WO2007014008A2 - Benzenesulfonamide inhibitor of ccr2 chemokine receptor - Google Patents

Abstract

The present invention is related to a compound of formula (I) or a salt thereof, or a solvate thereof, or a combination thereof. The present invention also includes a compositions including the compound of Formula I as well as a method of treating disorders mediated by the CCR2 receptor with the compound of Formula I.

Description

BENZENESULFONAMIDE INHIBITOR OF CCR2 CHEMOKINE RECEPTOR

Background of the Invention

The present invention relates to a compound useful as a modulator of chemokine receptors, particularly as CCR2 receptors.

Chemokines are chemotactic cytokines, of molecular weight 6-15 kDa, that are released by a wide variety of cells to attract and activate, among other cell types, macrophages, T and B lymphocytes, eosinophils, basophils and neutrophils (reviewed in: Luster, New Eng. J. Med. 1998, 338, 436-445 and Rollins, Blood 1997, 90, 909-928). There are two major classes of chemokines, CXC and CC, depending on whether the first two cysteines in the amino acid sequence are separated by a single amino acid (CXC) or are adjacent (CC). The CXC chemokines, such as interleukin-8 (IL-8), neutrophil-activating protein-2 (NAP-2) and melanoma growth stimulatory activity protein (MGSA) are chemotactic primarily for neutrophils and T lymphocytes, whereas the CC chemokines, such as RANTES, MIP-Ia, MlP-β, the monocyte chemotactic proteins (MCP-1 , MCP-2, MCP-3, MCP-4, and MCP-5) and the eotaxins (-1 and -2) are chemotactic for, among other cell types, macrophages, T lymphocytes, eosinophils, dendritic cells, and basophils. There also exist the chemokines lymphotactin-1 , lymphotactin-2 (both C chemokines), and fractalkine (a CX₃C chemokine) that do not fall into either of the major chemokine subfamilies.

The chemokines bind to specific cell-surface receptors belonging to the family of G- protein-coupled seven-transmembrane-domain proteins (reviewed in: Horuk, Trends Pharm. Sci. 1994, 15, 159-165) which are termed "chemokine receptors." On binding their cognate ligands, chemokine receptors transduce an intracellular signal through the associated trimeric G proteins, resulting in, among other responses, a rapid increase in intracellular calcium concentration, changes in cell shape, increased expression of cellular adhesion molecules, degranulation, and promotion of cell migration. There are at least ten human chemokine receptors that bind or respond to CC chemokines with following characteristic patterns (reviewed in Zlotnik and Oshie Immunity 2000, 12, 121): CCR-1 (or "CKR-1" or "CC-CKR-1 ") [MIP-1α, MCP-3, MCP-4, RANTES] (Ben-Barruch, et al., ce// 1993, 72, 415-425, and Luster, New Eng. J. Med. 1998, 338, 436-445; CCR2A and CCR2B (or "CKR-2AVCKR-2B" or "CC-CKR-2AVCC-CKR-2B") [MCP-1 , MCP-2, MCP-3, MCP-4, MCP-5] (Charo, et al., Proc. Natl. Acad. Sci. USA 1994, 91 , 2752-2756 Luster, New Eng. J Med. 1998, 338, 436-445); CCR-3 (or "CKR-3" or "CC-CKR-3") [eotaxin-1 , eotaxin-2, RANTES, MCP-3, MCP-4] (Combadiere, et al., J. Biol. Chem. 1995, 270, 16491-16494, and Luster, New Eng. J. Med. 1998, 338, 436-445); CCR-4 (or "CKR-4" or "CC-CKR-4") [TARC, MDC] (Power, et al., J. Biol. Chem. 1995, 270, 19495- 19500, and Luster, New Eng. J. Med. 1998, 338, 436-445); CCR-5 (or "CKR-5" or "CC- CKR-5") [MIP-1α, RANTES, MIP-1B] (Sanson, et al., Biochemistry 1996, 35, 3362-3367); CCR-6(or "CKR-6" [LARC] (Baba, et al., J. Biol. Chem. 1997, 272, 14893-14898); CCR-7 (or "CKR-7" or "CC-CKR-7") [ELC] (Yoshie et al., J. Leukoc. Biol. 1997, 62, 634-644); CCR-8 (or "CKR-8" or "CC-CKR-8") [1-309J (Napolitano et al., J. Immunol., 1996, 157, 2759-2763); CCR-10 (or "CKR-IO" or "CC-CKR-IO") [MCP-1 , MCP-3] (Bonini, et al., DNA and Cell Biol. 1997, 16, 1249-1256); and CCR-11 [MCP-1 , MCP-2, and MCP-4] (Schweickert, et al., J. biot Chem. 2000, 275, 90550).

In addition to the mammalian chemokine receptors, mammalian cytomegaloviruses, herpe's viruses and poxviruses have been shown to express, in infected cells, proteins with the binding properties of chemokine receptors (reviewed in: Wells and Schwartz, Curr. Opin. Biotech. 1997, 8, 741-748). Human CC chemokines, such as RANTES and MCF-3, can cause rapid mobilization of calcium via these virally encoded receptors. Receptor expression may be permissive for infection by allowing for the subversion of normal immune system surveillance and response to infection. Additionally, human chemokine receptors, such as CXCR4, CCR2, CCR3, CCR5 and CCR8, can act as co- receptors for the infection of mammalian cells by microbes as with, for example, the human immunodeficiency viruses (HIV).

The chemokines and their cognate receptors have been implicated as being important mediators of inflammatory, infectious, and immunoregulatory disorders and diseases, including asthma and allergic diseases, as well as autoimmune pathologies such as rheumatoid arthritis and atherosclerosis (reviewed in: F. H. Carter, Current Opinion in Chemical Biology 2002, 6, 510; Trivedi, et al, Ann. Reports Med. Chem. 2000, 35, 191 ; Saunders and Tarby, Drug Disc. Today 1999, 4, 80; Fremack and Schall, Nature Medicine 1996, 2, 1174). For example, the chemokine monocyte chemoattractant-l (MCP-1 ) and its receptor CC Chemokine Receptor 2 (CCR2) play a pivotal role in attracting leukocytes to sites of inflammation and in subsequently activating these cells. When the chemokine MCP-1 binds to CCR2, it induces a rapid increase in intracellular calcium concentration, increased expression of cellular adhesion molecules, cellular degranulation, and the promotion of leukocyte migration. Demonstration of the importance of the MCP-1/CCR2 interaction has been provided by experiments with genetically modified mice. MCP-1 -/- mice had normal numbers of leukocytes and macrophages, but were unable to recruit monocytes into sites of inflammation after several different types of immune challenge (Bao Lu, et al., J. Exp. Med. 1998, 187, 601). Likewise, CCR2 -/- mice were unable to recruit monocytes or produce interferon-γ when challenged with various exogenous agents; moreover, the leukocytes of CCR2 null mice did not migrate in response to MCP-1 (Landin Boring, et al., J. Clin. Invest. 1997, 100, 2552), thereby demonstrating the specificity of the MCP-1 /CCR2 interaction. Two other groups have independently reported equivalent results with different strains of CCR2 -/- mice (William A. Kuziel, et al., Proc. Natl. Acad. ScL USA 1997, 94, 12053, and Takao Kurihara, et al., J. Exp. Med. 1997, 186, 1757). The viability and generally normal health of the MCP-1 -/- and CCR2 -/- animals is noteworthy, in that disruption of the MCP-1 /CCR2 interaction does not induce physiological crisis. Taken together, these data lead one to the conclusion that molecules that block the actions of MCP-1 would be useful in treating a number of inflammatory and autoimmune disorders. This hypothesis has now been validated in a number of different animal disease models, as described below.

