Provided are compounds and pharmaceutically acceptable salts thereof, their pharmaceutical compositions, their methods of preparation, and methods for their use in the prevention and/or treatment of HIV; including the prevention of the progression of AIDS and general immunosuppression.

FIELD OF THE INVENTION

Compounds, methods and pharmaceutical compositions for the prevention and/or treatment of HIV; including the prevention of the progression of AIDS and general immunosuppression, by administering certain indoleamine 2,3-dioxygenase compounds in therapeutically effective amounts are disclosed. Methods for preparing such compounds and methods of using the compounds and pharmaceutical compositions thereof are also disclosed.

BACKGROUND OF THE INVENTION

Human immunodeficiency virus type 1 (HIV-1) leads to the contraction of acquired immune deficiency disease (AIDS). The number of cases of HIV continues to rise, and currently over twenty-five million individuals worldwide suffer from the virus. Presently, long-term suppression of viral replication with antiretroviral drugs is the only option for treating HIV-1 infection. Indeed, the U.S. Food and Drug Administration has approved twenty-five drugs over six different inhibitor classes, which have been shown to greatly increase patient survival and quality of life. However, additional therapies are still required due to a number of issues including but not limited to undesirable drug-drug interactions; drug-food interactions; non-adherence to therapy; drug resistance due to mutation of the enzyme target; and inflammation related to the immunologic damage caused by the HIV infection.

Currently, almost all HIV positive patients are treated with therapeutic regimens of antiretroviral drug combinations termed, highly active antiretroviral therapy (“HAART”). However, HAART therapies are often complex because a combination of different drugs must be administered often daily to the patient to avoid the rapid emergence of drug-resistant HIV-1 variants. Despite the positive impact of HAART on patient survival, drug resistance can still occur and the survival and quality of life are not normalized as compared to uninfected persons.1Indeed, the incidence of several non-AIDS morbidities and mortalities, such as cardiovascular disease, frailty, and neurocognitive impairment, are increased in HAART-suppressed, HIV-infected subjects.2This increased incidence of non-AIDS morbidity/mortality occurs in the context of, and is potentially caused by, elevated systemic inflammation related to the immunologic damage caused by HIV infection.3,4,5

Sustained successful treatment of the HIV-1-infected patient population with drugs will therefore require the continued development of new and improved drugs with new targets and mechanisms of action including antiretroviral and/or interventions aimed at restoration of the immune system and decreasing the systemic inflammation.

IDO is a monomeric 45 kDa extrahepatic heme-containing dioxygenase which catalyzes the oxidative pyrrole ring cleavage reaction of I-Trp to N-formylkynurenine utilizing molecular oxygen or reactive oxygen species via three proposed reaction mechanisms.6IDO is an enzyme that is the rate limiting step in the kynurenine pathway of tryptophan catabolism. IDO catalyzes the dioxidation of the indole ring of tryptophan (Trp), producing N-formyl-lynurenine (NFK), which is then metabolized by other enzymes into several downstream metabolites such as kynurenine (Kyn) and 3-hydroxy-anthranilate (HAA). The depletion of Trp and accumulation of Kyn and HAA have immunomodulatory activity, typically exemplified by decreased T cell activation and proliferation, enrichment of regulatory CD4+ T cells, and depletion of IL-17-producing CD4+ T cells. IDO activity therefore has a general immunosuppressive impact.

IDO is expressed in response to inflammation and is considered an important counter balance to prevent collateral tissue damaged during prolonged inflammation. IDO expression and activity are elevated during chronic viral infections such as HIV and HCV, chronic bacterial infections, as well as acute conditions such as sepsis. The IDO-mediated shift of Th17 to Treg differentiation of helper T cells likely plays a role in the intestinal immune dysfunction during HIV infection, likely related to the observed elevated systemic inflammation and increased incidence of non-AIDS morbidity/mortality. In addition, IDO activity likely also plays a role in the persistence of pathogens and cancer, and inhibition of IDO may improve clearance mechanism, potentially leading to cure of these chronic diseases. IDO may also play a role in neurological or neuropsychiatric diseases or disorders such as depression by modulating serotonin synthesis or production of excitatory neurotoxins such as kynurenine. As such, pharmacologic inhibition of IDO has application in a broad range of applications from neurology, oncology, and infectious diseases.

It would therefore be an advance in the art to discover IDO inhibitors that effective the balance of the aforementioned properties as a disease modifying therapy in chronic HIV infections to decrease the incidence of non-AIDS morbidity/mortality; and/or an immunotherapy to enhance the immune response to HIV, HBV, HCV and other chronic viral infections, chronic bacterial infections, chronic fungal infections, and to tumors; and/or for the treatment of depression or other neurological/neuropsychiatric disorders.

SUMMARY OF THE INVENTION

In accordance with one embodiment of the present invention, there is provided a compound having the structure of Formula (I):

or a pharmaceutically acceptable salt thereof, wherein:

W is CR4or N;

Y is CR5or N;

V is CR6or N;

A is selected from a group consisting of aryl, heteroaryl, and C3-C8cycloalkyl;

Provided that only one of R7or R8is H,

Or R7and R8are taken together with the nitrogen to which they are attached to form a group consisting of a 4- to 10-membered monocyclic, bicyclic or tricyclic heterocyclic ring, and a 5- to 7-membered monocyclic heteroaryl ring,

R10is selected from the group consisting of monocyclic and bicyclic heteroaryl ring, monocyclic and bicyclic heterocyclic ring, monocyclic and bicyclic aryl ring, (C3-C8)cycloalkyl, (C1-C6)alkyl, (C1-C6)alkyl-Q, wherein R10can be optionally substituted by one or more R16;

Q is selected from the group consisting of monocyclic and bicyclic aryl ring, monocyclic and bicyclic heterocyclic ring, monocyclic and bicyclic heteroaryl ring, —O(C1-C6)alkyl, —CN, and (C3-C8)cycloalkyl;

R11is selected from the group consisting of (C1-C10)alkyl, phenyl, —CF3, —CF2CF3, and —CH2CF3;

R12is selected from the group consisting of —CF3, (C1-C10)alkyl, and (C3-C8)cycloalkyl;

R13is selected from the group consisting of (C1-C6)alkyl, (C3-C8)cycloalkyl, (C2-C6)alkenyl, and (C2-C6)alkynyl;

R14is selected from the group consisting of (C1-C6)alkyl, (C3-C8)cycloalkyl, (C2-C6)alkenyl, and (C2-C6)alkynyl;

R15is selected from the group consisting of (C1-C10)alkyl, (C3-C8)cycloalkyl, (C2-C10)alkenyl, and (C2-C10)alkynyl;

R17is selected from the group consisting of —H, (C1-C6)alkyl, (C2-C6)alkenyl, and (C2-C6)alkynyl;

R18is selected from the group consisting of —H, (C1-C6)alkyl, halo, and trihaloalkyl.

DETAILED DESCRIPTION OF REPRESENTATIVE EMBODIMENTS

Throughout this application, references are made to various embodiments relating to compounds, compositions, and methods. The various embodiments described are meant to provide a variety of illustrative examples and should not be construed as descriptions of alternative species. Rather it should be noted that the descriptions of various embodiments provided herein may be of overlapping scope. The embodiments discussed herein are merely illustrative and are not meant to limit the scope of the present invention.

It is to be understood that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to limit the scope of the present invention. In this specification and in the claims that follow, reference will be made to a number of terms that shall be defined to have the following meanings.

As used herein unless otherwise specified, “alkyl” refers to a monovalent saturated aliphatic hydrocarbyl group having from 1 to 14 carbon atoms and, in some embodiments, from 1 to 6 carbon atoms. “(Cx-Cy)alkyl” refers to alkyl groups having from x to y carbon atoms. The term “alkyl” includes, by way of example, linear and branched hydrocarbyl groups such as methyl (CH3—), ethyl (CH3CH2—), n-propyl (CH3CH2CH2—), isopropyl ((CH3)2CH—), n-butyl (CH3CH2CH2CH2—), isobutyl ((CH3)2CHCH2—), sec-butyl ((CH3)(CH3CH2)CH—), t-butyl ((CH3)3C—), n-pentyl (CH3CH2CH2CH2CH2—), and neopentyl ((CH3)3CCH2—).

“Alkylene” or “alkylene” refers to divalent saturated aliphatic hydrocarbyl groups having from 1 to 10 carbon atoms and, in some embodiments, from 1 to 6 carbon atoms. “(Cu-Cv)alkylene” refers to alkylene groups having from u to v carbon atoms. The alkylene groups include branched and straight chain hydrocarbyl groups. For example, “(C1-C6)alkylene” is meant to include methylene, ethylene, propylene, 2-methypropylene, dimethylethylene, pentylene, and so forth. As such, the term “propylene” could be exemplified by the following structure:

Likewise, the term “dimethylbutylene” could be exemplified, for example, by any of the following structures:

Furthermore, the term “(C1-C6)alkylene” is meant to include such branched chain hydrocarbyl groups as cyclopropylmethylene, which could be exemplified by the following structure:

“Alkenyl” refers to a linear or branched hydrocarbyl group having from 2 to 10 carbon atoms and in some embodiments from 2 to 6 carbon atoms or 2 to 4 carbon atoms and having at least 1 site of vinyl unsaturation (>C═C<). For example, (Cx-Cy)alkenyl refers to alkenyl groups having from x to y carbon atoms and is meant to include for example, ethenyl, propenyl, isopropylene, 1,3-butadienyl, and the like.

“Alkynyl” refers to a linear monovalent hydrocarbon radical or a branched monovalent hydrocarbon radical containing at least one triple bond. The term “alkynyl” is also meant to include those hydrocarbyl groups having one triple bond and one double bond. For example, (C2-C6)alkynyl is meant to include ethynyl, propynyl, and the like.

“Amino” refers to the group —NR21R22, where R21and R22are independently selected from hydrogen, alkyl, alkenyl, alkynyl, aryl, cycloalkyl, heteroaryl, heterocyclic, —SO2-alkyl, —SO2-alkenyl, —SO2-cycloalkyl, —SO2-aryl, —SO2-heteroaryl, and —SO2-heterocyclic, and wherein R21and R22are optionally joined together with the nitrogen bound thereto to form a heterocyclic group. When R21is hydrogen and R22is alkyl, the amino group is sometimes referred to herein as alkylamino. When R21and R22are alkyl, the amino group is sometimes referred to herein as dialkylamino. When referring to a monosubstituted amino, it is meant that either R21or R22is hydrogen but not both. When referring to a disubstituted amino, it is meant that neither R21nor R22are hydrogen.

