritis varialoforme, ulcerative colitis, coeliac disease, regional ileitis, irritable bowel syndrome, inflammatory bowel disease, gastroesophageal reflux disease); organ transplantation; other conditions with an inflammatory component such as vascular disease, migraine, periarteritis nodosa, thyroiditis, aplastic anaemia, Hodgkin's disease, sclerodoma, myaesthenia gravis, multiple sclerosis, sorcoidosis, nephrotic syndrome, Bechet's syndrome, polymyositis, gingivitis, myocardial ischemia, pyrexia, systemic lupus erythematosus, tendinitis, bursitis, and Sjogren's syndrome.

Compounds of formula (I) and their pharmaceutically acceptable derivatives may also be useful in the treatment of bladder hyperrelexia following bladder inflammation. Compounds of formula (I) and their pharmaceutically acceptable derivatives may also be useful in the treatment of immunological diseases such as autoimmune diseases, immunological deficiency diseases or organ transplantation. Compounds of formula (I) and their pharmaceutically acceptable derivatives may also be effective in increasing the latency of HIV infection.

Compounds of formula (I) and their pharmaceutically acceptable derivatives may also be useful in the treatment of diseases of abnormal platelet function (e.g. occlusive vascular diseases).

Compounds of formula (I) and their pharmaceutically acceptable derivatives may also be useful in the treatment of neuritis, heart burn, dysphagia, pelvic hypersensitivity, urinary incontinence, cystitis or pruritis.

Compounds of formula (I) and their pharmaceutically acceptable derivatives may also have diuretic action.

Compounds of formula (I) and their pharmaceutically acceptable derivatives may also be useful in the treatment of impotence or erectile dysfunction.

Compounds of formula (I) and their pharmaceutically acceptable derivatives may also be useful for attenuating the hemodynamic side effects of non-steroidal anti-inflammatory drugs (NSAID's) and cyclooxygenase-2 (COX-2) inhibitors.

Compounds of formula (I) and their pharmaceutically acceptable derivatives may also be useful in the treatment of neurodegenerative diseases and neurodegeneration such as dementia, particularly degenerative dementia (including senile dementia, Alzheimer's disease, Pick's disease, Huntingdon's chorea, Parkinson's disease and Creutzfeldt-Jakob disease, motor neuron disease); vascular dementia (including multi-infarct dementia); as well as dementia associated with intracranial space occupying lesions; trauma; infections and related conditions (including HIV infection); dementia in Parkinson's disease ; metabolism; toxins; anoxia and vitamin deficiency; and mild cognitive impairment associated with ageing, particularly Age Associated Memory Impairment. The compounds may also be useful for the treatment of amyotrophic lateral sclerosis (ALS) and neuroinflamation.

Compounds of formula (I) and their pharmaceutically acceptable derivatives may also be useful in neuroprotection and in the treatment of neurodegeneration following stroke, cardiac arrest, pulmonary bypass, traumatic brain injury, spinal cord injury or the like.

Compounds of formula (I) and their pharmaceutically acceptable derivatives may also be useful in the treatment of tinnitus.

Compounds of formula (I) and their pharmaceutically acceptable derivatives may also be useful in the treatment of 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, sexual dysfunction, sleep disorders (including disturbances of circadian rhythm, dyssomnia, insomnia, sleep apnea and narcolepsy), withdrawal from abuse of drugs such as of cocaine, 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 of formula (I) and their pharmaceutically acceptable derivatives may also be useful in preventing or reducing dependence on, or preventing or reducing tolerance or reverse tolerance to, a dependence - inducing agent. Examples of dependence inducing agents include opioids (e.g. morphine), CNS depressants (e.g. ethanol), psychostimulants (e.g. cocaine) and nicotine.

Compounds of formula (T) and their pharmaceutically acceptable derivatives may also be useful in the treatment of kidney dysfunction (nephritis, particularly mesangial proliferative glomerulonephritis, nephritic syndrome), liver dysfunction (hepatitis, cirrhosis), gastrointestinal dysfunction (diarrhoea) and colon cancer. In one embodiment compounds of the invention may bind selectively to the CB2 receptor; such compounds may be particularly useful in treating CB2 receptor mediated diseases.

The term "treatment" or "treating" as used herein includes the treatment of established disorders and also includes the prophylaxis thereof. The term " prophylaxis" is used herein to mean preventing symptoms in an already afflicted subject or preventing recurrence of symptoms in an afflicted subject and is not limited to complete prevention of an affliction.

According to a further aspect of the invention, we provide a compound of formula (I) or a pharmaceutically acceptable derivative thereof for use in human or veterinary medicine. According to another aspect of the invention, we provide a compound of formula (I) or a pharmaceutically acceptable derivative thereof for use in the treatment of a condition which is mediated by the activity of cannabmoid 2 receptors.

According to a further aspect of the invention, we provide the use of a compound of formula (I) or a pharmaceutically acceptable derivative thereof for the manufacture of a therapeutic agent for the treatment of a condition which is mediated by the activity of cannabmoid 2 receptors. According to a further aspect of the invention, we provide a method of treating a mammal, for example a human suffering from a condition which is mediated by the activity of cannabmoid 2 receptors which comprises administering to said subject a non toxic, therapeutically effective amount of a compound of formula (I) or a pharmaceutically acceptable deπvative thereof.

According to a further aspect of the invention we provide a method of treating a mammal, for example a human suffering from an immune disorder, an inflammatory disorder, pain, rheumatoid arthritis, multiple sclerosis, osteoarthritis or osteoporosis which method comprises administering to said subject a non toxic, therapeutically effective amount of a compound of formula (I) or a pharmaceutically acceptable derivative thereof.

According to another aspect of the invention, we provide a compound of formula (I) or a pharmaceutically acceptable derivative thereof for use in the treatment of a condition such as an immune disorder, an inflammatory disorder, pain, rheumatoid arthritis, multiple sclerosis, osteoarthritis or osteoporosis. According to another aspect of the invention is provided the use of a compound of formula

(I) or a pharmaceutically acceptable deπvative thereof for the manufacture of a therapeutic agent for the treatment or prevention of a condition such as an immune disorder, an inflammatory disorder, pain, rheumatoid arthritis, multiple sclerosis, osteoarthritis or osteoporosis.

In one embodiment the condition is pain. In a further embodiment pain is selected from inflammatory pain, viseral pain, cancer pain, neuropathic pam, lower back pain, muscular sceletal, post operative pam, acute pain and migraine. For example, the inflammatory pain is pam associated with rheumatoid arthritis or osteoarthritis.

In order to use a compound of formula (I) or a pharmaceutically acceptable deπvative thereof for the treatment of humans and other mammals it is normally formulated in accordance with standard pharmaceutical practice as a pharmaceutical composition. Therefore in another aspect of the invention there is provided a pharmaceutical composition comprising a compound of formula (I) or a pharmaceutically acceptable derivative thereof adapted for use in human or veterinary medicine. In one embodiment the pharmaceutical composition further comprises a pharmaceutical carrier or diluent thereof.

As used herein, "modulator" means both antagonist, partial or full agonist and inverse agonist. In one embodiment the present modulators are agonists. In another embodiment the present modulators are antagonists. In one embodiment the compounds of the present invention are CB2 agonists. Compounds of formula (I) and their pharmaceutically acceptable deπvatives may be administered in a standard manner for the treatment of the indicated diseases, for example orally, parenterally, sub-hngually, dermally, intranasally, transdermally, rectally, via inhalation or via buccal administration. Compounds of formula (I) and their pharmaceutically acceptable derivatives which are active when given orally can be formulated as liquids, tablets, capsules and lozenges. A liquid formulation will generally consist of a suspension or solution of the compound or salt in a liquid carrier for example, ethanol, olive oil, glycerine, glucose (syrup) or water with a flavouring, suspending, or colouring agent. Where the composition is in the form of a tablet, any pharmaceutical carrier routinely used for preparing solid formulations may be used. Examples of such carriers include magnesium stearate, terra alba, talc, gelatin, acacia, steaπc acid, starch, lactose and sucrose. Where the composition is in the form of a capsule, any routine encapsulation is suitable, for example using the aforementioned earners or a semi solid e.g. mono di-glyceπdes of capric acid, Gelucire™ and Labrasol™, or a hard capsule shell e.g gelatin. Where the composition is in the form of a soft shell capsule e.g. gelatin, any pharmaceutical carrier routinely used for preparing dispersions or suspensions may be considered, for example aqueous gums or oils, and are incorporated in a soft capsule shell.

Typical parenteral compositions consist of a solution or suspension of a compound or derivative in a steπle aqueous or non-aqueous carrier optionally containing a parenterally acceptable oil, for example polyethylene glycol, polyvinylpyrrolidone, lecithin, arachis oil or sesame oil.

