WO2009029625A1

WO2009029625A1 - 4- [heterocyclyl-methyl] -8-fluoro-quinolin-2-ones useful as nitric oxide synthase inhibitors

Info

Publication number: WO2009029625A1
Application number: PCT/US2008/074334
Authority: WO
Inventors: Nicholas D. Smith; Celine Bonnefous; Sergio Duron
Original assignee: Kalypsys Inc
Current assignee: Kalypsys Inc
Priority date: 2007-08-27
Filing date: 2008-08-26
Publication date: 2009-03-05
Anticipated expiration: 2010-02-27

Abstract

Novel compounds of formulae (II, III) and pharmaceutical compositions have been found to inhibit inducible NOS synthase wherein: R4, R5, R6 and R7 are independently selected from the group consisting of hydrogen, lower alkyl, and halogen; and, R8 has the structure whrein X1, X2, X3, X4, X5, X6, R9, R13, R14 and n are as described herein.

Description

Attorney Docket No. KO 152-401 -PC

4- [HETEROCYCLYL-METHYL]-8-FLUORO-QUINOLIN-2-ONES USEFUL AS NITRIC OXIDE SYNTHASE INHIBITORS

[001] This application claims the benefit of priority of United States provisional application No. 60/968,177, filed August 27, 2007, the disclosure of which is hereby incorporated by reference as if written herein in its entirety.

[002] Disclosed herein are new heterobicyclic-substituted quinolone compounds and compositions and their application as pharmaceuticals for the treatment of disease. Methods of inhibition of nitric oxide synthase activity in a human or animal subject are also provided for the treatment disease.

[003] Nitric oxide (NO) is involved in the regulation of many physiological processes as well as the pathophysiology of a number of diseases. It is synthesized enzymatically from L-arginine in numerous tissues and cell types by three distinct isoforms of the enzyme NO synthase (NOS). Two of these isoforms, endothelial NOS (eNOS) and neuronal NOS (nNOS) are expressed in a constitutive manner and are calcium/calmodulin dependent. Endothelial NOS is expressed by endothelium and other cell types and is involved in cardiovascular homeostasis. Neuronal NOS is constitutively present in both the central and peripheral nervous system where NO acts a neurotransmitter. Under normal physiological conditions, these constitutive forms of NOS generate low, transient levels of NO in response to increases in intracellular calcium concentrations. These low levels of NO act to regulate blood pressure, platelet adhesion, gastrointestinal motility, bronchomotor tone and neurotransmission. [004] In contrast, the third isoform of NOS, inducible NOS (iNOS), a virtually calcium independent enzyme, is absent in resting cells, but is rapidly expressed in virtually all nucleated mammalian cells in response to stimuli such as endotoxins and/or cytokines. The inducible isoform is neither stimulated by calcium nor blocked by calmodulin antagonists. It contains several tightly bound co-factors, including FMN, FAD and tetrahydrobiopterin. The inducible isoform of nitric oxide synthase Attorney Docket No. KO 152-401 -PC

(NOS₂ or iNOS) is expressed in virtually all nucleated mammalian cells following exposure to inflammatory cytokines or lipopolysaccharide.

[005] The enzyme iNOS synthase is a homodimer composed of 130 kDa subunits. Each subunit comprises an oxygenase domain and a reductase domain. Importantly, dimerization of the iNOS synthase is required for enzyme activity. If the dimerization mechanism is disrupted, the production of nitric oxide via inducible NOS enzyme is inhibited.

[006] The presence of iNOS in macrophages and lung epithelial cells is significant. Once present, iNOS synthesizes 100-1000 times more NO than the constitutive enzymes synthesize and does so for prolonged periods. This excessive production of NO and resulting NO-derived metabolites (e.g., peroxynitrite) elicit cellular toxicity and tissue damage which contribute to the pathophysiology of a number of diseases, disorders and conditions.

[007] Nitric oxide generated by the inducible form of NOS has also been implicated in the pathogenesis of inflammatory diseases. In experimental animals, hypotension induced by lipopolysaccharide or tumor necrosis factor alpha can be reversed by NOS inhibitors. Conditions which lead to cytokine-induced hypotension include septic shock, hemodialysis and interleukin therapy in cancer patients. An iNOS inhibitor has been shown to be effective in treating cytokine-induced hypotension, inflammatory bowel disease, cerebral ischemia, osteoarthritis, asthma and neuropathies such as diabetic neuropathy and post-herpetic neuralgia. [008] In addition, nitric oxide localized in high amounts in inflamed tissues has been shown to induce pain locally and to enhance central as well as peripheral stimuli. Because nitric oxide produced by an inflammatory response is thought to be synthesized by iNOS, the inhibition of iNOS dimerization produces both prophylactic and remedial analgesia in patients. Nitric oxide and NOS activity and expression have been linked to neuropathic pain and postherpetic neuralgia, as well, conditions for which treatment options have been equivocal at best(Wu CL and Raja SN, J Pain. 2008 Jan;9(l Suppl l):S19-30). Inhibitors of iNOS show promise in the treatment of these debilitating diseases. By way of example, several recent studies in rats administered an iNOS inhibitor demonstrate a role for peripherally-expressed iNOS in Attorney Docket No. KO 152-401 -PC

pain conditions with an inflammatory component and the potential value of iNOS inhibitors in such conditions. In one, rats administered iNOS inhibitor GW274150 24 hours after Freund's Complete Adjuvant (FCA) injection in the hind paw demonstrated suppressed accumulation of nitrite in the inflamed paw indicating substantial iNOS inhibition and attenuated hypersensitivity to pain and edema in a dose-dependent manner (De Alba J et al, J Pain. 2006 Jan; 120(1 -2): 170-81). In another, evidence for local NOS expression and NO action in the chronic constriction injury (CCI) model of neuropathic pain was demonstrated (Levy D and Zochodne DW, Eur J Neurosci. 1998 May; 10 (5):1846-55), and administration of GW274150 following chronic constrictive injury surgery reversed significantly the CCI-associated hypersensitivity to pain (De Alba J et al., ] Pain. 2006 Jan; 120 (1-2): 170-81).

[009] INOS-selective and nonselective inhibitors have long been known in the art, and have been investigated for the treatment of iNOS-mediated diseases and conditions including pain, as described above. In situations where the overproduction of nitric oxide is deleterious, it makes logical sense to pursue an inhibitor of iNOS to reduce the production of NO, thereby relieving inflammation, pain, and other NO-mediated diseases. To date, however, no NOS inhibitor has made it to market, for reasons that will not be exhaustively addressed here. The earliest compounds tended to fall in to the category of active-site inhibitors, leading to unacceptable side effects. As the important physiological roles played by the constitutive NOS isoforms, particularly eNOS, became more clear, later compounds were designed to have isoform selectivity to ensure that the inhibition of iNOS has the least possible effect on the activity of eNOS. Other agents failed to have pharmacokinetic properties suitable for orally- administered agents that will have good patient compliance. What is clear is that a need still exists for a selective, effective, pharmacologically appropriate inhibitor of iNOS.

[010] Novel compounds and pharmaceutical compositions, certain of which have been found to inhibit inducible NOS synthase have been discovered, together with methods of synthesizing and using the compounds including methods for the treatment of iNOS-mediated diseases in a patient by administering the compounds. Certain of Attorney Docket No. KO 152-401 -PC

these compounds have additionally demonstrated efficacy in vivo as anti-inflammatory agents. Finally, certain compounds disclosed herein exhibit good metabolic stability. [Oi l] In certain embodiments of the present invention, compounds have structural Formula I:

wherein:

R¹ is selected from the group consisting of acyl, alkyl, alkylene, aminoalkyl, amidoalkyl, alkynyl, amido, amino, aminoalkyl, aryl, arylalkyl, arylalkoxy, arylamino, arylaminoalkyl, arylthio, carboxy, cycloalkyl, ester, ether, halo, haloalkoxy, haloalkyl, heteroaryl, heteroarylalkyl, heteroarylamino, heteroarylaminoalkyl, heterocycloalkyl, heterocycloalkylalkyl, hydrazinyl, hydrogen, imino, thio, sulfonate, sulfonylamino and sulfonylaminoalkyl, any of which may be optionally substituted;

R² is selected from the group consisting of acyl, alkoxy, alkoxyalkyl, alkyl, alkylene, alkylamino, alkynyl, alkylimino, amido, amino, aryl, carboxy, cyano, cycloalkyl, ester, halo, haloalkyl, heteroaryl, heterocycloalkyl and hydrogen, any of which may be optionally substituted; or, alternatively, R² may combine with R¹ to form heterocycloalkyl, which may be optionally substituted;

R³ is selected from the group consisting of alkyl, amino, arylalkyl, aryl, cycloalkyl, haloalkyl, heteroarylalkyl, heterocycloalkyl and hydrogen, any of which may be optionally substituted; and

A, B, C and D are each independently selected from the group consisting of acyl, alkoxy, alkyl, alkylene, alkylamino, alkynyl, amido, amino, aminosulfonyl, aryl, arylalkoxy, arylamino, arylthio, carboxy, cycloalkyl, ester, ether, halo, haloalkoxy, haloalkyl, heteroaryl, heteroarylamino, heterocycloalkyl, hydrazinyl, hydrogen, imino, thio, sulfonate and sulfonylamino, any of which may be optionally substituted; or, alternatively, any two or more A, B, C and D may combine to form aryl, cycloalkyl, heteroaryl or heterocycloalkyl, any of which may be optionally substituted. Attorney Docket No. KO 152-401 -PC

[012] Certain compounds disclosed herein may possess useful iNOS inhibitory activity, and may be used in the treatment or prophylaxis of a disease or condition in which iNOS plays an active role. Thus, in broad aspect, the certain embodiments also provide pharmaceutical compositions comprising one or more compounds disclosed herein together with a pharmaceutically acceptable carrier, as well as methods of making and using the compounds and compositions. Certain embodiments provide methods for inhibiting iNOS. Other embodiments provide methods for treating an iNOS-mediated disorder in a patient in need of such treatment, comprising administering to said patient a therapeutically effective amount of a compound or composition according to the present invention. Also provided is the use of certain compounds disclosed herein for use in the manufacture of a medicament for the treatment of a disease or condition ameliorated by the inhibition of iNOS. [013] In certain embodiments, the compounds have structural Formula II:

wherein:

R⁴, R⁵, R⁶ and R⁷ are independently selected from the group consisting of hydrogen, lower alkyl, and halogen; and

R⁸ is selected from the group consisting of heteroaryl and heterocycloalkyl, either of which may be optionally substituted. [014] In further embodiments, R⁶ and R⁷ are hydrogen. [015] In further embodiments, R⁴ is hydrogen.

[016] In further embodiments, R⁸ is selected from the group consisting of lower heteroaryl and lower heterocycloalkyl, either of which may be optionally substituted. [017] In further embodiments, R⁸ is selected from the group consisting of lower heteroaryl and lower heterocycloalkyl, either of which may be optionally substituted with one to three substituents selected from the group consisting of hydrogen, halogen, lower alkyl, lower monocyclic cycloalkyl, lower heteroalkyl, lower haloalkyl, lower amino, and lower alkoxy. Attorney Docket No. KO 152-401 -PC

[018] In further embodiments, R⁸ has the structure:

wherein at least one of X¹ - X⁶ is a heteroatom;

X¹, X², and X⁵ are each independently selected from the group consisting of C and N;

X³ is selected from the group consisting of C(R¹⁰) and N;

X⁴ is selected from the group consisting of C(R¹¹) and N

X⁶ is selected from the group consisting of C(R¹²), C(R¹²)-C(R¹³), C=C, C(R¹²)- N, N, O, and S;

R⁹ is selected from the group consisting of hydrogen, halogen, lower alkyl, and lower monocyclic cycloalkyl; or R⁹ may combine with X⁶ to form a cyclic moiety selected from the group consisting of aryl and heteroaryl; and

R¹⁰, R¹¹, and R¹² are each independently selected from the group consisting of a bond, hydrogen, halogen, lower alkyl, lower alkoxy, and lower monocyclic cycloalkyl; each R¹³ and R¹⁴ is independently selected from the group consisting of hydrogen, halogen, lower alkyl, and lower monocyclic cycloalkyl; and each n is independently 0 or 1 ; and with the provisos that: the fused ring formed by X¹, X², X⁵, X⁶ , and the carbon bearing R⁹ may be aromatic or aliphatic; and

X¹ and X⁶ may not both be N. Attorney Docket No. KO 152-401 -PC

[019] In yet further embodiments, R has the structure:

[020] In further embodiments, R⁸ has a structure selected from the group consisting of:

wherein R¹⁵ is selected from the group consisting of hydrogen, halogen, lower alkyl, and lower monocyclic cycloalkyl, and all other groups are as previously defined. Attorney Docket No. KO 152-401 -PC

[021] In certain embodiments, the compounds have structural Formula III:

wherein at least one of X¹ - X⁶ is a heteroatom;

X³ is selected from the group consisting of C(R¹⁰) and N;

X⁴ is selected from the group consisting of C(R¹¹) and N

X⁶ is selected from the group consisting of C(R¹²), C(R¹²)- C(R¹³), C=C, N, O, and S;

R⁴ and R⁵ are each independently selected from the group consisting of hydrogen, lower alkyl, and halogen;

R⁹ is selected from the group consisting of hydrogen, halogen, lower alkyl, and lower monocyclic cycloalkyl; or R⁹ may combine with X⁶ to form a cyclic moiety selected from the group consisting of aryl and heteroaryl;

R¹⁰, R¹¹, and R¹² are each independently selected from the group consisting of a bond, hydrogen, halogen, lower alkyl, lower alkoxy, and lower monocyclic cycloalkyl; each R¹³ and R¹⁴ is independently selected from the group consisting of hydrogen, halogen, lower alkyl, and lower monocyclic cycloalkyl; and each n is independently 0 or 1 ; and with the proviso that X¹ and X⁶ may not both be N. [022] In further embodiments, R⁴ is hydrogen. [023] In further embodiments, R⁵ is fluorine.

[024] In further embodiments is provided a compound selected from the group consisting of Examples 1 to 88 and 90-95. Attorney Docket No. KO 152-401 -PC

[025] In further embodiments is provided the use of a compound as recited in

Claim 1 as a medicament.

[026] In further embodiments is provided the use of a compound as recited in

Claim 1 as a medicament for the prevention or treatment of a disease or condition ameliorated by the inhibition of iNOS.

[027] Further provided is a pharmaceutical composition comprising a compound as recited in Claim 1 together with a pharmaceutically acceptable carrier.

[028] Further provided is a method of inhibition of iNOS comprising contacting iNOS with a compound as recited in Claim 1.

[029] Further provided is a method of treatment of a iNOS-mediated disease comprising the administration of a therapeutically effective amount of a compound as recited in Claim 1 to a patient in need thereof.

[030] Further provided is a method of treatment of a iNOS-mediated disease comprising the administration of: a therapeutically effective amount of a compound as recited in Claim 1; and another therapeutic agent.

[031] As used herein, the terms below have the meanings indicated. [032] When ranges of values are disclosed, and the notation "from ni ... to n₂" is used, where ni and n₂ are the numbers, then unless otherwise specified, this notation is intended to include the numbers themselves and the range between them. This range may be integral or continuous between and including the end values. By way of example, the range "from 2 to 6 carbons" is intended to include two, three, four, five, and six carbons, since carbons come in integer units. Compare, by way of example, the range "from 1 to 3 μM (micromolar)," which is intended to include 1 μM, 3 μM, and everything in between to any number of significant figures (e.g., 1.255 μM, 2.1 μM, 2.9999 μM, etc.).

[033] The term "about," as used herein, is intended to qualify the numerical values which it modifies, denoting such a value as variable within a margin of error. When no particular margin of error, such as a standard deviation to a mean value given in a chart or table of data, is recited, the term "about" should be understood to mean that range which would encompass the recited value and the range which would be Attorney Docket No. KO 152-401 -PC

included by rounding up or down to that figure as well, taking into account significant figures.

[034] The term "acyl," as used herein, alone or in combination, refers to a carbonyl attached to an alkenyl, alkyl, aryl, cycloalkyl, heteroaryl, heterocycle, or any other moiety were the atom attached to the carbonyl is carbon. An "acetyl" group refers to a -C(O)CH₃ group. An "alkylcarbonyl" or "alkanoyl" group refers to an alkyl group attached to the parent molecular moiety through a carbonyl group. Examples of such groups include methylcarbonyl and ethylcarbonyl. Examples of acyl groups include formyl, alkanoyl and aroyl.

[035] The term "alkenyl," as used herein, alone or in combination, refers to a straight-chain or branched-chain hydrocarbon group having one or more double bonds and containing from 2 to 20 carbon atoms. In certain embodiments, said alkenyl will comprise from 2 to 6 carbon atoms. The term "alkenylene" refers to a carbon-carbon double bond system attached at two or more positions such as ethenylene [(-CH=CH- ),(-C::C-)]. Examples of suitable alkenyl groups include ethenyl, propenyl, 2- methylpropenyl, 1 ,4-butadienyl and the like. Unless otherwise specified, the term "alkenyl" may include "alkenylene" groups.

[036] The term "alkoxy," as used herein, alone or in combination, refers to an alkyl ether group, wherein the term alkyl is as defined below. Examples of suitable alkyl ether groups include methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, iso- butoxy, sec-butoxy, tert-butoxy, and the like.

[037] The term "alkyl," as used herein, alone or in combination, refers to a straight-chain or branched-chain alkyl group containing from 1 to 20 carbon atoms. In certain embodiments, said alkyl will comprise from 1 to 10 carbon atoms. In further embodiments, said alkyl will comprise from 1 to 6 carbon atoms. Alkyl groups may be optionally substituted as defined herein. Examples of alkyl groups include methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, pentyl, iso-amyl, hexyl, octyl, noyl and the like. The term "alkylene," as used herein, alone or in combination, refers to a saturated aliphatic group derived from a straight or branched chain saturated hydrocarbon attached at two or more positions, such as methylene (- CH₂-). Unless otherwise specified, the term "alkyl" may include "alkylene" groups. Attorney Docket No. KO 152-401 -PC

[038] The term "alkylamino," as used herein, alone or in combination, refers to an alkyl group attached to the parent molecular moiety through an amino group. Suitable alkylamino groups may be mono- or dialkylated, forming groups such as, for example, N-methylamino, N-ethylamino, N,N-dimethylamino, N,N-ethylmethylamino and the like.

[039] The term "alkylidene," as used herein, alone or in combination, refers to an alkenyl group in which one carbon atom of the carbon-carbon double bond belongs to the moiety to which the alkenyl group is attached.

[040] The term "alkylthio," as used herein, alone or in combination, refers to an alkyl thioether (R-S-) group wherein the term alkyl is as defined above and wherein the sulfur may be singly or doubly oxidized. Examples of suitable alkyl thioether groups include methylthio, ethylthio, n-propylthio, isopropylthio, n-butylthio, iso- butylthio, sec-butylthio, tert-butylthio, methanesulfonyl, ethanesulfmyl, and the like. [041] The term "alkynyl," as used herein, alone or in combination, refers to a straight-chain or branched chain hydrocarbon group having one or more triple bonds and containing from 2 to 20 carbon atoms. In certain embodiments, said alkynyl comprises from 2 to 6 carbon atoms. In further embodiments, said alkynyl comprises from 2 to 4 carbon atoms. The term "alkynylene" refers to a carbon-carbon triple bond attached at two positions such as ethynylene (-C:::C-, -C≡C-). Examples of alkynyl groups include ethynyl, propynyl, hydroxypropynyl, butyn-1-yl, butyn-2-yl, pentyn-1- yl, 3-methylbutyn-l-yl, hexyn-2-yl, and the like. Unless otherwise specified, the term "alkynyl" may include "alkynylene" groups.

[042] The terms "amido" and "carbamoyl," as used herein, alone or in combination, refer to an amino group as described below attached to the parent molecular moiety through a carbonyl group, or vice versa. The term "C-amido" as used herein, alone or in combination, refers to a -Q=O)-NR₂ group with R as defined herein. The term "N-amido" as used herein, alone or in combination, refers to a RC(=O)NH- group, with R as defined herein. The term "acylamino" as used herein, alone or in combination, embraces an acyl group attached to the parent moiety through an amino group. An example of an "acylamino" group is acetylamino (CHsC(O)NH-). Attorney Docket No. KO 152-401 -PC

[043] The term "amino," as used herein, alone or in combination, refers to —

NRR , wherein R and R are independently selected from the group consisting of hydrogen, alkyl, acyl, heteroalkyl, aryl, cycloalkyl, heteroaryl, and heterocycloalkyl, any of which may themselves be optionally substituted. Additionally, R and R' may combine to form heterocycloalkyl, either of which may be optionally substituted.

