HK1164639B - C5ar antagonists - Google Patents

Description

C5aR antagonists

Cross Reference to Related Applications

This application claims the benefit of U.S. Pat. No. 61/139,919 filed on 22/12/2008; this application is incorporated by reference herein in its entirety.

Claims for inventions made with the assistance of federal research and development funds

Not applicable.

Reference to "sequence Listing" (attachment of computer program Table to form, or committed compact disk)

Not applicable.

Background

The complement system plays an important role in immune complex clearance and immune responses to infectious agents, foreign antigens, virus-infected cells and tumor cells. Inappropriate or excessive activation of the complement system can lead to deleterious and possibly even life-threatening consequences due to severe inflammation and to tissue destruction. These consequences are manifested clinically in a variety of conditions, including septic shock; myocardial and intestinal ischemia/reperfusion injury; transplant rejection; organ failure; nephritis; pathological inflammation; and autoimmune diseases.

The complement system consists of a group of proteins that are normally present in the serum in an inactive state. Activation of the complement system mainly involves three different pathways, namely the classical pathway, the alternative pathway and the lectin pathway (V.M. Holers, In Clinical Immunology: Principles and Practice, R.R. Rich code, Mosby Press; 1996, 363-391): 1) the classical pathway is a calcium/magnesium dependent cascade, which is usually activated by the formation of an antigen-antibody complex. It can also be activated in an antibody-independent manner by binding to C-reactive proteins complexed with ligands and by a variety of pathogens, including gram-negative bacteria. 2) An alternative pathway is the magnesium-dependent cascade, which is activated by deposition and activation of C3 on specific susceptible surfaces (e.g. muramyl polysaccharides of yeast and bacteria, and certain biopolymer species). 3) The lectin pathway involves first binding to mannose binding to lectins and subsequent activation of C2 and C4, which are common to the classical pathway (Matsushita, m. et al, j.exp.med.176: 1497-; suankratay, c, et al, j.immunol.160: 3006-3013(1998)).

Activation of the complement pathway produces biologically active fragments of complement proteins, such as C3a, C4a, and C5a anaphylatoxins, and C5b-9 Membrane Attack Complex (MAC), all by affecting leukocyte chemotaxis; activating macrophages, neutrophils, platelets, mast cells and endothelial cells; and to enhance vascular permeability, cytolysis, and tissue damage to mediate inflammatory responses.

Complement C5a is one of the most potent pro-inflammatory mediators of the complement system. (in terms of eliciting an inflammatory response, the allergenic C5a peptide is 100-fold more potent than C3a on a molar basis.) C5a is an activated form of C5 (190kD molecular weight). C5a was present at about 80. mu.g/ml in human serum (Kohler, P.F., et al, J.Immunol.99: 1211-1216 (1967)). It consists of two polypeptide chains, alpha and beta, which have respective molecular weights of about 115kD and 75kD (Tack, B.F. et al, Biochemistry 18: 1490-. Biosynthetic C5, a single-chain pre-molecule (promulule), is enzymatically cleaved into a double-stranded structure during processing and secretion. After cleavage, the two chains are held together by at least one disulfide bond and non-covalent interactions (Ooi, Y.M. et al, J.Immunol.124: 2494-2498 (1980)).

C5 is cleaved into C5a and C5b fragments during complement pathway activation. The convertases responsible for the activation of C5 are the C4b, C2a and C3b multi-subunit complexes of the classical pathway, and the (C3b)2, Bb and P multi-subunit complexes of the alternative pathway (Goldlust, M.B. et al, J.Immunol.113: 998-3970 (1974); Schreiber, R.D. et al, Proc. Natl.Acad.Sci.75: 3948-3952 (1978)). C5 is activated by cleavage at positions 74-75(Arg-Leu) in the alpha chain. Upon activation, the 74 amino acid peptide C5a, 11.2kD, is released from the amino-terminal portion of the alpha chain. C5a and C3a are potent activators of neutrophils and monocytes (Schinder, R. et al, Blood 76: 1631-.

In addition to their allergictoxic properties, C5a also induced chemotactic migration of neutrophils (Ward, P.A., et al, J.Immunol.102: 93-99(1969)), eosinophils (Kay, A.B., et al, Immunol.24: 969-976(1973)), basophils (Lett-Brown, M.A., et al, J.Immunol.117: 246-252(1976)), and monocytes (Snyderman, R.et al, Proc.Soc.exp.biol.Med.138: 387-390 (1971)). Both C5a and C5b-9 activate endothelial cells to express adhesion molecules necessary for the chelation of activated leukocytes, which mediate tissue Inflammation and injury (Foreman, K.E. et al, J.Clin.invest.94: 1147-containing 1155 (1994); Foreman, K.E. et al, Inflammation 20: 1-9 (1996); Rollins, S.A. et al, Transplantation 69: 1959-containing 1967 (2000)). C5a also mediates inflammatory responses by causing smooth muscle contraction, increasing vascular permeability, inducing degranulation of basophils and mast cells, and inducing lysosomal proteases and oxidative free radical release (Gerard, C. et al, Ann. Rev. Immunol.12: 775-808 (1994)). In addition, C5a regulates hepatic acute phase gene expression and potentiates The overall immune response by increasing The production of TNF- α, IL-1- β, IL-6, IL-8, prostaglandins, and leukotrienes (Lambris, J.D., et al, The Human comparative System in health and Disease, Volanakis, J.E., eds., Marcel Dekker, New York, pp 83-118).

It is believed that the allergic and chemotactic effects of C5a are mediated through their interaction with the C5a receptor. The human C5a receptor (C5aR) is a 52kD membrane-bound G protein-coupled receptor and is expressed on neutrophils, monocytes, basophils, eosinophils, hepatocytes, lung smooth muscle and endothelial cells, as well as glomerular tissue (Van-Epps, D.E. et al, J.Immunol.132: 2862-. The ligand binding site of C5aR is a complex and consists of at least two physically separable binding domains. One domain binds to the amino terminus of C5a (amino acids 1-20) and the disulfide core (amino acids 21-61), while the second domain binds to the carboxy terminus of C5a (amino acids 62-74) (Wetsel, R.A., curr. Opin. Immunol.7: 48-53 (1995)).

C5a plays an important role in inflammation and tissue injury. In cardiopulmonary bypass and hemodialysis, C5a forms as a result of activation of the alternative complement pathway when human blood contacts artificial surfaces of the heart-lung or kidney dialysis machines (Howard, R.J. et al, Arch.Surg.123: 1496-. C5a causes capillary permeability and edema, bronchoconstriction, pulmonary vasoconstriction, leukocyte and platelet activation and provision or increase of infiltration into tissues, especially the lungs (Czermak, b.j. et al, j.leukc.biol.64: 40-48 (1998)). Administration of an anti-C5 a monoclonal antibody was shown to reduce cardiopulmonary bypass and coronary endothelial cell dysfunction induced by cardioplegia (Tofukuji, M. et al, J.Thorac.Cardiovasc.Surg.116: 1060-1068 (1998)).

C5a is also implicated in Acute Respiratory Distress Syndrome (ARDS), Chronic Obstructive Pulmonary Disease (COPD) and Multiple Organ Failure (MOF) (Hack, C.E. et al, am.J.Med.1989: 86: 20-26; Hammerschmidt DE et al, Lancet 1980; 1: 947-. C5a potentiates the monocyte production of two important proinflammatory cytokines TNF-. alpha.and IL-1. C5a has also been shown to play an important role in the development of tissue damage, especially lung damage, in animal models of septic shock (Smedegard G et al, am. J. Pathol. 1989; 135: 489-. In a sepsis model using rats, pigs and non-human primates, administration of anti-C5 a antibody to animals prior to treatment with endotoxin or E.coli (E.coli) resulted in reduced tissue damage and reduced IL-6 production (Smedegard, G. et al, am. J. Pathol.135: 489-497 (1989); Hopken, U. et al, Eur. J. Immunol.26: 1103-1109 (1996); Stevens, J.H. et al, J.Clin. invest.77: 1812-1816 (1986)). More importantly, blocking C5a with polyclonal anti-C5 a antibody has been shown to significantly improve the survival rate in the caecal ligation/puncture model of sepsis in rats (Czermak, B.J., et al, nat. Med.5: 788-792 (1999)). This model shares many aspects with human clinical sepsis presentation (Parker, S.J., et al, Br.J.Surg.88: 22-30 (2001)). In this same sepsis model, anti-C5 a antibody was shown to inhibit thymocyte apoptosis (Guo, R.F. et al, J.Clin. invest.106: 1271-1190 (2000)) and prevent MOF (Huber-Lang, M. et al, J.Immunol.166: 1193-1199 (2001)). The anti-C5 a antibody was also protective in the cobra venom factor model of lung injury in rats and in immune complex-induced lung injury (Mulligan, M.S. et al, J.Clin. invest.98: 503-512 (1996)). The importance of C5a in immune complex-mediated lung injury was later demonstrated in mice (Bozic, C.R. et al, Science 26: 1103-1109 (1996)).

C5a was found to be the major mediator in myocardial ischemia-reperfusion injury. Complement depletion reduces myocardial infarct size in mice (Weisman, H.F. et al, Science 249: 146-151(1990)), and treatment with anti-C5 a antibody reduces injury in a rat model of hindlimb ischemia-reperfusion (Bless, N.M. et al, am.J. physiol.276: L57-L63 (1999)). Reperfusion injury during myocardial infarction was also significantly reduced in pigs retreated with monoclonal anti-C5 a IgG antibody (Amsterdam, E.A., et al, am.J.Physiol.268: H448-H457 (1995)). Recombinant human C5aR antagonists reduce infarct size in porcine models of revascularization surgery (Riley, R.D. et al, J.Thorac.Cardiovasc.Surg.120: 350-358 (2000)).

C5 a-driven neutrophils also contribute to a variety of bullous diseases (e.g., bullous pemphigoid, pemphigus vulgaris, and pemphigus foliaceus). These diseases are chronic, recurrent inflammatory conditions that are clinically characterized by the presence of sterile blisters in the sub-epidermal space of the skin and mucosa. While autoantibodies directed against keratinocytes located on the basal membrane of the skin are believed to form the basis of detachment of epidermal basal keratinocytes from the underlying basal membrane, blisters are also characterized by accumulation of neutrophils in the upper skin layers and within the vacuole cavity. In experimental models, the reduction of neutrophils or the absence of complement (total complement or C5 selective complement) can inhibit the formation of subcutaneous blisters even in the presence of high titer autoantibodies.

Complement levels are elevated in patients with rheumatoid arthritis (Jose, P.J. et al, Ann. Rheum. Dis.49: 747-752 (1990); Grant, E.P. et al, J.of exp. Med., 196 (11): 1461-1471, (2002)), lupus nephritis (Bao, L. et al, Eur.J. of Immunol, 35(8), 2496-2506, (2005)) and systemic lupus erythematosus (systemic lupus erythematous, SLE) (Porcel, J.M. et al, Clin. C5a levels correlate with the severity of the disease condition. Collagen-induced arthritis in mice and rats is similar to rheumatoid arthritis disease in humans. Mice deficient in the C5a receptor showed complete protection against arthritis induced by the injection of monoclonal anti-collagen Abs (Banda, N.K., et al, J.of Immunol., 2003, 171: 2109-2115). Therefore, inhibition of the C5a and/or C5a receptor (C5aR) may be useful in the treatment of these chronic diseases.

The complement system is believed to be activated in patients with Inflammatory Bowel Disease (IBD) and is thought to play a role in disease pathogenesis. Activated complement products are found in IBD patients on the luminal surface of epithelial cells, as well as in the muscularis mucosae and in submucosal vessels (Woodruff, T.M., et al, J of Immunol., 2003, 171: 5514-.

In the human central nervous system with inflammation, C5aR expression is upregulated on reactive astrocytes, microglia and endothelial cells (Gasque, P. et al, am. J. Pathol. 150: 31-41 (1997)). C5a may be involved in neurodegenerative diseases such as Alzheimer's disease (Mukherjee, P. et al, J. Neurommunol.105: 124-. Activation of neuronal C5aR induces apoptosis (Farkas I et al, J. Physiol.1998; 507: 679-687). Thus, inhibition of C5a and/or C5aR may also be useful for treating neurodegenerative diseases.

There is some evidence that the production of C5a exacerbates inflammation associated with atopic dermatitis (Neuber, K., et al, Immunology 73: 83-87, (1991)) and chronic urticaria (Kaplan, A.P., J.allergy Clin. Immunol.114; 465- & 474, (2004)).

Psoriasis is now known to be a T cell mediated disease (Gottlieb, E.L. et al, nat. Med.1: 442-447 (1995)). However, neutrophils and mast cells may also be involved in the pathogenesis of the disease (Terui, T. et al, exp. Dermatol.9: 1-10 (2000); Werfel, T. et al, Arch. Dermatol.Res.289: 83-86 (1997)). Accumulation of neutrophils under the stratum corneum was observed in highly inflamed areas of psoriatic plaques and psoriatic lesion (scales) extracts contained elevated levels of C5a and showed strong chemotactic activity for neutrophils, an effect that could be inhibited by the addition of C5a antibody. T cells and neutrophil lines were chemoattracted by C5a (Nataf, S. et al, J.Immunol.162: 4018-4023 (1999); Tsuji, R.F. et al, J.Immunol.165: 1588-1598 (2000); Cavaillon, J.M. et al, Eur.J.Immunol.20: 253-257 (1990)). In addition, expression of C5aR has been demonstrated in plasmacytoid dendritic cells (pdcs) isolated from the foci of cutaneous lupus erythematosus, and these cells were shown to exhibit chemotactic behavior for C5a, suggesting that blockade of C5aR on pdcs may be effective in reducing infiltration of pdcs into inflammatory skin in SLE and psoriasis. Thus, C5a may be an important target for the treatment of psoriasis.

Immunoglobulin G-containing Immune Complexes (IC) play a role in the pathophysiology of a variety of autoimmune diseases, such as systemic lupus erythematosus, rheumatoid arthritis, Sjogren's disease, Goodpasture's syndrome, and hypersensitivity pneumonitis (Madaio, M.P., Semin. Nephrol.19: 48-56 (1999); Korganow, A.S. et al, Immunity 10: 451-459 (1999); Bolten, W.K., Kidney Int.50: 1754-1760 (1996); Ando, M.et al, Current. Opin. palm. Med.3: 391-399 (399)). These diseases are highly heterogeneous and often affect one or more of the following organs: skin, blood vessels, joints, kidneys, heart, lungs, nervous system and liver (including cirrhosis and liver fibrosis). The classical animal model for the inflammatory response in these IC diseases is the Arthus reaction (Arthus reaction), which is characterized by polymorphonuclear cell infiltration, hemorrhage and plasma exudation (Arthus, M., C.R.Soc.biol.55: 817-824 (1903)). Recent studies have shown that C5aR deficient mice are protected from IC-induced tissue damage (Kohl, J. et al, mol. Immunol.36: 893-903 (1999); Baumann, U. et al, J. Immunol.164: 1065-1070 (2000)). The results are consistent with the observation that small peptide anti-C5 aR antagonists inhibit inflammatory responses caused by IC deposition (Strachan, A.J., et al, J.Immunol.164: 6560-. C5a, along with its receptor, plays an important role in the pathogenesis of IC disease. Inhibitors of C5a and C5aR may be useful in the treatment of these diseases.

Description of the Related Art：

Only recently have there been descriptions in the literature of non-peptide based antagonists of the C5a receptor (e.g. Sumichika, h. et al, j. biol. chem. (2002), 277, 49403-49407). Non-peptide-based C5a receptor antagonists have been reported to be effective in treating endotoxic shock in rats (Stracham, A.J., et al, J.of Immunol (2000), 164 (12): 6560-; and treatment of IBD in a rat model (Woodruff, T.M., et al, J of Immunol., 2003, 171: 5514-. Also in the patent literature by neurogenin corporation (e.g., WO2004/043925, WO2004/018460, WO2005/007087, WO03/082826, WO03/08828, WO02/49993, WO 03/084524); dompe S.P.A. (WO 02/029187); and The University of Queenland (WO2004/100975) describe non-peptide based modulators of The C5a receptor.

There is considerable experimental evidence in the literature suggesting that C5a levels are increased in a number of diseases and disorders, particularly autoimmune and inflammatory diseases and disorders. Thus, there remains a need in the art for novel small organic molecule modulators, e.g., agonists, preferably antagonists, partial agonists, of the C5a receptor (C5aR) that can be used to inhibit pathogenic events associated with elevated levels of anaphylatoxin activity, e.g., chemotaxis. The present invention fulfills this need and others.