It is known that MCP-1 is upregulated in patients with rheumatoid arthritis (Alisa Koch, et al., J. Clin. Invest. 1992, 90, 772 - 779). Moreover, several studies have demonstrated the potential therapeutic value of antagonism of the MCP-1 /CCR2 interaction in treating rheumatoid arthritis. A DNA vaccine encoding MCP-1 was shown recently to ameliorate chronic polyadjuvant-induced arthritis in rats (Sawsan Youssef, et al., J. Clin. Invest. 2000, 106, 361). Likewise, 'inflammatory disease symptoms could be controlled via direct administration of antibodies for MCP-1 to rats with collagen-induced arthritis (Hiroomi Ogata, et al., J. Pathol. 1997, 182, 106), or streptococcal cell wall-induced arthritis (Ralph C. Schimmer, et al., J. Immunol. 1998, 160, 1466). Perhaps most significantly, a peptide antagonist of MCP-1 , MCP-1 (9-76)1 was shown both to prevent disease onset and to reduce disease symptoms (depending on the time of administration) in the MRL-1pr mouse model of arthritis (Jiang-Hong Gong, et al., J. Exp. Med. 1997, 186, 131 ).

It is known that MCP-1 is upregulated in atherosclerotic lesions, and it has been shown that circulating levels of MCP-1 are reduced through treatment with therapeutic agents, plays a role in disease progression (Abdolreza Rezaie-Majd, et al, Arterioscler. Thromb. Vase. Biol. 2002, 22, 1194 - 1199). Four key studies have demonstrated the potential therapeutic value of antagonism of the MCP-1/CCR2 interaction in treating atherosclerosis. For example, when MCP-1 -/- mice are mated with LDL receptor- deficient mice, an 83% reduction in aortic lipid deposition was observed (Long Gu, et al., MoI. Cell 1998, 2, 275). Similarly, when MCP-1 was genetically ablated from mice which already overexpressed human apolipoprotein B, the resulting mice were protected from atherosclerotic lesion formation relative to the MCP-1 +/+ apoB control mice (Jennifa Gosling, et al., J. Clin. Invest. 1999, 103, 773). Likewise, when CCR2 -/- mice are crossed with apolipoprotein E -/- mice, a significant decrease in the incidence of atherosclerotic lesions was observed (Landin Boring, et al, Nature 1998, 394, 894). Finally, when apolipoprotein E -/- mice are administered a gene encoding a peptide antagonist of CCR2, then lesion size is decreased and plaque stability is increased (W Ni, et al. Circulation 2001, 103, 2096 -2101).

It is known that MCP-1 is upregulated in human multiple sclerosis, and it has been shown that effective therapy with interferon b-lb reduces MCP-1 expression in peripheral blood mononuclear cells, suggesting that MCP-1 plays a role in disease progression (Carla larlori, et al., J. Neuroimmunol. 2002, 123, 170 - 179). Other studies have demonstrated the potential therapeutic value of antagonism of the MCP-1 /CCR2 interaction in treating multiple sclerosis; all of these studies have been demonstrated in experimental autoimmune encephalomyelitis (EAE), the conventional animal model for multiple sclerosis. Administration of antibodies for MCP-1 to animals with EAE significantly diminished disease relapse (K. J. Kennedy, et al., J. Neuroimmunol. 1998, 92, 98). Furthermore, two recent reports have now shown that CCR2 -/- mice are resistant to EAB (Brian T. Fife, et al., J. Exp. Med. 2000, 192, 899; Leonid Izikson, et al., J. Exp. Med. 2000, 192, 1075).

It is known that MCP-1 is upregulated in patients who develop bronchiolitis obliterans syndrome after lung transplantation (Martine Reynaud-Gaubert, et al., J. of Heart and Lung Transplant, 2002, 21, 721 - 730; John Belperio, et al., J. Clin. Invest. 2001, 108, 547 - 556). In a murine model of bronchiolitis obliterans syndrome, administration of an antibody to MCP-1 led to attenuation of airway obliteration; likewise, CCR2 -/- mice were resistant to airway obliteration in this same model (John Belperio, et al., J. CHn. Invest. 2001, 108, 547 - 556). These data suggest that antagonism of MCP-1/CCR2 may be beneficial in treating rejection of organs following transplantation. Other studies have demonstrated the potential therapeutic value of antagonism of the MCP-1/CCR2 interaction in treating asthma. Sequestration of MCP-1 with a neutralizing antibody in ovalbumin-challenged mice resulted in marked decrease in bronchial hyperresponsiveness and inflammation (Jose-Angel Gonzalo, et al., J. Exp. Med. 1998, 188, 157). It proved possible to reduce allergic airway inflammation in Schistosoma mansoni egg-challenged mice through the administration of antibodies for MCP-1 (Nicholas W. Lukacs, et al., J. Immunol. 1997, 158, 4398). Consistent with this, MCP-1 - /- mice displayed a reduced response to challenge with Schistosoina mansoni egg (Bao Lu, et al., J. Exp. Med. 1998, 187, 601).

Other studies have demonstrated the potential therapeutic value of antagonism of the MCP-1 /CCR2 interaction in treating kidney disease. Administration of antibodies for MCP-1 in a murine model of glomerularnephritis resulted in a marked decrease in glomerular crescent formation and deposition of type I collagen (Clare M. Lloyd, et al., J. Exp. Med. 1997, 185, 1371). In addition, MCP-1 -/- mice with induced nephrotoxic serum nephritis showed significantly less tubular damage than their MCP-1 +/+ counterparts (Gregory H. Tesch, et al.7 J. Cur. Invest. 1999, 103, 73).

One study has demonstrated the potential therapeutic value of antagonism of the MCP- 1/CCR2 interaction in treating systemic lupus erythematosus. Crossing of MCP-1 -/- mice with MRL-FAS1 pr mice (the latter of which have a fatal autoimmune disease that is analogous to human systemic lupus erythematosus) results in mice that have less disease and longer survival than the wildtype MRLMSipr mice (Gregory H. Tesch, et al., J. Exp. Med. 1999, 190, 1813). One study has demonstrated the potential therapeutic value of antagonism of the MCP-1 /CCR2 interaction in treating colitis. CCR2 -/- mice were protected from the effects of dextran sodium sulfate-induced colitis (Pietro G. Andres, et al., J. Immunol. 2000, 164, 6303).

One study has demonstrated the potential therapeutic value of antagonism of the MCP- 1/CCR2 interaction in treating alveolitis. When rats with IgA immune complex lung injury were treated intravenously with antibodies raised against rat MCP-1 (JE), the symptoms of alveolitis were partially alleviated (Michael L. Jones, et al., J. Immunol. 1992, 149, 2147).

One study has demonstrated the potential therapeutic value of antagonism of the MCP- 1/CCR2 interaction in treating cancer. When immunodeficient mice bearing human breast carcinoma cells were treated with an anti-MCP-1 antibody, inhibition of lung micrometastases and increases in survival were observed (Rosalba Salcedo, et al, Blood 2000, 96, 34 - 40).

One study has demonstrated the potential therapeutic value of antagonism of the MGP- 1/CCR2 interaction in treating restinosis. Mice deficient in CCR2 showed reductions in the intimal area and in the intima/media ratio (relative to wildtype littermates) after injury of the femoral artery (Merce Roque, et al. Arterioscier. Thromb. Vase. PIoI. 2002, 22, 554 - 559).

