Imidazodiazepinedione and its method of use

Abstract

To provide compounds that have inhibitory action against Transient Receptor Potential Cation Channel, subfamily C, member 5 (TRPC5) and are effective for the treatment of kidney disease, diabetic retinopathy, anxiety, depression, or cancer.SOLUTION: Disclosed are compounds according to Formula (I) or (II), and pharmaceutical compositions comprising them.SELECTED DRAWING: None

Description

[Background technology]

[0001] Proteinuria is a condition in which excessive amounts of protein from the blood leak into the urine. Proteinuria can progress from a loss of 30 mg of protein in the urine per 24 hours (called microalbuminuria) to a loss of more than 300 mg per day (called macroalbuminuria) before reaching 3.5 grams or more, or 25 times the normal amount. Proteinuria occurs when the kidney's glomeruli malfunction, causing fluid accumulation in the body (edema). Long-term protein leakage has been shown to lead to kidney failure. Nephrotic syndrome (NS) accounts for approximately 12% of common end-stage renal disease cases in the United States, costing over $3 billion annually. Approximately 5 out of 100,000 children are diagnosed with NS each year, and 15 out of 100,000 children currently live with NS. Even in patients who respond well to treatment, the recurrence rate is very high. Approximately 90% of children with NS respond to treatment, but an estimated 75% experience recurrence. Therefore, there is a need for more effective methods of treating or reducing the risk of developing kidney disease, such as proteinuria.

[0002] Mammalian TRP channel proteins form six-spanning, cation-permeable channels that can be grouped into six subfamilies based on amino acid sequence homology (TRPC, TRPV, TRPM, TRPA, TRPP, and TRPML). Recent studies of TRP channels have demonstrated that they are involved in numerous fundamental cellular functions and are thought to play an important role in the pathophysiology of many diseases. Many TRPs are expressed in the kidney along various parts of the nephron, and increasing evidence suggests that these channels are involved in hereditary and acquired renal disorders. For example, TRPC6, TRPM6, and TRPP2 are involved in hereditary focal segmental glomerulosclerosis (FSGS), hypomagnesemia with secondary hypocalcemia (HSH), and polycystic kidney disease (PKD), respectively. TRPC5 has also been reported to contribute to the mechanisms underlying the regulation of innate fear responses (J Neurosci. 2014 Mar 5;34(10):3653-3667).

[0003] Therefore, there is a need for additional inhibitors of TRPC5. Summary of the Invention

[0004] The present invention is based, at least in part, on the discovery that transient receptor potential cation channel, subfamily C, member 5 (TRPC5) activity abolishes actin stress fibers and reduces focal adhesion formation, providing a motile, migratory podocyte phenotype.

[0005] In one aspect, the present invention relates to small molecule TRPC5 modulators.

[0006] In some embodiments, the present invention relates to small molecule TRPC5 inhibitors and the use of such inhibitors in methods of treating or reducing the risk of developing kidney disease (e.g., proteinuria, microalbuminuria, macroalbuminuria), anxiety, depression, or cancer, comprising administering to a subject in need thereof.

[0007] In some embodiments, the present invention relates to small molecule TRPC5 agonists and the use of such agonists in methods of treating obesity or reducing the risk of developing obesity.

[0008] The interaction of small molecule ligands with proteins can result in agonistic or antagonistic (inhibitory) activity. The structural determinants that result in agonistic or antagonistic activity are often poorly understood. The antagonistic effects of closely related molecules, even enantiomers, on the activity of their biological targets have been observed in multiple cases over decades of research. This is particularly common in membrane signaling proteins such as ion channels and GPCRs (X. Huang et al., ACS Med. Chem. Lett. 2018, 9, 679-684; R. Recio et al., Eur J Med Chem 2017, 138, 644-660; Y. Kim et al., Eur J Med Chem 2016, 123, 180-190). Examples of this behavior include the modulation of calcium channels such as the DHP receptor (GC Rovnyak et al., J Med Chem. 1995, 38(1):119-29, via Neil's email), calcium channels in cardiac cells (RS Kass, Circ Res 1987, 61(4 Pt 2), I1-5 and others (RP Hof et al., J Cardiovasc Pharmacol. 1985, 7(4):689-93). These references highlight how small structural features govern whether a compound can act as an agonist or antagonist. Very recently, such a phenomenon has been described for TRPC1 / 4 / 5 channels (HNRubaiy et al., Br J Pharmacol. 2018, 175(5):830-839. doi:10.1111 / bph.14128. Epub 2018 Jan 25). According to such literature reports, it has been found that Formula I and Formula II described herein include both agonists and inhibitors. Those skilled in the art can easily determine whether a compound of Formula I or Formula II is a TRPC5 agonist or inhibitor by testing it in the FLIPR assay described herein or any other assay that can determine whether a compound is a TRPC5 inhibitor or agonist.

[0009] The above-described methods of treatment are effective in a variety of subjects, including mammals, e.g., humans and other mammals, e.g., mice, rats, rabbits, and monkeys, as well as domestic and farm mammals, e.g., cats, dogs, goats, sheep, pigs, cows, or horses.

[0010] In some embodiments, the compound of the invention is a compound of formula (I) or (II), or a pharmaceutically acceptable salt thereof:

[0011] [ka] (In the formula, A and A' are independently selected from CR and N; R is LR 1 and L is absent, CH2, O, SO2, or NR 2 and R 1 is selected from optionally substituted alkyl, optionally substituted aryl, and optionally substituted heteroaryl; Each R 2 are independently H or alkyl; R 3 is an optionally substituted alkyl, an optionally substituted alkylene -OR 2 , optionally substituted cycloalkylene-OR 2 , optionally substituted alkylene-N(R 7 )2, optionally substituted cycloalkylene-N(R 7 )2, optionally substituted alkylene-C(O)N(R 2 )2, optionally substituted cycloalkylene-C(O)N(R 2 )2, optionally substituted alkylene-S(O)2N(R 2 )2, and optionally substituted cycloalkylene-S(O)2N(R 2 )2 is selected, R 4 is selected from alkyl, optionally substituted alkylene-aryl, and optionally substituted alkylene-heteroaryl; Each R 5is H,N(R 2 )2, OR 2 are independently selected from Each R 7 is independently selected from H, alkyl, (alkyl)C(O)—, (aryl)C(O)—, (alkyl)S(O)—, and (aryl)S(O)—; X is -C(O)-, CH2, CHR 6 , C(R 6 )2, Each R 6 is independently selected from H, alkyl, and optionally substituted alkylene-OH; X' is -C(O)-, CH2, CHR 3’ , C(R 3’ )2 or X' is R 3 together to form a five- or six-membered ring, Each R 3’ is an optionally substituted alkyl, an optionally substituted alkylene -OR 2 , optionally substituted cycloalkylene-OR 2 , optionally substituted alkylene-N(R 7 )2, optionally substituted cycloalkylene-N(R 7 )2, optionally substituted alkylene-C(O)N(R 2 )2, optionally substituted cycloalkylene-C(O)N(R 2 )2, optionally substituted alkylene-S(O)2N(R 2 )2, and optionally substituted cycloalkylene-S(O)2N(R 2 ) independently selected from Z does not exist, CH2, CHR 5 , O, -NR 2 -, or -SO2-, However, both X and X' cannot be -C(O)-, and when Z is O, NR, or SO2, R 5 is H).