Typical compositions for inhalation are in the form of a solution, suspension or emulsion that may be administered as a dry powder or in the form of an aerosol using a conventional propellant such as dichlorodifiuoromethane or tπchlorofluoromethane.

A typical suppository formulation comprises a compound of formula (I) or a pharmaceutically acceptable deπvative thereof which is active when administered in this way, with a binding and/or lubricating agent, for example polymeric glycols, gelatins, cocoa-butter or other low melting vegetable waxes or fats or their synthetic analogs. Typical dermal and transdermal formulations comprise a conventional aqueous or nonaqueous vehicle, for example a cream, ointment, lotion or paste or are in the form of a medicated plaster, patch or membrane.

In one embodiment the composition is m unit dosage form, for example a tablet, capsule or metered aerosol dose, so that the patient may administer a single dose. Each dosage unit for oral administration contains suitably from 0.001 mg to 500 mg, for example 0.01 mg to 500 mg such as from 0.01 mg to 100 mg, and each dosage unit for parenteral administration contains suitably from 0.001 mg to 100 mg, of a compound of formula (I) or a pharmaceutically acceptable deπvative thereof calculated as the free acid (undeπvatised compound). Each dosage unit for suppository administration contains suitably from 0.001 mg to 500 mg, for example 0.01 mg to 500 mg such as from 0.01 mg to 100 mg. Each dosage unit for intranasal administration contains suitably 1-400 mg and suitably 10 to 200 mg per person. A topical formulation contains suitably 0.01 to 5.0% of a compound of formula (I).

The daily dosage regimen for oral administration is suitably about 0.01 mg/Kg to 1000 mg/Kg, of a compound of formula(I) or a pharmaceutically acceptable deπvative thereof calculated as the free acid (undeπvatised compound). The daily dosage regimen for parenteral administration is suitably about 0.001 mg/Kg to 200 mg/Kg, of a compound of formula (I) or a pharmaceutically acceptable derivative thereof calculated as the free acid (undeπvatised compound). The daily dosage regimen for suppository administration is suitably about 0.01 mg/Kg to 1000 mg/Kg, of a compound of formula(I) or a pharmaceutically acceptable derivative thereof calculated as the free acid (underivatised compound). The daily dosage regimen for intranasal administration and oral inhalation is suitably about 10 to about 500 mg/person. The active ingredient may be administered from 1 to 6 times a day, sufficient to exhibit the desired activity. It may be advantageous to prepare the compounds of the present invention as nanoparticles. This may improve the oral bioavailability of the compounds. For the purposes of the present invention "nanoparticulate" is defined as solid particles with 50% of the particles having a particle size of less than lμm, for example less than 0.75μm

The particle size of the solid particles of a compound of formula (I) may be determined by laser diffraction. A suitable machine for determining particle size by laser diffraction is a Lecotrac laser particle size analyser, using an HELOS optical bench fitted with a QUDCEL dispersion unit.

Numerous processes for the synthesis of solid particles in nanoparticulate form are known. Typically these processes involve a milling process, for example a wet milling process in the presence of a surface modifying agent that inhibits aggregation and/or crystal growth of the nanoparticles once created. Alternatively these processes may involve a precipitation process, for example, a process of precipitation in an aqueous medium from a solution of the drug in a nonaqueous solvent.

Accordingly, in a further aspect, the present invention provides a process for preparing compounds of formula (I) and their pharmaceutically acceptable derivatives in nanoparticulate form as hereinbefore defined, which process comprises milling or precipitation.

Representative processes for the preparation of solid particles in nanoparticulate form are described in the patents and publications listed below.

U.S. Patent No. 4,826,689 to Violanto & Fischer, U. S. Patent No. 5,145,684 to Liversidge et al U.S Patent No. 5,298,262 to Na & Rajagopalan, U.S. Patent No. 5,302,401 Liversidge et al U.S. Patent No. 5,336,507 to Na & Rajagopalan, U.S. Patent No. 5,340,564 to Illig & Sarpotdar U.S. Patent No. 5,346,702 to Na Rajagopalan, U.S. Patent No. 5,352,459 to Hollister et al U.S. Patent No. 5,354,560 to Lovrecich, U.S. Patent No. 5,384,124 to Courteille et al, U.S. Patent No. 5,429,824 to June, U.S. Patent No. 5,503,723 to Ruddy et al, U.S. Patent No. 5,510 118 to Bosch et al, U.S. Patent No. 5,518 to Bruno et al, U.S. Patent No. 5,518,738 to Eickhoff et al, U.S. Patent No. 5,534,270 to De Castro, U.S. Patent No. 5,536,508 to Canal et al, U.S. Patent No. 5,552,160 to Liversidge et al, U.S. Patent No. 5,560,931 to Eickhoff et al, U.S. Patent No. 5,560,932 to Bagchi et al, U.S. Patent No. 5,565,188 to Wong et al, U.S. Patent No. 5,571,536 to Eickhoff et al, U.S. Patent No. 5,573,783 to Desieno & Stetsko, U.S Patent No. 5,580,579 to Ruddy et al, U.S. Patent No 5,585,108 to Ruddy et al, U.S. Patent No. 5,587,143 to Wong, U.S. Patent No. 5,591456 to Franson et al, U.S. Patent No. 5,622,938 to Wong, U.S. Patent No 5,662,883 to Bagchi et al, U.S. Patent No. 5,665,331 to Bagchi et al, U.S Patent No. 5,718,919 to Ruddy et al, U.S. Patent No. 5,747,001 to Wiedmann et al, WO93/25190, WO96/24336, WO 97/14407, WO 98/35666, WO 99/65469, WO 00/18374, WO 00/27369, WO 00/30615 and WO 01/41760. Such processes may be readily adapted for the preparation of compounds of formula (I) and their pharmaceutically acceptable derivatives in nanoparticulate form. Such processes form a further aspect of the invention. The process of the present invention may use a wet milling step carried out in a mill such as a dispersion mill in order to produce a nanoparticulate form of the compound. The present invention may be put into practice using a conventional wet milling technique, such as that described in Lachman et al., The Theory and Practice of Industrial Pharmacy, Chapter 2, "Milling" p.45 (1986).

In a further refinement, WO02/00196 (SmithKline Beecham pic) describes a wet milling procedure using a mill in which at least some of the surfaces are made of nylon (polyamide) comprising one or more internal lubricants, for use in the preparation of solid particles of a drug substance in nanoparticulate form. In another aspect the present invention provides a process for preparing compounds of the invention in nanoparticulate form comprising wet milling a suspension of the compound in a mill having at least one chamber and agitation means, said chamber(s) and/or said agitation means comprising a lubricated nylon, as described in WO02/00196.

The suspension of a compound of the invention for use in the wet milling is typically a liquid suspension of the coarse compound in a liquid medium. By "suspension" is meant that the compound is essentially insoluble in the liquid medium. Representative liquid media include an aqueous medium. Using the process of the present invention the average particle size of coarse compound of the invention may be up to lmm in diameter. This advantageously avoids the need to pre-process the compound. In a further aspect of the invention the aqueous medium to be subjected to the milling comprises a compound of formula (I) or a pharmaceutically acceptable derivative thereof present in from about 1% to about 40% w/w, suitably from about 10% to about 30% w/w, for example about 20% w/w.

The aqueous medium may further comprise one or more pharmaceutically acceptable water- soluble carriers which are suitable for steric stabilisation and the subsequent processing of a compound of formula (I) or a pharmaceutically acceptable derivative thereof after milling to a pharmaceutical composition, e.g. by spray drying. Pharmaceutically acceptable excipients most suitable for steric stabilisation and spray-drying are surfactants such as poloxamers, sodium lauryl sulphate and polysorbates etc; stabilisers such as celluloses e.g. hydroxypropylmethyl cellulose; and carriers such as carbohydrates e.g. mannitol.

In a further aspect of the invention the aqueous medium to be subjected to the milling may further comprise hydroxypropylmethyl cellulose (HPMC) present from about 0.1 to about 10% w/w.

The process of the present invention may comprise the subsequent step of drying a compound of the invention to yield a powder. Accordingly, in a further aspect, the present invention provides a process for preparing a pharmaceutical composition containing a compound of the present invention which process comprises producing a compound of formula (I) or a pharmaceutically acceptable derivative thereof in nanoparticulate form optionally followed by drying to yield a powder, and optionally admixing with one or more pharmaceutically acceptable carriers or excipients. A further aspect of the invention is a pharmaceutical composition comprising a compound of formula (I) or a pharmaceutically acceptable derivative thereof in which the compound of formula (I) or a pharmaceutically acceptable derivative thereof is present in solid particles in nanoparticulate form, in admixture with one or more pharmaceutically acceptable carriers or excipients.