Other studies have provided evidence that MCP-1 is overexpressed in various disease states not mentioned above. These reports provide correlative evidence that MCP-1 antagonists could be useful therapeutics for such diseases. Two reports described the overexpression of MCP-1 in the intestinal epithelial cells and bowel mucosa of patients with inflammatory bowel disease (H. C. Reinecker, et al., Gastroenterology 1995, 108, 40, and Michael C. Grimm, et al., J. Leukoc. Biol. 1996, 59, 804). Two reports describe the overexpression of MCP-1 rats with induced brain trauma (J. S. King, et al., J. Neuroimmunol. 1994, 56, 127, and Joan W. Berman, et al., J. lininunol. 1996, 756, 3017). Another study has demonstrated the overexpression of MCP-1 in rodent cardiac allografts, suggesting a role for MCP-1 in the pathogenesis of transplant arteriosclerosis (Mary E. Russell, et al. Proc. Natl. Acad. ScL USA 1993, 90, 6086). The overexpression of MCP-1 has been noted in the lung endothelial cells of patients with idiopathic pulmonary fibrosis (Harry N. Antoniades, et al., Proc. Natl. Acad. Sci. USA 1992, 89, 5371). Similarly, the overexpression of MCP-1 has been noted in the skin from patients with psoriasis (M. Deleuran, et al., J. Deήnatol. Sci. 1996, 13, 228, and R. Gillitzer, et al., J. Invest. Dermatol. 1993, 101, 127). Finally, a recent report has shown that MCP-1 is overexpressed in the brains and cerebrospinal fluid of patients with HIV-associated dementia (Alfredo Garzino-Demo, WO 99/46991).

It should also be noted that CCR2 has been implicated as a co-receptor for some strains of HIV (B. J. Doranz, et al., Cell 1996, 85, 1149). It has also been determined that the use of CCR2 as an HIV co-receptor can be correlated with disease progression (Ruth I. Connor, et al., J. Exp. Med. 1997, 185, 621). This finding is consistent with the recent finding that the presence of a CCR2 mutant, CCR2-641 , is positively correlated with delayed onset of HIV in the human population (Michael W. Smith, et al., Science 1997,

277,959). Although MCP-1 has not been implicated in these processes, it may be that MCP-1 antagonists that act via binding to CCR2 may have beneficial therapeutic effects in delaying the disease progression to AIDS in HIV-infected patients.

A new class of compounds which have affinity for chemokine receptors, in particular the CCR2 receptor, has now been found. These compounds have potential in the treatment of conditions wherein modulation, especially antagonism/inhibition, of the CCR2 receptor is thought to be beneficial.

There is a need in the art to discover compounds that antagonize MCP-1 by binding to the CCR2 receptor, more particularly the CCR2b receptor.

Summary of the Invention

In a first aspect, the present invention is a compound as represented in formula (I):

or a salt thereof, or a solvate thereof, or a combination thereof; wherein

one of D and E represents a nitrogen atom, and the other of D and E represents CH;

R¹ represents an aryl, a thienyl, a benzothienyl, an imidazolyl, a pyridyl, an isoquinolinyl, a piperonyl, a benzoxathiadiazolyl, or a benzoxadiazolyl group substituted by up to three R⁴ groups, wherein each R⁴ group is independently selected from the group consisting of C_1-6-alkyl, C_1-6-haloalkyl, C-|.₆-alkoxy, C^e-haloalkoxy, halo, -NH₂, -OH, -CN, -NO₂, CF₃, phenyl, phenoxy, phenyl-C(O)-, isoxazolyl, and -C(O)NR⁷R⁸, wherein R⁷ and R⁸ each independently represent hydrogen or Ci_-4 alkyl, or R⁷ and R⁸ together with the nitrogen atom to which they are attached form a 5- or 6-membered saturated heterocylic group optionally containing an additional heteroatom selected from nitrogen, oxygen and sulphur; m is 1 , 2, or 3;

each R² is independently selected from the group consisting of halo, -CN, and -CF₃-;

R³ is a heteroaryl group optionally substituted by one to three substituents independently selected from the group consisting of halo, hydroxy-C_1-4-alkyl-, C^alkoxy-C^alkyl-, C₁. ₄alkoxy-, R¹⁰R¹¹N-Ci.₄alkyl-, C_1-6-aikyl,

-CN, C_1-4alkylthio-, -CF₃, -CO₂H, - CONR⁷R⁸, and -CO-Ci_-4 alkyl, and R¹⁰ and R¹¹ each independently represent hydrogen or C_1-4 alkyl.

In a second aspect, the present invention is a method for treating or preventing a disease or condition mediated by CCR2 comprising administering to a patient in need thereof a pharmaceutically effective amount of the compound of formula (I), or a pharmaceutically acceptable salt thereof, or a solvate thereof, of a combination thereof.

In a third aspect, the present invention provides a pharmaceutical composition comprising a) a compound of formula (I) or or a pharmaceutically acceptable salt thereof, or a solvate thereof, of a combination thereof and b) a pharmaceutically acceptable carrier, diluents, or excipient or combination thereof and, optionally, c) another therapeutic or prophylactic agent or combination thereof.

The compounds of the present invention are believed to be effective in the treatment of diseases such as atherosclerosis, asthma, seasonal and perennial allergic rhinitis, sinusitis, conjunctivitis, food allergy, scombroid poisoning, pulmonary fibrosis, restenosis, including vascular restenosis, myocarditis, ulcerative colitis, psoriasis, urticaria, pruritis, eczema, atopic dermatitis, inflammatory bowel disease, chronic obstructive pulmonary disease, thrombotic disease, otis media, rheumatoid arthritis, nephritis (nephropathy), liver cirrhosis, multiple sclerosis and systemic sclerosis, lupus, erthematosis, hepatitis, pancreatitis, sarcoidosis, organ transplantation, Crohn's disease, endometriosis, cardiac disease, congestive heart failure, viral meningitis, cerebral infarction, neuropathy, Kawasaki disease, Alzheimer's disease, stroke, acute nerve injury, HIV infection, AIDS, autoimmune diseases, cancer, and sepsis. Detailed Description of the Invention

It should be noted that CCR2 is also the receptor for the chemokines MCP-2, MCP-3, MCP-4, and MCP-5 (Luster, New Eng. J. Med. 1998, 338, 436-445). Since the new compounds of formula (I) described herein antagonize MCP-1 by binding to the CCR2 receptor, it may be that these compounds of formula (I) are also effective antagonists of the actions of MCP-2, MCP-3, MCP-4, and MCP-5 that are mediated by CCR2.

For the avoidance of doubt, the term "independently" means that where more than one substituent is selected from a number of possible substituents, those substituents may be the same or different.

As used herein, the term "alkyl" refers to straight or branched hydrocarbon chains containing the specified number of carbon atoms. For example, Ci.₆alkyl means a straight or branched alkyl containing 1 to 6, carbon atoms. Examples of "alkyl" as used herein include, but are not limited to, methyl, ethyl, n-propyl, n-butyl, n-pentyl, n-hexyl, isobutyl, isopropyl, t-butyl and 1 ,1-dimethylpropyl.

As used herein, the term "alkoxy" refers to a straight or branched alkoxy group containing the specified number of carbon atoms. For example, d.₆alkoxy means a straight or branched alkoxy group containing 1 to 6, carbon atoms. Examples of "alkoxy" include, but are not limited to methoxy, ethoxy, propoxy, prop-2-oxy, butoxy, but-2-oxy, 2-methylprop-1 -oxy, 2-methylprop-2-oxy, pentoxy or hexyloxy.