[044] The term "aryl," as used herein, alone or in combination, means a carbocyclic aromatic system containing one, two or three rings wherein such polycyclic ring systems are fused together. The term "aryl" embraces aromatic groups such as phenyl, naphthyl, anthracenyl, and phenanthryl.

[045] The term "arylalkenyl" or "aralkenyl," as used herein, alone or in combination, refers to an aryl group attached to the parent molecular moiety through an alkenyl group.

[046] The term "arylalkoxy" or "aralkoxy," as used herein, alone or in combination, refers to an aryl group attached to the parent molecular moiety through an alkoxy group.

[047] The term "arylalkyl" or "aralkyl," as used herein, alone or in combination, refers to an aryl group attached to the parent molecular moiety through an alkyl group.

[048] The term "arylalkynyl" or "aralkynyl," as used herein, alone or in combination, refers to an aryl group attached to the parent molecular moiety through an alkynyl group.

[049] The term "arylalkanoyl" or "aralkanoyl" or "aroyl," as used herein, alone or in combination, refers to an acyl group derived from an aryl-substituted alkanecarboxylic acid such as benzoyl, naphthyl, phenylacetyl, 3-phenylpropionyl

(hydrocinnamoyl), 4-phenylbutyryl, (2-naphthyl)acetyl, 4-chlorohydrocinnamoyl, and the like.

[050] The term aryloxy as used herein, alone or in combination, refers to an aryl group attached to the parent molecular moiety through an oxy.

[051] The terms "benzo" and "benz," as used herein, alone or in combination, refer to the divalent group C₆H₄= derived from benzene. Examples include benzo thiophene and benzimidazole. Attorney Docket No. KO 152-401 -PC

[052] The term "carbamate," as used herein, alone or in combination, refers to an ester of carbamic acid (-NHCOO-) which may be attached to the parent molecular moiety from either the nitrogen or acid end, and which may be optionally substituted as defined herein.

[053] The term "O-carbamyl" as used herein, alone or in combination, refers to a

-OC(O)NRR', group with R and R' as defined herein.

[054] The term "N-carbamyl" as used herein, alone or in combination, refers to a

ROC(O)NR'- group, with R and R' as defined herein.

[055] The term "carbonyl," as used herein, when alone includes formyl [-C(O)H] and in combination is a -C(O)- group.

[056] The term "carboxyl" or "carboxy," as used herein, refers to -C(O)OH or the corresponding "carboxylate" anion, such as is in a carboxylic acid salt. An

"O-carboxy" group refers to a RC(O)O- group, where R is as defined herein. A

"C-carboxy" group refers to a -C(O)OR groups where R is as defined herein.

[057] The term "cyano," as used herein, alone or in combination, refers to -CN.

[058] The term "cycloalkyl," or, alternatively, "carbocycle," as used herein, alone or in combination, refers to a saturated or partially saturated monocyclic, bicyclic or tricyclic alkyl group wherein each cyclic moiety contains from 3 to 12 carbon atom ring members and which may optionally be a benzo fused ring system which is optionally substituted as defined herein. In certain embodiments, said cycloalkyl will comprise from 5 to 7 carbon atoms. Examples of such cycloalkyl groups include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, tetrahydronaphthyl, indanyl, octahydronaphthyl, 2,3-dihydro-lH-indenyl, adamantyl and the like.

"Bicyclic" and "tricyclic" as used herein are intended to include both fused ring systems, such as decahydronaphthalene, octahydronaphthalene as well as the multicyclic (multicentered) saturated or partially unsaturated type. The latter type of isomer is exemplified in general by, bicyclo[l,l,l]pentane, camphor, adamantane, and bicyclo[3,2,l]octane.

[059] The term "ester," as used herein, alone or in combination, refers to a carboxy group bridging two moieties linked at carbon atoms. Attorney Docket No. KO 152-401 -PC

[060] The term "ether," as used herein, alone or in combination, refers to an oxy group bridging two moieties linked at carbon atoms.

[061] The term "halo," or "halogen," as used herein, alone or in combination, refers to fluorine, chlorine, bromine, or iodine.

[062] The term "haloalkoxy," as used herein, alone or in combination, refers to a haloalkyl group attached to the parent molecular moiety through an oxygen atom. [063] The term "haloalkyl," as used herein, alone or in combination, refers to an alkyl group having the meaning as defined above wherein one or more hydrogens are replaced with a halogen. Specifically embraced are monohaloalkyl, dihaloalkyl and polyhaloalkyl groups. A monohaloalkyl group, for one example, may have an iodo, bromo, chloro or fluoro atom within the group. Dihalo and polyhaloalkyl groups may have two or more of the same halo atoms or a combination of different halo groups. Examples of haloalkyl groups include fluoromethyl, difluoromethyl, trifluoromethyl, chloromethyl, dichloromethyl, trichloromethyl, pentafluoroethyl, heptafluoropropyl, difluorochloromethyl, dichlorofluoromethyl, difluoroethyl, difluoropropyl, dichloroethyl and dichloropropyl. "Haloalkylene" refers to a haloalkyl group attached at two or more positions. Examples include fluoromethylene (-CFH-), difluoromethylene (-CF₂ -), chloromethylene (-CHC1-) and the like. [064] The term "heteroalkyl," as used herein, alone or in combination, refers to a stable straight or branched chain, or cyclic hydrocarbon group, or combinations thereof, fully saturated or containing from 1 to 3 degrees of unsaturation, consisting of the stated number of carbon atoms and from one to three heteroatoms selected from the group consisting of O, N, and S, and wherein the nitrogen and sulfur atoms may optionally be oxidized and the nitrogen heteroatom may optionally be quaternized. The heteroatom(s) O, N and S may be placed at any interior position of the heteroalkyl group. Up to two heteroatoms may be consecutive, such as, for example, -CH₂-NH- OCH₃.

[065] The term "heteroaryl," as used herein, alone or in combination, refers to a 3 to 7 membered unsaturated heteromonocyclic ring, or a fused monocyclic, bicyclic, or tricyclic ring system in which at least one of the fused rings is aromatic, which contains at least one atom selected from the group consisting of O, S, and N. In certain Attorney Docket No. KO 152-401 -PC

embodiments, said heteroaryl will comprise from 5 to 7 carbon atoms. The term also embraces fused polycyclic groups wherein heterocyclic rings are fused with aryl rings, wherein heteroaryl rings are fused with other heteroaryl rings, wherein heteroaryl rings are fused with heterocycloalkyl rings, or wherein heteroaryl rings are fused with cycloalkyl rings. Examples of heteroaryl groups include pyrrolyl, pyrrolinyl, imidazolyl, pyrazolyl, pyridyl, pyrimidinyl, pyrazinyl, pyridazinyl, triazolyl, pyranyl, furyl, thienyl, oxazolyl, isoxazolyl, oxadiazolyl, thiazolyl, thiadiazolyl, isothiazolyl, indolyl, isoindolyl, indolizinyl, benzimidazolyl, quinolyl, isoquinolyl, quinoxalinyl, quinazolinyl, indazolyl, benzotriazolyl, benzodioxolyl, benzopyranyl, benzoxazolyl, benzoxadiazolyl, benzothiazolyl, benzothiadiazolyl, benzofuryl, benzothienyl, chromonyl, coumarinyl, benzopyranyl, tetrahydroquinolinyl, tetrazolopyridazinyl, tetrahydroisoquinolinyl, thienopyridinyl, furopyridinyl, pyrrolopyridinyl and the like. Exemplary tricyclic heterocyclic groups include carbazolyl, benzidolyl, phenanthrolinyl, dibenzofuranyl, acridinyl, phenanthridinyl, xanthenyl and the like. [066] The terms "heterocycloalkyl" and, interchangeably, "heterocycle," as used herein, alone or in combination, each refer to a saturated, partially unsaturated, or fully unsaturated monocyclic, bicyclic, or tricyclic heterocyclic group containing at least one heteroatom as a ring member, wherein each said heteroatom may be independently selected from the group consisting of nitrogen, oxygen, and sulfur In certain embodiments, said heterocycloalkyl will comprise from 1 to 4 heteroatoms as ring members. In further embodiments, said heterocycloalkyl will comprise from 1 to 2 heteroatoms as ring members. In certain embodiments, said heterocycloalkyl will comprise from 3 to 8 ring members in each ring. In further embodiments, said heterocycloalkyl will comprise from 3 to 7 ring members in each ring. In yet further embodiments, said heterocycloalkyl will comprise from 5 to 6 ring members in each ring. "Heterocycloalkyl" and "heterocycle" are intended to include sulfones, sulfoxides, N-oxides of tertiary nitrogen ring members, and carbocyclic fused and benzo fused ring systems; additionally, both terms also include systems where a heterocycle ring is fused to an aryl group, as defined herein, or an additional heterocycle group. Examples of heterocycle groups include aziridinyl, azetidinyl, 1,3- benzodioxolyl, dihydroisoindolyl, dihydroisoquinolinyl, dihydrocinnolinyl, Attorney Docket No. KO 152-401 -PC

dihydrobenzodioxinyl, dihydro[ 1 ,3]oxazolo[4,5-b]pyridinyl, benzothiazolyl, dihydroindolyl, dihy-dropyridinyl, 1,3-dioxanyl, 1 ,4-dioxanyl, 1,3-dioxolanyl, isoindolinyl, morpholinyl, piperazinyl, pyrrolidinyl, tetrahydropyridinyl, piperidinyl, thiomorpholinyl, and the like. The heterocycle groups may be optionally substituted unless specifically prohibited.

[067] The term "hydrazinyl" as used herein, alone or in combination, refers to two amino groups joined by a single bond, i.e., -N-N-.

[068] The term "hydroxy," as used herein, alone or in combination, refers to -OH.

[069] The term "hydroxyalkyl," as used herein, alone or in combination, refers to a hydroxy group attached to the parent molecular moiety through an alkyl group.

[070] The term "imino," as used herein, alone or in combination, refers to =N-.

[071] The term "iminohydroxy," as used herein, alone or in combination, refers to

=N(OH) and =N-O-.

[072] The phrase "in the main chain" refers to the longest contiguous or adjacent chain of carbon atoms starting at the point of attachment of a group to the compounds of any one of the formulas disclosed herein.

[073] The term "isocyanato" refers to a -NCO group.

[074] The term "isothiocyanato" refers to a -NCS group.

[075] The phrase "linear chain of atoms" refers to the longest straight chain of atoms independently selected from carbon, nitrogen, oxygen and sulfur.

[076] The term "lower," as used herein, alone or in a combination, where not otherwise specifically defined, means containing from 1 to and including 6 carbon atoms.

[077] The term "lower aryl," as used herein, alone or in combination, means phenyl or naphthyl, which may be optionally substituted as provided.

[078] The term "lower heteroaryl," as used herein, alone or in combination, means either 1) monocyclic heteroaryl comprising five or six ring members, of which between one and four said members may be heteroatoms selected from the group consisting of

O, S, and N, or 2) bicyclic heteroaryl, wherein each of the fused rings comprises five or six ring members, comprising between them one to four heteroatoms selected from the group consisting of O, S, and N. Attorney Docket No. KO 152-401 -PC

[079] The term "lower cycloalkyl," as used herein, alone or in combination, means a monocyclic cycloalkyl having between three and six ring members. Lower cycloalkyls may be unsaturated. Examples of lower cycloalkyl include cyclopropyl, cyclobutyl, cyclopentyl, and cyclohexyl.

[080] The term "lower heterocycloalkyl," as used herein, alone or in combination, means a monocyclic heterocycloalkyl having between three and six ring members, of which between one and four may be heteroatoms selected from the group consisting of

O, S, and N. Examples of lower heterocycloalkyls include pyrrolidinyl, imidazolidinyl, pyrazolidinyl, piperidinyl, piperazinyl, and morpholinyl. Lower heterocycloalkyls may be unsaturated.

[081] The term "lower amino," as used herein, alone or in combination, refers to

— NRR , wherein R and R are independently selected from the group consisting of hydrogen, lower alkyl, and lower heteroalkyl, any of which may be optionally substituted. Additionally, the R and R' of a lower amino group may combine to form a five- or six-membered heterocycloalkyl, either of which may be optionally substituted.

[082] The term "mercaptyl" as used herein, alone or in combination, refers to an

RS- group, where R is as defined herein.

[083] The term "nitro," as used herein, alone or in combination, refers to -NO₂.

[084] The terms "oxy" or "oxa," as used herein, alone or in combination, refer to

-O-.

[085] The term "oxo," as used herein, alone or in combination, refers to =0.

[086] The term "perhaloalkoxy" refers to an alkoxy group where all of the hydrogen atoms are replaced by halogen atoms.

[087] The term "perhaloalkyl" as used herein, alone or in combination, refers to an alkyl group where all of the hydrogen atoms are replaced by halogen atoms.

[088] The terms "sulfonate," "sulfonic acid," and "sulfonic," as used herein, alone or in combination, refer to the -SO₃H group and its anion as the sulfonic acid is used in salt formation.

[089] The term "sulfanyl," as used herein, alone or in combination, refers to -S-.

[090] The term "sulfmyl," as used herein, alone or in combination, refers to

-S(O)-. Attorney Docket No. KO 152-401 -PC

[091] The term "sulfonyl," as used herein, alone or in combination, refers to -

S(O)₂-.

[092] The term "N-sulfonamido" refers to a RS(=O)₂NR'- group with R and R' as defined herein.

[093] The term "S-sulfonamido" refers to a -S(=O)₂NRR' , group, with R and R' as defined herein.

[094] The terms "thia" and "thio," as used herein, alone or in combination, refer to a -S- group or an ether wherein the oxygen is replaced with sulfur. The oxidized derivatives of the thio group, namely sulfmyl and sulfonyl, are included in the definition of thia and thio.

[095] The term "thiol," as used herein, alone or in combination, refers to an -SH group.

[096] The term "thiocarbonyl," as used herein, when alone includes thioformyl -

C(S)H and in combination is a -C(S)- group.

[097] The term "N-thiocarbamyl" refers to an ROC(S)NR'- group, with R and R' as defined herein.

[098] The term "O-thiocarbamyl" refers to a -OC(S)NRR', group with R and R' as defined herein.

[099] The term "thiocyanato" refers to a -CNS group.

[0100] The term "trihalomethoxy" refers to a X3CO- group where X is a halogen.

[0101] The term "trisubstituted silyl," as used herein, alone or in combination, refers to a silicone group substituted at its three free valences with groups as listed herein under the definition of substituted amino. Examples include trimethysilyl, tert- butyldimethylsilyl, triphenylsilyl and the like.

[0102] Any definition herein may be used in combination with any other definition to describe a composite structural group. By convention, the trailing element of any such definition is that which attaches to the parent moiety. For example, the composite group alkylamido would represent an alkyl group attached to the parent molecule through an amido group, and the term alkoxyalkyl would represent an alkoxy group attached to the parent molecule through an alkyl group. Attorney Docket No. KO 152-401 -PC

[0103] When a group is defined to be "null," what is meant is that said group is absent.

[0104] The term "optionally substituted" means the anteceding group may be substituted or unsubstituted. When substituted, the substituents of an "optionally substituted" group may include, without limitation, one or more substituents independently selected from the following groups or a particular designated set of groups, alone or in combination: lower alkyl, lower alkenyl, lower alkynyl, lower alkanoyl, lower heteroalkyl, lower heterocycloalkyl, lower haloalkyl, lower haloalkenyl, lower haloalkynyl, lower perhaloalkyl, lower perhaloalkoxy, lower cycloalkyl, phenyl, aryl, aryloxy, lower alkoxy, lower haloalkoxy, oxo, lower acyloxy, carbonyl, carboxyl, lower alkylcarbonyl, lower carboxyester, lower carboxamido, cyano, hydrogen, halogen, hydroxy, amino, lower alkylamino, arylamino, amido, nitro, thiol, lower alkylthio, lower haloalkylthio, lower perhaloalkylthio, arylthio, sulfonate, sulfonic acid, trisubstituted silyl, N₃, SH, SCH₃, C(O)CH₃, CO₂CH₃, CO₂H, pyridinyl, thiophene, furanyl, lower carbamate, and lower urea. Two substituents may be joined together to form a fused five-, six-, or seven-membered carbocyclic or heterocyclic ring consisting of zero to three heteroatoms, for example forming methylenedioxy or ethylenedioxy. An optionally substituted group may be unsubstituted (e.g., -CH₂CH₃), fully substituted (e.g., -CF₂CF₃), monosubstituted (e.g., -CH₂CH₂F) or substituted at a level anywhere in-between fully substituted and monosubstituted (e.g., -CH₂CF₃). Where substituents are recited without qualification as to substitution, both substituted and unsubstituted forms are encompassed. Where a substituent is qualified as "substituted," the substituted form is specifically intended. Additionally, different sets of optional substituents to a particular moiety may be defined as needed; in these cases, the optional substitution will be as defined, often immediately following the phrase, "optionally substituted with."

[0105] The term R or the term R' , appearing by itself and without a number designation, unless otherwise defined, refers to a moiety selected from the group consisting of hydrogen, alkyl, cycloalkyl, heteroalkyl, aryl, heteroaryl and heterocycloalkyl, any of which may be optionally substituted. Such R and R' groups should be understood to be optionally substituted as defined herein. Whether an R Attorney Docket No. KO 152-401 -PC

group has a number designation or not, every R group, including R, R' and Rⁿ where n=(l, 2, 3, ...n), every substituent, and every term should be understood to be independent of every other in terms of selection from a group. Should any variable, substituent, or term (e.g. aryl, heterocycle, R, etc.) occur more than one time in a formula or generic structure, its definition at each occurrence is independent of the definition at every other occurrence. Those of skill in the art will further recognize that certain groups may be attached to a parent molecule or may occupy a position in a chain of elements from either end as written. Thus, by way of example only, an unsymmetrical group such as -C(O)N(R)- may be attached to the parent moiety at either the carbon or the nitrogen.

[0106] Asymmetric centers exist in the compounds disclosed herein. These centers are designated by the symbols "R" or "S," depending on the configuration of substituents around the chiral carbon atom. It should be understood that the invention encompasses all stereochemical isomeric forms, including diastereomeric, enantiomeric, and epimeric forms, as well as d-isomers and 1 -isomers, and mixtures thereof. Individual stereoisomers of compounds can be prepared synthetically from commercially available starting materials which contain chiral centers or by preparation of mixtures of enantiomeric products followed by separation such as conversion to a mixture of diastereomers followed by separation or recrystallization, chromatographic techniques, direct separation of enantiomers on chiral chromatographic columns, or any other appropriate method known in the art. Starting compounds of particular stereochemistry are either commercially available or can be made and resolved by techniques known in the art. Additionally, the compounds disclosed herein may exist as geometric isomers. The present invention includes all cis, trans, syn, anti, entgegen (E), and zusammen (Z) isomers as well as the appropriate mixtures thereof. Additionally, compounds may exist as tautomers; all tautomeric isomers are provided by this invention. Additionally, the compounds disclosed herein can exist in unsolvated as well as solvated forms with pharmaceutically acceptable solvents such as water, ethanol, and the like. In general, the solvated forms are considered equivalent to the unsolvated forms. Attorney Docket No. KO 152-401 -PC

[0107] The term "bond" refers to a covalent linkage between two atoms, or two moieties when the atoms joined by the bond are considered to be part of larger substructure. A bond may be single, double, or triple unless otherwise specified. A dashed line between two atoms in a drawing of a molecule indicates that an additional bond may be present or absent at that position.

[0108] The term "disease" as used herein is intended to be generally synonymous, and is used interchangeably with, the terms "disorder" and "condition" (as in medical condition), in that all reflect an abnormal condition of the human or animal body or of one of its parts that impairs normal functioning, is typically manifested by distinguishing signs and symptoms, and causes the human or animal to have a reduced duration or quality of life.

[0109] The term "combination therapy" means the administration of two or more therapeutic agents to treat a therapeutic condition or disorder described in the present disclosure. Such administration encompasses co-administration of these therapeutic agents in a substantially simultaneous manner, such as in a single capsule having a fixed ratio of active ingredients or in multiple, separate capsules for each active ingredient. In addition, such administration also encompasses use of each type of therapeutic agent in a sequential manner. In either case, the treatment regimen will provide beneficial effects of the drug combination in treating the conditions or disorders described herein.

[0110] "iNOS inhibitor" is used herein to refer to a compound that exhibits an IC50 with respect to iNOS activity of no more than about 100 μM and more typically not more than about 50 μM, as measured in the iNOS assay described generally hereinbelow. "IC50" is that concentration of inhibitor which reduces the activity of an enzyme (e.g., iNOS) to half-maximal level. Certain compounds disclosed herein have been discovered to exhibit inhibition against iNOS. In certain embodiments, compounds will exhibit an EC₅₀ with respect to iNOS of no more than about 10 μM; in further embodiments, compounds will exhibit an EC50 with respect to iNOS of no more than about 5 μM; in yet further embodiments, compounds will exhibit an EC50 with respect to iNOS of not more than about 1 μM; in yet further embodiments, compounds Attorney Docket No. KO 152-401 -PC

will exhibit an EC50 with respect to iNOS of not more than about 200 nM, as measured in the iNOS assay described herein.

