WO2012163456A1

WO2012163456A1 - Novel p2x7r antagonists and their use

Info

Publication number: WO2012163456A1
Application number: PCT/EP2012/001869
Authority: WO
Inventors: Michael Boes
Original assignee: Affectis Pharmaceuticals AG
Current assignee: Affectis Pharmaceuticals AG
Priority date: 2011-05-27
Filing date: 2012-04-30
Publication date: 2012-12-06
Anticipated expiration: 2013-11-27
Also published as: AR086555A1

Abstract

The present application is directed to novel P2X7R antagonists that are N- indol-3-yl-acetamide and N-azaindol-3-yl-acetamide compounds, pharmaceutical compositions comprising the same and their use for the prophylactic or therapeutic treatment of diseases mediated by P2X7R activity.

Description

NOVEL P2X7R ANTAGONISTS AND THEIR USE

The present application relates to novel P2X7R antagonists that are N-indol- 3-yl-acetamide and N-azaindol-3-yl-acetamide compounds, pharmaceutical compositions comprising these compounds and to their use in the

prophylactic and therapeutic treatment of diseases and disorders mediated by P2X7R.

BACKGROUND

P2X7R is an ATP-gated ion channel belonging to the P2X ionotropic channel family. The gene was first isolated from rat brain (Surprenant et al. (1996) 272:735-738) and subsequently from a human monocyte library (Rassendren et al. (1997) J. Biol. Chem. 272:5482-5486; Genbank accession numbers NM_002562, Y09561) by virtue of its sequence homology with the other members of the P2X family. It was later found that P2X7R corresponded to the unidentified P2Z receptor which mediates the permeabilising action of ATP on mast cells and macrophages (Dahlqvist and Diamant (1974) Acta Physiol. Scand. 34:368-384; Steinberg and Silverstein (1987) J. Biol. Chem. 262:3118-3122; Gordon (1986) Biochem. J. 233:309-319). The P2X7R has two hydrophobic membrane-spanning domains, an extracellular loop, and forms transmembrane ion channels. P2X7R bears a pharmacological profile markedly different from other P2X homo- or heteromers (North and

Surprenant (2000) Annual Rev. Pharmacology Toxicology 40:563-580).

P2X7R requires levels of ATP in excess of 1 mM to achieve activation, whereas other P2X receptors activate at ATP concentrations of <100 μΜ (Steinberg et al. (1987) J. Biol. Chem. 262:8884-8888; Greenberg et al.

(1988) J. Biol. Chem. 263:10337-10343). While all P2X receptors

demonstrate non-selective channel-like properties following ligation, the channels formed by the P2X7R can rapidly transform into pores that can allow the passage of molecules of up to 900 Dalton (Virginio et al. (1999) J. Physiol. 519:335-346). P2X7R is expressed in haematopoietic cells, mast cells, lymphocytes, erythrocytes, fibroblast, Langerhans cells, and macrophages (Surprenant et al., 1996, Science 272:3118-3122). In the central nervous system, P2X7R expression has been reported in glial cells, Schwann cells, astrocytes, as well as in neurons (Ferrari et al. (1996) J. Immunol 156:1531-1539; Collo et al. (1997) Neuropharmacology 36: 1277-1283; Anderson and Nedergaard (2006) Trends Neuroscien 29: 257-262). P2X7R is involved in the regulation of the immune function and inflammatory response. Activation of P2X7R by ATP in macrophages is associated with mitogenic stimulation of T cells (Baricordi et al. (1996) Blood 87:682-690), the release of cytokines (Griffiths et al. (1995) J. Immol. 154:2821-2828), and formation of macrophage polykarions (Falzoni et al. (1995) J. Clin. Invest. 95:1207-1216). P2X7R is involved in the processing and release of active interleukin-1beta (IL-1 β) from proinflammatory cells (Perregaux and Gabel (1998) J Biol Chem 269:15195-15203; Ferrari et al., (2006) J Immunol 176: 3877-3883). Stimulation of the P2X7R by ATP can also result in apoptosis and cell death by triggering the formation of non-selective plasma membrane pores (Di Virgilio et al. (1998) Cell Death Differ. 5:191-199).

Upregulation of P2X7R has been observed during ischemic damage and necrosis induced by occlusion of middle cerebral artery in rat brain (Collo et al. (1997) Neuropharmacol 36:1277-1283). Recent studies indicate a role of P2X7R in the generation of superoxide in microglia, and upregulation of P2X7R has been detected around amyloid plaques in a transgenic mouse models for Alzheimer's disease (Parvathenani et al. (2003) J Biol Chem 278:13300-13317) and in multiple sclerosis lesions from autopsy brain sections (Narcisse et al. (2005) Glia, 49:245-258). Inhibition of P2X7 can improve symptoms in a model of multiple sclerosis (Matute et al. (2007) J Neurosci 27:9525-9533). Studies from mice lacking P2X7R resulted in absence of inflammatory and neuropathic hypersensitivity to mechanical and thermal stimuli, indicating a link between P2X7R and inflammatory and neuropathic pain (Chessell et al. (2005) Pain 114:386-396). Antagonists of P2X7R significantly improved functional recovery and decreased cell death in spinal cord injury in animal models (Wang et al. (2004) Nature Med 10:B21-B27). P2X7 has also been shown to be involved in depression and anxiety (Basso et al. (2009) Behav Brain Res 198:83-90; WO 2009/019503); bipolar affective disorders (Barden et al. (2006) Am J Med Genet B 141 B:374-382); glaucoma (Resta et al.

(2007) Eur J Neurosci 25:2741-2754); inflammatory bowel diseases (Cesaro et al. (2010) Am J Physiol-Gastr L 299:G32-42); cancer (Sun (2010) Mol Neurobiol 41:351-355; Di Virgilio et al. (2009) Purinergic Signal 5:251-256); and graft-versus-host disease (Wilhelm et al. (2010) Nat Med 16:1434-1438).

Compounds which modulate P2X7R have been reported. For example, Brilliant Blue (Jiang et al., Mol. Phamacol. 58 (2000), 82-88), the

isoquinolines 1-[N,O-Bis(5-isoquinolinesulfonyl)-N-methyl-L-tyrosyl]-4- phenylpiperazine and N-[1-[N-methyl-p-(5 isoquinolinesulfonyl) benzyl]-2-(4- phenylpiperazine)ethyl]-5-isoquinolinesulfonamide (Humphreys et al., Mol. Pharmacol., 54 (1998), 22-32), adamantane derivatives (WO 99/29660, WO 99/29661 , WO 00/61569, WO 01/42194, WO 01/44170, WO 01/44213, WO 01/94338, WO 03/041707, WO 03/042190, WO 03/080579, WO 04/074224, WO 05/014529, WO 06/025783, WO 06/059945), piperidine and piperazine compounds (WO 01/44213, WO 01/46200, WO 08/005368;

WO 2010/125103), benzamide and heteroarylamide compounds (WO

03/042191 , WO 04/058731 , WO 04/058270, WO 04/099146, WO 05/019182, WO 06/003500, WO 06/003513, WO 06/067444), substituted tyrosine derivatives (WO 00/71529, WO 03/047515, WO 03/059353), imidazole compounds (WO 05/014555), amino-tetrazoles compounds (WO 05/111003), cyanoamidine (WO 06/017406), bicycloheteroaryl derivatives

(WO 05/009968, WO 06/102588, WO 06/102610, WO 07/028022, WO

07/109154, WO 07/109160, WO 07/109172, WO 07/109182, WO 07/109192, O 07/109201), acylhydrazide (WO 06/110516), and other examples (WO 9/29686, WO 04/106305, WO 05/039590, WO 06/080884, WO 06/086229, O 06/136004, WO 07/025366, WO 07/056046, WO 07/056091 , WO

07/141267, WO 07/141269, WO 08/003697; WO 2008/138876;

WO 2008/125600; WO 2008/124153; WO 2008/119825; WO 2008/013494; WO 2009/070116; WO 2009/053459 WO 2009/077559; WO 2009/132000; WO 2009/118175; WO 2010/118921) are antagonists of P2X7R while

Oxidized ATP (oATP) acts as an irreversible inhibitor of the receptor (Chen et al., J. Biol. Chem., 268 (1993), 8199-8203). Methods for synthesising oxetanes are disclosed in Wuitschik et al., J. Med. Chem., 53 (2010) 3227- 3246.

Consequently, there is strong evidence that compounds acting on P2X7R can be used in the treatment of pain, inflammatory processes, and

degenerative conditions associated with disease states such as rheumatoid arthritis, osteoarthritis, psoriasis, allergic dermatitis, asthma, chronic obstructive pulmonary disease, airways hyper-responsiveness, septic shock, glomerulonephritis, irritable bowel disease, inflammatory bowel disease, Crohn's disease, ulcerative colitis, atherosclerosis, growth and metastases of malignant cells, myoblastic leukaemia, diabetes, Alzheimer's disease, Parkinson's disease, multiple sclerosis, glaucoma, age-related macular degeneration, uveitis, neuropathic pain, depression, bipolar affective disorders, anxiety, meningitis, traumatic brain injury, acute spinal cord injury, neuropathic pain, osteoporosis, burn injury, ischemic heart disease, myocardial infarction, stroke, and varicose veins.

Thus, the object on the present invention is to provide a novel series of compound which can inhibit P2X7R activity and can be used in the treatment of the above-mentioned diseases. DETAILED DESCRIPTION OF THE INVENTION

The present invention relates to novel P2X7R antagonists that are N-indol-3- yl-acetamide and N-azaindol-3-yl-acetamide compounds represented by the general formula (I):

R-i is a mono- or bicycloalkylalkyl group;

R₂ is selected from hydrogen, -OH, -NH2, or halogen (i.e., CI, F, Br or i);

R3. R , R5, F?6 are at each occurrence independently selected from hydrogen, halogen (i.e., CI, F, Br or I), d-C₆ alkyl, hydroxy, Ci-C₆ alkoxy, cyano, or trifluoromethyl; n equals 1 or 2; a, b, c, d, x are at each occurrence independently selected from carbon, or nitrogen; or a pharmaceutically acceptable salt or solvate thereof. Compounds of Formula (I), wherein R₃, R4, Rs, R6 are at each occurrence independently selected from hydrogen, halogen, methyl, hydroxy, methoxy, cyano, or trifluoromethyl. Compounds of Formula (I), wherein Ri is a mono- or bicycloalkylalkyi group selected from cyclopentylmethyl, cyclopentylethyl, cyclohexylmethyl, cyclohexylethyl, cycloheptylmethyl, cycloheptylethyl, adamantanyl-methyl, or adamantanyl-ethyl thereof are preferred. Preferred are also compounds, wherein R₂ is selected from -OH or fluoride. Furthermore, it is preferred that at least two of R₃, R₄, R₅ and R₆ are hydrogen. In further preferred embodiments, R₄ and R₆ are H, wherein R₃ and R₅ are defined as above, or R₄-R₆ are H, wherein R₃ is defined as above, wherein Ci-C₆ alkyl is preferably methyl and C C₆ alkoxy is preferably methoxy. If necessitated by valency, R₃-R6 may also be absent.

Additionally, it is preferred that a, b, c, and d are CH or one of a, b, c and d is N. Preferred are also compounds wherein n=2 and x is N.

Examples of novel N-indol-3-yl-acetamide and N-azaindol-3-yl-acetamide compounds are disclosed in examples 1-66. The invention further relates to a compound of Formula (I) or a

pharmaceutically acceptable salt or solvate thereof, being:

N-(4-chloro-1-((3-hydroxyoxetan-3-yl)methyl)-1 H-indol-3-yl)-2- cyclopentylacetamide

- N-(4-bromo-1 -((3-hydroxyoxetan-3-yl)methyl)-1 H-indol-3-yl)-2- cyclopentylacetamide N-(4-chloro-1 -((3-hydroxyoxetan-3-yl)methyl)-1 H-indol-3-yl)-2- cyclohexylacetamide

N-(4-bromo-1 -((3-hydroxyoxetan-3-yl)methyl)-1 H-indol-3-yl)-2- cyclohexylacetamide

N-(4-chloro-1 -((3-hydroxyoxetan-3-yl)methyl)-1 H-indol-3-yl)-2- cycloheptylacetamide

N-(4-bromo-1-((3-hydroxyoxetan-3-yl)methyl)-1 H-indol-3-yl)-2- cycloheptylacetamide

N-(4-chloro-1-((3-hydroxyoxetan-3-yl)methyl)-1H-indol-3-yl)-2- adamantane-1yl-acetamide

N-(4-bromo-1-((3-hydroxyoxetan-3-yl)methyl)-1 H-indol-3-yl)-2- adamantane-1yl-acetamide

N-(4-chloro-1 -((3-hydroxyoxetan-3-yl)methyl)-1 H-indol-3-yl)-3- cyclopentylpropanamide

N-(4-bromo-1-((3-hydroxyoxetan-3-yl)methyl)-1 H-indol-3-yl)-3- cyclopentylpropanamide

N-(4-chloro-1 -((3-hydroxyoxetan-3-yl)methyl)-1 H-indol-3-yl)-3- cyclohexylpropanamide

N-(4-bromo-1 -((3-hydroxyoxetan-3-yl)methyl)-1 H-indol-3-yl)-3- cyclohexylpropanamide

N-(4-chloro-1 -((3-hydroxyoxetan-3-yl)methyl)-1 H-indol-3-yl)-3- cycloheptylpropanamide

N-(4-bromo-1 -((3-hydroxyoxetan-3-yl)methyl)-1 H-indol-3-yl)-3- cycloheptylpropanamide

N-(4-chloro-1-((3-hydroxyoxetan-3-yl)methyl)-1H-indol-3-yl)-3- adamantane-1 yl-propanamide

N-(4-bromo-1-((3-hydroxyoxetan-3-yl)methyl)-1H-indol-3-yl)-3- adamantane-1yl-propanamide 2-cycloheptyl-N-(1-((3-hydrpxyoxetan-3-yl)methyl)-4-methyl-1H-indol-3- yl)acetamide

2- cycloheptyl-N-(4-hydroxy-1-((3-hydroxyoxetan-3-yl)methyl)-1H-indol-

3- yl)acetamide

2- cycloheptyl-N-(1-((3-hydroxyoxetan-3-yl)methyl)-4-methoxy-1 H-indol-

3- yl)acetamide

2-cycloheptyl-N-(4-(trifluoromethyl)-1-((3-hydroxyoxetan-3-yl)methyl)- 1 H-indol-3-yl)acetamide

N-(4-cyano-1 -((3-hydroxyoxetan-3-yl)methyl)-1 H-indol-3-yl)-2- cycloheptylacetamide

N-(4-chloro-1-(2-(3-hydroxyoxetan-3-yl)ethyl)-1H-indol-3-yl)-2- cyclopentylacetamide

N-(4-bromo-1 -(2-(3-hydroxyoxetan-3-yl)ethyl)-1 H-indol-3-yl)-2- cyclopentylacetamide

N-(4-chloro-1-(2-(3-hydroxyoxetan-3-yl)ethyl)-1 H-indol-3-yl)-2- cyclohexylacetamide

N-(4-bromo-1-(2-(3-hydroxyoxetan-3-yl)ethyl)-1 H-indol-3-yl)-2- cyclohexylacetamide