By "drying" is meant the removal of any water or other liquid vehicle used during the process to keep a compound of formula (I) in liquid suspension or solution. This drying step may be any process for drying known in the art, including freeze drying, spray granulation or spray drying. Of these methods spray drying is particularly preferred. All of these techniques are well known in the art. Spray drymg/fiuid bed granulation of milled compositions is earned out most suitably using a spray dryer such as a Mobile Minor Spray Dryer [Niro, Denmark], or a fluid bed drier, such as those manufactured by Glatt, Germany. In a further aspect the invention provides a pharmaceutical composition as hereinbefore defined, in the form of a dπed powder, obtainable by wet milling solid particles of a compound of formula (I) followed by spray-drymg the resultant suspension.

In one embodiment, the pharmaceutical composition as hereinbefore defined, further compπses HPMC present in less than 15% w/w, for example, in the range 0.1 to 10% w/w. The CB2 receptor modulators for use in the instant invention may be used in combination with other therapeutic agents, for example COX-2 inhibitors, such as celecoxib, deracoxib, rofecoxib, valdecoxib, parecoxib or COX-189; 5 -lipoxygenase inhibitors; NSAID's, such as aspirin, diclofenac, lndomethacin, nabumetone or lbuprofen; leukotπene receptor antagonists; DMARD' s such as methotrexate; adenosine Al receptor agonists; sodium channel blockers, such as lamotrigine; NMDA receptor modulators, such as glycine receptor antagonists; gabapentin and related compounds; tricyclic antidepressants such as amitriptyhne; neurone stabilising antiepileptic drugs; mono-aminergic uptake inhibitors such as venlafaxme; opioid analgesics; local anaesthetics; 5HT, agonists, such as tπptans, for example sumatriptan, naratriptan, zolmitπptan, eletπptan, frovatriptan, almotπptan or rizatriptan; EPi receptor hgands, EP₄ receptor hgands; EP₂ receptor hgands; EP₃ receptor hgands; EP₄ antagonists; EP₂ antagonists and EP₃ antagonists; bradykinin receptor hgands and vamlloid receptor hgand, antirheumatoid arthritis drugs, for example anti TNF drugs e.g. enbrel, remicade, anti-IL-1 drugs, DMARDS e.g. leflunamide or 5HT₆ compounds. When the compounds are used in combination with other therapeutic agents, the compounds may be administered either sequentially or simultaneously by any convenient route. Additional COX-2 inhibitors are disclosed in US Patent Nos. 5,474,995 US5,633,272;

US5,466,823, US6,310,099 and US6.291.523; and in WO 96/25405, WO 97/38986, WO 98/03484, WO 97/14691, WO99/12930, WO00/26216, WO00/52008, WOOO/38311, WO01/58881 and WO02/18374.

Suitable 5HT6 compounds for a combination suitable for the treatment of e.g Alzheimer's disease or cognitive enhancement, may be selected from SGS518 (Saegis), BGC20 761 (BTG disclosed in WO00/34242), WAY466 (Wyeth), PO4368554 (Hoffman Ie Roche), BVT5182 (Biovitron) and LY483518 (Lily), SB742457 (GSK) and/or compounds disclosed as Example 1 to 50 in WO03/080580.

Compounds of the present invention may be administered in combination with other active substances such as 5HT3 antagonists, NK-I antagonists, serotonin agonists, selective serotonin reuptake inhibitors (SSRI), noradrenaline re-uptake inhibitors (SNRI), tricyclic antidepressants and/or dopaminergic antidepressants. Suitable 5HT3 antagonists which may be used in combination with a compound of the invention include for example ondansetron, granisetron, metoclopramide.

Suitable NKl antagonists which may be used in combination with a compound of the invention include for example aprepitant. Suitable serotonin agonists which may be used in combination with a compound of the invention include for example sumatriptan, rauwolscine, yohimbine, metoclopramide.

Suitable SSRIs which may be used in combination with a compound of the invention include for example fluoxetine, citalopram, femoxetine, fluvoxamine, paroxetine, indalpine, sertraline, zimeldine. Suitable SNRIs which may be used in combination with a compound of the invention include for example venlafaxine and reboxetine.

Suitable tricyclic antidepressants which may be used in combination with a compound of the invention include for example imipramine, amitriptiline, chlomipramine and nortriptiline.

Suitable dopaminergic antidepressants which may be used in combination with a compound of the invention include for example bupropion and amineptine.

Compounds of the present invention may used in combination with PDE4 inhibitors. PDE4 inhibitors useful in this invention may be any compound that is known to inhibit the PDE4 enzyme or which is discovered to act in as PDE4 inhibitor, and which is only or essentially only a PDE4 inhibitor, not compounds which inhibit to a degree of exhibiting a therapeutic effect other members of the PDE family as well as PDE4. Generally it is preferred to use a PDE4 antagonist which has an IC₅₀ ratio of about 0.1 or greater as regards the IC₅₀ for the PDE4 catalytic form which binds rolipram with a high affinity divided by the IC₅₀ for the form which binds rolipram with a low affinity. Compounds of the present invention or combinations with PDE4 can be used in treating inflammation and as bronchodilators. There are at least two binding forms on human monocyte recombinant PDE 4 (hPDE 4) at which inhibitors bind. One explanation for these observations is that hPDE 4 exists in two distinct forms. One binds the likes of rolipram and denbufylline with a high affinity while the other binds these compounds with a low affinity. The preferred PDE4 inhibitors of for use in this invention will be those compounds which have a salutary therapeutic ratio, i.e., compounds which preferentially inhibit cAMP catalytic activity where the enzyme is in the form that binds rolipram with a low affinity, thereby reducing the side effects which apparently are linked to inhibiting the form which binds rolipram with a high affinity. Another way to state this is that the preferred compounds will have an IC₅₀ ratio of about 0.1 or greater as regards the IC₅₀ for the PDE 4 catalytic form which binds rolipram with a high affinity divided by the IC₅₀ for the form which binds rolipram with a low affinity.

Reference is made to U.S. patent 5,998,428, which describes these methods in more detail. It is incorporated herein in full as though set forth herein.

Suitably the PDE4 inhibitors are those PDE4 inhibitors which have an IC₅₀ ratio of greater than 0.5, and particularly those compounds having a ratio of greater than 1.0. A further aspect of the invention is a CB2 modulator (a compound of formula (I) and pharmaceutically acceptable derivatives thereof) in combination with a PDE4 inhibitor and pharmaceutical compositions comprising said combination. A further aspect of the invention is a method of treating lung disorders for example asthma, bronchitis, emphysema, allergic rhinitis, respiratory distress syndrome, pigeon fancier's disease, farmer's lung, chronic obstructive pulmonary disease, (COPD) and cough or a disorder which can be treated with a broncodilator which comprises administering to a mammal including man, an effective amount of a CB modulator (compounds of formula (I) and their pharmaceutically acceptable derivatives) and an effective amount of a PDE4 inhibitor or a pharmaceutically acceptable derivative thereof.

An additional aspect of the invention is the use of an effective amount of a CB2 modulator of formula (I) or a pharmaceutically acceptable derivative thereof and an effective^" amount of a PDE4 inhibitor or a pharmaceutically acceptable derivative thereof in the manufacture of a medicament in the treatment of lung disorders for example asthma, bronchitis, emphysema, allergic rhinitis, respiratory distress syndrome, pigeon fancier's disease, farmer's lung, chronic obstructive pulmonary disease, (COPD) and cough or for the manufacture of a bronchodilator.

When used herein cough can have a number of forms and includes productive, non- productive, hyper-reactive, asthma and COPD associated.

A further aspect of the invention is a patient pack comprising an effective amount of a CB 2 modulator of formula (I) or a pharmaceutically acceptable derivative thereof and an effective amount of a PDE4 inhibitor or a pharmaceutically acceptable derivative thereof.