[0111] The phrase "therapeutically effective" is intended to qualify the amount of active ingredients used in the treatment of a disease or disorder. This amount will achieve the goal of reducing or eliminating the said disease or disorder. [0112] The term "therapeutically acceptable" refers to those compounds (or salts, prodrugs, tautomers, zwitterionic forms, etc.) which are suitable for use in contact with the tissues of patients without undue toxicity, irritation, and allergic response, are commensurate with a reasonable benefit/risk ratio, and are effective for their intended use.

[0113] As used herein, reference to "treatment" of a patient is intended to include prophylaxis. The term "patient" means all mammals including humans. Examples of patients include humans, cows, dogs, cats, goats, sheep, pigs, and rabbits. Preferably, the patient is a human.

[0114] The term "prodrug" refers to a compound that is made more active in vivo. Certain compounds disclosed herein may also exist as prodrugs, as described in Hydrolysis in Drug and Prodrug Metabolism : Chemistry, Biochemistry, and Enzymology (Testa, Bernard and Mayer, Joachim M. Wiley- VHC A, Zurich, Switzerland 2003). Prodrugs of the compounds described herein are structurally modified forms of the compound that readily undergo chemical changes under physiological conditions to provide the compound. Additionally, prodrugs can be converted to the compound by chemical or biochemical methods in an ex vivo environment. For example, prodrugs can be slowly converted to a compound when placed in a transdermal patch reservoir with a suitable enzyme or chemical reagent. Prodrugs are often useful because, in some situations, they may be easier to administer than the compound, or parent drug. They may, for instance, be bioavailable by oral administration whereas the parent drug is not. The prodrug may also have improved solubility in pharmaceutical compositions over the parent drug. A wide variety of prodrug derivatives are known in the art, such as those that rely on hydrolytic cleavage or oxidative activation of the prodrug. An example, without limitation, of a prodrug would be a compound which is administered as an ester (the "prodrug"), but then is Attorney Docket No. KO 152-401 -PC

metabolically hydrolyzed to the carboxylic acid, the active entity. Additional examples include peptidyl derivatives of a compound.

[0115] The compounds disclosed herein can exist as therapeutically acceptable salts. The present invention includes compounds listed above in the form of salts, including acid addition salts. Suitable salts include those formed with both organic and inorganic acids. Such acid addition salts will normally be pharmaceutically acceptable. However, salts of non-pharmaceutically acceptable salts may be of utility in the preparation and purification of the compound in question. Basic addition salts may also be formed and be pharmaceutically acceptable. For a more complete discussion of the preparation and selection of salts, refer to Pharmaceutical Salts: Properties, Selection, and Use (Stahl, P. Heinrich. Wiley- VCHA, Zurich, Switzerland, 2002). [0116] The term "therapeutically acceptable salt," as used herein, represents salts or zwitterionic forms of the compounds disclosed herein which are water or oil-soluble or dispersible and therapeutically acceptable as defined herein. The salts can be prepared during the final isolation and purification of the compounds or separately by reacting the appropriate compound in the form of the free base with a suitable acid. Representative acid addition salts include acetate, adipate, alginate, L-ascorbate, aspartate, benzoate, benzenesulfonate (besylate), bisulfate, butyrate, camphorate, camphorsulfonate, citrate, digluconate, formate, fumarate, gentisate, glutarate, glycerophosphate, glycolate, hemisulfate, heptanoate, hexanoate, hippurate, hydrochloride, hydrobromide, hydroiodide, 2-hydroxyethansulfonate (isethionate), lactate, maleate, malonate, DL-mandelate, mesitylenesulfonate, methanesulfonate, naphthylenesulfonate, nicotinate, 2-naphthalenesulfonate, oxalate, pamoate, pectinate, persulfate, 3-phenylproprionate, phosphonate, picrate, pivalate, propionate, pyroglutamate, succinate, sulfonate, tartrate, L-tartrate, trichloroacetate, trifluoroacetate, phosphate, glutamate, bicarbonate, para-toluenesulfonate (p-tosylate), and undecanoate. Also, basic groups in the compounds disclosed herein can be quaternized with methyl, ethyl, propyl, and butyl chlorides, bromides, and iodides; dimethyl, diethyl, dibutyl, and diamyl sulfates; decyl, lauryl, myristyl, and steryl chlorides, bromides, and iodides; and benzyl and phenethyl bromides. Examples of acids which can be employed to form therapeutically acceptable addition salts include Attorney Docket No. KO 152-401 -PC

inorganic acids such as hydrochloric, hydrobromic, sulfuric, and phosphoric, and organic acids such as oxalic, maleic, succinic, and citric. Salts can also be formed by coordination of the compounds with an alkali metal or alkaline earth ion. Hence, the present invention contemplates sodium, potassium, magnesium, and calcium salts of the compounds disclosed herein, and the like.

[0117] Basic addition salts can be prepared during the final isolation and purification of the compounds by reacting a carboxy group with a suitable base such as the hydroxide, carbonate, or bicarbonate of a metal cation or with ammonia or an organic primary, secondary, or tertiary amine. The cations of therapeutically acceptable salts include lithium, sodium, potassium, calcium, magnesium, and aluminum, as well as nontoxic quaternary amine cations such as ammonium, tetramethylammonium, tetraethylammonium, methylamine, dimethylamine, trimethylamine, triethylamine, diethylamine, ethylamine, tributylamine, pyridine, JV,iV-dimethylaniline, N- methylpiperidine, JV-methylmorpholine, dicyclohexylamine, procaine, dibenzylamine, Λ/,Λ/-dibenzylphenethylamine, 1-ephenamine, and iV.Λ/'-dibenzylethylenediamme. Other representative organic amines useful for the formation of base addition salts include ethylenediamine, ethanolamine, diethanolamine, piperidine, and piperazine. [0118] While it may be possible for the compounds of the subject invention to be administered as the raw chemical, it is also possible to present them as a pharmaceutical formulation. Accordingly, provided herein are pharmaceutical formulations which comprise one or more of certain compounds disclosed herein, or one or more pharmaceutically acceptable salts, esters, prodrugs, amides, or solvates thereof, together with one or more pharmaceutically acceptable carriers thereof and optionally one or more other therapeutic ingredients. The carrier(s) must be "acceptable" in the sense of being compatible with the other ingredients of the formulation and not deleterious to the recipient thereof. Proper formulation is dependent upon the route of administration chosen. Any of the well-known techniques, carriers, and excipients may be used as suitable and as understood in the art; e.g., in Remington's Pharmaceutical Sciences. The pharmaceutical compositions disclosed herein may be manufactured in any manner known in the art, e.g. , by means Attorney Docket No. KO 152-401 -PC

of conventional mixing, dissolving, granulating, dragee-making, levigating, emulsifying, encapsulating, entrapping or compression processes. [0119] The formulations include those suitable for oral, parenteral (including subcutaneous, intradermal, intramuscular, intravenous, intraarticular, and intramedullary), intraperitoneal, transmucosal, transdermal, rectal and topical (including dermal, buccal, sublingual and intraocular) administration although the most suitable route may depend upon for example the condition and disorder of the recipient. The formulations may conveniently be presented in unit dosage form and may be prepared by any of the methods well known in the art of pharmacy. Typically, these methods include the step of bringing into association a compound of the subject invention or a pharmaceutically acceptable salt, ester, amide, prodrug or solvate thereof ("active ingredient") with the carrier which constitutes one or more accessory ingredients. In general, the formulations 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.

[0120] Formulations of the compounds disclosed herein suitable for oral administration may be presented as discrete units such as capsules, cachets or tablets each containing a predetermined amount of the active ingredient; as a powder or granules; as a solution or a suspension in an aqueous liquid or a non-aqueous liquid; or as an oil-in- water liquid emulsion or a water-in-oil liquid emulsion. The active ingredient may also be presented as a bolus, electuary or paste. [0121] Pharmaceutical preparations which can be used orally include tablets, push-fit capsules made of gelatin, as well as soft, sealed capsules made of gelatin and a plasticizer, such as glycerol or sorbitol. Tablets may be made by compression or molding, optionally with one or more accessory ingredients. Compressed tablets may be prepared by compressing in a suitable machine the active ingredient in a free- flowing form such as a powder or granules, optionally mixed with binders, inert diluents, or lubricating, surface active or dispersing agents. Molded tablets may be made by molding in a suitable machine a mixture of the powdered compound moistened with an inert liquid diluent. The tablets may optionally be coated or scored Attorney Docket No. KO 152-401 -PC

and may be formulated so as to provide slow or controlled release of the active ingredient therein. All formulations for oral administration should be in dosages suitable for such administration. The push- fit capsules can contain the active ingredients in admixture with filler such as lactose, binders such as starches, and/or lubricants such as talc or magnesium stearate and, optionally, stabilizers. In soft capsules, the active compounds may be dissolved or suspended in suitable liquids, such as fatty oils, liquid paraffin, or liquid polyethylene glycols. In addition, stabilizers may be added. Dragee cores are provided with suitable coatings. For this purpose, concentrated sugar solutions may be used, which may optionally contain gum arabic, talc, polyvinyl pyrrolidone, carbopol gel, polyethylene glycol, and/or titanium dioxide, lacquer solutions, and suitable organic solvents or solvent mixtures. Dyestuffs or pigments may be added to the tablets or dragee coatings for identification or to characterize different combinations of active compound doses. [0122] The compounds may be formulated for parenteral administration by injection, e.g., by bolus injection or continuous infusion. Formulations for injection may be presented in unit dosage form, e.g., in ampoules or in multi-dose containers, with an added preservative. The compositions may take such forms as suspensions, solutions or emulsions in oily or aqueous vehicles, and may contain formulatory agents such as suspending, stabilizing and/or dispersing agents. The formulations may be presented in unit-dose or multi-dose containers, for example sealed ampoules and vials, and may be stored in powder form or in a freeze-dried (lyophilized) condition requiring only the addition of the sterile liquid carrier, for example, saline or sterile pyrogen- free water, immediately prior to use. Extemporaneous injection solutions and suspensions may be prepared from sterile powders, granules and tablets of the kind previously described.

[0123] Formulations for parenteral administration include aqueous and nonaqueous (oily) sterile injection solutions of the active compounds which may contain antioxidants, buffers, bacteriostats and solutes which render the formulation isotonic with the blood of the intended recipient; and aqueous and non-aqueous sterile suspensions which may include suspending agents and thickening agents. Suitable lipophilic solvents or vehicles include fatty oils such as sesame oil, or synthetic fatty Attorney Docket No. KO 152-401 -PC

acid esters, such as ethyl oleate or triglycerides, or liposomes. Aqueous injection suspensions may contain substances which increase the viscosity of the suspension, such as sodium carboxymethyl cellulose, sorbitol, or dextran. Optionally, the suspension may also contain suitable stabilizers or agents which increase the solubility of the compounds to allow for the preparation of highly concentrated solutions. [0124] In addition to the formulations described previously, the compounds may also be formulated as a depot preparation. Such long acting formulations may be administered by implantation (for example subcutaneously or intramuscularly) or by intramuscular injection. Thus, for example, the compounds may be formulated with suitable polymeric or hydrophobic materials (for example as an emulsion in an acceptable oil) or ion exchange resins, or as sparingly soluble derivatives, for example, as a sparingly soluble salt.

[0125] For buccal or sublingual administration, the compositions may take the form of tablets, lozenges, pastilles, or gels formulated in conventional manner. Such compositions may comprise the active ingredient in a flavored basis such as sucrose and acacia or tragacanth.

[0126] The compounds may also be formulated in rectal compositions such as suppositories or retention enemas, e.g., containing conventional suppository bases such as cocoa butter, polyethylene glycol, or other glycerides.

[0127] Certain compounds disclosed herein may be administered topically, that is by non-systemic administration. This includes the application of a compound disclosed herein externally to the epidermis or the buccal cavity and the instillation of such a compound into the ear, eye and nose, such that the compound does not significantly enter the blood stream. In contrast, systemic administration refers to oral, intravenous, intraperitoneal and intramuscular administration.

[0128] Formulations suitable for topical administration include liquid or semi- liquid preparations suitable for penetration through the skin to the site of inflammation such as gels, liniments, lotions, creams, ointments or pastes, and drops suitable for administration to the eye, ear or nose. The active ingredient for topical administration may comprise, for example, from 0.001% to 10% w/w (by weight) of the formulation. In certain embodiments, the active ingredient may comprise as much as 10% w/w. In Attorney Docket No. KO 152-401 -PC

other embodiments, it may comprise less than 5% w/w. In certain embodiments, the active ingredient may comprise from 2% w/w to 5% w/w. In other embodiments, it may comprise from 0.1% to 1% w/w of the formulation.

[0129] Gels for topical or transdermal administration may comprise, generally, a mixture of volatile solvents, nonvolatile solvents, and water. In certain embodiments, the volatile solvent component of the buffered solvent system may include lower (Cl- C6) alkyl alcohols, lower alkyl glycols and lower glycol polymers. In further embodiments, the volatile solvent is ethanol. The volatile solvent component is thought to act as a penetration enhancer, while also producing a cooling effect on the skin as it evaporates. The nonvolatile solvent portion of the buffered solvent system is selected from lower alkylene glycols and lower glycol polymers. In certain embodiments, propylene glycol is used. The nonvolatile solvent slows the evaporation of the volatile solvent and reduces the vapor pressure of the buffered solvent system. The amount of this nonvolatile solvent component, as with the volatile solvent, is determined by the pharmaceutical compound or drug being used. When too little of the nonvolatile solvent is in the system, the pharmaceutical compound may crystallize due to evaporation of volatile solvent, while an excess may result in a lack of bioavailability due to poor release of drug from solvent mixture. The buffer component of the buffered solvent system may be selected from any buffer commonly used in the art; in certain embodiments, water is used. A common ratio of ingredients is about 20% of the nonvolatile solvent, about 40% of the volatile solvent, and about 40% water. There are several optional ingredients which can be added to the topical composition. These include, but are not limited to, chelators and gelling agents. Appropriate gelling agents can include, but are not limited to, semisynthetic cellulose derivatives (such as hydroxypropylmethylcellulose) and synthetic polymers, and cosmetic agents.

[0130] Lotions include those suitable for application to the skin or eye. An eye lotion may comprise a sterile aqueous solution optionally containing a bactericide and may be prepared by methods similar to those for the preparation of drops. Lotions or liniments for application to the skin may also include an agent to hasten drying and to Attorney Docket No. KO 152-401 -PC

cool the skin, such as an alcohol or acetone, and/or a moisturizer such as glycerol or an oil such as castor oil or arachis oil.

[0131] Creams, ointments or pastes are semi-solid formulations of the active ingredient for external application. They may be made by mixing the active ingredient in finely-divided or powdered form, alone or in solution or suspension in an aqueous or non-aqueous fluid, with the aid of suitable machinery, with a greasy or non-greasy base. The base may comprise hydrocarbons such as hard, soft or liquid paraffin, glycerol, beeswax, a metallic soap; a mucilage; an oil of natural origin such as almond, corn, arachis, castor or olive oil; wool fat or its derivatives or a fatty acid such as stearic or oleic acid together with an alcohol such as propylene glycol or a macrogel. The formulation may incorporate any suitable surface active agent such as an anionic, cationic or non-ionic surfactant such as a sorbitan ester or a polyoxyethylene derivative thereof. Suspending agents such as natural gums, cellulose derivatives or inorganic materials such as silicaceous silicas, and other ingredients such as lanolin, may also be included.

[0132] Drops may comprise sterile aqueous or oily solutions or suspensions and may be prepared by dissolving the active ingredient in a suitable aqueous solution of a bactericidal and/or fungicidal agent and/or any other suitable preservative, and, in certain embodiments, including a surface active agent. The resulting solution may then be clarified by filtration, transferred to a suitable container which is then sealed and sterilized by autoclaving or maintaining at 98-100⁰C for half an hour. Alternatively, the solution may be sterilized by filtration and transferred to the container by an aseptic technique. Examples of bactericidal and fungicidal agents suitable for inclusion in the drops are phenylmercuric nitrate or acetate (0.002%), benzalkonium chloride (0.01%) and chlorhexidine acetate (0.01%). Suitable solvents for the preparation of an oily solution include glycerol, diluted alcohol and propylene glycol. [0133] Formulations for topical administration in the mouth, for example buccally or sublingually, include lozenges comprising the active ingredient in a flavored basis such as sucrose and acacia or tragacanth, and pastilles comprising the active ingredient in a basis such as gelatin and glycerin or sucrose and acacia. Attorney Docket No. KO 152-401 -PC

[0134] For administration by inhalation, compounds may be conveniently delivered from an insufflator, nebulizer pressurized packs or other convenient means of delivering an aerosol spray. Pressurized packs may comprise a suitable propellant such as dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide or other suitable gas. In the case of a pressurized aerosol, the dosage unit may be determined by providing a valve to deliver a metered amount. Alternatively, for administration by inhalation or insufflation, the compounds according to the invention may take the form of a dry powder composition, for example a powder mix of the compound and a suitable powder base such as lactose or starch. The powder composition may be presented in unit dosage form, in for example, capsules, cartridges, gelatin or blister packs from which the powder may be administered with the aid of an inhalator or insufflator.

[0135] Preferred unit dosage formulations are those containing an effective dose, as herein below recited, or an appropriate fraction thereof, of the active ingredient. [0136] It should be understood that in addition to the ingredients particularly mentioned above, the formulations described above may include other agents conventional in the art having regard to the type of formulation in question, for example those suitable for oral administration may include flavoring agents. [0137] Compounds may be administered orally or via injection at a dose of from 0.1 to 500 mg/kg per day. Tablets or other forms of presentation provided in discrete units may conveniently contain an amount of one or more compounds which is effective at such dosage or as a multiple of the same, for instance, units containing 5 mg to 500 mg, usually around 10 mg to 200 mg.

[0138] 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.

[0139] The compounds can be administered in various modes, e.g. orally, topically, or by injection. The precise amount of compound administered to a patient will be the responsibility of the attendant physician. 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, diets, time of Attorney Docket No. KO 152-401 -PC

administration, route of administration, rate of excretion, drug combination, the precise disorder being treated, and the severity of the indication or condition being treated. Also, the route of administration may vary depending on the condition and its severity. [0140] In certain instances, it may be appropriate to administer at least one of the compounds described herein (or a pharmaceutically acceptable salt, ester, or prodrug thereof) in combination with another therapeutic agent. By way of example only, if one of the side effects experienced by a patient upon receiving one of the compounds herein is hypertension, then it may be appropriate to administer an anti-hypertensive agent in combination with the initial therapeutic agent. Or, by way of example only, the therapeutic effectiveness of one of the compounds described herein may be enhanced by administration of an adjuvant (i.e., by itself the adjuvant may only have minimal therapeutic benefit, but in combination with another therapeutic agent, the overall therapeutic benefit to the patient is enhanced). Or, by way of example only, the benefit of experienced by a patient may be increased by administering one of the compounds described herein with another therapeutic agent (which also includes a therapeutic regimen) that also has therapeutic benefit. By way of example only, in a treatment for diabetes involving administration of one of the compounds described herein, increased therapeutic benefit may result by also providing the patient with another therapeutic agent for diabetes. In any case, regardless of the disease, disorder or condition being treated, the overall benefit experienced by the patient may simply be additive of the two therapeutic agents or the patient may experience a synergistic benefit.