Summary of the invention

In one aspect, the invention provides a compound having the formula:

and pharmaceutically acceptable salts, hydrates, and rotamers thereof; wherein

C¹Selected from aryl and heteroaryl, wherein said heteroaryl has 1-3 heteroatoms selected from N, O and S as ring members; and wherein said aryl and heteroaryl are optionally substituted with 1 to 3R¹Substituent group substitution;

C²selected from aryl and heteroaryl, wherein said heteroaryl has 1-3 heteroatoms selected from N, O and S as ring members; and wherein said aryl and heteroaryl are optionally substituted with 1 to 3R²Substituent group substitution;

C³is selected from C_1-8Alkyl radical, C_3-8Cycloalkyl radical, C_3-8cycloalkyl-C_1-4Alkyl, aryl-C_1-4Alkyl, heteroaryl-C_1-4Alkyl, heterocycloalkyl or heterocycloalkyl-C_1-4Alkyl, wherein the heterocycloalkyl or moiety has 1-3 heteroatoms selected from N, O and S, and wherein the heteroaryl has 1-3 heteroatoms selected from N, O and S as ring members, and each C³Optionally substituted by 1-3R³Substituent group substitution;

each R¹Independently selected from: halogen, -CN, -R^c、-CO₂R^a、-CONR^aR^b、-C(O)R^a、-OC(O)NR^aR^b、-NR^bC(O)R^a、-NR^bC(O)₂R^c、-NR^a-C(O)NR^aR^b、-NR^aC(O)NR^aR^b、-NR^aR^b、-OR^aand-S (O)₂NR^aR^b(ii) a Wherein R is^aAnd R^bEach independently selected from hydrogen and C_1-8Alkyl and C_1-8Haloalkyl, or when attached to the same nitrogen atom, can combine with that nitrogen atom to form a five-or six-membered ring having from 0 to 2 additional heteroatoms selected from N, O or S as ring members; each R^cIndependently selected from C_1-8Alkyl radical, C_1-8Haloalkyl, C_3-6Cycloalkyl, heterocycloalkyl, aryl and heteroaryl, and wherein R^a、R^bAnd R^cOptionally further substituted with one to three halogens, hydroxy, methyl, amino, alkylamino and dialkylamino; and optionally, when two R are¹When the substituents are on adjacent atoms, they combine to form a fused five-or six-membered carbocyclic ring;

each R²Independently selected from: halogen, -CN, -R^f、-CO₂R^d、-CONR^dR^e、-C(O)R^d、-OC(O)NR^dR^e、-NR^eC(O)R^d、-NR^eC(O)₂R^f、-NR^dC(O)NR^dR^e、-NR^dC(O)NR^dR^e、-NR^dR^e、-OR^dand-S (O)₂NR^dR^e(ii) a Wherein R is^dAnd R^eEach independently selected from hydrogen and C_1-8Alkyl and C_1-8Haloalkyl, or when attached to the same nitrogen atom, can combine with that nitrogen atom to form a five-or six-membered ring having from 0 to 2 additional heteroatoms selected from N, O or S as ring members; each R^fIndependently selected from C_1-8Alkyl radical, C_1-8Haloalkyl, C_3-6Cycloalkyl, heterocycloalkyl, aryl and heteroaryl, and wherein R^d、R^eAnd R^fOptionally further substituted with one to three halogens, hydroxy, methyl, amino, alkylamino and dialkylamino;

each R³Independently selected from: halogen, -CN, -Rⁱ、-CO₂R^g、-CONR^gR^h、-C(O)R^g、-OC(O)NR^gR^h、-NR^hC(O)R^g、-NR^hC(O)₂Rⁱ、-NR^gC(O)NR^gR^h、-NR^gR^h、-OR^g、-S(O)₂NR^gR^h、-X⁴-R^j、-X⁴-NR^gR^h、-X⁴-CONR^gR^h、-X⁴-NR^hC(O)R^g、-NHR^jand-NHCH₂R^jWherein X is⁴Is C_1-4An alkylene group; r^gAnd R^hEach independently selected from hydrogen and C_1-8Alkyl radical, C_3-6Cycloalkyl and C_1-8Haloalkyl, or when attached to the same nitrogen atom, may combine with that nitrogen atom to form a five-or six-membered ring having 0 to 2 additional heteroatoms selected from N, O or S as ring members, and optionally substituted with one or two oxo groups (oxo); each RⁱIndependently selected from C_1-8Alkyl radical, C_1-8Haloalkyl, C_3-6Cycloalkyl, heterocycloalkyl, aryl, and heteroaryl; and each R^jIs selected from C_3-6Cycloalkyl, pyrrolinyl, piperidinyl, morpholinyl, tetrahydrofuryl and tetrahydropyranyl, and wherein R is^g、R^h、RⁱAnd R^jOptionally further substituted with one to three halogens, methyl, CF₃Hydroxy, amino, alkylamino and dialkylamino; and is

X is hydrogen or CH₃。

In addition to the compounds provided herein, the invention further provides pharmaceutical compositions containing one or more of these compounds, as well as methods of using these compounds in therapeutic methods, primarily for the treatment of diseases associated with C5a signaling activity.

In yet another aspect, the invention provides a method of diagnosing a disease in an individual. In these methods, a compound provided herein is administered to an individual in labeled form, followed by diagnostic imaging to determine the presence or absence of C5aR 7. In a related aspect, a method of diagnosing a disease is performed by contacting a tissue or blood sample with a labeled compound as provided herein and determining the presence, absence, or amount of C5aR in the sample.

Drawings

Figure 1 provides the structure and activity of representative compounds of the invention. The compounds are generally prepared by the methods as generally described below and as provided in the examples.

Detailed Description

I. Abbreviations and Definitions

Unless otherwise specified, the term "alkyl", alone or as part of another substituent, means a straight or branched chain hydrocarbon radical (i.e., C) having the indicated number of carbon atoms_1-8Meaning one to eight carbons). Examples of alkyl groups include methyl, ethyl, n-propyl, isopropyl, n-butyl, tert-butyl, isobutyl, sec-butyl, n-pentyl, n-hexyl, n-heptyl, n-octyl, and the like. The term "alkenyl" refers to an unsaturated alkyl group having one or more double bonds. Similarly, the term "alkynyl" refers to an unsaturated alkyl group having one or more triple bonds. Examples of such unsaturated alkyl groups include ethenyl, 2-propenyl, crotyl, 2-isopentenyl, 2- (butadienyl), 2, 4-pentadienyl, 3- (1, 4-pentadienyl), ethynyl, 1-and 3-propynyl, 3-butynyl, and higher homologs and isomers. The term "cycloalkyl" refers to a ring having the indicated number of ring atoms (e.g., C)_3-6Cycloalkyl) and is fully saturated or has not more than one ring apexA hydrocarbon ring of double bonds. "cycloalkyl" also means bicyclic and polycyclic hydrocarbon rings, e.g. bicyclo [2.2.1]Heptane, bicyclo [2.2.2]Octane, and the like. The term "heterocycloalkyl" refers to a cycloalkyl group containing one to five heteroatoms selected from N, O and S, wherein the nitrogen and sulfur atoms are optionally oxidized, and the nitrogen atom is optionally quaternized. The heterocycloalkyl group can be a monocyclic, bicyclic, or polycyclic ring system. Non-limiting examples of heterocycloalkyl groups include pyrrolidine, imidazolidine, pyrazolidine, butyrolactam, valerolactam, imidazolidinone, hydantoin, dioxolane, phthalimide, piperidine, 1, 4-dioxane, morpholine, thiomorpholine-S-oxide, thiomorpholine-S, S-oxide, piperazine, pyran, pyridone, 3-pyrroline, thiopyran, pyrone, tetrahydrofuran, tetrahydrothiophene, quinuclidine, and the like. The heterocycloalkyl group can be attached to the rest of the molecule through a ring carbon or a heteroatom.

The term "alkylene" alone or as part of another substituent means a divalent radical derived from an alkane, an example being-CH₂CH₂CH₂CH₂-. Generally, alkyl (or alkylene) groups have 1 to 24 carbon atoms, and those having 10 or less carbon atoms are preferred in the present invention. "lower alkyl" or "lower alkylene" is a shorter chain alkyl or alkylene group typically having four or fewer carbon atoms. Similarly, "alkenylene" and "alkynylene" refer to unsaturated forms of "alkylene" having double or triple bonds, respectively.

Unless otherwise specified, the term "heteroalkyl," alone or in combination with another term, means a stable straight or branched chain or cyclic hydrocarbon radical consisting of the stated number of carbon atoms and one to three heteroatoms selected from O, N, Si and S, or a combination thereof, and wherein the nitrogen and sulfur atoms may optionally be oxidized and the nitrogen heteroatom may optionally be quaternized. The heteroatoms O, N and S can be located anywhere within the heteroalkyl group. The heteroatom Si may be located anywhere in the heteroalkyl group, including the position where the alkyl group is attached to the remainder of the molecule. Examples include-CH₂-CH₂-O-CH₃、-CH₂-CH₂-NH-CH₃、-CH₂-CH₂-N(CH₃)-CH₃、-CH₂-S-CH₂-CH₃、-CH₂-CH₂、-S(O)-CH₃、-CH₂-CH₂-S(O)₂-CH₃、-CH＝CH-O-CH₃、-Si(CH₃)₃、-CH₂-CH＝N-OCH₃and-CH ═ CH-N (CH)₃)-CH₃. Up to two hetero atoms may be attached, e.g. -CH₂-NH-OCH₃and-CH₂-O-Si(CH₃)₃. Similarly, unless otherwise specified, the terms "heteroalkenyl" and "heteroalkynyl" alone or in combination with another term, respectively, mean an alkenyl or alkynyl group containing the recited number of carbons and having one to three heteroatoms selected from O, N, Si and S, and wherein the nitrogen and sulfur atoms may optionally be oxidized and the nitrogen heteroatom may optionally be quaternized. The heteroatoms O, N and S can be located anywhere within the heteroalkyl group.

The term "heteroalkylene" alone or as part of another substituent means a saturated or unsaturated or polyunsaturated divalent radical derived from a heteroalkyl radical, an example being-CH₂-CH₂-S-CH₂CH₂-and-CH₂-S-CH₂-CH₂-NH-CH₂-、-O-CH₂-CH＝CH-、-CH₂-CH＝C(H)CH₂-O-CH₂-and-S-CH₂-C ≡ C-. For heteroalkylene groups, heteroatoms can also occupy one or both of the chain termini (e.g., alkyleneoxy, alkylenedioxy, alkyleneamino, alkylenediamino, and the like).

The terms "alkoxy", "alkylamino" and "alkylthio" (or thioalkoxy) are used in their conventional sense and refer to an alkyl group attached to the remainder of the molecule through an oxygen atom, an amino group, or a sulfur atom, respectively. In addition, for dialkylamino groups, the alkyl moieties can be the same or different and can also be combined with the nitrogen atom to which each is attached to form a 3-7 membered ring. Thus, it is represented as-NR^aR^bThe groups of (a) are intended to include piperidinyl, pyrrolidinyl, morpholinyl, azetidinyl, and the like.

Unless otherwise specified, the terms "halo" or "halogen", alone or as part of another substituent, mean a fluorine, chlorine, bromine or iodine atom. Additionally, terms such as "haloalkyl" are intended to include monohaloalkyl as well as polyhaloalkyl. For example, the term "C_1-4Haloalkyl "is intended to include trifluoromethyl, 2, 2, 2-trifluoroethyl, 4-chlorobutyl, 3-bromopropyl, and the like.

Unless otherwise specified, the term "aryl" means a polyunsaturated, usually aromatic, hydrocarbon group that can be a single ring, or multiple rings (up to three rings) that are fused together or covalently bonded. The term "heteroaryl" refers to an aryl (or aromatic ring) containing one to five heteroatoms selected from N, O and S, wherein the nitrogen and sulfur atoms are optionally oxidized, and the nitrogen atoms are optionally quaternized. The heteroaryl group may be attached to the rest of the molecule through a heteroatom. Non-limiting examples of aryl groups include phenyl, naphthyl, and biphenyl groups, while non-limiting examples of heteroaryl groups include pyridyl, pyridazinyl, pyrazinyl, pyrimidinyl, triazinyl, quinolinyl, quinoxalinyl, quinazolinyl, cinnolinyl, phthalazinyl, benzotriazinyl, purinyl, benzimidazolyl, benzopyrazolyl, benzotriazolyl, benzisoxazolyl, and the likeOxazolyl, isobenzofuranyl, isoindolyl, indolizinyl, benzotriazinyl, thienopyridinyl, thienopyrimidinyl, pyrazolopyrimidinyl, imidazopyridinyl, benzothiazolyl, benzofuranyl, benzothienyl, indolyl, quinolinyl, isoquinolinyl, isothiazolyl, pyrazolyl, indazolyl, pteridinyl, imidazolyl, triazolyl, tetrazolyl, and the like,Azolyl radical, isoOxazolyl, thiadiazolyl, pyrrolyl, thiazolyl, furyl, thienyl, and the like. Each of the above aryl and heteroarylThe substituents of the ring system are selected from the following acceptable substituents.

For convenience, the term "aryl" when used in combination with other terms (e.g., aryloxy, arylsulfenoxy, arylalkyl) includes both aromatic and heteroaromatic rings as defined above. Thus, the term "arylalkyl" is intended to include groups in which the aryl group is attached to an alkyl group (e.g., benzyl, phenethyl, pyridylmethyl, and the like).

In some embodiments, the above terms (e.g., "alkyl," "aryl," and "heteroaryl") are intended to include both substituted and unsubstituted forms of the indicated group. Preferred substituents for each type of group are provided below. For simplicity, the terms aryl and heteroaryl will refer to the substituted or unsubstituted forms as provided below, while the term "alkyl" and related aliphatic groups are intended to refer to the unsubstituted forms unless indicated to be substituted.

Substituents for alkyl groups (including those groups commonly referred to as alkylene, alkenyl, alkynyl, and cycloalkyl) can be a variety of groups selected from: -halogen, -OR ', -NR ' R ', -SR ', -SiR ' R ' R ', -OC (O) R ', -C (O) R ', -CO₂R′、-CONR′R″、-OC(O)NR′R″、-NR″C(O)R′、-NR′-C(O)NR″R″′、-NR″C(O)₂R′、-NH-C(NH₂)＝NH、-NR′C(NH₂)＝NH、-NH-C(NH₂)＝NR′、-S(O)R′、-S(O)₂R′、-S(O)₂NR′R″、-NR′S(O)₂R', -CN and-NO₂The number is in the range of zero to (2m '+ 1), where m' is the total number of carbon atoms in these groups. R ', R ' and R ' each independently mean hydrogen, unsubstituted C_1-8Alkyl, unsubstituted heteroalkyl, unsubstituted aryl, aryl substituted with 1-3 halogens, unsubstituted C_1-8Alkyl radical, C_1-8Alkoxy or C_1-8Thioalkoxy, or unsubstituted aryl-C_1-4An alkyl group. When R 'and R' are attached to the same nitrogen atom, they may combine with the nitrogen atom to form a 3-, 4-, 5-, 6-or 7-membered ring. For example, -NR' R "is intended to include 1-pyrrolidinyl and 4-morpholinyl. The term "acyl" monoBy itself or when used as part of another group is meant an alkyl group: wherein the two substituents on the carbon closest to the point of attachment of the group are replaced by a substituent ═ O (e.g., -c (O) CH₃、-C(O)CH₂CH₂OR', etc.).

Similarly, substituents for aryl and heteroaryl are numerous and are typically selected from: -halogen, -OR ', -OC (O) R', -NR 'R', -SR ', -R', -CN, -NO₂、-CO₂R′、-CONR′R″、-C(O)R′、-OC(O)NR′R″、-NR″C(O)R′、-NR″C(O)₂R′、-NR′-C(O)NR″R″′、-NH-C(NH₂)＝NH、-NR′C(NH₂)＝NH、-NH-C(NH₂)＝NR′、-S(O)R′、-S(O)₂R′、-S(O)₂NR′R″、-NR′S(O)₂R″、-N₃Perfluoro (C)₁-C₄) Alkoxy and perfluoro (C)₁-C₄) Alkyl groups in a number ranging from zero to the sum of the open valences on the aromatic ring system; wherein R ', R ' and R ' are independently selected from hydrogen, C_1-8Alkyl radical, C_3-6Cycloalkyl radical, C_2-8Alkenyl radical, C_2-8Alkynyl, unsubstituted aryl and heteroaryl, (unsubstituted aryl) -C_1-4Alkyl, and unsubstituted aryloxy-C_1-4An alkyl group. Other suitable substituents include each of the aryl substituents described above attached to a ring atom through an alkylene chain having from 1 to 4 carbon atoms.

Two substituents on adjacent atoms of the aromatic or heteroaromatic ring may optionally be replaced by a substituent of the formula-T-C (O) - (CH)₂)_q-substitution of substituents for U-, wherein T and U are independently-NH-, -O-, -CH₂-or a single bond, and q is an integer from 0 to 2. Alternatively, two substituents on adjacent atoms of the aromatic or heteroaromatic ring may optionally be replaced by a group of formula-A- (CH)₂)_rA substituent replacement of-B-, wherein A and B are independently-CH₂-、-O-、-NH-、-S-、-S(O)-、-S(O)₂-、-S(O)₂NR' -or a single bond, and r is an integer of 1 to 3. One of the single bonds of the new ring so formed may optionally be replaced by a double bond. Alternatively, two substituents on adjacent atoms of the aromatic or heteroaromatic ring may optionally be substituted by a group of formula- (CH)₂)_s-X-(CH₂)_t-wherein S and t are independently integers from 0 to 3, and X is-O-, -NR' -, -S (O)₂-or-S (O)₂NR' -. -NR' -and-S (O)₂The substituents R 'in NR' are selected from hydrogen or unsubstituted C_1-6An alkyl group.

The term "heteroatom" as used herein is intended to include oxygen (O), nitrogen (N), sulfur (S), and silicon (Si).

The term "ionic liquid" refers to any liquid that contains primarily ions. Preferably, in the present invention, "ionic liquid" refers to a salt having a relatively low melting point (e.g., less than 250 ℃). Examples of ionic liquids include, but are not limited to, 1-butyl-3-methylimidazolium tetrafluoroborate, 1-hexyl-3-methylimidazolium tetrafluoroborate, 1-octyl-3-methylimidazolium tetrafluoroborate, 1-nonyl-3-methylimidazolium tetrafluoroborate, 1-decyl 3-methylimidazolium tetrafluoroborate, 1-hexyl-3-methylimidazolium hexafluorophosphate, 1-hexyl-3-methylimidazolium bromide, and the like.