“Hydroxyamino” refers to the group —NHOH.

“Alkoxyamino” refers to the group —NHO-alkyl wherein alkyl is defined herein.

“Aminocarbonyl” refers to the group —C(O)NR26R27where R26and R27are independently selected from hydrogen, alkyl, alkenyl, alkynyl, aryl, cycloalkyl, heteroaryl, heterocyclic, hydroxy, alkoxy, amino, and acylamino, and where R26and R27are optionally joined together with the nitrogen bound thereto to form a heterocyclic group.

“Aryl” refers to an aromatic group of from 6 to 14 carbon atoms and no ring heteroatoms and having a single ring (e.g., phenyl) or multiple condensed (fused) rings (e.g., naphthyl or anthryl). For multiple ring systems, including fused, bridged, and spiro ring systems having aromatic and non-aromatic rings that have no ring heteroatoms, the term “Aryl” or “Ar” applies when the point of attachment is at an aromatic carbon atom (e.g., 5,6,7,8 tetrahydronaphthalene-2-yl is an aryl group as its point of attachment is at the 2-position of the aromatic phenyl ring).

“AUC” refers to the area under the plot of plasma concentration of drug (not logarithm of the concentration) against time after drug administration.

“EC50” refers to the concentration of a drug that gives half-maximal response.

“IC50” refers to the half-maximal inhibitory concentration of a drug. Sometimes, it is also converted to the pIC50scale (−log IC50), in which higher values indicate exponentially greater potency.

“Clade” refers to a hypothetical construct based on experimental data. Clades are found using multiple (sometimes hundreds) of traits from a number of species (or specimens) and analyzing them statistically to find the most likely phylogenetic tree for the group.

“Cyano” or “nitrile” refers to the group —CN.

“Cycloalkyl” refers to a saturated or partially saturated cyclic group of from 3 to 14 carbon atoms and no ring heteroatoms and having a single ring or multiple rings including fused, bridged, and spiro ring systems. For multiple ring systems having aromatic and non-aromatic rings that have no ring heteroatoms, the term “cycloalkyl” applies when the point of attachment is at a non-aromatic carbon atom (e.g. 5,6,7,8,-tetrahydronaphthalene-5-yl). The term “cycloalkyl” includes cycloalkenyl groups, such as cyclohexenyl. Examples of cycloalkyl groups include, for instance, adamantyl, cyclopropyl, cyclobutyl, cyclohexyl, cyclopentyl, cyclooctyl, cyclopentenyl, and cyclohexenyl. Examples of cycloalkyl groups that include multiple bicycloalkyl ring systems are bicyclohexyl, bicyclopentyl, bicyclooctyl, and the like. Two such bicycloalkyl multiple ring structures are exemplified and named below:

“(Cu-Cv)cycloalkyl” refers to cycloalkyl groups having u to v carbon atoms.

“Spiro cycloalkyl” refers to a 3 to 10 member cyclic substituent formed by replacement of two hydrogen atoms at a common carbon atom in a cyclic ring structure or in an alkylene group having 2 to 9 carbon atoms, as exemplified by the following structure wherein the group shown here attached to bonds marked with wavy lines is substituted with a spiro cycloalkyl group:

“Fused cycloalkyl” refers to a 3 to 10 member cyclic substituent formed by the replacement of two hydrogen atoms at different carbon atoms in a cycloalkyl ring structure, as exemplified by the following structure wherein the cycloalkyl group shown here contains bonds marked with wavy lines which are bonded to carbon atoms that are substituted with a fused cycloalkyl group:

“Haloalkyl” refers to substitution of an alkyl group with 1 to 3 halo groups (e.g., bifluoromethyl or trifluoromethyl).

“Haloalkoxy” refers to substitution of alkoxy groups with 1 to 5 (e.g. when the alkoxy group has at least 2 carbon atoms) or in some embodiments 1 to 3 halo groups (e.g. trifluoromethoxy).

“Human Serum Protein Shift Assay” refers to an HIV assay using a Luciferase Reporter to determine percent inhibition—plC50. The HIV assay makes use of a two-cell co-culture system. In this assay, an infected cell line J4HxB2 and an indicator cell line HOS (delta LTR+luciferase) are co-cultured in the presence and absence of compound. The assay is designed to find inhibitors that prevent the infection of HOS cells by the J4HxB2 cell line. The assay can detect inhibitors of any stage of the HIV infection cycle.

“Hydroxy” or “hydroxyl” refers to the group —OH.

“Heterocyclic” or “heterocycle” or “heterocycloalkyl” or “heterocyclyl” refers to a saturated or partially saturated cyclic group having from 1 to 14 carbon atoms and from 1 to 6 heteroatoms selected from, for example, boron, silicon, nitrogen, sulfur, phosphorus or oxygen and includes single ring and multiple ring systems including fused, bridged, and spiro ring systems. For multiple ring systems having aromatic and/or non-aromatic rings, the terms “heterocyclic”, “heterocycle”, “heterocycloalkyl”, or “heterocyclyl” apply when there is at least one ring heteroatom and the point of attachment is at an atom of a non-aromatic ring (e.g. 1,2,3,4-tetrahydroquinoline-3-yl, 5,6,7,8-tetrahydroquinoline-6-yl, and decahydroquinolin-6-yl). In one embodiment, for example, the nitrogen, phosphorus and/or sulfur atom(s) of the heterocyclic group are optionally oxidized to provide for the N-oxide, phosphinane oxide, sulfinyl, sulfonyl moieties. More specifically the heterocyclyl includes, but is not limited to, tetrahydropyranyl, piperidinyl, piperazinyl, 3-pyrrolidinyl, 2-pyrrolidon-1-yl, morpholinyl, and pyrrolidinyl. A prefix indicating the number of carbon atoms (e.g., C3-C10) refers to the total number of carbon atoms in the portion of the heterocyclyl group exclusive of the number of heteroatoms.

“Fused heterocyclic” or “fused heterocycle” refer to a 3 to 10 member cyclic substituent formed by the replacement of two hydrogen atoms at different carbon atoms in a cycloalkyl ring structure, as exemplified by the following structure wherein the cycloalkyl group shown here contains bonds marked with wavy lines which are bonded to carbon atoms that are substituted with a fused heterocyclic group:

“Compound”, “compounds”, “chemical entity”, and “chemical entities” as used herein refers to a compound encompassed by the generic formulae disclosed herein, any subgenus of those generic formulae, and any forms of the compounds within the generic and subgeneric formulae, including the racemates, stereoisomers, and tautomers of the compound or compounds.

The term “heteroatom” means such atoms as, for example, boron, silicon, nitrogen, oxygen, phosphorous, or sulfur and includes any oxidized form of nitrogen, such as N(O) {N+—O−}, phosphorous, and sulfur such as S(O) and S(O)2, and the quaternized form of any basic nitrogen.

“Oxazolidinone” refers to a 5-membered heterocyclic ring containing one nitrogen and one oxygen as heteroatoms and also contains two carbons and is substituted at one of the two carbons by a carbonyl group as exemplified by any of the following structures, wherein the oxazolidinone groups shown here are bonded to a parent molecule, which is indicated by a wavy line in the bond to the parent molecule:

Oxo” refers to a (═O) group.

“Polymorphism” refers to when two or more clearly different phenotypes exist in the same population of a species where the occurrence of more than one form or morph. In order to be classified as such, morphs must occupy the same habitat at the same time and belong to a panmictic population (one with random mating).

“Protein binding” refers to the binding of a drug to proteins in blood plasma, tissue membranes, red blood cells and other components of blood.

“Protein shift” refers to determining a binding shift by comparing the EC50values determined in the absence and presence of human serum.

“QVT” refers to the amino acids at positions 369, 370, and 371, respectively in the Sp1 fragment of HIV-1 Gag.

“Racemates” refers to a mixture of enantiomers. In an embodiment of the invention, the compounds recited within, or pharmaceutically acceptable salts thereof, are enantiomerically enriched with one enantiomer wherein all of the chiral carbons referred to are in one configuration. In general, reference to an enantiomerically enriched compound or salt, is meant to indicate that the specified enantiomer will comprise more than 50% by weight of the total weight of all enantiomers of the compound or salt.

“Solvate” or “solvates” of a compound refer to those compounds, as defined above, which are bound to a stoichiometric or non-stoichiometric amount of a solvent. Solvates of a compound includes solvates of all forms of the compound. In certain embodiments, solvents are volatile, non-toxic, and/or acceptable for administration to humans in trace amounts. Suitable solvates include water.

“Stereoisomer” or “stereoisomers” refer to compounds that differ in the chirality of one or more stereocenters. Stereoisomers include enantiomers and diastereomers.

“Tautomer” refer to alternate forms of a compound that differ in the position of a proton, such as enol-keto and imine-enamine tautomers, or the tautomeric forms of heteroaryl groups containing a ring atom attached to both a ring —NH— moiety and a ring ═N— moiety such as pyrazoles, imidazoles, benzimidazoles, triazoles, and tetrazoles.

The term ‘atropisomer’ refers to a stereoisomer resulting from an axis of asymmetry. This can result from restricted rotation about a single bond where the rotational barrier is high enough to allow differentiation of the isomeric species up to and including complete isolation of stable non-interconverting diastereomer or enantiomeric species. One skilled in the art will recognize that upon installing a nonsymmetrical Rxto core, the formation of atropisomers is possible. In addition, once a second chiral center is installed in a given molecule containing an atropisomer, the two chiral elements taken together can create diastereomeric and enantiomeric stereochemical species. Depending upon the substitution about the Cx axis, interconversion between the atropisomers may or may not be possible and may depend on temperature. In some instances, the atropisomers may interconvert rapidly at room temperature and not resolve under ambient conditions. Other situations may allow for resolution and isolation but interconversion can occur over a period of seconds to hours or even days or months such that optical purity is degraded measurably over time. Yet other species may be completely restricted from interconversion under ambient and/or elevated temperatures such that resolution and isolation is possible and yields stable species. When known, the resolved atropisomers were named using the helical nomenclature. For this designation, only the two ligands of highest priority in front and behind the axis are considered. When the turn priority from the front ligand 1 to the rear ligand 1 is clockwise, the configuration is P, if counterclockwise it is M.