N-(4-chloro-1-(2-(3-hydroxyoxetan-3-yl)ethyl)-1H-indol-3-yl)-2- cycloheptylacetamide

N-(4-bromo-1-(2-(3-hydroxyoxetan-3-yl)ethyl)-1H-indol-3-yl)-2- cycloheptylacetamide

N-(4-chloro-1-(2-(3-hydroxyoxetan-3-yl)ethyl)-1H-indol-3-yl)-3- cyclopentylpropanamide

N-(4-bromo-1 -(2-(3-hydroxyoxetan-3-yl)ethyl)-1 H-indol-3-yl)-2- cycloheptylacetamide

N-(4-chloro-1-(2-(3-hydroxyoxetan-3-yl)ethyl)-1H-indol-3-yl)-2- adamantane-1yl-acetamide

N-(4-bromo-1-(2-(3-hydroxyoxetan-3-yl)ethyl)-1 H-indol-3-yl)-2- adamantane-1yl-acetamide

N-(4-chloro-1-(2-(3-hydroxyoxetan-3-yl)ethyl)-1H-indol-3-yl)-3- cyclopentylpropanamide N-(4-bromo-1-(2-(3-hydroxyoxetan-3-yl)ethyl)-1 H-indol-3-yl)-3- cyclopentylpropanamide

N-(4-chloro-1-(2-(3-hydroxyoxetan-3-yl)ethyl)-1 H-indol-3-yl)-3- cyclohexylpropanamide

N-(4-bromo-1-(2-(3-hydroxyoxetan-3-yl)ethyl)-1 H-indol-3-yl)-3- cyclohexylpropanamide

N-(4-chloro-1-(2-(3-hydroxyoxetan-3-yl)ethyl)-1 H-indol-3-yl)-3- cycloheptylpropanamide

N-(4-bromo-1-(2-(3-hydroxyoxetan-3-yl)ethyl)-1 H-indol-3-yl)-3- cycloheptylpropanamide

N-(4-chloro-1-(2-(3-hydroxyoxetan-3-yl)ethyl)-1 H-indol-3-yl)-3- adamantane-1 yl-propanamide

N-(4-bromo-1-(2-(3-hydroxyoxetan-3-yl)ethyl)-1 H-indol-3-yl)-3- adamantane-1yl-propanamide

3-cycloheptyl-N-(1-(2-(3-hydroxyoxetan-3-yl)ethyl)-4-methyl-1H-indol-3 yl)propanamide

3-cycloheptyl-N-(4-hydroxy-1-(2-(3-hydroxyoxetan-3-yl)ethyl)-1 H-indol- 3-yl)propanamide

3-cycloheptyl-N-(1-(2-(3-hydroxyoxetan-3-yl)ethyl)-4-methoxy-1 H-indol- 3-yl)propanamide

3-cycloheptyl-N-(4-(trifluoromethyl)-1-(2-(3-hydroxyoxetan-3-yl)ethyl)- 1 H-indol-3-yl)propanamide

N-(4-cyano-1-(2-(3-hydroxyoxetan-3-yl)ethyl)-1 H-indol-3-yl)-3- cycloheptylpropanamide

N-(4-chloro-1 -((oxetan-3-yl)methyl)-1 H-indol-3-yl)-2- cycloheptylacetamide

N-(4-bromo-1 -((oxetan-3-yl)methyl)-1 H-indol-3-yl)-2- cycloheptylacetamide

N-(4-chloro-1 -((oxetan-3-yl)methyl)-1 H-indol-3-yl)-3- cycloheptylpropanamide

N-(4-bromo-1-((oxetan-3-yl)methyl)-1 H-indol-3-yl)-3- cycloheptylpropanamide N-(4-chloro-1 -((3-fluorooxetan-3-yl)methyl)-1 H-indoI-3-yl)-2- cycloheptylacetamide

N-(4-bromo-1-((3-fluorooxetan-3-yl)methyl)-1H-indol-3-yl)-2- cycloheptylacetamide

N-(4-chloro-1 -((3-fIuorooxetan-3-yl)methyl)-1 H-indol-3-yl)-3- cycloheptylpropanamide

N-(4-bromo-1 -((3-fluorooxetan-3-yl)methyl)-1 H-indol-3-yl)-3- cycloheptylpropanamide

N-(4-chloro-1-((3-hydroxyoxetan-3-yl)methyl)-1 H-pyrrolo[2,3-b]pyridin-3 yl)-2-cycloheptylacetamide

N-(4-bromo-1-((3-hydroxyoxetan-3-yl)methyl)-1H-pyrrolo[2,3-b]pyridin- 3-yl)-2-cycloheptylacetamide

N-(6-bromo-4-chloro-1-((3-hydroxyoxetan-3-yl)methyl)-1 H-indol-3-yl)-2- cycloheptylacetamide

N-(4-bromo-6-chloro-1-((3-hydroxyoxetan-3-yl)methyl)-1H-indol-3-yl)-2- cycloheptylacetamide

N-(4-chloro-1-(2-(oxetan-3-yl)ethyl)-1H-indol-3-yl)-2- cycloheptylacetamide

N-(4-bromo-1-(2-(oxetan-3-yl)ethyl)-1 H-indol-3-yl)-2- cycloheptylacetamide

N-(4-chloro-1 -(2-(oxetan-3-yl)ethyl)-1 H-indol-3-yl)-3- cycloheptylpropanamide

N-(4-bromo-1-(2-(oxetan-3-yl)ethyl)-1 H-indol-3-yl)-3- cycloheptylpropanamide

N-(4-chloro-1 -(2-(3-fluorooxetan-3-yl)ethyl)-1 H-indol-3-yl)-2- cycloheptylacetamide

N-(4-bromo-1 -(2-(3-fluorooxetan-3-yl)ethyl)-1 H-indol-3-yl)-2- cycloheptylacetamide

N-(4-chloro-1-(2-(3-fluorooxetan-3-yl)ethyl)-1 H-indol-3-yl)-3- cycloheptylpropanamide

N-(4-bromo-1-(2-(3-fluorooxetan-3-yl)ethyl)-1 H-indol-3-yl)-3- cycloheptylpropanamide N-(4-chloro-1 -(2-(3-hydroxyoxetan-3-yl)ethyl)-1 H-indazol-3-yl)-2- cycloheptylacetamide

N-(4-bromo-1-(2-(3-hydroxyoxetan-3-yl)ethyl)- H-indazol-3-yl)-2- cycloheptylacetamide

The present invention also includes isotopically-labelled compounds, which are identical to those recited in Formula (I), 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 25 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 and chlorine, such as ²H, ³H, ¹³C, ¹⁴C, ¹⁵N, ¹⁸0, ¹⁷0, 31 p 32p 35_{S >} i8p _and 35^ _reSp_ectj_ve|_y Compounds of the present invention, prodrugs thereof, and pharmaceutically acceptable salts of said compounds or of said prodrugs which contain the aforementioned isotopes and/or other isotopes of other atoms are within the scope of this invention. Certain isotopically-labelled compounds of the present invention, for example those into which radioactive isotopes such as ³H and 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. 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-labelled compounds of Formula (I) of this invention and prodrugs thereof can generally be prepared by carrying out the procedures disclosed in the Examples below, by substituting a readily available isotopically-labelled reagent for a non-isotopically-labelled reagent. Pharmaceutically acceptable salts include those formed with anions such as those derived from hydrochloric, phosphoric, acetic, oxalic, tartaric acids, etc., and those formed with cations such as those derived from sodium, potassium, ammonium, calcium, ferric hydroxides, isopropylamine, triethylamine, 2-ethylamino ethanol, histidine and procaine.

Examples of further pharmaceutically acceptable, nontoxic acid addition salts are salts formed with inorganic acids such as hydrochloric acid, hydrobromic acid, phosphoric acid, sulphuric acid and perchloric acid or with organic acids such as acetic acid, oxalic acid, maleic acid, tartaric acid, citric acid, succinic acid or malonic acid or by using other methods used in the art such as ion exchange. Other pharmaceutically acceptable salts include adipate, alginate, ascorbate, aspartate, benzenesulfonate, benzoate, bisulfate, borate, butyrate, camphorate, camphorsulfonate, citrate, cyclopentaneproprionate, digluconate, dodecylsulfate, ethanesulfonate, formate, fumarate,

glucoheptonate, glycerophosphate, gluconate, hernisulfate, heptanoate, hexanoate, hydroiodide, 2-hydroxy-ethanesulfonate, lactobionate, lactate, laurate, lauryl sulphate, malate, maleate, malonate, methanesulfonate, 2- naphthalenesulfonate, nicotinate, nitrate, oieate, oxalate, palmitate, pamoate, pectinate, persulfate, 3-phenylpropionate, phosphate, picrate, pivalate, propionate, stearate, succinate, sulphate, tartrate, thiocyanate, p- toluenesulfonate, undecanoate, valerate salts, and the like. Representative alkali or alkaline earth metal salts include sodium, lithium, potassium, calcium, magnesium, and the like. Further pharmaceutically acceptable salts include, when appropriate, nontoxic ammonium, quaternary ammonium, and amine cations formed using counterions such as halide, hydroxide, carboxylate, sulphate, phosphate, nitrate, loweralkyl sulfonate and aryl sulfonate. In a further embodiment the present application is directed to a

pharmaceutical composition comprising a compound of Formula (I) of the present invention. The pharmaceutical composition according to the present invention may further comprise an additional active compound in separate or unit dosage form for simultaneous or sequential administration.

The compounds of Formula (I) or a pharmaceutically acceptable salt thereof can be used in the manufacture of a medicament for the prophylactic or therapeutic treatment of any disease state in a human, or other mammal, which is exacerbated or caused by excessive or unregulated cytokine production by such mammal's cells, such as but not limited to monocytes and/or macrophages.

The present invention also relates to the treatment of an IL-1 or cytokine mediated condition.

As defined herein, an "IL-1 mediated condition" and "cytokine mediated condition" includes, but is not limited to, a disease or disorder selected from the group consisting of arthritis (including psoriatic arthritis, Reiter's syndrome, rheumatoid arthritis, gout, traumatic arthritis, rubella arthritis, rheumatoid spondylitis, osteoarthritis, gouty arthritis and acute synovitis), inflammatory bowel disease, Crohn's disease, emphysema, acute respiratory distress syndrome, adult respiratory distress syndrome, asthma, bronchitis, chronic obstructive pulmonary disease, chronic pulmonary inflammatory disease, silicosis, pulmonary sarcoidosis, allergic reactions, allergic contact hypersensitivity, eczema, contact dermatitis, psoriasis, sunburn, cancer, tissue ulceration, restenosis, periodontal disease, epidermolysis bullosa, osteoporosis, bone resorption disease, loosening of artificial joint implants, atherosclerosis, aortic aneurysm, congestive heart failure, myocardial infarction, stroke, cerebral ischemia, head trauma, neurotrauma, spinal cord injury, neuro- degenerative disorders, Alzheimer's disease, Parkinson's disease, glaucoma, age-related macular degeneration, uveitis, neuropathic pain, migraine, depression, peripheral neuropathy, pain, cerebral amyloid angiopathy, nootropic or cognition enhancement, amyotrophic lateral sclerosis, multiple sclerosis, ocular angiogenesis, corneal injury, macular degeneration, corneal scarring, scleritis, abnormal wound healing, burns, autoimmune disorders, Huntington's disease, diabetes, AIDS, cachexia, sepsis, septic shock, endotoxic shock, conjunctivitis shock, gram negative sepsis, toxic shock syndrome, cerebral malaria, cardiac and renal reperfusion injury, thrombosis, glomerularonephritis, graft vs. host reaction, allograft rejection, organ transplant toxicity, ulcerative colitis, or muscle degeneration, in a mammal, including a human, comprising administering to said mammal an amount of a compound to Formula (I), effective in treating such a condition.

The present invention relates to a pharmaceutical composition for the treatment of an IL-1 mediated condition in a mammal, including a human, comprising an amount of a compound of Formula (I), effective in treating such a condition and a pharmaceutically acceptable carrier.

The compounds of the invention are useful for the treatment of rheumatoid arthritis, osteoarthritis, psoriasis, allergic dermatitis, asthma, chronic obstructive pulmonary disease (COPD), hyperresponsiveness of the airway, septic shock, glomerulonephritis, irritable bowel disease, Crohn's disease, ulcerative colitis, atherosclerosis, growth and metastases of malignant cells, myoblastic leukemia, diabetes, Alzheimer's disease, meningitis,

osteoporosis, burn injury, ischemic heart disease, stroke and varicose veins.

In another aspect, the invention further provides a pharmaceutical

composition for treating osteoarthritis which comprises a therapeutically effective amount of a compound of Formula (I), or a pharmaceutically acceptable salt or solvate thereof, as hereinbefore defined.

The invention further provides a pharmaceutical composition for effecting immunosuppression (e.g., in the treatment of rheumatoid arthritis, irritable bowel disease, atherosclerosis or psoriasis) which comprises a

therapeutically effective amount of a compound of Formula (I), or a pharmaceutically acceptable salt or solvate thereof, as hereinbefore defined. The invention also provides a pharmaceutical composition for treating an obstructive airways disease (e.g. asthma or COPD) which comprises a therapeutically effective amount of a compound of Formula (I), or a pharmaceutically acceptable salt or solvate thereof, as hereinbefore defined. The present invention yet further provides a pharmaceutical composition for treating a mammal susceptible to or afflicted with conditions that are causally related to abnormal activity of the P2X7 receptor, such as neurodegenerative diseases and disorders including, for example, Parkinson's disease, multiple sclerosis, glaucoma, age-related macular degeneration, uveitis, neuropathic pain, diseases and disorders which are mediated by or result in

neuromfiammation such as, for example traumatic brain injury and

encephalitis; centrally-mediated neuropsychiatric diseases and disorders such as, for example depression mania, bipolar disease, anxiety,

schizophrenia, eating disorders, sleep disorders and cognition disorders, epilepsy and seizure disorders comprising a therapeutically effective amount of a compound of Formula (I), or a pharmaceutically acceptable salt or solvate thereof, as hereinbefore defined.

In particular embodiment the pharmaceutical composition according to the present invention may be used for the treatment of affective disorders. In a preferred embodiment the affective disorder is selected from depression, anxiety, bipolar disorder and schizophrenia.

In an alternative embodiment the pharmaceutical composition according to the present invention is useful for the treatment of neurodegenerative diseases and disorders, diseases and disorders which are mediated by or result in neuroinflammation and centrally-mediated neuropsychiatric diseases and disorders.

Furthermore, the pharmaceutical composition according to the present invention may particular/ be useful for the treatment of pain, inflammatory processes, and degenerative conditions. In a more preferred embodiment the inflammatory process is selected from rheumatoid arthritis, osteoporosis and chronic obstructive pulmonary disease.

Moreover, the pharmaceutical composition according to the present invention may be used for the treatment of neuropathic pain.

Dosage, pharmaceutical preparation and delivery of a compound of Formula (I) for use in accordance with the present invention can be formulated in conventional manner according to methods found in the art, using one or more physiological carriers or excipient, see, for example Ansel et al.,

"Pharmaceutical Dosage Forms and Drug Delivery Systems", 7th edition, Lippincott Williams & Wilkins Publishers, 1999. Thus, the P2X7R modulating agent and its physiologically acceptable salts and solvates can be formulated for administration by inhalation, insufflation (either through the mouth, or nose), oral, buccal, parenteral, or rectal administration.