The term "pharmaceutically acceptable salt" is intended to include salts of the active compounds prepared with relatively nontoxic acids or bases, depending on the particular substituents present on the compounds described herein. When the compounds of the present invention contain relatively acidic functional groups, base addition salts can be obtained by contacting the neutral forms of these compounds with a sufficient amount of the desired base, either neat or in a suitable inert solvent. Examples of salts derived from pharmaceutically acceptable inorganic bases include aluminum, ammonium, calcium, copper, ferric, ferrous, lithium, magnesium, manganic, manganous, potassium, sodium, zinc, and the like. Salts derived from pharmaceutically acceptable organic bases include salts of primary, secondary, and tertiary amines, including substituted amines, cyclic amines, naturally occurring amines, and the like, such as arginine, betaine, caffeine, choline, N' -dibenzylethylenediamine, diethylamine, 2-diethylaminoethanol, 2-dimethylaminoethanol, ethanolamine, ethylenediamine, N-ethylmorpholine, N-ethylpiperidine, glucosamine (glucamine), glucosamine, histidine, rapamycin V (hydrabamine), isopropylamine, lysine, methylglucamine, morpholine, piperazine, piperidine, polyamine resins, procaine, purines, theobromine, triethylamine, trimethylamine, tripropylamine, tromethamine, and the like. When the compounds of the present invention contain relatively basic functional groups, acid addition salts can be obtained by contacting the neutral forms of these compounds with a sufficient amount of the desired acid, either neat or in a suitable inert solvent. Examples of pharmaceutically acceptable acid addition salts include those derived from inorganic acids such as hydrochloric, hydrobromic, nitric, carbonic, monohydrogencarbonic acid, phosphoric, monohydrogenphosphoric acid, dihydrogenphosphoric acid, sulfuric, monohydrogensulfuric acid, hydroiodic or phosphorous acid, and the like, as well as those derived from relatively nontoxic organic acids such as acetic, propionic, isobutyric, malonic, benzoic, succinic, suberic, fumaric, mandelic, phthalic, benzenesulfonic, p-toluenesulfonic, citric, tartaric, methanesulfonic, and the like. Also included are Salts of amino acids (e.g., arginine, etc.), and Salts of organic acids (e.g., glucuronic acid or galacturonic acid), etc. (see, e.g., Berge, s.m., et al, "Pharmaceutical Salts", Journal of Pharmaceutical science, 1977, 66, 1-19). Certain specific compounds of the invention contain both basic and acidic functional groups that allow the compounds to be converted into base addition salts or acid addition salts.

The neutral form of the compound may be regenerated by contacting the salt with a base or acid and isolating the parent compound in a conventional manner. The parent form of the compound differs from the various salt forms in certain physical properties, such as solubility in polar solvents, but for the purposes of this invention the salts are otherwise equivalent to the parent form of the compound.

In addition to salt forms, the present invention also provides compounds in prodrug form. Prodrugs of the compounds described herein are those compounds that readily undergo chemical changes under physiological conditions to provide the compounds of the present invention. Alternatively, prodrugs can be converted to compounds of the invention in an ex vivo environment by chemical or biochemical means. For example, a prodrug can be slowly converted to a compound of the invention when placed in a transdermal patch reservoir containing a suitable enzyme or chemical agent.

Certain compounds of the present invention may exist in unsolvated forms as well as solvated forms (including hydrated forms). In general, the solvated forms are equivalent to unsolvated forms and are intended to be encompassed within the scope of the present invention. Certain compounds of the invention may exist in polycrystalline (multiple crystaline) or amorphous form. In general, all physical forms are equivalent for the uses contemplated by the present invention and are intended to be within the scope of the present invention.

Certain compounds of the present invention have asymmetric carbon atoms (optical centers) or double bonds; racemates, diastereomers, geometric isomers, regioisomers (regioisomers), and individual isomers (e.g., separate enantiomers) are intended to be within the scope of the present invention. The compounds of the invention may also contain unnatural proportions of isotopes at one or more of the atoms that constitute such compounds. For example, the compounds may be used with radioactive isotopes such as tritium (A), (B), (C), (³H) Iodine-125 (¹²⁵I) Or carbon-14 (¹⁴C) For radiolabelling. All isotopic variations of the compounds of the present invention, whether radioactive or non-radioactive, are intended to be encompassed within the scope of the present invention.

II. Compound

In one aspect, the present invention provides compounds having formula I:

and pharmaceutically acceptable salts, hydrates, and rotamers thereof; wherein

C¹Selected from aryl and heteroaryl, wherein said heteroaryl has 1-3 heteroatoms selected from N, O and S as ring members; and wherein said aryl and heteroaryl are optionally substituted with 1 to 3R¹Substituent group substitution;

C²selected from aryl and heteroaryl, wherein said heteroaryl has 1-3 heteroatoms selected from N, O and S as ring members; and wherein said aryl and heteroaryl are optionally substituted with 1 to 3R²Substituent group substitution;

C³is selected from C_1-8Alkyl radical, C_3-8Cycloalkyl radical, C_3-8cycloalkyl-C_1-4Alkyl, aryl-C_1-4Alkyl, heteroaryl-C_1-4Alkyl, heterocycloalkyl or heterocycloalkyl-C_1-4Alkyl, wherein the heterocycloalkyl or moiety has 1-3 heteroatoms selected from N, O and S, and wherein the heteroaryl has 1-3 heteroatoms selected from N, O and S as ring members, and each C³Optionally substituted by 1-3R³Substituent group substitution;

each R¹Independently selected from: halogen, -CN, -R^c、-CO₂R^a、-CONR^aR^b、-C(O)R^a、-OC(O)NR^aR^b、-NR^bC(O)R^a、-NR^bC(O)₂R^c、-NR^a-C(O)NR^aR^b、-NR^aC(O)NR^aR^b、-NR^aR^b、-OR^aand-S (O)₂NR^aR^b(ii) a Wherein R is^aAnd R^bEach independently selected from hydrogen and C_1-8Alkyl and C_1-8Haloalkyl, or when attached to the same nitrogen atom, can combine with that nitrogen atom to form a five-or six-membered ring having from 0 to 2 additional heteroatoms selected from N, O or S as ring members; each R^cIndependently selected from C_1-8Alkyl radical, C_1-8Haloalkyl, C_3-6Cycloalkyl, heterocycloalkyl, aryl and heteroaryl, and wherein R^a、R^bAnd R^cOptionally further substituted with one to three halogens, hydroxy, methyl, amino, alkylamino and dialkylamino; and optionally when two R are¹When the substituents are on adjacent atoms, they combine to form a fused five-or six-membered carbocyclic ring;

each R²Independently selected from: halogen, -CN, -R^f、-CO₂R^d、-CONR^dR^e、-C(O)R^d、-OC(O)NR^dR^e、-NR^eC(O)R^d、-NR^eC(O)₂R^f、-NR^dC(O)NR^dR^e、-NR^dC(O)NR^dR^e、-NR^dR^e、-OR^dand-S (O)₂NR^dR^e(ii) a Wherein R is^dAnd R^eEach independently selected from hydrogen and C_1-8Alkyl and C_1-8Haloalkyl, or when attached to the same nitrogen atom, can combine with that nitrogen atom to form a five-or six-membered ring having from 0 to 2 additional heteroatoms selected from N, O or S as ring members; each R^fIndependently selected from C_1-8Alkyl radical, C_1-8Haloalkyl, C_3-6Cycloalkyl, heterocycloalkyl, aryl and heteroaryl, and wherein R^d、R^eAnd R^fOptionally further substituted with one to three halogens, hydroxy, methyl, amino, alkylamino and dialkylamino;

each R³Independently selected from: halogen, -CN, -Rⁱ、-CO₂R^g、-CONR^gR^h、-C(O)R^g、-OC(O)NR^gR^h、-NR^hC(O)R^g、-NR^hC(O)₂Rⁱ、-NR^gC(O)NR^gR^h、-NR^gR^h、-OR^g、-S(O)₂NR^gR^h、-X⁴-R^j、-X⁴-NR^gR^h、-X⁴-CONR^gR^h、-X⁴-NR^hC(O)R^g、-NHR^jand-NHCH₂R^jWherein X is⁴Is C_1-4An alkylene group; r^gAnd R^hEach independently selected from hydrogen and C_1-8Alkyl radical, C_3-6Cycloalkyl and C_1-8Haloalkyl groups, or when attached to the same nitrogen atom, may combine with the nitrogen atom to form a halogen having 0 to2 five-or six-membered ring with N, O or S as additional heteroatoms as ring members, and optionally substituted by one or two oxo groups; each RⁱIndependently selected from C_1-8Alkyl radical, C_1-8Haloalkyl, C_3-6Cycloalkyl, heterocycloalkyl, aryl, and heteroaryl; and each R^jIs selected from C_3-6Cycloalkyl, pyrrolinyl, piperidinyl, morpholinyl, tetrahydrofuryl and tetrahydropyranyl, and wherein R is^g、R^h、RⁱAnd R^jOptionally further substituted with one to three halogens, methyl, CF₃Hydroxy, amino, alkylamino and dialkylamino; and is

X is hydrogen or CH₃。

In one embodiment, in formula I, substituent C¹Selected from phenyl, pyridyl, indolyl and thiazolyl, each optionally substituted with 1 to 3R¹And (4) substituent substitution. Preferably, each R¹Independently selected from halogen, -CN, -R^c、-NR^aR^band-OR^aAnd wherein R is^aAnd R^bEach independently selected from hydrogen and C_1-8Alkyl and C_1-8Haloalkyl groups, or when attached to the same nitrogen atom, may combine with the nitrogen atom to form a pyrrolidine ring; each R^cIndependently selected from C_1-8Alkyl radical, C_1-8Haloalkyl and C_3-6Cycloalkyl, and wherein R^a、R^bAnd R^cOptionally further substituted with one to three hydroxy, methyl, amino, alkylamino and dialkylamino groups; and optionally when two R are¹When the substituents are on adjacent atoms, they combine to form a fused five-or six-membered carbocyclic ring. In selected embodiments of the invention, C¹Selected from:

returning to formula I, in one embodiment, substituent C²Selected from phenyl, naphthyl, pyridyl and indolyl, each optionally substituted with 1 to 3R²And (4) substituent substitution. Preferably, each R²Independently selected from halogen, -R^fand-OR^d(ii) a Wherein each R^dIndependently selected from hydrogen, C_1-8Alkyl and C_1-8A haloalkyl group; each R^fIndependently selected from C_1-8Alkyl radical, C_1-8Haloalkyl, C_3-6Cycloalkyl, heterocycloalkyl and heteroaryl, and wherein R^dAnd R^fOptionally further substituted with one to three halogens, hydroxy, methyl, amino, alkylamino and dialkylamino. In selected embodiments of the invention, C²Selected from:

in some embodiments, substituent C³Is selected from C_3-6Alkyl radical, C_3-6Cycloalkyl radical, C_3-6Cycloalkyl radical C_1-2Alkyl, phenyl, pyridyl, pyrazolyl, piperidinyl, pyrrolidinyl, piperidinylmethyl and pyrrolidinylmethyl, each optionally substituted with 1 to 3R³And (4) substituent substitution. Preferably, each R³Independently selected from: halogen, -Rⁱ、-CO₂R^g、-CONR^gR^h、-NR^hC(O)R^g、-NR^hC(O)₂Rⁱ、-NR^gR^h、-OR^g、-X⁴-R^j、-X⁴-NR^gR^h、-X⁴-CONR^gR^h、-X⁴-NR^hC(O)R^g、-NHR^jand-NHCH₂R^jWherein X is⁴Is C_1-3An alkylene group; r^gAnd R^hEach independently selected from hydrogen and C_1-8Alkyl radical, C_3-6Cycloalkyl and C_1-8Haloalkyl, or when attached to the same nitrogen atom, can combine with that nitrogen atom to form a five-or six-membered ring having from 0 to 1 additional heteroatom selected from N, O or S as a ring member, and optionally substituted with one or two oxo groups; each RⁱIndependently selected from C_1-8Alkyl radical, C_1-8Haloalkyl, C_3-6Cycloalkyl, heterocycloalkyl, aryl, and heteroaryl; and each R^jIs selected from C_3-6Cycloalkyl, pyrrolinyl, piperidinyl, morpholinyl, tetrahydrofuryl and tetrahydropyranyl, and wherein R is^g、R^h、RⁱAnd R^jOptionally further substituted with one to three halogens, methyl, CF₃Hydroxyl, amino, alkylamino and dialkylamino. In selected embodiments of the invention, C³Selected from:

in other embodiments, C³Selected from:

returning to formula I, X is preferably H.

A subformula of formula I:

in one embodiment of the invention, the compounds of formula I have the formula Ia:

in a second embodiment of the invention, the compounds of formula I have the formula Ib:

in a third embodiment of the invention, the compounds of formula I have the sub-formula Ic:

wherein X¹Selected from N, CH and CR¹(ii) a Subscript n is an integer of 0 to 2; x²Selected from N, CH and CR²；

Subscript m is an integer of 0 to 2.

In a fourth embodiment of the invention, the compound of formula I has the sub-formula Id:

wherein X¹Selected from N, CH and CR¹(ii) a Subscript n is an integer of 0 to 2; x²Selected from N, CH and CR²；

And subscript m is an integer of 0 to 2.

In a fifth embodiment of the invention, the compound of formula I has the sub-formula Ie:

wherein subscript p is an integer of 0 to 3; x¹Selected from N, CH and CR¹(ii) a Subscript n is an integer of 0 to 2; x²Is selected from N,CH and CR²(ii) a And subscript m is an integer of 0 to 2.

In other selected embodiments, the compounds of the present invention are represented by the formula:

wherein the substituent R¹、R²And R³And subscript p has the meaning provided for formula I.

In other selected embodiments, the compounds of the present invention are represented by the formula:

wherein the substituent R¹And R³And subscript p has the meaning provided for formula I.

In a particularly preferred group of embodiments, the compounds of the present invention are represented by formula (Ie)⁵) Is represented by, wherein R³Is selected from-NR^gR^h、-NHR^jand-NHCH₂R^jAnd R is^g、R^hAnd R^jEach having the meaning provided for in formula I.

In another particularly preferred group of embodiments, the compounds of the present invention are represented by formula (Ie)⁵) Is represented by, wherein R³Is selected from-X⁴-NR^gR^h、-X⁴-R^jand-X⁴-NR^hCOR^gIs a member of, and X⁴、R^g、R^hAnd R^jEach having the meaning provided for in formula I.

The compounds of the invention having the formula I can exist in different diastereoisomeric forms, for example as substituents C in the sub-formulae Ia and Ic¹And C²May be cis to each other or trans to each other. The terms cis or trans as used herein are used in their conventional meaning in the chemical arts, i.e., the positions of substituents relative to a reference plane (e.g., a double bond, or a ring system such as a decalin-type ring system or a hydroquinone (hydroquinolone) ring system) with respect to each other: in the cis isomer, the substituents are on the same side of the reference plane, and in the trans isomer, the substituents are on opposite sides. In addition, the invention also encompasses different conformers, as well as different rotamers. Conformers are those that can adopt conformations that differ by rotation of one or more sigma bonds. Rotamers are conformational isomers that differ by rotation of only one sigma bond.

Preparation of the Compounds

One skilled in the art will recognize that there are a variety of methods that can be used to synthesize the molecules shown in the claims. In general, the method that can be used to synthesize the molecules shown in the claims consists of four parts, which can be carried out in any order as follows: formation of a piperidine ring, installation of two amide bonds, and installation and/or modification of C¹、C²And C³A functional group of (a).

Several methods for preparing the claimed compounds are illustrated below (equations 1-6).

Equations 1-4 show some of the methods for forming the piperidine ring. Coupling at the 2-position of the pyridine ring can be achieved by transition metal mediated coupling as shown in equations 1-2, or by metal catalyzed addition of organometallic species (e.g., zincate or magnesium salt) (equation 3). After coupling at the 2-position, transition metal mediated hydrogenation of the pyridine ring yields the piperidine ring system (equations 1-3). Another method allows the piperidine ring to be made from the beta-amino acid as described in equation 4. One skilled in the art will recognize that there are many synthetic methods to produce substituted piperidines, including C-C or C-N cyclization by alkylation or ring closing metathesis with acyclic precursors. The relative stereochemistry during the hydrogenation step can be set by a variety of methods, including surface selectivity. Absolute stereochemistry may also be set by various methods, by using chiral ligands or chiral auxiliaries, separating chiral diastereomers, using chiral starting materials, or classical resolution. Compounds with 2, 3-trans stereochemistry may set the relative stereochemistry during piperidine formation, or may be obtained by epimerization of 2, 3-cis piperidine, as illustrated in equation 5.

Acylation of the piperidine ring is described in equation 6. In the case of equation 6, X may be selected from suitable groups such as OH, Cl, and F, or from any group capable of activating the carbonyl group for amine addition (e.g., OSu, imidazole, etc.). These couplings can be facilitated by the use of inorganic or organic bases, activators such as HBTU, and by catalysts, particularly those known in the art to facilitate formation of amide bonds such as DMAP, HOBT, and the like. Suitable coupling partners include carboxylic acids and piperidines, acid fluorides and amines, and the like. One skilled in the art will recognize that there are other possible combinations that produce the desired product.

Various methods have been used to prepare the compounds of the present invention, some of which are described in the examples.

A particular family of compounds of particular interest having formula I consists of the compounds as shown in figure 1, their pharmaceutically acceptable salts, hydrates and rotamers.

Pharmaceutical composition

Compositions for modulating C5a activity in humans and animals generally contain a pharmaceutically acceptable carrier or diluent in addition to the compounds provided above.

The term "composition" as used herein is intended to encompass a product comprising the specified ingredients in the specified amounts, as well as any product which results, directly or indirectly, from combination of the specified ingredients in the specified amounts. By "pharmaceutically acceptable" it is meant that the carrier, diluent or excipient must be compatible with the other ingredients of the formulation and not deleterious to the recipient thereof. Pharmaceutical compositions for administration of the compounds of the present invention may conveniently be presented in unit dosage form and may be prepared by any of the methods well known in the art of pharmacy and drug delivery. All methods include the step of bringing into association the active ingredient with the carrier which constitutes one or more additional ingredients. Generally, pharmaceutical compositions are prepared by uniformly and intimately bringing into association the active ingredient with liquid carriers or finely divided solid carriers or both, and then, if necessary, shaping the product into the desired formulation. In a pharmaceutical composition, the active object compound is included in an amount sufficient to produce the desired effect on the process or condition of the disease.