[0141] Specific, non- limiting examples of possible combination therapies include use of the compounds of the invention with: a) corticosteroids including betamethasone dipropionate (augmented and nonaugemented), betamethasone valerate, clobetasol propionate, diflorasone diacetate, halobetasol propionate, amcinonide, dexosimethasone, fluocinolone acetononide, fluocinonide, halocinonide, clocortalone pivalate, and flurandrenalide; b) non-steroidal anti-inflammatory drugs including diclofenac, ketoprofen, and piroxicam; c) muscle relaxants and combinations thereof with other agents, including cyclobenzaprine, baclofen, cyclobenzaprine/lidocaine, baclofen/cyclobenzaprine, and cyclobenzaprine/lidocaine/ketoprofen; d) anaesthetics Attorney Docket No. KO 152-401 -PC

and combinations thereof with other agents, including lidocaine, lidocaine/deoxy-D- glucose (an antiviral), prilocaine, and EMLA Cream [Eutectic Mixture of Local Anesthetics (lidocaine 2.5% and prilocaine 2.5%; an emulsion in which the oil phase is a eutectic mixture of lidocaine and prilocaine in a ratio of 1 : 1 by weight. This eutectic mixture has a melting point below room temperature and therefore both local anesthetics exist as a liquid oil rather then as crystals)]; e) expectorants and combinations thereof with other agents, including guaifenesin and guaifenesin/ketoprofen/cyclobenzaprine; f) antidepressants including tricyclic antidepressants (e.g., amitryptiline, doxepin, desipramine, imipramine, amoxapine, clomipramine, nortriptyline, and protriptyline), selective serotonin/norepinephrine reuptake inhibitors including (e.g., duloxetine and mirtazepine), and selective norepinephrine reuptake inhibitors (e.g., nisoxetine, maprotiline, and reboxetine), selective serotonin reuptake inhibitors (e.g., fluoxetine and fluvoxamine); g) anticonvulsants and combinations thereof, including gabapentin, carbamazepine, felbamate, lamotrigine, topiramate, tiagabine, oxcarbazepine, carbamezipine, zonisamide, mexiletine, gabapentin/clonidine, gabapentin/carbamazepine, and carbamazepine/cyclobenzaprine; h) antihypertensives including clonidine; i) opioids including loperamide, tramadol, morphine, fentanyl, oxycodone, levorphanol, and butorphanol; j) topical counter-irritants including menthol, oil of wintergreen, camphor, eucalyptus oil and turpentine oil; k) topical cannabinoids including selective and non-selective CB1/CB2 ligands; 1) histamine receptor antagonists including selective and non-selective HiR and H₄R ligands; and other agents, such as TRVPl antagonists and capsaicin.

[0142] In any case, the multiple therapeutic agents (at least one of which is a compound disclosed herein) may be administered in any order or even simultaneously. If simultaneously, the multiple therapeutic agents may be provided in a single, unified form, or in multiple forms (by way of example only, either as a single pill or as two separate pills). One of the therapeutic agents may be given in multiple doses, or both may be given as multiple doses. If not simultaneous, the timing between the multiple doses may be any duration of time ranging from a few minutes to four weeks. Attorney Docket No. KO 152-401 -PC

[0143] Thus, in another aspect, certain embodiments provide methods for treating iNOS-mediated disorders in a human or animal subject in need of such treatment comprising administering to said subject an amount of a compound disclosed herein effective to reduce or prevent said disorder in the subject, in combination with at least one additional agent for the treatment of said disorder that is known in the art. In a related aspect, certain embodiments provide therapeutic compositions comprising at least one compound disclosed herein in combination with one or more additional agents for the treatment of iNOS-mediated disorders.

[0144] Compounds of the subject invention are useful in treating nitric oxide synthase-mediated disease, disorders and conditions, and are particularly suitable as inhibitors of nitric oxide synthase. A recent study in iNOS knockout mice found that during microvascular changes following injury, axon and myelin breakdown, and "clearance" prior to regeneration, nitric oxide contributes to the development of neuropathic pain (Levy D, Kubes P, and Zochodne DW, J Neuropathol Exp Neurol. 2001 May;60(5):411-21). The compounds of the present invention are useful to treat patients with neuropathy or inflammatory pain such as reflex sympathetic dystrophy/causalgia (nerve injury), peripheral neuropathy (including diabetic neuropathy; Kern TS, Exp Diabetes Res. 2007;2007:95-103), intractable cancer pain, complex regional pain syndrome, chemotherapy-induced neuropathy, HIV neuropathy, or entrapment neuropathy (carpel tunnel syndrome). The compounds are also useful in the treatment of pain associated with acute herpes zoster (shingles), postherpetic neuralgia (PHN), and associated pain syndromes such as ocular pain. The compounds are further useful as analgesics in the treatment of pain such as surgical analgesia, or as an antipyretic for the treatment of fever. Pain indications include, but are not limited to, treatment or prophylaxis of surgical or post-surgical pain for various surgical procedures including amputation, post-cardiac surgery, dental pain/dental extraction, pain resulting from cancer, muscular pain, mastalgia, pain resulting from dermal injuries, lower back pain, headaches of various etiologies, including migraine, and the like. The compounds are also useful for the treatment of pain-related disorders such as tactile allodynia and hyperalgesia. The pain may be somatogenic (either nociceptive or neuropathic), acute and/or chronic. The nitric oxide inhibitors of the subject invention Attorney Docket No. KO 152-401 -PC

are also useful in conditions where NSAIDs, morphine or fentanyl opiates and/or other opioid analgesics would traditionally be administered.

[0145] Furthermore, the compounds of the subject invention can be used in the treatment or prevention of opiate tolerance in patients needing protracted opiate analgesics, and benzodiazepine tolerance in patients taking benzodiazepines, and other addictive behavior, for example, nicotine addiction, alcoholism, and eating disorders. Moreover, the compounds and methods of the present invention are useful in the treatment or prevention of drug withdrawal symptoms, for example treatment or prevention of symptoms of withdrawal from opiate, alcohol, or tobacco addiction. [0146] In addition, the compounds of the subject invention can be used to treat insulin resistance and other metabolic disorders such as atherosclerosis that are typically associated with an exaggerated inflammatory signaling. [0147] The present invention encompasses therapeutic methods using novel selective iNOS inhibitors to treat or prevent respiratory disease or conditions, including therapeutic methods of use in medicine for preventing and treating a respiratory disease or condition including: asthmatic conditions including allergen-induced asthma, exercise-induced asthma, pollution-induced asthma, cold-induced asthma, and viral- induced-asthma; chronic obstructive pulmonary diseases including chronic bronchitis with normal airflow, chronic bronchitis with airway obstruction (chronic obstructive bronchitis), emphysema, asthmatic bronchitis, and bullous disease; and other pulmonary diseases involving inflammation including bronchioectasis cystic fibrosis, pigeon fancier's disease, farmer's lung, acute respiratory distress syndrome, pneumonia, aspiration or inhalation injury, fat embolism in the lung, acidosis inflammation of the lung, acute pulmonary edema, acute mountain sickness, acute pulmonary hypertension, persistent pulmonary hypertension of the newborn, perinatal aspiration syndrome, hyaline membrane disease, acute pulmonary thromboembolism, heparin-protamine reactions, sepsis, status asthamticus, hypoxia, hyperoxic lung injuries (Respiratory Research 2004, 5:1), and injury induced by inhalation of certain injurious agents including cigarette smoking, leading up to complications thereof such as lung carcinoma. Attorney Docket No. KO 152-401 -PC

[0148] Other disorders or conditions which can be advantageously treated by the compounds of the present invention include inflammation. The compounds of the present invention are useful as anti-inflammatory agents with the additional benefit of having significantly less harmful side effects. The compounds are useful to treat arthritis (Cuzzocrea S., Curr Pharm Des. 2006;12(27):3551-70, including but not limited to rheumatoid arthritis (Maki-Petaja KM et al, MJ Cardiol. 2008 Jun 18. [Epub ahead of print]), spondyloarthropathies, gouty arthritis, osteoarthritis, systemic lupus erythematosus, juvenile arthritis, acute rheumatic arthritis, enteropathic arthritis, neuropathic arthritis, psoriatic arthritis, and pyogenic arthritis. The compounds are also useful in treating osteoporosis and other related bone disorders. These compounds can also be used to treat gastrointestinal conditions such as reflux esophagitis, diarrhea, inflammatory bowel disease, Crohn's disease, gastritis, irritable bowel syndrome, ulcerative colitis, and acute and chronic inflammation of the pancreas. The compounds may also be used in the treatment of pulmonary inflammation, such as that associated with viral infections and cystic fibrosis. In addition, compounds of invention are also useful in organ transplant patients either alone or in combination with conventional immunomodulators. Yet further, the compounds of the invention are useful in the treatment of pruritis and vitaligo.

[0149] The compounds of the present invention are also useful in treating organ and tissue injury injury associated with severe burns, sepsis, trauma, and hemorrhage- or resuscitation-induced hypotension, and also in such diseases as vascular diseases, migraine headaches, periarteritis nodosa, thyroiditis, aplastic anemia, Hodgkin's disease, sclerodoma, rheumatic fever, type I diabetes, neuromuscular junction disease including myasthenia gravis, white matter disease including multiple sclerosis, sarcoidosis, nephritis, nephrotic syndrome, Behcet's syndrome, polymyositis, gingivitis, periodontis, swelling occurring after injury, ischemias including myocardial ischemia, cardiovascular ischemia, and ischemia secondary to cardiac arrest, and the like.

[0150] The compounds of the subject invention are also be useful for the treatment of certain diseases and disorders of the nervous system. Central nervous system disorders in which nitric oxide inhibition is useful include cortical dementias including Attorney Docket No. KO 152-401 -PC

Alzheimer's disease, central nervous system damage resulting from stroke, ischemias including cerebral ischemia (both focal ischemia, thrombotic stroke and global ischemia (for example, secondary to cardiac arrest), and trauma. Neurodegenerative disorders in which nitric oxide inhibition is useful include nerve degeneration or nerve necrosis in disorders such as hypoxia, hypoglycemia, epilepsy, and in cases of central nervous system (CNS) trauma (such as spinal cord and head injury), hyperbaric oxygen-induced convulsions and toxicity, dementia e.g. pre-senile dementia, and AIDS-related dementia, cachexia, Sydenham's chorea, Huntington's disease, Parkinson's Disease, amyotrophic lateral sclerosis (ALS), Korsakoff s disease, cognitive disorders relating to a cerebral vessel disorder, hypersensitivity, sleeping disorders, schizophrenia, depression, depression or other symptoms associated with Premenstrual Syndrome (PMS), and anxiety.

[0151] Furthermore, the compounds of the present invention are also useful in inhibiting NO production from L-arginine including systemic hypotension associated with septic and/or toxic hemorrhagic shock induced by a wide variety of agents; therapy with cytokines such as TNF, IL-I and IL-2; and as an adjuvant to short term immunosuppression in transplant therapy. These compounds can also be used to treat allergic rhinitis, respiratory distress syndrome, endotoxic shock syndrome, and atherosclerosis.

[0152] Still other disorders or conditions advantageously treated by the compounds of the subject invention include the prevention or treatment of hyperproliferative diseases, especially cancers, either alone or incombination of standards of care especially those agents that target tumor growth by re-instating the aberrant apoptotic machinery in the malignant cells. Hematological and non-hematological malignancies which may be treated or prevented include but are not limited to multiple myeloma, acute and chronic leukemias including Acute Lymphocytic Leukemia (ALL), Chronic Lymphocytic Leukemia (CLL), and Chronic Myelogenous Leukemia(CLL), lymphomas, including Hodgkin's lymphoma and non-Hodgkin's lymphoma (low, intermediate, and high grade), as well as solid tumors and malignancies of the brain, head and neck, breast, lung, reproductive tract, upper digestive tract, pancreas, liver, renal, bladder, prostate and colorectal. The present compounds and methods can also Attorney Docket No. KO 152-401 -PC

be used to treat the fibrosis, such as that which occurs with radiation therapy. The present compounds and methods can be used to treat subjects having adenomatous polyps, including those with familial adenomatous polyposis (FAP). Additionally, the present compounds and methods can be used to prevent polyps from forming in patients at risk of FAP.

[0153] The compounds of the subject invention can be used in the treatment of ophthalmic diseases, such as glaucoma, retinal ganglion degeneration, ocular ischemia, corneal neovascularization, optic neuritis, retinitis, retinopathies such as glaucomatous retinopathy and/or diabetic retinopathy, uveitis, ocular photophobia, dry eye, Sjogren's syndrome, seasonal and chronic allergic conjunctivitis, and of inflammation and pain associated with chronic ocular disorders and acute injury to the eye tissue. The compounds can also be used to treat post-operative inflammation or pain as from ophthalmic surgery such as cataract surgery and refractive surgery. [0154] Moreover, compounds of the subject invention may be used in the treatment of menstrual cramps, dysmenorrhea, premature labor, tendonitis, bursitis, skin-related conditions such as psoriasis, eczema, burns, sunburn, dermatitis, pancreatitis, hepatitis, and the like. Other conditions in which the compounds of the subject invention provide an advantage in inhibiting nitric oxide inhibition include diabetes (type I or type II), congestive heart failure, myocarditis, atherosclerosis, and aortic aneurysm. [0155] The present compounds may also be used in co-therapies, partially or completely, in place of other conventional anti-inflammatory therapies, such as together with steroids, NSAIDs, COX-2 selective inhibitors, 5 -lipoxygenase inhibitors, LTB₄ antagonists and LTA₄ hydrolase inhibitors. The compounds of the subject invention may also be used to prevent tissue damage when therapeutically combined with antibacterial or antiviral agents.

[0156] Besides being useful for human treatment, certain compounds and formulations disclosed herein may also be useful for veterinary treatment of companion animals, exotic animals and farm animals, including mammals, rodents, and the like. More preferred animals include horses, dogs, and cats. Attorney Docket No. KO 152-401 -PC

[0157] All references, patents or applications, U.S. or foreign, cited in the application are hereby incorporated by reference as if written herein in their entireties. Where any inconsistencies arise, material literally disclosed herein controls.

Attorney Docket No. KO 152-401 -PC

GENERAL SYNTHETIC METHODS FOR PREPARING COMPOUNDS

[0158] The following schemes can be used to practice the present invention.

Scheme 1

Reagents: (a) 3-oxobutanoate, xylene/pyridine, reflux, 3Oh. (b) (i) Br₂, AcOH, 25°C, 22h. (ii) Acetone, (c) H₂SO₄, 40⁰C, 2Oh. (d) POCl₃, 80⁰C, 3h. (e) HCOOK, MeOH, H₂O, reflux, 18h. (f) MeONa, MeOH, reflux, 2h. (g) MnO₂, CHCl₃, 50⁰C, 1.5h.

Scheme 2

Reagents: (a) (i) 2,2,2-trichloroethane-l,l-diol, Na₂SO₄, H₂O, 25°C, Ih. (ii) NH₂OKHCl 60⁰C, 2h. (b) and H₂SO₄, 80⁰C, 3h. (c) (i) Ac₂O, 130⁰C, 3h. Attorney Docket No. KO 152-401 -PC

(ii) NaOH, reflux, 3h. (d) POCl₃, 100⁰C, 2h. (e) NaOMe, MeOH, reflux, 18h. (f) SOCl₂, 80⁰C, 4h.

Scheme 3

Reagents: (a) NBS, AIBN, CCl₄, reflux, 3h. (b) Potassium isoindoline-1,3- dione, DMF, 25°C, 12h. (c) hydrazine hydrate, 50⁰C, 2h. (d) Isobutyryl chloride, Et₃N, THF, 0⁰C, Ih. (e) Fe, NH₄Cl, H₂O, EtOH, reflux, 2h. (f) 4- (Bromomethyl)-2-chloro-7,8-difluoroquinoline, Na₂CO₃, DMF, 50⁰C, 2h. (g) Titanium(IV) chloride, toluene, ACN, reflux, 18h. (h) AcOH, reflux, 3h.

Attorney Docket No. KO 152-401 -PC

Scheme 4

Reagents: (a) 3-Methylbutan-2-one, H₂O, KOH/EtOH, reflux, 2days. (b) 1- phenoxybenzene, 270⁰C, Ih. (c) NaBH₄, NiCl₂.6H₂O, MeOH, 25°C, Ih. (d) Titanium(IV) chloride, toluene, ACN, reflux, 18h. (e) AcOH, reflux, 3h.

Attorney Docket No. KO 152-401 -PC

Scheme 5

Reagents: (a) NaH, DMF, 25°C, 2- 1Oh.

Attorney Docket No. KO 152-401 -PC

Scheme 6

OEt

O

Reagents: (a) Sodium azide, DMF (500 niL), 50⁰C, 12h. (b) (i) Na, EtOH, 0⁰C. (ii) Picolinaldehyde, EtOH, 0⁰C, 4h. (c) Xylene, 110⁰C, Ih. (d) (i) MeI, Mg, Et₂O, reflux, 1.5h. (ii) ethyl H-pyrazolo[l,5-a]pyridine-2- carboxylate,0°C, lO min. (e) H₂, Pd/C, 33°C, 2h. (f) 7,8-Difluoro-2- methoxyquinoline-4-carbonyl chloride, 150⁰C, 3h. (g) AlLiH₄, AICI3, Et₂O, 0⁰C to 25°C, 15 min. (h) (i) n-BuLi, THF, -78 ⁰C to RT (ii) N-methoxy-N- methylcyclobutanecarboxamide, THF, -78 ⁰C to RT, 25 min. (i) O- Attorney Docket No. KO 152-401 -PC

(mesitylsulfonyl)hydroxylamine DCM, 0 ⁰C to RT, 5.5h. (j) 4- (bromomethyl)-7,8-difluoroquinolin-2(lH)-one, 230⁰C, 4 min.

Scheme 7

Reagents: (a) Isobutyryl chloride, Et₃N, toluene, reflux, 12h. (b) AlCl₃, 7,8- difluoro-2-methoxyquinoline-4-carbonyl chloride, DCM, -78°C to -10⁰C, 2.5h. (c) AlLiH₄₅ AlCl₃, Et₂O, 0⁰C to 25°C, 15 min. (d) HBr/AcOH, 50⁰C, Ih.

Attorney Docket No. KO 152-401 -PC

Scheme 8

j

Reagents: (a) Hydrazine, EtOH, reflux, 4h. (b) Raney Ni, H₂, EtOH, 18h. (c) Cyclobutylethanone, EtOH, reflux, 18h. (d) NH₃.H₂O, 125°C, 18h. (e) Pd/C, H₂, MeOH, RT, 1.5h. (f) NBS, CHCl₃, RT, 30 min or NBS, AIBN, 55°C, 20 min. (g) (i) THF, -78°C, 30 min (ii) 7,8-difluoro-2- methoxyquinoline-4-carbaldehyde, -78°C, 2h. (h) Et₃SiH, TFA, 85°C, 18h. (i) HBr/AcOH, 50⁰C, Ih. (j) NaI, TMSCl, ACN, 75°C, Ih. Attorney Docket No. KO 152-401 -PC

INTERMEDIATE 1

4-(Bromomethyl)-7,8-difluoroquinolin-2(lH)-one

Step 1 : N-(2,3-Difluorophenyl)-3-oxobutanamide

[0159] A solution of methyl 3-oxobutanoate (300 g, 2.4 mol) in xylene/pyridine (900mL/20mL) was heated to gentle reflux for 0.5h. Then 2,3-difluoroaniline (145 g, 1.1 mol) was added dropwise into above reaction mixture. The mixture was stirred at reflux for 30h. No further progress was observed by HPLC at this point. The solution was allowed to cool and was extracted with 500 mL 20% aqueous NaOH. At this point precipitated salts were removed by filtration. The aqueous layer was separated and made weakly acidic with concentrated HCl. The resulting precipitate was collected by filtration and air dried to give 149 g (62%) of N-(2,3-difluorophenyl)-3-oxobutanamide as a brown solid, 90% pure by HPLC. ¹H NMR (300 MHz, DMSO-d₆) δ 9.48 (s, IH), 8.10 (m, IH), 7.04 (m, IH), 6.92 (m, IH), 3.64 (s, 2H), 2.34 (s, 3H).

Step 2: 4-Bromo-N-(2,3-difluorophenyl)-3-oxobutanamide

[0160] To a solution of N-(2,3-difluorophenyl)-3-oxobutanamide (111 g, .52 mol) in AcOH (500 mL) was added dropwise Br₂ (80 mL, 1.5 mol). The mixture was stirred for 22h at RT. To it was then added 50 mL acetone. Once the reaction was complete by HPLC, to the mixture was added dropwise 170 mL of water. The mixture was seeded and crystals rapidly formed. The mixture was cooled and the solid was Attorney Docket No. KO 152-401 -PC

collected by filtration and air dried to give 80 g (52%) of 4-bromo-N-(2,3- difluorophenyl)-3-oxobutanamide as a light tan solid, 99% pure by HPLC.

Step 3: 4-(Bromomethyl)-7,8-difluoroquinolin-2(lH)-one

[0161] 4-Bromo-N-(2,3-difluorophenyl)-3-oxobutanamide (79g) was dissolved in concentrated H₂SO₄ (400 mL) heated to 40°C.The solution was stirred for 2Oh at 40⁰C. After cooling to RT, the solution was poured into 200Og of H₂OZiCe. The solid was filtered and washed with H₂O. It was then washed with 5% aqueous sodium bicarbonate, again with water, and dried under vacuum to give 59g of 4- (bromomethyl)-7,8-difluoroquinolin-2(lH)-one as an off-white solid (80 %). ¹H NMR (300 MHz, DMSO-de) δ 12.07 (s, IH), 7.72 (m, IH), 7.37 (m, IH), 6.78 (s, IH), 4.89 (s, 2H). LCMS: 274 (M+H)⁺.