[0012] In one aspect, the invention features a composition including a compound of any one of Formula (I) or (II), or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable excipient.

[0013] In one aspect, the invention features a method for treating or reducing the risk of developing kidney disease, diabetic retinopathy, anxiety, depression, or cancer, comprising administering to a subject in need thereof a therapeutically effective amount of a compound of Formula (I) or (II). In certain embodiments, kidney disease is treated or the risk of developing kidney disease is reduced. In certain embodiments, kidney disease is treated. In certain embodiments, the renal disease is selected from the group consisting of focal segmental glomerulosclerosis (FSGS), diabetic nephropathy, Alport syndrome, hypertensive renal disease, nephrotic syndrome, steroid-resistant nephrotic syndrome, minimal change disease, membranous nephropathy, idiopathic membranous nephropathy, membranoproliferative glomerulonephritis (MPGN), immune complex-mediated MPGN, complement-mediated MPGN, lupus nephritis, post-infectious glomerulonephritis, thin basement membrane disease, mesangial proliferative glomerulonephritis, amyloidosis (primary), c1q nephropathy, rapidly progressive GN, anti-GBM disease, C3 glomerulonephritis, hypertensive nephrosclerosis, and IgA nephropathy. In certain embodiments, the renal disease is proteinuria. In certain embodiments, the renal disease is microalbuminuria or macroalbuminuria.

[0014] In some embodiments, the invention features methods of treating or reducing the risk of developing obesity.

[0015] In certain embodiments, the subject is a mammal. In certain embodiments, the mammal is a human.

[0016] In some embodiments, the invention comprises administering a compound of Formula (I) or (II) to a mammal and assessing the effect of the compound on calcium transport, wherein a compound that reduces or inhibits calcium transport is a therapeutic agent for treating or reducing the risk of developing kidney disease, anxiety, depression, or cancer.

[0017] The present invention provides several advantages. The prophylactic and therapeutic methods described herein are effective in treating kidney disease, e.g., proteinuria, with minimal, if any, side effects. Furthermore, the methods described herein are useful for identifying compounds that treat or reduce the risk of developing kidney disease, anxiety, depression, or cancer.

[0018] Other features, objects, and advantages of the invention will become apparent from the detailed description and claims. DETAILED DESCRIPTION OF THE INVENTION

[0019] definition The term "acyl" is art-recognized and refers to a group represented by the general formula hydrocarbylC(O)-, preferably alkylC(O)-.

[0020] The term "acylamino" is art-recognized and refers to an amino group substituted with an acyl group and may be represented, for example, by the formula hydrocarbylC(O)NH-.

[0021] The term "acyloxy" is art-recognized and refers to a group represented by the general formula hydrocarbylC(O)O-, preferably alkylC(O)O-.

[0022] The term "alkoxy" refers to an alkyl group, preferably a lower alkyl group, having an oxygen attached thereto. Representative alkoxy groups include methoxy, trifluoromethoxy, ethoxy, propoxy, tert-butoxy, and the like.

[0023] The term "alkoxyalkyl" refers to an alkyl group substituted with an alkoxy group and may be represented by the general formula alkyl-O-alkyl.

[0024] The term "alkenyl," as used herein, refers to an aliphatic group containing at least one double bond and is intended to include both "unsubstituted alkenyl" and "substituted alkenyl," the latter of which refers to an alkenyl moiety having substituents replacing a hydrogen on one or more carbons of the alkenyl group. Such substitutions may occur on one or more carbons included or not included in one or more double bonds. Furthermore, such substitutions include all of those contemplated for alkyl groups, as described below, except where stability would be inhibited. For example, substitution of alkenyl groups with one or more alkyl, carbocyclyl, aryl, heterocyclyl, or heteroaryl groups is contemplated.

[0025] An "alkyl" group or "alkane" is a fully saturated, straight-chain or branched non-aromatic hydrocarbon. Typically, a straight-chain or branched alkyl group has 1 to about 20 carbon atoms, preferably 1 to about 10, unless otherwise defined. Examples of straight-chain and branched alkyl groups include methyl, ethyl, n-propyl, iso-propyl, n-butyl, sec-butyl, tert-butyl, pentyl, hexyl, pentyl, and octyl. C1-C6 straight-chain or branched alkyl groups are also referred to as "lower alkyl" groups.

[0026] Furthermore, the term "alkyl" (or "lower alkyl"), as used throughout the specification, examples, and claims, is intended to include both "unsubstituted alkyl" and "substituted alkyl," the latter of which refers to an alkyl moiety having substituents replacing a hydrogen on one or more carbons of the hydrocarbon backbone. Such substituents, unless otherwise specified, can include, for example, halogen (e.g., fluoro), hydroxyl, carbonyl (e.g., carboxyl, alkoxycarbonyl, formyl, or acyl), thiocarbonyl (e.g., thioester, thioacetate, or thioformate), alkoxy, phosphoryl, phosphate, phosphonate, phosphinate, amino, amido, amidine, imine, cyano, nitro, azido, sulfhydryl, alkylthio, sulfate, sulfonate, sulfamoyl, sulfonamido, sulfonyl, heterocyclyl, aralkyl, or aromatic or heteroaromatic moieties. In a preferred embodiment, the substituents on a substituted alkyl are C 1-6 Alkyl, C 3-6 The substituents of the substituted alkyl are selected from cycloalkyl, halogen, carbonyl, cyano, or hydroxyl. In a more preferred embodiment, the substituents of the substituted alkyl are selected from fluoro, carbonyl, cyano, or hydroxyl. Those skilled in the art will understand that the moieties substituted on the hydrocarbon chain may themselves be substituted, if appropriate. For example, the substituents of the substituted alkyl may include amino, azido, imino, amido, phosphoryl (including phosphonates and phosphinates), sulfonyl (including sulfates, sulfonamides, sulfamoyl, and sulfonates), and silyl groups, as well as substituted and unsubstituted forms of ethers, alkylthio, carbonyl (including ketones, aldehydes, carboxylates, and esters), -CF3, -CN, and the like. Exemplary substituted alkyls are described below. The cycloalkyl may be further substituted with alkyl, alkenyl, alkoxy, alkylthio, aminoalkyl, carbonyl-substituted alkyl, -CF3, -CN, and the like.

[0027] Unless otherwise stated, "alkylene," by itself or as part of another substituent, refers to a saturated, straight-chain or branched divalent radical derived from the removal of two hydrogen atoms from the corresponding alkane, having the designated number of carbon atoms. Examples of straight-chain and branched alkylene groups include -CH- (methylene), -CH-CH- (ethylene), -CH-CH-CH- (propylene), -C(CH)-, -CH-CH(CH)-, -CH-CH-CH-CH-, -CH-CH-CH-CH- (pentylene), -CH-CH(CH)-CH-, and -CH-C(CH)-CH-.