As used herein, the term "alkylthio" refers to a straight or branched alkylthio group containing the specified number of carbon atoms. For example, Ci-₄alkylthio means a straight or branched alylthio group containing 1 to 6, carbon atoms. Examples of "alkylthio" as used herein include, but are not limited to methylthio, ethylthio, propylthio, prop-2-thio, butylthio, but-2-thio, 2-methylprop-i-thio, 2-methylprop-2-thio.

As used herein the term C₃.₆ cycloalkyl includes cyclopropyl, cyclobutyl, cyclopentyl, and cyclohexyl groups.

The term halogen or halo refers to fluoro, chloro, bromo and iodo. Examples of heterocycloalkyl groups include, but are not limited to pyrrolidinyl, imidazolidinyl, pyrazolidinyl, piperidinyl, morpholino, thiomorpholino, piperazinyl, oxopiperidinyl, oxophenyltriazaspirodecyl, oxopyrazolidinyl, oxopyrazolyl, oxooxazolidinyl, oxoimidazolidinyl, dioxaazaspirodecyl groups.

Heteroaryl groups include, but are not limited to, pyrrolyl, oxazolyl, isoxazolyl, thiazolyl, isothiazolyl, imidazolyl, pyrazolyl, furazanyl, oxadiazolyl, thiadiazolyl, triazolyl, tetrazolyl, pyridyl, pyridazinyl, pyrimidinyl, pyrazinyl, triazinyl, benzofuranyl, isobenzofuranyl, benzothienyl, indolyl, isoindolyl, benzoxazolyl, benzthiazolyl, indazolyl, indolazinyl, benzimidazolyl, benzotriazolyl, purinyl, coumarinyl, isocoumarinyl, chromonyl, quinolinyl, isoquinolinyl, cinnolinyl, quinazolinyl, phthalazinly, quinoxalinyl, pteridinyl, 1 ,8- naphthyridinyl, oxo- or dioxo- benzoxazolyl, -benzotriazolyl, -pyridinyl, -pyrazolyl, - benzopiperidinyl and -benzopiperazinyl.

As used herein, the term "pharmaceutically acceptable" means suitable for pharmaceutical use.

As used herein, the term "solvate" refers to a complex of variable stoichiometry formed by a solute (in this invention, a compound of formula (I) or a salt or physiologically acceptable derivative thereof) and a solvent. Such solvents for the purpose of the invention may not interfere with the biological activity of the solute. Examples of suitable solvents include water, methanol, ethanol and acetic acid. Most preferably the solvent used is water and the solvate may also be referred to as a hydrate.

The compounds of formula (I) as defined above contain a basic grouping and may also contain an acidic grouping and therefore may form salts with physiologically acceptable acids or bases.

As used herein, the term "salt" refers to any salt of a compound according to the present invention prepared from an inorganic or organic acid or base, quaternary ammonium salts and internally formed salts.

Physiologically acceptable salts are particularly suitable for medical applications because of their greater aqueous solubility relative to the parent compounds. Such salts must clearly have a physiologically acceptable anion or cation. Suitably physiologically acceptable salts of the compounds of the present invention include acid addition salts formed with inorganic acids such as hydrochloric, hydrobromic, hydroiodic, phosphoric, metaphosphoric, nitric and sulfuric acids, and with organic acids, such as tartaric, acetic, trifluoroacetic, citric, malic, lactic, fumaric, benzoic, formic, propionic, glycolic, gluconic, maleic, succinic, camphorsulfuric, isothionic, mucic, gentisic, isonicotinic, saccharic, glucuronic, furoic, glutamic, ascorbic, anthranilic, salicylic, phenylacetic, mandelic, embonic (pamoic), methanesulfonic, ethanesulfonic, pantothenic, stearic, sulfinilic, alginic, galacturonic and arylsulfonic, for example benzenesulfonic and p-toluenesulfonic, acids; base addition salts formed with alkali metals and alkaline earth metals and organic bases such as N.N-dibenzylethylenediamine, chloroprocaine, choline, diethanolamine, ethylenediamine, meglumaine (N-methylglucamine), lysine and procaine; and internally formed salts. Salts having a non-physiologically acceptable anion or cation are within the scope of the invention as useful intermediates for the preparation of physiologically acceptable salts and/or for use in non-therapeutic, for example, in vitro, situations.

Certain of the compounds of the invention may form acid addition salts with one or more equivalents of the acid. Certain of the compounds of the invention may form acid addition salts with less than one equivalent of the acid. The present invention includes within its scope all possible stoichiometric and non-stoichiometric forms.

Certain compounds of formula (I) may exist in stereoisomeric forms (e.g. they may contain one or more asymmetric carbon atoms). The individual stereoisomers (enantiomers and diastereomers) and mixtures of these are included within the scope of the present invention. The present invention also covers the individual isomers of the compounds represented by formula (I) as mixtures with isomers thereof in which one or more chiral centres are inverted. Likewise, it is understood that compounds of formula (I) may exist in tautomeric forms other than that shown in the formula and these are also included within the scope of the present invention.

When a specific enantiomer of a compound of general formula (I) is required, this may be obtained for example by resolution of a corresponding enantiomeric mixture of a compound of formula (I) using conventional methods. Thus the required enantiomer may be obtained from the racemic compound of formula (I) by use of chiral HPLC procedure.

Compounds of the present invention include the compounds of Examples 1 to 8 referred to below, inter alia including all compounds referenced in Table 1. It will be appreciated by those skilled in the art that reference herein to treatment extends to prophylaxis as well as the treatment of established diseases or symptoms.

While it is possible that, for use in therapy, a compound of the invention may be administered as the raw chemical, it is preferable to present the active ingredient as a pharmaceutical formulation.

The carrier, diluent and excipient must be 'acceptable' in the sense of being compatible with the other ingredients of the formulation and not deleterious to the patient.

The compounds of the invention may be administered in conventional dosage forms prepared by combining a compound of the invention with standard pharmaceutical carriers, diluents or excipients according to conventional procedures well known in the art. These procedures may involve mixing, granulating and compressing or dissolving the ingredients as appropriate to the desired preparation.

The pharmaceutical compositions of the invention may be formulated for administration by any route, and include those in a form adapted for oral, buccal, topical, inhalation or insufflation, implant, rectal or parenteral administration to mammals including humans.

The compositions may be in the form of tablets, capsules, powders, granules, lozenges, creams or liquid preparations, such as oral or sterile parenteral solutions or suspensions.

Tablets and capsules for oral administration may contain conventional excipients such as binding agents, for example, syrup, acacia, gelatin, sorbitol, tragacanth, mucilage of starch or polyvinylpyrrolidone; fillers, for example, lactose, sugar, microcystalline cellulose, maize-starch, calcium phosphate, glycine or sorbitol; lubricants, for example, magnesium stearate, stearic acid, talc, polyethylene glycol or silica; disintegrants, for example, potato starch or sodium starch glycollate, or wetting agents such as sodium lauryl sulphate. The tablets may be coated according to methods well known in the art. Oral liquid preparations may be in the form of, for example, aqueous or oily suspensions, solutions emulsions, syrups or elixirs, or may be presented as a dry product for constitution with water or other suitable vehicle before use. Such liquid preparations may contain conventional additives such as suspending agents, for example, sorbitol syrup, methyl cellulose, glucose/sugar syrup, gelatin, hydroxyethylcellulose, carboxymethyl cellulose, aluminium stearate gel or hydrogenated edible fats; emulsifying agents, for example, lecithin, sorbitan mono-oleate or acacia; non-aqueous vehicles (which may include edible oils), for example, almond oil, fractionated coconut oil, oily esters, propylene glycol or ethyl alcohol; solubilizers such as surfactants for example polysorbates or other agents such as cyclodextrins; and preservatives, for example, methyl or propyl p-hydroxybenzoates or ascorbic acid; and, if desired, conventional flavouring or colouring agents. The compositions may also be formulated as suppositories, e.g. containing conventional suppository bases such as cocoa butter or other glycerides.