INTERMEDIATE 2 4-(Bromomethyl)-2-chloro-7,8-difluoroquinoline

[0162] POCl₃ (50 mL) was added to 4-(bromomethyl)-7,8-difluoroquinolin-2(lH)- one (4 g, 14.65 mmol) and the resulting mixture was stirred at 80⁰C for 3h. The reaction progress was monitored by TLC (EtOAc/PE = 1 :5). The mixture was concentrated by evaporation under vacuum and the resulting residue was diluted with 200 mL of EtOAc. Adjustment of the pH to 8 was accomplished by the addition of NaHCO₃ and the aqueous layer was extracted with EtOAc (2x100 mL). The organics were combined, dried over Na₂SO₄, and concentrated by evaporation under vacuum to Attorney Docket No. KO 152-401 -PC

afford 3.5 g (80%) of 4-(bromomethyl)-2-chloro-7,8-difluoroquinoline as a yellow solid.

INTERMEDIATE 3 7,8-Difluoro-2-methoxyquinoline-4-carbonyl chloride

N-(2,3-Difluorophenyl)-2-(hydroxyimino)acetamide

[0163] 2,3-Difluoroaniline (15 g, 116.28 mmol) in HCl (10 mL).was added to a solution of 2,2,2-trichloroethane-l,l-diol (25 g, 164.47 mmol) and Na₂SO₄ (92 g, 647.89 mmol) in H₂O (100 mL) and the resulting mixture was stirred at RT for Ih. NH₂OH.HC1 (11.3 g, 163.77 mmol) was then added and the resulting solution was heated to 6O⁰C for 2h. The precipitate was filtered to afford in 15 g (crude) of N-(2,3- difluorophenyl)-2-(hydroxyimino)acetamide as a light yellow solid.

Step 2: 6,7-Difluoroindoline-2,3-dione

[0164] A mixture of N-(2,3-difluorophenyl)-2-(hydroxyimino)acetamide (5 g, 25.00 mmol) and H₂SO₄ (cone, 30 mL) was heated to 8O⁰C for 3h. After cooling, the mixture was poured into ice-H₂O (20OmL) and the precipitate was filtered and dried to afford 2.5 g (crude) of 6,7-difluoroindoline-2,3-dione as a brown solid. Attorney Docket No. KO 152-401 -PC

Step 3 : 7,8-Difluoro-2-hydroxyquinoline-4-carboxylic acid

[0165] A mixture of 6,7-difluoroindoline-2,3-dione (10 g, 54.64 mmol) and Ac₂O (100 rnL) was heated to 130⁰C for 3h. The mixture was concentrated by evaporation under vacuum, the residue obtained was dissolved in NaOH (2M, 100 mL), and the solution was refluxed for 3h. The resulting solution was cooled to RT and washed with EtOAc (2x50 mL). Adjustment of the pH to 2 was accomplished by the addition of HCl (4N, 50 mL) and the precipitate was filtered and dried to afford 5.6 g of 7,8- difluoro-2-hydroxyquinoline-4-carboxylic acid(crude) as a yellow solid.

Step 4: 2-Chloro-7,8-difluoroquinoline-4-carboxylic acid

[0166] A mixture of 7,8-difluoro-2-hydroxyquinoline-4-carboxylic acid (5.6 g, 24.89 mmol) and POCl₃ (132.2 g) was heated to 100⁰C for 2h. The reaction progress was monitored by TLC (EtOAc/PE = 1 :1). The mixture was concentrated by evaporation under vacuum, the resulting residue was diluted with 500 mL of EtOAc, and it was washed with H₂O (2x100 mL). The organics were combined, dried, and concentrated to give a residue that was purified by column chromatography on silica gel (eluting with 1 :10 EtOAc/PE) to afford 3 g (50%) of 2-chloro-7,8- difluoroquinoline-4-carboxylic acid as a white solid.

Step 5: 7,8-Difluoro-2-methoxyquinoline-4-carboxylic acid

Attorney Docket No. KO 152-401 -PC

[0167] A mixture of 2-chloro-7,8-difluoroquinoline-4-carboxylic acid (2.8 g, 11.52 mmol)and NaOMe (6.3 g, 116.67 mmol) in MeOH (150 rnL) was refluxed for 18h. The reaction progress was monitored by TLC (EtOAc/PE = 1 :2). The mixture was concentrated by evaporation under vacuum and the resulting residue was diluted with 300 mL OfH₂O. Adjustment of the pH to <3 was accomplished by the addition of HCl (4N) and the precipitate was filtered, washed with H₂O, and dried to afford in 2.58 g (94%) of 7,8-difluoro-2-methoxyquinoline-4-carboxylic acid as a white solid.

Step 6: 7,8-Difluoro-2-methoxyquinoline-4-carbonyl chloride

[0168] A solution of 7,8-difluoro-2-methoxyquinoline-4-carboxylic acid (960 mg, 4.02 mmol) in SOCl₂ (15 mL) was heated to 80⁰C for 4h. The mixture was concentrated by evaporation under vacuum to afford 950 mg (crude) of 7,8-difluoro-2- methoxyquinoline-4-carbonyl chloride as a light yellow solid.

INTERMEDIATE 4

4-(Bromomethyl)-8-fluoroquinolin-2(lH)-one

[0169] 4-(Bromomethyl)-8-fluoroquinolin-2(lH)-one was synthesized as described for INTERMEDIATE 1 using 2-fiuoroaniline as a starting material. LCMS: 255 (M+H)⁺. Attorney Docket No. KO 152-401 -PC

INTERMEDIATE 5

7,8-Difluoro-2-methoxyquinoline-4-carbaldehyde

Step 1 : (2-Chloro-7,8-difluoroquinolin-4-yl)methanol

[0170] HCOOK (45 g) in H₂O (100 mL) was added to 4-(bromomethyl)-2-chloro- 7,8-difluoroquinoline (30 g) in MeOH (200 mL) and it was refluxed for 18h. The reaction mixture was cooled in an ice bath and the precipitate was filtered to afford 21 g (crude) of (2-chloro-7,8-difluoroquinolin-4-yl)methanol as a yellow solid.

Step 2: (7,8-Difluoro-2-methoxyquinolin-4-yl)methanol

[0171] A mixture of (2-chloro-7,8-difluoroquinolin-4-yl)methanol (1 g) and MeONa (1.08 g) in MeOH (50 mL) was refluxed for 2h. The cooled reaction mixture was slowly poured into H₂O (2OmL) of H2O and the precipitate was filtered to afford 0.5 g (55%) of (7,8-difluoro-2-methoxyquinolin-4-yl)methanol as a yellow solid. LCMS: 226 (M+H)⁺.

Step 3: 7,8-Difluoro-2-methoxyquinoline-4-carbaldehyde

Attorney Docket No. KO 152-401 -PC

[0172] A mixture of (7,8-difluoro-2-methoxyquinolin-4-yl)methanol (2.5 g, 11.1 mmol) and MnO₂ (9.8 g, 107.01 mmol,) in CHCl₃ (160 niL) was stirred at 50⁰C for 1.5h. The reaction progress was monitored by TLC (EtOAc/PE = 1 :5). The precipitate was filtered, washed with EtOAc and purified by flash chromatography on silica gel (eluting with 1 :80 EtOAc/PE solvent system) to afford 1.5 g (61%) of 7,8-difluoro-2- methoxyquinoline-4-carbaldehyde as a white solid. LCMS: 224 (M+H)⁺.

[0173] The invention is further illustrated by the following Examples, which can be made by a skilled practitioner using methods well know in the art, optionally using the methods and/or intermediates above, or optionally by the methods disclosed in United States Application No. 11/678,572, filed February 23, 2007, the disclosure of which is hereby incorporated by reference as if written herein in its entirety.

EXAMPLE 1

7,8-Difluoro-4-((2-isopropylquinazolin-l(4H)-yl)methyl)quinolin-2(lH)-one

Step 1 : l-(Bromomethyl)-2-nitrobenzene

[0174] A mixture of l-methyl-2-nitrobenzene (20 g, 144.53 mmol), NBS (28.4 g, 158.85 mmol), and AIBN (0.2 g) in CCl₄ (150 mL) was refiuxed for 3h, then cooled to RT, filtered, and concentrated under vacuum. The residue was partitioned between EtOAc and H₂O and the aqueous phase was extracted with EtOAc (3x300ml). The organics were combined, dried over Na₂SO₄, and concentrated under vacuum. The residue was purified by column chromatography on silica gel (eluting with 1 :20 Attorney Docket No. KO 152-401 -PC

EtOAc/PE) to afford 1O g (29%) of l-(bromomethyl)-2-nitrobenzene as a white solid. 1HNMR (300MHz, DMSOd₆) δ 8.08 (s, IH), 7.75 (m, 2H), 7.60 (m, IH), 4.94 (s, 2H)

Step 2: 2-(2-Nitrobenzyl)isoindoline-l,3-dione

[0175] A mixture of l-(bromomethyl)-2 -nitrobenzene (4.3 g, 19.80 mmol) and potassium isoindoline-l,3-dione (3.7 g, 19.80 mmol) in DMF (100 mL) was stirred at RT for 12h. The resulting solution was diluted with H₂O (100 mL) and the precipitate was filtered and dried to afford 4 g (64%) of 2-(2-nitrobenzyl)isoindoline-l,3-dione as a white solid.

Step 3: (2-Nitrophenyl)methanamine

[0176] Amixture of 2-(2-nitrobenzyl)isoindoline-l,3-dione (10 g, 35.11 mmol) and hydrazine hydrate (100 mL) was stirred at 50⁰C for 2h. The reaction progress was monitored by TLC (DCM:MeOH = 15:1). The resulting solution was diluted with H₂O(IOO mL) and it was extracted with EtOAc (3x200 mL), the organics were combined, dried over Na₂SO₄, and concentrated under vacuum. The residue was purified by column chromatography on silica gel (eluting 100:1 DCM:MeOH) to afford 4 g (67%) of (2-nitrophenyl)methanamine as yellow oil.

Step 4: N-(2-Nitrobenzyl)isobutyramide

[0177] Isobutyryl chloride (5.6 g, 52.30 mmol) in THF (20 mL) was added dropwise to a cooled (0⁰C) solution of (2-nitrophenyl)methanamine (4 g, 26.05 mmol) and Et₃N (3.2 g, 31.37 mmol) in THF (100 mL) and it was stirred at 0⁰C for Ih. The Attorney Docket No. KO 152-401 -PC

reaction progress was monitored by TLC (DCM:MeOH = 20: 1). The resulting solution was diluted with 100 rnL of iced H₂O. Adjustment of the pH to 8 was accomplished by the addition of NaHCO₃ and the aqueous layer was extracted with EtOAc (2x200 mL). The organics were combined, dried over Na₂SO₄, and concentrated under vacuum. The residue was purified by column chromatography on silica gel (eluting with 100:1 DCM:MeOH) to afford 5 g (78%) of N-(2-nitrobenzyl)isobutyramide as yellow oil. LCMS: 223 (M+H)⁺.

Step 5: N-(2-Aminobenzyl)isobutyramide

[0178] A mixture of N-(2-nitrobenzyl)isobutyramide (5 g, 22.30 mmol), Fe (20 g, 353.57 mmol), and NH₄Cl in H₂O (20 mL, 30%) in EtOH (150 mL) was refluxed for 2h. The reaction progress was monitored by TLC (DCM:MeOH = 20: 1). The insolubles were removed by filtration and the filtrate was concentrated under vacuum. The residue obtained was diluted with H₂O(200 mL), it was extracted with EtOAc (3x500 mL), and the combined organics were dried over Na₂SO₄, and concentrated under vacuum to give 4 g (84%) of N-(2-aminobenzyl)isobutyramide as a white solid. LCMS: 193 (M+H)⁺.

Step 6: N-(2-((2-Chloro-7,8-difluoroquinolin-4- yl)methylamino)benzyl)isobutyramide

[0179] A mixture of 4-(bromomethyl)-2-chloro-7,8-difluoroquinoline (INTERMEDIATE 2, 1 g, 3.40 mmol), N-(2-aminobenzyl)isobutyramide (730 mg, Attorney Docket No. KO 152-401 -PC

3.76 mmol), and Na₂CO₃ (370 mg, 3.46 mmol) in DMF (50 niL) was heated to 50⁰C for 2h. The reaction progress was monitored by TLC (EtOAc/PE = 1 :3). A filtration was performed and the filtrate was concentrated under vacuum. The residue was diluted with EtOAc (30 mL) and the precipitate was filtered, washed with H₂O (20 mL), and dried to give 1 g (66%) of N-(2-((2-chloro-7,8-difluoroquinolin-4- yl)methylamino)benzyl)isobutyramide as a white solid. LCMS: 404 (M+H)⁺.

Step 7: l-((2-Chloro-7,8-difluoroquinolin-3-yl)methyl)-2-isopr<)pyl-l,2- dihydroquinazoline

[0180] Titanium(IV) chloride was added dropwise to a cooled solution (10⁰C) of N-(2-((2-chloro-7,8-difluoroquinolin-3-yl)methylamino)benzyl)isobutyramide (900 mg, 2.12 mmol) in toluene and ACN (60 mL) and the resulting solution was refluxed for 18h. The reaction progress was monitored by TLC (DCM:MeOH = 15:1). The mixture was concentrated under vacuum. The reaction mixture was then quenched by the adding of ice/salt. A filtration was performed. The resulting solution was extracted with EtOAc (2x100 mL) and the combined organic layers was concentrated under vacuum. The residue was purified by column chromatography on silica gel (eluting with 1 :3 EtOAc/PE) to afford 0.6 g(68.32%) of l-((2-chloro-7,8-difluoroquinolin-3- yl)methyl)-2-isopropyl-l,2-dihydroquinazoline as a yellow solid. LCMS: 386 (M+H)⁺.

Attorney Docket No. KO 152-401 -PC

Step 8: 7,8-Difluoro-4-((2-isopropylquinazolin-l(2H)-yl)methyl)quinolin-2(lH)- one

[0181] A solution of 1 -((2-chloro-7,8-difluoroquinolin-4-yl)methyl)-2-isopropyl- 1 ,2-dihydroquinazoline (600 mg, 1.48 mmol) in AcOH (60 rnL) was refluxed for 3h. The reaction progress was monitored by TLC (DCM:MeOH = 1 :5). The mixture was concentrated under vacuum and the final product was purified by recrystallization from EtOH (2OmL) and hexane(20mL) to afford 0.35 g (64%) of 7,8-difluoro-4-((2- isopropylquinazolin-l(2H)-yl)methyl)quinolin-2(lH)-one as a white solid. ¹HNMR (300MHz, DMSO-de) δ 12.14 (s, IH), 10.60 (s, IH), 7.79 (m, IH), 7.51 (m, IH), 7.40 (m, 2H), 6.99 (s, IH), 6.32 (s, IH), 5.57 (s, 2H), 4.82 (s, 2H), 3.04 (m, IH), 1.29 (d, 6H). LCMS: 368 (M+H)⁺.

EXAMPLE 2

7,8-Difluoro-4-((2-isopropyl-3 ,4-dihydroquinolin- 1 (2H)-yl)methyl)quinolin-2( 1 H)-one

Step 1 : 2-Isopropylquinoline-4-carboxylic acid

Attorney Docket No. KO 152-401 -PC

[0182] A mixture of indoline-2,3-dione (8 g, 54.42 mmol), 3-methylbutan-2-one (9.36 g, 108.84 mmol), H₂O (100 mL), and KOH/EtOH (35 mL, 34%) was refluxed for 2days. EtOH was removed by evaporation and the aqueous layer was washed with EtOAc (200 mL). The pH of the aqueous layer was adjusted to 4 by the addition of HCl (4M) and the solid obtained was filtered to give 6 g (crude) of 2- isopropylquinoline-4-carboxylic acid as a light yellow solid. LCMS: 216 (M+H)⁺.

_2: 2-Isopropylquinoline

[0183] A solution of 2-isopropylquinoline-4-carboxylic acid (4.3 g, 20.00 mmol) in 1-phenoxybenzene (80 mL) was heated to 270⁰C for Ih. After cooling to RT, the solution was diluted with 80 mL of EtOAc and washed with HCl (4M, 2x60 mL). The pH of the aqueous layer was adjusted to 10 by the addition of KOH and then the mixture was extracted with EtOAc (2x100 mL). The organics were combined, washed with brine, dried, and evaporated. The crude product was purified by flash chromatography on silica gel (EtOAc/PE =1 :100) to afford 1.2 g (35%) of 2- isopropylquinoline as yellow oil. LCMS: 172 (M+H)⁺.

2-Isopropyl-l,2,3,4-tetrahydroquinoline

[0184] NaBH₄ (450 mg, 11.84 mmol) was added to a cooled (0⁰C) solution of NiCl₂.6H₂O (121 mg, 0.51 mmol) in MeOH (50 mL). 2-Isopropylquinoline (500 mg, 2.92 mmol) was then added and the resulting mixture was stirred at RT for Ih. MeOH was evaporated and 10%HCl (5mL) was added to dissolve the residue. The PH of the solution was adjusted to 11 and the mixture was extracted with Et₂O (2x10OmL). The organics were combined, dried, and evaporated to give 0.4 g (78%) of 2-isopropyl- 1,2,3,4-tetrahydroquinoline as a colorless liquid. ¹HNMR (300MHz, DMSO-d₆) δ 6.99 Attorney Docket No. KO 152-401 -PC

(m, 2H), 6.64 (m, IH), 6.52 (d, IH), 3.06 (m, IH), 2.81 (m, 2H), 1.93 (m, IH), 1.71 (m, 2H), 1.04 (d, 6H). LCMS: 176 (M+H)⁺.

Step 4: 2-Chloro-7,8-difluoro-4-((2-isopropyl-3,4-dihydroquinolin-l(2H)- yl)methyl)quinoline

[0185] 2-Chloro-7,8-difluoro-4-((2-isopropyl-3,4-dihydroquinolin-l(2H)- yl)methyl)quinoline was synthesized as described in EXAMPLE 1 , Step 6 using 2- isopropyl-1, 2,3, 4-tetrahydroquino line and 4-(bromomethyl)-2-chloro-7,8- difluoroquinoline (INTERMEDIATE 2) as starting materials. LCMS: 3896 (M+H)⁺.

Step 5: 7,8-Difluoro-4-((2-isopropyl-3,4-dihydroquinolin-l(2H)- yl)methyl)quinolin-2(lH)-one

[0186] 7,8-Difluoro-4-((2-isopropyl-3,4-dihydroquinolin-l(2H)- yl)methyl)quinolin-2(lH)-one was synthesized as described in EXAMPLE 1, Step 8 using 2-chloro-7,8-difluoro-4-((2-isopropyl-3,4-dihydroquinolin-l(2H)- yl)methyl)quinoline as a starting material. ¹HNMR (300MHz, DMSO-d₆) δ 11.92 (s, IH), 7.73 (m, IH), 7.30 (m, IH), 6.96-6.85 (m, 2H), 6.51 (m, IH), 6.33 (d, IH), 6.20 (s, IH), 4.55 (d, IH), 4.08 (d, IH), 3.12 (m, IH), 2.81 (m, 2H), 2.04 (m, IH), 1.88 (m, 2H), 1.00 (d, 6H). LCMS: 369 (M+H)⁺. Attorney Docket No. KO 152-401 -PC

EXAMPLE 3

4-((lH-Pyrrolo[2,3-b]pyridin-l-yl)methyl)-7,8-difluoroquinolin-2(lH)-one

[0187] NaH (131 mg, 3.3 mmol) was added to a solution of lH-pyrrolo[2,3- b]pyridine (168 mg, 1.43 mmol) in DMF (5 mL). 4-(Bromomethyl)-7,8- difluoroquinolin-2(lH)-one (INTERMEDIATE 1, 300 mg, 1.09 mmol) was then added as a solid and the resulting mixture was stirred at RT for Ih. The crude reaction mixture was diluted with MeOH (2 mL) and it was purified by reverse phase chromatography to afford 4-((lH-pyrrolo[2,3-b]pyridin-l-yl)methyl)-7,8- difluoroquinolin-2(lH)-one as white solid. ¹HNMR (400MHz, DMSO-d₆, TFA salt) δ 12.00 (s, IH), 8.25 (d, IH), 8.03 (d, IH), 7.84-7.81 (m, IH), 7.62 (d, IH), 7.32-7.27 (m, IH), 7.13 (m, IH), 6.60 (d, IH), 5.75 (s, 2H), 5.46 (s, IH). LCMS: 312 (M+H)⁺.

EXAMPLE 4

7,8-Difluoro-4-((2-isopropylpyrazolo[l,5-a]pyridin-3-yl)methyl)quinolin-2(lH)-one

Step 1 : Ethyl 2-azidoacetate

O

[0188] Ethyl 2-bromoacetate (171.4 g) in DMF (200 mL) was added dropwise to a solution of sodium azide (99.9 g, 1.52 mol) in DMF (500 mL) and the resulting Attorney Docket No. KO 152-401 -PC

solution was heated to 50⁰C for 12h. The reaction mixture was then quenched by adding 500 rnL of H₂O/ice and it was extracted with EtOAc (2x500 rnL). The organics were combined, washed with brine, dried, and concentrated under vacuum to afford HO g (75%) of ethyl 2-azidoacetate as yellow oil.