[0028] "C x-y The term "C" when used with a chemical moiety such as acyl, acyloxy, alkyl, alkenyl, alkynyl, or alkoxy is meant to include groups containing x to y carbons in the chain. For example, "C x-y The term "alkyl" refers to substituted or unsubstituted saturated hydrocarbon groups, including straight-chain alkyl and branched-chain alkyl groups, containing x to y carbons in the chain, including haloalkyl groups. Preferred haloalkyl groups include trifluoromethyl, difluoromethyl, 2,2,2-trifluoroethyl, and pentafluoroethyl. CO alkyl indicates a hydrogen atom in the terminal position of the group, or a bond if the group is internal. "C 2-y alkenyl" and "C 2-y The term "alkynyl" refers to substituted or unsubstituted unsaturated aliphatic groups analogous in length and possible substitution to the alkyls described above, but which contain at least one double or triple bond respectively.

[0029] The term "alkylamino," as used herein, refers to an amino group substituted with at least one alkyl group.

[0030] The term "alkylthio," as used herein, refers to a thiol group substituted with an alkyl group and may be represented by the general formula alkylS-.

[0031] The term "alkynyl," as used herein, refers to an aliphatic group containing at least one triple bond and is intended to include both "unsubstituted alkynyl" and "substituted alkynyl," the latter of which refers to an alkynyl moiety having substituents replacing hydrogen on one or more carbons of the alkynyl group. Such substitutions may occur on one or more carbons included or not included in one or more triple bonds. Furthermore, such substitutions include all of those contemplated for alkyl groups as described above, except where stability would be inhibited. For example, substitution of alkynyl groups with one or more alkyl, carbocyclyl, aryl, heterocyclyl, or heteroaryl groups is contemplated.

[0032] The term "amide" as used herein refers to the group

[0033] [ka] In the formula, each R A independently represent hydrogen or a hydrocarbyl group, or two R A together with the N atom to which they are attached complete a heterocycle with 4 to 8 atoms in the ring structure.

[0034] The terms "amine" and "amino" are art-recognized and refer to both unsubstituted and substituted amines, as well as salts thereof,

[0035] [ka] For example, it refers to a moiety that can be represented by A independently represent hydrogen or a hydrocarbyl group, or two R A together with the N atom to which they are attached complete a heterocycle with 4 to 8 atoms in the ring structure.

[0036] The term "aminoalkyl," as used herein, refers to an alkyl group substituted with an amino group.

[0037] The term "aralkyl," as used herein, refers to an alkyl group substituted with an aryl group.

[0038] The term "aryl," as used herein, includes substituted or unsubstituted monocyclic aromatic groups in which each atom of the ring is carbon. Preferably, the ring is a 6- to 10-membered ring, more preferably a 6-membered ring. The term "aryl" also includes polycyclic ring systems having two or more cyclic rings in which two or more carbons are common to two adjacent rings, and at least one of the rings is aromatic; for example, the other cyclic rings may be cycloalkyl, cycloalkenyl, aryl, heteroaryl, and / or heterocyclyl. Aryl groups include benzene, naphthalene, phenanthrene, phenol, aniline, and the like.

[0039] The term "carbamate" is art-recognized and refers to a group

[0040] [ka] In the formula, each R A independently represent hydrogen, or a hydrocarbyl group such as an alkyl group, or both R A together with the intervening atom(s) complete a heterocycle having from 4 to 8 atoms in the ring structure.

[0041] The terms "carbocycle" and "carbocyclic," as used herein, refer to a saturated or unsaturated ring in which each atom of the ring is carbon. The term carbocycle includes both aromatic and non-aromatic carbocycles. Non-aromatic carbocycles include both cycloalkane rings in which all carbon atoms are saturated and cycloalkene rings that contain at least one double bond. "Carbocycle" includes 5- to 7-membered monocyclic rings and 8- to 12-membered bicyclic rings. Each ring of a bicyclic carbocycle may be selected from saturated, unsaturated, and aromatic rings. Carbocycles include bicyclic molecules in which one, two, or three or more atoms are shared between the two rings. The term "fused carbocycle" refers to a bicyclic carbocycle in which each ring shares two adjacent atoms with the other ring. Each ring of a fused carbocycle may be selected from saturated, unsaturated, and aromatic rings. In exemplary embodiments, an aromatic ring, e.g., phenyl, can be fused to a saturated or unsaturated ring, e.g., cyclohexane, cyclopentane, or cyclohexene. Any combination of saturated, unsaturated, and aromatic bicyclic rings is included in the definition of carbocycle, where valences permit. Exemplary "carbocycles" include cyclopentane, cyclohexane, bicyclo[2.2.1]heptane, 1,5-cyclooctadiene, 1,2,3,4-tetrahydronaphthalene, bicyclo[4.2.0]oct-3-ene, naphthalene, and adamantane. Exemplary fused carbocycles include decalin, naphthalene, 1,2,3,4-tetrahydronaphthalene, bicyclo[4.2.0]octane, 4,5,6,7-tetrahydro-1H-indene, and bicyclo[4.1.0]hept-3-ene. A "carbocycle" can be substituted at any one or more positions capable of bearing a hydrogen atom.

[0042] A "cycloalkyl" group is a fully saturated cyclic hydrocarbon. "Cycloalkyl" includes monocyclic and bicyclic rings. Typically, a monocyclic cycloalkyl group has 3 to about 10 carbon atoms, more typically 3 to 8 carbon atoms, unless otherwise defined. The second ring of a bicyclic cycloalkyl can be selected from saturated, unsaturated, and aromatic rings. Cycloalkyl includes bicyclic molecules in which one, two, or three or more atoms are shared between the two rings. The term "fused cycloalkyl" refers to a bicyclic cycloalkyl in which each ring shares two adjacent atoms with the other ring. The second ring of a fused bicyclic cycloalkyl can be selected from saturated, unsaturated, and aromatic rings. A "cycloalkenyl" group is a cyclic hydrocarbon containing one or more double bonds.

[0043] The term "carbocyclylalkyl," as used herein, refers to an alkyl group substituted with a carbocyclic group.

[0044] The term "carbonate" is art-recognized and refers to the group -OCO-R A refers to a group, wherein R A represents a hydrocarbyl group.

[0045] The term "carboxy," as used herein, refers to a group represented by the formula -CO2H.

[0046] The term "ester" as used herein refers to an ester of -C(O)OR A refers to a group, wherein R A represents a hydrocarbyl group.

[0047] The term "ether," as used herein, refers to a hydrocarbyl group bonded to another hydrocarbyl group via an oxygen atom. Thus, the ether substituent of a hydrocarbyl group can be hydrocarbyl-O-. Ethers can be symmetrical or asymmetrical. Examples of ethers include, but are not limited to, heterocycle-O-heterocycle and aryl-O-heterocycle. Ethers include "alkoxyalkyl" groups, which can be represented by the general formula alkyl-O-alkyl.

[0048] The terms "halo" and "halogen" as used herein mean halogen and include chloro, fluoro, bromo, and iodo.

[0049] The terms "hetaralkyl" and "heteroaralkyl," as used herein, refer to an alkyl group substituted with a hetaryl group.

[0050] The term "heteroalkyl," as used herein, refers to a saturated or unsaturated chain of carbon atoms and at least one heteroatom, wherein no two heteroatoms are adjacent.