For oral administration, the pharmaceutical composition of a compound of Formula (I) can take the form of, for example, tablets or capsules prepared by conventional means with pharmaceutical acceptable excipients such as binding agents (e.g., pregelatinised maize starch, polyvinylpyrrolidone, hydroxypropyl methylcellulose), fillers (e.g., lactose, microcrystalline cellulose, calcium hydrogen phosphate), lubricants (e.g., magnesium stearate, talc, silica), disintegrants (e.g., potato starch, sodium starch glycolate), or wetting agents (e.g., sodium lauryl sulphate). The

pharmaceutical composition can be administered with a physiologically acceptable carrier to a patient, as described herein. In a specific

embodiment, the term "pharmaceutically acceptable" means approved by a regulatory agency or other generally recognized pharmacopoeia for use in animals, and more particularly in humans. The term "carrier" refers to a diluent, adjuvant, excipient, or vehicle with which the therapeutic is

administered. Such pharmaceutical carriers can be sterile liquids, such as water and oils, including those of petroleum, animal, vegetable or synthetic origin, such as peanut oil, soybean oil, mineral oil, sesame oil and the like. Water is a preferred carrier when the pharmaceutical composition is administered intravenously. Saline solutions and aqueous dextrose and glycerol solutions can also be employed as liquid carriers, particularly for injectable solutions. Suitable pharmaceutical excipients include starch, glucose, lactose, sucrose, gelatin, malt, rice, flour, chalk, silica gel, sodium stearate, glycerol monostearate, talc, sodium ion, dried skim milk, glycerol, propylene, glycol, water, ethanol and the like. The composition, if desired, can also contain minor amounts of wetting or emulsifying agents, or pH buffering agents. These compositions can be in the form of solutions, suspensions, emulsion, tablets, pills, capsules, powders, sustained-release formulations and the like. The composition can be formulated as a

suppository, with traditional binders and carriers such as triglycerides. Oral formulation can include standard carriers such as pharmaceutical grades of mannitol, lactose, starch, magnesium stearate, sodium saccharine, cellulose, magnesium carbonate, etc. Examples of suitable pharmaceutical carriers are described in "Remington's Pharmaceutical Sciences" by E.W. Martin. Such compositions will contain a therapeutically effective amount of the

aforementioned compounds, preferably in purified form, together with a suitable amount of carrier so as to provide the form for proper administration to the patient. The formulation should suit the mode of administration.

Liquid preparations for oral administration can be in the form of, for example, solutions, syrups, or suspensions, or can be presented as a dry product for constitution with water or other suitable vehicle before use. Such liquid preparation can be prepared by conventional means with pharmaceutically acceptable additives such as suspending agents (e.g., sorbitol, syrup, cellulose derivatives, hydrogenated edible fats), emulsifying agents (e.g., lecithin, acacia), non-aqueous vehicles (e.g., almond oil, oily esters, ethyl alcohol, fractionated vegetable oils), preservatives (e.g., methyl or propyl-p- hydroxycarbonates, soric acids). The preparations can also contain buffer salts, flavouring, coloring and sweetening agents as deemed appropriate. Preparations for oral administration can be suitably formulated to give controlled release of a compound of Formula (I).

For administration by inhalation, a compound of Formula (I) of the present invention is conveniently delivered in the form of an aerosol spray

presentation from a pressurised pack or a nebulizer, with the use of a suitable propellant (e.g., dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide or other suitable gas). In the case of a pressurised aerosol, the dosage unit can be determined by providing a valve to deliver a metered amount. Capsules and cartridges of, for example, gelatine, for use in an inhaler or insufflator can be formulated containing a powder mix of a compound of Formula (I) and a suitable powder base such as lactose or starch.

A compound of Formula (I) of the present invention can be formulated for parenteral administration by injection, for example, by bolus injection or continuous infusion. Site of injections include intra-venous, intra-peritoneal or sub-cutaneous. Formulations for injection can be presented in units dosage form (e.g., in phial, in multi-dose container), and with an added preservative. A compound of Formula (I) of the present invention can take such forms as suspensions, solutions or emulsions in oily or aqueous vehicles, and can contain formulatory agents such as suspending, stabilizing, or dispersing agents. Alternatively, the agent can be in powder form for constitution with a suitable vehicle (e.g., sterile pyrogen-free water) before use. Typically, compositions for intravenous administration are solutions in sterile isotonic aqueous buffer. Where necessary, the composition can also include a solubilizing agent and a local anesthetic such as lignocaine to ease pain at the site of the injection. Generally, the ingredients are supplied either separately or mixed together in unit dosage form, for example, as a dry lyophilised powder or water free concentrate in a hermetically sealed container such as an ampoule or sachette indicating the quantity of active agent. Where the composition is to be administered by infusion, it can be dispensed with an infusion bottle containing sterile pharmaceutical grade water or saline. Where the composition is administered by injection, an ampoule of sterile water for injection or saline can be provided so that the ingredients can be mixed prior to administration.

A compound of Formula (I) of the present invention can be formulated for transdermal administration. Transdermal compositions are typically formulated as a topical ointment or cream containing the active ingredient(s), generally in an amount ranging from about 0.01 to about 20% by weight, preferably from about 0.1 to about 20% by weight, preferably from about 0.1 to about 10% by weight, and more preferably from about 0.5 to about 15% by weight. When formulated as a ointment, the active ingredients will typically be combined with either a paraffinic or a water-miscible ointment base.

Alternatively, the active ingredients may be formulated in a cream with, for example an oil-in-water cream base. Such transdermal formulations are well- known in the art and generally include additional ingredients to enhance the dermal penetration of stability of the active ingredients or the formulation. All such known transdermal formulations and ingredients are included within the scope of this invention. The compounds of this invention can also be administered by a transdermal device. Accordingly, transdermal

administration can be accomplished using a patch either of the reservoir or porous membrane type, or of a solid matrix variety.

The pharmaceutical composition of the invention can be formulated as neutral or salt forms. Pharmaceutically acceptable salts include those formed with anions such as those derived from hydrochloric, phosphoric, acetic, oxalic, tartaric acids, etc., and those formed with cations such as those derived from sodium, potassium, ammonium, calcium, ferric hydroxides, isopropylamine, triethylamine, 2-ethylamino ethanol, histidine, procaine, etc.

A compound of Formula (I) of the present invention can also, if desired, be presented in a pack, or dispenser device which cancontain one or more unit dosage forms containing the said agent. The pack can for example comprise metal or plastic foil, such as blister pack. The pack or dispenser device can be accompanied with instruction for administration.

A compound of Formula (I) of the present invention can be administered as sole active agent or can be adminstered in combination with other agents. These agents include non-steroidal anti-inflammatory drug (NSAIDS) such as celecoxib, rofecoxib, cimicoxib, etoricoxib, lumiracoxib, valdecoxib, deracoxib, N-(2-cyclohexyloxynitrophenyl)methane sulphonamide, COX189, ABT963, JTE-522, GW-406381 , LAS-34475, CS-706, PAC-10649, SVT- 2016, GW-644784, tenidap, acetylsalicylic acid (aspirin), amoxiprin, benorilate, choline magnesium salicylate, diflunisal, faislamine, methyl salicylate, magnesium salicylate, salicyl salicylate (salsalatee), diclofenac, aceclofenac, acemetacin, bromfenac, etodolac, indometacin, nabumetone, sulindac, tolmetin, ibuprofen, carprofen, fenbufen, fenoprofen, flurbiprofen, ketoprofen, ketorolac, loxoprofen, naproxen, oxaprozin, tiaprofenic acid, suprofen, mefenamic acid, meclofenamic acid, phenylbutazone,

azapropazone, metamizole, oxyphenbutazone, sulfinpyrazone, piroxicam, lornoxicam, meloxicam, tenoxicam, nimesulide, licofelone, paracetamol.

A compound of Formula (I) of the present invention can be combined with agents such as TNF-a inhibitors such as anti-TNF monoclonal antibodies (such as Remicade, CDP-870 and D2E7) and TNF receptor immunoglobulin molecules (such as Enbrel), low dose methotrexate, lefunomide; ciclesonide; hydroxychloroquine, d-penicillamine, auranofin or parenteral or oral gold. A compound of Formula (I) of the present invention can also be administered in combination with an inhibitor of proTNFalpha convertase enzyme (TACE) such as 3-Amino-N-hydroxy-a-(2-methylpropyl)-3-[4-[(2-methyl-4- quinolinyl)methoxy]phenyl]-2-oxo-1-pyrrolidineacetamide, 2(S),3(S)- Piperidinedicarboxamide, N3-hydroxy-1-methyl-N-2-[4-[(2-methyl-4- quinolinyl)methoxy]phenyl], 3-Thiomorpholinecarboxamide, 4-[[4-(2- butynyloxy)phenyl]sulfonyl]-N-hydroxy-2,2-dimethyl, 5-Hexenoic acid, 3- [(hydroxyamino)carbonyl]-2-(2-methylpropyl)-6-phenyl-, 2-(2-methylpropyl)-2- (methylsulfonyl)hydrazide, (2R,3S,5E), 2-Piperidinecarboxamide, N,5- dihydroxy-1-[[4-(1-naphthalenylmethoxy)phenyl]sulfonyl]-, (2R.5R),

Pentanamide, 3-(formylhydroxyamino)-4-methyl-2-(2-methylpropyl)-N- [(1S,2S)-2-methyl-1-[(2-pyridinylamino)carbonyl]butyl]-, (2R.3S), 2- Propenamide, N-hydroxy-3-[3-[[(4-methoxyphenyl)sulfonyl](1- methylethyl)amino]phenyl]-3-(3-pyridinyl)-, (2E), Benzamide, N-(2,4-dioxo- 1 ,3,7-triazaspiro[4.4]non-9-yl)-4-[(2-methyl-4-quinolinyl)methoxy],

Benzamide, N-[(1-acetyl-4-piperidinyl)(2,5-dioxo-4-imidazolidinyl)methyl]-4- [(2-meth- yl-4-quinolinyl)methoxy], or 2,4-lmidazolidinedione, 5-methyl-5-[[[4- [(2-methyl-4-quinolinyl)methoxy]phenyl]sulfonyl]methyl]. Other examples of TACE inhibitors are described in WO 99/18074, WO 99/65867, U.S. Pat. No. 6,225,311 , WO 00/00465, WO 00/09485, WO 98/38179, WO 02/18326, WO 02/096426, WO 03/079986, WO 03/055856, WO 03/053941 , WO

03/040103, WO 03/031431, WO 03/024899, WO 03/016248, WO 04/096206, WO 04/033632, WO 04/108086, WO 04/043349, WO 04/032846, WO

04/012663, WO 04/006925, WO 07/016597.

A compound of Formula (I) of the present invention can also be administered in combination with a corticosteroid such as budesonide, corticosterone, Cortisol, cortisone acetate, prednisone, prednisolone, methylprednisolone, dexamethasone, betamethasone, triamcinolone, beclometasone,

fludrocortisone acetate, deoxycorticosterone acetate (doca), aldosterone. A compound of Formula (I) of the present invention can further be

administered in combination with a p2-adrenergic receptor agonist such as formoterol, salbutamol (albuterol), levalbuterol, terbutaline, pirbuterol, procaterol, metaproterenol, fenoterol, bitolterol mesylate, salmeterol, bambuterol, clenbuterol.

A compound of Formula (I) of the present invention can further be

administered in combination with an antidepressant drug such as sertraline, escitalopram, fluoxetine, bupropion, paroxetine, venlafaxine, trazodone, amitriptyline, citalopram, duloxetine, mirtazapine, nortriptyline, imipramine, lithium.

A compound of Formula (I) of the present invention can further be

administered in combination with an antipsychotic drug such as

chlorpromazine, fluphenazine, perphenazine, prochlorperazine, thioridazine, trifluoperazine, mesoridazine, promazine, triflupromazine, levomepromazine, promethazine, chlorprothixene, flupenthixol, thiothixene, zuclopenthixol, haloperidol, droperidol, pimozide, melperone, benperidol, triperidol, clozapine , olanzapine, risperidone, quetiapine, ziprasidone, amisulpride, paliperidone , bifeprunox, aripiprazole.

A compound of Formula (I) of the present invention can also be administered in combination with a leukotriene biosynthesis inhibitor, 5-lipoxygenase (5- LO) inhibitor or 5-lipoxygenase activating protein (FLAP) antagonist, for example, zileuton; ABT-761 ; fenleuton; tepoxalin; nicaraven; VIA-2291 ;

etalocib; ketoprofen, Abt-79175; Abt-8576 ; N-(5-substituted) thiophene-2- alkylsulfonamides; TDT-070; licofelone; PEP-03; tenoxicam; 2,6-di-tert- butylphenol hydrazones; methoxytetrahydropyrans such as Zeneca ZD-2138; the compound SB- 210661 ; pyridinyl-substituted 2-cyanonaphthalene compounds such as L-739-010; 2-cyanoquinoline compounds such as L-746- 530; indole and quinoline compounds such as MK-591 , MK-886, and BAY x 1005. A compound of Formula (I) of the present invention can be administered in combination with a receptor antagonists for leukotrienes LTB4, LTC4, LTD4, and LTE, for example, phenothiazin-3-ones such as L-651 ,392; amidino compounds such as CGS-25019c; benzoxalamines such as ontezolast; benzenecarboximidamides such as BIIL 284/260; and compounds such as zafirlukast, ablukast, montelukast, praniukast, verlukast (MK-679), RG- 12525, Ro-245913, iralukast (CGP 45715A), and BAY x 7195; masilukast. A compound of Formula (I) of the present invention can also be administered in combination with a PDE4 inhibitor including inhibitors of the isoform PDE4D.

A compound of Formula (I) of the present invention can also be administered in combination with a antihistaminic Hi receptor antagonists including cetirizine, loratadine, desloratadine, fexofenadine, astemizole, azelastine, and chlorpheniramine.

A compound of Formula (I) of the present invention can further be

administered in combination with with a gastroprotective H₂ receptor antagonist.

A compound of Formula (I) of the present invention can yet further be administered in combination with an a1- and a2-adrenoceptor agonist vasoconstrictor sympathomimetic agent, including propylhexedrine, phenylephrine, phenylpropanolamine, pseudoephedrine, naphazoline hydrochloride, oxymetazoline hydrochloride, tetrahydrozoline hydrochloride, xylometazoline hydrochloride, and ethylnorepinephrine hydrochloride. A compound of Formula (I) of the present invention can be administered in combination with anticholinergic agents including ipratropium bromide;

tiotropium bromide; oxitropium bromide; pirenzepine; and telenzepine The present invention still further relates to the combination of a compound of the invention together with a βι- to p₄-adrenoceptor agonists including

metaproterenol, isoproterenol, isoprenaline, albuterol, salbutamol, formoterol, salmeterol, terbutaline, orciprenaline, bitolterol mesylate, and pirbuterol; or methylxanthanines including theophylline and aminophylline; sodium cromoglycate; or muscarinic receptor (M1 , M2, and M3) antagonist.