For buccal administration the composition may take the form of tablets or lozenges formulated in conventional manner.

The topical formulations of the present invention may be presented as, for instance, ointments, creams or lotions, eye ointments and eye or ear drops, impregnated dressings and aerosols, and may contain appropriate conventional additives such as preservatives, solvents to assist drug penetration and emollients in ointments and creams.

The formulations may also contain compatible conventional carriers, such as cream or ointment bases and ethanol or oleyl alcohol for lotions. Such carriers may be present as from about 1% up to about 98% of the formulation. More usually they will form up to about 80% of the formulation.

The composition according to the invention may be formulated for parenteral administration by injection or continuous infusion. Formulations for injection may be presented in unit dose form in ampoules, or in multi-dose containers with an added preservative. The compositions may take such forms as suspensions, solutions, or emulsions in oily or aqueous vehicles, and may contain formulatory agents such as suspending, stabilising and/or dispersing agents. Alternatively the active ingredient may be in powder form for constitution with a suitable vehicle, e.g. sterile, pyrogen-free water, before use.

For parenteral administration, fluid unit dosage forms are prepared utilising the compound and a sterile vehicle, water being preferred. The compound, depending on the vehicle and concentration used, can be either suspended or dissolved in the vehicle. In preparing solutions the compound can be dissolved in water for injection and filter sterilised before filling into a suitable vial or ampoule and sealing.

Advantageously, agents such as a local anaesthetic, preservative and buffering agents can be dissolved in the vehicle. To enhance the stability, the composition can be frozen after filling into the vial and the water removed under vacuum. The dry lyophilised powder is then sealed in the vial and an accompanying vial of water for injection may be supplied to reconstitute the liquid prior to use. Parenteral suspensions are prepared in substantially the same manner except that the compound is suspended in the vehicle instead of being dissolved and sterilisation cannot be accomplished by filtration. The compound can be sterilised by exposure to ethylene oxide before suspending in the sterile vehicle. Advantageously, a surfactant or wetting agent is included in the composition to facilitate uniform distribution of the compound.

The compositions according to the invention may contain between 0.1-99% of the active ingredient depending on the method of administration, conveniently from 30-95% for tablets and capsules and 3-50% for liquid preparations.

Since the compounds of the invention are intended for use in pharmaceutical compositions it will readily be understood that they are each preferably provided in substantially pure form, for example at least 60% pure, more suitably at least 75% pure and preferably at least 85%, especially at least 98% pure (% are on a weight for weight basis). Impure preparations of the compounds may be used for preparing the more pure forms used in the pharmaceutical compositions; these less pure preparations of the compounds should contain at least 1 %, more suitably at least 5% and preferably from 10 to 59% of a compound of the invention.

It will further be appreciated that the amount of a compound of the invention required for use in treatment will vary with the nature of the condition being treated, the route of administration and the age and the condition of the patient and will be ultimately at the discretion of the attendant physician. In general however doses employed for adult human treatment will typically be in the range of 1 to 10OOmg per day, dependent upon the route of administration. Thus for parenteral administration a daily dose will typically be in the range 1 to 10Omg, or 5 to 50mg per day. For oral administration a daily dose will typically be within the range 1 to ^"lOOOmg, e.g. 5 to 500 mg per day.

The desired dose may conveniently be presented in a single dose or as divided doses administered at appropriate intervals, for example as two, three, four or more sub-doses per day.

Where the compositions comprise dosage units, each unit will typically contain from 1 -1000 mg of the active ingredient.

Compounds of the invention may be prepared, in known manner, in a variety of ways. In the following reaction schemes and hereafter, unless otherwise stated R1 to R3 etc. are as defined above. These processes form further aspects of the invention.

Throughout the specification, general formulae are designated by Roman numerals (I), (II), (III), (IV) etc.

Further details for the preparation of compounds of formula (I) are found in the Examples section hereinafter.

The compounds of the invention may be prepared singly or as compound libraries comprising at least 2, for example 5 to 1 ,000 compounds, and more preferably 10 to 100 compounds. Libraries of compounds of the invention may be prepared by a combinatorial 'split and mix¹ approach or by multiple parallel synthesis using either solution phase or solid phase chemistry, by procedures known to those skilled in the art. Thus according to a further aspect there is provided a compound library comprising at least 2 compounds of the invention.

Those skilled in the art will appreciate that in the preparation of the compounds of the invention or a solvate thereof it may be necessary and/or desirable to protect one or more sensitive groups in the molecule to prevent undesirable side reactions. Suitable protecting groups for use according to the present invention are well known to those skilled in the art and may be used in a conventional manner. See, for example, "Protective groups in organic synthesis" by T.W. Greene and P.G.M. Wuts (John Wiley & sons 1991 ) or "Protecting Groups" by PJ. Kocienski (Georg Thieme Verlag 1994). Examples of suitable amino protecting groups include acyl type protecting groups (e.g. formyl, trifluoroacetyl, acetyl), aromatic urethane type protecting groups (e.g. benzyloxycarbonyl (Cbz) and substituted Cbz), aliphatic urethane protecting groups (e.g. 9-fluorenylmethoxycarbonyl (Fmoc), t-butyloxycarbonyl (Boc), isopropyloxycarbonyl, cyclohexyloxycarbonyl) and alkyl type protecting groups (e.g. benzyl, trityl, chlorotrityl). Examples of suitable oxygen protecting groups may include for example alkyl silyl groups, such as trimethylsilyl or tert-butyldimethylsilyl; alkyl ethers such as tetrahydropyranyl or tert-butyl; or esters such as acetate.

Compounds of formula (I) may be prepared by the reaction of a compound of formula (II)

wherein D, E, G, m, R² and R³ are as defined above, with a compound of formula (III)

L-SO₂-R¹ (III)

wherein L is a suitable leaving group, and R¹ is as defined above;

and thereafter optionally,

(i) removing any protecting group(s); and/or

(ii) forming a salt; and/or

(iii) converting a compound of formula (I) or a salt thereof to another compound of formula (I) or a salt thereof.

Suitable leaving groups L include chloro, bromo or pentafluorophenoxy.

Typically, where L represents chloro, such a reaction may be carried out by dissolving the compound of formula (II) in a suitable solvent, for example pyridine optionally mixed with a second solvent, such as chloroform or tetrahydrofuran, and reacting it with the compound of formula (III) also in a suitable solvent, for example pyridine. The addition of a catalytic quantity of dimethylaminopyridine may also be used. The reaction would generally be carried out at elevated temperature in the region of 80-250^eC, for example at about 200^eC, for a period of 30 minutes to 1 hour or at 80⁰C for a period of 5-24 hours.