Pharmaceutical compositions containing the active ingredient may be in a form suitable for oral use, for example, as tablets, dragees, lozenges, aqueous or oily suspensions, dispersible powders or granules, emulsions and self-emulsions (as described in U.S. patent application 2002-0012680), hard or soft capsules, syrups, elixirs, solutions, buccal patches, oral gels, chewing gums (chewing gum), chewable tablets, effervescent powders and effervescent tablets. Compositions intended for oral use may be prepared according to any method known to the art for the manufacture of pharmaceutical compositions, and such compositions may contain one or more agents selected from the group consisting of: sweetening agents, flavouring agents, colouring agents, antioxidants and preservatives to provide pharmaceutically elegant and palatable preparations. Tablets contain the active ingredient in admixture with non-toxic pharmaceutically acceptable excipients which are suitable for the manufacture of tablets. These excipients may be, for example, inert diluents, such as cellulose, silica, alumina, calcium carbonate, sodium carbonate, glucose, mannitol, sorbitol, lactose, calcium phosphate or sodium phosphate; granulating and disintegrating agents, such as corn starch or alginic acid; binding agents, for example PVP, cellulose, PEG, starch, gelatin or acacia, and lubricating agents, for example magnesium stearate, stearic acid or talc. The tablets may be uncoated or they may be coated by known techniques with an enteric coating or other form of coating to delay disintegration and absorption in the gastrointestinal tract and thereby provide a long-lasting effect. For example, a time delay material such as glyceryl monostearate or glyceryl distearate may be employed. It may also be found in U.S. Pat. Nos. 4,256,108; coating by the techniques described in U.S. Pat. Nos. 4,166,452 and 4,265,874 to form osmotic therapeutic tablets for controlled release.

Formulations for oral use may also be presented as hard gelatin capsules wherein the active ingredient is mixed with an inert solid diluent, for example, calcium carbonate, calcium phosphate or kaolin, or as soft gelatin capsules wherein the active ingredient is mixed with water or an oil medium, for example peanut oil, liquid paraffin or olive oil. Additionally, emulsions can be prepared with water-immiscible ingredients (e.g., oils) and stabilized with surfactants (e.g., mono-glycerides, di-glycerides, PEG esters, and the like).

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

Oily suspensions may be formulated by suspending the active ingredient in a vegetable oil, for example arachis oil, olive oil, sesame oil or coconut oil, or in a mineral oil such as liquid paraffin. Oily suspensions may contain a thickening agent, for example beeswax, hard paraffin or cetyl alcohol. Sweetening agents such as those set forth above, and flavoring agents may be added to provide a palatable oral preparation. These compositions may be preserved by the addition of an antioxidant such as ascorbic acid.

Dispersible powders and granules suitable for preparation of an aqueous suspension by the addition of water provide the active ingredient in admixture with a dispersing or wetting agent, suspending agent and one or more preservatives. Examples of suitable dispersing or wetting agents and suspending agents are described above. Other excipients, for example sweetening, flavouring and colouring agents, may also be present.

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

Syrups and elixirs may be formulated with sweetening agents, for example glycerol, propylene glycol, sorbitol or sucrose. These preparations may also contain a demulcent (demulcent), a preservative and flavouring and colouring agents. Oral solutions can be prepared with, for example, cyclodextrin, PEG, and surfactant.

The pharmaceutical compositions may be in the form of a sterile injectable aqueous or oleaginous suspension. This suspension may be formulated according to the known art using those suitable dispersing or wetting agents and suspending agents which have been mentioned above. The sterile injectable preparation may also be a sterile injectable solution or suspension in a non-toxic parenterally-acceptable diluent or solvent, for example as a solution in 1, 3-butanediol. Acceptable carriers and solvents that may be employed include water, ringer's solution and isotonic sodium chloride solution. In addition, sterile fixed oils are conventionally employed as a solvent or suspending medium. For secondary purposes, any bland fixed oil may be employed including synthetic mono-or diglycerides. In addition, fatty acids (e.g., oleic acid) may be used in the preparation of injectables.

The compounds of the present invention may also be administered in the form of suppositories for rectal administration of the drug. These compositions can be prepared by mixing the drug with a suitable non-irritating excipient which is solid at ordinary temperatures but liquid at the rectal temperature and will therefore melt in the rectum to release the drug. These materials include cocoa butter and polyethylene glycols. In addition, the compounds may be administered by ocular delivery via a solution or ointment. In addition, transdermal delivery of the compound of interest may be achieved by iontophoresis patches and the like. For topical use, creams, ointments, jellies, solutions or suspensions, etc., containing the compounds of the present invention are employed. Topical application as used herein is also intended to include the use of mouthwashes and gargles (gargle).

The compounds of the invention may also be coupled to carriers which are suitable polymers as carriers for targetable drugs. These polymers may include polyvinylpyrrolidone, pyran copolymer, polyhydroxypropyl-methacrylamide-phenol, polyhydroxyethyl-asparagine-phenol, or polyoxyethylene-polylysine substituted with palmitoyl residues. In addition, the compounds of the present invention may be coupled to a carrier which is a type of biodegradable polymer that may be used to achieve controlled release of the drug, such as polylactic acid, polyglycolic acid, copolymers of polylactic and polyglycolic acid, polyepsilon caprolactone, polyhydroxy butyric acid, polyorthoesters, polyacetals, polydihydropyrans, polycyanoacrylates, and crosslinked or amphiphilic block copolymers of hydrogels. The polymer and semipermeable polymer matrices can be formed into shaped articles, such as valves, stents, tubes, prostheses, and the like. In one embodiment of the invention, the compounds of the invention are coupled to a polymeric or semipermeable polymeric matrix shaped as a stent or stent-graft device.

Methods of treating diseases and disorders modulated by C5a

The compounds of the present invention may be used as agonists, (preferably) antagonists, partial agonists, inverse agonists (inverse agonst) of the C5a receptor in a variety of contexts, both in vitro and in vivo. In one embodiment, the compounds of the invention are C5aR antagonists, which are useful for inhibiting the binding of C5a receptor ligands (e.g., C5a) to the C5a receptor in vitro or in vivo. Generally, these methods comprise the step of contacting the C5a receptor with a sufficient amount of one or more C5a receptor modulators as provided herein in an aqueous solution in the presence of a C5a receptor ligand or under other conditions suitable for binding the ligand to the C5a receptor. The C5a receptor may be present in suspension (e.g., in an isolated membrane or cell preparation), in cultured or isolated cells, or in a tissue or organ.

Preferably, the amount of C5a receptor modulator contacted with the receptor should be sufficient to inhibit the in vitro binding of C5a to C5a receptor as measured, for example, using a radioligand binding assay, a calcium mobilization assay, or a chemotaxis assay as described herein.

In one embodiment of the invention, a C5a modulator of the invention is used to modulate (preferably inhibit) the signaling activity of the C5a receptor, for example by contacting one or more compounds of the invention with the C5a receptor under conditions (in vitro or in vivo) suitable for the modulator to bind to the receptor. The receptor may be present in solution or suspension, in a cultured or isolated cell preparation, or in the patient. Any modulation of signaling activity can be assessed by measuring the effect on calcium mobilization by calcium ions or by measuring the effect on C5a receptor-mediated cell chemotaxis. Generally, an effective amount of a C5a modulator is an amount sufficient to modulate the in vitro signaling activity of the C5a receptor in a calcium mobilization assay or modulate the chemotaxis of cells mediated by the C5a receptor in a migration assay.

When the compounds of the invention are used to inhibit C5a receptor-mediated cell chemotaxis, preferably chemotaxis of leukocytes (e.g., neutrophils), in an in vitro chemotaxis assay, these methods comprise contacting leukocytes, particularly primate leukocytes, particularly human leukocytes, with one or more compounds of the invention. Preferably, the concentration is sufficient to inhibit leukocyte chemotaxis in an in vitro chemotaxis assay such that the level of chemotaxis observed in a control assay is significantly higher than that observed in an assay to which a compound of the invention has been added, as described above.

In another embodiment, the compounds of the present invention are also useful for treating patients suffering from conditions responsive to modulation of the C5a receptor. The term "treatment" as used herein encompasses both treatments that alter the disease and symptomatic treatments, either of which can be prophylactic (i.e., prevent, delay, or reduce the severity of symptoms prior to onset of symptoms) or therapeutic (i.e., reduce the severity and/or duration of symptoms after onset of symptoms). As used herein, a condition is considered "responsive to modulation of the C5a receptor" if modulation of the activity of the C5a receptor results in a reduction in the inappropriate activity of the C5a receptor. The term "patient" as used herein includes primates (especially humans), domesticated companion animals (e.g., dogs, cats, horses, etc.), and livestock (e.g., cattle, pigs, sheep, etc.), in dosages as described herein.

Disorders treatable by C5a modulation:

autoimmune diseasesFor example, rheumatoid arthritis, systemic lupus erythematosus, Guillain-Barre syndrome, pancreatitis, lupus nephritis, lupus glomerulonephritis, psoriasis, Crohn's disease, vasculitis, irritable bowel syndrome, dermatomyositis, multiple sclerosis, bronchial asthma, pemphigus, pemphigoid, scleroderma, myasthenia gravis, autoimmune hemolysis and thrombocytopenia states, Goodpasture's syndrome (and associated glomerulonephritis and pulmonary hemorrhage), immune vascular diseaseInflammation, tissue transplant rejection, hyperacute rejection of transplanted organs, and the like.

Inflammatory disorders and related conditionsFor example, neutropenia, sepsis, septic shock, alzheimer's disease, multiple sclerosis, stroke, Inflammatory Bowel Disease (IBD), inflammation associated with severe burns, lung injury and ischemia-reperfusion injury, osteoarthritis, as well as acute (adult) respiratory distress syndrome (ARDS), Chronic Obstructive Pulmonary Disease (COPD), Systemic Inflammatory Response Syndrome (SIRS), allergic dermatitis, psoriasis, chronic urticaria and Multiple Organ Dysfunction Syndrome (MODS). Also included are pathological sequelae associated with: insulin-dependent diabetes mellitus (including diabetic retinopathy), lupus nephropathy, Heyman nephritis (Heyman nephritis), membranous nephritis (membrane nephritis) and other forms of glomerulonephritis, contact hypersensitivity, and inflammation caused by blood contact with artificial surfaces that can lead to complement activation, such as occurs during extracorporeal blood circulation (e.g., during hemodialysis, or by heart-lung machines (e.g., associated with vascular procedures such as coronary artery bypass grafting or heart valve replacement)) or inflammation associated with contact with other artificial blood vessels or container surfaces (e.g., ventricular assist devices, artificial heart machines, blood transfusion vessels, blood storage bags, plasmapheresis, thrombopheresis, etc.). Also included are diseases associated with ischemia/reperfusion injury, for example, diseases caused by transplantation (including solid organ transplantation), and syndromes such as ischemia reperfusion injury, ischemic colitis, and cardiac ischemia. The compounds of the invention are also useful in the treatment of age-related macular degeneration (Hageman et al, P.N.A.S.102: 7227-7232, 2005).

Cardiovascular and cerebrovascular disordersFor example, myocardial infarction, coronary artery embolism, vessel occlusion, surgical vascular reocclusion, atherosclerosis, traumatic central nervous system injury and ischemic heart disease. In one embodiment, patients at risk for myocardial infarction or embolism (i.e., having one or more of the following) may be treatedA recognized risk factor for myocardial infarction or embolism (such as, but not limited to, obesity, smoking, hypertension, hypercholesterolemia, previous or genetic history of myocardial infarction or embolism) is administered an effective amount of a compound of the invention to reduce the risk of myocardial infarction or embolism.

Vasculitis diseaseVasculitis diseases are characterized by inflammation of the blood vessels. Leukocyte infiltration causes vascular wall destruction, and the complement pathway is believed to play an important role in initiating leukocyte migration and resultant damage manifested at sites of inflammation (Vasculitis, 2 nd edition, Ball and Bridges, eds., oxford university Press, pages 47-53, 2008). The compounds provided by the invention can be used for treating leucocyte disruptive vasculitis (leukocytic vasculitis), Wegener's granulomatosis (Wegener's granulomatosis), microscopic polyangiitis (microscopical polyangiitis), Scherger-Strauss syndrome (Churg-Strauss syndrome), Henoch-Schonenopurpa, polyarteritis nodosa, Rapidly progressive glomerulonephritis (Rapid progressive glomeloropephritis, RPGN), cryoglobulinemia, Giant Cell Arteritis (GCA), Behcet's disease and Takayasu's Arteritis (TAK).

HIV infection and AIDSModulators of the C5a receptor provided herein are useful for inhibiting HIV infection, delaying the progression of AIDS or reducing the symptoms or severity of HIV infection and AIDS.

Neurodegenerative disorders and related diseases-among other aspects, the C5a antagonists provided herein can be used to treat alzheimer's disease, multiple sclerosis, and cognitive decline associated with cardiopulmonary bypass surgery and related processes.

In one embodiment of the invention, the compounds of the invention are useful for the treatment of a disease selected from: sepsis (and related conditions), COPD, rheumatoid arthritis, lupus nephritis, and multiple sclerosis.

The methods of treatment provided herein generally comprise administering to a patient an effective amount of one or more compounds provided herein. Suitable patients include patients suffering from or susceptible to (i.e., prophylactically treated) a condition or disease identified herein. Typical patients for treatment as described herein include mammals, particularly primates, and particularly humans. Other suitable patients include domesticated companion animals such as dogs, cats, horses, etc., or livestock animals such as cattle, pigs, sheep, etc.

In general, the methods of treatment provided herein comprise administering to a patient an effective amount of one or more compounds provided herein. In a preferred embodiment, the compounds of the invention are preferably administered to a patient (e.g., a human) orally or in a topical manner. The effective amount may be an amount sufficient to modulate the activity of the C5a receptor and/or an amount sufficient to reduce or alleviate symptoms exhibited by the patient. Preferably, the amount administered is an amount sufficient to produce a sufficiently high plasma concentration of the compound (or its active metabolite if the compound is a prodrug)) to detectably inhibit chemotaxis of leukocytes (e.g., neutrophils) in vitro. The treatment regimen may vary depending upon the compound used and the particular condition being treated; for the treatment of most conditions, a frequency of administration of 4 times daily or less is preferred. In general, a 2-time daily dosing regimen is more preferred, with once daily dosing being particularly preferred. It will be understood, however, that the specific dose level and treatment regimen for any particular patient will depend upon a variety of factors including the activity of the specific compound employed, the age, body weight, general health, sex, diet, time of administration, route of administration, rate of excretion, drug combination (i.e., other drugs administered to that patient), and the severity of the particular disease undergoing therapy, and the judgment of the attending medical personnel. In general, it is preferred to use the minimum dose sufficient to provide effective treatment. The effectiveness of a treatment in a patient can generally be monitored using medical or veterinary standards appropriate to the condition being treated or prevented.

Dosage levels of about 0.1mg to about 140mg per kilogram of body weight per day are useful for treating or preventing conditions involving pathogenic C5a activity (about 0.5mg to about 7g per day per human patient). The amount of active ingredient that may be combined with the carrier materials to produce a single dosage form will vary depending upon the host treated and the particular mode of administration. Unit dosage forms typically contain from about 1mg to about 500mg of the active ingredient. For compounds to be administered orally, transdermally, intravenously or subcutaneously, preferably a sufficient amount of the compound to be administered should be administered to achieve a serum concentration of 5ng (nanogram)/mL-10 μ g (microgram)/mL serum, more preferably a sufficient amount of the compound should be administered to achieve a serum concentration of 20ng-1 μ g/mL serum, and most preferably a sufficient amount of the compound should be administered to achieve a serum concentration of 50ng/mL-200ng/mL serum. For direct injection into the synovium (for the treatment of arthritis), sufficient compound should be administered to achieve a local concentration of about 1 micromole per liter.

The frequency of administration may also vary depending on the compound used and the particular disease being treated. However, for the treatment of most conditions, a regimen of 4 times daily, 3 times daily or less is preferred, with a regimen of 1 or 2 times daily being more preferred. It will be understood, however, that the specific dose level for any particular patient will depend upon a variety of factors including the activity of the specific compound employed, the age, body weight, general health, sex, diet, time of administration, route of administration and rate of excretion, drug combination (i.e., the other drugs administered to that patient), the severity of the particular disease undergoing therapy, and other factors including the judgment of the attending medical personnel.

In another aspect of the invention, the compounds of the invention are useful in a variety of non-pharmaceutical in vitro and in vivo applications. For example, the compounds of the present invention can be labeled and used as probes for detecting and localizing the C5a receptor (cell preparation or tissue slice specimen). The compounds of the invention are also useful as positive controls in assays for C5a receptor activity, i.e., as standards for determining the ability of a candidate agent to bind to the C5a receptor, or as radiotracers for Positron Emission Tomography (PET) imaging or Single Photon Emission Computed Tomography (SPECT). These methods can be used to characterize the C5a receptor in living subjects. For example, the C5a receptor modulator can be labeled using any of a variety of well-known techniques (e.g., radiolabeling with a radionuclide such as tritium) and incubated with the sample for an appropriate incubation time (e.g., as determined by first determining the time course of binding). After incubation, unbound compounds are removed (e.g., by washing) and bound compounds are detected using any method appropriate to the label used (e.g., autoradiography or scintillation counting of radiolabeled compounds; spectroscopy can be used to detect luminophores and fluorophores). As a control, a matched sample containing a labeled compound and a greater (e.g., 10-fold greater) amount of unlabeled compound can be treated in the same manner. The retention of a greater amount of detectable label in the test sample than in the control indicates the presence of C5a receptor in the sample. Assays for detection of the C5a receptor, including receptor autoradiography (receptor localization), can be performed in cultured cell or tissue samples as described by Kuhar in Current Protocols in Pharmacology (1998) John Wiley & Sons, New York, sections 8.1.1 to 8.1.9.