A compound of Formula (I) of the present invention can be administered in combination with an insulin-like growth factor type I (IGF-1) mimetic.

A compound of Formula (I) of the present invention can be administered in combination with an inhaled glucocorticoid with reduced systemic side effects, including, prednisone, prednisolone, flunisolide, triamcinolone acetonide, beclomethasone dipropionate, budesonide, fluticasone

propionate, and mometasone furoate.

A compound of Formula (I) of the present invention can be administered in combination with (a) tryptase inhibitors; (b) platelet activating factor (PAF) antagonists; (c) interleukin converting enzyme (ICE) inhibitors; (d) IMPDH inhibitors; (e) adhesion molecule inhibitors including VLA-4 antagonists; (f) cathepsins; (g) MAP kinase inhibitors; (h) glucose-6 phosphate

dehydrogenase inhibitors; (i) kinin-Bi- and B2-receptor antagonists; j) anti- gout agents, e.g., colchicine; (k) xanthine oxidase inhibitors, e.g., allopurinol; (I) uricosuric agents, e. g., probenecid, sulfinpyrazone, and benzbromarone; (m) growth hormone secretagogues; (n) transforming growth factor (TGF ); (o) platelet- derived growth factor (PDGF); (p) fibroblast growth factor, e.g., basic fibroblast growth factor (bFGF); (q) granulocyte macrophage colony stimulating factor (GM-CSF); (r) capsaicin cream; (s) Tachykinin NI | and NK₃ receptor antagonists such as NKP-608C; SB-233412 (talnetant); and D- 4418; and (t) elastase inhibitors such as UT-77 and ZD-0892. A compound of Formula (I) of the present invention can be administered in combination with an inhibitor of matrix metalloproteases (MMPs), i.e., the stromelysins, the collagenases, and the gelatinases, as well as aggrecanase; especially collagenase-1 ( MP-1), collagenase-2 (MMP-8), collagenase-3 (MMP-13), stromelysin-1 (MMP-3), stromelysin-2 (MMP-10), and stromelysin- 3 (MMP-11).

A compound of Formula (I) of the present invention can be administered in combination with anticancer agents such as endostatin and angiostatin or cytotoxic drugs such as adriamycin, daunomycin, cis-platinum, etoposide, taxol, taxotere and farnesyl transferase inhibitors, VEGF inhibitors, COX-2 inhibitors and antimetabolites such as methotrexate antineoplastic agents, especially antimitotic drugs including the vinca alkaloids such as vinblastine and vincristine.

A compound of Formula (I) of the present invention can be administered in combination with antiviral agents such as Viracept, AZT, aciclovir and famciclovir, and antisepsis compounds such as Valant. A compound of Formula (I) of the present invention can be administered in combination with cardiovascular agents such as calcium channel blockers, lipid lowering agents such as stating, fibrates, beta-blockers, ACE inhibitors, Angiotensin-2 receptor antagonists and platelet aggregation inhibitors. A compound of Formula (I) of the present invention can be administered in combination with CNS agents such as antidepressants (such as sertraline), anti-Parkinsonian drugs (such as deprenyl, L-dopa, Requip, Mirapex, MAOB inhibitors such as selegine and rasagiline, comP inhibitors such as Tasmar, A-2 inhibitors, dopamine reuptake inhibitors, NMDA antagonists, Nicotine agonists, Dopamine agonists and inhibitors of neuronal nitric oxide

synthase), and anti-Alzheimer's drugs such as donepezil, tacrine, COX-2 inhibitors, propentofylline or metryfonate. A compound of Formula (I) of the present invention can be administered in combination with osteoporosis agents such as roloxifene, droloxifene, lasofoxifene or fosomax and immunosuppressant agents such as FK-506, rapamycin, cyclosporine, azathioprine, and methotrexate.

EXAMPLES Example 1

General Synthetic Procedure I

General Procedure for the preparation ofXIntOI

To a solution of an indole or azaindole derivative X in acetic acid was added sodium cyano borohydride (NaBH₃CN) at 0°C and the solution was stirred at room temperature for 16 hours. The excess of acetic acid was distilled off and the residue was diluted with water and extracted with ethyl acetate. The combined organic layer was washed with aqueous sodium bicarbonate solution, brine, dried over anhydrous sodium sulfate and concentrated under reduced pressure to obtain a mixture of desired product and N-acetyl compound in almost 1 :1 ratio. This material was subjected to hydrolysis by treating it with 6N HCI at reflux temperature for 12 hours. The reaction mixture was neutralized with aqueous sodium bicarbonate to pH=8 and extracted with ethyl acetate. The combined organic layer is washed with brine solution, dried over anhydrous sodium sulfate and concentrated under reduced pressure. XlntOI was purified by flash column chromatography using 60-120 mesh silica gel and 3% ethyl acetate/hexane as eluent.

General Procedure for the preparation ofXYInt02:

A solution of XlntOI and an oxetane aldehyde derivative Y in methanol was stirred for 3 hours at room temperature. Sodium cyano borohydride was added at 0°C and the mixture was stirred for another 3 hours at room temperature. The solvent was removed and the residue was diluted with water and extracted with ethyl acetate. The combined organic layer was washed with water, brine solution, dried over sodium sulfate and

concentrated under reduced pressure to give the crude compound.

Purification by column chromatography with 3% ethyl acetate / hexane yielded XYInt02.

General Procedure for the preparation ofXYInt03:

To a solution of XYInt02 in 1 ,4-dioxane was added 2,3-dichloro-5,6-dicyano- 1 ,4-benzoquinone (DDQ) at room temperature and the solution was allowed to stir for 1 hour. The solvent was removed under vacuum and the residue was purified by column chromatography with 4% ethyl acetate / hexane yielding XYInt03.

General Procedure for the preparation ofXYInt04:

To a solution of XYInt03 in acetic anhydride cupric nitrate was added at 0°C and the solution was allowed to stir for 1 hour at room temperature. Water was added and the reaction mixture was extracted with ethyl acetate. The organic layer was washed with water and with brine solution, dried over sodium sulfate and concentrated under vacuum. The residue was purified by silica gel column chromatography to yield XYInt04. General Procedure for the preparation ofXYInt05:

To a solution of XYInt04 in 1,4-dioxane and water was added Zn and stirred for 5 minutes. Ammonium chloride was added and after 30 minutes the reaction mixture was filtered through celite. The filtrate was diluted with ethyl acetate, washed with water, brine and dried over sodium sulfate. The solvent was removed under reduced pressure to yield XYInt05.

General Procedure for the preparation ofXYZ:

To a solution of cyclo-alkyl acetic acid derivative Z in N.N-dimethylformamide (DMF) was added 0-(7-Azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (HATU) followed by Ν,Ν-diisopropylethylamine (DIPEA) and stirred for 5 minutes. To this a solution of XYInt05 in DMF was added and stirred for another 15 minutes. The reaction mixture was diluted with ice water and extracted with ethyl acetate. The combined organic layer was washed with brine solution, dried over sodium sulfate and the solvent was removed. The residue was purified by chromatography with 5%

methanol/chloroform to yield XYZ.

Example 2

General Synthetic Procedure II

General Procedure for the preparation of XYIntOI

To a solution of an indole or azaindole derivative X in DMF was added the activated (L=tosylate, mesylate, bromide, chloride or iodide) oxetane derivative Y and the mixture was heated to 60°C. After completion of the reaction water was added and the layers were extracted with

dichloromethane. The organic phase was dried over sodium sulfate and the solvent was removed under reduced pressure. Purification by column chromatography with 4% ethyl acetate / hexane yielded XYIntOI .

General Procedure for the preparation of XYIntOI:

To a solution of XYIntOI in acetic anhydride cupric nitrate was added at 0°C and the solution was allowed to stir for 1 hour at room temperature. Water was added and the reaction mixture was extracted with ethyl acetate. The organic layer was washed with water and with brine solution, dried over sodium sulfate and concentrated under vacuum. The residue was purified by silica gel column chromatography to yield XYInt02.

General Procedure for the preparation ofXYInt03: To a solution of XYInt02 in 1 ,4-dioxane and water was added Zn and stirred for 5 minutes. Ammonium chloride was added and after 30 minutes the reaction mixture was filtered through celite. The filtrate was diluted with ethyl acetate, washed with water, brine and dried over sodium sulfate. The solvent was removed under reduced pressure to yield XYInt03.

General Procedure for the preparation ofXYZ:

To a solution of cyclo-alkyl acetic acid derivative Z in N,N-dimethylformamide (DMF) was added 0-(7-Azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (HATU) followed by Ν,Ν-diisopropylethylamine (DIPEA) and stirred for 5 minutes. To this a solution of XYInt03 in DMF was added and stirred for another 5 minutes. The reaction mixture was diluted with ice water and extracted with ethyl acetate. The combined organic layer was washed with brine solution, dried over sodium sulfate and the solvent was removed. The residue was purified by chromatography with 5%

methanol/chloroform to yield XYZ.

Example 3

N-(4-chloro-1-((3-hydroxyoxetan-3-yl)methyl)-1H-indol-3-yl)-2- cyclopentylacetamide

Synthesised according to the procedure disclosed in Example 1 where X is 4- chloro indole, Y is 3-hydroxyoxetane-3-carbaldehyde, and Z is 2- cyclopentylacetic acid. Formula: C₁₉H₂₃CIN₂0₃; Molecular Weight: 362.85; Mass/charge ratio: 362.14 (100.0%), 364.14 (32.7%), 363.14 (21.4%), 365.14 (6.7%), 364.15 (2.1 %); Elemental analysis: C, 62.89; H, 6.39; CI, 9.77; N, 7.72; O, 13.23.

Example 4

N-(4-bromo-1 -((3-hydroxyoxetan-3-yl)methyl)-1 H-indol-3-yl)-2- cyclopentylacetamide

Synthesised according to the procedure disclosed in Example 1 where X is 4- bromo indole, Y is 3-hydroxyoxetane-3-carbaldehyde, and Z is 2- cyclopentylacetic acid. Formula: C₁₉H₂₃BrN₂0₃; Molecular Weight: 407.3; Mass/charge ratio: 406.09 (100.0%), 408.09 (98.0%), 407.09 (21.4%), 409.09 (20.4%), 410.09 (2.7%), 408.10 (2.1%); Elemental analysis: C, 56.03; H, 5.69; Br, 19.62; N, 6.88; O, 11.78.

Example 5

N-(4-chloro-1-((3-hydroxyoxetan-3-yl)methyl)-1H-indol-3-yl)-2- cyclohexylacetamide

Synthesised according to the procedure disclosed in Example 1 where X is 4- chloro indole, Y is 3-hydroxyoxetane-3-carbaldehyde, and Z is 2- cyclohexylacetic acid. Formula: C20H25CIN2O3; Molecular Weight: 376.88; Mass/charge ratio: 376.16 (100.0%), 378.15 (32.0%), 377.16 (22.0%), 379.16 (7.2%), 378.16 (3.0%); Elemental analysis: C, 63.74; H, 6.69; CI, 9.41; N, 7.43; O, 12.74.

Example 6

N-(4-bromo-1 -((3-hydroxyoxetan-3-yl)methy H-indol-3-y l)-2- cyclohexylacetamide

Synthesised according to the procedure disclosed in Example 1 where X is 4- bromo indole, Y is 3-hydroxyoxetane-3-carbaldehyde, and Z is 2- cyclohexylacetic acid. Formula: C₂oH₂5BrN₂03; Molecular Weight: 421.33; Mass/charge ratio: 420.10 (100.0%), 422.10 (97.3%), 421.11 (22.0%), 423.11 (21.7%), 422.11 (3.1%), 424.11 (2.9%); Elemental analysis: C, 57.01 ; H, 5.98; Br, 18.96; N, 6.65; O, 11.39.

Example 7

N-(4-chloro-1-((3-hydroxyoxetan-3-yl)methyl)-1 H-indol-3-yl)-2- cycloheptylacetamide

Synthesised according to the procedure disclosed in Example 1 where X is 4- chloro indole, Y is 3-hydroxyoxetane-3-carbaldehyde, and Z is 2- cycloheptylacetic acid. Formula: C2iH₂7CIN₂0₃; Molecular Weight: 390.9; Mass/charge ratio: 390.17 (100.0%), 392.17 (32.1%), 391.17 (23.5%), 393.17 (7.7%), 392.18 (3.2%), 394.17 (1.0%); Elemental analysis: C, 64.52; H, 6.96; CI, 9.07; N, 7.17; O, 12.28.

Example 8

N-(4-brotno-1-((3-hydroxyoxetan-3-yl)methyl)-1H-indol-3-yl)-2- cycloheptylacetamide

Synthesised according to the procedure disclosed in Example 1 where X is 4- bromo indole, Y is 3-hydroxyoxetane-3-carbaldehyde, and Z is 2- cycloheptylacetic acid. Formula: C₂iH₂₇BrN₂0₃; Molecular Weight: 435.35; Mass/charge ratio: 434.12 (100.0%), 436.12 (98.1%), 435.12 (23.6%), 437.12 (23.2%), 436.13 (2.6%), 438.13 (2.5%); Elemental analysis: C, 57.94; H, 6.25; Br, 18.35; N, 6.43; O, 11.03.

Example 9

N-(4-chloro-1 -((3-hydroxyoxetan-3-yl)methyl)-1 H-indol-3-yl)-2- adamantane-1yl-acetamide

Synthesised according to the procedure disclosed in Example 1 where X is 4- chloro indole, Y is 3-hydroxyoxetane-3-carbaldehyde, and Z is adamantane- 1yl-acetic acid. Formula: C24H29CIN2O3; Molecular Weight: 428.95;

Mass/charge ratio: 428.19 (100.0%), 430.18 (32.0%), 429.19 (26.4%), 431.19 (8.6%), 430.19 (4.1 %), 432.19 (1.3%); Elemental analysis: C, 67.20; H, 6.81 ; CI, 8.27; N, 6.53; O, 11.19.

Example 10

N-(4-bromo-1 -((3-hydroxyoxetan-3-yl)methy l)-1 H-indol-3-yl)-2- adamantane-1yl-acetamide

Synthesised according to the procedure disclosed in Example 1 where X is 4- bromo indole, Y is 3-hydroxyoxetane-3-carbaldehyde, and Z is adamantane- 1yl-acetic acid. Formula: C2₄H29BrN₂0₃; Molecular Weight: 473.4;

Mass/charge ratio: 472.14 (100.0%), 474.13 (97.3%), 473.14 (26.4%), 475.14 (25.9%), 474.14 (4.1%), 476.14 (3.9%); Elemental analysis: C, 60.89; H, 6.17; Br, 16.88; N, 5.92; O, 10.14.

Example 11

N-(4-chloro-1 -((3-hydroxyoxetan-3-yl)met yl)-1 H-indol-3-y l)-3- cyclopentylpropanamide

Synthesised according to the procedure disclosed in Example 1 where X is 4- chloro indole, Y is 3-hydroxyoxetane-3-carbaldehyde, and Z is 3- cyclopentylpropanoic acid. Formula: C20H25CIN2O3; Molecular Weight:

376.88; Mass/charge ratio: 376.16 (100.0%), 378.15 (32.0%), 377.16

(22.0%), 379.16 (7.2%), 378.16 (3.0%); Elemental analysis: C, 63.74; H, 6.69; CI, 9.41 ; N, 7.43; O, 12.74.