Possible PDE4 compounds are cis [cyano-4-(3-cyclopentyloxy-4- methoxyphenytycyclohexan-l-carboxylate] also known as cilomilast or Ariflo®, 2-carbomethoxy-

4-cyano-4-(3-cyclopropylmethoxy-4-difluoromethoxyphenyl)cyclohexan-l-one, and cis [4-cyano-

4-(3-cyclopropylmethoxy-4-difluoromethoxyphenyl)cyclohexan-l-ol]. They can be made by the processed described in US patents 5,449,686 and 5,552,438. Other PDE4 inhibitors, specific inhibitors, which can be used in this invention are AWD- 12-281 from ASTA MEDICA (Hofgen, N. et al. 15th EFMC Int Symp Med Chem (Sept 6-10, Edinburgh) 1998, Abst P.98); a 9- benzyladenine derivative nominated NCS-613 (INSERM); D-4418 from Chiroscience and

Schering-Plough; a benzodiazepine PDE4 inhibitor identified as CI-1018 (PD-168787; Parke-

Davis/Warner-Lambert); a benzodioxole derivative Kyowa Hakko disclosed in WO 9916766; 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 (WO 99/47505) from Byk-Gulden (now Altana); or a compound identified as T-

440 (Tanabe Seiyaku; Fuji, K. et al. J Pharmacol Exp Ther, 1998, 284(1): 162).

Additional PDE4 inhibitors are disclosed on pages 2 to 15 of WO01/13953. Specifically selected are arofylline, atizoram, BAY-19-8004, benafentrine, BYK-33043, CC-3052, CDP-840, cipamfylline, CP-220629, CP-293121, D-22888, D-4396, denbufylline, filaminast, GW-3600, ibudilast, KF-17625, KS-506-G, laprafylline, NA-0226A, NA-23063A, ORG-20241, ORG-30029,

PDB-093, pentoxifylline, piclamilast, rolipram, RPR-117658, RPR-122818, RPR-132294, RPR-

132703, RS-17597, RS-25344-000, SB-207499, SB210667, SB211572, SB-211600, SB212066,

SB212179, SDZ-ISQ-844, SDZ-MNS-949, SKF-107806, SQ-20006, T-2585, tibenelast, tolafentrine, UCB-29646, V-11294A, YM-58997, YM-976 and zardaverine.

In one embodiment the PDE4 inhibitor is selected from cilomilast, AWD-12-281, NCS-613, D- 4418, CI-1018, V-11294A, roflumilast or T-440. Compounds of the present invention may also be of use in treating atherosclerosis in combination with an anti-hyperlipidaemic, anti-atherosclerotic, anti-diabetic, anti-anginal, anti- hypertension agent or an agent for lowering Lp(a). Examples of the above include cholesterol synthesis inhibitors such as statins, anti-oxidants such as probucol, insulin sensitisers, calcium channel antagonists. Examples of agents for lowering Lp(a) include the aminophosphonates described in WO 97/02037, WO 98/28310, WO 98/28311 and WO 98/28312 (Symphar SA and SmithKline Beecham). Examples of antihyerpertension agents are angiotensin-converting enzyme inhibitors, angiotensin-II receptor antagonists, ACE / NEP inhibitors, -blockers, calcium channel blockers, PDE inhibitors, aldosterone blockers A possible combination therapy will be the use of a compound of the present invention and a statin. The statins are a well known class of cholesterol lowering agents and include atorvastatin, simvarstatin, pravastatin, cerivastatin, fluvastatin, lovastatin and ZD 4522 (also referred to as S- 4522, Astra Zeneca). The two agents may be administered at substantially the same time or at different times, according to the discretion of the physician. A further possible combination therapy will be the use of a compound of the present invention and an anti-diabetic agent or an insulin sensitiser. Within this class, possible compounds for use with a compound of the present invention include the PPARgamma activators, for instance G 1262570 (Glaxo Wellcome) and also the glitazone class of compounds such as rosiglitazone (Avandia, SmithKline Beecham), troglitazone and pioglitazone. It will be appreciated that the compounds of any of the above combinations or compositions may be administered simultaneously (either in the same or different pharmaceutical formulations), separately or sequentially.

The invention thus provides, in a further aspect, a combination comprising a compound of formula (I) or a pharmaceutically acceptable derivative thereof together with a further therapeutic agent or agents.

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 carrier or excipient comprise a further aspect of the invention. The individual components of such combinations may be administered either sequentially or simultaneously in separate or combined pharmaceutical formulations.

When a compound of formula (I) or a pharmaceutically acceptable derivative thereof is used in combination with a second therapeutic agent active against the same disease state the dose of each compound may differ from that when the compound is used alone. Appropriate doses will be readily appreciated by those skilled in the art.

Determination of cannabinoid CBl Receptor Agonist Activity

The cannabinoid CBl receptor agonist activity of compounds of formula (I) was determined in accordance with the following experimental method.

Experimental Method

Yeast (Saccharomyces cerevisiae) cells expressing the human cannabinoid CBl receptor were generated by integration of an expression cassette into the ura3 chromosomal locus of yeast strain MMY23. This cassette consisted of DNA sequence encoding the human CBl receptor flanked by the yeast GPD promoter to the 5' end of CBl and a yeast transcriptional terminator sequence to the 3' end of CBl. MMY23 expresses a yeast/mammalian chimeric G-protein alpha subunit in which the C-terminal 5 amino acids of Gpal are replaced with the C-terminal 5 amino acids of human Gαil/2 (as described in Brown et al. (2000), Yeast 16:11-22). Cells were grown at 30⁰C in liquid Synthetic Complete (SC) yeast media (Guthrie and Fink (1991), Methods in Enzymology, Vol. 194) lacking uracil, tryptophan, adenine and leucine to late logarithmic phase (approximately 6 OD₆₀o/ml).

Agonists were prepared as 10 mM stocks in DMSO. EC₅₀ values (the concentration required to produce 50% maximal response) were estimated using 4 fold dilutions (BiomekFX, Beckman) into DMSO. Agonist solutions in DMSO (1% final assay volume) were transferred into black microtitre plates from Gremer (384-well). Cells were suspended at a density of 0.2 OO₆₀JnU in SC media lacking histidine, uracil, tryptophan, adenine and leucine and supplemented with

1OmM 3-aminotriazole, 0.1M sodium phosphate pH 7.0, and lOμM fluorescein di-β-D- glucopyranoside (FDGIu). This mixture (50ul per well) was added to agonist in the assay plates (Multidrop 384, Labsystems). After incubation at 30⁰C for 24 hours, fluorescence resulting from degradation of FDGIu to fluorescein due to exoglucanase, an endogenous yeast enzyme produced during agonist-stimulated cell growth, was determined using a fluorescence microtitre plate reader

(Tecan Spectrofluor or LJL analyst excitation wavelength: 485nm; emission wavelength: 535nm).

Fluorescence was plotted against compound concentration and iteratively curve fitted using a four parameter fit to generate a concentration effect value. Efficacy (E_n13x) was calculated from the equation

Emax = Max_[comp_{Ound X]} - Min_[compound x] / Max_[Hu2io] - Mm[HU₂IO] X 100% where Max_[Compoun(i _X] and Min_{[COmpound X]} are the fitted maximum and minimum respectively from the concentration effect curve for compound X, and Max_[Hu₂io_] and Min_[Hu₂io_] are the fitted maximum and minimum respectively from the concentration effect curve for (6aR,10aR)-3-(l,l'- Dimethylheptyl)-6a,7, 10, 1 Oa-tetrahydro- 1 -hydroxy-6,6-dimethyl-6H-dibenzo[b,d]pyran-9- methanol (HU210; available from Tocπs). Equieffective molar ratio (EMR) values were calculated from the equation

EMR = EC50 [compound X] / EC50 [HU210] Where EC_{50 [}compound x] is the EC₅₀ of compound X and EC_{5O [H}U₂io_] is the EC₅₀ofHU210. pEC₅₀ is the negative log of the EC₅₀

Compounds of Examples 2, 3 and 18 as tested according to this method had pEC₅₀ values of between 5 and 5.6, all other examples had pEC50 values of less than or equal to 5 and/or efficacy of <30% at the cloned human cannabinoid CBl receptor.

Determination of cannabinoid CB2 Receptor Agonist Activity

The cannabinoid CB2 receptor agonist activity of compounds of formula (I) was determined in accordance with the following expeπmental method.

Experimental Method

Yeast (Saccharomyces cerevisiae) cells expressing the human cannabinoid CB2 receptor were generated by integration of an expression cassette into the ura3 chromosomal locus of yeast strain MMY23. This cassette consisted of DNA sequence encoding the human CB2 receptor flanked by the yeast GPD promoter to the 5' end of CB2 and a yeast transcriptional terminator sequence to the 3' end of CB2. MMY23 expresses a yeast/mammalian chimeric G-protem alpha subunit in which the C-terminal 5 ammo acids of Gpal are replaced with the C-terminal 5 amino acids of human Gαil/2 (as described in Brown et al. (2000), Yeast 16:11-22). Cells were grown at 30⁰C in liquid Synthetic Complete (SC) yeast media (Guthrie and Fink (1991), Methods in Enzymology, Vol. 194) lacking uracil, tryptophan, adenine and leucine to late logarithmic phase (approximately 6 OD₆₀₀/ml).