“Pharmaceutically acceptable salt” refers to pharmaceutically acceptable salts derived from a variety of organic and inorganic counter ions well known in the art and include, by way of example only, sodium, potassium, calcium, magnesium, ammonium, and tetraalkylammonium, and when the molecule contains a basic functionality, salts of organic or inorganic acids, such as hydrochloride, hydrobromide, tartrate, mesylate, acetate, maleate, and oxalate. Suitable salts include those described in P. Heinrich Stahl, Camille G. Wermuth (Eds.), Handbook of Pharmaceutical Salts Properties, Selection, and Use; 2002.

“Patient” or “subject” refers to mammals and includes humans and non-human mammals.

“Treating” or “treatment” of a disease in a patient refers to 1) preventing the disease from occurring in a patient that is predisposed or does not yet display symptoms of the disease; 2) inhibiting the disease or arresting its development; or 3) ameliorating or causing regression of the disease.

Wherever dashed lines occur adjacent to single bonds denoted by solid lines, then the dashed line represents an optional double bond at that position. Likewise, wherever dashed circles appear within ring structures denoted by solid lines or solid circles, then the dashed circles represent one to three optional double bonds arranged according to their proper valence taking into account whether the ring has any optional substitutions around the ring as will be known by one of skill in the art. For example, the dashed line in the structure below could either indicate a double bond at that position or a single bond at that position:

Similarly, ring A below could be a cyclohexyl ring without any double bonds or it could also be a phenyl ring having three double bonds arranged in any position that still depicts the proper valence for a phenyl ring. Likewise, in ring B below, any of X1-X5could be selected from: C, CH, or CH2, N, or NH, and the dashed circle means that ring B could be a cyclohexyl or phenyl ring or a N-containing heterocycle with no double bonds or a N-containing heteroaryl ring with one to three double bonds arranged in any position that still depicts the proper valence:

Where specific compounds or generic formulas are drawn that have aromatic rings, such as aryl or heteroaryl rings, then it will understood by one of still in the art that the particular aromatic location of any double bonds are a blend of equivalent positions even if they are drawn in different locations from compound to compound or from formula to formula. For example, in the two pyridine rings (A and B) below, the double bonds are drawn in different locations, however, they are known to be the same structure and compound:

The present invention includes compounds as well as their pharmaceutically acceptable salts. Accordingly, the word “or” in the context of “a compound or a pharmaceutically acceptable salt thereof” is understood to refer to either: 1) a compound alone or a compound and a pharmaceutically acceptable salt thereof (alternative), or 2) a compound and a pharmaceutically acceptable salt thereof (in combination).

Unless indicated otherwise, the nomenclature of substituents that are not explicitly defined herein are arrived at by naming the terminal portion of the functionality followed by the adjacent functionality toward the point of attachment. For example, the substituent “arylalkyloxycarbonyl” refers to the group (aryl)-(alkyl)-O—C(O)—. In a term such as “—C(Rx)2”, it should be understood that the two Rxgroups can be the same, or they can be different if Rxis defined as having more than one possible identity. In addition, certain substituents are drawn as —RxRy, where the “—” indicates a bond adjacent to the parent molecule and Rybeing the terminal portion of the functionality. Similarly, it is understood that the above definitions are not intended to include impermissible substitution patterns (e.g., methyl substituted with 5 fluoro groups). Such impermissible substitution patterns are well known to the skilled artisan.

Throughout this application, references are made to various embodiments relating to compounds, compositions, and methods. The various embodiments described are meant to provide a variety of illustrative examples and should not be construed as descriptions of alternative species. Rather it should be noted that the descriptions of various embodiments provided herein may be of overlapping scope. The embodiments discussed herein are merely illustrative and are not meant to limit the scope of the present invention.

It is to be understood that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to limit the scope of the present invention. In this specification and in the claims that follow, reference will be made to a number of terms that shall be defined to have the following meanings.

It is further intended that the compounds of the invention are stable. As used herein “stable” refers to a compound that is sufficiently robust to survive isolation to a useful degree of purity from a reaction mixture, and preferably capable of formulation into an efficacious therapeutic agent.

The present invention also includes isomers, or mixed isomers, which by definition are the molecules of identical atomic compositions, but with different bonding arrangements of atoms or orientations of their atoms in space i.e., isomers are two or more different substances with the same molecular formula. Cis and trans geometic isomers of the compound of the present invention are described and may be isolated as a mixture of isomers or as separated isomeric forms. A bond in a structure diagram represented by a wavy line “” or a crossed line “” is intended to indicate that the structure represents the cis or the trans isomer, or a mixture of the cis and trans isomer in any proportion. Isomerism, in the field of clinical pharmacology and pharmacotherapeutics, can differ in their pharmacokinetic and pharmacodynamic which may provide introducing safer and more effective drug alternatives of newer as well as existing drugs.

In accordance with one embodiment of the present invention, there is provided a compound having the structure of Formula (I):

or a pharmaceutically acceptable salt thereof, wherein:

W is CR4or N;

Y is CR5or N;

V is CR6or N;

A is selected from a group consisting of aryl, heteroaryl, and C3-C8cycloalkyl;

Provided that only one of R7or R8is H,

Or R7and R8are taken together with the nitrogen to which they are attached to form a group consisting of a 4- to 10-membered monocyclic, bicyclic or tricyclic heterocyclic ring, and a 5- to 7-membered monocyclic heteroaryl ring,

R10is selected from the group consisting of monocyclic and bicyclic heteroaryl ring, monocyclic and bicyclic heterocyclic ring, monocyclic and bicyclic aryl ring, (C3-C8)cycloalkyl, (C1-C6)alkyl, (C1-C6)alkyl-Q, wherein R10can be optionally substituted by one or more R16;

Q is selected from the group consisting of monocyclic and bicyclic aryl ring, monocyclic and bicyclic heterocyclic ring, monocyclic and bicyclic heteroaryl ring, —O(C1-C6)alkyl, —CN, and (C3-C8)cycloalkyl;

R11is selected from the group consisting of (C1-C10)alkyl, phenyl, —CF3, —CF2CF3, and —CH2CF3;

R12is selected from the group consisting of —CF3, (C1-C10)alkyl, and (C3-C8)cycloalkyl;

R13is selected from the group consisting of (C1-C6)alkyl, (C3-C8)cycloalkyl, (C2-C6)alkenyl, and (C2-C6)alkynyl;

R14is selected from the group consisting of (C1-C6)alkyl, (C3-C8)cycloalkyl, (C2-C6)alkenyl, and (C2-C6)alkynyl;

R15is selected from the group consisting of (C1-C10)alkyl, (C3-C8)cycloalkyl, (C2-C10)alkenyl, and (C2-C10)alkynyl;

R17is selected from the group consisting of —H, (C1-C6)alkyl, (C2-C6)alkenyl, and (C2-C6)alkynyl;

R18is selected from the group consisting of —H, (C1-C6)alkyl, halo, and trihaloalkyl.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I:

or a pharmaceutically acceptable salt thereof, wherein:

W is CR4;

Y is CR5;

V is CR6or N;

A is selected from a group consisting of phenyl, a 5- to 6-membered monocyclic heteroaryl, and (C3—O5)cycloalkyl;

R1is selected from the group consisting of —CO2H, tetrazol-5-yl, —NHSO2R11, —CONHSO2R12,

R2is selected from the group consisting of —H, hydroxyl, halo, (C1-C6)alkyl, and (C1-C6)alkoxy;

R3is selected from the group consisting of —H, halo, (C1-C6)alkyl, and (C1-C6)alkoxy;

R4and R5are independently selected from the group consisting of —H, —CN, —OH, and halo;

R10is selected from the group consisting of a 5- or 6-membered heteroaryl ring, a [5,6]-bicyclic heteroaryl ring, a 4- to 6-membered heterocylic ring, 6-membered aryl ring, (C3-C6)cycloalkyl, (C1-C6)alkyl, and (C1-C6)alkyl-Q, wherein R10can be optionally substituted by one or more R16;

Q is selected from the group consisting of 5- or 6-membered aryl ring, 4- to 6-membered heterocyclic ring, 5- or 6-membered heteroaryl ring, —O(C1-C6)alkyl, —CN, and (C3-C8)cycloalkyl;

R11is selected from the group consisting of (C1-C10)alkyl, phenyl, —CF3, —CF2CF3, and —CH2CF3;

R12is selected from the group consisting of —CF3, (C1-C10)alkyl, and (C3-C8)cycloalkyl;

R17is selected from the group consisting of —H, (C1-C6)alkyl, (C2-C6)alkenyl, and (C2-C6)alkynyl;

R18is selected from the group consisting of —H, (C1-C6)alkyl, halo, and trihaloalkyl.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I:

or a pharmaceutically acceptable salt thereof, wherein:

W is CR4;

Y is CR5;

V is CR6;

is selected from a group consisting of the following structures:

R1is selected from the group consisting of —CO2H, tetrazol-5-yl, —NHSO2R11, —CONHSO2R12, and

R3is selected from the group consisting of —H, —OCH3, —F, —CH3, and —C2C5;

R5is selected from the group consisting of —H and halo;

Q is selected from the group consisting of phenyl ring, thiazole ring, thiophene ring, and furan ring;

R11is selected from the group consisting of (C1-C10)alkyl and —CF3;

R12is selected from the group consisting of (C1-C10)alkyl, (C3-C8)cycloalkyl, and —CF3;

R17is selected from the group consisting of —H, (C1-C6)alkyl, (C2-C6)alkenyl, and (C2-C6)alkynyl.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I:

or a pharmaceutically acceptable salt thereof, wherein:

W is CR4;

Y is CR5;

V is CR6;

is selected from a group consisting of the following structures:

R1is selected from the group consisting of —CO2H, tetrazol-5-yl, —NHSO2R11, and —CONHSO2R12;

R2is selected from the group consisting of —H, —OH, —Cl, —F, and —OCH3;

R6is selected from the group consisting of the following structures:

R9is selected from the group consisting of the following structures:

R10is selected from the group consisting of the following structures:

R12is selected from the group consisting of -Me, —CF3, and -cC3H5.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I:

or a pharmaceutically acceptable salt thereof, wherein:

W is CR4;

Y is CR5;

V is CR6or N;

A is selected from a group consisting of phenyl, a 5- to 6-membered monocyclic heteroaryl, and (C3-C5)cycloalkyl;

R1is selected from the group consisting of —CO2H, tetrazol-5-yl, —NHSO2R11, —CONHSO2R12, and

R2is selected from the group consisting of —H, hydroxyl, halo, (C1-C6)alkyl, and (C1-C6)alkoxy;

R3is selected from the group consisting of —H, halo, (C1-C6)alkyl, and (C1-C6)alkoxy;

Or R7and R8are taken together with the nitrogen to which they are attached to form a group consisting of a 7- to 10-membered bicyclic heterocyclic ring, a 5-to 7-membered monocyclic heterocyclic, and a 5- to 7 membered monocyclic heteroaryl,

R10is selected from the group consisting of a 5- or 6-membered heteroaryl ring, a [5,6]-bicyclic heteroaryl ring, a 4- to 6-membered heterocylic ring, 6-membered aryl ring, (C3-C6)cycloalkyl, (C1-C6)alkyl, and (C1-C6)alkyl-Q, wherein R10can be optionally substituted by one or more R16;

Q is selected from the group consisting of 5- or 6-membered aryl ring, 4- to 6-membered heterocyclic ring, 5- or 6-membered heteroaryl ring, —O(C1-C6)alkyl, —CN, and (C3-C8)cycloalkyl;

R11is selected from the group consisting of (C1-C10)alkyl, phenyl, —CF3, —CF2CF3, and —CH2CF3;

R12is selected from the group consisting of —CF3, (C1-C10)alkyl, and (C3-C8)cycloalkyl;

R18is selected from the group consisting of —H, (C1-C6)alkyl, halo, and trihaloalkyl.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I:

or a pharmaceutically acceptable salt thereof, wherein:

W is CR4;

Y is CR5;

V is CR6;

is selected from a group consisting of the following structures:

R1is selected from the group consisting of —CO2H, tetrazol-5-yl, —NHSO2R11, —CONHSO2R12, and

R3is selected from the group consisting of —H, —OCH3, —F, —CH3, and —C2C5;

R4is selected from the group consisting of —H, halo, and (C1-C6)alkyl;

R5is selected from the group consisting of —H, halo, and (C1-C6)alkyl;

R6is selected from the group consisting of —H and halo;

R7and R8are independently selected from the group consisting of the following structures:

Or R7and R8are taken together with the nitrogen to which they are attached to form a group selected from the following structures:

Q is selected from the group consisting of phenyl ring, thiazole ring, thiophene ring, and furan ring;

R11is selected from the group consisting of (C1-C10)alkyl and —CF3;

R12is selected from the group consisting of (C1-C10)alkyl, (C3-C8)cycloalkyl, and —CF3;

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I:

or a pharmaceutically acceptable salt thereof, wherein:

W is CR4;

Y is CR5;

V is CR6;

is selected from a group consisting of the following structures:

R1is selected from the group consisting of —CO2H, tetrazol-5-yl, —NHSO2R11, and —CONHSO2R12;

R2is selected from the group consisting of —H, —OH, —Cl, —F, and —OCH3;

R4is selected from the group consisting of —H and —F;

R5is selected from the group consisting of —H and —F;

R6is selected from the group consisting of —H, —F, and —Cl;

R7and R8are independently selected from the group consisting of the following structures:

R10is selected from the group consisting of the following structures:

R12is selected from the group consisting of —CH3, —CF3, and -cC3H5.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein A is selected from phenyl or (C3-C5)cycloalkyl.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein A is phenyl.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein A is C3-cycloalkyl.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein A is

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R1is selected from —CO2H or tetrazol-5-yl.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R1is —CO2H

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R1is tetrazol-5-yl.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R2is selected from —H, hydroxyl, halo, (C1-C6)alkyl, or (C1-C6)alkoxy.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R2is selected from —H, —OH, —Cl, —F, —OCH3, —OCF3, —CH3, or —C2C5.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R2is selected from —H, —OH, —Cl, —F, or —OCH3.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R2is —H.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R3is selected from —H, halo, (C1-C6)alkyl, or (C1-C6)alkoxy.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R3is selected from —H, —OCH3, —F, —CH3, or —C2C5.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R3is —H.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein

is selected from

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein W is CR4.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R4is selected from —H or halo.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R4is selected from —H or —F.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R4is —H.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R4is —F.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein W is C—F.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein W is C—H.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein Y is CR5.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R5is selected from —H or halo.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R5is selected from —H or —F.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R5is —H.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R5is —F.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein Y is C—H.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein Y is C—F.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein V is CR6.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R6is selected from —H, (C1-C6)alkyl, or (C2-C6)alkenyl.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R6is selected from —H, -nC4H9, —CH2CHCHCH3or —CHCHCH2CH2CH3.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R6is —H.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R6is -nC4H9.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R6is —CH2CHCHCH3.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R6is —CHCHCH2CH2CH3.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein V is C—H.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein V is C-nC4H9.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein V is C—CH2CHCHCH3.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein V is C—CHCHCH2CH2CH3.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein X is selected from

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein X is

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R7and R8are independently selected from (C1-C6)alkyl or (C3-C8)cycloalkyl.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R7and R8are independently selected from

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R7is

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R7is

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R8is

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R8is

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R7and R8are both

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein X is

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein X is

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein X

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R9is (C1-C10)alkyl.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R9is —CH2CH2CH3.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R9is —CH(CH2CH2CH3)2.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein X is —OCH2CH2CH3.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein X is —OCH(CH2CH2CH3)2.

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R10is selected from

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R10is

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R10

In accordance with another embodiment of the present invention, there is provided a compound having the structure of Formula I above, wherein R10is

The present invention further provides compositions comprising a compound of the invention, or pharmaceutically acceptable salt thereof, and at least one pharmaceutically acceptable carrier.

The present invention further provides methods of modulating activity of indoleamine 2,3-dioxygenase by contacting the indoleamine 2,3-dioxygenase with a compound of the invention, or pharmaceutically acceptable salt thereof.

The present invention further provides methods for the prevention and/or treatment of HIV; including the prevention of the progression of AIDS and general immunosuppression in a patient by administering to the patient an effective amount of a compound of the invention, or pharmaceutically acceptable salt thereof.

The present invention further provides methods of treating cancer, viral infection, bacterial infection, sepsis, macular degeneration, wounds, depression, a neurodegenerative disorder, trauma, age-related cataracts, organ transplant rejection, an autoimmune disease, or the like, in a patient comprising administering to the patient a therapeutically effective amount of a compound of the invention, or pharmaceutically acceptable salt thereof.

The present invention further provides use of the compounds herein for the production of a medicament for use in therapy.

Such compounds of the present invention can exist in particular geometric or stereoisomeric forms. The invention contemplates all such compounds, including cis- and trans-isomers, (−)- and (+)-enantiomers, (R)- and (S)-enantiomers, diastereomers, (D)-isomers, (L)-isomers, the racemic mixtures thereof, and other mixtures thereof, such as enantiomerically or diastereomerically enriched mixtures, as falling within the scope of the invention. Additional asymmetric carbon atoms can be present in a substituent such as an alkyl group. All such isomers, as well as mixtures thereof, are intended to be included in this invention.

Optically active (R)- and (S)-isomers and d and l isomers can be prepared using chiral synthons or chiral reagents, or resolved using conventional techniques. If, for instance, a particular enantiomer of a compound of the present invention is desired, it can be prepared by asymmetric synthesis, or by derivatization with a chiral auxiliary, where the resulting diastereomeric mixture is separated and the auxiliary group cleaved to provide the pure desired enantiomers. Alternatively, where the molecule contains a basic functional group, such as an amino group, or an acidic functional group, such as a carboxyl group, diastereomeric salts can be formed with an appropriate optically active acid or base, followed by resolution of the diastereomers thus formed by fractional crystallization or chromatographic means known in the art, and subsequent recovery of the pure enantiomers. In addition, separation of enantiomers and diastereomers is frequently accomplished using chromatography employing chiral, stationary phases, optionally in combination with chemical derivatization (e.g., formation of carbamates from amines).

In another embodiment of the invention, there is provided a compound of Formula I, wherein the compound or salt of the compound is used in the manufacture of a medicament for use in the treatment immunosuppression in a human.

In another embodiment of the invention, there is provided a pharmaceutical composition comprising a pharmaceutically acceptable diluent and a therapeutically effective amount of a compound as defined in Formula I.

In one embodiment, the pharmaceutical formulation containing a compound of Formula I or a salt thereof is a formulation adapted for parenteral administration. In another embodiment, the formulation is a long-acting parenteral formulation. In a further embodiment, the formulation is a nano-particle formulation.

The present invention is directed to compounds, compositions and pharmaceutical compositions that have utility as novel treatments for immunosuppresion. While not wanting to be bound by any particular theory, it is thought that the present compounds are able to inhibit the enzyme that catalyzes the oxidative pyrrole ring cleavage reaction of I-Trp to N-formylkynurenine utilizing molecular oxygen or reactive oxygen species.

Therefore, in another embodiment of the present invention, there is provided a method for the prevention and/or treatment of HIV; including the prevention of the progression of AIDS and general immunosuppression.

The compounds of Table 1 were synthesized according to the Synthetic Methods, General Schemes, and the Examples described below. Any chemical not directly described are readily prepared by one skilled in the art using available starting materials. In certain embodiments, the compound(s) of the present invention, or a pharmaceutically acceptable salt thereof, is chosen from the compounds set forth in Table 1. Wherein a salt is indicated in Table 1, the present invention also encompasses the free base.

Synthetic Methods

Additionally, the methods of this invention may employ protecting groups which prevent certain functional groups from undergoing undesired reactions. Suitable protecting groups for various functional groups as well as suitable conditions for protecting and deprotecting particular functional groups are well known in the art. For example, numerous protecting groups are described in T. W. Greene and G. M. Wuts, Protecting Groups in Organic Synthesis, Third Edition, Wiley, New York, 1999, and references cited therein.