Step 2: Ethyl 2-azido-3-(pyridin-2-yl)acrylate

[0189] Na (19.6 g, 843.65 mmol) was added in several batches to EtOH (1000 mL) at 0⁰C. Picolinaldehyde (30.7 g, 284.05 mmol) in EtOH (200 mL) was then added dropwise at 0⁰C. This was followed by the dropwise addition of a solution of ethyl 2- azidoacetate (110 g, 844.19 mmol) in EtOH (200 mL). The resulting solution was stirred at 0⁰C for 4h. The reaction progress was monitored by TLC (EtOAc/PE = 1 :2). The reaction mixture was then quenched by adding 500 OfNH₄Cl in H₂O and it was extracted with EtOAc 2x1000 mL). The organics were combined, dried, and concentrated under vacuum to afford 27.8 g (40%) of ethyl 2-azido-3-(pyridin-2- yl)acrylate as a yellow solid. LCMS: 219 (M+H)⁺.

Step 3 : Ethyl H-pyrazolo[l,5-a]pyridine-2-carboxylate

[0190] Ethyl 2-azido-3-(pyridin-2-yl)acrylate (27.5 g, 124.89 mmol) in xylene (200 mL) was heated to 110⁰C for Ih. The reaction progress was monitored by TLC (EtOAc/PE = 1 :2). The mixture was concentrated by evaporation under vacuum and the residue was purified by column chromatography on silica gel (eluting with 1 :10 EtOAc/PE) to aford 5.3 g (20%) of ethyl H-pyrazolo[l,5-a]pyridine-2-carboxylate as a yellow solid. Attorney Docket No. KO 152-401 -PC

2-(Prop- l-en-2-yl)H-pyrazolo [ 1 ,5-a] pyridine

[0191] MeI (41.2 g, 287.24 mmol) in Et₂O (50 mL) was added dropwise to a solution of Mg (6.7 g, 276.38 mmol) in Et₂O (200 mL) and the mixture was refluxed for 1.5h. The reaction mixture was cooled to 0⁰C and a solution of ethyl H- pyrazolo[l,5-a]pyridine-2-carboxylate (5.3 g, 27.62 mmol) in Et₂O (50 mL) was added dropwise. The mixture was reacted for 10 min at 0⁰C and was then quenched by adding 200 mL of NH₄C1/H₂O. It was then extracted with EtOAc (3x200 mL) and the organics were combined, dried, and concentrated by evaporation under vacuum. A mixture of the obtained residue and P₂Os(Ig) in benzene (20OmL) was refluxed for Ih. Adjustment of the pH to 9 was accomplished by the addition of NaHCOs/H₂O and the aqueous layer was extracted with EtOAc (2x500 mL). The organics were combined, dried, and concentrated by evaporation under vacuum to afford 4.3 g (89%) of 2-(prop- l-en-2-yl)H-pyrazolo[l,5-a]pyridine as red oil.

2-IsopropylH-pyr azolo [ 1 ,5-a] pyridine

[0192] 2-(Prop-l-en-2-yl)H-pyrazolo[l,5-a]pyridine (4.3 g, 26.94 mmol) in ethanol (50 mL) was hydrogenated with a balloon of hydrogen in the presence of Pd/C (1.1 g) for 2h at 33°C. The reaction progress was monitored by TLC (EtOAc/PE = 1 :2). A filtration was performed to remove the Pd/C and the filtrate was concentrated by evaporation under vacuum. The residue was purified by column chromatography on silica gel (eluting with 1 :20 EtOAc/PE) to afford 2 g (42%) of 2-isopropylH- pyrazolo[l,5-a]pyridine as yellow oil. LCMS: 1619 (M+H)⁺. Attorney Docket No. KO 152-401 -PC

Step 6: 7,8-Difluoro-4-(2-isopropylpyrazolo[l,5-a]pyridine-3-carbonyl)quinolin- 2(lH)-one

[0193] A mixture of 2-isopropylH-pyrazolo[l,5-a]pyridine (160 mg, 0.99 mmol) and 7,8-difluoro-2-methoxyquinoline-4-carbonyl chloride (INTERMEDIATE 3, 510 mg, 1.96 mmol) was heated to 150⁰C for 3h. The reaction progress was monitored by TLC (DCM/MeOH = 20: 1). Adjustment of the pH to 9 was accomplished by the addition OfNaOHZH₂O and the aqueous layer was extracted with EtOAc (3x100 mL). The organics were combined, dried, and concentrated under vacuum to afford a residue that was purified by column chromatography on silica gel (eluting with 100:1 DCM/MeOH) to give 150 mg (37%) of 7,8-difluoro-4-(2-isopropylpyrazolo[l,5- a]pyridine-3-carbonyl)quinolin-2(lH)-one as a yellow solid. ¹H NMR (300 MHz, DMSO-de) δ 12.35 (s, IH), 8.89 (d, IH), 7.54 (m, IH), 7.43 (d, IH), 7.20 (m, 3H), 6.55 (s, IH), 3.46 (m, IH), 1.22 (d, 6H). LCMS: 368 (M+H)⁺.

Step 7: 7,8-Difluoro-4-((2-isopropylpyrazolo[l,5-a]pyridin-3-yl)methyl)quinolin- 2(lH)-one

[0194] AlLiH₄ (129 mg, 3.36 mmol) was added in several batches to a solution of AlCl₃ (223 mg, 1.67 mmol) in Et₂O (20 mL) at 0⁰C. This was followed by the dropwise addition of a solution of 7,8-difluoro-4-(2-isopropylpyrazolo[l,5-a]pyridine- 3-carbonyl)quinolin-2(lH)-one (207 mg, 0.56 mmol) in Et₂O (15 mL). The resulting Attorney Docket No. KO 152-401 -PC

solution was stirred at RT for 15 min. The reaction progress was monitored by TLC (DCM:MeOH = 20:1). The reaction mixture was then quenched by the addition of 0.13 mL OfH₂O, 0.3 mL of NaOH (15%) and 0.13 mL of H₂O. A filtration was performed and the filtrate was concentrated by evaporation under vacuum. The residue was purified by column chromatography on silica gel (eluting with 100:1 DCM:MeOH) to afford in 90 mg (45%) of 7,8-Difluoro-4-((2-isopropylpyrazolo[l,5- a]pyridin-3-yl)methyl)quinolin-2(lH)-one as a white solid. ¹H NMR (300 MHz, DMSO-de) δ 12.9 (s, IH), 8.66 (d, IH), 7.86 (m, IH), 7.51 (d, IH), 7.34 (d, IH), 7.15 (d, IH), 6.85 (d, IH), 5.81 (s, IH), 4.35 (s, 2H), 3.13 (m, IH), 1.27 (d, 6H). LCMS: 353 (M+H)⁺.

EXAMPLE 5

7,8-Difluoro-4-((2-isopropylbenzofuran-3-yl)methyl)quinolin-2(lH)-one

2-Isopropylbenzofuran

[0195] Isobutyryl chloride (4 g, 37.74 mmol) was added to a solution of (2- hydroxybenzyl)triphenylphosphonium bromide (10 g, 22.32 mmol), and Et₃N (10 g, 99.01 mmol) in toluene (150 mL) and it was refluxed for 12h. The reaction progress was monitored by TLC (hexanes). The reaction mixture was cooled and filtrated. The filtrate was diluted with EtOAc (300 mL) and the solvent was removed under vacuum. The residue was purified by flash chromatography on silica gel (PE) to afford 2 g (55%) of 2-isopropylbenzofuran as a light yellow oil. ¹H NMR (300 MHz, DMSO-d₆) δ 7.54-7.47 (m, 2H), 7.21-7.17 (m, 2H), 6.55 (s, IH), 3.05 (m, IH), 1.29 (d, 6H). Attorney Docket No. KO 152-401 -PC

Step 2: (7,8-Difluoro-2-methoxyquinolin-4-yl)(2-isopropylbenzofuran-3-yl)methanone

[0196] AICI₃ (350 mg, 2.63 mmol) was added dropwise to a solution of 7,8- difluoro-2-methoxyquinoline-4-carbonyl chloride (INTERMEDIATE 3, 260 mg, 1.01 mmol) and 2-isopropylbenzofuran (160 mg, 1.00 mmol, 1.00 equiv) in DCM (25 mL) cooled to -78°C. . The mixture was stirred for 30 min at -78°C, then warmed up to - 10⁰C and stirred for another 2h. The mixture was then quenched by adding 10 mL of HCl (2N) and diluted with 150 mL of EtOAc. The organic layer was washed with HCl (2N, 2x100 mL) and sat. NaHCO₃ (2x100 mL), dried, and concentrated. The residue obtained was purified by flash chromatography on silica gel (EtOAc/PE : 1 : 100) to afford 100 mg (26%) of 7,8-difluoro-2-methoxyquinolin-4-yl)(2-isopropylbenzofuran- 3-yl)methanone as a light yellow oil. ¹U NMR (300 MHz, DMSO-d₆) δ 7.63-7.19 (m, 7H), 4.02 (s, 3H), 3.05 (m, IH), 1.12 (d, 6H). LCMS: 382 (M+H)⁺.

7,8-Difluoro-4-((2-isopropylbenzofuran-3-yl)methyl)-2-methoxyquinoline

[0197] 7,8-difluoro-4-((2-isopropylbenzofuran-3-yl)methyl)-2-methoxyquinoline was synthesized as described in EXAMPLE 4, Step 7 using 7,8-difluoro-2- methoxyquinolin-4-yl)(2-isopropylbenzofuran-3-yl)methanone as a starting material. LCMS: 368 (M+H)⁺. Attorney Docket No. KO 152-401 -PC

Step 4: 7,8-Difluoro-4-((2-isopropylbenzofuran-3-yl)methyl)quinolin-2(lH)-one

[0198] A solution of 7,8-difluoro-4-((2-isopropylbenzofuran-3-yl)methyl)-2- methoxyquinoline (160 mg, 0.44 mmol) in HBr/ AcOH (15 rnL) was heated to 50⁰C for Ih. The mixture was then diluted with H₂O (50 rnL) and the pH value was adjusted to 9 with NaOH (20 %). The resulting aqueous solution was extracted with EtOAc (2x150 mL), the organics were combined, dried, and concentrated. The residue was purified by column chromatography on silica gel (DCM/MeOH = 200:1) to afford 35 mg (22%) 7,8-difluoro-4-((2-isopropylbenzofuran-3-yl)methyl)quinolin-2(lH)-one as a white solid. . ¹H NMR (300 MHz, DMSO-d₆) δ 11.9 (s, IH), 7.77 (m, IH), 7.50 (m, IH), 7.43-7.09 (m, 4H), 5.91 (s, IH), 4.23 (s, 2H), 3.26 (m, IH), 1.21 (d, 6H). LCMS: 354 (M+H)⁺.

EXAMPLE 6

4-((9H-Pyrido[3,4-b]indol-9-yl)methyl)-7,8-difiuoroquinolin-2(lH)-one

[0199] 4-((9H-Pyrido[3,4-b]indol-9-yl)methyl)-7,8-difluoroquinolin-2(lH)-one was synthesized as described in EXAMPLE 3 using 9H-pyrido[3,4-b]indole and 4- (bromomethyl)-7,8-difluoroquinolin-2(lH)-one (INTERMEDIATE 1) as starting materials. ¹H NMR (400 MHz, DMSO-d₆) δ 12.10 (br s, IH), 9.59 (s, IH), 8.95 (d, Attorney Docket No. KO 152-401 -PC

IH), 8.75 (d, IH), 8.69 (d, IH), 7.89-7.85 (m, 3H), 7.61-7.48 (m, 2H), 6.31 (s, 2H), 5.13 (s, IH). LCMS: 362.1 (M+H)⁺.

EXAMPLE 7

7, 8-Difluoro-4-((2-isopropyl- 1 H-indol- 1 -yl)methyl)quinolin-2( 1 H)-one

[0200] 7,8-Difluoro-4-((2-isopropyl- 1 H-indol- l-yl)methyl)quinolin-2(lH)-one was synthesized as described in EXAMPLE 3 using 2-isopropyl-lH-indole (J.Org. Chem. 1980, 45(8), 1546) and 4-(bromomethyl)-7,8-difluoroquinolin-2(lH)-one (INTERMEDIATE 1) as starting materials. LCMS: 353 (M+H)⁺.

EXAMPLE 8

7, 8-Difluoro-4-((3 -isopropyl- 1 H-indol- 1 -yl)methyl)quinolin-2( 1 H)-one

[0201 ] 7,8-Difluoro-4-((3-isopropyl- 1 H-indol- 1 -yl)methyl)quinolin-2( 1 H)-one was synthesized as described in EXAMPLE 3 using 3 -isopropyl- lH-indole (J.Org. Chem. 1980, 45(13), 2709) and 4-(bromomethyl)-7,8-difluoroquinolin-2(lH)-one (INTERMEDIATE 1) as starting materials. LCMS: 353 (M+H)⁺. Attorney Docket No. KO 152-401 -PC

EXAMPLE 9

4-((6-Cyclobutyl-5H-pyrrolo[2,3-b]pyrazin-5-yl)methyl)-7,8-difluoroquinolin-2(lH)- one

Step 1 : 3-Methylpyrazin-2-amine

[0202] A mixture of 2-chloro-3-methylpyrazine (7 g, 54.69 mmol) and 25% NH₃.H₂O (100 mL) was heated to 160⁰C for 24h in a sealed tube. The resulting solution was cooled to RT and extracted with EtOAc (3x200 mL). The organics were combined, dried over Na₂SO₄, and concentrated under vacuum to afford 2.18 g (89%) of 3-methylpyrazin-2-amine as a yellow solid. LCMS: 110 (M+H)⁺.

Step 2: tert-Butyl 3-methylpyrazin-2-ylcarbamate

N .

N NH Boc

[0203] A mixture of 3-methylpyrazin-2-amine (2.18 g, 20.00 mmol) and BoC₂O (8.72 g, 40.00 mmol) in THF (70 mL) was refluxed for 18h. The reaction mixture was cooled to RT and concentrated under vacuum. The residue obtained was purified by column chromatography on silica gel (eluting with 10:1 PE:EA) to afford 1 g (23%) of tert-butyi 3-methylpyrazin-2-ylcarbamate as a white solid. Attorney Docket No. KO 152-401 -PC

tert-Butyl 3 -^-cyclobutyl^-oxoethy^pyrazin^-ylcarbamate

[0204] sec-BuLi (7.5 ml, 4.00 equiv) was added dropwise to a solution of tert- butyl 3-methylpyrazin-2-ylcarbamate (510 mg, 2.44 mmol) in THF (15 rnL) maintained at -50⁰C. The resulting solution was stirred for Ih at -50⁰C and N- methoxy-N-methylcyclobutanecarboxamide (580 mg, 4.06 mmol) in THF (5 mL) was added drpwise. The resulting solution was stirred for an additional 2h at -50⁰C. The reaction mixture was then quenched by the adding of H₂OZiCe (20 mL) and the aqueous layer was extracted with CHCI3 (4x160 mL). The organics were combined, dried over Na₂Sθ4,and concentrated under vacuum. The residue obtained was purified by column chromatography on silica gel (eluting with 500:1 DCM:MeOH) to afford 0.6 g (crude) of tert-butyi 3-(2-cyclobutyl-2-oxoethyl)pyrazin-2-ylcarbamate as yellow oil.

Step 4: 6-Cyclobutyl-5H-pyrrolo[2,3-b]pyrazine

[0205] A mixture of tert-butyl 3-(2-cyclobutyl-2-oxoethyl)pyrazin-2-ylcarbamate (600 mg, 2.06 mmol) and HCl (4 mL) in ethanol (16 mL) was refluxed for Ih. The reaction progress was monitored by TLC (EtOAc/PE = 1 :1). The mixture was concentrated under vacuum. Adjustment of the pH to 8 was accomplished by the addition of Na₂COs. A filtration was performed and the filter cake was washed with H₂O (2x10 mL) and with PE (2x20 mL). This resulted in 300 mg (crude) of 6- cyclobutyl-5H-pyrrolo[2,3-b]pyrazine as a yellow solid. LCMS: 174 (M+H)⁺. Attorney Docket No. KO 152-401 -PC

Step 5 : 4-((6-Cyclobutyl-5H-pyrrolo[2,3-b]pyrazin-5-yl)methyl)-7,8-difluoroquinolin- 2(lH)-one

[0206] 4-((6-cyclobutyl-5H-pyrrolo[2,3-b]pyrazin-5-yl)methyl)-7,8- difluoroquinolin-2(lH)-one was synthesized as described in EXAMPLE 3 using 6- cyclobutyl-5H-pyrrolo[2,3-b]pyrazine and 4-(bromomethyl)-7,8-difluoroquinolin- 2(lH)-one (INTERMEDIATE 1) as starting materials. LCMS: 367 (M+H)⁺.

EXAMPLE 10

4-((6-cyclobutyl-5H-pyrrolo[2,3-b]pyrazin-5-yl)methyl)-8-fluoroquinolin-2(lH)-one

[0207] 4-((6-cyclobutyl-5H-pyrrolo[2,3-b]pyrazin-5-yl)methyl)-8-fluoroquinolin- 2(lH)-one was synthesized as described in EXAMPLE 3 using 6-cyclobutyl-5H- pyrrolo[2,3-b]pyrazine and 4-(bromomethyl)-8-fluoroquinolin-2(lH)-one (INTERMEDIATE 4) as starting materials. ¹H NMR (400 MHz, CDCl₃) δ 8.46 (s, IH), 8.26 (d, IH), 7.60 (d, IH), 7.39 (t, IH), 7.33-7.28 (m, IH), 6.83 (s, IH), 5.66 (s, 2H), 5.63 (s, IH), 3.58 (pent., IH), 2.38-2.29 (m, 4H), 2.13-2.02 (m, IH), 2.01-1.40 (m, IH). LCMS: 349.2 (M+H)⁺. Attorney Docket No. KO 152-401 -PC

EXAMPLE 11

4-((2-cyclobutylpyrazolo[l,5-a]pyridin-3-yl)methyl)-7,8-difluoroquinolin-2(lH)-one

Stepl : 1 -Cyclobutyl-2-(pyridin-2-yl)ethanone

[0208] A solution of n-BuLi (1.9 rnL, 1.6M hexanes, 3.0 mmol) was added dropwise to a solution of 2-methylpyridine (0.3 rnL, 3.0 mmol) and anhydrous THF (3 mL) at -78°C under N₂. The bath was pulled, and the reaction was allowed to warm to rt. The reaction was then cooled to -78 ⁰C, and a solution of JV-methoxy-iV- methylcyclobutanecarboxamide (435 mg, 3.0 mmol) and anhydrous THF (0.6 mL) was added dropwise. The bath was pulled, and the reaction was allowed to warm to rt. After 25 min at rt, silica gel (5 mL) was added, and the reaction was concentrated. Purification by silica gel chromatography (l :0→l :l; hexanes: ethyl acetate) gave 1- cyclobutyl-2-(pyridin-2-yl)ethanone. LCMS: 176.1 (M+H)⁺.

Step 2: 2-Cyclobutylpyrazolo[ 1 ,5-a]pyridine

[0209] A solution of O-(mesitylsulfonyl)hydroxylamine (480 mg, 2.23 mmol) and dichloromethane (2 mL) was added dropwise over 3 min to a solution of 1-cyclobutyl- 2-(pyridin-2-yl)ethanone (390 mg, 2.23mmol) and dichloromethane (4 mL) at 0 ⁰C under N₂. The bath was pulled, and the reaction was allowed to warm to rt. After 5.5 h at rt, silica gel (5 mL) was added, and the reaction was concentrated. Purification by silica gel chromatography (1 :0 — >7:3; hexanes:ethyl acetate) gave 2- cyclobutylpyrazolo[l,5-a]pyridine. LCMS: 173.2 (M+H)⁺. Attorney Docket No. KO 152-401 -PC

Step 3 : 4-((2-Cyclobutylpyrazolo[ 1 ,5-a]pyridin-3-yl)methyl)-7,8-difluoroquinolin- 2(lH)-one

[0210] A suspension of 2-cyclobutylpyrazolo[l,5-a]pyridine (35 mg, 0.20 mmol), 4-(bromomethyl)-7,8-difluoroquinolin-2(lH)-one (INTERMEDIATE 1, 100 mg, 0.36 mmol), and dichloromethane (1.5 mL) in a small vial was agitated for 5 min and then concentrated to a fine powder. This powder was heated in the vial with a standard laboratory heat gun for 4 min. Purification by reverse-phase HPLC (7:3→3:7; water: acetonitrile) gave 4-((2-cyclobutylpyrazolo[l ,5-a]pyridin-3-yl)methyl)-7,8- difluoroquinolin-2(lH)-one. ¹U NMR (400 MHz, DMSO-d₆) δ 11.90 (br s, IH), 8.63 (d, IH), 7.76 (m, IH), 7.48 (d, IH), 7.28 (m, IH), 7.11 (t, IH), 6.79 (t, IH), 5.77 (s, IH), 4.22 (s, 2H), 3.62 (m, IH), 2.36-2.14 (m, 4H), 2.00-1.78 (m, 2H). LCMS: 366.0 (M+H)⁺.