[0051] The terms "heteroaryl" and "hetaryl" include substituted or unsubstituted aromatic monocyclic ring structures, preferably 5- to 7-membered rings, more preferably 5- or 6-membered rings, which ring structures contain at least one heteroatom, preferably 1 to 4 heteroatoms, more preferably 1 or 2 heteroatoms. The terms "heteroaryl" and "hetaryl" also include polycyclic ring systems having two or more cyclic rings in which two or more carbons are common to two adjacent rings, at least one of which is heteroaromatic; for example, the other cyclic rings may be cycloalkyl, cycloalkenyl, aryl, heteroaryl, and / or heterocyclyl. Heteroaryl groups include, for example, pyrrole, furan, thiophene, imidazole, oxazole, thiazole, pyrazole, pyridine, pyrazine, pyridazine, and pyrimidine.

[0052] The term "heteroatom," as used herein, means an atom of any element other than carbon or hydrogen. Preferred heteroatoms are nitrogen, oxygen, and sulfur.

[0053] The terms "heterocyclyl," "heterocycle," and "heterocyclic" refer to substituted or unsubstituted non-aromatic ring structures, preferably 3- to 10-membered rings, more preferably 3- to 7-membered rings, which contain at least one heteroatom, preferably 1 to 4 heteroatoms, more preferably 1 or 2 heteroatoms. The terms "heterocyclyl" and "heterocyclic" also include polycyclic ring systems having two or more cyclic rings in which two or more carbons are common to two adjacent rings, at least one of which is heterocyclic; for example, the other cyclic rings may be cycloalkyl, cycloalkenyl, aryl, heteroaryl, and / or heterocyclyl. Heterocyclyl groups include, for example, piperidine, piperazine, pyrrolidine, tetrahydropyran, tetrahydrofuran, morpholine, lactones, lactams, and the like.

[0054] The terms "heterocyclylalkyl" or "heterocycloalkyl," as used herein, refers to an alkyl group substituted with a heterocycle group.

[0055] The term "hydrocarbyl," as used herein, refers to a group having at least one carbon-hydrogen bond and a primarily carbon backbone, but which may optionally contain heteroatoms, that does not have =0 or =S substituents and is bonded through carbon atoms. Thus, groups such as methyl, ethoxyethyl, 2-pyridyl, and trifluoromethyl are considered hydrocarbyl for purposes of this application, while substituents such as acetyl (which has an =0 substituent on the bonded carbon) and ethoxy (which is bonded through an oxygen rather than a carbon) are not. Hydrocarbyl groups include, but are not limited to, aryl, heteroaryl, carbocycle, heterocyclyl, alkyl, alkenyl, alkynyl, and combinations thereof.

[0056] The term "hydroxyalkyl," as used herein, refers to an alkyl group substituted with a hydroxy group.

[0057] When used in conjunction with chemical moieties such as acyl, acyloxy, alkyl, alkenyl, alkynyl, or alkoxy, the term "lower" is meant to include groups in which there are 10 or fewer, preferably 6 or fewer, non-hydrogen atoms in the substituent. "Lower alkyl," for example, refers to alkyl groups containing 10 or fewer, preferably 6 or fewer, carbon atoms. In certain embodiments, acyl, acyloxy, alkyl, alkenyl, alkynyl, or alkoxy substituents defined herein, whether they appear alone or in combination with other substituents, are lower acyl, lower acyloxy, lower alkyl, lower alkenyl, lower alkynyl, or lower alkoxy, respectively, including, for example, hydroxyalkyl and aralkyl (in which case, for example, atoms in aryl groups are not counted when counting the carbon atoms of an alkyl substituent).

[0058] The terms "polycyclyl," "polycycle," and "polycyclic" refer to two or more rings (e.g., cycloalkyl, cycloalkenyl, aryl, heteroaryl, and / or heterocyclyl) in which two or more atoms are common to two adjacent rings, e.g., the rings are "fused rings." Each of the rings of a polycycle can be substituted or unsubstituted. In certain embodiments, each ring of a polycycle contains 3 to 10 atoms in the ring, preferably 5 to 7 atoms.

[0059] The term "silyl" refers to a silicon moiety having three hydrocarbyl moieties attached thereto.

[0060] The term "substituted" refers to moieties having substituents replacing a hydrogen on one or more backbone carbons. It is understood that "substituted" or "substituted with" includes the implicit proviso that such substitution is subject to the permissible valences of the replacing atom and substituent, as well as the implicit proviso that the substitution results in a stable compound that does not spontaneously undergo transformation, e.g., by rearrangement, cyclization, elimination, and the like. As used herein, the term "substituted" is contemplated to include all permissible substituents of organic compounds. In a broad aspect, the permissible substituents include acyclic and cyclic, branched and unbranched, carbocyclic and heterocyclic, aromatic and nonaromatic substituents of organic compounds. The permissible substituents can be one or more and the same or different for appropriate organic compounds. For purposes of this invention, heteroatoms, such as nitrogen, can have hydrogen substituents and / or any permissible substituents of organic compounds described herein that satisfy the valence of the heteroatom. The substituents may include any of the substituents described herein, for example, halogen, hydroxyl, carbonyl (e.g., carboxyl, alkoxycarbonyl, formyl, or acyl), thiocarbonyl (e.g., thioester, thioacetate, or thioformate), alkoxy, phosphoryl, phosphate, phosphonate, phosphinate, amino, amido, amidine, imine, cyano, nitro, azido, sulfhydryl, alkylthio, sulfate, sulfonate, sulfamoyl, sulfonamido, sulfonyl, heterocyclyl, aralkyl, or aromatic or heteroaromatic moiety. In a preferred embodiment, the substituent of the substituted alkyl is C 1-6 Alkyl, C 3-6 In a more preferred embodiment, the substituents of the substituted alkyl are selected from fluoro, carbonyl, cyano, or hydroxyl. Those skilled in the art will understand that the substituents themselves can be substituted, where appropriate. Unless specifically stated as "unsubstituted," references to chemical moieties herein are understood to include substituted variants. For example, references to "aryl" groups or moieties implicitly include both substituted and unsubstituted variants.

[0061] The term "sulfate" is art-recognized and refers to the group -OSO3H, or a pharmaceutically acceptable salt thereof.

[0062] The term "sulfonamide" is art-recognized and can be represented by the general formula

[0063] [ka] wherein each R A independently represent hydrogen or hydrocarbyl such as alkyl, or both R A together with the intervening atom(s) complete a heterocycle having from 4 to 8 atoms in the ring structure.

[0064] The term "sulfoxide" is art-recognized and refers to an -S(O)-R A refers to a group, wherein R A represents a hydrocarbyl.

[0065] The term "sulfonate" is art-recognized and refers to a compound selected from the group consisting of sulfonates, sulfonates, and sulfonates. 3H group, or a pharmaceutically acceptable salt thereof.

[0066] The term “sulfone” is art-recognized and refers to the group —S(O)—R A refers to a group, wherein R A represents a hydrocarbyl.

[0067] The term "thioalkyl," as used herein, refers to an alkyl group substituted with a thiol group.

[0068] The term "thioester" as used herein refers to a group selected from the group consisting of -C(O)SR A or -SC(O)R A where R A represents a hydrocarbyl.

[0069] The term "thioether," as used herein, is equivalent to an ether, where the oxygen has been replaced with a sulfur.

[0070] The term "urea" is art-recognized and has the general formula

[0071] [ka] wherein each R A independently represent hydrogen or hydrocarbyl such as alkyl, or R taken together with another and intervening atom(s) A Any occurrence of completes a heterocycle having from 4 to 8 atoms in the ring structure.