The starting materials for the following reactions are generally known compounds or can be prepared by known procedures or obvious modifications thereof. For example, many of the starting materials are available from commercial suppliers such as Aldrich Chemical Co. (Milwaukee, Wis., USA), Bachem (Torrance, Calif., USA), Ernka-Chemce or Sigma (St. Louis, Mo., USA). Others may be prepared by procedures, or obvious modifications thereof, described in standard reference texts such as Fieser and Fieser's Reagents for Organic Synthesis, Volumes 1-15 (John Wiley and Sons, 1991), Rodd's Chemistry of Carbon Compounds, Volumes 1-5 and Supplementals (Elsevier Science Publishers, 1989), Organic Reactions, Volumes 1-40 (John Wiley and Sons, 1991), March's Advanced Organic Chemistry, (John Wiley and Sons, 4thEdition), and Larock's Comprehensive Organic Transformations (VCH Publishers Inc., 1989).

Unless specified to the contrary, the reactions described herein take place at atmospheric pressure, generally within a temperature range from −78° C. to 200° C. Further, except as employed in the Examples or as otherwise specified, reaction times and conditions are intended to be approximate, e.g., taking place at about atmospheric pressure within a temperature range of about −78° C. to about 110° C. over a period of about 1 to about 24 hours; reactions left to run overnight average a period of about 16 hours.

Isolation and purification of the chemical entities and intermediates described herein can be effected, if desired, by any suitable separation or purification procedure such as, for example, filtration, extraction, crystallization, column chromatography, thin-layer chromatography or thick-layer chromatography, or a combination of these procedures. Specific illustrations of suitable separation and isolation procedures can be had by reference to the examples herein below. However, other equivalent separation or isolation procedures can also be used.

When desired, the (R)- and (S)-isomers may be resolved by methods known to those skilled in the art, for example by formation of diastereoisomeric salts or complexes which may be separated, for example, by crystallization; via formation of diastereoisomeric derivatives which may be separated, for example, by crystallization, gas-liquid or liquid chromatography; selective reaction of one enantiomer with an enantiomer-specific reagent, for example enzymatic oxidation or reduction, followed by separation of the modified and unmodified enantiomers; or gas-liquid or liquid chromatography in a chiral environment, for example on a chiral support, such as silica with a bound chiral ligand or in the presence of a chiral solvent. Alternatively, a specific enantiomer may be synthesized by asymmetric synthesis using optically active reagents, substrates, catalysts or solvents, or by converting one enantiomer to the other by asymmetric transformation.

EXAMPLES

The following examples serve to more fully describe the manner of making and using the above-described invention. It is understood that these examples in no way serve to limit the true scope of the invention, but rather are presented for illustrative purposes. In the examples and the synthetic schemes below, the following abbreviations have the following meanings. If an abbreviation is not defined, it has its generally accepted meaning.ACN=acetonitrileAIBN=azobisisobutyronitrileaq.=aqueousμL or uL=microlitersμM or uM=micromolarNMR=nuclear magnetic resonanceboc=tert-butoxycarbonylbr=broadCbz=BenzyloxycarbonylCDI=1,1′-carbonyldiimidazoled=doubletδ=chemical shift° C.=degrees celciusDCM=dichloromethanedd=doublet of doubletsDHP=dihydropyranDIAD=diisopropyl azodicarboxylateDIEA or DIPEA=N,N-diisopropylethylamineDMAP=4-(dimethylamino)pyridineDMEM=Dulbeco's Modified Eagle's MediumDMF=dimethylformamideEtOAc=ethyl acetateh or hr=hoursHATU=1-[Bis(dimethylamino)methylene]-1H-1,2,3-triazolo[4,5-b]pyridinium 3-oxid hexafluorophosphateHCV=hepatitus C virusHPLC=high performance liquid chromatographyHz=hertzIU=International UnitsIC50=inhibitory concentration at 50% inhibitionJ=coupling constant (given in Hz unless otherwise indicated)KHMDS=potassium bis(trimethylsilyl)amideLCMS=liquid chromatography-mass spectrometrym=multipletM=molarM+H+=parent mass spectrum peak plus H+MeOH=methanolmg=milligrammin=minutesmL=millilitermM=millimolarmmol=millimoleMS=mass spectrumMTBE=methyl tert-butyl etherN=normalNFK=N-formylkynurenineNBS=N-bromosuccinimidenm=nanomolarNMP=N-methyl-2-pyrrolidonePE=petroleum etherppm=parts per millionq.s.=sufficient amounts=singletRT=room temperatureRf=retardation factorsat.=saturatedt=tripletTEA=triethylaminetetrakis=tetrakis(triphenylphosphine)palladium(0)TFA=trifluoroacetic acidTFAA=trifluoroacetic anhydrideTHF=tetrahydrofuran

Equipment Description

1H NMR spectra were recorded on a Bruker Ascend 400 spectrometer or Varian 400 spectrometer. Chemical shifts are expressed in parts per million (ppm, δ units). Coupling constants are in units of hertz (Hz). Splitting patterns describe apparent multiplicities and are designated as s (singlet), d (doublet), t (triplet), q (quartet), quint (quintet), m (multiplet), br (broad).

The analytical low-resolution mass spectra (MS) were recorded on Waters ACQUITY UPLC with SQ Detectors using a Waters BEH C18, 2.1×50 mm, 1.7 μm using a gradient elution method.

30% B for 0.5 min followed by 30-100% B over 2.5 min.

Schemes and Experimental Procedures

The following schemes and procedures illustrate how compounds of the present invention can be prepared. The specific solvents and reaction conditions referred to are also illustrative and are not intended to be limiting. Compounds not described are either commercially available or are readily prepared by one skilled in the art using available starting materials. The Examples disclosed herein are for illustrative purposes only and are not intended to limit the scope of the invention. All examples exhibited IDO EC50values between 700 nM and 1 nM using the assay disclosed herein.

Step A

Step B

Step C

Step D

To a solution of cis-ethyl 2-(4-(cyclohexyl(isobutyl)amino)-3-nitrophenyl)cyclopropanecarboxylate (750 mg, 1.9 mmol) in EtOH/H2O (30 mL/30 mL) at 0° C. was added NH4Cl (2.08 g, 38.8 mmol), followed by the addition of zinc powder (1.27 g, 19.4 mmol) in one portion. The resulting suspension was stirred at room temperature for 2 hrs. Excess zinc powder was filtered off and the filtrate was partitioned between EtOAc (100 mL) and water (30 mL). The layers were separated and the organic layer was washed with brine, dried over Na2SO4and concentrated to give cis-ethyl 2-(3-amino-4-(cyclohexyl(isobutyl)amino)phenyl)cyclopropanecarboxylate (700 mg, quant.) as a brown oil. LCMS (M+H)+: m/z=359.1.

Step A

Step B

Synthesis of methyl 3′-amino-4′-(cyclohexyl(isobutyl)amino)-[1,1′-biphenyl]-2-carboxylate

Step A

To a solution of 4-bromo-1-fluoro-2-nitrobenzene (60 g, 0.27 mol), N-isobutylcyclohexanamine (52 g, 0.33 mol) in 1-methyl-2-pyrrolidinone (300 mL) was added DIEA (70 g, 0.55 mol) at room temperature. The resulting mixture was stirred at 120° C. overnight, then cooled and quenched with water (350 mL). The layers were separated and the aqueous layer was extracted with EtOAc (3×300 mL). The combined organic layers were washed with brine (800 mL), dried over Na2SO4and concentrated under reduced pressure to give a red solid which was subject to trituration with petroleum ether (200 mL). The precipitate was collected by filtration to give 4-bromo-N-cyclohexyl-N-isobutyl-2-nitroaniline (74 g, 77%) as a red solid.

Step B

Step C

Synthesis of N4-cyclohexyl-N4-isobutyl-2′-(1H-tetrazol-5-yl)-[1,1′-biphenyl]-3,4-diamine

Step A

A mixture of 4-bromo-N-cyclohexyl-N-isobutyl-2-nitroaniline (20.0 g, 56 mmol), (2-(1H-tetrazol-5-yl)phenyl)boronic acid (10.6 g, 56 mmol), Pd(PPh3)4(6.4 g, 5.6 mmol), K2CO3(23.4 g, 168 mmol) in DMF/H2O (400 mL/40 mL) was purged with N2(3×) and stirred at 95° C. under N2atmosphere overnight. The reaction was cooled to room temperature and filtered through celite to remove the solids. The filtrate was concentrated and was partitioned between EtOAc (70 mL) and water (25 mL). The layers were separated and the organic layer was washed with brine (25 mL), dried over Na2SO4, and concentrated. The residue was purified by silica gel chromatography (5-30% EtOAc/PE) to give N-cyclohexyl-N-isobutyl-3-nitro-2′-(1H-tetrazol-5-yl)-[1,1′-biphenyl]-4-amine (15 g, 64%) as yellow solid. LCMS (M−H)−: m/z=421.2.

Step B

Step A

Step B

Step C

Step D

Step E

Synthesis of (E)-5-bromo-3-(but-2-en-1-yl)-2-propoxyaniline

Step A

Step B

A solution of 4-bromo-1-(but-3-en-2-yloxy)-2-nitrobenzene (30 g, 0.11 mol) in diglyme (160 mL) was stirred at 150° C. overnight, then cooled down to room temperature and partitioned between DCM (300 mL) and water (100 mL). The layers were separated and the organic layer was washed with brine (100 mL), dried over Na2SO4, and concentrated. The residue was purified by silica gel chromatography (5-15% EtOAc/PE) to give (E)-4-bromo-2-(but-2-en-1-yl)-6-nitrophenol (27 g, 90%) as a yellow oil. LCMS (M+H)+: m/z=272.1.

Step C

Step D

Step A

Step B

Step A

Step B

Step C

Step A

To a solution of 2,4-dibromo-6-nitrophenol (5.0 g, 17 mmol), 4-heptanol (79 mg, 17 mmol), PPh3(215 mg, 20.5 mml) in THF (50 mL) at 0° C. was added diisopropyl azodicarboxylate (4.15 g, 20.5 mmol) in THF (10 mL) dropwise. The resulting mixture was stirred at 50° C. overnight under N2. After the reaction was cooled to room temperature, it was quenched with water (40 mL), and extracted with EtOAc (50 mL×2). The combined organics were washed with brine (40 mL), dried over Na2SO4, and concentrated. The residue was purified by silica gel chromatography (2-10% EtOAc/PE) to give 1,5-dibromo-2-(heptan-4-yloxy)-3-nitrobenzene (3.7 g, 77%) as a yellow oil.