Agonists were prepared as 10 mM solutions in DMSO. EC₅₀ values (the concentration required to produce 50% maximal response) were estimated using 4 fold dilutions (BiomekFX, Beckman) into DMSO. Agonist solutions in DMSO (1% final assay volume) were transferred into black microtitre plates from Greiner (384-well). Cells were suspended at a density of 0.2 OD₆oo/ml in SC media lacking histidine, uracil, tryptophan, adenine and leucine and supplemented with

1OmM 3-aminotriazole, 0.1M sodium phosphate pH 7.0, and lOμM fluorescein di-β-D- glucopyranoside (FDGIu). This mixture (50ul per well) was added to agonist in the assay plates (Multidrop 384, Labsystems). After incubation at 30⁰C for 24 hours, fluorescence resulting from degradation of FDGIu to fluorescein due to exoglucanase, an endogenous yeast enzyme produced during agonist-stimulated cell growth, was determined using a fluorescence microtitre plate reader

(Tecan Spectrofluor or LJL Analyst excitation wavelength: 485nm; emission wavelength: 535nm).

Fluorescence was plotted against compound concentration and iteratively curve fitted using a four parameter fit to generate a concentration effect value. Efficacy (E_m2x) was calculated from the equation

Emax = Max_{[compound X]} - Min_{[compound X]}/ Max_[Hu2io] - Mm_[Hu2io] X 100% where Max_{[compound X]} and Min_[compOunti _X] are the fitted maximum and minimum respectively from the concentration effect curve for compound X, and MaX[HU2io] and Mm_[Hu₂io_] are the fitted maximum and minimum respectively from the concentration effect curve for (6aR,10aR)-3-(l,l'- Dimethylheptyl)-6a,7, 10,1 Oa-tetrahydro- 1 -hydroxy-6,6-dimethyl-6H-dibenzo[b,d]pyran-9- methanol (HU210; available from Tocπs). Equieffective molar ratio (EMR) values were calculated from the equation

EMR = EC₅₀ [compound X] / EC₅₀ [HU210] Where EC_{50 [}compound x] is the EC₅₀ of compound X and EC₅₀ [HU2_IOJ IS the EC₅₀ of HU210. pECso is the negative log of the EC₅₀

The compounds of Examples 1 to 27 as tested according to this method had pEC₅₀ values of greater than 6.5 units and efficacy of >50% at the cloned human cannabinoid CB2 receptor. The compounds of Examples 28 to 37 had pEC₅₀ values of 6 to 6.5 units and the compounds of Examples 38 to 54 had pEC₅₀ values of up to 6 units at the cloned human cannabinoid CB2 receptor.

Experimental Method Measurement of CB2 agonist effects in a reporter gene assay

CB2 agonist effects may be determined using a reporter gene assay. These studies are performed using a CHO-Kl cell line expressing human recombinant CB2 receptors (CHO-Kl CB2 CRE-LUC cells). These cells additionally express a "CRE-LUC" reporter gene construct comprising the gene for luciferase under the control of multiple cAMP response element binding protein promoters. In these cells, increases in intracellular cAMP levels leads to transcription of the luciferase gene and the subsequent production of luciferase. The expression of luciferase is measured by addition to the cells of a proprietary mixture containing luciferin, the substrate for luciferase (Luclite, Perkin Elmer, Cat No 6016919). The resultant reaction leads to the generation of light which is measured in a TopCount scintillation counter. In the CHO-Kl CB2 CRE-LUC cells, forskolin produces a marked increase in luciferase expression and CB2 agonists inhibit this response. The CHO-Kl CB2 CRE-LUC cells routinely express a high level of constitutive CB2 receptor activity. This was overcome in these experiments by pre-treating the cells with the inverse agonist, SR144528, for 30-60mins before use. This treatment has been shown to eliminate constitutive CB2 receptor activity (Bouaboula et al., 1999).

Methods

CHO-Kl CB2 CRE-LUC cells are grown in DMEM/F12 plus glutamax I medium (Gibco Cat. No. 31331-028), supplemented with 9% FBS (Gibco, Cat. No. 16000-040) and O.Smg.ml^"1 G418

(Gibco, Cat. No. 10131-027) and 0.5mg.mr' Hygromycin (Invitrogen, Cat. No. 10687-010). Cells are grown as a monolayer culture in 162cm² vented Nunclon flasks (NUNC, Cat. No. 178883) in 27.5ml of media in a humidified 95% air and 5% CO₂ atmosphere at 37⁰C. When confluent, the growth media is replaced with DMEM/F12 medium (Gibco, Cat. No. 31331-028) containing 10OnM of the CB2 inverse agonist, SR144528, and the cells are incubated at 37⁰C for 30-60mins. Flasks are rinsed twice with 25ml Dulbecco's phosphate buffered saline (PBS, Gibco Cat. No. 14190-094) and then harvested by incubation for lOmins in 10ml of Versene (Gibco, Cat. No. 15040-033). Cells are detached by a sharp blow to the flask and the cell suspension made up to 50ml with PBS and centrifuged at 250xg for 5mins. The cell pellet is re-suspended in 24mls of phenol-red free DMEM/F12 assay buffer (Gibco, Cat. No. 11039-021) and 50μl of cell suspension (approximately 50,000 cells) is added to 96 well plates (Costar, Cat. No. 3904 - clear bottomed black well plates) containing 50μl of test agonist in 2μM forskolin (final assay concentration of lμM FSK). Test agonists are prepared as 1OmM solutions in DMSO and diluted into phenol-red free DMEM/F12 assay buffer containing 2μM forskolin to produce a 20μM solution of test agonist. Subsequent serial dilutions of test agonist are prepared in the assay buffer containing forskolin and each test agonist is routinely examined over a final assay concentration range of lOμM to 1OnM (or lower if required). The plates are mixed on a plate shaker for 5mins (800-1000 rpm) and then centrifuged briefly (5-1Os) at 250xg, placed in a Bioplate without their lids, and incubated for 4-5hr in a humidified 95% air and 5% CO₂ atmosphere at 37⁰C. The 96 well plates are removed from the incubator and placed at RT for 10-15mins before addition of 25μl of Luclite solution, prepared according to the manufacturer's instructions. The plates are sealed with Topseal A (Perkin Elmer, Cat. No. 6005185), mixed on a plate shaker for 5mins (800-1000 rpm) and then centrifuged briefly (5-1Os) at 250xg. Finally, luminescence is measured using a Packard TopCount scintillation counter.

Data Analysis

For each compound maximal inhibition of the forsklin response and the EC50 for this effect is determined. In each experiment the reference agonist HU210 is included and the maximal effect of each test agonist is expressed relative to the maximal effect produced by HU210 to provide an estimate of intrinsic activity. In addition the EC50 of each compound is divided by the EC50 for HU210 to calculate the equipotent molar ratio (EMR) for the test compound.

Reference

Bouaboula M. Dussossoy D. Casellas P. Regulation of peripheral cannabinoid receptor CB2 phosphorylation by the inverse agonist SR 144528. Implications for receptor biological responses. Journal of Biological Chemistry. 274(29):20397-405, 1999

The following examples are illustrative, but not limiting of the embodiments of the present invention.

Abbreviations:

AcOH (acetic acid), Bn (benzyl), Bu, Pr, Me, Et (butyl, propyl, methyl ethyl), DMSO (dimethyl sulfoxide), DCM (dichloromethane), DME (1 ,2-dimethoxyethane), DMF (N,N- dimethylformamide), EDC (l-(3-dimethylaminopropyl)-3-ethylcarbodiimide), EtOAc (ethyl acetate), EtOH (ethanol), HPLC (High pressure liquid chromatography), LC/MS (Liquid chromatography/Mass spectroscopy), MDAP (Mass Directed AutoPurification), MeCN (acetonitrile), MeOH (methanol), NMR (Nuclear Magnetic Resonance (spectrum)), NMP (N- methyl pyrrolidone), SCX (strong cation exchanger e.g. Isolute SCX-2 cartridges), SPE (Solid

Phase Extraction), TFA (Trifluoroacetic acid), THF (tetrahydrofuran), s, d, t, q, quin, m, br (singlet, doublet, triplet, quartet, quintet, multiplet, broad.) Coupling constants (J) are given in Hz. Petroleum ether 40:60 refers to the fraction boiling between 40 and 60⁰C.