The compounds provided herein can also be used in a variety of well-known cell separation methods. For example, modulators may be attached to the inner surface of a tissue culture plate or other support to serve as immobilized affinity ligands and thereby isolate the C5a receptor in vitro (e.g., to isolate receptor-expressing cells). In a preferred application, a modulator linked to a fluorescent marker (e.g., fluorescein) is contacted with the cells, which are then analyzed (or isolated) by Fluorescence Activated Cell Sorting (FACS).

In fig. 1, the structures and activities of representative compounds described herein are provided. For the binding assays described herein, the activities are provided as follows: IC + 500nM < 1 >₅₀＜2000nM；++，50nM≤IC₅₀＜500nM；+++，5nM≤IC₅₀< 50 nM; and + + + +, IC₅₀＜5nM。

V. examples

The following examples are provided to illustrate, but not to limit, the claimed invention. The reagents and solvents used hereinafter are available from commercial sources, such as Aldrich Chemical Co. (Milwaukee, Wisconsi)n, USA). Recording with a Varian Mercury 400MHz NMR spectrometer¹H-NMR spectrum. Significant peaks were provided relative to TMS and are shown in the table in the following order: multiplicity (s, singlet; d, doublet; t, triplet; q, quartet; m, multiplet) and number of protons. Mass spectrometry results are reported as mass-to-charge ratios, followed by the relative abundance of each ion (in parentheses). In the examples, a single M/e value is reported for the M + H (or M-H as indicated) ion containing the most common atomic isotope. In all cases, the isotopic pattern corresponds to the expected formula. Electrospray ionization (ESI) mass spectrometry was performed on a Hewlett-Packard MSD electrospray mass spectrometer using HP1100 HPLC for sample delivery. Typically the analyte is dissolved in methanol at 0.1mg/mL and 1 microliter is infused with the delivery solvent into a mass spectrometer scanning from 100 to 1500 daltons. All compounds can be analyzed in positive ESI mode using acetonitrile/water with 1% formic acid as the delivery solvent. The compounds provided below can also be used in negative ESI mode, using 2mM NH in acetonitrile/water₄The OAc solution was analyzed as a delivery system.

The following abbreviations are used in the examples and throughout the present specification:

compounds within the scope of the present invention may be synthesized as described below using a variety of reactions known to those skilled in the art. It will also be recognized by those skilled in the art that alternative methods may be employed to synthesize the subject compounds of the present invention, and that the methods described in this document are not exhaustive, but rather provide a widely applicable and practical route to the subject compounds.

Certain molecules claimed in this patent may exist in different enantiomeric and diastereomeric forms, and all such variations of these compounds are claimed.

The molecules are obtained herein through a detailed description of the experimental procedures used to synthesize key compounds, which are described by the physical data identifying them and the structural descriptions associated with them.

Those skilled in the art will also recognize that acids and bases are often used during standard processing procedures in organic chemistry. During the experimental procedures described in this patent, salts are sometimes prepared if the parent compound has the requisite inherent acidity or basicity.

Example 1

Synthesis of cis-1- (2-fluoro-6-methylbenzoyl) -2-phenylpiperidine-3-carboxylic acid (3-trifluoromethylphenyl) amide

a) Pd (PPh)₃)₄(3.0g, 2.6mmol) was added to 2-chloro-3-carboxyethylpyridine (25g, 134.7mmol), phenylboronic acid (21.04g, 172.6mmol) and K₂CO₃(55.1g, 399mmol) in 1, 4-bisAlkane (200mL) and water (200 mL). The reaction mixture was heated at 100 ℃ for 2 hours. The solution was then cooled to room temperature and the di-s removed under reduced pressureAn alkane. The resulting aqueous layer was extracted with ethyl acetate and the combined organic layers were dried (Na)₂SO₄) Filtered through celite and concentrated under reduced pressure. By flash chromatography (SiO)₂10-100% EtOAc/hexanes) to afford the 2-phenylpyridine derivative (91% yield, 27.98 g). LC-MS R_t(retention time): 2.45 min, MS: (ES) M/z 228(M + H)⁺)。

b) To PtO₂(800mg，352mmol) was added to a solution of 2-phenyl-nicotinic acid ethyl ester (20g, 88mmol, prepared in step a above) in EtOH (60mL) and concentrated hydrochloric acid (15 mL). The reaction mixture was hydrogenated using a Parr shaker at 40-45psi for 1 hour. The reaction mixture was then filtered through celite, washed with EtOH, and the filtrate was concentrated under reduced pressure. The residue is treated with CH₂Cl₂Diluted and saturated NaHCO₃And (6) washing. By flash chromatography (SiO)₂，0-20％MeOH/CH₂Cl₂) Purification gave the desired product (85% yield, 17.4 g). LC-MS R_t(retention time): 1.73 min, MS: (ES) M/z 234(M + H)⁺)。

c) Oxalyl chloride (3.2mL, 30.75mmol) was added to 2-fluoro-6-methylbenzoic acid (3.79g, 24.6mmol) in the reaction flask in CH at room temperature₂Cl₂(20mL) and then a catalytic amount of DMF was added. The reaction solution was kept stirred at room temperature for 2 hours. The solvent and excess oxalyl chloride were removed in vacuo and the residue was dried under high vacuum for 20 min. Dissolving the obtained acyl chloride in anhydrous CH₂Cl₂(20mL) and cooled to 0 deg.C, then piperidine (5.56g, 20.5mmol) from step b and Et were added₃N (8.6mL, 61.5 mmol). The mixture was then allowed to warm to room temperature and stirred overnight. With CH₂Cl₂The reaction mixture was diluted and water was added. Separating the layers with CH₂Cl₂The aqueous layer was extracted. The combined organic layers were dried (MgSO)₄) And concentrated under reduced pressure. By flash chromatography (SiO)₂10-35% EtOAc/hexanes) to yield 7.47g of the desired compound (99% yield). LC-MS R_t(retention time): 2.50 min and 2.58 min (two rotamers), MS: (ES) M/z 370(M + H)⁺)。

d) Lithium aluminum hydride solution (2.0M in THF, 8.2mL, 16.4mmol) was added to a solution of the ester from step c (2.98g, 8.06mmol) in THF (100mL) at 0 deg.C. The resulting solution was stirred for 2 hours while maintaining at 0 ℃ at which point the reaction was complete. 15% aqueous NaOH (625 μ L) was added dropwise to quench the reaction, followed by H₂O (625. mu.L). To the cloudy colloidal mixture was added water (1.85mL) andthe mixture was kept stirring at room temperature for 1 hour. The mixture was then filtered through a plug of celite and the filtrate was concentrated under reduced pressure. By flash chromatography (SiO)₂33-67% EtOAc/hexanes) to yield 2.46g of the desired product (93% yield). LC-MS: r_t(retention time): 1.90 min and 2.09 min (two rotamers), MS: (ES) M/z 328(M + H)⁺)。

e) A solution of the alcohol (1.42g, 4.33mmol) from step d in acetic acid (65ml) was added to CrO at room temperature₃(2.61g, 26.1mmol) in H₂O (16ml) in a slurry. The resulting mixture was kept stirring at room temperature until the reaction was complete (90 min). The mixture was filtered through a plug of celite and the filtrate was concentrated under reduced pressure. By flash chromatography (SiO)₂，3-10％CH₂Cl₂MeOH then 50-67% EtOAc/hexanes) to give 1.03g of the desired product (70% yield). LC-MS: r_t(retention time): 1.88 min and 2.12 min (two rotamers), MS: (ES) M/z 342(M + H)⁺)。

f) 3-Trifluoromethylaniline (16.2mg, 0.1mmol, 1.0 equiv.) was added to the CH solution of the acid (34.2mg, 0.1mmol) prepared above and triethylamine (6 equiv.)₂Cl₂(1mL) in solution. T3P (95.5mg, 0.15mmol) was then added slowly and the solution was stirred at room temperature for 1.5 h. Reacting the reaction mixture with CH₂Cl₂(1mL) diluted sequentially with 1N aqueous HCl and saturated NaHCO₃And (4) washing with an aqueous solution. The organic layer was separated and dried over anhydrous MgSO₄Dried and concentrated under reduced pressure. By flash chromatography (SiO)₂5-40% EtOAc/hexanes) to yield 35mg (73% yield) of the product as a white solid.¹H NMR(400MHz，CDCl₃)δ1.22-2.45(m，8H)，2.93-3.32(m，3H)，6.77-7.82(m，12H)，9.10(s，0.38H)，9.30(s，0.62H)。LC-MS：R_t(retention time) 2.88 min, MS: (ES) M/z 485(M + H)⁺)。

Example 2

Synthesis of N- (3-tert-butylphenyl) -1- (5-chloro-3-methylpyridinoyl) -2-phenylpiperidine-3-carboxamide

a) 2-Chloronicotinyl chloride (1.05 equiv.) dissolved in anhydrous dichloromethane (0.5M) was added to 3-tert-butylaniline (1 equiv.) and 2M K over 30 minutes at 0 deg.C₂CO₃Aqueous (2.2 equiv.) solution in anhydrous dichloromethane (0.5M) and the reaction mixture stirred at room temperature for an additional 1.5 hours. The layers were separated and the aqueous layer was extracted with dichloromethane. The combined organic layers were washed with brine and dried (MgSO)₄) Filtered and concentrated to give the desired amide as a foamy solid, which was used in the next step without further purification. MS: (ES) M/z 289.1(M + H)⁺)。

b) Pd (PPh)₃)₄(2-5 mol%) was added to picolinamide (1 equivalent), phenylboronic acid (1.4 equivalents), and 2M K above₂CO₃Aqueous (2.4 equiv.) solution in toluene (0.7M) and the reaction mixture was heated at 100 ℃ overnight (about 12 hours). After cooling to room temperature, the reaction mixture was filtered through celite and the celite plug was washed with EtOAc. The filtrate was diluted with water and extracted with EtOAc and dried (MgSO)₄) Filtered and concentrated under reduced pressure. By automated flash chromatography (SiO)₂10% to 100% gradient EtOAc-hexanes) and dried in vacuo to give 2-phenyl-3-carboxyamide pyridine in 60-75% yield, MS: (ES) M/z 331.2(M + H)⁺)。

c) To PtO₂(10 mol%) was added to a solution of the 2-phenylpyridine derivative prepared above (1 eq) in EtOH and concentrated hydrochloric acid (excess, 4: 1 ratio) and the reaction mixture was hydrogenated using a Parr shaker at 40-45psi for 1.5 hours. It was filtered through celite, washed with EtOH, and the filtrate was concentrated. With CH₂Cl₂The residue was diluted and washed with saturated NaHCO₃And (4) washing with an aqueous solution. Then through automationFlash chromatography (SiO)₂Gradient CH of 1% to 30%₂Cl₂MeOH) and dried in vacuo to afford the title compound as a foamy solid (yield about 85%). MS: (ES) M/z 337.2(M + H)⁺)。

d) 5-chloro-3-methylpyridinecarboxylic acid (30mg, 0.16mmol) and N- (3-tert-butylphenyl) -2-phenylpiperidine-3-carboxamide (50mg, 0.15mmol, prepared in step c above) were dissolved in anhydrous DMF (1 mL). N, N-diisopropylethylamine (0.15mL) was added at room temperature, followed by HCTU (67mg, 0.16 mmol). After stirring for 2 hours at ambient temperature, LC-MS and TLC indicated the reaction was complete. The reaction mixture was diluted with EtOAc (50mL), and saturated NaHCO with 1N HCl (20mL)₃(30mL) and brine (30mL), and the resulting solution was concentrated under reduced pressure. By preparative HPLC (20 → 95% gradient of MeCN-H₂O, containing 0.1% TFA) and the pure fractions were lyophilized to give the title compound (50mg, 67% yield). HPLC retention time 2.88 mins.¹H NMR(400MHz，CDCl₃) δ 8.42(d, 1H, J ═ 0.8Hz), 7.97(br, 1H), 7.59(d, 1H, J ═ 0.8Hz), 7.56(d, 1H, J ═ 7.6Hz), 7.34(m, 3H), 7.20(m, 3H), 7.10(d, 1H, J ═ 7.6Hz), 6.61 (two sets of br, 1H), 3.12 (two sets of m, 2H), 2.94 (three sets of m, 1H), 2.36(s, 3H), 2.20 (two sets of br, 2H), 1.74 (composite broad peak, 2H), 1.29(s, 9H). MS: (ES) M/z 490.2(M + H)⁺)。

Example 3

Synthesis of cis-1- (2-methylbenzoyl) -2- (3-fluorophenyl) piperidine-3-carboxylic acid (3-tert-butylphenyl) amide

a) To a solution of N- (3-tert-butylphenyl) -2-chloronicotinamide (570.2mg, 2mmol), 3-fluorophenylboronic acid (401.2mg, 2.8mmol), 3mL of toluene and 1mL of 2N potassium carbonate in waterTo the mixture in (1) was added tetrakis (triphenylphosphine) palladium (0) (234.5mg, 0.2 mmol). The mixture was then heated at 90 ℃ under nitrogen for 3 hours, and subsequently cooled to room temperature. The reaction mixture was then diluted with 30mL of water and 150mL of letoac. The organic layer was separated, washed with brine and dried (Na)₂SO₄). The organic solvent was removed under reduced pressure and the residue was purified by column on silica gel (40% EtOAc in hexane) to give N- (3-tert-butylphenyl) -2- (3-fluorophenyl) nicotinamide (691.4mg, 99%). MS: (ES) M/z394.5(M + H)⁺)。

b) A mixture of N- (3-tert-butylphenyl) -2- (3-fluorophenyl) nicotinamide (501.2mg, 1.4mmol), platinum oxide (51.9mg, 0.21mmol) and concentrated hydrochloric acid (400. mu.l, 5.2mmol) in 5mL of ethanol was stirred vigorously under a hydrogen balloon overnight. The mixture was filtered and the solid was washed three times with 25mL methanol. The combined solutions were dried under reduced pressure. To the residue was added 30mL of saturated sodium bicarbonate and 150mL of EtOAc. The organic layer was separated and dried over sodium sulfate. The solvent was evaporated to give crude 2- (3-fluorophenyl) piperidine-3-carboxylic acid (3-tert-butylphenyl) amide as a brown solid, which was used directly in the next step. MS: (ES) M/z 355.7(M + H)⁺)。

c) To a solution of 2- (3-fluorophenyl) piperidine-3-carboxylic acid (3-tert-butylphenyl) amide (prepared above, 177.3mg, 0.5mmol) in 2mL of dichloromethane at room temperature was added Et₃N (100. mu.l, excess) and 2-methylbenzoyl chloride (92.3mg, 0.6 mmol). The resulting solution was then stirred at this temperature until the reaction was complete (10 minutes). The reaction mixture was then loaded directly onto a silica gel column and purified by using ISCO (30% EtOAc in hexanes to give the final product 2- (3-fluorophenyl) -1- (2-methylbenzoyl) piperidine-3-carboxylic acid (3-tert-butylphenyl) amide (151.2mg, 64% yield).¹HNMR(400MHZ，CDCl₃Mixtures of rotamers): δ 7.91(s, 0.6H), 7.85(s, 0.4H), 7.18 to 7.46(m, 9H), 7.11(m, 1H), 6.95(m, 1H), 6.67(d, J ═ 1.2Hz, 1H), 3.36(d, J ═ 1.6Hz, 0.4H), 3.26(d, J ═ 1.6Hz, 1H), 3.05(m, 1H), 2.89(t, J ═ 1.2Hz, 1H), 2.45(s, 1H), 2.02 to 2.40(m, 4H), 1.70 to 1.84(m, 3H), 1.44 to 1.64(s, 1H), 1.32(s,6H)，1.25(s，1H)。MS：(ES)m/z 473.2(M+H⁺)。

example 4

Synthesis of cis-1- (2-methylbenzoyl) -2- (2, 2-dimethylpropyl) piperidine-3-carboxylic acid (3-tert-butylphenyl) amide

a) To a stirred solution of 2-bromonicotinic acid (1.01g, 5mmol) dissolved in anhydrous dichloromethane (8mL) was added EDCI (1.34g, 7mmol) and 3-tert-butylaniline (0.74g, 5mmol) at room temperature and the reaction mixture was stirred for 12 hours. The mixture was then diluted with dichloromethane and washed with saturated sodium bicarbonate and water. The dichloromethane layer was dried over anhydrous magnesium sulfate, filtered and concentrated under reduced pressure. The residue was purified by flash chromatography to give 2-bromo-N- (3-tert-butylphenyl) nicotinamide (59% yield, 950 mg). R_t: 2.44 minutes (20-100-5 method). MS: (ES) M/z 333, 335(M + H)⁺)。

b)2, 2-Dimethylpropylmagnesium chloride (1M diethyl ether, 4.8mL, 4.8mmol) was added to a suspension of copper cyanide (215mg, 2.40mmol) in THF (6mL) at-78 deg.C. After stirring at the same temperature for 1 hour, 2-bromo-N- (3-tert-butylphenyl) nicotinamide (200mg, 0.601mmol) as a solid was added in one portion. The reaction mixture was gradually warmed to room temperature and the reaction was stirred overnight. Saturated ammonium chloride solution and ethyl acetate were added and the reaction mixture was filtered through celite and washed with ethyl acetate. The layers were separated and the product was extracted again with ethyl acetate. The combined organic layers were washed with brine and dried over anhydrous sodium sulfate. After removing the solvent under reduced pressure, the crude material was purified by silica gel column chromatography (gradient using 20% to 50% ethyl acetate in hexane) to give N- (3-tert-butylphenyl) -2- (2, 2-dimethylpropyl) nicotinamide (168mg, 0.517mmol, 86%). Rf 0.45 (toluene: ethyl acetate 2: 1).