Step B

At 0° C., to a solution of 1,5-dibromo-2-(heptan-4-yloxy)-3-nitrobenzene (300 mg, 0.76 mol) in EtOH (5 mL) and H2O (5 mL) were added NH4Cl (812 mg, 15 mol) and zinc powder (497 mg, 7.59 mol). The resulting mixture was stirred at 25° C. for 2 hrs. After completion, the excess zinc powder was filtered off. The filtrate was concentrated and partitioned between EtOAc (20 mL) and water (6 mL). The layers were separated and the organic layer was washed with brine (10 mL), dried over Na2SO4, and concentrated. The residue was purified by silica gel chromatography (5-20% EtOAc/PE) to give 3,5-dibromo-2-(heptan-4-yloxy)aniline (283 mg, quantitative) as a yellow oil. LCMS (M+2)+: m/z=366.5.

Step C

To a solution of 3,5-dibromo-2-(heptan-4-yloxy)aniline (283 mg, 0.77 mmol) in o-xylene (2 mL) was added 2-chlorobenzo[d]oxazole (118 mg, 0.77 mmol). The resulting mixture was stirred at 100° C. for 1 hr. After the reaction was cooled down to room temperature, it was partitioned between EtOAc (10 mL) and water (5 mL). The layers were separated and the organic layer was washed with brine (5 mL), dried over Na2SO4, and concentrated. The residue was purified by silica gel chromatography (10-30% EtOAc/PE) to give N-(3,5-dibromo-2-(heptan-4-yloxy)phenyl)benzo[d]oxazol-2-amine (260 mg, 70% yield). LCMS (M+H)+: m/z=481.3.

Step D

Step A

To methyl 3′-amino-4′-(cyclohexyl(isobutyl)amino)-[1,1′-biphenyl]-2-carboxylate (160 mg, 0.420 mmol) in DCM (2 mL) was added O,O-di(pyridin-2-yl) carbonothioate (117 mg, 0.505 mmol) and the mixture was stirred at room temperature for 2 hours. The reaction was concentrated and used in the next step without further purification.

Step B

To a mixture of methyl 4′-(cyclohexyl(isobutyl)amino)-3′-isothiocyanato-[1,1′-biphenyl]-2-carboxylate in ethanol (1 ml) was added cyclopropanecarbohydrazide (50.5 mg, 0.505 mmol) and the mixture was stirred at 50° C. overnight. The reaction mixture was concentrated and used in the next step without further purification.

Step C

Methyl 4′-(cyclohexyl(isobutyl)amino)-3′-(2-(cyclopropanecarbonyl)hydrazinecarbothioamido)-[1,1′-biphenyl]-2-carboxylate was treated with sulfuric acid (0.701 mL, 8.41 mmol) and the mixture was stirred for 1 h. After 1 h, reaction mixture was cooled and poured over ice-water and extracted with ethyl acetate, dried over sodium sulfate and concentrate. Crude product was used in the next step.

Step D

Step A

To N4-cyclohexyl-N4-isobutyl-2′-(1H-tetrazol-5-yl)-[1,1′-biphenyl]-3,4-diamine (150 mg, 0.384 mmol) in ACN (4 mL) was added di(1H-imidazol-1-yl)methanethione (137 mg, 0.768 mmol) and stirred at room temperature for 2 hours. The reaction mixture was concentrated and used in the next step without further purification.

Step B

A mixture of N-cyclohexyl-N-isobutyl-3-isothiocyanato-2′-(1H-tetrazol-5-yl)-[1,1′-biphenyl]-4-amine in ethanol (4.0 mL) was treated with cyclopropanecarbohydrazide (77 mg, 0.768 mmol) and the mixture was stirred at 50° C. overnight. The reaction mixture was concentrated and used in the next step without further purification.

Step C

Administration and Formulation

In another embodiment, there is provided a pharmaceutical composition comprising a pharmaceutically acceptable diluent and a therapeutically effective amount of a compound of Formula I or a pharmaceutically acceptable salt thereof.

The compounds of the present invention can be supplied in the form of a pharmaceutically acceptable salt. The terms “pharmaceutically acceptable salt” refer to salts prepared from pharmaceutically acceptable inorganic and organic acids and bases. Accordingly, the word “or” in the context of “a compound or a pharmaceutically acceptable salt thereof” is understood to refer to either a compound or a pharmaceutically acceptable salt thereof (alternative), or a compound and a pharmaceutically acceptable salt thereof (in combination).

As used herein, the term “pharmaceutically acceptable” refers to those compounds, materials, compositions, and dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, or other problem or complication. The skilled artisan will appreciate that pharmaceutically acceptable salts of compounds according to Formula I may be prepared. These pharmaceutically acceptable salts may be prepared in situ during the final isolation and purification of the compound, or by separately reacting the purified compound in its free acid or free base form with a suitable base or acid, respectively.

Illustrative pharmaceutically acceptable inorganic base salts of the compounds of the present invention include metallic ions. More preferred metallic ions include, but are not limited to, appropriate alkali metal salts, alkaline earth metal salts and other physiological acceptable metal ions. Salts derived from inorganic bases include aluminum, ammonium, calcium, copper, ferric, ferrous, lithium, magnesium, manganic salts, manganous, potassium, sodium, zinc, and the like and in their usual valences. Exemplary base salts include aluminum, calcium, lithium, magnesium, potassium, sodium and zinc. Other exemplary base salts include the ammonium, calcium, magnesium, potassium, and sodium salts. Still other exemplary base salts include, for example, hydroxides, carbonates, hydrides, and alkoxides including NaOH, KOH, Na2CO3, K2CO3, NaH, and potassium-t-butoxide.

All of the above salts can be prepared by those skilled in the art by conventional means from the corresponding compound of the present invention. For example, the pharmaceutically acceptable salts of the present invention can be synthesized from the parent compound which contains a basic or acidic moiety by conventional chemical methods. Generally, such salts can be prepared by reacting the free acid or base forms of these compounds with a stoichiometric amount of the appropriate base or acid in water or in an organic solvent, or in a mixture of the two; generally, nonaqueous media like ether, ethyl acetate, ethanol, isopropanol, or acetonitrile are preferred. The salt may precipitate from solution and be collected by filtration or may be recovered by evaporation of the solvent. The degree of ionisation in the salt may vary from completely ionised to almost non-ionised. Lists of suitable salts are found inRemington's Pharmaceutical Sciences,17th ed., Mack Publishing Company, Easton, Pa., 1985, p. 1418, the disclosure of which is hereby incorporated by reference only with regards to the lists of suitable salts.

The compounds of the invention may exist in both unsolvated and solvated forms. The term ‘solvate’ is used herein to describe a molecular complex comprising the compound of the invention and one or more pharmaceutically acceptable solvent molecules, for example, ethanol. The term ‘hydrate’ is employed when said solvent is water. Pharmaceutically acceptable solvates include hydrates and other solvates wherein the solvent of crystallization may be isotopically substituted, e.g. D2O, d6-acetone, d6-DMSO.

Compounds of Formula I containing one or more asymmetric carbon atoms can exist as two or more stereoisomers. Where a compound of Formula I contains an amidoxime or alkenyl or alkenylene group or a cycloalkyl group, geometric cis/trans (or Z/E) isomers are possible. Where the compound contains, for example, a keto or oxime group or an aromatic moiety, tautomeric isomerism (‘tautomerism’) can occur. It follows that a single compound may exhibit more than one type of isomerism.

Included within the scope in some embodiments or alternate embodiements of the claimed compounds in the present invention are all stereoisomers, geometric isomers and tautomeric forms of the compounds of Formula I, including compounds exhibiting more than one type of isomerism, and mixtures of one or more thereof. Also included are acid addition or base salts wherein the counterion is optically active, for example, D-lactate or L-lysine, or racemic, for example, DL-tartrate or DL-arginine.

Cis/trans isomers may be separated by conventional techniques well known to those skilled in the art, for example, chromatography and fractional crystallisation.

Conventional techniques for the preparation/isolation of individual enantiomers include chiral synthesis from a suitable optically pure precursor or resolution of the racemate (or the racemate of a salt or derivative) using, for example, chiral high pressure liquid chromatography (HPLC).

Alternatively, the racemate (or a racemic precursor) may be reacted with a suitable optically active compound, for example, an alcohol, or, in the case where in some embodiements or alternate embodiments the compounds of Formula I contains an acidic or basic moiety, an acid or base such as tartaric acid or 1-phenylethylamine. The resulting diastereomeric mixture may be separated by chromatography and/or fractional crystallization and one or both of the diastereoisomers converted to the corresponding pure enantiomer(s) by means well known to a skilled person.

Chiral compounds of the invention (and chiral precursors thereof) may be obtained in enantiomerically-enriched form using chromatography, typically HPLC, on a resin with an asymmetric stationary phase and with a mobile phase consisting of a hydrocarbon, typically heptane or hexane, containing from 0 to 50% isopropanol, typically from 2 to 20%, and from 0 to 5% of an alkylamine, typically 0.1% diethylamine. Concentration of the eluate affords the enriched mixture.

Mixtures of stereoisomers may be separated by conventional techniques known to those skilled in the art. [see, for example, “Stereochemistry of Organic Compounds” by E L Eliel (Wiley, New York, 1994).]

The present invention includes all pharmaceutically acceptable isotopically-labelled compounds of Formula I wherein one or more atoms are replaced by atoms having the same atomic number, but an atomic mass or mass number different from the atomic mass or mass number usually found in nature.

Certain isotopically-labelled compounds of Formula I, for example, those incorporating a radioactive isotope, are useful in drug and/or substrate tissue distribution studies. The radioactive isotopes tritium, i.e.3H, and carbon-14, i.e.14C, are particularly useful for this purpose in view of their ease of incorporation and ready means of detection.