Mass Directed Auto-Purification System Hardware

Waters 2525 Binary Gradient Module

Waters 515 Makeup Pump

Waters Pump Control Module Waters 2767 Inject Collect

Waters Column Fluidics Manager

Waters 2996 Photodiode Array Detector

Waters ZQ Mass Spectrometer

Gilson 202 fraction collector Gilson Aspec waste collector

Software

Waters Masslynx version 4 SP2

Column

The columns used are Waters Atlantis, the dimensions of which are 19mm x 100mm (small scale) and 30mm x 100mm (large scale). The stationary phase particle size is 5μm. Solvents

A : Aqueous solvent = Water + 0.1% Formic Acid B : Organic solvent = Acetonitrile + 0.1% Formic Acid Make up solvent = Methanol : Water 80:20 Needle rinse solvent = Methanol

Methods

There are four methods used depending on the analytical retention time of the compound of interest. They all have a 13.5-minute runtime, which comprises of a 10-minute gradient followed by a 3.5 minute column flush and re-equilibration step.

Large/Small Scale 1.0-1.5 = 5-30% B Large/Small Scale 1.5-2.2 = 15-55% B Large/Small Scale 2.2-2.9 = 30-85% B Large/Small Scale 2.9-3.6 = 50-99% B

Large/Small Scale 3.6-5.0 = 80-99% B (in 6 mins)

Flow rate

All of the above methods have a flow rate of either 20mls/min (Small Scale) or 40mls/min (Large Scale)

Analytical LCMS Systems Hardware

Agilent 1100 Gradient Pump Agilent 1100 Autosampler

Agilent 1100 DAD Detector

Agilent 1100 Degasser

Agilent 1100 Oven

Agilent 1100 Controller Waters ZQ Mass Spectrometer

Sedere Sedex 75 or Sedere Sedex 85 or Polymer Labs PL-ELS-2100

Software

Waters MassLynx version 4.0 SP2

Column

The column used is a Waters Atlantis, the dimensions of which are 4.6mm x 50mm. The stationary phase particle size is 3μm.

Solvents

A : Aqueous solvent = Water + 0.05% Formic Acid

B : Organic solvent = Acetonitrile + 0.05% Formic Acid Method

The generic method used has a 5 minute runtir

Time/min %B

0 3

0.1 3

4 97

4.8 97

4.9 3

5.0 3

Flow rate

The above method has a flow rate of 3ml/mins

Conditions used for NMR Hardware

Bruker 400MHz Ultrashield Bruker B-ACS60 Autosampler Bruker Advance 400 Console Software

User interface - NMR Kiosk Controlling software - XWin NMR version 3.0

Conditions used for the Microwave Hardware

Biotage Initiator

Specifications Heating temperature up to 25O⁰C

Microwave radiation 50-300W at 2.45GHz Intermediate 1 : 4-(6-Bromo-3-pyridazinyl)isoquinoline.

To a suspension of 3,6-dibromopyridazine (828mg, 3.5mmol) in ethylene glycol dimethyl ether (10ml) was added isoquinoline-4-boronic acid (675mg, 3.9mmol), tetrakis(triphenylphosphine)palladium(0) (202mg, 175μmol) and sodium carbonate (2N, aqueous) (3ml), the mixture was then heated to 80⁰C for 16 hours whilst under argon. The dark crude reaction mixture was then evaporated to dryness. The solid was suspended in ethyl acetate (20ml) and poured through cellite and again evaporated to dryness. The sample was dissolved in methanol (20ml) and loaded to an SCX cartridge (50g cartridge) and the sample was eluted from the cartridge with 2N ammonia in methanol to yield the title compound as a dark brown solid (620mg). LC/MS m/z [MH⁺] 287 consistent with molecular formula C₁₃H₈ ⁸¹BrN₃

¹H-NMR (CD₃OD) δ 7.80 (IH, dt, J = 9, 1), 7.87-7.91 (IH, m), 7.99 (IH, d, J = 9), 8.10-8.16 (2H, m), 8.26 (IH. D, J = 8), 8.64 (IH, s), 9.41 (IH, s).

Intermediate 2: 3-Bromo-6-(2,3-dichlorophenyl)pyridazine.

To a suspension of 3,6-dibromopyridazine (5g, 21mmol) in ethylene glycol dimethyl ether (100ml) was added 2,3-dichlorophenyl boronic acid (4.03g, 21mmol), tetrakis(triphenylphosphine)palladium(0) (1.2g, 1.05mmol) and sodium carbonate (2N, aqueous) (50ml), the mixture was then heated to 80⁰C for 17 hours whilst under argon. The dark crude reaction mixture was then evaporated to dryness. The residue was partitioned between water (200ml) and dichloromethane (300ml). The aqueous layer was separated and further extracted with dichloromethane (200ml). The combined dichloromethane layers were evaporated and the residue was purified by chromatography (90g of silica) eluting with 75% dichloromethane/ petroleum ether 40:60. The title compound was obtained as a pink coloured solid (680mg). ¹H-NMR (CDCl₃) δ 7.38 (IH, X, J= 8), 7.58-7.63 (2H, m), 7.70-7.76 (2H, m) LC/MS m/z [MH+] 305 consistent with molecular formula C₁₀H₅ ⁸¹Br³⁵Cl₂N₂

Intermediate 3: 3-(2,3-Dichlorophenyl)-6-iodopyridazine.

Copper (I) iodide (9.6mg, 0.05mmol), sodium iodide (300mg, 2mmol), and intermediate 2 (300mg, lmmol) were placed under argon before the addition of 1,4-dioxane (ImI) and røcew/c-trans-N,N- dimethyl-l,2-cyclohexanediamine (16μl, 0. lmmol) The reaction was heated to 100⁰C and stirred for 18 hours. The sample was then allowed to cool and diluted with 0.88 ammonia (5ml) and poured into water (20ml). The suspension was extracted with dichloromethane (2^χ 20ml) which was collected and evaporated. The crude material was purified using chromatography (2Og of silica) eluting with dichloromethane to afford the title compound as a pale pink solid (320mg). ¹H-NMR (CDCl₃) δ 7.35-7.39 (IH, m), 7.51 (IH, d, J = 9), 7.57-7.63 (2H, m), 7.94 (IH, d, J= 9)

Intermediate 4: S-Chloro-ό-iodopyridazine.

To 3,6-dichloropyridazine (4g, 26.8mmol) and sodium iodide (5.4g, 35.9mmol) was added hydriodic acid (20ml) and the mixture was heated to 40⁰C for 18 hours. After cooling to room temperature the material was poured onto ice and stirred in a beaker (500ml). The aqueous acid was basified (pH>12) by addition of sodium hydroxide (cone, 15ml) and water (20ml). Dichloromethane (40ml) was added to the aqueous and collected. The dichloromethane layer was then dried filtered and evaporated to obtain the title compound as a brown solid (6.3g). ¹H-NMR (CDCl₃) 8 7.21 (IH, ά, J= 9), 7.82 (IH, ά, J= 8)

LC/MS m/z [MH⁺] 241 consistent with molecular formula C₄H₂ ³⁵Cl¹²⁷IN₂

Intermediate 5: 3-Chloro-6-(2,3-dichlorophenyl)pyridazine.

To a suspension of intermediate 4 (3.2g, 13.2mmol) in 1,4-dioxane (40ml) was added 2,3- dichlorophenyl boronic acid (2.5g, 13.2mmol), tris(dibenzylideneacetone)-di-palladium(0)- chloroform adduct (725mg, 0.79mmol), potassium fluoride (2.5g, 43.5mmol) and tri-tert- butylphosphine-tetra-fluoroborate (458mg, 1.58mmol), the mixture was then heated to 100⁰C for 1 hour whilst under argon. The dark crude reaction mixture was then evaporated to dryness. The solid was suspended in ethyl acetate (50ml) and poured through cellite and again evaporated to dryness. The sample was then purified by chromatography (9Og of silica) eluting with 10% ethyl acetate/ petroleum ether 40:60. The title compound was obtained as a white solid (2.2g). ¹H-NMR (CDCl₃) δ 7.38 (IH, t, J= 8), 7.59-7.63 (3H, m), 7.83 (IH, d, J= 9) LC/MS m/z [MH+] 259 consistent with molecular formula C₁₀H₅ ³⁵Cl₃N₂

Intermediate 6: 4-(6-Bromo-3-pyridazinyl)hexahydro-l,4-oxazepine.

3,6-Dibromopyridazine (Ig, 4.3mmol) was dissolved in acetonitrile (8ml) in a microwave vial (10ml capacity).To this homomorpholine hydrochloride (1.2g, 8.5mmol) and triethylamine (1.2ml, 8.6mmol) were added and the vial was irradiated for 2 hours at 160⁰C in the microwave. The sample was then extracted between water (150ml) and dichloromethane (150ml). The mixture was poured through a hydrophobic frit collecting the dichloromethane layer which was evaporated to as dry as possible. The sample was then purified by chromatography (40g of silica) eluting with 40% ethyl acetate/ petroleum ether 40:60. (R_f = 0.32 from 50% ethyl acetate/ petroleum ether 40:60) The title compound was obtained as an off white solid (970mg)

¹H-NMR (CDCl₃) δ 1.99-2.05 (2H, m), 3.71-3.74 (2H, m), 3.79 (2H, t, J= 6), 3.84-3.90 (4H, m),

6.71 (IH, d, J= 10), 7.28 (IH, d, J= 10).