Example 12

N-(4-bromo-1 -((3-hydroxyoxetan-3-yl)methyl)-1 H-indol-3-yl)-3- cyclopentylpropanamide

Synthesised according to the procedure disclosed in Example 1 where X is 4- bromo indole, Y is 3-hydroxyoxetane-3-carbaldehyde, and Z is 3- cyclopentylpropanoic acid. Formula: C₂oH25BrN₂0₃; Molecular Weight:

421.33; Mass/charge ratio: 420.10 (100.0%), 422.10 (97.3%), 421.11

(22.0%), 423.11 (21.7%), 422.11 (3.1%), 424.11 (2.9%); Elemental analysis: C, 57.01 ; H, 5.98; Br, 18.96; N, 6.65; O, 11.39.

Example 13

Synthesised according to the procedure disclosed in Example 1 where X is 4- chloro indole, Y is 3-hydroxyoxetane-3-carbaldehyde, and Z is 3- cyclohexylpropanoic acid. Formula: C2₁H2₇CIN2O3; Molecular Weight: 390.9; Mass/charge ratio: 390.17 (100.0%), 392.17 (32.1%), 391.17 (23.5%), 393.17 (7.7%), 392.18 (3.2%), 394.17 (1.0%); Elemental analysis: C, 64.52; H, 6.96; CI, 9.07; N, 7.17; O, 12.28.

Example 14

N-(4-bromo-1-((3-hydroxyoxetan-3-yl)methyl)-1H-indol-3-yl)-3- cyclohexylpropanamide

Synthesised according to the procedure disclosed in Example 1 where X is 4- bromo indole, Y is 3-hydroxyoxetane-3-carbaldehyde, and Z is 3- cyclohexylpropanoic acid. Formula: C₂iH₂₇BrN₂03; Molecular Weight: 435.35; Mass/charge ratio: 434.12 (100.0%), 436.12 (98.1%), 435.12 (23.6%), 437.12 (23.2%), 436.13 (2.6%), 438.13 (2.5%); Elemental analysis: C, 57.94; H, 6.25; Br, 18.35; N, 6.43; O, 11.03. Example 15

N-(4-chloro-1-((3-hydroxyoxetan-3-yl)methyl)-1H-indol-3-yl)-3- cycloheptylpropanamide

Synthesised according to the procedure disclosed in Example 1 where X is 4- chloro indole, Y is 3-hydroxyoxetane-3-carbaldehyde, and Z is 3- cycloheptylpropanoic acid. Formula: C22H29CIN2O3; Molecular Weight:

404.93; Mass/charge ratio: 404.19 (100.0%), 406.18 (32.0%), 405.19

(24.2%), 407.19 (7.9%), 406.19 (3.5%), 408.19 (1.1%); Elemental analysis: C, 65.25; H, 7.22; CI, 8.76; N, 6.92; O, 11.85.

Example 16

N-(4-bromo-1 -((3-hydroxyoxetan-3-yl)methy l)-1 H

cycloheptylpropanamide

Synthesised according to the procedure disclosed in Example 1 where X is 4- bromo indole, Y is 3-hydroxyoxetane-3-carbaldehyde, and Z is 3- cycloheptylpropanoic acid. Formula: C₂2H₂9BrN₂0₃; Molecular Weight:

449.38; Mass/charge ratio: 448.14 (100.0%), 450.13 (97.3%), 449.14

(24.2%), 451.14 (23.8%), 450.14 (3.5%), 452.14 (3.3%); Elemental ana C, 58.80; H, 6.50; Br, 17.78; N, 6.23; O, 10.68.

Example 17

N-(4-chloro-1-((3-hydroxyoxetan-3-yl)methyl)-1 H-indol-3-yl)-3- adamantane-1yl-propanamide

Synthesised according to the procedure disclosed in Example 1 where X is 4- chloro indole, Y is 3-hydroxyoxetane-3-carbaldehyde, and Z is adamantane- 1yl-propanoic acid. Formula: C25H31CIN2O3; Molecular Weight: 442.98;

Mass/charge ratio: 442.20 (100.0%), 444.20 (32.2%), 443.21 (27.5%), 445.20 (8.9%), 444.21 (4.3%), 446.21 (1.2%); Elemental analysis: C, 67.78; H, 7.05; CI, 8.00; N, 6.32; O, 10.84.

Example 18

N-(4-bromo-1 -((3-hydroxyoxetan-3-yl)methyl)-1 H-indol-3-yl)-3- adamantane-1yl-propanamide

Synthesised according to the procedure disclosed in Example 1 where X is 4- bromo indole, Y is 3-hydroxyoxetane-3-carbaldehyde, and Z is adamantane- 1yl-propanoic acid. Formula: C25H₃₁BrN₂0₃; Molecular Weight: 487.43;

Mass/charge ratio: 486.15 (100.0%), 488.15 (97.5%), 487.16 (27.5%), 489.15 (27.1%), 488.16 (4.3%), 490.16 (3.6%); Elemental analysis: C, 61.60; H, 6.41 ; Br, 16.39; N, 5.75; O, 9.85.

Example 19

2-cycloheptyl-N-(1-((3-hydroxyoxetan-3-yl)methyl)-4-methyl-1H-indol-3- yl)acetamide

Synthesised according to the procedure disclosed in Example 1 where X is 4- methyl indole, Y is 3-hydroxyoxetane-3-carbaldehyde, and Z is 2- cycloheptylacetic acid. Formula: C22H30N2O3; Molecular Weight: 370.49; Mass/charge ratio: 370.23 (100.0%), 371.23 (24.3%), 372.23 (3.5%);

Elemental analysis: C, 71.32; H, 8.16; N, 7.56; O, 12.96.

Example 20

2-cyclohepty l-N-(4-hydroxy-1 -((3-hydroxyoxetan-3-yl)methy H-indol- 3-yl)acetamide

Synthesised according to the procedure disclosed in Example 1 where X is 4- hydroxy indole, Y is 3-hydroxyoxetane-3-carbaldehyde, and Z is 2- cycloheptylacetic acid. Formula: C21 H28N2O4; Molecular Weight: 372.46; Mass/charge ratio: 372.20 (100.0%), 373.21 (23.2%), 374.21 (3.6%);

Elemental analysis: C, 67.72; H, 7.58; N, 7.52; O, 17.18.

Example 21

2- cycloheptyl-N-(1-((3-hydroxyoxetan-3-yl)methyl)-4-methoxy-1H-indol-

3- yl)acetamide

Synthesised according to the procedure disclosed in Example 1 where X is 4- methoxy indole, Y is 3-hydroxyoxetane-3-carbaldehyde, and Z is 2- cycloheptylacetic acid. Formula: C₂₂H₃₀N2O₄; Molecular Weight: 386.48; Mass/charge ratio: 386.22 (100.0%), 387.22 (24.7%), 388.23 (2.8%), 388.22 (1.0%); Elemental analysis: C, 68.37; H, 7.82; N, 7.25; O, 16.56.

Example 22

Synthesised according to the procedure disclosed in Example 1 where X is 4- trifluoromethyl indole, Y is 3-hydroxyoxetane-3-carbaldehyde, and Z is 2- cycloheptylacetic acid. Formula: C22H2₇F3N2O3; Molecular Weight: 424.46; Mass/charge ratio: 424.20 (100.0%), 425.20 (24.2%), 426.20 (3.5%);

Elemental analysis: C, 62.25; H, 6.41 ; F, 13.43; N, 6.60; O, 11.31.

Example 23

N-(4-cyano-1-((3-hydroxyoxetan-3-yl)methyl)-1H-indol-3-yl)-2- cycloheptylacetamide

Synthesised according to the procedure disclosed in Example 1 where X is 4- cyano indole, Y is 3-hydroxyoxetane-3-carbaldehyde, and Z is 2- cycloheptylacetic acid. Formula: C2₂H27N3O3; Molecular Weight: 381.47; Mass/charge ratio: 381.21 (100.0%), 382.21 (24.2%), 383.21 (3.7%), 382.20 (1.1 %); Elemental analysis: C, 69.27; H, 7.13; N, 11.02; O, 12.58.

Example 24

Synthesised according to the procedure disclosed in Example 1 where X is 4- chloro indole, Y is 2-(3-hydroxyoxetan-3-yl)acetaldehyde, and Z is 2- cyclopentylacetic acid. Formula: C20H25CIN2O3; Molecular Weight: 376.88; Mass/charge ratio: 376.16 (100.0%), 378.15 (32.0%), 377.16 (22.0%), 379.16 (7.2%), 378.16 (3.0%); Elemental analysis: C, 63.74; H, 6.69; CI, 9.41 ; N, 7.43; O, 12.74.

Example 25

Synthesised according to the procedure disclosed in Example 1 where X is 4- bromo indole, Y is 2-(3-hydroxyoxetan-3-yl)acetaldehyde, and Z is 2- cyclopentylacetic acid. Formula: C2oH25BrN₂03; Molecular Weight: 421.33; Mass/charge ratio: 420.10 (100.0%), 422.10 (97.3%), 421.11 (22.0%), 423.11 (21.7%), 422.11 (3.1%), 424.11 (2.9%); Elemental analysis: C, 57.01; H, 5.98; Br, 18.96; N, 6.65; O, 11.39.

Example 26

N-(4-chloro-1-(2-(3-hydroxyoxetan-3-yl)ethyl)-1H-indol-3-yl)-2- cyclohexylacetamide

Synthesised according to the procedure disclosed in Example 1 where X is 4- chloro indole, Y is 2-(3-hydroxyoxetan-3-yl)acetaldehyde, and Z is 2- cyclohexylacetic acid. Formula: C2₁H2₇CIN2O3; Molecular Weight: 390.9;

Mass/charge ratio: 390.17 (100.0%), 392.17 (32.1 %), 391.17 (23.5%), 393.17 (7.7%), 392.18 (3.2%), 394.17 (1.0%); Elemental analysis

H, 6.96; CI, 9.07; N, 7.17; O, 12.28.

Example 27

N-(4-bromo-1 -(2-(3-hydroxyoxetan-3-yl)ethy H-indol-3-yl)-2

cyclohexylacetamide

Synthesised according to the procedure disclosed in Example 1 where X is 4- bromo indole, Y is 2-(3-hydroxyoxetan-3-yl)acetaldehyde, and Z is 2- cyclohexylacetic acid. Formula: C₂iH₂₇BrN₂0₃; Molecular Weight: 435.35; Mass/charge ratio: 434.12 (100.0%), 436.12 (98.1%), 435.12 (23.6%), 437.12 (23.2%), 436.13 (2.6%), 438.13 (2.5%); Elemental analysis: C, 57.94; H, 6.25; Br, 18.35; N, 6.43; O, 11.03.

Example 28

Synthesised according to the procedure disclosed in Example 1 where X is 4- chloro indole, Y is 2-(3-hydroxyoxetan-3-yl)acetaldehyde, and Z is 2- cycloheptylacetic acid. Formula: C22H29CIN2O3; Molecular Weight: 404.93; Mass/charge ratio: 404.19 (100.0%), 406.18 (32.0%), 405.19 (24.2%), 407.19 (7.9%), 406.19 (3.5%), 408.19 (1.1%); Elemental analysis: C, 65.25; H, 7.22; CI, 8.76; N, 6.92; O, 11.85.

Example 29

Synthesised according to the procedure disclosed in Example 1 where X is 4- bromo indole, Y is 2-(3-hydroxyoxetan-3-yl)acetaldehyde, and Z is 2- cycloheptylacetic acid. Formula: C₂2H29BrN₂0₃; Molecular Weight: 449.38; Mass/charge ratio: 448.14 (100.0%), 450.13 (97.3%), 449.14 (24.2%), 451.14 (23.8%), 450.14 (3.5%), 452.14 (3.3%); Elemental analysis: C, 58.80; H, 6.50; Br, 17.78; N, 6.23; O, 10.68.

Example 30

N-(4-chloro-1-(2-(3-hydroxyoxetan-3-yl)ethyl)-1 H-indol-3-yl)-2- adamantane-1yl-acetamide

Synthesised according to the procedure disclosed in Example 1 where X is 4- chloro indole, Y is 2-(3-hydroxyoxetan-3-yl)acetaldehyde, and Z is

adamantane-1yl-acetic acid. Formula: C25H31CIN2O3; Molecular Weight:

442.98; Mass/charge ratio: 442.20 (100.0%), 444.20 (32.2%), 443.21

(27.5%), 445.20 (8.9%), 444.21 (4.3%), 446.21 (1.2%); Elemental analysis: C, 67.78; H, 7.05; CI, 8.00; N, 6.32; O, 10.84.

Example 31

N-(4-bromo-1 -(2-(3-hydroxyoxetan-3-yl)ethyl)-1 H-indol-3-yl)-2- adamantane-1yl-acetamide

Synthesised according to the procedure disclosed in Example 1 where X is 4- bromo indole, Y is 2-(3-hydroxyoxetan-3-yl)acetaldehyde, and Z is

adamantane-1yl-acetic acid. Formula: C25H3iBrN₂0₃; Molecular Weight:

487.43; Mass/charge ratio: 486.15 (100.0%), 488.15 (97.5%), 487.16 (27.5%), 489.15 (27.1%), 488.16 (4.3%), 490.16 (3.6%); Elemental analysis: C, 61.60; H, 6.41 ; Br, 16.39; N, 5.75; O, 9.85.

Example 32

-(4-chloro-1-(2-(3-hydroxyoxetan-3-yl)ethyl)-1H-indol-3-yl)-3-

Synthesised according to the procedure disclosed in Example 1 where X is 4- chloro indole, Y is 2-(3-hydroxyoxetan-3-yl)acetaldehyde, and Z is 3- cyclopentylpropanoic acid. Formula: C21 H27CIN2O3; Molecular Weight: 390.9; Mass/charge ratio: 390.17 (100.0%), 392.17 (32.1 %), 391.17 (23.5%), 393.17 (7.7%), 392.18 (3.2%), 394.17 (1.0%); Elemental analysis: C, 64.52; H, 6.96; CI, 9.07; N, 7.17; O, 12.28.

Example 33

N-(4-bromo-1 -(2-(3-hydroxyoxetan-3-yl)ethyl)-1 H-indol-3-yl)-3- cyclopentylpropanamide --^ Synthesised according to the procedure disclosed in Example 1 where X is 4- bromo indole, Y is 2-(3-hydroxyoxetan-3-yl)acetaldehyde, and Z is 3- cyclopentylpropanoic acid. Formula: C2iH2₇BrN₂0₃; Molecular Weight:

435.35; Mass/charge ratio: 434.12 (100.0%), 436.12 (98.1%), 435.12

(23.6%), 437.12 (23.2%), 436.13 (2.6%), 438.13 (2.5%); Elemental analysis: C, 57.94; H, 6.25; Br, 18.35; N, 6.43; O, 11.03.