c) N- (3-tert-butylphenyl) -2- (2, 2-dimethylpropyl) nicotinamide (168mg, 0.517mmol) was dissolved in ethanol (5 mL). Platinum oxide (11.6mg, 0.0511mmol) was added followed by concentrated hydrochloric acid (250. mu.L). The reaction mixture was hydrogenated using a Parr apparatus at 45psi for 1.5 hours. Analysis of the reaction mixture showed incomplete conversion and the sequence was repeated once more. The platinum oxide was filtered off and the solvent was removed under reduced pressure. The crude material was neutralized with saturated sodium bicarbonate solution and extracted with ethyl acetate. The organic layer was then washed with brine and dried over anhydrous magnesium sulfate. The solvent was removed under reduced pressure to give crude 2, 3-cis-2- (2, 2-dimethylpropyl) piperidine-3-carboxylic acid- (3-tert-butylphenyl) amide (153mg), which was used in the next step without further purification.

d) To a solution of 2, 3-cis-2- (2, 2-dimethylpropyl) piperidine-3-carboxylic acid- (3-tert-butylphenyl) amide (84.8mg, 0.257mmol) in pyridine (415. mu.L, 5.13mmol) at room temperature was added a solution of 2-methylbenzoyl chloride (81.6mg, 0.528mmol) in chloroform (415. mu.L). A catalytic amount (unweighted) of dimethylaminopyridine was added to enhance the reaction and the mixture was stirred for three days. Ethyl acetate and water were then added to the reaction mixture, and the product was extracted three times with ethyl acetate. The combined organic layers were dried over anhydrous magnesium sulfate. After removal of the solvent under reduced pressure, the crude material was purified by silica gel chromatography using 10% to 20% ethyl acetate in hexane to give 2, 3-cis-2- (2, 2-dimethylpropyl) -1- (2-methylbenzoyl) piperidine-3-carboxylic acid- (3-tert-butylphenyl) amide (47.0mg, 0.105mmol, 41%). Rf 0.6 (hexane: ethyl acetate 2: 1). Rt 3.16 min, 3.26 min. (the compound exists as a mixture of several conformational isomers. 20-100-5 method).¹H NMR(CDCl₃)δ9.68(s，1H)，9.43(s，1H)，8.33(s，1H)，8.28(s，1H)，6.97-7.79(m，8H)，5.48(br，1H)，5.39(dd，J＝4，10Hz，1H)，5.33(dd，J＝6，6Hz，1H)，3.38(ddd，J＝4，14，14Hz，2H)，3.25(dd，J＝13，13Hz，2H)，2.66(dd，J＝4，8.4Hz，1H)，2.63(ddd，J＝2.8，2.8，8Hz，1H)，2.50(s，9H)，2.40(s，9H)，2.25(s，9H)，2.13(s，9H)，1.79-1.99(m，2H)，1.23-1.56(m，2H)，1.32(s，9H)，1.07(s，9H)，1.06(s，9H)，0.97(s，9H)，0.95(s，9H)。MS：(ES)m/z 449(M+H⁺)。

Example 5

Synthesis of cis-2-cyclopentyl-1- (2-methylbenzoyl) piperidine-3-carboxylic acid (3-tert-butylphenyl) amide

a) Cyclopentylzinc bromide (0.5M, 6.5mL, 3.26mmol) was added under nitrogen to room temperature stirred methyl 2-chloronicotinate (400mg, 2.33mmol), CuI (19mg, 0.1mmol) and Pd (dppf) Cl₂(42mg, 0.06mmol) in anhydrous dimethylacetamide (1.7 mL). The reaction mixture was heated to 70 ℃ for 3.5 hours, cooled to room temperature, filtered through celite, and the filter cake was rinsed with ethyl acetate. The filtrate was washed with water, brine and dried (MgSO)₄) Filtered and concentrated under reduced pressure. By flash chromatography (SiO)₂10-100% EtOAc/hexanes) to afford the desired compound (83% yield, 400 mg). LC-MS R_t(retention time): 1.87 minutes; MS: (ES) M/z 206(M + H)⁺)。

b) n-BuLi (1.47mL, 3.68mmol) was added to a solution of 3-tert-butylaniline (580mg, 3.89mmol) in anhydrous THF (2mL) at-78 deg.C under nitrogen and the solution was stirred at 0 deg.C for 10 min. The reaction mixture was cooled again to-78 ℃ and 2-cyclopentyl-nicotinic acid methyl ester (400mg, 1.94mmol) dissolved in anhydrous THF (2mL) was added thereto. The reaction mixture was brought to 0 ℃ over 2 hours to saturate the NH₄The reaction was quenched with aqueous Cl and extracted with ethyl acetate. The combined organic layers were dried (MgSO)₄) Filtered and concentrated under reduced pressure. By flash chromatography (SiO)₂10-100% EtOAc/hexanes) to afford the pure compound (91% yield, 57%)2mg)。LC-MS R_t(retention time): 2.61 minutes; MS: (ES) M/z 323(M + H)⁺)。

c) To a solution of N- (3-tert-butylphenyl) -2-cyclopentylnicotinamide (570mg, 1.77mmol) in ethanol (10mL) containing concentrated hydrochloric acid (1mL) was added platinum oxide (40mg, 0.17mmol), and the solution was hydrogenated using a Parr shaker at 40psi for 1.5 hours. The reaction mixture was filtered through celite and the filter cake was washed with ethanol. The filtrate was concentrated and the residue was dried under high vacuum for 2 hours to give the desired piperidine as the hydrochloride salt in quantitative yield. LC-MS R_t(retention time): 1.97 minutes; MS: (ES) M/z 329(M + H)⁺)。

d) To the above prepared cis-2-cyclopentylpiperidine-3-carboxylic acid (3-tert-butylphenyl) amide (123mg, 0.34mmol) in Et-containing solution₃N (142. mu.L, 1.02mmol) in anhydrous CH₂Cl₂To the solution in (1mL) was added 2-methylbenzoyl chloride (53mg, 0.34mmol), and the mixture was stirred at room temperature for 2 hours. The reaction mixture was then diluted with ethyl acetate (20mL) and washed with 1N aqueous HCl, water and brine. Drying (MgSO)₄) The organic layer was filtered and concentrated under reduced pressure. By reverse phase preparative HPLC (20-95% gradient of CH)₃CN-H₂O) the residue was purified and dried (lyophilizer) to give the title compound (yield 65%, 109 mg).¹H NMR(400MHz，CDCl₃)：δ1.22-1.48(m，11H)，1.56-1.80(m，5H)，1.84-2.06(m，4H)，2.10-2.23(m，1H)，2.30(s，1.6H)，2.39(s，1.4H)，2.41-2.50(m，1H)，2.71-2.76(m，1H)，3.02-3.09(m，1H)，3.25-3.39(m，1H)，5.11(bs，1H)，7.05-7.30(m，6H)，7.47-7.55(m，2H)，8.32(bs，1H)。LC-MS R_t(retention time): 3.16 minutes; MS: (ES) M/z 447(M + H)⁺. LC-MS method: agilent Zorbax SB-C18, 2.1X 50mm, 5. mu.g, 35 ℃, flow rate 1mL/min, gradient of 20% to 100% B for 2.5 min, wash for 1.0 min at 100% B; a ═ 0.1% formic acid/5% acetonitrile/94.9 water, B ═ 0.1% formic acid/5% water/94.9 acetonitrile.

Example 6

Synthesis of (2R, 3S) -2- (4-cyclopentylaminophenyl) -1- (2-methylbenzoyl) piperidine-3-carboxylic acid (3-chloro-4-methylphenyl) amide

b) In analogy to the procedure described in example 1, cis-2- (4-tert-butoxycarbonylaminophenyl) piperidine-3-carboxylic acid ethyl ester was synthesized.

c: 1) cis-2- (4-tert-Butoxycarbonylaminophenyl) piperidine-3-carboxylic acid ethyl ester (61g, 174.8mmol) and di-p-toluoyl-L-tartaric acid (62g, 174.8mmol) were dissolved in EtOH (500 ml). The clear solution was concentrated and sucked to dryness. The resulting white salt was then dissolved in 250ml ethyl acetate to form a clear solution. To this solution was slowly added 500ml of TBME. The resulting solution was left undisturbed at room temperature for 3 days. At this time, a large amount of white crystals were formed. It was then filtered and washed with 100ml TBME to give a white solid (60 g).

The above salt was redissolved in ethanol, concentrated and sucked to dryness. The resulting salt was dissolved in 500ml THF, followed by the addition of TBME (500 ml). The resulting clear solution was then left undisturbed at room temperature for 2.5 days. The resulting white crystals were filtered to give 20.5g (64: 1 enriched) of the salt.

c: 2) to the salt (16.7g) in CH at 0 deg.C₂Cl₂(150mL) to the stirred suspension saturated NaHCO was added₃Aqueous solution (100mL) and the reaction mixture was stirred at room temperature for 30 minutes. Separating the layers with CH₂Cl₂The aqueous layer was extracted (50 mL). The combined organic layers were washed with saturated NaHCO₃The aqueous solution (2X 100mL) was washed, dried and concentrated to give ethyl (2R, 3S) -2- (4-tert-butoxycarbonylaminophenyl) piperidine-3-carboxylate (90% yield, about 97% ee).

d) To the ethyl (2R, 3S) -2- (4-tert-butoxycarbonylaminophenyl) -piperidine-3-carboxylate (600) prepared abovemg, 1.72mmol) in Et₃N (480. mu.l, 3.44mmol) anhydrous CH₂Cl₂To a 0 ℃ solution in (5mL) was added 2-methylbenzoyl chloride (266mg, 1.72mmol) and the mixture was stirred at room temperature overnight. Then with CH₂Cl₂The reaction mixture was diluted (20mL) and washed with 1N aqueous HCl, water and brine. The organic layer was dried (MgSO₄) Filtered and concentrated under reduced pressure to give a quantitative yield of ethyl (2R, 3S) -2- (4-tert-butoxycarbonylaminophenyl) -1- (2-methylbenzoyl) piperidine-3-carboxylate and the crude product was used as such in the next step.

e) Reacting 4N HCl 1, 4-bisA solution of an alkane (5mL, 20mmol) was added slowly to the above crude product ethyl (2R, 3S) -2- (4-tert-butoxycarbonylaminophenyl) -1- (2-methylbenzoyl) piperidine-3-carboxylate (840mg, 1.72mmol) in dry CH₂Cl₂(4mL) in 0 ℃ solution. After addition of HCl, the reaction mixture was allowed to reach room temperature and stirred for 1 hour. Adding it with CH₂Cl₂(30mL), cooled to 0 deg.C and saturated NaHCO₃The aqueous solution was neutralized to give ethyl (2R, 3S) -2- (4-aminophenyl) -1- (2-methylbenzoyl) piperidine-3-carboxylate (612mg) in 97% yield over two steps. f) Reacting Na (OAC) at room temperature₃BH (495mg, 2.33mmol) was added to a solution of ethyl (2R, 3S) -2- (4-aminophenyl) -1- (2-methylbenzoyl) piperidine-3-carboxylate (612mg, 1.67mmol), cyclopentanone (140mg, 1.67mmol) and acetic acid (100mg, 1.67mmol) in anhydrous dichloroethane and the reaction mixture was heated to 50 ℃ for 4 hours, cooled to room temperature and stirred for 48 hours. Then adding it with CH₂Cl₂Diluted (30mL) with saturated NaHCO₃The aqueous solution was washed, dried and concentrated under vacuum. The residue was purified by ISCO flash column using ethyl acetate and hexane as mobile phases (40g column, 0-40% gradient) to give ethyl (2R, 3S) -2- (4-cyclopentylaminophenyl) -1- (2-methylbenzoyl) piperidine-3-carboxylate (450 mg).

g) At ambient temperature Me₃Al(290μl，0.57mmol, 2M in toluene) was added to a solution of 3-chloro-4-methylaniline (65mg, 0.46mmol) in dry dichloroethane (1 mL). After stirring for 20 minutes, (2R, 3S) -2- (4-cyclopentylaminophenyl) -1- (2-methylbenzoyl) piperidine-3-carboxylic acid ethyl ester (100mg, 0.23mmol) dissolved in anhydrous dichloroethane (1mL) was added thereto. The reaction mixture was then heated to 85 ℃ for 3 hours, cooled to room temperature, and charged with CH₂Cl₂Diluted (20mL) with saturated NaHCO₃And (4) washing with an aqueous solution. The aqueous layer was washed with CH₂Cl₂(20mL) extraction and drying of the combined organic layers (MgSO₄) And concentrated. By reverse phase preparative HPLC (20-95% gradient of CH)₃CN-H₂O, containing 0.1% TFA as additive) the residue was purified and the fractions containing the product were combined together and concentrated. With CH₂Cl₂The residue was diluted (30mL) with saturated NaHCO₃And (4) washing with an aqueous solution. Drying (MgSO)₄)CH₂Cl₂Layer and concentrated to give pure (2R, 3S) -2- (4-cyclopentylaminophenyl) -1- (2-methylbenzoyl) piperidine-3-carboxylic acid (3-chloro-4-methylphenyl) amide (50% yield).

¹H NMR(400MHz，CDCl₃)δ8.4(bs，1H)，7.55(s，1H)，7.37-7.05(m，9H)，6.55-6.52(m，2H)，3.77-3.70(m，1H)，3.30-3.16(m，1H)，3.04-2.91(m，2H)，2.43-1.94(m，8H)，1.71-1.46(m，11H)。

Example 7

The following are representative compounds prepared and evaluated using methods analogous to the examples herein. Characterization data for the compounds are provided below. Biological evaluation of these compounds and other compounds prepared as described herein is shown in figure 1.

(2R, 3S) -2- (4-Cyclopentylaminophenyl) -1- (2-fluoro-6-methylbenzoyl) piperidine-3-carboxylic acid (4-methyl-3-trifluoromethylphenyl) amide

¹H NMR(400MHz，TFA-d)δ7.91(d，J＝8.6Hz，1H)，7.84(d，J＝8.6Hz，1H)，7.58-6.82(m，8H)，6.75(t，J＝8.6Hz，1H)，4.10-4.00(m，1H)，3.60-3.47(m，1H)，3.45-3.41(m，1H)，3.33-3.25(m，1H)，2.44-2.22(m，7H)，2.04-1.92(m，4H)，1.82-.169(m，7H)。

(2R, 3S) -1- (2-chlorobenzoyl) -2- (4-cyclopentylaminophenyl) piperidine-3-carboxylic acid (4-methyl-3-trifluoromethylphenyl) amide

¹H NMR(400MHz，CDCl₃)δ9.41(bs，0.5H)，9.03(bs，0.5H)，7.55(s，1H)，7.49-7.39(m，3H)，7.31-7.27(m，2H)，7.18-7.04(m，2H)，6.83-6.74(m，3H)，3.76-3.64(m，1H)，3.22-2.90(m，5H)，2.39(s，3H)，2.32-2.20(m，1H)，2.16-2.04(m，1H)，2.0-1.86(m，2H)1.80-1.72(m，3H)，1.56(bs，5H)。

(2R, 3S) -2- (4-Cyclopentylaminophenyl) -1- (2-fluoro-6-methylbenzoyl) piperidine-3-carboxylic acid (3-chloro-4-methylphenyl) amide

¹H NMR(DMSO-d₆)δ10.22(s，1H)，7.67(dd，J＝1.8Hz，J＝11.0Hz，1H)，7.04-7.33(m，9H)，6.30(dd，J＝5.8Hz，J＝9.4Hz，1H)，5.52(br，1H)，3.56-3.64(m，1H)，3.00-3.17(m，2H)，2.90-2.98(m，1H)，2.23(2.24)(s，3H)，1.97(2.33)(s，3H)，1.32-2.22(m，12H)。

(2R, 3S) -1- (4-chlorobenzoyl) -2- (4-cyclopentylaminophenyl) piperidine-3-carboxylic acid (4-methyl-3-trifluoromethylphenyl) amide

¹H NMR(400MHz，CDCl₃)δ8.79(bs，1H)，7.62(s，1H)，7.52-7.48(m，1H)，7.37-7.30(m，5H)，7.13(d，J＝8.4Hz，1H)，6.52-6.50(m，3H)，3.75-3.69(m，1H)，3.44(bs，1H)，3.09-2.97(m，2H)，2.39(s，3H)，2.37-2.30(m，1H)，2.13-2.08(m，1H)，2.10-1.93(m，2H)，1.80-1.59(m，7H)，1.48-1.42(m，2H)。

(2R, 3S) -2- (4-Cyclohexylaminophenyl) -1- (2-fluoro-6-methylbenzoyl) piperidine-3-carboxylic acid (3-tert-butylphenyl) amide

¹H NMR(400MHz，CDCl₃)：δ8.24(m，1H)，7.40-6.85(m，8H)，6.65-6.40(m，3H)，3.57(s，1H)，3.30-2.90(m，4H)，2.50-1.85(m，9H)，1.80-1.50(m，5H)，1.40-1.00(m，13H)。

(2R, 3S) -2- (4-Cyclopentylaminophenyl) -1- (2-fluoro-6-methylbenzoyl) piperidine-3-carboxylic acid (4-methyl-3-pyrrolidin-1-yl-phenyl) amide