Compounds of the formula (II), wherein D represents N, may be prepared according to the chemistry detailed in Scheme 1. Treatment of acetic-acid esters bearing the relevant group R³, readily prepared by a person skilled in the art according to literature methods, may be treated with an appropriate base, for example sodium hydride, in a solvent such as dimethyl formamide and a nitro-pyridine bearing an appropriate leaving group L, such as chlorine. The coupled product may then be decarboxylated upon treatment with aqueous acid, such as hydrochloric acid, in ethanol or similar solvent. The product nitro- pyridine may then be reduced using standard literature techniques; for example, hydrogenation in the presence of a metal catalyst such as platinum or palladium.

Scheme 1

Similar processes would apply to the compounds of formula (I) wherein E or G represented a nitrogen atom.

Interconversion reactions between compounds of formula (I) may be performed using methods well known in the art. Included are conversions of the substituent(s) within the group R³ and/or conversions within groups R² and R¹ for example:

(i) converting an acid to an acid amide;

(ii) converting an acid to an acid ester;

(iii) converting an acid amide to a cyano.

(iv) converting an aniline to a cyano

(v) converting an aniline to a chloro.

Compounds of formula (I) wherein R¹ represents Ar, R² represents -CN in the 5-position, R³ represents tetrazole and D represents N can be prepared according to Scheme 2 below: Scheme 2

LiOH

The nitro intermediate may be converted into the corresponding amine by, for example hydrogenation in the presence of a suitable catalyst in an appropriate solvent such as a mixture of DCM and an alcohol such as ethanol in the presence of ammonium formate at an elevated temperature, for example heating to 6O⁰C for 18 hours, although the reaction may be run at any temperature over 50⁰C for up to 24 hours. The reduction can be driven to completion by filtration and re-exposing the intermediate to similar reaction conditions. The amine coupling step may be effected in a suitable solvent such as pyridine at an elevated temperature such as 90⁰C for 24 hours, although the reaction may be conducted at any temperature up to the point of reflux and may be complete in less than 24 hours.

The deprotection step to provide the free carboxylic acid may be effected by treatment with lithium hydroxide at an elevated temperate such as 55°C, for example for a period less than 5 hours such as 2 hours, wherein the reaction is performed in a suitable solvent, for example an alcohol such as methanol.

In the last step the carboxylic acid substituent may be converted into the required CN group by treatment with phosphorus oxychloride in a suitable solvent, for example chloroform for a period such as 18 to 24 hours at an elevated temperature, such as reflux.

The following non-limiting examples illustrate the present invention. Abbreviations

HPLC - high-performance liquid chromatography

DCM - dichloromethane

DMF - dimethylformamide

DMSO - dimethylsulfoxide

IMS - industrial methylated spirit

HCI - hydrochloric acid

Mass spectra were obtained using either a Waters ZQ mass spectrometer or Micromass Platform 2 mass spectrometer and use electro-spray ionisoation to observe either MH+ or M-. Proton Nuclear Magnetic Resonance (1 H-NMR) spectra were recorded at 400 MHz unless otherwise stated, chemical shifts are reported in ppm downfield from Me4Si, used as internal standard, and are assigned as singlets (s), doublets (d), doublets of doublets (dd), triplets (t), doublet of triplets (dt), quartets (q) multiplets (m) or are otherwise described in full. The prefix "br" refers to a broad peak; for example, a broad single may appear as br.s (or br s).

Examples

Intermediates 1 and 2: 5-chloro-2-f(2-methyl-2H-tetrazol-5-yl)methyl1-3-nitropyridine and 5-chloro-2-r(1-methyl-1 H-tetrazol-5-v0methyl1-3-nitropyridine

(+lsomer)

NaH (60% in mineral oil) (1.2 g) was washed with hexane (1OmL); and mixed with stirring with a solution of (2-methyl-2H-tetrazol-5-yl)acetate (3.25 g, see Chθm. Pharm. Bull., 1991, 1099. - contains other tetrazole isomer also) in anhydrous DMF (20 mL) at 20 ⁰C. The mixture was stirred for 2 minutes at 20 ⁰C under argon, whereupon a solution of 2,5-dichloronitropyridine (3.6 g) dissolved in DMF (10 mL) was added in one portion. The mixture turned dark purple and an exotherm was observed. The mixture was stirred for a further 20 minutes, quenched with 2M HCI (5 ml_) and the DMF was removed in vacuo.

The residue was dissolved in ethyl acetate (150 mL), washed with 2M HCI (100 ml_), water (100 mL) and brine (100 mL). The solvent was removed in vacuo to give 5.50 g of crude product, which was dissolved in concentrated HCI (25 mL) and IMS (75 mL) and heated under reflux for 20 hours. The mixture was cooled and extracted into DCM (350 mL); the DCM fraction was washed with water (2 x 300 mL) and brine (100 mL). The solvent was removed in vacuo and the resulting oil purified by column chromatography eluting with 6-35% ethyl acetate in hexane give the products that eluted in the order described.

5-chloro-2-[(2-methyl-2H-tetrazol-5-yl)methyl]-3-nitropyridine (0.93g, contaminated with tetrazole starting material) Cf₆-DMSO δ 8.90 (1 H, d), 8.74 (1 H,d), 4.31 (3H, s), 4.27 (2H, s).

5-chloro-2-[(1-methyl-1 H-tetrazol-5-yl)methyl]-3-nitropyridine as a yellow solid (1.2 g,

23%) CZ₆-DMSO δ 8.85 (1 H, d), 8.77 (1 H, d), 4.90 (2H, s), 3.97 (3H₁ s).

Intermediate 3: 5-chloro-2-f(1-methyl-1 H-tetrazol-5-yl)methvH-3-pyridinamine

5-chloro-2-[(1 -methyl-1 H-tetrazol-5-yl)methyl]-3-nitropyridine (1.1 g), 1% platinum on carbon (1 mol %), ammonium formate (2.3 g) and DCM (15 mL) were heated under reflux with vigorous stirring for 48 h under an atmosphere of argon. The mixture was cooled and the mixture was filtered through a pad of Dicalite eluting with a further portion of DCM (100 mL). The solvent was removed in vacuo to give the title compound (724 mg, 73%). ¹H NMR (c/₆-DMSO) δ 7.59 (1 H₁ d), 7.04 (1 H, d), 5.71 (2H, br.s), 4.29 (2H, s), 3.89 (3H, s).

Intermediate 4: 5-chloro-2-r(2-methyl-2H-tetrazol-5-yl)methyl1-3-pyridinamine

The title compound was prepared by a similar procedure to that described for Intermediate 3 from 0.9 g of 5-chloro-2-[(2-methyl-2H-tetrazol-5-yl)methyl]-3- nitropyridine. An aqueous extraction with HCI/Sodium hydroxide was included to remove impurities and give the title compound (111 mg). ¹H NMR (d₆-DMSO) δ 7.60 (1 H, d), 7.02 (1 H, d), 5.56 (2H, s.br), 4.31 (3H, s), 4.15 (2H, s).