[0072] A "protecting group" refers to a group of atoms that, when attached to a reactive functional group in a molecule, masks, reduces, or prevents the reactivity of the functional group. Typically, a protecting group can be selectively removed as desired during the course of synthesis. Examples of protecting groups can be found in Greene and Wuts, Protective Groups in Organic Chemistry, 3rd Ed., 1999, John Wiley & Sons, NY, and Harrison et al., Compendium of Synthetic Organic Methods, Vols. 1-8, 1971-1996, John Wiley & Sons, NY. Representative nitrogen protecting groups include, but are not limited to, formyl, acetyl, trifluoroacetyl, benzyl, benzyloxycarbonyl ("CBZ"), tert-butoxycarbonyl ("Boc"), trimethylsilyl ("TMS"), 2-trimethylsilyl-ethanesulfonyl ("TES"), trityl and substituted trityl groups, allyloxycarbonyl, 9-fluorenylmethyloxycarbonyl ("FMOC"), nitro-veratryloxycarbonyl ("NVOC"), etc. Representative hydroxyl protecting groups include, but are not limited to, those in which the hydroxyl group is acylated (esterified) or alkylated, such as benzyl and trityl ethers, as well as alkyl ethers, tetrahydropyranyl ethers, trialkylsilyl ethers (e.g., TMS or TIPS groups), glycol ethers (e.g., ethylene glycol and propylene glycol derivatives), and allyl ethers.

[0073] As used herein, a therapeutic agent that "prevents" a disorder or condition refers to a compound that, in a statistical sample, reduces or delays the onset of the disorder or condition in a treated sample relative to an untreated control sample, or reduces the severity of one or more symptoms of the disorder or condition relative to an untreated control sample.

[0074] The term "treating" includes prophylactic and / or therapeutic treatment. The term "prophylactic or therapeutic" treatment is art-recognized and includes administration of one or more of the subject compositions to a host. If it is administered prior to clinical manifestation of an undesired condition (e.g., a disease or other undesired condition in a host animal), the treatment is prophylactic (i.e., it protects the host from the onset of the undesired condition), whereas if it is administered after the undesired condition is manifested, the treatment is therapeutic (i.e., aimed at reducing, ameliorating, or stabilizing an existing undesired condition or its side effects).

[0075] The phrases "co-administration" and "administered in combination" refer to any administration form of two or more different therapeutic compounds in which a second compound is administered while a previously administered therapeutic compound is still effective in the body (e.g., the two compounds are effective on a patient simultaneously, which may involve a synergistic effect of the two compounds). For example, the different therapeutic compounds can be administered simultaneously or sequentially, in the same formulation or in separate formulations. In certain embodiments, the different therapeutic compounds can be administered within 1 hour, 12 hours, 24 hours, 36 hours, 48 ​​hours, 72 hours, or 1 week of each other. Thus, an individual receiving such treatment can benefit from the combined effects of the different therapeutic compounds.

[0076] The term "prodrug" is intended to encompass compounds that are converted under physiological conditions to the therapeutically active agents of the present invention. A common method for making prodrugs is to include one or more selected moieties that are hydrolyzed under physiological conditions to reveal the desired molecule. In other embodiments, the prodrug is converted by the enzymatic activity of the host animal. For example, esters or carbonates (e.g., esters or carbonates of alcohols or carboxylic acids) are preferred prodrugs of the present invention. In certain embodiments, some or all of the compounds of the present invention in the above formulations can be replaced with the corresponding suitable prodrug; for example, a hydroxyl in the parent compound is presented as an ester or carbonate, or a carboxylic acid present in the parent compound is presented as an ester.

[0077] As used herein, "small molecule" refers to a small organic or inorganic molecule having a molecular weight of less than about 3,000 daltons. Generally, small molecules useful in the present invention have a molecular weight of less than 3,000 daltons (Da). A small molecule can be, for example, at least about 100 Da to about 3,000 Da (e.g., about 100 to about 3,000 Da, about 100 to about 2,500 Da, about 100 to about 2,000 Da, about 100 to about 1,750 Da, about 100 to about 1,500 Da, about 100 to about 1,250 Da, about 100 to about 1,000 Da, about 100 to about 750 Da, about 100 to about 500 Da, about 200 to about 1,500, about 500 to about 1,000, about 300 to about 1,000 Da, or about 100 to about 250 Da).

[0078] In some embodiments, "small molecule" refers to an organic, inorganic, or organometallic compound, typically having a molecular weight of less than about 1000. In some embodiments, small molecules are organic compounds about 1 nm in size. In some embodiments, small molecule drugs of the present invention include oligopeptides and other biomolecules having a molecular weight of less than about 1000.

[0079] An "effective amount" is an amount sufficient to produce a beneficial or desired result. For example, a therapeutic amount is an amount that achieves a desired therapeutic effect. This amount may be the same as or different from a prophylactically effective amount, which is an amount necessary to prevent the onset of a disease or disease symptoms. An effective amount may be administered in one or more administrations, applications, or doses. The therapeutically effective amount of a composition will depend on the composition selected. The composition may be administered once or more times daily to once or more times weekly (including every other day). One of ordinary skill in the art will appreciate that certain factors, including, but not limited to, the severity of the disease or disorder, previous treatments, general health, and / or age of the subject, as well as other diseases present, may affect the dosage and timing required to effectively treat a subject. Furthermore, treatment of a subject with a therapeutically effective amount of a composition described herein may include a single treatment or a series of treatments.

[0080] Compounds of the Invention One aspect of the present invention provides small molecule modulators of TRPC5. In some embodiments, the present invention provides small molecule inhibitors of TRPC5. In some embodiments, the present invention provides small molecule agonists of TRPC5.

[0081] In some embodiments, the compound of the invention is a compound of formula (I) or (II), or a pharmaceutically acceptable salt thereof:

[0082] [ka] (In the formula, A and A' are independently selected from CR and N; R is LR 1 and L is absent, CH2, O, SO2, or NR 2 and R 1 is selected from optionally substituted alkyl, optionally substituted aryl, and optionally substituted heteroaryl; if L is absent, R 1 is further selected from H, Each R2 are independently H or alkyl; R 3 is an optionally substituted alkyl, an optionally substituted alkylene -OR 2 , optionally substituted cycloalkylene-OR 2 , optionally substituted alkylene-N(R 7 )2, optionally substituted cycloalkylene-N(R 7 )2, optionally substituted alkylene-C(O)N(R 2 )2, optionally substituted cycloalkylene-C(O)N(R 2 )2, optionally substituted alkylene-S(O)2N(R 2 )2, and optionally substituted cycloalkylene-S(O)2N(R 2 )2 is selected, R 4 is selected from alkyl, optionally substituted alkylene-aryl, and optionally substituted alkylene-heteroaryl; Each R 5 is H,N(R 2 )2, OR 2 are independently selected from Each R 7 is independently selected from H, alkyl, (alkyl)C(O)—, (aryl)C(O)—, (alkyl)S(O)—, and (aryl)S(O)—; X is -C(O)-, CH2, CHR 6 , C(R 6 )2, Each R 6 is independently selected from H, alkyl, and optionally substituted alkylene-OH; X' is -C(O)-, CH2, CHR 3’ , C(R 3’ )2 or X' is R 3 together to form a five- or six-membered ring, Each R 3’ is an optionally substituted alkyl, an optionally substituted alkylene -OR 2 , optionally substituted cycloalkylene-OR 2 , optionally substituted alkylene-N(R 7)2, optionally substituted cycloalkylene-N(R 7 )2, optionally substituted alkylene-C(O)N(R 2 )2, optionally substituted cycloalkylene-C(O)N(R 2 )2, optionally substituted alkylene-S(O)2N(R 2 )2, and optionally substituted cycloalkylene-S(O)2N(R 2 ) independently selected from Z does not exist, CH2, CHR 5 , O, -NR 2 -, or -SO2-, However, both X and X' cannot be -C(O)-, and when Z is O, NR, or SO2, R 5 is H).