LC/MS m/z [MH+] 260 consistent with molecular formula C₉H₁₂ ⁸¹BrN₃O

Intermediate 7: 3-Bromo-6-[(c«)-2,5-dimethyl-l-pyrrolidinyl] pyridazine.

3,6-Dibromopyridazine (3g, 12.7mmol) was dissolved in acetonitrile (7ml) in a microwave vial (10ml capacity). To this 2,5-dimethylpyrrolidine (mixture of cis and trans) (2g, 20.2mmol) and triethylamine (3ml, 20.2mmol) were added and the vial was irradiated for 2 hours at 160⁰C in the microwave. The sample was then extracted between water (100ml) and dichloromethane (100ml).

The mixture was poured through a hydrophobic frit collecting the dichloromethane layer which was evaporated to as dry as possible. The sample was then purified by chromatography (50g of silica) eluting with 30% ethyl acetate/ petroleum ether 40:60 to give the title compound (1.79g)

¹H-NMR (CDCl₃) 1.32 (6H, d, J= 6), 1.75-1.85 (2H, m), 2.07-2.17 (2H, m), 4.05-4.13 (2H, m),

6.54 (IH, t, J= 10), 7.21-7.28 (IH, m).

LC/MS m/z [MH+] 258 consistent with molecular formula C₁₀Hi₄ ⁸¹BrN₃

Intermediate 8: 4-(6-Bromo-3-pyridazinyl)morpholine.

3,6-Dibromopyridazine (1.34g, 5.63mmol) was dissolved in acetonitrile (8ml), to this moφholine

(0.74ml, 8.45mmol) and triethylamine (1.22ml, 8.45mmol) were added and the suspension was irradiated in the microwave to 160⁰C for 80 minutes. The sample was then extracted between water

(100ml) and dichloromethane (100ml). The mixture was poured through a hydrophobic frit collecting the dichloromethane layer which was evaporated to as dry as possible. The sample was then purified by chromatography (4* 10g of silica) eluting with 10% ethyl acetate/ dichloromethane to obtain the title compound (1.23g)

¹H-NMR (CDCl₃) δ 3.59-3.61 (4H, m), 3.82-3.85 (4H, m), 6.79 (IH, d, J= 10), 7.35 (IH, d, J =

10).

LC/MS m/z [MH+] 246 consistent with molecular formula C₈H₁₀ ⁸¹BrN₃O

Example 1: 4-{6-[(cis)-2,5-Dimethyl-l-pyrrolidinyl]-3-pyridazinyl} isoquinoline hydrochloride salt.

Intermediate 1 (200mg 0.7mmol) was dissolved in acetonitrile (3ml) in a microwave vial (5ml capacity), To this 2,5-dimethylpyrrolidine (mixture of cis and trans) (170μl, 1.4mmol) and triethylamine (0.3ml, 2.1mmol) were added and the vial was irradiated for 2 hours at 160⁰C in the microwave. The sample was then evaporated to dryness and purified by chromatography (1Og of silica) eluting with 40% ethyl acetate/ petroleum ether 40:60. (R_f = 0.25 from 50% ethyl acetate/ petroleum ether 40:60) to give the free base of the title compound as an off white solid ¹H-NMR (CDCl₃) δl .35 (6H, d, J = 6), 1.80-1.88 (2H, m), 2.12-2.30 (2H, m), 4.25-4.28 (2H, m), 7.40 (IH, d, J = 9), 7.79-7.82 (IH, m), 7.86-7.95 (2H, m), 8.20-8.31 (2H, m), 8.72 (IH, s), 9.48 (IH, s). The sample was dissolved in 1,4-dioxane (5ml) and treated with hydrogen chloride (IM) solution in diethyl ether (0.5ml) the sample was frozen and dried using freeze drying apparatus to give the title compound an off white solid (120mg).

¹H-NMR ((CD₃)₂SO) δ 1.37 (6H, d, J = 6), 1.87-1.93 (2H, m), 2.18-2.26 (2H, m), 4.34-4.36 (2H, m), 7.83 (IH, d, J= 10), 7.93 (IH, t, J= 8), 8.01-8.06 (IH, m), 8.15-8.33 (2H, m), 8.45 (IH, t, J = 7), 8.81 (IH, s), 9.69 (IH, s) LC/MS m/z [MH+] 305 consistent with molecular formula Ci₉H₂₀N₄

Example 2: 3-(2,3-dichlorophenyl)-6-[(c/s)-2,5-dimethyl-2,5-dihydro-lH-pyrrol-l- yl]pyridazine.

Intermediate 5 (300mg 1.15mmol) was dissolved in acetonitrile (4ml) in a microwave vial (5ml capacity) to this 2,5-dimethyl-3-pyrroline (mixture of cis and trans) (270μl, 2.4mmol) and triethylamine (0.5ml, 3.45mmol) were added and the vial was irradiated for 2 hours at 160⁰C in the microwave. The sample was then extracted between dichloromethane (10ml) and water (10ml) the dichloromethane layer was collected, dried with Na₂SO₄, filtered and evaporated. The crude yellow gummy material was dissolved in methanol (20ml) and loaded to an SCX cartridge (1Og cartridge) the sample was eluted from the cartridge with 2N ammonia in methanol to yield the title compound (70mg).

¹H-NMR (CDCl₃) δ 1.46 (6H, d, J = 6), 4.84-4.89 (2H, m), 5.88 (2H, s), 6.72 (IH, d, J = 9), 7.30- 7.36 (IH, m), 7.48-7.51 (IH, m), 7.60-7.66 (2H, m)

LC/MS m/z [MH+] 320 consistent with molecular formula Ci₆H₁₅ Cl₂N₃

Example 12: 4-{6-[2,3-Bis(trifluoromethyl)phenyl]-3-pyridazinyl} hexahydro-l,4-oxazepine.

To a suspension of intermediate 6 (70mg, 0.27mmol) in ethylene glycol dimethyl ether (5ml) was added 2,3-di-trifluoromethylphenyl-boronic acid (72mg, 0.27mmol), tetrakis(triphenylphosphine)palladium(0) (16mg, 13.5μmol) and sodium carbonate (2N, aqueous) (1.5ml), the mixture was then heated to 80⁰C for overnight whilst under argon. The dark crude reaction mixture was then evaporated to dryness. The solid was suspended in ethyl acetate (50ml) and poured through cellite and again evaporated to dryness. The sample was purified using the mass directed auto-purification system to obtain the title compound (13.7mg). ¹H-NMR (CDCl₃) δ 2.05-2.11 (2H, m), 3.78 (2H, t, J = 6), 3.88-3.94 (4H, m), 3.99-4.02 (2H, m), 6.85 (IH, d, J = 9), 7.31 (IH, d, J = 9), 7.74 (IH, t, J= 8), 7.81 (IH, d, J= 8), 7.93 (IH, d, J= 8). LC/MS m/z [MH+] 392 consistent with molecular formula C₁₇H₁₅ ¹⁹F₆N₃O

Example 13: 5-{6-[(cis)-2,5-Dimethyl-l-pyrrolidinyl]-3-pyridazinyl} isoquinoline.

To a suspension of intermediate 7 (50mg, 0.19mmol) in ethylene glycol dimethyl ether (5ml) was added 5-isoquinolinylboronic acid (69mg, 0.4mmol), tetrakis(triphenylphosphine)palladium(0) (1 lmg, lOμmol) and sodium carbonate (2N, aqueous) (1.5ml), the mixture was then heated to 80⁰C for overnight whilst under argon. The dark crude reaction mixture was then evaporated to dryness. The solid was suspended in ethyl acetate (50ml) and poured through cellite and again evaporated to dryness. The sample was purified using the mass directed auto-purification system and evaporated to obtain the title compound (14mg).

¹H-NMR (CDCl₃) δ 1.43 (6H, d, J = 6), 1.83-1.89 (2H, m), 2.15-2.22 (2H, m), 4.24-4.30 (2H, m), 6.83 (IH, d, J= 10), 7.48 (IH, d, J= 10), 7.71 (IH, t, J= 8), 7.88 (IH, dd, J= 7, 1), 8.04 (IH, d, J = 8), 8.13-8.22 (IH, m), 8.52 (IH, d, J= 6), 9.32 (IH, s).