Example 34

Synthesised according to the procedure disclosed in Example 1 where X is 4- chloro indole, Y is 2-(3-hydroxyoxetan-3-yl)acetaldehyde, and Z is 3- cyclohexylpropanoic acid. Formula: C22H29CIN₂O3; Molecular Weight: 404.93; Mass/charge ratio: 404.19 (100.0%), 406.18 (32.0%), 405.19 (24.2%), 407.19 (7.9%), 406.19 (3.5%), 408.19 (1.1 %); Elemental analysis: C, 65.25; H, 7.22; CI, 8.76; N, 6.92; O, 11.85.

Example 35

N-(4-bromo-1 -(2-(3-hydroxyoxetan-3-yl)ethy l)-1 H-indol-3-yl)-3- cyclohexylpropanamide

Synthesised according to the procedure disclosed in Example 1 where X is 4- bromo indole, Y is 2-(3-hydroxyoxetan-3-yl)acetaldehyde, and Z is 3- cyclohexylpropanoic acid. Formula: C22H29BrN₂0₃; Molecular Weight: 449.38; Mass/charge ratio: 448.14 (100.0%), 450.13 (97.3%), 449.14 (24.2%), 451.14 (23.8%), 450.14 (3.5%), 452.14 (3.3%); Elemental analysis: C, 58.80; H, 6.50; Br, 17.78; N, 6.23; O, 10.68.

Example 36

N-(4-chloro-1-(2-(3-hydroxyoxetan-3-yl)ethyl)-1H-indol-3-yl)-3- cycloheptylpropanamide

Synthesised according to the procedure disclosed in Example 1 where X is 4- chloro indole, Y is 2-(3-hydroxyoxetan-3-yl)acetaldehyde, and Z is 3- cycloheptylpropanoic acid. Formula: C23H₃₁CIN₂0₃; Molecular Weight:

418.96; Mass/charge ratio: 418.20 (100.0%), 420.20 (32.1%), 419.21

(25.3%), 421.20 (8.2%), 420.21 (3.7%), 422.21 (1.0%); Elemental analysis: C, 65.94; H, 7.46; CI, 8.46; N, 6.69; O, 11.46.

Example 37

N-(4-bromo-1 -(2-(3-hydroxyoxetan-3-y l)ethy l)-1 H-indol-3-y l)-3- cycloheptylpropanamide

Synthesised according to the procedure disclosed in Example 1 where X is 4- bromo indole, Y is 2-(3-hydroxyoxetan-3-yl)acetaldehyde, and Z is 3- cycloheptylpropanoic acid. Formula: C23H₃₁BrN₂O3; Molecular Weight:

463.41 ; Mass/charge ratio: 462.15 (100.0%), 464.15 (97.5%), 463.16

(25.3%), 465.15 (25.0%), 464.16 (3.7%), 466.16 (3.0%); Elemental analysis: C, 59.61 ; H, 6.74; Br, 17.24; N, 6.05; O, 10.36.

Example 38

N-(4-chloro-1-(2-(3-hydroxyoxetan-3-yl)ethyl)-1H-indol-3-yl)-3- adamantane-1yl-propanamide

adamantane-1yl-propanoic acid. Formula: C26H33CIN2O3; Molecular Weight: 457; Mass/charge ratio: 56.22 (100.0%), 458.22 (36.6%), 457.22 (29.4%), 459.22 (9.2%), 460.22 (1.5%); Elemental analysis: C, 68.33; H, 7.28; CI, 7.76; N, 6.13; O, 10.50.

Example 39

N-(4-bromo-1 -(2-(3-hydroxyoxetan-3-yl)ethyl)-1 H-indol-3-yl)-3- adamantane-1yl-propanamide

adamantane-1yl-propanoic acid. Formula: C₂6H33BrN₂0₃; Molecular Weight: 501.46; Mass/charge ratio: 502.17 (100.0%), 500.17 (98.1 %), 501.17

(28.1%), 503.17 (27.3%), 504.17 (4.5%); Elemental analysis: C, 62.27; H, 6.63; Br, 15.93; N, 5.59; O, 9.57.

Example 40

3-cycloheptyl-N-(1-(2-(3-hydroxyoxetan-3-yl)ethyl)-4-methyl-1 H-indol-3- yl)propanamide

Synthesised according to the procedure disclosed in Example 1 where X is 4- methyl indole, Y is 2-(3-hydroxyoxetan-3-yl)acetaldehyde, and Z is 3- cycloheptylpropanoic acid. Formula: C24H₃4N₂0₃; Molecular Weight: 398.54; Mass/charge ratio: 398.26 (100.0%), 399.26 (26.5%), 400.26 (4.1 %);

Elemental analysis: C, 72.33; H, 8.60; N, 7.03; O, 12.04.

Example 41

Synthesised according to the procedure disclosed in Example 1 where X is 4- hydroxy indole, Y is 2-(3-hydroxyoxetan-3-yl)acetaldehyde, and Z is 3- cycloheptylpropanoic acid. Formula: C23H32N2O4; Molecular Weight: 400.51 ; Mass/charge ratio: 400.24 (100.0%), 401.24 (25.4%), 402.24 (4.0%);

Elemental analysis: C, 68.97; H, 8.05; N, 6.99; O, 15.98.

Example 42

3-cycloheptyl-N-(1-(2-(3-hydroxyoxetan-3-yl)ethyl)-4-methoxy-1H-indol- 3-yl)propanamide

Synthesised according to the procedure disclosed in Example 1 where X is 4- methoxy indole, Y is 2-(3-hydroxyoxetan-3-yl)acetaldehyde, and Z is 3- cycloheptylpropanoic acid. Formula: C24H34N2O4; Molecular Weight: 414.54; Mass/charge ratio: 414.25 (100.0%), 415.26 (26.5%), 416.26 (4.2%);

Elemental analysis: C, 69.54; H, 8.27; N, 6.76; O, 15.44. Example 43

Synthesised according to the procedure disclosed in Example 1 where X is 4- trifluoromethyl indole, Y is 2-(3-hydroxyoxetan-3-yl)acetaldehyde, and Z is 3- cycloheptylpropanoic acid. Formula: C2₄H31 F3N2O3; Molecular Weight:

452.51 ; Mass/charge ratio: 452.23 (100.0%), 453.23 (27.2%), 454.24 (3.4%); Elemental analysis: C, 63.70; H, 6.91 ; F, 12.60; N, 6.19; O, 10.61.

Example 44

N-(4-cyano-1-(2-(3-hydroxyoxetan-3-yl)ethyl)-1H-indol-3-yl)-3- cycloheptylpropanamide

Synthesised according to the procedure disclosed in Example 1 where X is 4- cyano indole, Y is 2-(3-hydroxyoxetan-3-yl)acetaldehyde, and Z is 3- cycloheptylpropanoic acid. Formula: C24H31 N3O3; Molecular Weight: 409.52; Mass/charge ratio: 409.24 (100.0%), 410.24 (26.4%), 411.24 (4.2%), 410.23 (1.1%); Elemental analysis: C, 70.39; H, 7.63; N, 10.26; O, 11.72.

Example 45

N-(4-chloro-1 -((oxetan-3-yl)methyl)-1 H-indol-3-yl)-2- cycloheptylacetamide

Synthesised according to the procedure disclosed in Example 1 where X is 4- chloro indole, Y is oxetane-3-carbaldehyde, and Z is 2-cycloheptylacetic acid.

Formula: C21 H27CIN2O2; Molecular Weight: 374.9; Mass/charge ratio: 374.18 (100.0%), 376.17 (32.0%), 375.18 (23.1%), 377.18 (7.5%), 376.18 (3.1%); Elemental analysis: C, 67.28; H, 7.26; CI, 9.46; N, 7.47; O, 8.54. Example 46

N-(4-bromo-1-((oxetan-3-yl)methyl)-1H-indol-3-yl)-2- cycloheptylacetamide

Synthesised according to the procedure disclosed in Example 1 where X is 4- bromo indole, Y is oxetane-3-carbaldehyde, and Z is 2-cycloheptylacetic acid. Formula: C₂iH27BrN₂02; Molecular Weight: 419.36; Mass/charge ratio: 418.13 (100.0%), 420.12 (97.3%), 419.13 (23.1%), 421.13 (22.6%), 420.13 (3.1 %), 422.13 (2.9%); Elemental analysis: C, 60.15; H, 6.49; Br, 19.05; N, 6.68; O, 7.63.

Example 47

N-(4-chloro-1 -((oxetan-3-yl)methyl)-1 H-indol-3-yl)-3-

Synthesised according to the procedure disclosed in Example 1 where X is 4- chloro indole, Y is oxetane-3-carbaldehyde, and Z is 3-cycloheptylpropanoic acid. Formula: C22H29CIN2O2; Molecular Weight: 388.93; Mass/charge ratio: 388.19 (100.0%), 390.19 (32.1%), 389.20 (24.2%), 391.19 (7.9%), 390.20 (3.2%); Elemental analysis: C, 67.94; H, 7.52; CI, 9.12; N, 7.20; O, 8.23.

Example 48

N-(4-bromo-1-((oxetan-3-yl)methyl)-1H-indol-3-yl)-3-

Synthesised according to the procedure disclosed in Example 1 where X is 4- bromo indole, Y is oxetane-3-carbaldehyde, and Z is 3-cycloheptylpropanoic acid. Formula: C₂2H29BrN₂0₂; Molecular Weight: 433.38; Mass/charge ratio: 432.14 (100.0%), 434.14 (97.5%), 433.14 (24.5%), 435.14 (24.0%), 434.15 (3.2%), 436.15 (2.7%); Elemental analysis: C, 60.97; H, 6.74; Br, 18.44; N, 6.46; O, 7.38.

Example 49

N-(4-chloro-1 -((3-fluorooxetan-3-yl)methyl)-1 H-indol-3-yl)-2- cycloheptylacetamide

Synthesised according to the procedure disclosed in Example 1 where X is 4- chloro indole, Y is 3-fluorooxetane-3-carbaldehyde, and Z is 2- cycloheptylacetic acid. Formula: C21H26CIFN2O2; Molecular Weight: 392.89; Mass/charge ratio: 392.17 (100.0%), 394.16 (32.0%), 393.17 (23.1 %), 395.17 (7.5%), 394.17 (3.1 %); Elemental analysis: C, 64.20; H, 6.67; CI, 9.02; F. 4.84; N, 7.13; O, 8.14.

Example 50

Synthesised according to the procedure disclosed in Example 1 where X is 4- bromo indole, Y is 3-fluorooxetane-3-carbaldehyde, and Z is 2- cycloheptylacetic acid. Formula: C₂iH₂₆BrFN₂O₂; Molecular Weight: 437.35; Mass/charge ratio: 436.12 (100.0%), 438.11 (97.3%), 437.12 (23.1 %), 439.12 (22.6%), 438.12 (3.1%), 440.12 (2.9%); Elemental analysis: C, 57.67; H, 5.99; Br, 18.27; F, 4.34; N, 6.41 ; O, 7.32.

Example 51

N-(4-chloro-1 -((3-fluorooxetan-3-yl)methyl)-1 H-indol-3-yl)-3- cycloheptylpropanamide

Synthesised according to the procedure disclosed in Example 1 where X is 4- chloro indole, Y is 3-fluorooxetane-3-carbaldehyde, and Z is 3- cycloheptylpropanoic acid. Formula: C22H28CIFN2O2; Molecular Weight:

406.92; Mass/charge ratio: 406.18 (100.0%), 408.18 (32.1%), 407.19

(24.2%), 409.18 (7.9%), 408.19 (3.2%); Elemental analysis: C, 64.94; H, 6.94; CI, 8.71 ; F, 4.67; N, 6.88; O, 7.86.

Example 52

Synthesised according to the procedure disclosed in Example 1 where X is 4- bromo indole, Y is 3-fluorooxetane-3-carbaldehyde, and Z is 3- cycloheptylpropanoic acid. Formula: C22H28BrFN₂0₂; Molecular Weight: 451 .37; Mass/charge ratio: 450.13 (100.0%), 452.13 (97.5%), 451.14

(24.2%), 453.13 (23.9%), 452.14 (3.2%), 454.14 (2.7%); Elemental analysis: C, 58.54; H, 6.25; Br, 17.70; F, 4.21 ; N, 6.21 ; O, 7.09. Example 53

N-(4-chloro-1-((3-hydroxyoxetan-3-yl)methyl)-1 H-pyrrolo[2,3-b]pyridin-3- yl)-2-cycloheptylacetami

Synthesised according to the procedure disclosed in Example 1 where X is 4- chloro azaindole, Y is 3-hydroxyoxetane-3-carbaldehyde, and Z is 2- cycloheptylacetic acid. Formula: C20H26CIN3O3; Molecular Weight: 391.89; Mass/charge ratio: 391.17 (100.0%), 393.16 (32.0%), 392.17 (22.0%), 394.17 (7.2%), 393.17 (3.1 %), 392.16 (1.1 %); Elemental analysis: C, 61.30; H, 6.69; CI, 9.05; N, 10.72; O, 12.25.

Example 54

N-(4-bromo-1-((3-hydroxyoxetan-3-yl)methyl)-1 H-pyrrolo[2,3-b]pyridin-3- yl)-2-cycloheptylacetamide

Synthesised according to the procedure disclosed in Example 1 where X is 4- bromo azaindole, Y is 3-hydroxyoxetane-3-carbaldehyde, and Z is 2- cycloheptylacetic acid. Formula: C₂oH26BrN₃03; Molecular Weight: 436.34; Mass/charge ratio: 435.12 (100.0%), 437.11 (97.3%), 436.12 (22.0%), 438.12 (21.6%), 437.12 (3.1%), 439.12 (2.9%), 436.11 (1.1%), 438.11

(1.1%); Elemental analysis: C, 55.05; H, 6.01 ; Br, 18.31; N, 9.63; O, 11.00.

Example 55

N-(6-bromo-4-chloro-1-((3-hydroxyoxetan-3-yl)methyl)-1H-indol-3-yl)-2- cycloheptylacetamide

Synthesised according to the procedure disclosed in Example 1 where X is 6- bromo-4-chloro indole, Y is 3-hydroxyoxetane-3-carbaldehyde, and Z is 2- cycloheptylacetic acid. Formula: C₂iH₂6BrCI ₂03; Molecular Weight: 469.8; Mass/charge ratio: 470.08 (100.0%), 468.08 (77.3%), 472.08 (24.8%), 471.08 (23.6%), 469.08 (18.1%), 473.08 (5.6%), 472.09 (2.6%), 470.09 (2.4%); Elemental analysis: C, 53.69; H, 5.58; Br, 17.01 ; CI, 7.55; N, 5.96; O, 10.22.

Example 56

N-(4-bromo-6-chloro-1-((3-hydroxyoxetan-3-yl)methyl)-1H-indol-3-yl)-2- cycloheptylacetamlde

Synthesised according to the procedure disclosed in Example 1 where X is 4- bromo-6-chloro indole, Y is 3-hydroxyoxetane-3-carbaldehyde, and Z is 2- cycloheptylacetic acid. Formula: C₂iH₂₆BrCIN₂0₃; Molecular Weight: 469.8; Mass/charge ratio: 470.08 (100.0%), 468.08 (77.3%), 472.08 (24.8%), 471.08 (23.6%), 469.08 (18.1 %), 473.08 (5.6%), 472.09 (2.6%), 470.09 (2.4%); Elemental analysis: C, 53.69; H, 5.58; Br, 17.01 ; CI, 7.55; N, 5.96; O, 10.22.