¹H NMR(400MHz，CDCl₃)：δ7.98(m，1H)，7.40-7.18(m，3H)，7.10-6.80(m，4H)，6.64-6.40(m，3H)，3.80-3.50(m，2H)，3.30-2.90(m，6H)，2.50-2.10(m，7H)，2.10-1.80(m，8H)，1.80-1.20(m，9H)。

(2R, 3S) -2- [4- (cyclopentyloxy) phenyl ] -1- (2-fluoro-6-methylbenzoyl) piperidine-3-carboxylic acid (3-chloro-4-methylphenyl) amide

¹H NMR(400MHz，CDCl₃)δ8.68(bs，0.6H)，8.58(bs，0.4H)，7.59-7.40(m，3H)，7.29-6.90(m，4H)，6.80(m，2H)，6.65(m，1H)，4.72(m，1H)，3.30-2.92(m，3H)，2.44(s，1H)，2.42-2.30(m，1H)，2.30(s，1H)，2.29(s，2H)，2.20(s，2H)，2.19-2.12(m，1H)，2.08-1.92(m，2H)，1.90-1.72(m，7H)1.60(m，2H)。

(±) - (2R, 3S) -2- (4-cyclopentylaminophenyl) -1- (2-fluoro-6-methylbenzoyl) piperidine-3-carboxylic acid (4-chloro-3-methylphenyl) amide

¹H NMR(400MHz，CDCl₃)δ8.25(bs，0.4H)，8.16(bs，0.6H)，7.44-7.20(m，6H)，7.06-6.84(m，2H)，6.59-6.50(m，2H)，3.75(m，1H)，3.66(bs，1H)，3.26-2.92(m，3H)，2.43(s，1H)，2.42-2.30(m，1H)，2.30(s，1H)，2.29(s，2H)，2.20(s，2H)，2.19-2.12(m，1H)，2.08-1.92(m，2H)，1.80-1.58(m，7H)1.45(m，2H)。

(2R, 3S) -2- (4-Cyclobutylaminophenyl) -1- (2-fluoro-6-methylbenzoyl) piperidine-3-carboxylic acid (3-tert-butylphenyl) amide

¹H NMR(400MHz，CDCl₃)δ8.20(s，0.6H)，8.39(s，0.4H)，7.44-6.88(m，10H)，6.25(dd，J＝12Hz，J＝6Hz，1H)，6.45(t，J＝8.4Hz，1H)，3.87(m，1H)，3.26-2.95(m，3H)，2.46-2.05(m，8H)，1.86-1.61(m，5H)，1.34-1.11(m，9H)。

(2R, 3S) -1- (2-fluoro-6-methylbenzoyl) -2- [4- (tetrahydropyran-4-ylamino) phenyl ] piperidine-3-carboxylic acid (3-morpholin-4-yl-phenyl) amide

¹H NMR(400MHz，CDCl₃)δ7.61(s，1H)，7.34-6.92(m，10H)，6.78-6.65(m，1H)，6.62-6.53(m，1H)，3.98-3.85(m，4H)，3.83-3.70(m，1H)，3.55-3.30(m，3H)，3.27-2.98(M，4H)，2.42-1.92(m，8H)，1.81-1.45(m，7H)。

(2R, 3S) -1- (2-fluoro-6-methylbenzoyl) -2- [4- ((R) -2-trifluoromethylpyrrolidin-1-ylmethyl) phenyl ] piperidine-3-carboxylic acid (3-tert-butylphenyl) amide

¹H NMR(400MHz，CDCl₃)δ8.01(bs，0.5H)，7.96(bs，0.5H)，7.55-7.37(m，3H)，7.30-7.19(m，6H)，7.13-7.06(m，1H)，7.01-6.90(m，1H)，6.85-6.64(m，1H)，4.15-4.11(m，1H)，3.58-3.54(m，1H)，3.30-3.20(m，2H)，3.17-2.80(m，2H)，2.45-2.17(m，4H)，2.00-1.94(m，2H)，1.86-1.60(m，8H)，1.31-1.26(m，7H)。

Example 8

Materials andmethod of producing a composite material

A. Cells

1.Cells expressing the C5a receptor

a) U937 cells

U937 cells are a C5aR expressing monocyte cell line available from atcc (va). These cells were cultured in suspension in RPMI-1640 medium supplemented with 2mM L-glutamine, 1.5g/L sodium bicarbonate, 4.5g/L glucose, 10mM HEPES, 1mM sodium pyruvate and 10% FBS. Growing cells on 5% CO₂Per 95% air, 100% humidity and 37 ℃ and passaged twice 1: 6 weekly (at 1X 10)⁵To 2X 10⁶Cells cultured at a density range of 1 × 10/mL) and⁶individual cells/mL were collected. Cells were treated with 0.5mM cAMP (Sigma, OH) overnight and washed once before use prior to assay. cAMP treated U937 cells can be used in C5aR ligand binding and function assays.

b) Isolated human neutrophils

Optionally, human or murine neutrophils may be used for compound activity assays. Neutrophils can be isolated from fresh human blood using density separation and centrifugation. Briefly, whole blood was incubated with equal portions of 3% dextran and allowed to separate for 45 minutes. After separation, the top layer was layered on top of 15ml of Ficoll (15ml of Ficoll per 30ml of blood suspension) and centrifuged at 400Xg for 30 minutes (no brake). The pellet at the bottom of the tube was then separated and resuspended in PharmLyse RBC lysis buffer (BD Biosciences, San Jose, Calif.), after which the sample was centrifuged again at 400Xg for 10 min (with brake). The remaining cell pellet is resuspended, if necessary, and consists of isolated neutrophils.

B.Measurement of

1.Inhibition of C5aR ligand binding

cAMP-treated U937 cells expressing C5aR were centrifuged and resuspended in assay buffer (20mM HEPES (pH 7.1), 140mM NaCl、1mM CaCl₂、5mM MgCl₂And containing 0.1% bovine serum albumin) to 3X 10⁶Concentration of individual cells/mL. Binding assays were set up as follows. 0.1mL of cells was added to an assay plate containing 5. mu.L of compound to give a final concentration of each screening compound (or compound IC) of about 2-10. mu.M₅₀A portion of the dose response measured). Then 0.1mL of a buffer diluted to a final concentration of about 50pM (resulting in about 30,000cpm per well) in assay buffer is added¹²⁵I-labeled C5a (obtained from Perkin Elmer Life Sciences, Boston, MA), plates were sealed and incubated on a shaker platform at 4 ℃ for about 3 hours. The reaction solution was pipetted onto a GF/B glass filter on a vacuum cell harvester (Packard Instruments; Meriden, CT) pre-soaked in a 0.3% Polyethyleneimine (PEI) solution. Scintillation fluid (40 μ l; Microscint 20, Packard Instruments) was added to each well, the plate sealed and the radioactivity measured in a Topcount scintillation counter (Packard Instruments). Control wells containing either diluent only (for total counts) or excess C5a (1 μ g/mL for non-specific binding) were used to calculate the percent total inhibition of the compound. IC was calculated using a computer program Prism from GraphPad, Inc. (San Diego, Ca)₅₀The value is obtained. IC (integrated circuit)₅₀Values are the concentration required to reduce binding of radiolabeled C5a to the receptor by 50%. (for further description of ligand binding and other functional assays, see Dairaghi et al, J.biol.chem.274: 21569-21574(1999), Penfold et al, Proc.Natl.Acad.Sci.USA.96: 9839-9844(1999), and Dairaghi et al, J.biol.chem.272: 28206-28209 (1997)).

2.Calcium mobilization

Optionally, the ability of the compound to inhibit calcium flux in a cell can be further determined. To detect the release of intracellular calcium stores, cells (e.g., cAMP-stimulated U937 OR neutrophils) were incubated with 3 μ M INDO-1AM dye (Molecular Probes; Eugene, OR) in cell culture medium for 45 minutes at room temperature and washed with Phosphate Buffered Saline (PBS). After loading of INDO-1AM, cells were resuspended in flux buffer (Henkel's Balanced salt solution (H)ank's saline solution, HBSS) and 1% FBS). Calcium mobilization was measured using a Photon Technology International spectrophotometer (Photon Technology International; New Jersey) (excitation at 350 nm) with simultaneous dual recording of fluorescence emissions at 400nm and 490 nm. The relative intracellular calcium levels are expressed as the 400nm/490nm emission ratio. At 37 ℃ in an absorption cell (each containing 10 in 2mL of flux buffer)⁶Individual cells) were run with constant mixing. Chemokine ligands in the range of 1 to 100nM may be used. The emission ratio is plotted as a function of time (typically 2-3 minutes). Candidate ligand blocking compounds (up to 10. mu.M) were added at 10 seconds, followed by chemokine (i.e., C5 a; R) at 60 seconds&DSystems; minneapolis, MN) and at 150 seconds the control chemokine (i.e., SDF-1 α; r&D Systems；Minneapolis，MN)。

3.Chemotaxis assay

Optionally, the ability of the compound to inhibit chemotaxis in cells can be further determined. Chemotaxis assays were performed using chemotaxis buffer (Henkel's Balanced salt solution (HBSS) and 1% FBS) in a 96-well chemotaxis chamber (Neuroprobe; Gaithersburg, Md.) using 5 μm-well polycarbonate, polyvinylpyrrolidone coated filters. C5aR ligand (i.e., C5a, R & DSystems; Minneapolis, MN) was used to evaluate compound-mediated inhibition of C5 aR-mediated migration. Other chemokines (i.e., SDF-1 α; R & D Systems; Minneapolis, MN) were used as specificity controls. The lower chamber was loaded with 29 μ l of chemokine (i.e., 0.03nM C5a) and varying amounts of compound; the upper chamber contained 100,000U 937 or neutrophils in 20 μ l. The chamber was incubated at 37 ℃ for 1.5 hours and the number of cells in the lower chamber was quantified either by direct cell counting in five high power zones per well or by CyQuant assays (Molecular Probes), a fluorescent staining method to measure nucleic acid content and microscopic observation.

Identification of C.C5aR inhibitors

1.Measurement of

To evaluate small organic molecules that prevent binding of the C5a receptor to the ligand, assays were used that detect binding of the radioligand (i.e., C5a) to cells that express C5aR on the cell surface (e.g., cAMP-stimulated U937 cells or isolated human neutrophils). For compounds that inhibit binding (whether competitive or non-competitive), a lower radioactive count is observed when compared to uninhibited controls.

Equal numbers of cells were added to each well in the plate. The cells were then incubated with radiolabeled C5 a. Unbound ligand is removed by washing the cells and bound ligand is determined by quantifying the radioactive counts. Cells not incubated with any organic compound were counted; nonspecific binding was determined by incubating cells with unlabeled ligand and labeled ligand. Percent inhibition was determined by the following equation:

2.dose response curve

To determine the affinity of candidate compounds for C5aR and to determine their ability to inhibit ligand binding, the affinity was determined at 1X 10^-10M to 1X 10^-4Titration inhibition activity over a range of compound concentrations of M. In the assay, the amount of compound varied; while the cell number and ligand concentration remained constant.

D. In vivo efficacy model

The potential efficacy of a compound in treating a C5 a-mediated disorder can be assessed by determining the efficacy of the compound of interest in an animal model. In addition to the models described below, other animal models suitable for studying the compound of interest can be found in Mizuno, M.et al, Expert Opin Investig.drugs (2005), 14(7), 807-821, which is incorporated herein by reference in its entirety.

Model of C5 a-induced leukopenia

a) C5 a-induced leukopenia in a mouse model knock-in with the human C5aR gene to investigate the efficacy of the compounds of the invention in animal models, standard techniques can be used to generate recombinant mice in which the genetic sequence encoding mouse C5aR is replaced with the sequence encoding human C5aR to generate hC5aR-KI mice. In this mouse, administration of hC5a resulted in upregulation of adhesion molecules on the vessel wall that bound blood leukocytes, thereby isolating blood leukocytes from the blood stream. Animals were administered 20 μ g/kg hC5a and leukocytes in peripheral blood were quantified by standard techniques after 1 minute. Pretreatment of mice with various doses of the compounds of the invention almost completely blocked hC5 a-induced leukopenia.

b) C5 a-induced leukopenia in macaque model

To study the efficacy of the compounds of the invention in non-human primate models, C5 a-induced leukopenia was studied in a cynomolgus monkey model. In this model, administration of hC5a resulted in upregulation of adhesion molecules on the vessel wall that bound blood leukocytes, thus isolating blood leukocytes from the blood stream. Animals were administered 10 μ g/kg hC5a and leukocytes in peripheral blood were quantified after 1 minute.

Mouse model of ANCA induced vasculitis

On day 0, hC5aR-KI mice were injected intravenously with 50mg/kg of purified antibody against myeloperoxidase (myeloperoxidase) (Xiao et al, J.Clin.invest.110: 955-963 (2002)). Mice were further given an oral daily dose of the compound of the invention or vehicle for seven days, after which the mice were sacrificed and the kidneys were collected for histological examination. Kidney section analysis may indicate: the number and severity of crescent and necrotic lesions in the glomeruli was significantly reduced compared to vehicle-treated animals.

2. Mouse model of choroidal neovascularization

To investigate the efficacy of the compounds of the invention in the treatment of age-related macular degeneration (AMD), brucella membrane (Bruchmembrane) was ruptured in the eyes of hC5aR-KI mice by laser photocoagulation (Nozika et al, PNAS 103: 2328-2333 (2006)). Mice are treated with the compounds of the invention either orally or by suitable intravitreal administration in a vehicle for one to two weeks. Repair of laser-induced damage and neovascularization were assessed by histology and angiography.

3. Rheumatoid arthritis model

a) Rabbit model of destructive arthritis

To investigate the effect of candidate compounds on inhibiting the inflammatory response of rabbits to intra-articular injection of the bacterial membrane fraction Lipopolysaccharide (LPS), a rabbit model of destructive joint inflammation was used. This study was designed to mimic the destructive inflammation of joints seen in arthritis. Intra-articular injection of LPS causes an acute inflammatory response characterized by the release of cytokines and chemokines, many of which have been identified in rheumatoid arthritis joints. In response to these increases in chemotactic mediators, a significant increase in leukocytes occurs in synovial fluid and membranes. Selective antagonists of chemokine receptors have shown efficacy in this model (see Podolin et al, J.Immunol.169 (11): 6435-6444 (2002)).

A rabbit LPS study was performed essentially as described in Podolin et al (above), female new zealand rabbits (approximately 2 kg) were treated intra-articularly in one knee with LPS (10ng) together with vehicle only (phosphate buffered saline with 1% DMSO) or with the addition of candidate compound (1 ═ 50 μ M dose or 2 ═ 100 μ M dose) (total volume 1.0 mL). Sixteen hours after LPS injection, knees were lavaged and cells were counted. The beneficial effects of treatment were determined by histopathological evaluation of synovial inflammation. Histopathological evaluation using inflammation scores: 1-minimal, 2-mild, 3-moderate, 4-moderate-significant.

b) Evaluation of compounds in a rat model of collagen-induced arthritis

A 17 day progressive collagen II arthritis study was performed to evaluate the effect of candidate compounds on arthritis-induced clinical ankle swelling. Rat collagen Arthritis is an experimental model of polyarthritis, which has been widely used in preclinical testing for a variety of anti-arthritic agents (see Trentham et al, J.exp. Med.146 (3): 857-868 (1977); Bendle et al, clinical Pathol.27: 134-142 (1999); Bendle et al, Arthritis Rheum.42: 498-506 (1999)). This model is characterized by a reliable and well-developed onset, easy measurement of polyarthritis, significant cartilage destruction associated with pannus formation, and mild to moderate bone resorption and periosteal bone hyperplasia.

On days 0 and 6 of this 17-day study, female lewis rats (approximately 0.2 kg) were anesthetized with isoflurane and injected with freund's incomplete adjuvant containing 2mg/mLII type bovine collagen at both sites on the base of the tail and back. The candidate compound is administered subcutaneously daily at an effective dose from day 0 until day 17. Ankle joint diameter was measured with calipers and joint swelling reduction was taken as a measure of efficacy.

4. Septicemia rat model

To investigate the effect of the compounds of interest on inhibiting the generalized inflammatory response associated with sepsis-like diseases, the Cecal Ligation and Puncture (CLP) rat model of sepsis was used. The rat CLP study was performed essentially as described in Fujimura N et al (American Journal respiratory clinical Care Medicine 2000; 161: 440-. Briefly, male and female Wister albino rats weighing between 200-250g were fasted for twelve hours prior to the experiment. Animals were kept on normal 12-hour light and dark cycles and fed standard rat chow until 12 hours prior to the experiment. The animals were then divided into four groups: (i) two sham groups and (ii) two CLP groups. Each of these two groups, i.e. (i) and (ii), was divided into a vehicle control group and a test compound group. Sepsis was induced by CLP method. Under transient anesthesia, midline laparotomy was performed with minimal dissection and the cecum was ligated with 3-0 wire just below the ileocecal valve to maintain intestinal continuity. The cecum was perforated with an 18-gauge needle on the mesenteric surface of the cecum at two locations 1cm apart and gently squeezed until fecal material was excreted. The intestine is then returned to the abdomen and the incision is closed. At the end of the surgery, all rats were resuscitated by subcutaneous administration of 3ml/100g body weight of saline. After surgery, rats were fasted, but water was freely available within the next 16 hours until they were sacrificed. The sham group was subjected to a laparotomy and the cecum was manipulated but not ligated or perforated. The beneficial effects of treatment are determined by histopathological scoring of tissues and organs and measuring several key indicators of liver function, kidney function and lipid peroxidation. To test for liver function, aspartate Aminotransferase (AST) and alanine Aminotransferase (ALT) were measured. Blood urea nitrogen and creatinine concentrations were studied to assess renal function. Serum levels of proinflammatory cytokines (e.g., TNF- α and IL-1 β) were also determined by ELISA.