Isotopically-labelled compounds of Formula I can generally be prepared by conventional techniques known to those skilled in the art or by processes analogous to those described in the accompanying Examples and Preparations using an appropriate isotopically-labelled reagents in place of the non-labelled reagent previously employed.

The compounds of the present invention may be administered as prodrugs. Thus, certain derivatives of compounds of Formula I, which may have little or no pharmacological activity themselves can, when administered into or onto the body, be converted into compounds of Formula I as ‘prodrugs’.

Administration of the chemical entities described herein can be via any of the accepted modes of administration for agents that serve similar utilities including, but not limited to, orally, sublingually, subcutaneously, intravenously, intranasally, topically, transdermally, intraperitoneally, intramuscularly, intrapulmonarilly, vaginally, rectally, or intraocularly. In some embodiments, oral or parenteral administration is used.

Pharmaceutical compositions or formulations include solid, semi-solid, liquid and aerosol dosage forms, such as, e.g., tablets, capsules, powders, liquids, suspensions, suppositories, aerosols or the like. The chemical entities can also be administered in sustained or controlled release dosage forms, including depot injections, osmotic pumps, pills, transdermal (including electrotransport) patches, and the like, for prolonged and/or timed, pulsed administration at a predetermined rate. In certain embodiments, the compositions are provided in unit dosage forms suitable for single administration of a precise dose.

The chemical entities described herein can be administered either alone or more typically in combination with a conventional pharmaceutical carrier, excipient or the like (e.g., mannitol, lactose, starch, magnesium stearate, sodium saccharine, talcum, cellulose, sodium crosscarmellose, glucose, gelatin, sucrose, magnesium carbonate, and the like). If desired, the pharmaceutical composition can also contain minor amounts of nontoxic auxiliary substances such as wetting agents, emulsifying agents, solubilizing agents, pH buffering agents and the like (e.g., sodium acetate, sodium citrate, cyclodextrine derivatives, sorbitan monolaurate, triethanolamine acetate, triethanolamine oleate, and the like). Generally, depending on the intended mode of administration, the pharmaceutical composition will contain about 0.005% to 95%; in certain embodiments, about 0.5% to 50% by weight of a chemical entity. Actual methods of preparing such dosage forms are known, or will be apparent, to those skilled in this art; for example, seeRemington's Pharmaceutical Sciences, Mack Publishing Company, Easton, Pa.

In certain embodiments, the compositions will take the form of a pill or tablet and thus the composition will contain, along with the active ingredient, a diluent such as lactose, sucrose, dicalcium phosphate, or the like; a lubricant such as magnesium stearate or the like; and a binder such as starch, gum acacia, polyvinylpyrrolidine, gelatin, cellulose, cellulose derivatives or the like. In another solid dosage form, a powder, marume, solution or suspension (e.g., in propylene carbonate, vegetable oils or triglycerides) is encapsulated in a gelatin capsule.

Liquid pharmaceutically administrable compositions can, for example, be prepared by dissolving, dispersing, etc. at least one chemical entity and optional pharmaceutical adjuvants in a carrier (e.g., water, saline, aqueous dextrose, glycerol, glycols, ethanol or the like) to form a solution or suspension. Injectables can be prepared in conventional forms, either as liquid solutions or suspensions, as emulsions, or in solid forms suitable for dissolution or suspension in liquid prior to injection. The percentage of chemical entities contained in such parenteral compositions is highly dependent on the specific nature thereof, as well as the activity of the chemical entities and the needs of the subject. However, percentages of active ingredient of 0.01% to 10% in solution are employable, and will be higher if the composition is a solid which will be subsequently diluted to the above percentages. In certain embodiments, the composition will comprise from about 0.2 to 2% of the active agent in solution.

Pharmaceutical compositions of the chemical entities described herein may also be administered to the respiratory tract as an aerosol or solution for a nebulizer, or as a microfine powder for insufflation, alone or in combination with an inert carrier such as lactose. In such a case, the particles of the pharmaceutical composition have diameters of less than 50 microns, in certain embodiments, less than 10 microns.

In general, the chemical entities provided will be administered in a therapeutically effective amount by any of the accepted modes of administration for agents that serve similar utilities. The actual amount of the chemical entity, i.e., the active ingredient, will depend upon numerous factors such as the severity of the disease to be treated, the age and relative health of the subject, the potency of the chemical entity used the route and form of administration, and other factors. The drug can be administered more than once a day, such as once or twice a day.

Therapeutically effective amounts of the chemical entities described herein may range from approximately 0.01 to 200 mg per kilogram body weight of the recipient per day; such as about 0.01-100 mg/kg/day, for example, from about 0.1 to 50 mg/kg/day. Thus, for administration to a 70 kg person, the dosage range may be about 7-3500 mg per day.

In general, the chemical entities will be administered as pharmaceutical compositions by any one of the following routes: oral, systemic (e.g., transdermal, intranasal or by suppository), or parenteral (e.g., intramuscular, intravenous or subcutaneous) administration. In certain embodiments, oral administration with a convenient daily dosage regimen that can be adjusted according to the degree of affliction may be used. Compositions can take the form of tablets, pills, capsules, semisolids, powders, sustained release formulations, solutions, suspensions, elixirs, aerosols, or any other appropriate compositions. Another manner for administering the provided chemical entities is inhalation.

The choice of formulation depends on various factors such as the mode of drug administration and bioavailability of the drug substance. For delivery via inhalation the chemical entity can be formulated as liquid solution, suspensions, aerosol propellants or dry powder and loaded into a suitable dispenser for administration. There are several types of pharmaceutical inhalation devices-nebulizer inhalers, metered dose inhalers (MDI) and dry powder inhalers (DPI). Nebulizer devices produce a stream of high velocity air that causes the therapeutic agents (which are formulated in a liquid form) to spray as a mist that is carried into the patient's respiratory tract. MDIs typically are formulation packaged with a compressed gas. Upon actuation, the device discharges a measured amount of therapeutic agent by compressed gas, thus affording a reliable method of administering a set amount of agent. DPI dispenses therapeutic agents in the form of a free flowing powder that can be dispersed in the patient's inspiratory air-stream during breathing by the device. In order to achieve a free flowing powder, the therapeutic agent is formulated with an excipient such as lactose. A measured amount of the therapeutic agent is stored in a capsule form and is dispensed with each actuation.

Recently, pharmaceutical compositions have been developed for drugs that show poor bioavailability based upon the principle that bioavailability can be increased by increasing the surface area i.e., decreasing particle size. For example, U.S. Pat. No. 4,107,288 describes a pharmaceutical formulation having particles in the size range from 10 to 1,000 nm in which the active material is supported on a cross-linked matrix of macromolecules. U.S. Pat. No. 5,145,684 describes the production of a pharmaceutical formulation in which the drug substance is pulverized to nanoparticles (average particle size of 400 nm) in the presence of a surface modifier and then dispersed in a liquid medium to give a pharmaceutical formulation that exhibits remarkably high bioavailability.

The compositions are comprised of, in general, at least one chemical entity described herein in combination with at least one pharmaceutically acceptable excipient. Acceptable excipients are non-toxic, aid administration, and do not adversely affect the therapeutic benefit of the at least one chemical entity described herein. Such excipient may be any solid, liquid, semi-solid or, in the case of an aerosol composition, gaseous excipient that is generally available to one of skill in the art.

Compressed gases may be used to disperse a chemical entity described herein in aerosol form. Inert gases suitable for this purpose are nitrogen, carbon dioxide, etc. Other suitable pharmaceutical excipients and their formulations are described in Remington's Pharmaceutical Sciences, edited by E. W. Martin (Mack Publishing Company, 18th ed., 1990).

The amount of the chemical entity in a composition can vary within the full range employed by those skilled in the art. Typically, the composition will contain, on a weight percent (wt %) basis, from about 0.01-99.99 wt % of at least one chemical entity described herein based on the total composition, with the balance being one or more suitable pharmaceutical excipients. In certain embodiments, the at least one chemical entity described herein is present at a level of about 1-80 wt %.

Compound Data

Human indoleamine 2,3-dioxgenase (IDO) cellular data is presented in Table 2 below. Brief descriptions of the cellular assays are provided following the table.

HeLa IDOi assay: Data shown in Table 2. Compounds of the present invention were tested via high-throughput cellular assays utilizing detection of kynurenine via mass spectrometry and cytotoxicity as end-points. For the mass spectrometry and cytotoxicity assays, human epithelial HeLa cells (CCL-2; ATCC®, Manassas, Va.) were stimulated with human interferon-γ (IFN-γ) (Sigma-Aldrich Corporation, St. Louis, Mo.) to induce the expression of indoleamine 2, 3-dioxygenase (IDO1). Compounds with IDO1 inhibitory properties decreased the amount of kynurenine produced by the cells via the tryptophan catabolic pathway. Cellular toxicity due to the effect of compound treatment was measured using CellTiter-Glo® reagent (CTG) (Promega Corporation, Madison, Wis.), which is based on luminescent detection of ATP, an indicator of metabolically active cells.

In preparation for the assays, test compounds were serially diluted 3-fold in DMSO from a typical top concentration of 5 mM and plated at 0.5 μL in 384-well, polystyrene, clear bottom, tissue culture treated plates with lids (Greiner Bio-One, Kremsmünster, Austria) to generate 11-point dose response curves. Low control wells (0% kynurenine or 100% cytotoxicity) contained either 0.5 μL of DMSO in the presence of unstimulated (−IFN-γ) HeLa cells for the mass spectrometry assay or 0.5 μL of DMSO in the absence of cells for the cytotoxicity assay, and high control wells (100% kynurenine or 0% cytotoxicity) contained 0.5 μL of DMSO in the presence of stimulated (+IFN-γ) HeLa cells for both the mass spectrometry and cytotoxicity assays.

Frozen stocks of HeLa cells were washed and recovered in DMEM high glucose medium with HEPES (Thermo Fisher Scientific, Inc., Waltham, Mass.) supplemented with 10% v/v certified fetal bovine serum (FBS) (Thermo Fisher Scientific, Inc., Waltham, Mass.), and 1× penicillin-streptomycin antibiotic solution (Thermo Fisher Scientific, Inc., Waltham, Mass.). The cells were diluted to 100,000 cells/mL in the supplemented DMEM medium. 50 μL of either the cell suspension, for the mass spectrometry assay, or medium alone, for the cytotoxicity assay, were added to the low control wells, on the previously prepared 384-well compound plates, resulting in 5,000 cells/well or 0 cells/well respectively. IFN-γ was added to the remaining cell suspension at a final concentration of 10 nM, and 50 μL of the stimulated cells were added to all remaining wells on the 384-well compound plates. The plates, with lids, were then placed in a 37° C., 5% CO2humidified incubator for 2 days.