[0083] In some embodiments, the compound is a compound of formula (I): In some embodiments, the compound is a compound of formula (II):

[0084] In some embodiments, at least one of A and A' is CR.

[0085] In some embodiments, A is N. In some embodiments, A is CR.

[0086] In some embodiments, A' is N. In some embodiments, A' is CR.

[0087] In some embodiments, A' is N and A is CR.

[0088] In some embodiments, R is LR 1 is.

[0089] In some embodiments, L is absent. In some embodiments, when L is absent, R 1is further selected from H. In some embodiments, L is CH. In some embodiments, L is O. In some embodiments, L is SO. In some embodiments, L is NR 2 is.

[0090] In some embodiments, R 1 is optionally substituted aryl. In some embodiments, R 1 is optionally substituted phenyl. In some embodiments, R 1 is a substituted phenyl. In some embodiments, the substituted phenyl is substituted with one or more substituents independently selected from halogen, —CF 3 , —C(H)F 2 , and —OCF 3 .

[0091] In some embodiments, R 1 is optionally substituted alkyl. In some embodiments, alkyl is chosen from methyl, ethyl, n-propyl, iso-propyl, n-butyl, iso-butyl, and tert-butyl.

[0092] In some embodiments, R 1 is optionally substituted heteroaryl. In some embodiments, R 1 is a substituted heteroaryl substituted with one or more substituents independently selected from halogen, —CF 3 , —C(H)F 2 , and —OCF 3 .

[0093] In some embodiments, L is O and R 1 is 3-chlorophenyl, 3-fluorophenyl, 3-trifluoromethoxyphenyl, isopropyl, or n-propyl.

[0094] In some embodiments, R 2 is H. In some embodiments, R 2 is alkyl. In some embodiments, R 2is selected from methyl, ethyl, n-propyl, iso-propyl, n-butyl, iso-butyl, and tert-butyl. 2 is methyl.

[0095] In some embodiments, R 3 is optionally substituted alkyl. In some embodiments, R 3 is an optionally substituted alkylene -OR 2 In some embodiments, R 3 is an optionally substituted cycloalkylene -OR 2 In some embodiments, R 3 is an optionally substituted alkylene-N(R 7 )2. In some embodiments, R 3 is an optionally substituted cycloalkylene-N(R 7 )2. In some embodiments, R 3 is an optionally substituted alkylene -C(O)N(R 2 )2. In some embodiments, R 3 is an optionally substituted cycloalkylene -C(O)N(R 2 )2. In some embodiments, R 3 is an optionally substituted alkylene -S(O)N(R 2 )2. In some embodiments, R 3 is an optionally substituted cycloalkylene -S(O)N(R 2 )2. In some embodiments, R 3 is methyl, 2-hydroxyethyl, 2,3-dihydroxypropyl, 2,2-difluoro-3-hydroxypropyl, 3-hydroxypropyl, 3-methoxypropyl, 3-hydroxycyclobutyl, or 3-hydroxycyclopentyl.

[0096] In some embodiments, R 7 One example of is H, and another example is R 7A second example of is alkyl, (alkyl)C(O)—, (aryl)C(O)—, (alkyl)S(O)—, or (aryl)S(O)—. In some embodiments, R 7 One example of is alkyl, and R 7 A second example of is H, (alkyl)C(O)—, (aryl)C(O)—, (alkyl)S(O)—, or (aryl)S(O)—. In some embodiments, R 7 Both examples of R are H. In some embodiments, R 7 Both examples of are alkyl.

[0097] In some embodiments, R 3 teeth,

[0098] [ka] In some embodiments, R 3 teeth,

[0099] [ka] is.

[0100] In some embodiments, R 3 teeth,

[0101] [ka] is selected from.

[0102] In some embodiments, R 3 teeth,

[0103] [ka] is selected from.

[0104] In some embodiments, R 4 is alkyl. In some embodiments, R 4is selected from methyl, ethyl, n-propyl, iso-propyl, n-butyl, iso-butyl, and tert-butyl. 4 is selected from n-butyl, iso-butyl, and tert-butyl. In some embodiments, R 4 is n-butyl.

[0105] In some embodiments, R 4 is an optionally substituted alkylene-aryl. In some embodiments, the alkylene of the alkylene-aryl is substituted. In some embodiments, the aryl of the alkylene-aryl is substituted. In some embodiments, the substituted aryl is substituted with halogen, —CF3, —C(H)F2, or —OCF3. In some embodiments, the aryl of the alkylene-aryl is an optionally substituted phenyl. In some embodiments, the phenyl is substituted with one or more instances of halogen. In some embodiments, the alkylene of the alkylene-aryl is methylene. In some embodiments, R 4 is an optionally substituted alkylene-heteroaryl.

[0106] In some embodiments, R 4 is n-butyl, 4-chlorobenzyl, or 2-(4-chlorophenyl)ethan-2-yl.

[0107] In some embodiments, R 4 teeth

[0108] [ka] In some embodiments, R 4 teeth

[0109] [ka] In some embodiments, R 4 teeth

[0110] [ka] is.

[0111] In some embodiments, each R 5 is H. In some embodiments, one R 5 is hydrogen, and the other R 5 is —O-alkyl. In some embodiments, one R 5 is hydrogen, and the other R 5 is -OMe. In some embodiments, one R 5 is hydrogen, and the other R 5 is —OH. In some embodiments, one R 5 is hydrogen, and the other R 5 is -NMe2. In some embodiments, one R 5 is hydrogen, and the other R 5 is -NH2.

[0112] In some embodiments, X is -C(O)-. In some embodiments, X is CH. In some embodiments, X is -CHR 6 In some embodiments, X is -C(R 6 )2-. In some embodiments, R 6 is alkyl. In some embodiments, R 6 is selected from methyl, ethyl, n-propyl, iso-propyl, n-butyl, iso-butyl, and tert-butyl. 6 is methyl. In some embodiments, R 6 is optionally substituted alkylene-OH. In some embodiments, R 6 is optionally substituted ethylene-OH. In some embodiments, R 6 is a substituted ethylene-OH. In some embodiments, R 6 is H.

[0113] In some embodiments, X' is -C(O)-. In some embodiments, X' is CH2.

[0114] In some embodiments, X' is -CHR 3’ In some embodiments, X' is -C(R 3’ )2-. In some embodiments, R 3’ is alkyl. In some embodiments, R 3’ is selected from methyl, ethyl, n-propyl, iso-propyl, n-butyl, iso-butyl, and tert-butyl. 3’ is methyl. In some embodiments, R 3’ is optionally substituted alkylene-OH. In some embodiments, R 3’ is optionally substituted ethylene-OH. In some embodiments, R 3’ is a substituted ethylene -OH.