Intermediate 5 : ethyl 6-r(2-methyl-2H-tetrazol-5-yl)methyll-5-nitro-3- pyridinecarboxylate

NaH (60% in mineral oil, 3.8 g) was washed with hexane (15 ml_) and mixed with stirring with a solution of (2-methyl-2W-tetrazol-5-yl)acetate (9.8 g, see Chem. Pharm. Bull., 1991, 1099- also contains other tetrazole isomer) in anhydrous DMF (60 ml_) at 0 ⁰C. The mixture was stirred for 5 minutes at 0 ⁰C under argon. A solution of ethyl 6-chloro-5- nitro-3-pyridinecarboxylate (17.4 g, see J. Chem. Soc, 1951 , 2590.) was dissolved in DMF (60 ml_) and added in one portion at 0 ⁰C. The mixture turned red and an exotherm was observed. The mixture was stirred for a further 20 minutes then quenched with 2M HCI (6 ml_), whereupon the DMF was removed in vacuo. The residue was dissolved in ethyl acetate (250 ml_), washed with 2M HCI (150 mL), water (2x150 ml_) and brine (100 mL). The solvent was removed in vacuo to give 18.4 g of crude product, which was dissolved in concentrated HCI (50 mL) and IMS (150 mL) and heated under reflux for 66 hours. The mixture was cooled, poured onto ice (300 g) and extracted into DCM (300 mL); the DCM fraction was washed with brine (200 mL). The solvent was removed in vacuo and the resulting oil purified by column chromatography eluting with 10-40% ethyl acetate in hexane give the title compound, partially contaminated with starting material but ca 75% pure.

¹H NMR (DMSO-Gf₆) δ 8.79 (s, 1 H), 8.62 (d, 1 H), 5.01 (s, 2H), 4.38 (q, 2H), 3.95 (s, 3H), 1.32 (t, 3H).

Intermediate 6 ethyl 5-amino-6-f(2-methyl-2H-tetrazol-5-yl)methyl1-3- pyridinecarboxylate

Ethyl 6-[(2-methyl-2H-tetrazol-5-yI)methyl]-5-nitro-3-pyridinecarboxylate (1.17 g) was dissolved in DCM/ethanol (1 :1 , 1OmL). 5% Platinum on carbon (624 mg, Degussa-type, 50% wet) and ammonium formate (2.5 g) were added and the mixture was heated to 60 ⁰C for 18 hours. The mixture was then cooled to room temperature and filtered through a celite pad, washing the filter pad with ethyl acetate. The solvent was removed in vacuo and the residue dissolved in ethyl acetate/water (50:30 ml_); the organic layer was washed with brine, dried over sodium sulfate and reduced in vacuo to provide the title compound (0.95 g). HPLC Rt = 4.3 minutes; m/z [MH]⁺= 263

Intermediate 7 ethyl 5-{r(3,4-dichlorophenyl)sulfonyl1amino)-6-f(2-methyl-

2H-tetrazol-5-vDmethvP-3-pyridinecarboχylate

Ethyl 5-amino-6-[(2-methyl-2H-tetrazol-5-yl)methyl]-3-pyridinecarboxylate (0.88 g), 4- dimethylaminopyridine (10 mg) and 3,4-dichlorobenzenesulfonyl chloride (415 uL) were heated in pyridine (5 ml_) overnight at 90 ⁰C. An additional portion of 3,4- dichlorobenzesulfonyl chloride (100 uL) was added and the mixture heated for one additional hour. The reaction mixture was concentrated in vacuo and purified by column chromatography; repeated 20 g Si SPE cartridge eluting with ethyl acetate/cyclohexane gave 720 mg of the title compound. HPLC Rt = 3.2 minutes; m/z [MH]⁺= 471

Intermediate 8 5-{r(3,4-dichlorophenvπsulfonvnamino)-6-r(2-methyl-2H- tetrazol-δ-vPmethvπ-S-pyridinecarboxylic acid

A solution of ethyl 5-{[(3,4-dichlorophenyl)sulfonyl]amino}-6-[(2-methyl-2/-/-tetrazol-5- yl)methyl]-3-pyridinecarboxylate (700 mg) in methanol (5 mL) was treated with lithium hydroxide (108 mg) in water (2 mL) and stirred at 55 ⁰C for 2 hours. Solvent was removed in vacuo and the residue dissolved in water (15 mL) and acidified with 2M HCI (3 ml_). The product was extracted into ethyl acetate (20 ml_), washed with brine (15 mL) and dried over sodium sulfate. Removal of the solvent gave the title compound (650 mg). HPLC Rt = 3.1 minutes; m/z [MH]⁺ = 443

Intermediate 9: 5-(r(3,4-dichlorophenyl)sulfonvnamino)-6-f(2-methyl-2H-tetrazol-5- vOmethvπ-3-pyridinecarboxamide

5-{[(3,4-dichlorophenyl)sulfonyl]amino}-6-[(2-methyl-2/-/-tetrazol-5-yl)methyl]-3- pyridinecarboxylic acid (650 mg) was suspended in acetonitrile/DCM (1 :1 , 10 mL) and triethylamine (246 uL) added. Upon dissolution TBtU (566 mg) was added and the mixture aged for 5 minutes. 2M ammonia in methanol (2.94 mL) was added and the mixture was stirred for a further 18 hours. Solvent was removed in vacuo, whereupon the crude product was dissolved in ethyl acetate and washed with saturated sodium bicarbonate solution. The residue obtained upon removing the solvent was purified using reverse phase chromatography with 10 g SPE C-18 cartridge eluting with 0-10% acetonitrile+0.05% formic acid:water+0.1 % formic acid to provide 140 mg of the title compound. HPLC Rt = 2.8 minutes; m/z [MH]⁺ = 442

Example Compounds

Example 1: 3.4-dichloro-Λ/-(5-chloro-2-f(1-methyl-1 H-tetrazol-5-vnmethvn-3- pyridinyllbenzenesulfonamide

A mixture of 5-chloro-2-[(1 -methyl-1 H-tetrazol-5-yl)methyl]-3-pyridinamine (100 mg), 3,4- dichlorobenzenesulfonyl chloride (60 mg) and 4-(dimethylamino)-pyridine (5 mg) in pyridine (2 mL) was stirred overnight at 80 ⁰C under an atmosphere of argon. The mixture was cooled and concentrated. The residue was partitioned between ethyl acetate (20 ml_) and 2M HCI (3 ml_), the organic layer was washed with brine (3 mL) and concentrated. The resulting residue was purified by column chromatography (10-30% ethyl acetate in hexane) and the product slurried in methanol, cooled and filtered to give the title compound, 120 mg. HPLC Rt = 3.4 minutes; m/z [M]^" = 433. ¹H NMR (d₆-DMSO) δ 10.65 (br.s, 1 H), 8.38 (br.d, 1 H), 7.91 (d, 1 H), 7.85 (d, 1 H), 7.66 (dd, 1 H), 7.52 (d, 1 H), 4.36 (s, 2H), 3.85 (s, 3H).

The following compounds of formula (I) set out in Table 1 were prepared by a similar procedure to that described for Example 1 using the appropriate starting materials.

Table 1

Example 8: 3,4-dichloro-Λ/-(5-cvano-2-r(2-methyl-2H-tθtrazol-5-vhnnethvn-3- pyridinvDbenzenesulfonamide

5-{[(3,4-dichlorophenyl)sulfonyl]amino}-6-[(2-methyl-2/-/-tetrazol-5-yl)methyl]-3- pyridinecarboxamide (22 mg), phosphorous oxychloride (47 uL) and chloroform were heated at reflux for 18 hours, cooled and reduced in vacuo. Mass directed autopreparative HPLC gave 2.8 mg of the title compound.

HPLC Rt = 3.3 minutes; m/z [MH]⁺= 424

¹H NMR (CD₃OD) δ 8.47 (d, 1 H), 7.87 (d, 1 H), 7.74 (d, 1 H)₁ 7.69 (d, 1 H), 7.60 (dd, 1 H),

4.55 (s, 2H), 4.02 (s, 3H). Biological Data

Compounds of the present invention have been found to exhibit affinity for cytokine receptors, in particular the CCR2 receptor. Such affinity is typically calculated from the IC₅₀ as the concentration of a compound necessary to displace 50% of the radiolabeled ligand from the receptor in an appropriate assay, and is reported as a "K" value calculated by the following equation: _{κ =} IC₅₀ ¹ 1 + L/ K_D where L = radioligand and K₀ = affinity of radioligand for receptor (Cheng and Prusoff, Biochem. Pharmacol. 22:3099, 1973).