Example 57

N-(4-chloro-1 -(2-(oxetan-3-yl)ethy l)-1 H-indol-3-y l)-2-

Synthesised according to the procedure disclosed in Example 1 where X is 4- chloro indole, Y is 2-(oxetan-3-yl)acetaldehyde, and Z is 2-cycloheptylacetic acid. Formula: C₂2H29CIN₂0₂; Molecular Weight: 388.93; Mass/charge ratio: 388.19 (100.0%), 390.19 (32.1%), 389.20 (24.2%), 391.19 (7.9%), 390.20 (3.2%); Elemental analysis: C, 67.94; H, 7.52; CI, 9.12; N, 7.20; O, 8.23.

Example 58

N-(4-bromo-1 -(2-(oxetan-3-y l)ethyl)-1 H-indol-3-y l)-2- cycloheptylacetamide

Synthesised according to the procedure disclosed in Example 1 where X is 4- bromo indole, Y is 2-(oxetan-3-yl)acetaldehyde, and Z is 2-cycloheptylacetic acid. Formula: C₂2H29BrN₂0₂; Molecular Weight: 433.38; Mass/charge ratio: 432.14 (100.0%), 434.14 (97.5%), 433.14 (24.5%), 435.14 (24.0%), 434.15 (3.2%), 436.15 (2.7%); Elemental analysis: C, 60.97; H, 6.74; Br, 18.44; N, 6.46; O, 7.38.

Example 59

N-(4-chloro-1-(2-(oxetan-3-yl)ethyl)-1H-indol-3-yl)-3- cycloheptylpropanamide Synthesised according to the procedure disclosed in Example 1 where X is 4- chloro indole, Y is 2-(oxetan-3-yl)acetaldehyde, and Z is 3- cycloheptylpropanoic acid. Formula: C23H3₁CIN2O2; Molecular Weight:

402.96; Mass/charge ratio: 402.21 (100.0%), 404.20 (32.0%), 403.21

(25.3%), 405.21 (8.1%), 404.21 (3.6%), 406.21 (1.1%); Elemental analysis: C, 68.55; H, 7.75; CI, 8.80; N, 6.95; O, 7.94.

Example 60

N-(4-bromo-1 -(2-(oxetan-3-yl)ethy l)-1 H-indol-3-yl)-3- cycloheptylpropanamide

Synthesised according to the procedure disclosed in Example 1 where X is 4- bromo indole, Y is 2-(oxetan-3-yl)acetaldehyde, and Z is 3- cycloheptylpropanoic acid. Formula: C₂3H3iBrN₂0₂; Molecular Weight:

447.41 ; Mass/charge ratio: 446.16 (100.0%), 448.15 (97.3%), 447.16

(25.3%), 449.16 (24.7%), 448.16 (3.6%), 450.16 (3.6%); Elemental analysis: C, 61.74; H, 6.98; Br, 17.86; N, 6.26; O, 7.15.

Example 61

Synthesised according to the procedure disclosed in Example 1 where X is 4- chloro indole, Y is 2-(3-fluorooxetan-3-yl)acetaldehyde, and Z is 2- cycloheptylacetic acid. Formula: C22H28CIFN2O2; Molecular Weight: 406.92; Mass/charge ratio: 406. 8 (100.0%), 408.18 (32.1%), 407.19 (24.2%), 409.18 (7.9%), 408.19 (3.2%); Elemental analysis: C, 64.94; H, 6.94; CI, 8.71 ; F. 4.67; N, 6.88; O, 7.86.

Example 62

Synthesised according to the procedure disclosed in Example 1 where X is 4- bromo indole, Y is 2-(3-fluorooxetan-3-yl)acetaldehyde, and Z is 2- cycloheptylacetic acid. Formula: C22H28BrFN₂0₂; Molecular Weight: 451.37; Mass/charge ratio: 450.13 (100.0%), 452.13 (97.5%), 451.14 (24.2%), 453.13 (23.9%), 452.14 (3.2%), 454.14 (2.7%); Elemental analysis: C, 58.54; H, 6.25; Br, 17.70; F, 4.21 ; N, 6.21; O, 7.09.

Example 63

-(4-chloro-1-(2-(3-fluorooxetan-3-yl)ethyl)-1H-indol-3-yl)-3-

Synthesised according to the procedure disclosed in Example 1 where X is 4- chloro indole, Y is 2-(3-fluorooxetan-3-yl)acetaldehyde, and Z is 3- cycloheptylpropanoic acid. Formula: C23H30CIFN2O2; Molecular Weight:

420.95; Mass/charge ratio: 420.20 (100.0%), 422.20 (35.5%), 421.20

(26.0%), 423.20 (8.1%), 424.20 (1.2%); Elemental analysis: C, 65.62; H, 7.18; CI, 8.42; F, 4.51 ; N, 6.65; O, 7.60.

Example 64

N-(4-bromo-1 -(2-(3-fluorooxetan-3-yl)ethyl)-1 H-indol-3-yl)-3- cycloheptylpropanamide Synthesised according to the procedure disclosed in Example 1 where X is 4- bromo indole, Y is 2-(3-fluorooxetan-3-yl)acetaldehyde, and Z is 3- cycloheptylpropanoic acid. Formula: C₂3H₃oBrFN₂O₂; Molecular Weight:

465.4; Mass/charge ratio: 466.15 (100.0%), 464.15 (99.2%), 465.15 (25.1%), 467.15 (24.4%), 468.15 (3.5%); Elemental analysis: C, 59.36; H, 6.50; Br, 17.17; F, 4.08; N, 6.02; O, 6.88.

Example 65

N-(4-chloro-1-(2-(3-hydroxyoxetan-3-yl)ethyl)-1 H-indazol-3-yl)-2- cycloheptylacetamide

Synthesised according to the procedure disclosed in Example 2 where X is 4- chloro azaindole, Y is 2-(3-hydroxyoxetan-3-yl)acetaldehyde, and Z is 2- cycloheptylacetic acid. Formula: C₂ H28CIN3O3; Molecular Weight: 405.92; Mass/charge ratio: 405.18 (100.0%), 407.18 (32.2%), 406.19 (23.1%), 408.18 (7.7%), 407.19 (3.2%), 406.18 (1.1 %); Elemental analysis: C, 62.14; H, 6.95; CI, 8.73; N, 10.35; O, 11.82.

Example 66

N-(4-bromo-1-(2-(3-hydroxyoxetan-3-yl)ethyl)-1 H-indazol-3-yl)-2- cycloheptylacetamide

Synthesised according to the procedure disclosed in Example 2 where X is 4- chloro azaindole, Y is 2-(3-hydroxyoxetan-3-yl)acetaldehyde, and Z is 2- cycloheptylacetic acid. Formula: C₂iH₂₈Br ₃03; Molecular Weight: 450.37; Mass/charge ratio: 449.13 (100.0%), 451.13 (97.5%), 450.13 (23.8%), 452.13 (23.3%), 451.14 (3.2%), 453.14 (2.5%); Elemental analysis: C, 56.00; H, 6.27; Br, 17.74; N, 9.33; O, 10.66.

Example 67

N-indol-3-yl-acetamide and N-azaindol-3-yl-acetamide compounds antagonise P2X7R activity

Inhibition of P2X7R activity by the compounds of the present invention is assessed by measuring calcium influx in Hek293 cells (ECACC No.

85120602) which have been stably transfected with a cDNA for the human P2X7R.

The Hek293 cells are human embryo kidney cells that do not express endogenous P2X7R (Surprenant et al. (1996) Science 272:735-738). Hek293 cells expressing P2X7R were generated by lipofectamine transfection of the human P2X7R cDNA (Genbank accession number BC011913) under the control of the human cytomegalovirus immediate-early (CMV) promoter and inserted into the pcDNA3.1 vector (Invitrogen). Cells were cultivated at 37°C with 8.5% C0₂ in Dulbecco's modified eagles medium (DMEM; GibcoBRUInvitrogen) supplemented with heat-inactivated foetal calf serum (10% v/v), 2 mM L-glutamine, 100 units/ml penicillin, 0.1 mg/ml streptomycin, and 750 pg/ml Geneticin G418 (GibcoBRUInvitrogen).

Inhibition of Bz-ATP-stimulated P2X7R by test compounds was monitored by measuring changes in calcium influx using the Fluo-4-AM fluorescent dye according to the manufacturer's recommendations (Molecular Devices Corporation, U.S.A.). Briefly, Hek293 cells expressing P2X7R were cultured in 96-well plates at a final density of approximately 10,000 cells per well. On the day of the experiment, the culture medium was completely removed from the wells and cells were washed one time in assay buffer (1X Hank's

Balanced Salt (HBSS) solution containing 20 mM Hepes buffer pH 7.4 and 250 mM Probenecid; GibcoBRUInvitrogen). The cells were incubated in 50 μΙ of assay buffer containing 100 μΜ Fluo-4 AM fluorescent dye per well for 1 hour at room temperature. The assay buffer containing the Fluo-4 AM fluorescent dye was then removed, the cells were washed once with assay buffer (without Fluo-4 AM), 100 μΙ of assay buffer (without Fluo-4 AM) containing the test compounds was then added per well. After a 15 minute incubation, 100 μΜ Bz-ATP was added and fluorescence was measured in a FlexStation II (Molecular Devices, U.S.A.) according to the following parameters: 485 nm Excitation Wavelength; 525 nm Emission Wavelength; 515 nm Emission Cut-off; 100 μΙ Pipette Height; 25 μΙ Transfer Volume; 5 fold Compound Concentration; 3 rate Addition Speed. Test compounds were added at concentrations of 0.001 μΜ up to 60 μΜ. The fluorescence data were processed using a lag time of 15 seconds, recording 45 seconds, zero baseline calibrated using 2 points, and % baseline multiplier set at 3. Then, the area of the resulting curve was calculated and the half-maximal inhibitory concentration (IC50) for each test compound was determined using SoftMax Pro software (Molecular Devices, U.S.A.). Compounds of the present invention can inhibit P2X7R activity with an IC50 between 1 μΜ and 0.001 μΜ. For example, the IC50 of compound described in Example 7 is

approximately 0.018 μΜ. Example 68

In vitro efficacy of N-(4-chloro-1-((3-hydroxyoxetan-3-yl)methyl)-1 H- indol-3-yl)-2-cyclopentylacetamide in reducing LPS/ATP induced-IL-Ιβ release in human whole blood

Methods

Whole Blood Stimulation with LPS 026:B6:

Dilution of LPS (stock solution 1 mg/ml): 1 μΙ of stock solution in 1ml of PBS (concentration at 1 Mg/ml). In a BD polypropylene tube, 80 μΙ of diluted LPS are added to 8 ml of blood to reach the concentration of 10 ng/ml as stimulation probe, and 1 ml of unstimulated whole blood probe are kept as control. The tubes are incubated for 4 hours in the incubator (37°C, 5% CO₂) on a shaker. Compound preparation:

The test compound is diluted from stock concentration (10 mM in 100% DMSO). A serial dilution (1/3) is prepared in 100% DMSO, 10

concentrations/compound, then all previous solutions are diluted 10x in PBS and again 10 times in PBS to reach concentrations from 100 μΜ to 10 nM. The final concentration in blood is 10 μΜ to 1 nM.

5 μΙ of the diluted compound are dispensed in a polypropylene 96-well plate 5 μΙ of PBS-1% DMSO for "no compound control" (final 0.1% of DMSO in whole blood). 50 μΙ of whole blood are dispensed on the top of the compound dilution and incubated for 30 minutes in an incubator (37°C, 5% CO₂) on a shaker.

ATP stimulation:

ATP Sigma A2383, stock 100 mM in water, frozen at -20°C aliquot 200 μΙ (each aliquot is frozen and thawed once) is diluted ¼ in PBS and 5 μΙ/well are added in each well (final concentration at 2.1 mM) according to the plate layout and incubated for 30 minutes at 37°C on a plate shaker. End of stimulation:

The stimulation is stopped by adding 90 μΙ of cold PBS and by cetrifugation at 1500 rpm (400 g) for 5 min at 4°C. 30 μΙ of the supernatant (plasma-like, diluted 3 times) are taken and transferred into a new 96-well plate. Taking red blood cells, which interfere with the IL-1 β ELISA, should be avoided.

Cytokine measurement:

The diluted plasma is freezed at -80°C until cytokine measurement. The cytokine level in diluted plasma is analysed with a dilution of 1/25 in dilution buffer (see supplier instruction) with IL-1 beta ELISA kit R&D DY201.

Data analysis:

Optima excel tables are generated with OD₄₅onm and export for IC₅₀ calculation.

IC₅₀ values are calulated using GraphPad Prism software and a four- parameter log fit log (agonist) vs. response - Variable slope (four

parameters) as given by the following formula: Y=Bottom + (Top- Bottom)/(1+10^A((LoglC50-X)^*HillSlope)) where X is the logarithm of concentration and Y is the IL-1 beta concentration. Y starts at Bottom and goes to Top with a sigmoid shape.

Results and conclusion:

P2X7 has a role in secretion of mature IL-1 beta and has been shown, that monocytes express P2X7. In this assay, human whole blood is incubated with LPS in order to stimulate the production of pro-IL-1beta, and treated with ATP to initiate mature IL-1 β secretion. This result demonstrates that N-(4- chloro-1 -((3-hydroxyoxetan-3-yl)methyl)-1 H-indol-3-yl)-2- cyclopentylacetamide (example 3), through the blocking of the P2X7 receptor, is a potent inhibitor of LPS/ATP induced-IL-1beta release (see table 1 and figure 1). IL-1 beta is a very potent pro-inflammatory cytokine in chronic inflammatory conditions and is involved in pathological conditions such as neuropathic pain, rheumatoid arthritis, multiple sclerosis and other

inflammatory / neurodegenerative diseases. N-(4-chloro-1-((3- hydroxyoxetan-3-yl)methyl)-1 H-indol-3-yl)-2-cyclopentylacetamide has therefore been shown to exhibit anti-inflammatory properties on human blood samples.

Table 1 : IC50 values of four different donors

Legend of figure 1 :

Example of the IL-1 beta secretion dose response curves to N-(4-chloro-1-((3- hydroxyoxetan-3-yl)methyl)-1 H-indol-3-yl)-2-cyclopentylacetamide. The IC₅₀ values were determined on four different donors. Two dose response curves are shown. The IC₅₀ values for the four donors are noted in table 1 above. Two donors showed an IC₅₀ value of 3.8 μΜ while the other two donors showed IC50 vaues of 5.3 μΜ and 6.1 μΜ respectively.