EXAMPLE 12

4-((2-Cyclobutyl-8-methylimidazo[l,2-b]pyridazin-3-yl)methyl)-7,8-difluoroquinolin-

2(lH)-one

Attorney Docket No. KO 152-401 -PC

1 -(4-Methylpyridazin-3 -yl)hydrazine

[0211] A mixture of 3-chloro-4-methylpyridazine (8 g, 62.50 mmol)and hydrazine (2.7 g, 71.72 mmol) in EtOH (100 rnL) was refluxed for 4h. The reaction progress was monitored by TLC (DCM/MeOH = 10:1). The mixture was concentrated under vacuum to afford 5 g of l-(4-methylpyridazin-3-yl)hydrazine as black oil.

4-Methylpyridazin-3 -amine

[0212] l-(4-Methylpyridazin-3-yl)hydrazine (5 g, 40.32 mmol) in EtOH (150 mL) was hydrogenated at RT for 18h in the presence of Raney Ni (2 g). The reaction progress was monitored by TLC (DCM/MeOH = 20: 1). A filtration was performed and the filtrate was concentrated under vacuum. The residue was purified by flash chromatography on silica gel (eluting with 100:1 DCM/MeOH solvent system) to afford 3 g of 4-methylpyridazin-3 -amine as a red solid.

2-Cyclobutyl-8-methylimidazo[ 1 ,2-b]pyridazine

[0213] A mixture of 4-methylpyridazin-3 -amine (2 g, 18.35 mmol) and 2-bromo-l- cyclobutylethanone (3 g, 17.05 mmol, obtained by brominating 1-cyclobutylethanone with Br₂) in EtOH (60 mL) was refluxed for 18 h. The reaction progress was monitored by TLC (EtOAc/PE = 1 :2). The mixture was concentrated by evaporation under vacuum to afford a residue that was purified by flash chromatography on silica gel (eluting with 1 :20 EtOAc/PE solvent system). This resulted in 340 mg of 2- cyclobutyl-8-methylimidazo[l,2-b]pyridazine as light yellow oil. LCMS: 188 (M+H)⁺. Attorney Docket No. KO 152-401 -PC

3-Bromo-2-cyclobutyl-8-methylimidazo[ 1 ,2-b]pyridazine

[0214] A mixture of 2-cyclobutyl-8-methylimidazo[l,2-b]pyridazine (350 mg, 1.87 mmol) and NBS (350 mg, 1.97 mmol) in CHCl₃ (20 mL) was stirred at RT for 0.5h. The reaction progress was monitored by TLC (EtOAc/PE = 1 :5). The solvent was removed and the residue was purified by flash chromatography on silica gel (eluting with 1 :50 EtOAc/PE solvent system) to afford 270 mg of 3-bromo-2-cyclobutyl-8- methylimidazo[l,2-b]pyridazine as a light yellow solid. LCMS: 267 (M+H)⁺.

Step 5: (2-Cyclobutyl-8-methylimidazo[ 1 ,2-b]pyridazin-3-yl)(7,8-difluoro-2- methoxyquinolin-4-yl)methanol

[0215] n-BuLi (2.5M, 0.5 mL) was slowly added to a solution of 3-bromo-2- cyclobutyl-8-methylimidazo[l,2-b]pyridazine (150 mg, 0.57 mmol) in THF (20 mL) cooled -78°C. The resulting solution was stirred at -78°C for 0.5h followed by addition of 7,8-difluoro-2-methoxyquinoline-4-carbaldehyde (INTERMEDIATE 5, 120 mg, 0.54 mmol). The resulting solution was stirred at -78°C for another 2h. The reaction progress was monitored by TLC (EtOAc/PE = 1 :5). The mixture was concentrated under vacuum and the residue was purified by flash chromatography on silica gel (eluting with 1 :20 EtOAc/PE solvent system) to afford 80 mg of (2-cyclobutyl-8- methylimidazo[l,2-b]pyridazin-3-yl)(7,8-difluoro-2-methoxyquinolin-4-yl)methanol as a light yellow solid. LCMS: 411 (M+H)⁺. Attorney Docket No. KO 152-401 -PC

Step 6: 4-((2-Cyclobutyl-8-methylimidazo[ 1 ,2-b]pyridazin-3-yl)methyl)-7,8-difluoro- 2-methoxyquinoline

[0216] A mixture of (2-cyclobutyl-8-methylimidazo[ 1 ,2-b]pyridazin-3-yl)(7,8- difluoro-2-methoxyquinolin-4-yl)methanol (120 mg, 0.29 mmol) and Et₃SiH (15 rnL) in CF3COOH (15 mL) was stirred at 85°C for 18h. The reaction progress was monitored by TLC (EtOAc/PE = 1 :2). The reaction mixture was then quenched by the addition of 150 mL of H₂OZiCe. The pH was adjusted to ~9 by the addition of NH₃ ^»H₂O and the aqueous solution was extracted with EtOAc (3x100 mL). The organics were combined and they were washed with brine (2x100 mL), dried, and evaporated to dryness. The residue was purified by flash chromatography on silica gel (eluting with 1 :20 EtOAc:PE solvent system) to afford 80 mg of 4-((2-cyclobutyl-8- methylimidazo[l ,2-b]pyridazin-3-yl)methyl)-7,8-difluoro-2-methoxyquinoline as a white solid. LCMS: 395 (M+H)⁺.

Step 7: 4-((2-Cyclobutyl-8-methylimidazo[ 1 ,2-b]pyridazin-3-yl)methyl)-7,8- difluoroquinolin-2( 1 H)-one

[0217] 4-((2-Cyclobutyl-8-methylimidazo[l,2-b]pyridazin-3-yl)methyl)-7,8- difluoroquinolin-2(lH)-one was synthesized as described in EXAMPLE 5, Step 4 using 4-((2-cyclobutyl-8-methylimidazo[ 1 ,2-b]pyridazin-3-yl)methyl)-7,8-difluoro-2- methoxyquinoline as a starting material. ¹H NMR (300 MHz, DMSO-d₆) δ 9.17 (s, Attorney Docket No. KO 152-401 -PC

IH), 8.12 (s, IH), 7.66 (s, IH), 7.13-7.08 (m, IH), 6.86 (s, IH), 5.94 (s, IH), 4.48 (s, 2H), 3.74-3.68 (m, IH), 2.75 (s, 3H), 2.58 (m, 2H), 2.31 (m, 2H), 2.00 (m, 2H). LCMS: 3815 (M+H)⁺.

EXAMPLE 13

4-((6-Cyclobutylimidazo[ 1 ,2-b] [ 1 ,2,4]triazin-7-yl)methyl)-7,8-difluoroquinolin-2( 1 H)- one

Step 1 : l,2,4-Triazin-3-amine

[0218] Oxalaldehyde (600 mg, 10.34 mmol) in H₂O (2 rnL) was added to a solution of aminoguanidine bicarbonate (1.4 g, 9.26 mmol) in phosphate buffer (230 mL) and it was stirred at RT for 9h. The reaction progress was monitored by TLC (EtOAc/PE = 3:1). The resulting solution was extracted with EtOAc (8x50 mL) and the organics were combined and concentrated by evaporation under vacuum. The final product was purified by recrystallization from EtOAc/PE (3:1) to afford 0.3 g of l,2,4-triazin-3- amine as a pale yellow solid.

Step 2: 6-Cyclobutylimidazo[ 1 ,2-b] [ 1 ,2,4]triazine

[0219] 6-Cyclobutylimidazo[l,2-b][l,2,4]triazine was synthesized as described in EXAMPLE 12, Step 3 using l,2,4-triazin-3-amine as a starting material. LCMS: 175 (M+H)⁺. Attorney Docket No. KO 152-401 -PC

Step 3: 7-Bromo-6-cyclobutylimidazo[ 1 ,2-b] [ 1 ,2,4]triazine

[0220] In mixture of 6-cyclobutylimidazo[l,2-b][l,2,4]triazine (2.5 g, 12.93 mmol), NBS (2.5 g, 14.04 mmol), and AIBN (70 mg) in CCl₄ (100 niL) was heated to 55°C for 20 min. The reaction progress was monitored by TLC (DCM/MeOH = 50:1). A filtration was performed to remove insolubles. The filtrate was washed with Na₂CO₃ (2x) and with H₂O (2x), dried, and evaporated to dryness. The residue was purified by flash chromatography on silica gel (eluting with 150:1 DCM:MeOH solvent system) to afford 2.5 g of 7-bromo-6-cyclobutylimidazo[l,2-b][l,2,4]triazine as a yellow solid. LCMS: 253 (M+H)⁺.

Step 4: (6-Cyclobutylimidazo[l,2-b][l,2,4]triazin-7-yl)(7,8-difluoro-2- methoxyquinolin-4-yl)methanol

[0221 ] (6-Cyclobutylimidazo[ 1 ,2-b] [1 ,2,4]triazin-7-yl)(7,8-difiuoro-2- methoxyquinolin-4-yl)methanol was synthesized as described in EXAMPLE 12, Step 5 using 7,8-difluoro-2-methoxyquinoline-4-carbaldehyde (INTERMEDIATE 5) and 7- bromo-6-cyclobutylimidazo[l,2-b][l,2,4]triazine as starting materials. LCMS: 398 (M+H)⁺. Attorney Docket No. KO 152-401 -PC

Step 5: 4-((6-Cyclobutylimidazo[l,2-b][l,2,4]triazin-7-yl)methyl)-7,8- difluoroquinolin-2( 1 H)-one

[0222] A mixture of (6-cyclobutylimidazo[l,2-b][l,2,4]triazin-7-yl)(7,8-difluoro-2- methoxyquinolin-4-yl)methanol (250 mg, 0.60 mmol), NaI (950 mg, 6.33 mmol), and TMSCl (680 mg, 6.30 mmol) in ACN (3 ml) was heated to 75°C for Ih. The reaction progress was monitored by TLC (EtOAc/PE = 1 :1). The resulting solution was diluted with 100 mL OfH₂O and it was extracted with EtOAc (4x). The organics were combined, dried over Na₂SO₄, and evaporated to dryness. The residue was purified by flash chromatography on silica gel (eluting with 100:1 DCM/MeOH solvent system) to afford 62 mg of 4-((6-cyclobutylimidazo[l,2-b][l,2,4]triazin-7-yl)methyl)-7,8- difluoroquinolin-2(lH)-one as a yellow solid. ¹U NMR (300 MHz, DMSO-d₆) δ 11.95 (s, IH), 8.60 (d, IH), 8.53 (d, IH), 7.81 (m, IH), 7.33 (m, IH), 5.74 (s, IH), 4.57 (s, 2H), 3.82 (m, IH), 2.42 (m, 2H), 2.24 (m, 2H), 1.95 (m, 2H). LCMS: 368 (M+H)⁺.

EXAMPLE 14

4-((2-Cyclobutylimidazo[l,2-b]pyridazin-3-yl)methyl)-7,8-difluoroquinolin-2(lH)-one

Step 1 : 6-Chloropyridazin-3 -amine

Attorney Docket No. KO 152-401 -PC

[0223] A mixture of 3,6-dichloropyridazine (200 mg, 1.34 mmol) and NH₃.H₂O (25%, 15 rnL) was heated to 125°C for 18h in a sealed tube. The reaction progress was monitored by TLC (DCM:MeOH=10: 1). A filtration was performed and the filter cake was washed with H₂O and it was dried to result in 150 mg (crude) of 6- chloropyridazin-3 -amine as a yellow solid.

Step 2: 6-Chloro-2-cyclobutylimidazo[ 1 ,2-b]pyridazine

[0224] 6-Chloro-2-cyclobutylimidazo[l,2-b]pyridazine was synthesized as described in EXAMPLE 12, Step 3 using 2-bromo-l-cyclobutylethanone (obtained by brominating 1-cyclobutylethanone with Br₂) and 6-chloropyridazin-3 -amine as starting materials. LCMS: 208 (M+H)⁺.

Step 3 : 2-Cyclobutylimidazo[ 1 ,2-b]pyridazine

[0225] 6-Chloro-2-cyclobutylimidazo[l,2-b]pyridazine (2.4 g, 11.48 mmol) in MeOH (40 mL) was hydrogenated in the presence of Pd/C (0.5 g) for 1.5h at RT. The reaction progress was monitored by TLC (DCM/MeOH = 20: 1). The Pd/C was removed and the filtrate was evaporated to dryness. The residue was purified by flash chromatography on silica gel (eluting with 50:1 DCM/MeOH solvent system) to afford 1.9 g of 2-cyclobutylimidazo[l,2-b]pyridazine as a yellow solid. LCMS: 1748 (M+H)⁺.

Step 4: 3-Bromo-2-cyclobutylimidazo[ 1 ,2-b]pyridazine

Attorney Docket No. KO 152-401 -PC

[0227] 3-Bromo-2-cyclobutylimidazo[l,2-b]pyridazine was synthesized as described in EXAMPLE 13, Step 3 using 2-cyclobutylimidazo[l,2-b]pyridazine as a starting material. LCMS: 252 (M+H)⁺.

Step 5: 4-((2-Cyclobutylimidazo[ 1 ,2-b]pyridazin-3-yl)methyl)-7,8-difluoroquinolin- 2(lH)-one

[0228] 4-((2-Cyclobutylimidazo[l,2-b]pyridazin-3-yl)methyl)-7,8- difluoroquinolin-2(lH)-one was synthesized as described in EXAMPLE 12 using 7,8- difluoro-2-methoxyquinoline-4-carbaldehyde (INTERMEDIATE 5) and 3-bromo-2- cyclobutylimidazo[l,2-b]pyridazine in Step 5, (2-cyclobutylimidazo[l,2-b]pyridazin-3- yl)(7,8-difluoro-2-methoxyquinolin-4-yl)methanol in Step 6, and 4-((2- cyclobutylimidazo[ 1 ,2-b]pyridazin-3-yl)methyl)-7,8-difluoro-2-methoxyquinoline in Step 7. ¹H NMR (300 MHz, DMSO-d₆) δ 11.93 (s, IH), 8.44 (m, IH), 7.83 (m, IH), 7.30 (m, IH), 7.19 (m, 2H), 5.60 (s, IH), 4.53 (s, 2H), 3.75 (m, IH), 2.39 (m, 2H), 2.20 (m, 2H), 1.93 (m, 2H). LCMS: 367 (M+H)⁺.

EXAMPLE 15

4-((2-Cyclobutyl-lH-pyrrolo[2,3-b]pyridin-l-yl)methyl)-7,8-difluoroquinolin-2(lH)- one

Attorney Docket No. KO 152-401 -PC

[0229] 4-((2-Cyclobutyl-lH-pyrrolo[2,3-b]pyridin-l-yl)methyl)-7,8- difluoroquinolin-2(lH)-one was synthesized as describe in EXAMPLE 9 using 3- methylpyridin-2-amine as a starting material. ¹H NMR (400 MHz, CD₃OD; HCl salt) δ 8.67 (d, IH), 8.34 (d, IH), 7.84 (m, IH), 7.62 (m, IH), 7.40 (m, IH), 6.97 (s, IH), 5.90 (d, 2H), 5.28 (m, IH), 3.78 (m, IH), 1.85-2.40 (m, 6H). LCMS: 366.0 (M+H)⁺.

EXAMPLE 16

4-((7-bromo-6-cyclobutyl-5H-pyrrolo[2,3-b]pyrazin-5-yl)methyl)-7,8- difluoroquinolin-2( 1 H)-one

Step 1 : 7-bromo-6-cyclobutyl-5H-pyrrolo[2,3-b]pyrazine

[0230] Br₂ (213 mg, 1.33 mmol) in DMF (3 rnL) was added dropwise to a solution of 6-cyclobutyl-5H-pyrrolo[2,3-b]pyrazine (from EXAMPLE 9, Step 4, 232 mg, 1.34 mmol) in DMF (9 mL) and it was stirred at RT for 1.5h. The reaction mixture was then quenched by adding 50 mL of H₂OZiCe. The precipitate was filtered and dried to afford 275 mg of 7-bromo-6-cyclobutyl-5H-pyrrolo[2,3-b]pyrazine as a white solid.

Attorney Docket No. KO 152-401 -PC

Step 2: 4-((7-bromo-6-cyclobutyl-5H-pyrrolo[2,3-b]pyrazin-5-yl)methyl)-7,8- difluoroquinolin-2( 1 H)-one

[0231] 4-((7-bromo-6-cyclobutyl-5H-pyrrolo[2,3-b]pyrazin-5-yl)methyl)-7,8- difluoroquinolin-2(lH)-one was synthesized as described in EXAMPLE 3 using 7- bromo-6-cyclobutyl-5H-pyrrolo[2,3-b]pyrazine and 4-(bromomethyl)-7,8- difluoroquinolin-2(lH)-one (INTERMEDIATE 1) as starting materials. ¹H NMR (400 MHz, DMSO-de) δ 12.07 (s, IH), 8.47 (d, IH), 8.35 (d, IH), 7.84 (m, IH), 7.31 (m, IH), 6.10 (s, 2H), 6.03 (s, IH), 3.95 (m, 3H), 1.80-2.45 (m, 6H). LCMS: 445.4 (M+H)⁺.

EXAMPLE 17

4-((6-Cyclobutyl-7-methyl-5H-pyrrolo[2,3-b]pyrazin-5-yl)methyl)-7,8- difluoroquinolin-2( 1 H)-one

6-Cyclobutyl-7-methyl-5H-pyrrolo[2,3-b]pyrazine

[0232] A mixture of 7-bromo-6-cyclobutyl-5H-pyrrolo[2,3-b]pyrazine (from EXAMPLE 16, Step 1, 300 mg, 1.19 mmol), Pd(dppf)Cl₂ (Cat.),and Zn(CH₃)₂ (2.97 Attorney Docket No. KO 152-401 -PC

rnL, 1.2M) in 1,4-dioxane (20 mL) was heated to 100⁰C for 18h. The reaction mixture was quenched by adding 50 mL of H₂O/ice and it was extracted with EtOAc (2x100 mL) and the organic layers were combined. The residue was purified by flash chromatography on silica gel (eluting with 1 :10 EtOAc:PE solvent system) to afford 170 mg of 6-cyclobutyl-7-methyl-5H-pyrrolo[2,3-b]pyrazine as a white solid.

Step 2: 4-((6-Cyclobutyl-7-methyl-5H-pyrrolo[2,3-b]pyrazin-5-yl)methyl)-7,8- difluoroquinolin-2( 1 H)-one

[0233] 4-((6-Cyclobutyl-7-methyl-5H-pyrrolo[2,3-b]pyrazin-5-yl)methyl)-7,8- difluoroquinolin-2(lH)-one was synthesized as described in EXAMPLE 3 using 6- cyclobutyl-7-methyl-5H-pyrrolo[2,3-b]pyrazine and 4-(bromomethyl)-7,8- difluoroquinolin-2(lH)-one (INTERMEDIATE 1) as starting materials. ¹H NMR (400 MHz, DMSO-d₆; TFA salt) δ 12.02 (s, IH), 8.39 (d, IH), 8.13 (d, IH), 7.87 (m, IH), 7.37 (m, IH), 5.70 (s, 2H), 4.98 (s, IH), 3.80 (m, IH), 2.46 (s, 3H), 1.78-2.30 (m, 6H), 1.35 (t, 3H). LCMS: 381.6 (M+H)⁺.

EXAMPLE 18

4-((2-Cyclobutyl- 1 H-imidazo[4,5 -b]pyridin- 1 -yl)methyl)-7,8-difiuoroquinolin-2( 1 H)- one

Attorney Docket No. KO 152-401 -PC

Step 1 : 2-Cyclobutyl-lH-imidazo[4,5-b]pyridine

[0234] To 1 ,2-diaminopyridine (820 mg, 7.5 mmol) in a 20 mL vial, is added cyclobutanecarboxylic acid (0.73 mL, 7.5 mmol), followed by PPA (3.0 mL, 115% H₃PO₄). The yellow reaction slurry was stirred and heated at 100⁰C for Ih. The resulting dark brown reaction mixture was poured into a stirred, cold sodium hydroxide (30 mL of 2.5 N NaOH) solution. Extractions with DCM (2 x 10 mL) are combined, dried (MgSO₄), filtered, and concentrated. Silica gel chromatography (Gradient: 5% to 10% MeOH/DCM) gave 1.0 g of 2-cyclobutyl-lH-imidazo[4,5-b]pyridine as a beige solid (77%). ¹H NMR (400 MHz, DMSO-d₆) δ 12.40 (s, IH), 8.22 (m, IH), 7.84 (m, IH), 7.14 (m, IH), 3.71 (m, IH), 1.80-2.45 (m, 6H). LCMS: 174.2 (M+H)⁺.