5. Mouse SLE model of Experimental lupus nephritis

To study the effect of the compounds of interest on Systemic Lupus Erythematosus (SLE), the MRL/lpr mouse SLE model was used. MRL/Mp-Tfrsf 6^lpr/lprLine (MRL/lpr) is a commonly used mouse model of human SLE. To test the efficacy of the compounds in this model, male MRL/lpr mice were aliquoted at 13 weeks of age into a control group and a C5aR antagonist group. The compound or vehicle is then administered to the animal via an osmotic pump over the next 6 weeks to maintain the range of action and minimize the stress effects on the animal. Serum and urine samples were collected every two weeks during the six weeks of disease onset and progression. A minority of these mice develop glomerulosclerosis, which results in death of the animal from renal failure. With mortality as an indicator of renal failure being one of the measurement criteria, successful treatment will generally result in delayed onset of sudden death in the test group. In addition, Blood Urea Nitrogen (BUN) and proteinuria measurements can be used to continuously monitor the presence and level of renal disease. Tissues and organs were also collected at 19 weeks and subjected to histopathology and immunohistochemistry and scored based on tissue damage and cellular infiltration.

Rat model of COPD

The efficacy of compounds in Chronic Obstructive Pulmonary Disease (COPD) can be assessed using smoke-induced airway inflammation in rodent models. Selective antagonists of chemokines have been shown to have efficacy in this model (see Stevenson et al, am. J. physiol Lung Cell Mol physiol.288L514-L522, (2005)). An acute rat model of COPD was performed as described by Stevenson et al. The subject compound is administered systemically either orally or by Intravenous (IV) administration; or topically applied as a spray compound. Male Sprague-Dawley rats (350-. Rats were exposed for a total period of 32 minutes. Rats were sacrificed up to 7 days after the initial exposure. Any beneficial effects of treatment are assessed by decreased inflammatory cell infiltration, decreased levels of chemokines and cytokines. In the chronic model, mice or rats were subjected to daily tobacco smoke exposure for up to 12 months. The compounds are administered systemically by once daily oral administration, or possibly topically by spraying the compound. In addition to the inflammation observed for the acute model (Stevensen et al), animals may also show other pathologies similar to those seen in human COPD, such as emphysema (as indicated by the increase in mean linear intercept) and pulmonary chemistry alterations (see Martorana et al, am.J.Respir.Crit Care Med.172 (7): 848-53).

7. Mouse EAE model of multiple sclerosis

Experimental Autoimmune Encephalomyelitis (EAE) is a model of human multiple sclerosis. Variations of the model have been disclosed and are well known in the art. In a typical protocol, C57BL/6(Charles River Laboratories) mice were used in the EAE model. Mice were immunized on day 0 subcutaneously with 200 μ g Myelin Oligodendrocyte Glycoprotein (MOG) 35-55(Peptide International) emulsified in Complete Freund's Adjuvant (CFA) containing 4mg/ml mycobacterium tuberculosis (Sigma-Aldrich). Additionally, animals were given 200ng of pertussis toxin (Calbiochem) intravenously on days 0 and 2. Clinical scores were based on a scale of 0-5: 0, no signs of disease; 1, weakness of the tail; 2, hind limb weakness; 3, hind limb paralysis; 4, forelimb weakness or paralysis; and 5, dying. Administration of the compound of interest to be evaluated may be initiated on day 0 (prophylactic) or on day 7 (therapeutic, when there are histological signs of the disease, but few animals show clinical signs) and administered once or more times daily at a concentration suitable for its activity and pharmacokinetic properties (e.g. 100mg/kg, subcutaneously). Compound efficacy can be assessed by comparing severity (maximum mean clinical score in the presence of compound compared to vehicle) or by measuring a reduction in the number of macrophages isolated from the spinal cord (F4/80 positive). Spinal cord mononuclear cells can be isolated by a discontinuous Percoll gradient. Cells can be stained using rat anti-mouse F4/80-PE or rat IgG2b-PE (Caltag laboratories) and quantified by FACS analysis using 10. mu.l of beads (Polybeads) (Polysciences) per sample.

8. Mouse model for kidney transplantation

The transplantation model can be performed in mice, for example, an allogenic kidney transplantation model from C57BL/6 to BALB/C mice as described in Faikah Gueler et al, JASN Express, 2008, 8/27. Briefly, mice were anesthetized and the left donor kidney was connected to a cuff of the aorta and renal veins using a small vena cava cuff, and the ureters were removed en bloc. After a left nephrectomy is performed on the recipient, the vascular cuff is anastomosed to the recipient's abdominal aorta and vena cava, respectively, below the level of the native renal blood vessels. The ureter is directly anastomosed in the bladder. The cold ischemia time was 60 minutes, and the warm ischemia time was 30 minutes. The right native kidney can be removed at the time of allograft transplantation or at day 4 post-transplantation for long-term survival studies. General physical condition of the mice was monitored for signs of rejection. Treatment of animals with compounds can be initiated prior to surgery or immediately after transplantation, e.g., by once daily subcutaneous injection. Mice were studied for renal function and survival. Serum creatinine levels were measured by an automated method (Beckman Analyzer, Krefeld, Germany).

9. Mouse model of ischemia/reperfusion

Mouse models of ischemia/reperfusion injury can be found in Xiufen Zheng et al, am.J. Pathol, 173: volume 4, as described in 10 months 2008. Briefly, 6-8 week old CD1 mice were anesthetized and placed on a heating pad to remain warm during surgery. After abdominal incision, the renal pedicles are dissected with a blunt instrument and the microvascular clamp is placed on the left renal pedicle for 25-30 minutes. The clip was removed after ischemia and the right kidney incision was closed and the animal was allowed to recover. Blood was collected for serum creatinine and BUN analysis (as an indicator of kidney health). Or monitoring the survival of the animal over time. The compound can be administered to the animal before and/or after surgery and the effect on serum creatinine, BUN or animal survival can be used as an indicator of compound efficacy.

10. Mouse model of tumor growth

6-16 week-old C57BL/6 mice were injected subcutaneously on the right posterior flank or left posterior flank by 1X 10 injections⁵TC-1 cells (ATCC, VA). Mice were sacrificed starting at about 2 weeks after cell injection, every 2-4 days with caliper measurements of tumors until the desired tumor size was reached. At sacrifice, animals were subjected to a complete necropsy and spleens and tumors were removed. Excised tumors were measured and weighed. The compounds can be administered before and/or after tumor injection and the delay or inhibition of tumor growth used to assess compound efficacy.

Claims (36)

1. A compound having the formula

Or a pharmaceutically acceptable salt, hydrate or rotamer thereof; wherein

C¹Selected from phenyl, pyridyl, indolyl and thiazolyl, each of which is optionally substituted with 1 to 3R¹Substituent group substitution;

C²selected from phenyl, naphthyl, pyridyl and indolyl, each of which is optionally substituted with 1 to 3R²Substituent group substitution;

C³is selected from C_3-6Alkyl radical, C_3-6Cycloalkyl radical, C_3-6Cycloalkyl radical C_1-2Alkyl, phenyl, pyridyl, pyrazolyl, piperidinyl, pyrrolidinyl, piperidinylmethyl, and pyrrolidinylmethyl, each of which is optionally substituted with 1 to 3R³Substituent group substitution;

each R¹Independently selected from: halogen, -CN, -R^c、-CO₂R^a、-CONR^aR^b、-C(O)R^a、-OC(O)NR^aR^b、-NR^bC(O)R^a、-NR^bC(O)₂R^c、-NR^a-C(O)NR^aR^b、-NR^aC(O)NR^aR^b、-NR^aR^b、-OR^aand-S (O)₂NR^aR^b(ii) a Wherein R is^aAnd R^bEach independently selected from hydrogen and C_1-8Alkyl and C_1-8Haloalkyl, or when attached to the same nitrogen atom, can combine with that nitrogen atom to form a five-or six-membered ring having from 0 to 2 additional heteroatoms selected from N, O or S as ring members; each R^cIndependently selected from C_1-8Alkyl radical, C_1-8Haloalkyl, C_3-6Cycloalkyl, heterocycloalkyl, aryl and heteroaryl, and wherein R^a、R^bAnd R^cOptionally further substituted with one to three halogens, hydroxy, methyl, amino, alkylamino and dialkylamino; and optionally, when two R are¹When the substituents are on adjacent atoms, they combine to form a fused five-or six-membered carbocyclic ring;

each R²Independently selected from: halogen, -CN, -R^f、-CO₂R^d、-CONR^dR^e、-C(O)R^d、-OC(O)NR^dR^e、-NR^eC(O)R^d、-NR^eC(O)₂R^f、-NR^dC(O)NR^dR^e、-NR^dC(O)NR^dR^e、-NR^dR^e、-OR^dand-S (O)₂NR^dR^e(ii) a Wherein R is^dAnd R^eEach independently selected from hydrogen and C_1-8Alkyl and C_1-8Haloalkyl, or when attached to the same nitrogen atom, can combine with that nitrogen atom to form a five-or six-membered ring having from 0 to 2 additional heteroatoms selected from N, O or S as ring members; each R^fIndependently selected from C_1-8Alkyl radical, C_1-8Haloalkyl, C_3-6Cycloalkyl, heterocycloalkyl, aryl and heteroaryl, and wherein R^d、R^eAnd R^fOptionally further substituted with one to three halogens, hydroxy, methyl, amino, alkylamino and dialkylamino;

each R³Independently selected from: halogen, -CN, -Rⁱ、-CO₂R^g、-CONR^gR^h、-C(O)R^g、-OC(O)NR^gR^h、-NR^hC(O)R^g、-NR^hC(O)₂Rⁱ、-NR^gC(O)NR^gR^h、-NR^gR^h、-OR^g、-S(O)₂NR^gR^h、-X⁴-R^j、-X⁴-NR^gR^h、-X⁴-CONR^gR^h、-X⁴-NR^hC(O)R^g、-NHR^jand-NHCH₂R^jWherein X is⁴Is C_1-4An alkylene group; r^gAnd R^hEach independently selected from hydrogen and C_1-8Alkyl radical, C_3-6Cycloalkyl and C_1-8Haloalkyl, or when attached to the same nitrogen atom, can combine with that nitrogen atom to form a five-or six-membered ring having from 0 to 2 additional heteroatoms selected from N, O or S as ring members, and optionally substituted with one or two oxo groups; each RⁱIndependently selected from C_1-8Alkyl radical, C_1-8Haloalkyl, C_3-6Cycloalkyl, heterocycloalkyl, aryl, and heteroaryl; and each R^jIs selected from C_3-6Cycloalkyl, pyrrolinyl, piperidinyl, morpholinyl, tetrahydrofuranAnd tetrahydropyranyl, and wherein R^g、R^h、RⁱAnd R^jOptionally further substituted with one to three halogens, methyl, CF₃Hydroxy, amino, alkylamino and dialkylamino; and X is hydrogen or CH₃。

2. The compound of claim 1, wherein X is hydrogen.

3. The compound of claim 1, having the formula:

4. the compound of claim 1, having the formula:

5. the compound of claim 1, having the formula:

wherein

X¹Selected from N, CH and CR¹；

Subscript n is an integer of 0 to 2;

X²selected from N, CH and CR²(ii) a And is

Subscript m is an integer of 0 to 2.

6. The compound of claim 1, having the formula:

wherein

X¹Selected from N, CH and CR¹；

Subscript n is an integer of 0 to 2;

X²selected from N, CH and CR²(ii) a And is

Subscript m is an integer of 0 to 2.

7. The compound of claim 1, having the formula:

wherein

Subscript p is an integer of 0 to 3;

X¹selected from N, CH and CR¹；

Subscript n is an integer of 0 to 2;

X²selected from N, CH and CR²(ii) a And is

Subscript m is an integer of 0 to 2.

8. The compound of claim 1, having the formula:

9. the compound of claim 1, having the formula:

10. the compound of claim 1, having the formula:

11. the compound of claim 1, having the formula:

12. the compound of claim 1, having the formula:

13. the compound of claim 1, having the formula:

14. the compound of claim 1, having the formula:

wherein R is³Is selected from-NR^gR^h、-NHR^jand-NHCH₂R^j。

15. The compound of claim 1, having the formula:

wherein R is³Is selected from-X⁴-NR^gR^h、-X⁴-R^jand-X⁴-NR^hCOR^g。

16. The compound of claim 1, wherein each R¹Independently selected from halogen, -CN, -R^c、-NR^aR^band-OR^aAnd wherein R is^aAnd R^bEach independently selected from hydrogen and C_1-8Alkyl and C_1-8Haloalkyl groups, or when attached to the same nitrogen atom, may combine with the nitrogen atom to form a pyrrolidine ring; each R^cIndependently selected from C_1-8Alkyl radical, C_1-8Haloalkyl and C_3-6Cycloalkyl, and wherein R^a、R^bAnd R^cOptionally further substituted with one to three hydroxy, methyl, amino, alkylamino and dialkylamino groups; and optionally, when two R are¹When the substituents are on adjacent atoms, they combine to form a fused five-or six-membered carbocyclic ring.

17. The compound of claim 1, wherein each R²Independently selected from halogen, -R^fand-OR^d(ii) a Wherein each R^dIndependently selected from hydrogen, C_1-8Alkyl and C_1-8A haloalkyl group; each R^fIndependently selected from C_1-8Alkyl radical, C_1-8Haloalkyl, C_3-6Cycloalkyl, heterocycloalkyl and heteroaryl, and wherein R^dAnd R^fOptionally further substituted with one to three halogens, hydroxy, methyl, amino, alkylamino and dialkylamino.

18. The compound of claim 1, wherein each R³Independently selected from: halogen, -Rⁱ、-CO₂R^g、-CONR^gR^h、-NR^hC(O)R^g、-NR^hC(O)₂Rⁱ、-NR^gR^h、-OR^g、-X⁴-R^j、-X⁴-NR^gR^h、-X⁴-CONR^gR^h、-X⁴-NR^hC(O)R^g、-NHR^jand-NHCH₂R^jWherein X is⁴Is C_1-3An alkylene group; r^gAnd R^hEach independently selected from hydrogen and C_1-8Alkyl radical, C_3-6Cycloalkyl and C_1-8Haloalkyl, or when attached to the same nitrogen atom, can combine with that nitrogen atom to form a five-or six-membered ring having from 0 to 1 additional heteroatom selected from N, O or S as a ring member, and optionally substituted with one or two oxo groups; each RⁱIndependently selected from C_1-8Alkyl radical, C_1-8Haloalkyl, C_3-6Cycloalkyl, heterocycloalkyl, aryl, and heteroaryl; and each R^jIs selected from C_3-6Cycloalkyl, pyrrolinyl, piperidinyl, morpholinyl, tetrahydrofuryl and tetrahydropyranyl, and wherein R is^g、R^h、RⁱAnd R^jOptionally further substituted with one to three halogens, methyl, CF₃Hydroxy, amino, alkylamino and dialkylamino.

19. The compound of claim 17, wherein C²Selected from:

20. the compound of claim 16, wherein C¹Selected from:

21. the compound of claim 1, wherein C³Selected from:

22. the compound of claim 1, wherein C³Selected from:

23. the compound of claim 1, wherein the compound is selected from the group consisting of:

24. a pharmaceutical composition comprising a pharmaceutically acceptable carrier and a compound of claim 1.

25. The use of a compound of claim 1 in the manufacture of a medicament for the therapeutic or prophylactic treatment of a disease or condition in a mammal involving pathological activation of the C5a receptor.

26. The use of a compound of claim 1 in the manufacture of a medicament for inhibiting C5a receptor mediated cell chemotaxis.

27. The use of claim 25, wherein the disease or disorder is an inflammatory disease or disorder.

28. The use of claim 27, wherein the disease or condition is selected from the group consisting of: neutropenia, sepsis, septic shock, alzheimer's disease, multiple sclerosis, stroke, inflammatory bowel disease, chronic obstructive pulmonary disease, inflammation associated with burns, lung injury, osteoarthritis, atopic dermatitis, chronic urticaria, ischemia reperfusion injury, acute respiratory distress syndrome, systemic inflammatory response syndrome, multiple organ dysfunction syndrome, tissue transplant rejection, and hyperacute rejection of transplanted organs.

29. The use of claim 25, wherein the disease or disorder is a cardiovascular or cerebrovascular disorder.

30. The use of claim 29, wherein the disease or condition is selected from the group consisting of myocardial infarction, coronary artery embolism, vascular occlusion, reocclusion of blood vessels following surgical procedures, atherosclerosis, traumatic central nervous system injury, and ischemic heart disease.

31. The use of claim 25, wherein the disease or disorder is an autoimmune disorder.

32. The use of claim 31, wherein the disease or condition is selected from the group consisting of: rheumatoid arthritis, systemic lupus erythematosus, Guillain-Barre syndrome, pancreatitis, lupus nephritis, lupus glomerulonephritis, psoriasis, Crohn's disease, vasculitis, irritable bowel syndrome, dermatomyositis, multiple sclerosis, bronchial asthma, pemphigus, pemphigoid, scleroderma, myasthenia gravis, autoimmune hemolysis and thrombocytopenia, Goodpasture's syndrome, immune vasculitis, tissue transplant rejection and hyperacute rejection of transplanted organs.

33. The use of claim 25, wherein the disease or condition is a pathological sequelae associated with a condition selected from the group consisting of: insulin-dependent diabetes mellitus, lupus nephropathy, Heimanychis, membranous nephritis, glomerulonephritis, contact hypersensitivity, and inflammation caused by blood contact with artificial surfaces.

34. The compound of claim 1, wherein said compound is

Or a pharmaceutically acceptable salt thereof.

35. The compound of claim 1, wherein said compound is

Or a pharmaceutically acceptable salt thereof.

36. The compound of claim 1, wherein said compound is

Or a pharmaceutically acceptable salt thereof.