Example 69

In vivo efficacy of N-(4-chloro-1-((3-hydroxyoxetan-3-yl)methyl)-1 H- indol-3-yl)-2-cyclopentylacetamide to reduce LPS/ATP induced-IL-Ιβ release in mouse

Protocol:

Five groups of 8 female CH3 mice (Janvier) were enrolled in the study. The first group (control group) was treated with the vehicle (Kleptose 40% in water) intraperitoneally at 10 ml/kg. The second group was treated orally with Dexamethasone (Memphamoson-4) at 1mg/kg. In the third, fourth and fifth group, animals were treated with N-(4-chloro-1-((3-hydroxyoxetan-3- yl)methyl)-1 H-indol-3-yl)-2-cyclopentylacetamide at 15, 50 and 150 mg/kg respectively. The compound was dissolved in the vehicle at a final concentration of 1.5, 5 and 15 mg/ml respectively and the different solutions were administrated intraperitoneally at 10 ml/kg. Ten minutes after the different treatments, animals in the five groups were injected with Lipopolysaccharide (LPS, Sigma) intraperitoneally at 50 pg/ml. Two hours after LPS injection, animals were subsequently injected intraperitoneally with 10 mM ATP (Sigma). Fifteen minutes later, animals were sacrificed and 3 ml of PBS containing protease inhibitors (Roche) and Heparin (Liquemin) were injected into the peritoneal cavity and peritoneal lavages were collected. 500 μΙ of blood was collected through a retro-orbital puncture in heparinised (Liquemin) test tubes. Peritoneal lavages and blood samples were centrifuged at 6000 rpm for 10 min at 4°C and supernatants were collected. Peritoneal lavage samples were diluted 4 times whereas plasma samples were diluted 2 times in PBS and IL-Ι β was measured using an ELISA kit Duoset® (R&D) for peritoneal lavage samples and an ELISA kit Quantikine® (R&D) following the manufacturer instructions. ELISA plates were analyzed by spectrophotometry (BMG labtech).

Results and conclusion:

Treatment with 50 mg/kg of N-(4-chloro-1-((3-hydroxyoxetan-3-yl)methyl)-1H- indol-3-yl)-2-cyclopentylacetamide reduced IL- β release by 55% (p value = 0.01) in the plasma and 52% (p value=0.05) in the peritoneal lavage, whereas treatment with 150 mg/kg of N-(4-chloro-1-((3-hydroxyoxetan-3- yl)methyl)-1H-indol-3-yl)-2-cyclopentylacetamide reduced IL-1 β release by 81% (p value=0.001) in the plasma and 92% (p value=0.001) in the

peritoneal lavage. Treatment with 15 mg/kg did not induce a statistically significant reduction of IL-1 β release. This result demonstrates that N-(4- chloro-1-((3-hydroxyoxetan-3-yl)methyl)-1 H-indol-3-yl)-2- cyclopentylacetamide, through the blocking of the P2X7 receptor, is a potent inhibitor of LPS/ATP induced-IL-Ιβ release. IL-Ιβ is a very potent proinflammatory cytokine in chronic inflammatory conditions and is involved in pathological conditions such as naturopathic pain, rheumatoid arthritis, multiple sclerosis and other neurodegenerative diseases. Legend of figure 2:

Animals (N=8/group) pre-treated with vehicle (ctrl group), dexamethasone at 1 mg/kg (Dexa group), N-(4-chloro-1-((3-hydroxyoxetan-3-yl)methyl)-1 H- indol-3-yl)-2-cyclopentylacetamide (compound) at 15, 50 or 150 mg/kg were injected 10 minutes later with LPS (50pg/mL). Two hours later, animal were injected with ATP (10 mM) and peritoneal lavages and blood samples were collected 15 minutes later. IL-Ιβ release in the peritoneal lavages and plasma were measured by ELISA. ^*: P value = 0.05 **: P value = 0.01. ^***: P value = 0.005.

Claims

1. A compound of the eneral formula:

Ri is a mono- or bicycloalkylalkyl group;

R₂ is selected from hydrogen, -OH, -NH₂, or halogen; 3, R4, R5, e are at each occurrence independently selected from hydrogen, halogen, C1-C6 alkyl, hydroxy, C1-C6 alkoxy, cyano, or trifluoromethyl; n equals 1 or 2; a, b, c, d, x are at each occurrence independently selected from carbon, or nitrogen; or a pharmaceutically acceptable salt or solvate thereof.

2. A compound of claim 1 , wherein R₃, R₄, R₅, R₆ are at each occurrence independently selected from hydrogen, halogen, methyl, hydroxy, methoxy, cyano, or trifluoromethyl. A compound according to claim 1 or 2, wherein Ri is a mono- or bicycloalkylalkyl group selected from cyclopentylmethyl,

cyclopentylethyl, cyclohexylmethyl, cyclohexylethyl, cycloheptylmethyl, cycloheptylethyl, adamantanyl-methyl, or adamantanyl-ethyl.

A compound according to any one of claims 1 to 3, wherein R₂ is -OH or fluoride.

A compound according to any one of claims 1 to 4, wherein at least two of R3, R₄, 5 and R₆ are hydrogen.

A compound of claim 5, wherein R and R6 are H and R3 and R5 are defined as in claim 1 or 2.

A compound of claim 5, wherein R4-R6 are H and R₃ is as defined in claim 1 or 2.

A compound according to any of claims 1 to 7 , wherein x is C.

A compound according to any of claims 1 to 7, wherein n=2 and x is N.

A compound according to any one of claims 1 to 9, wherein a, b, c, and d are C. 11. A compound according to any one of claims 1 to 9, wherein one of a, b, c and d is N.

A compound according to any one claims 1 to 11 selected from

N-(4-chloro-1-((3-hydroxyoxetan-3-yl)methyl)-1 H-indol-3-yl)-2- cyclopentylacetamide N-(4-bromo-1-((3-hydroxyoxetan-3-yl)methyl )-1 H-indol-3-yl)-2- cyclopentylacetamide

N-(4-chloro-1-((3-hydroxyoxetan-3-yl)methyl )-1 H-indol-3-yl)-2- cyclohexylacetamide

N-(4-bromo-1-((3-hydroxyoxetan-3-yl)methyl )-1 H-indol-3-yl)-2- cyclohexylacetamide

N-(4-chloro-1-((3-hydroxyoxetan-3-yl)methyl )-1 H-indol-3-yl)-2- cycloheptylacetamide

N-(4-bromo-1-((3-hydroxyoxetan-3-yl)methyl )-1 H-indol-3-yl)-2- cycloheptylacetamide

N-(4-chloro-1-((3-hydroxyoxetan-3-yl)methyl )-1 H-indol-3-yl)-2- adamantane-1yl-acetamide

N-(4-bromo-1-((3-hydroxyoxetan-3-yl)methyl )-1 H-indol-3-yl)-2- adamantane-1 yl-acetamide

N-(4-chloro-1-((3-hydroxyoxetan-3-yl)methyl )-1 H-indol-3-yl)-3- cyclopentylpropanamide

N-(4-bromo-1-((3-hydroxyoxetan-3-yl)methyl )-1 H-indol-3-yl)-3- cyclopentylpropanamide

N-(4-chloro-1-((3-hydroxyoxetan-3-yl)methyl )-1 H-indol-3-yl)-3- cyclohexylpropanamide

N-(4-bromo-1-((3-hydroxyoxetan-3-yl)methyl )-1 H-indol-3-yl)-3- cyclohexylpropanamide

N-(4-chloro-1-((3-hydroxyoxetan-3-yl)methyl )-1 H-indol-3-yl)-3- cycloheptylpropanamide

N-(4-bromo-1-((3-hydroxyoxetan-3-yl)methyl )-1 H-indol-3-yl)-3- cycloheptylpropanamide

N-(4-chloro-1-((3-hydroxyoxetan-3-yl)methyl )-1 H-indol-3-yl)-3- adamantane-1 yl-propanamide N-(4-bromo-1 -((3-hydroxyoxetan-3-yl)methyl)-1 H-indol-3-yl)-3- adamantane-1 yl-propanamide

2-cycloheptyl-N-(1 -((3-hydroxyoxetan-3-yl)methyl)-4-methyl-1 H- indol-3-yl)acetamide

2-cycloheptyl-N-(4-hydroxy-1 -((3-hydroxyoxetan-3-yl)methyl)-1 H- indol-3-yl)acetamide

2-cycloheptyl-N-(1 -((3-hydroxyoxetan-3-yl)methyl)-4-methoxy-1 H indol-3-yl)acetamide

2-cycloheptyl-N-(4-(trifluoromethyl)-1-((3-hydroxyoxetan-3- yl)methyl)-1 H-indol-3-yl)acetamide

N-(4-chloro-1-(2-(3-hydroxyoxetan-3-yl)ethyl)-1 H-indol-3-yl)-2- cyclopentylacetamide

N-(4-chloro-1-(2-(3-hydroxyoxetan-3-yl)ethyl)-1 H-indol-3-yl)-2- cycloheptylacetamide

N-(4-bromo-1-(2-(3-hydroxyoxetan-3-yl)ethyl)-1 H-indol-3-yl)-2- cycloheptylacetamide

N-(4-chloro-1-(2-(3-hydroxyoxetan-3-yl)ethyl)-1 H-indol-3-yl)-3- cyclopentylpropanamide

N-(4-bromo-1-(2-(3-hydroxyoxetan-3-yl)ethyl)-1 H-indol-3-yl)-2- adamantane-1yl-acetamide N-(4-c loro-1-(2-(3-hydroxyoxetan-3-yl)ethyl)-1 H-indol-3-yl)-3- cyclopentylpropanamide

N-(4-bromo-1-(2-(3-hydroxyoxetan-3-yl)ethyl)-1H-indol-3-yl)-3- cyclopentylpropanamide

N-(4-chloro-1-(2-(3-hydroxyoxetan-3-yl)ethyl)-1H-indol-3-yl)-3- adamantane-1 yl-propanamide

N-(4-bromo-1 -(2-(3-hydroxyoxetan-3-yl)ethyl)-1 H-indol-3-yl)-3- adamantane-1 yl-propanamide

3-cyclohepty l-N-(1 -(2-(3-hydroxyoxetan-3-yl)ethyl)-4-methyl-1 H- indol-3-yl)propanamide

3-cycloheptyl-N-(4-hydroxy-1-(2-(3-hydroxyoxetan-3-yl)ethyl)-1 H- indol-3-yl)propanamide

3-cycloheptyl-N-(1-(2-(3-hydroxyoxetan-3-yl)ethyl)-4-methoxy-1 H indol-3-yl)propanamide

3-cyclohepty l-N-(4-(trifluoromethyl)-1-(2-(3-hydroxyoxetan-3- yl)ethyl)-1 H-indol-3-yl)propanamide

N-(4-chloro-1-((oxetan-3-yl)methyl)-1H-indol-3-yl)-2- cycloheptylacetamide

N-(4-bromo-1 -((oxetan-3-yl)methyl)-1 H-indol-3-yl)-2- cycloheptylacetamide

N-(4-chloro-1 -((oxetan-3-yl)methyl)-1 H-indol-3-yl)-3- cycloheptylpropanamide N-(4-bromo-1-((oxetan-3-yl)methyl)-1H-indol-3-yl)-3- cycloheptylpropanamide

N-(4-chloro-1-((3-fluorooxetan-3-yl)methyl)-1H-indol-3-yl)-2- cycloheptylacetamide

N-(4-chloro-1 -((3-hydroxyoxetan-3-yl)methyl)-1 H-pyrrolo[2,3- b]pyridin-3-yl)-2-cycloheptylacetamide

N-(4-bromo-1-((3-hydroxyoxetan-3-yl)methyl)-1 H-pyrrolo[2,3- b]pyridin-3-yl)-2-cycloheptylacetamide

N-(6-bromo-4-chloro-1 -((3-hydroxyoxetan-3-yl)methyl)-1 H-indol-3 yl)-2-cycloheptylacetamide

N-(4-bromo-6-chloro-1-((3-hydroxyoxetan-3-yl)methyl)-1 H-indol-3 yl)-2-cycloheptylacetamide

N-(4-chloro-1 -(2-(oxetan-3-yl)ethyl)-1 H-indol-3-yl)-2- cycloheptylacetamide

N-(4-bromo-1-(2-(oxetan-3-yl)ethyl)-1H-indol-3-yl)-2- cycloheptylacetamide

N-(4-chloro-1-(2-(oxetan-3-yl)ethyl)-1 H-indol-3-yl)-3- cycloheptylpropanamide

N-(4-bromo-1 -(2-(oxetan-3-yl)ethyl)-1 H-indol-3-yl)-3- cycloheptylpropanamide

N-(4-chloro-1-(2-(3-fluorooxetan-3-yl)ethyl)-1H-indol-3-yl)-2- cycloheptylacetamide

N-(4-chloro-1-(2-(3-fluorooxetan-3-yl)ethyl)-1 H-indol-3-yl)-3- cycloheptylpropanamide N-(4-bromo-1-(2-(3-fluorooxetan-3-yl)ethyl)-1 H-indol-3-yl)-3- cycloheptylpropanamide

- N-(4-bromo-1-(2-(3-hydroxyoxetan-3-yl)ethyl)-1 H-indazol-3-yl)-2- cycloheptylacetamide

A pharmaceutical composition comprising a compound according to any one of claims 1 to 12.

The pharmaceutical composition according to claim 13 further comprising an additional active compound in separate or unit dosage form for simultaneous or sequential administration.

The pharmaceutical composition according to claim 13 or 14 for use in the prophylactic or therapeutic treatment of any disease state in a human, or other mammal, which is exacerbated or caused by excessive or unregulated cytokine production by such mammal's cells, such as but not limited to monocytes and/or macrophages.

The pharmaceutical composition according to claims 13 to 15 for use in the prophylactic or therapeutic treatment of an IL-1 or cytokine mediated condition. 17. The pharmaceutical composition according to claims 13 to 16 for use in the prophylactic or therapeutic treatment of a disease or disorder selected from the group consisting of arthritis (including psoriatic arthritis, Reiter's syndrome, rheumatoid arthritis, gout, traumatic arthritis, rubella arthritis, rheumatoid spondylitis, osteoarthritis, gouty arthritis and acute synovitis), inflammatory bowel disease, Crohn's disease, emphysema, acute respiratory distress syndrome, adult respiratory distress syndrome, asthma, bronchitis, chronic obstructive pulmonary disease, chronic pulmonary inflammatory disease, hyperresponsiveness of the airway, silicosis, pulmonary sarcoidosis, allergic reactions, allergic contact hypersensitivity, eczema, contact dermatitis, allergic dermatitis psoriasis, sunburn, cancer, myoblastic leukemia, diabetes, tissue ulceration, restenosis, periodontal disease, epidermolysis bullosa, osteoporosis, bone resorption disease, loosening of artificial joint implants, atherosclerosis, aortic aneurysm, congestive heart failure, myocardial infarction, stroke, cerebral ischemia, ischemic heart disease, head trauma, neurotrauma, spinal cord injury, neurodegenerative disorders, Alzheimer's disease, Parkinson's disease, glaucoma, age-related macular degeneration, uveitis, neuropathic pain, migraine, depression, peripheral neuropathy, pain, cerebral amyloid angiopathy, nootropic or cognition enhancement, amyotrophic lateral sclerosis, multiple sclerosis, ocular angiogenesis, corneal injury, macular degeneration, corneal scarring, scleritis, abnormal wound healing, burns, autoimmune disorders, Huntington's disease, diabetes, AIDS, cachexia, sepsis, septic shock, endotoxic shock, conjunctivitis shock, gram negative sepsis, toxic shock syndrome, cerebral malaria, cardiac and renal reperfusion injury, thrombosis, glomerularonephritis, graft vs. host reaction, allograft rejection, organ transplant toxicity, ulcerative colitis, or muscle degeneration, in a mammal, including a human.