Step 2: 4-((2-Cyclobutyl-lH-imidazo[4,5-b]pyridin-l-yl)methyl)-7,8-difluoroquinolin- 2(lH)-one

[0235] 4-((2-cyclobutyl-lH-imidazo[4,5-b]pyridin-l-yl)methyl)-7,8- difluoroquinolin-2(lH)-one was synthesized as described in EXAMPLE 3 using 2- cyclobutyl-lH-imidazo[4,5-b]pyridine and 4-(bromomethyl)-7,8-difluoroquinolin- 2(lH)-one (INTERMEDIATE 1) as starting materials. Second isomer eluting off HPLC column. ¹H NMR (400 MHz, CD₃OD; TFA salt) δ 8.63 (d, IH), 8.41 (m, IH), 7.80 (m, IH), 7.65 (m, IH), 7.32 (m, IH), 5.97 (s, 2H), 5.52 (s, IH), 4.03 (m, 3H), 1.95-2.70 (m, 6H). LCMS: 366.8 (M+H)⁺. [0236] Attorney Docket No. KO 152-401 -PC

[0237] The following compounds can generally be made using the methods described above. It is expected that these compounds when made will have activity similar to those that have been made in the examples above. [0238] The following compounds are represented herein using the Simplified Molecular Input Line Entry System, or SMILES. SMILES is a modern chemical notation system, developed by David Weininger and Daylight Chemical Information Systems, Inc., that is built into all major commercial chemical structure drawing software packages. Software is not needed to interpret SMILES text strings, and an explanation of how to translate SMILES into structures can be found in Weininger, D., J. Chem. Inf. Comput. Sci. 1988, 28, 31-36. AU SMILES strings used herein, as well as many IUPAC names, were generated using CambridgeSoft's ChemDraw 10.0.

FC1=C(NC(C=C2CN3C(C4CCC4)=NN5C3=NC=C5)=O)C2=CC=C1F

FC1=C(NC(C=C2CN3C(C4CCC4)=CN5C3=NC=C5)=O)C2=CC=C1F

FC1=C(NC(C=C2CN3C(N=CC=C4)=C4C=C3C5CC(F)(F)C5)=O)C2=CC=C1F

FC 1 =C(NC(C=C2CC3=C(C=CC=C4)N4N=C3 C5CC(F)(F)C5)=O)C2=CC=C 1 F

FC1=C(NC(C=C2CC3=C(N=CC=N4)N4N=C3C5CCC5)=O)C2=CC=C1F

FC 1 =C(NC(C=C2CN3C(CC(C)C)=NN4C3=NC=C4)=O)C2=CC=C 1 F

FC1=C(NC(C=C2CN3C(C4CCC4)=CN5C3=NC=C5)=O)C2=CC=C1

FC1=C(NC(C=C2CN3C(N=CC=C4)=C4C=C3C5=CC=CC=C5)=O)C2=CC=C1F

FC1=C(NC(C=C2CC3=C(C=CC=C4C1)N4N=C3C5CC(F)(F)C5)=O)C2=CC=C1F

FC1=C(NC(C=C2CC3=C(N=CC=N4)N4N=C3C5CCC5)=O)C2=CC=C1C

FC1=C(NC(C=C2CN3C(C4CCC4)=NN5C3=NC=C5C)=O)C2=CC=C1F

FC1=C(NC(C(C1)=C2CN3C(C4CCC4)=CN5C3=NC=C5)=O)C2=CC=C1F

FC1=C(NC(C=C2CN3C(N=CC=C4N(C)C)=C4C=C3C5CC(F)(F)C5)=O)C2=CC=C1

F FC1=C(NC(C=C2CC3=C(C=C(C4=CC=CC=C4)C=C5)N5N=C3C6CC(F)(F)C6)=O)

C2=CC=C1F

FC1=C(NC(C=C2CC3=C(N=CC=N4)N4N=C3C5CCC5)=O)C2=C(F)C=C1 FC 1 =C(NC(C=C2CN3C(CCC(C)C)=NN4C3=NC=C4)=O)C2=CC=C 1 F FC1=C(NC(C=C2CN3C(C4CCC4)=CN5C3=NC=C5OC)=O)C2=CC=C1F Attorney Docket No. KO 152-401 -PC

FC1=C(NC(C=C2CN3C(C=CC=N4)=C4CC3C5CC(F)(F)C5)=O)C2=CC=C1F

FC1=C(NC(C=C2CC3=C(CCCC4(C)C)N4N=C3C5CC(F)(F)C5)=O)C2=CC=C1F

FC1=C(NC(C=C2CC3=C(C=CC=N4)N4N=C3C5CCC5)=O)C2=CC=C1

[0239] The activity of the compounds in Examples 1-18 as iNOS inhibitors is illustrated in the following assays. The other compounds listed above, which have not yet been made and/or tested, are predicted to have activity in these assays as well.

Biological Activity Assay

Enzyme Source

[0240] The source of nitric oxide synthase (NOS) enzyme can be generated in several ways including induction of endogenous iNOS using cytokines and/or lipopolysaccharide (LPS) in various cell types known in the art. Alternatively, the gene encoding the enzyme can be cloned and the enzyme can be generated in cells via heterologous expression from a transient or stable expression plasmid with suitable features for protein expression as are known in the art. Enzymatic activity (nitric oxide production) is calcium independent for iNOS, while the constitutive NOS isoforms, nNOS and eNOS, become active with the addition of various cofactors added to cellular media or extract as are well known in the art. Enzymes specified in Table 1 were expressed in HEK293 cells transiently transfected with human iNOS. DAN Assay

[0241] A major metabolic pathway for nitric oxide is to nitrate and nitrite, which are stable metabolites within tissue culture, tissue, plasma, and urine (S Moncada, A Higgs, N Eng J Med 329, 2002 (1993)). Tracer studies in humans have demonstrated that perhaps 50% of the total body nitrate/nitrite originates from the substrate for NO synthesis, L-arginine (PM Rhodes, AM Leone, PL Francis, AD Struthers, S Moncada, Biomed Biophys Res. Commun. 209, 590 (1995); L. Castillo et al, Proc Natl Acad Sci USA 90, 193 (1993). Although nitrate and nitrite are not measures of biologically active NO, plasma and urine samples obtained from subjects after a suitable period of fasting, and optionally after administration of a controlled diet (low nitrate/low Attorney Docket No. KO 152-401 -PC

arginine), allow the use of nitrate and nitrite as an index of NO activity (C Baylis, P Vallance, Curr Opin Nephrol Hypertens 7, 59 (1998)).

[0242] The level of nitrate or nitrite in the specimen can be quantified by any method known in the art which provides adequate sensitivity and reproducibility. A variety of protocols have also been described for detecting and quantifying nitrite and nitrate levels in biological fluids by ion chromatography (e.g., SA Everett et al., J. Chromatogr. 706, 437 (1995); JM Monaghan et al., J. Chromatogr. 770, 143 (1997)), high-performance liquid chromatography (e.g., M KeIm et al., Cardiovasc. Res. 41, 765 (1999)), and capillary electrophoresis (MA Friedberg et al., J. Chromatogr. 781, 491 (1997)). For example, 2,3-diaminonaphthalene reacts with the nitrosonium cation that forms spontaneously from NO to form the fluorescent product lH-naphthotriazole. Using 2,3-diaminonaphthalene ("DAN"), researchers have developed a rapid, quantitative fluorometric assay that can detect from 10 nM to 10 μM nitrite and is compatible with a multi-well microplate format. DAN is a highly selective photometric and fluorometric reagent for Se and nitrite ion. DAN reacts with nitrite ion and gives fluorescent naphthotriazole (MC Carre et al., Analusis 27, 835-838 (1999)). Table 1 provides the test results of various compounds of the subject invention using the DAN assay.

[0243] A specimen can be processed prior to determination of nitrate or nitrite as required by the quantification method, or in order to improve the results, or for the convenience of the investigator. For example, processing can involve centrifuging, filtering, or homogenizing the sample. If the sample is whole blood, the blood can be centrifuged to remove cells and the nitrate or nitrite assay performed on the plasma or serum fraction. If the sample is tissue, the tissue can be dispersed or homogenized by any method known in the art prior to determination of nitrate or nitrite. It may be preferable to remove cells and other debris by centrifugation or another method and to determine the nitrate or nitrite level using only the fluid portion of the sample, or the extracellular fluid fraction of the sample. The sample can also be preserved for later determination, for example by freezing of urine or plasma samples. When appropriate, additives may be introduced into the specimen to preserve or improve its characteristics for use in the nitrate or nitrite assay. Attorney Docket No. KO 152-401 -PC

[0244] The "level" of nitrate, nitrite, or other NO-related product usually refers to the concentration (in moles per liter, micromoles per liter, or other suitable units) of nitrate or nitrite in the specimen, or in the fluid portion of the specimen. However, other units of measure can also be used to express the level of nitrate or nitrite. For example, an absolute amount (in micrograms, milligrams, nanomoles, moles, or other suitable units) can be used, particularly if the amount refers back to a constant amount (e.g., grams, kilograms, milliliters, liters, or other suitable units) of the specimens under consideration. A number of commercially available kits can be used. In certain instances two regioisomers correspond to a single example #, whereby the example # is given in Table 1 with two hiNOS activity values respectively.

Table 1. Biological Activity

Attorney Docket No. KO 152-401 -PC

Pharmacokinetic Assay

Metabolic Stability of Compounds Following Incubation with Male Mouse Liver

Microsomes

[0245] Liver microsomes are subcellular fractions that contain drug-metabolizing enzymes including the cytochrome P450s (CYPs), flavin monooxygenases and UDP glucuronyl transferases. These enzymes require exogenous cofactors for activity in this test system. The co-factors used consist of an NADPH regenerating system (supporting Phase I oxidation) and UDP-GA (supporting Phase II glucuronidation). The following protocol, employing murine liver microsomes, may be used to determine the in vitro metabolic stability of compounds in terms of half life. Preparation:

[0246] Potassium phosphate buffer at pH 7.4 (incubation buffer) was prepared from 110 mL IM KPO₄ monobasic and 390 mL of IM KPO₄ dibasic (500 mL total volume), which was diluted with water to 100 mM prior to addition ofl mM EDTA, 3 mM MgCl₂ and 1.25 mM D-saccharic acid 1,4-lactone monohydrate. May be stored for up to one month at 4°C. One day prior to study, to 200 mL of incubation buffer was added 5 mg alamethicin and 0.21 g bovine serum albumin (BSA), with 2 min of sonication and several hours of stirring. In 15 mL polypropylene screw-cap tubes containing 8.5 mL incubation buffer, 40 μL of compound in DMSO at 1.25 mM was added to yield a 5.88 μM solution (test article). Liver microsomes (male CD-I mouse liver microsomes from, e.g., BD Gentest, Woburn, MA; may be stored at -70⁰C until use) at a concentration of 20 mg/mL were diluted to 5 mg/mL in incubation buffer in a separate tube. Attorney Docket No. KO 152-401 -PC

[0247] A NADPH-generating system was prepared by combining one aliquot each of the following premade ingredients:

1. 0.5 mL/ aliquot of 10 mL of 600 mM of Glucose-6-phosphate stock solution in 100 mM phosphate buffer;

2. 0.5 mL/ aliquot of 10 mL of 120 mM of NADP stock solution in 100 mM phosphate buffer; and

3. 0.5 mL/ aliquot of 10 mL of 120 Units/ml Glucose-6-phosphate dehydrogenase stock solution in 100 mM phosphate buffer, with an equal volume of 200 mM UDP-GA in 10OmM phosphate buffer stock solution prepared the same day. Each of 1-3 may be stored at -8O⁰C in aliquot vial until use.

Half-life determination:

[0248] In an appropriate incubation tube, 425 μL of the incubation buffer containing compound test article at 5.88 μM was combined with 50 μL of the 5 mg/mL diluted liver microsomes and pre-incubated in a 37°C water bath for at least 5 minutes. 25 μL NADPH Generating System was added to the incubation vial, and a 100 μL aliquot immediately removed from each incubation tube and combined with 300 μL of chilled quench reagent (1 :2 water: acetonitrile) containing internal standard. At each subsequent incubation time point (15, 30 or 60 min), an additional 100 μL was removed and quenched. The final concentration of test articles were 5 μM, and final concentration of DMSO solvent in the incubation were 0.4% (v/v). [0249] Metabolic stability was determined based on disappearance half-life. The percent (%) test article remaining was determined at each incubation time-point, relative to the zero-time sample. These results were transformed by natural log, such that first-order decline of substrate concentrations with time produced a straight line. This line was fit with a standard linear regression algorithm, and the slope determined. If data points suggested higher order elimination kinetics (non linear), then the earliest data points were used for slope calculation. Disappearance half-life in minutes (Ty₂) was calculated by multiplying the inverse of this slope by the natural log of 2 {In∑). All data analysis was performed with Microsoft Excel (ver 2007). Results are reported Attorney Docket No. KO 152-401 -PC

in Table 2 below, in which A represents a half life of < 30 min, B represents a half life of >30 min but <120 min, and C represents a half life of > 120 min, wherein said values are generally representative of a mean value across several runs of the same test. NT indicates not tested.

Table 2. Metabolic Stability

Attorney Docket No. KO 152-401 -PC

In Vivo Assays

Murine LPS Challenge

[0250] Inhibition of induction of iNOS can be quantified via the LPS challenge. Inflammation, edema, and the onset of sepsis can be observed following an injection of lipopolysaccharide (LPS), a substance produced by Gram-negative bacteria. Injection of LPS has been shown to induce iNOS transcription, leading to measureable increases in both iNOS and NO. (Iuvone T et al., Evidence that inducible nitric oxide synthase is involved in LPS-mediated plasma leakage in rat skin through the activation of nuclear factor-κB, Br J Pharm 1998:123 1325-1330.) As described above, the level of nitric oxide in the specimen can be quantified by correlation with plasma nitrate or nitrite levels via chemiluminescence, fluorescence, spectophotometric assays, or by any method known in the art which provides adequate sensitivity and reproducibility, including those described above.

[0251] Male Lewis rats weighing 150-250 g are often used in the studies. Rats may be fasted for up to 16 hours prior to the administration of LPS. Free access to water is maintained. Test compounds are administered with LPS or alone. Compounds are dissolved in the vehicle of 0.5% methycele/0.025% Tween 20 or 20% encapsin for oral administration. For the intravenous dosing, compounds are dissolved in saline or 0.5-3%DMSO/20% encapsin. Compounds may also be dissolved in 10% stock solution of 90% PGE-400, 5% Tween 80, 5%PVP + 90% CMC. The dosing volumes are 1-2 ml for oral and 0.3-1 ml for intravenous administration. [0252] LPS is injected intravenously (under anesthesia) or intraperitoneally in sterile 0.9% saline (sodium chloride ) at a dose between 0.1-10 mg/kg in a volume not excess to 1 ml. The needle is 26-30 gauge. Following LPS injection, rats usually exhibit flu-like symptoms, principally involving lack of activity and diarrhea. In routine screening experiments, rats are sacrificed 1.5-6 hr after LPS injection and a terminal bleeding is performed under anesthesia to collect 1-3 ml blood samples and then animals are then euthanized by CO₂.

[0253] A protocol using mice instead of rats was adapted from the methods disclosed above, adjusting for species and weight differences, for example, using Attorney Docket No. KO 152-401 -PC

modifications well known in the art. Compound 9 was administered at 30 mpk, and yielded 41% inhibition of NO production versus control.

[0254] From the foregoing description, one skilled in the art can easily ascertain the essential characteristics of this invention, and without departing from the spirit and scope thereof, can make various changes and modifications of the invention to adapt it to various usages and conditions.

Claims

Attorney Docket No. KO 152-401 -PCCLAIMSWhat is claimed is:

1. A compound of structural Formula II

II or a salt, ester, or prodrug thereof, wherein:

R⁸ has the structure:

at least one of X¹ - X⁶ is a heteroatom;

X³ is selected from the group consisting of C(R¹⁰) and N;

X⁴ is selected from the group consisting of C(R¹¹) and N

X⁶ is selected from the group consisting of C(R¹²), C(R¹²)-C(R¹³), C=C, C(R¹²)-N, N, O, and S;

R¹⁰, R¹¹, and R¹² are each independently selected from the group consisting of a bond, hydrogen, halogen, lower alkyl, lower alkoxy, and lower monocyclic cycloalkyl; Attorney Docket No. KO 152-401 -PC

each R¹³ and R¹⁴ is independently selected from the group consisting of hydrogen, halogen, lower alkyl, and lower monocyclic cycloalkyl; and each n is independently 0 or 1 ; and with the provisos that: the fused ring formed by X¹, X², X⁵, X⁶ , and the carbon bearing R⁹ may be aromatic or aliphatic; and

X¹ and X⁶ may not both be N.

2. The compound as recited in claim 1, having structural Formula III

in or a salt, ester, or prodrug thereof, wherein: at least one of X¹ - X⁶ is a heteroatom;

X³ is selected from the group consisting of C(R¹⁰) and N;

X⁴ is selected from the group consisting of C(R¹¹) and N

X⁶ is selected from the group consisting of C(R¹²), C(R¹²)-C(R¹³), C=C, N, O, and S;

each R¹³ and R¹⁴ is independently selected from the group consisting of hydrogen, halogen, lower alkyl, and lower monocyclic cycloalkyl; and each n is independently 0 or 1 ; and with the proviso that X¹ and X⁶ may not both be N.

3. The compound as recited in claim 1, wherein R⁴ is hydrogen.

4. The compound as recited in claim 3, wherein R⁸ has a structure selected from the group consisting of:

wherein R¹⁵ is selected from the group consisting of hydrogen, halogen, lower alkyl, and lower monocyclic cycloalkyl.

5. A compound selected from the group consisting of Examples 1 to 18.

6. Use of a compound as recited in Claim 1 as a medicament.

7. Use of a compound as recited in Claim 1 as a medicament for the prevention or treatment of a disease or condition ameliorated by the inhibition of iNOS. Attorney Docket No. KO 152-401 -PC

8. A pharmaceutical composition comprising a compound as recited in Claim 1 together with a pharmaceutically acceptable carrier.

9. A method of inhibition of iNOS comprising contacting iNOS with a compound as recited in Claim 1.

10. A method of treatment of a iNOS-mediated disease comprising the administration of a therapeutically effective amount of a compound as recited in Claim 1 to a patient in need thereof.

11. The method as recited in claim 10, wherein said disease is selected from the group consisting of pain, an inflammatory disease, an autoimmune disease, and a respiratory disease.

12. The method as recited in claim 11, wherein said disease is selected from the group consisting of neuropathy, inflammatory pain, nerve injury, peripheral neuropathy, diabetic neuropathy, chronic pain, intractable cancer pain, complex regional pain syndrome, and entrapment neuropathy (carpel tunnel syndrome), neuropathic pain, acute herpes zoster (shingles), postherpetic neuralgia (PHN), chemotherapy- induced neuropathy, HIV neuropathy, ocular pain, post-surgical pain, dental pain/dental extraction, pain resulting from dermal injury, lower back pain, headaches, migraine, tactile allodynia, and hyperalgesia.

13. The method as recited in claim 18, wherein said pain is inflammatory pain.

14. The method as recited in claim 18, wherein said pain is a result of dermal injury, and said dermal injury is a burn.

15. The method as recited in claim 12, wherein said disease is selected from the group consisting of neuropathic pain and postherpetic neuralgia.

16. The method as recited in claim 11, wherein said disease is is an inflammatory disease selected from the group consisting of sepsis, arthritis, rheumatoid arthritis, osteoarthritis, osteoporosis, systemic lupus erythematosus, reflux esophagitis, diarrhea, inflammatory bowel disease, Crohn's disease, gastritis, irritable bowel syndrome, ulcerative colitis, and psoriasis.

17. The method as recited in claim 11, wherein said disease is is a respiratory disease selected from the group consisting of asthma, chronic obstructive pulmonary Attorney Docket No. KO 152-401 -PC

disease, including chronic bronchitis, acute respiratory distress syndrome, pneumonia, pulmonary hypertension, and status asthamticus.

18. A method of treatment of a iNOS-mediated disease comprising the administration of: a. a therapeutically effective amount of a compound as recited in Claim 1; and b. at least one other therapeutic agent.