In the context of the present invention pKi (corresponding to the antilogarithm of Ki) is used instead of Ki.

The above examples all have a pKi from 5.8 to 9.0 as derived from such an assay.

CCR2 T³⁵Sl GTPaS SPA binding assay

Membrane preparation

CHO cells expressing the human CCR2 receptor were grown in DMEM F12 media supplemented with 10% foetal calf serum, 2mM L-glutamine, G418 at 37°C 5% CO₂. Confluent cells were harvested using Hanks buffered salt solution (HBSS, Ca²⁺, Mg²⁺ free) containing 0.6mM EDTA. The resulting cell suspension was centrifuged at 30Og at 4⁰C for 10 min, cell pellet resuspended in 100ml HBSS+EDTA and respun at 30Og for 5 min. The resulting cell pellet was resuspended in 5OmM HEPES containing 10OmM leupeptin, 25μg/ml bacitracin, 1 mM EDTA, 1 mM PMSF and 2μM pepstain A, at pH7.4. The suspension was homogenised using an ice cold blender and centrifuged at 50Og for 20mins. The supernatant is withdrawn and spun at 4800Og for 30mins. This cell pellet is resuspended in the above buffer minus the pepstatin A and PMSF and stored in aliquots at -70°C.

Assay

For the assay, membranes are thawed and resuspended in assay buffer (2OmM HEPES, 1OmM MgCI₂, 10OmM NaCI, pH7.4, containing 1 mg/ml saponin, 1 OmM GDP) to give final concentration of 5μg/well. These are pre-coupled with LEADseeker SPA beads (0.25mg/well) for 30 min at room temperature whilst mixing. Assay plates containing 0.5μl of various test compounds (30μM-30pM) in 100% DMSO as 11 point, four fold dilutions across a 384 well plate are used in the assay which have been prepared on a Biomek FX. The plate also contains 16 wells of DMSO and 16 wells of a high concentration of a standard antagonist to produce high and low controls in the experiment. To this 15μl of bead and membrane mix are added with, 15μl [³⁵S] GTPgS (0.2nM final assay concentration) and 15μl of an EC_8O (4OnM) of MCP-1. This concentration has been pre-determined from agonist curves run against this receptor. All additions are made using a multidrop. Plates are then sealed and centrifuged for 5 min at 300rpm before they are left to incubate at room temperature for 3 hours. After this time they are read on a Viewlux imaging system. For data handling the high and low controls wells are used to normalise the data which is then fitted using a 4 parameter kit in Excel.

The assay described above is believed to have an effective limit of detection of a pKi in the region of 5.0-5.5. Accordingly, a compound exhibiting a pKi value within this range from such an assay may indeed have a reasonable affinity for the receptor, but equally it may also have a lower affinity, including a considerably lower affinity.

Claims

Claims

1. A compound as represented in formula (I):

or a salt thereof, or a solvate thereof, or a combination thereof; wherein one of D and E represents a nitrogen atom, and the other of D and E represents CH;

R¹ represents an aryl, a thienyl, a benzothienyl, an imidazolyl, a pyridyl, an isoquinolinyl, , a piperonyl, a benzoxathiadiazolyl, or a benzoxadiazolyl group substituted by up to three R⁴ groups, wherein each R⁴ group is independently selected from the group consisting of d-e-alkyl, C_1-6-haloalkyl, C_1-6-alkoxy, Cm-haloalkoxy, halo, -NH₂, -OH, -CN, -NO₂, CF₃, phenyl, phenoxy, phenyl-C(O)-, isoxazolyl, and -C(O)NR⁷R⁸, wherein R⁷ and R⁸ each independently represent hydrogen or CM alky], or R⁷ and R⁸ together with the nitrogen atom to which they are attached form a 5- or 6-membered saturated heterocylic group optionally containing an additional heteroatom selected from nitrogen, oxygen and sulphur;

m is 1 , 2, or 3;

each R² is independently selected from the group consisting of halo, -CN, and -CF₃-;

R³ is a heteroaryl group optionally substituted by one to three substituents independently selected from the group consisting of halo, hydroxy-C-_M-alkyl-, C_1-4alkoxy-Ci_-4alkyl-, Ci- ₄alkoxy-, R¹⁰R¹¹N-C₁.₄alkyl-, C_1-6-alkyl, C₃.₆-cycloalkyl, -CN, d^alkylthio-, -CF₃, -CO₂H, - CONR⁷R⁸, and -CO-Ci_-4 alkyl, and R¹⁰ and R¹¹ each independently represent hydrogen or C_M alkyl.

2. The compound of Claim 1 wherein D is N and E is CH; m is 1 ; R¹ is chlorophenyl, dichlorophenyl, dichlorothienyl, trifluoromethylphenyl, or cyanophenyl; R² is chloro or cyano; and R³ is tetrazolyl.

3. The compound of Claim 2 wherein R¹ is 4-chlorophenyl, 3,4-dichlorophenyl, 4,5- dichlorothienyl, 3-trifluoromethylphenyl, or 3-cyanophenyl; R² is 5-chloro or 5-cyano; and R³ is 1 -methyl-1 H-tetrazol-5-yl or 2-methyl-2H-tetrazol-5-yl.

4. A compound selected from the group consisting of:

3,4-dichloro-Λ/-{5-chloro-2-[(1-methyl-1 H-tetrazol-5-yl)methyl]-3- pyridinyl}benzenesulfonamide;

3,4-dichloro-Λ/-{5-chloro-2-[(2-methyl-2H-tetrazol-5-yl)methyl]-3- pyridinyl}benzenesulfonamide;

4,5-dichloro-Λ/-{5-chloro-2-[(1 -methyl-1 H-tetrazol-5-yl)methyl]-3-pyridinyl}-2- thiophenesulfonamide;

4,5-dichloro-Λ/-{5-chloro-2-[(2-methyl-2H-tetrazol-5-yl)methyl]-3-pyridinyl}-2- thiophenesulfonamide;

Λ/-{5-chloro-2-[(1-methyl-1 H-tetrazol-5-yl)methyl]-3-pyridinyl}-3- (trifluoromethyl)benzenesulfonamide;

Λ/-{5-chloro-2-[(1 -methyl-1 H-tetrazol-5-yl)methyl]-3-pyridinyl}-3- cyanobenzenesulfonamide;

4-chloro-/V-{5-chloro-2-[(1 -methyl-1 H-tetrazol-5-yl)methyl]-3- pyridinyl}benzenesulfonamide; or

3,4-dichloro-N-{5-cyano-2-[(2-methyl-2H-tetrazol-5-yl)methyl]-3- pyridinyl}benzenesulfonamide ;

or a pharmaceutically acceptable salt thereof, or a solvate thereof, or a combination thereof.

5. A method for treating or preventing a disease or condition mediated by CCR2 comprising administering to a patient in need thereof a pharmaceutically effective amount of the compound of Claim 1 , or a pharmaceutically acceptable salt thereof, or a solvate thereof, of a combination thereof.

6. A pharmaceutical composition comprising a) a compound of formula (I) or a pharmaceutically acceptable salt thereof, or a solvate thereof, or a combination thereof and b) a pharmaceutically acceptable carrier, diluents, or excipient or combination thereof.