LC/MS m/z [MH+] 305 consistent with molecular formula Ci₉H₂₀N₄

Example 15: 4-[6-(2,3-Dichlorophenyl)-3-pyridazinyl]morpholine hydrochloride salt.

To a suspension of intermediate 8 (500mg, 2.04mmol) in ethylene glycol dimethyl ether (10ml) was added 2,3-dichlorophenyl-boronic acid (428mg, 2.24mmol), tetrakis(triphenylphosphine)palladium(0) (120mg, 1 lOμmol) and sodium carbonate (2N, aqueous) (5ml), the mixture was then heated to 80⁰C for 16 hours whilst under argon. The dark crude reaction mixture was then evaporated to dryness. The residue was partitioned between water (200ml) and dichloromethane (300ml). The aqueous layer was separated and further extracted with dichloromethane (200ml). The combined dichloromethane layers were dried (Na₂SO₄), filtered and evaporated. The residue was purified by chromatography (50g of silica) eluting with (30%) ethyl acetate/ petroleum ether 40:60 to give the free base of the title compound as an off white solid ¹H- NMR (CDCl₃.) δ 3.70-3.73 (4H, m), 3.87-3.89 (4H, m), 6.95 (IH, d, J= 9), 7.32 (IH, t, J= 8), 7.52- 7.54 (IH, m), 7.60-7.65 (2H, m). The sample was dissolved in 1,4-dioxane (5ml) and treated with hydrogen chloride (IM) solution in diethyl ether (ImI) which was then frozen and dried using freeze drying apparatus to obtain the title compound as an off white solid (160mg). LC retention time 2.47 mins, MS m/z 310 consistent with [MH+] for molecular formula C₁₄H₁₃ ³⁵Cl₂N₃O

Example 20: 3-(2,3-Dichlorophenyl)-6-(lH-pyrrol-l-yl)pyridazine.

Copper (I) iodide (1.5mg, 7.5μmol), potassium phosphate (tribasic) (67mg, 0.3mmol), and intermediate 3 (55mg, 0.15mmol) were placed under argon before the addition of toluene (4ml) and røcew/c-frans-N,N-dimethyl-l,2-cyclohexanediamine (5mg, 30 μmol) and pyrrole (34.6μl 0.5mmol) The reaction was heated to 110⁰C and stirred for 12 hours. The sample was then allowed to cool and diluted with 0.88 ammonia (2ml) and poured into water (10ml). The suspension was extracted with dichloromethane (2* 10ml) which was collected and evaporated. The crude material was purified using chromatography (5g of silica) eluting with (10%) ethyl acetate/ hexane to obtain the title compound as a white solid (30mg).

LC retention time 3.11 mins, MS m/z 290 consistent with [MH+] for molecular formula C₁₄H₉ ³⁵Cl₂N₃

Example 49: Inhalant Formulation

A compound of formula (I) or a pharmaceutically acceptable derivative thereof, (1 mg to 100 mg) is aerosolized from a metered dose inhaler to deliver the desired amount of drug per use.

Example 50 : Tablet Formulation

Tablets/Ingredients Per Tablet

1. Active ingredient 40 mg

(Compound of formula (I) or pharmaceutically acceptable derivative)

2. Corn Starch 20 mg

3. Alginic acid 20 mg

4. Sodium Alginate 20 mg

5. Mg stearate 1.3 mg

Procedure for tablet formulation:

Ingredients 1, 2, 3 and 4 are blended in a suitable mixer/blender. Sufficient water is added portion-wise to the blend with careful mixing after each addition until the mass is of a consistency to permit its conversion to wet granules. The wet mass is converted to granules by passing it through an oscillating granulator using a No. 8 mesh (2.38 mm) screen. The wet granules are then dried in an oven at 140⁰F (6O⁰C) until dry. The dry granules are lubricated with ingredient No. 5, and the lubricated granules are compressed on a suitable tablet press.

Example 51: Parenteral Formulation

A pharmaceutical composition for parenteral administration is prepared by dissolving an appropriate amount of a compound of formula (I) in polyethylene glycol with heating. This solution is then diluted with water for injections Ph Eur. (to 100 ml). The solution is then rendered sterile by filtration through a 0.22 micron membrane filter and sealed in sterile containers.

Claims

Claims

1. A compound selected from a compound of formula (I):

(I) wherein

Ar represents quinolinyl, lsoquinohnyl, either of which can be unsubstituted or substituted with 1, 2 or 3 substituents selected from halo, Ci_-6alkyl, Ci.₆alkoxy, halosubstituted Ci_-6alkyl, halosubstitued C_1-6alkoxy, NH₂, -SO₂C,.₆alkyl, -NHSO₂Ci.₆alkyl, -C_{1 -6} alkylNR³R⁴, or Ar represents phenyl substituted with 1, 2 or 3 substituents at the 2, 3, 5 or 6 positions, which substituents are selected from halo, C].₆alkyl, C_1-6alkoxy, halosubstituted Ci_-6alkyl, halosubstitued C,.₆alkoxy, NH₂, -SO₂C,.₆alkyl, -NHSO₂Ci_-6alkyl, -C_{1 -6} alkylNR³R⁴;

R¹ and R² together with the nitrogen to which they are attached form a 5- to 8- membered heterocyclyl or a 5-membered heteroaryl πng which can be unsubstituted or substituted with 1, 2 or 3 substituents selected from halo, C_1-6 alkyl, cyano, -C_{1 -6} alkylNR³R⁴;

R³ and R⁴ are independently selected from hydrogen, C_1-6 alkyl or together with the N to which they are attached form a pyrrolidinyl, pipeπdmyl or morpholinyl ring; and a pharmaceutically acceptable deπvative thereof.

2. A compound as claimed in claim 1 wherein the substituents for Ar are selected from halo, Ci_-6 alkyl, Ci_-6alkoxy, halosubstituted Ci_-6 alkyl, halosubstituted Ci_₆alkoxy, -SO₂Ci. ₆alkyl, -NHSO₂Ci_-6alkyl and NH₂

3. A compound as claimed in claim 1 or claim 2 wherein R¹ and R² together with the nitrogen to which they are attached form a pyrrolidinyl, pyrrolyl, 2,5,dihydro-lH-pyrrolyl, morpholinyl, thiomorphohnyl, pipendinyl, pyrazolyl, hexahydro-lH-azepmyl, hexahydro-1,4- oxazepinyl or 3-oxa-8-azabicyclo[3.2.1]octanyl, any of which can be unsubstituted or substituted with 1, 2 or 3 substituents selected from halo, Ci_-6 alkyl, ethylpypyrrolidmyl, or cyano.

4. A compound as claimed in any previous claim wherein R³ and R⁴ are independently selected from hydrogen, methyl or together with the N to which they are attached form a pyrohdinyl, pipendinyl or morpholinyl πng.

5. A pharmaceutical composition comprising a compound as claimed in any preceding claim or a pharmaceutically acceptable deπvative thereof.

6. A pharmaceutical composition as claimed in claim 5 further compπsmg a pharmaceutical carrier or diluent thereof.

7. A pharmaceutical composition as claimed in claim 5 or 6 further comprising a second therapeutic agent.

8. A compound of formula (I) as claimed in any one of claims 1-4 or a pharmaceutically acceptable derivative thereof for use in human or veterinary medicine.

9. A compound of formula (1) as claimed in any one of claims 1-4 or a pharmaceutically acceptable derivative thereof for use in the treatment of a condition which is mediated by the activity of cannabinoid 2 receptors.

10. The use of a compound of formula (I) as claimed in any one of claims 1-4 or a pharmaceutically acceptable derivative thereof for the manufacture of a therapeutic agent for the treatment of a condition which is mediated by the activity of cannabinoid 2 receptors.

11. A method of treating mammal for example a human suffering from a condition which is mediated by the activity of cannabinoid 2 receptor which comprises administering to said subject a therapeutically effective amount of a compound of formula (I) as claimed in any one of claims 1-4 or a pharmaceutically acceptable derivative thereof.

12. The compound as claimed in claim 9 or the use as claimed in claim 10 or the method of treatment as claimed in claim 11 wherein the condition which is mediated by the activity of cannabinoid 2 receptor is an immune disorder, an inflammatory disorder, pain, rheumatoid arthritis, multiple sclerosis, osteoarthritis or osteoporosis.

13. The compound as claimed in claim 9 or the use as claimed in claim 10 or the method of treatment as claimed in claim 11 , wherein the pain is selected from inflammatory pain, visceral pain, cancer pain, neuropathic pain, lower back pain, musculoskeletal, post operative pain, acute pain and migraine.