[0115] In some embodiments, Z is absent. In some embodiments, Z is CH. In some embodiments, Z is -N(alkyl)-. In some embodiments, Z is selected from -N(n-butyl)-, -N(iso-butyl)-, and -N(tert-butyl)-. In some embodiments, Z is -SO-. In some embodiments, Z is absent and each R 5 is hydrogen.

[0116] In some embodiments, the compound is

[0117] [ka] is selected from.

[0118] In some embodiments, the compound is

[0119] [ka] is.

[0120] In some embodiments, the compound is

[0121] [ka] is.

[0122] In some embodiments, the compound is

[0123] [ka] is selected from.

[0124] In some embodiments, the compound is

[0125] [ka]

[0126] [ka] is selected from.

[0127] In some embodiments, the compound is

[0128] [ka] is selected from.

[0129] In some embodiments, the compound is

[0130] [ka] is.

[0131] In some embodiments, the compound is

[0132] [ka] is.

[0133] In some embodiments, the compound is

[0134] [ka] is.

[0135] In some embodiments, the compound is

[0136] [ka] is.

[0137] In some embodiments, the compound is

[0138] [ka] is.

[0139] In certain embodiments, the compound of the present invention can be racemic.In certain embodiments, the compound of the present invention can be enriched in one enantiomer.For example, the compound of the present invention can have an ee of more than 30%, an ee of more than 40%, an ee of more than 50%, an ee of more than 60%, an ee of more than 70%, an ee of more than 80%, an ee of more than 90%, or even an ee of more than 95%.

[0140] The compounds of the present invention have multiple stereocenters. Therefore, the compounds of the present invention can be enriched in one or more diastereomers. For example, the compounds of the present invention can have more than 30% de, more than 40% de, more than 50% de, more than 60% de, more than 70% de, more than 80% de, more than 90% de, or even more than 95% de. In certain embodiments, the compounds of the present invention have substantially one isomeric configuration at one or more stereocenters, and multiple isomeric configurations at the remaining stereocenters.

[0141] In certain embodiments, the enantiomeric excess of the stereocenter is at least 40% ee, 50% ee, 60% ee, 70% ee, 80% ee, 90% ee, 92% ee, 94% ee, 95% ee, 96% ee, 98% ee or more ee.

[0142] As used herein, a single bond drawn without stereochemistry does not represent the stereochemistry of the compound.

[0143] As used herein, hashed or bolded non-wedged bonds indicate relative but not absolute stereochemical configurations (eg, do not distinguish between enantiomers of a given diastereomer).

[0144] As used herein, hashed or bold wedge bonds indicate absolute stereochemical configuration.

[0145] In certain embodiments, therapeutic preparations of the compounds of the present invention can be enriched to provide predominantly one enantiomer of the compound. An enantiomer-enriched mixture can, for example, contain at least 60 mole percent of one enantiomer, or more preferably at least 75, 90, 95, or even 99 mole percent. In certain embodiments, a compound enriched in one enantiomer is substantially free of other enantiomers, where substantially free means that the substance in question accounts for, for example, less than 10%, or less than 5%, or less than 4%, or less than 3%, or less than 2%, or less than 1% of the amount of other enantiomers in the composition or compound mixture. For example, if a composition or compound mixture contains 98 grams of a first enantiomer and 2 grams of a second enantiomer, it is considered to contain 98 mole percent of the first enantiomer and only 2% of the second enantiomer.

[0146] In certain embodiments, therapeutic preparations can be enriched to provide predominantly one diastereomer of a compound of the invention. A diastereomerically enriched mixture can contain, for example, at least 60 mole percent, or more preferably at least 75, 90, 95, or even 99 mole percent of one diastereomer.

[0147] Treatment method Non-selective Ca 2+ Transient receptor potential (TRP) channels function as sensors that transduce extracellular cues into the intracellular environment in diverse cellular processes, including actin remodeling and cell migration (Greka et al., Nat Neurosci 6, 837-845, 2003; Ramsey et al., Annu Rev Physiol 68, 619-647, 2006; Montell, Pflugers Arch 451, 19-28, 2005; Clapham, Nature 426, 517-524, 2003). Dynamic reorganization of the actin cytoskeleton is mediated by spatiotemporally regulated Ca2+ transport. 2+ These changes are dependent on cell influx (Zheng and Poo, Annu Rev Cell Dev Biol 23, 375-404, 2007; Brandman and Meyer, Science 322, 390-395, 2008; Collins and Meyer, Dev Cell 16, 160-161, 2009), and the small GTPases RhoA and Rac1 function as key modulators of these changes (Etienne-Manneville and Hall, Nature 420, 629-635, 2002; Raftopoulou and Hall, Dev Biol 265, 23-32, 2004). RhoA induces the formation of stress fibers and focal adhesions, while Rac1 mediates the formation of lamellipodia (Etienne-Manneville and Hall, Nature 420, 629-635, 2002). Transient receptor potential cation channel, subfamily C, member 5 (TRPC5) cooperates with TRPC6 to regulate Ca(2+) levels in renal podocytes and fibroblasts. 2+Regulating Ca influx, actin remodeling, and cell motility. 2+ Ca influx increases Rac1 activity, but TRPC6-mediated Ca 2+ Influx promotes RhoA activity. Genetic silencing of TRPC6 channels abolishes stress fibers and reduces focal contacts, resulting in a motile, migratory cell phenotype. In contrast, genetic silencing of TRPC5 channels favors stress fiber formation and results in a contractile cell phenotype. The results described herein reveal a conserved signaling mechanism by which TRPC5 and TRPC6 channels control a tightly regulated balance of cytoskeletal dynamics through differential binding to Rac1 and RhoA.

[0148] Actin cytoskeleton Ca 2+ Ca-dependent remodeling is a dynamic process that drives cell migration (Wei et al., Nature 457, 901-905, 2009). RhoA and Rac1 function as switches involved in the rearrangement of the cytoskeleton in migrating cells (Etienne-Manneville and Hall, Nature 420, 629-635, 2002; Raftopoulou and Hall, Dev Biol 265, 23-32, 2004). Rac1 activation mediates a motile cell phenotype, whereas RhoA activity promotes a contractile phenotype (Etienne-Manneville and Hall, Nature 420, 629-635, 2002). 2+ Ca plays a central role in small GTPase regulation (Aspenstrom et al., Biochem J 377, 327-337, 2004). 2+ Spatially and temporally restricted flickering of Ca is abundant near the leading edge of migrating cells (Wei et al., Nature 457, 901-905, 2009). 2+Microdomains participate in localized bursts of Rac1 activity (Gardiner et al., Curr Biol 12, 2029-2034, 2002; Machacek et al., Nature 461, 99-103, 2009) as a key apical event. To date, no studies have shown that Ca2+ is involved in GTPase regulation. 2+ The source of this influx is largely unknown. TRP (Transient Receptor Potential) channels mediate the temporally and spatially restricted Ca influx associated with cell migration in fibroblasts and neuronal growth cones. 2+ Specifically, TRPC5 channels are known regulators of neuronal growth cone guidance 1, and their activity in neurons depends on PI3K and Rac1 activity (Bezzerides et al., Nat Cell Biol 6, 709-720, 2004).