Following incubation, the 384-well plates were removed from the incubator and allowed to equilibrate to room temperature for 30 minutes. For the cytotoxicity assay, CellTiter-Glo® was prepared according to the manufacturer's instructions, and 10 μL were added to each plate well. After a twenty minute incubation at room temperature, luminescence was read on an EnVision® Multilabel Reader (PerkinElmer Inc., Waltham, Mass.). For the mass spectrometry assay, 10 μL of supernatant from each well of the compound-treated plates were added to 40 μL of acetonitrile, containing 10 μM of an internal standard for normalization, in 384-well, polypropylene, V-bottom plates (Greiner Bio-One, Kremsmünster, Austria) to extract the organic analytes. Following centrifugation at 2000 rpm for 10 minutes, 10 μL from each well of the acetonitrile extraction plates were added to 90 μL of sterile, distilled H2O in 384-well, polypropylene, V-bottom plates for analysis of kynurenine and the internal standard on the RapidFire 300 (Agilent Technologies, Santa Clara, Calif.) and 4000 QTRAP MS (SCIEX, Framingham, Mass.). MS data were integrated using Agilent Technologies' RapidFire Integrator software, and data were normalized for analysis as a ratio of kynurenine to the internal standard.

The data for dose responses in the mass spectrometry assay were plotted as % IDO1 inhibition versus compound concentration following normalization using the formula 100−(100*((U−C2)/(C1−C2))), where U was the unknown value, C1 was the average of the high (100% kynurenine; 0% inhibition) control wells and C2 was the average of the low (0% kynurenine; 100% inhibition) control wells. The data for dose responses in the cytotoxicity assay were plotted as % cytotoxicity versus compound concentration following normalization using the formula 100−(100*((U−C2)/(C1−C2))), where U was the unknown value, C1 was the average of the high (0% cytotoxicity) control wells and C2 was the average of the low (100% cytotoxicity) control wells.

Curve fitting was performed with the equation y=A+((B−A)/(1+(10x/10C)D)), where A was the minimum response, B was the maximum response, C was the log(XC50) and D was the Hill slope. The results for each test compound were recorded as pIC50 values for the mass spectrometry assay and as pCC50 values for the cytoxicity assay (−C in the above equation).

PBMC IDOi assay: Data shown in Table 2. Compounds of the present invention were tested via high-throughput cellular assays utilizing detection of kynurenine via mass spectrometry and cytotoxicity as end-points. For the mass spectrometry and cytotoxicity assays, human peripheral blood mononuclear cells (PBMC) (PB003F; AlICells®, Alameda, Calif.) were stimulated with human interferon-γ (IFN-γ) (Sigma-Aldrich Corporation, St. Louis, Mo.) and lipopolysaccharide fromSalmonella minnesota(LPS) (Invivogen, San Diego, Calif.) to induce the expression of indoleamine 2, 3-dioxygenase (IDO1). Compounds with IDO1 inhibitory properties decreased the amount of kynurenine produced by the cells via the tryptophan catabolic pathway. Cellular toxicity due to the effect of compound treatment was measured using CellTiter-Glo® reagent (CTG) (Promega Corporation, Madison, Wis.), which is based on luminescent detection of ATP, an indicator of metabolically active cells.

In preparation for the assays, test compounds were serially diluted 3-fold in DMSO from a typical top concentration of 5 mM and plated at 0.5 μL in 384-well, polystyrene, clear bottom, tissue culture treated plates with lids (Greiner Bio-One, Kremsmünster, Austria) to generate 11-point dose response curves. Low control wells (0% kynurenine or 100% cytotoxicity) contained either 0.5 μL of DMSO in the presence of unstimulated (−IFN-γ/−LPS) PBMCs for the mass spectrometry assay or 0.5 μL of DMSO in the absence of cells for the cytotoxicity assay, and high control wells (100% kynurenine or 0% cytotoxicity) contained 0.5 μL of DMSO in the presence of stimulated (+IFN-γ/+LPS) PBMCs for both the mass spectrometry and cytotoxicity assays.

Frozen stocks of PBMCs were washed and recovered in RPMI 1640 medium (Thermo Fisher Scientific, Inc., Waltham, Mass.) supplemented with 10% v/v heat-inactivated fetal bovine serum (FBS) (Thermo Fisher Scientific, Inc., Waltham, Mass.), and 1× penicillin-streptomycin antibiotic solution (Thermo Fisher Scientific, Inc., Waltham, Mass.). The cells were diluted to 1,000,000 cells/mL in the supplemented RPMI 1640 medium. 50 μL of either the cell suspension, for the mass spectrometry assay, or medium alone, for the cytotoxicity assay, were added to the low control wells, on the previously prepared 384-well compound plates, resulting in 50,000 cells/well or 0 cells/well respectively. IFN-γ and LPS were added to the remaining cell suspension at final concentrations of 100 ng/ml and 50 ng/ml respectively, and 50 μL of the stimulated cells were added to all remaining wells on the 384-well compound plates. The plates, with lids, were then placed in a 37° C., 5% CO2humidified incubator for 2 days.

Following incubation, the 384-well plates were removed from the incubator and allowed to equilibrate to room temperature for 30 minutes. For the cytotoxicity assay, CellTiter-Glo® was prepared according to the manufacturer's instructions, and 40 μL were added to each plate well. After a twenty minute incubation at room temperature, luminescence was read on an EnVision® Multilabel Reader (PerkinElmer Inc., Waltham, Mass.). For the mass spectrometry assay, 10 μL of supernatant from each well of the compound-treated plates were added to 40 μL of acetonitrile, containing 10 μM of an internal standard for normalization, in 384-well, polypropylene, V-bottom plates (Greiner Bio-One, Kremsmünster, Austria) to extract the organic analytes. Following centrifugation at 2000 rpm for 10 minutes, 10 μL from each well of the acetonitrile extraction plates were added to 90 μL of sterile, distilled H2O in 384-well, polypropylene, V-bottom plates for analysis of kynurenine and the internal standard on the RapidFire 300 (Agilent Technologies, Santa Clara, Calif.) and 4000 QTRAP MS (SCIEX, Framingham, Mass.). MS data were integrated using Agilent Technologies' RapidFire Integrator software, and data were normalized for analysis as a ratio of kynurenine to the internal standard.

The data for dose responses in the mass spectrometry assay were plotted as % IDO1 inhibition versus compound concentration following normalization using the formula 100−(100*((U−C2)/(C1−C2))), where U was the unknown value, C1 was the average of the high (100% kynurenine; 0% inhibition) control wells and C2 was the average of the low (0% kynurenine; 100% inhibition) control wells. The data for dose responses in the cytotoxicity assay were plotted as % cytotoxicity versus compound concentration following normalization using the formula 100−(100*((U−C2)/(C1−C2))), where U was the unknown value, C1 was the average of the high (0% cytotoxicity) control wells and C2 was the average of the low (100% cytotoxicity) control wells.

Curve fitting was performed with the equation y=A+((B−A)/(1+(10x/10C)D)), where A was the minimum response, B was the maximum response, C was the log(XC50) and D was the Hill slope. The results for each test compound were recorded as pIC50 values for the mass spectrometry assay and as pCC50 values for the cytoxicity assay (−C in the above equation).

Although the invention has been shown and described above with reference to some embodiments, those skilled in the art will readily appreciate that the specific experiments detailed are only illustrative of the invention. It should be understood that various modifications can be made without departing from the spirit of the invention.

For example, for claim construction purposes, it is not intended that the claims set forth hereinafter be construed in any way narrower than the literal language thereof, and it is thus not intended that exemplary embodiments from the specification be read into the claims. Accordingly, it is to be understood that the present invention has been described by way of illustration and not limitations on the scope of the claims. Accordingly, the invention is limited only by the following claims. All publications, issued patents, patent applications, books and journal articles, cited in this application are each herein incorporated by reference in their entirety.1) Lohse N, Hansen A B, Pedersen G, Kronborg G, Gerstoft J, Sørensen H T, Vaeth M, Obel N. Survival of persons with and without HIV infection in Denmark, 1995-2005. Ann Intern Med. 2007 Jan. 16; 146(2):87-95.2) Deeks S G. HIV infection, inflammation, immunosenescence, and aging. Annu Rev Med. 2011; 62:141-55.3) Hunt P W, Sinclair E, Rodriguez B, Shive C, Clagett B, Funderburg N, Robinson J, Huang Y, Epling L, Martin J N, Deeks S G, Meinert C L, Van Natta M L, Jabs D A, Lederman M M. Gut epithelial barrier dysfunction and innate immune activation predict mortality in treated HIV infection. J Infect Dis. 2014 Oct. 15; 210(8):1228-38.4) Tenorio A R, Zheng Y, Bosch R J, Krishnan S, Rodriguez B, Hunt P W, Plants J, Seth A, Wilson C C, Deeks S G, Lederman M M, Landay A L. Soluble markers of inflammation and coagulation but not T-cell activation predict non-AIDS-defining morbid events during suppressive antiretroviral treatment. J Infect Dis. 2014 Oct. 15; 210(8):1248-59.5) Byakwaga H, Bourn Y 2nd, Huang Y, Muzoora C, Kembabazi A, Weiser S D, Bennett J, Cao H, Haberer J E, Deeks S G, Bangsberg D R, McCune J M, Martin J N, Hunt P W. The kynurenine pathway of tryptophan catabolism, CD4+ T-cell recovery, and mortality among HIV-infected Ugandans initiating antiretroviral therapy. J Infect Dis. 2014 Aug. 1; 210(3):383-91.6) Pearson J T, Siu S, Meininger D P, Wienkers L C, Rock D A. In vitro modulation of cytochrome P450 reductase supportedindoleamine 2,3-dioxygenase activity by allosteric effectors cytochrome b(5) and methylene blue. Biochemistry 49, 2647-2656 (2010).