[0149] Podocytes are neuron-like cells derived from the metanephric mesenchyme of the renal glomerulus and are essential for the formation of the renal filtration apparatus (Somlo and Mundel, Nat Genet. 24, 333-335, 2000; Fukasawa et al., J Am Soc Nephrol 20, 1491-1503, 2009). Podocytes have an exquisitely refined repertoire of cytoskeletal adaptations to environmental cues (Somlo and Mundel, Nat Genet 24, 333-335, 2000; Garg et al., Mol Cell Biol 27, 8698-8712, 2007; Verma et al., J Clin Invest 116, 1346-1359, 2006; Verma et al., J Biol Chem 278, 20716-20723, 2003; Barletta et al., J Biol Chem 278, 19266-19271, 2003; Holzman et al., Kidney Int 56, 1481-1491, 1999; Ahola et al., Am J Pathol 155, 907-913, 1999; Tryggvason and Wartiovaara, N Engl J Med 354, 1387-1401, 2006; Schnabel and Farquhar, J Cell Biol 111, 1255-1263, 1990; Kurihara et al., Proc Natl Acad Sci USA 89, 7075-7079, 1992). Early events in podocyte injury include dysregulation of the actin cytoskeleton (Faul et al., Trends Cell Biol 17, 428-437, 2007; Takeda et al., J Clin Invest 108, 289-301, 2001; Asanuma et al., Nat Cell Biol 8, 485-491, 2006) and Ca 2+The vasoactive hormone angiotensin II induces a decrease in Ca2+ homeostasis in podocytes (Hunt et al., J Am Soc Nephrol 16, 1593-1602, 2005; Faul et al., Nat Med 14, 931-938, 2008). These changes are associated with the development of proteinuria, loss of albumin into the urinary cavity, and ultimately renal failure (Tryggvason and Wartiovaara, N Engl J Med 354, 1387-1401, 2006). 2+ Ca influx is induced, and prolonged treatment leads to the loss of stress fibers (Hsu et al., J Mol Med 86, 1379-1394, 2008). 2+ Although a link between influx and cytoskeletal reorganization has been recognized, the mechanisms by which podocytes sense and transduce extracellular cues that regulate cell shape and motility are unknown. Although TRP canonical 6 (TRPC6) channel mutations have been associated with podocyte damage (Winn et al., Science 308, 1801-1804, 2005; Reiser et al., Nat Genet 37, 739-744, 2005; Moller et al., J Am Soc Nephrol 18, 29-36, 2007; Hsu et al., Biochim Biophys Acta 1772, 928-936, 2007), little is known about the specific pathways that control this process. Furthermore, TRPC6 is closely homologous to six other members of the TRPC channel family (Ramsey et al., Annu Rev Physiol 68, 619-647, 2006; Clapham, Nature 426, 517-524, 2003). TRPC5 channels antagonize the activity of TRPC6 channels, controlling a tightly regulated balance of cytoskeletal dynamics through differential binding to distinct small GTPases.

[0150] Proteinuria Proteinuria is a pathological condition in which protein is present in the urine. Albuminuria is a type of proteinuria. Microalbuminuria occurs when the kidneys leak small amounts of albumin into the urine. In a normally functioning body, albumin is retained in the bloodstream by the kidneys and is therefore not normally present in the urine. Microalbuminuria is diagnosed by a 24-hour urine collection (20-200 μg / min) or, more commonly, by at least two high concentrations (30-300 mg / L). Microalbuminuria can be a precursor to diabetic nephropathy. Albumin levels exceeding these values ​​are called macroalbuminuria. For example, subjects with certain conditions, such as diabetic nephropathy, can progress from microalbuminuria to macroalbuminuria, potentially reaching the nephrotic range (>3.5 g / 24 hours) as kidney disease reaches advanced stages.

[0151] Causes of Proteinuria Proteinuria can be associated with many conditions, including focal segmental glomerulosclerosis, IgA nephropathy, diabetic nephropathy, lupus nephritis, membranoproliferative glomerulonephritis, progressive (crescentic) glomerulonephritis, and membranous glomerulonephritis.

[0152] A. Focal segmental glomerulosclerosis (FSGS) Focal segmental glomerulosclerosis (FSGS) is a disease that attacks the kidney's filtering system (glomeruli), causing severe scarring. FSGS is one of many causes of a condition known as nephrotic syndrome, which occurs when protein from the blood leaks into the urine (proteinuria).

[0153] Few treatments are available for patients with FSGS. Many patients are treated with steroid therapy, most of which have severe side effects. Some patients respond positively to immunosuppressants as well as blood pressure suppressants, which have been shown to reduce urinary protein levels. To date, there is no generally accepted, effective treatment or cure, and no FDA-approved drugs to treat FSGS. Therefore, more effective methods to reduce or suppress proteinuria are desirable.

[0154] B. IgA nephropathy IgA nephropathy (also known as IgA nephritis, IgAN, Berger's disease, and synpharyngitic glomerulonephritis) is a type of glomerulonephritis (inflammation of the kidney's glomeruli). IgA nephropathy is the most common form of glomerulonephritis worldwide. Primary IgA nephropathy is characterized by the deposition of IgA antibodies in the glomeruli. There are other diseases associated with glomerular IgA deposition, the most common of which is Henoch-Schönlein purpura (HSP), which many consider to be a systemic form of IgA nephropathy. Henoch-Schönlein purpura presents with a characteristic purpuric skin rash, arthritis, and abdominal pain and occurs more commonly in young adults (ages 16–35). HSP is associated with a more benign prognosis than IgA nephropathy. IgA nephropathy progresses slowly to chronic renal failure in 25–30% of cases over a 20-year period.

[0155] C. Diabetic nephropathy Diabetic nephropathy, also known as Kimmel-Stiel-Wilson syndrome and capillary glomerulonephritis, is a progressive kidney disease caused by vascular damage to the capillaries of the renal glomeruli. It is characterized by nephrotic syndrome and diffuse glomerulosclerosis. It results from long-term diabetes and is the primary cause of dialysis. The earliest detectable change in the course of diabetic nephropathy is thickening of the glomeruli. At this stage, the kidneys may begin to produce more serum albumin than normal in the urine. As diabetic nephropathy progresses, an increasing number of glomeruli are destroyed by nodular glomerulosclerosis, leading to increased amounts of albumin excreted in the urine.

[0156] D. Lupus nephritis Lupus nephritis is a kidney disorder that is a complication of systemic lupus erythematosus (SLE). It occurs when antibodies and complement build up in the kidneys, causing inflammation. It often leads to proteinuria and can rapidly progress to kidney failure. Nitrogenous waste products build up in the bloodstream. SLE causes various disorders of the internal structures of the kidneys, including interstitial nephritis. Lupus nephritis affects approximately 3 in 10,000 people.

[0157] E. Membranoproliferative glomerulonephritis I / II / III Membranoproliferative glomerulonephritis (MPN) is a type of glomerulonephritis caused by deposits in the glomerular mesangium and basement membrane, complement activation, and glomerular damage. There are three types of MNPN: Type I, which is caused by immune complex deposition in the kidney and is thought to be associated with the classical complement pathway; Type II, which is similar to MNPN but is thought to be associated with the alternative complement pathway; and Type III, which is very rare, is characterized by admixture of subepithelial deposits and the typical pathological findings of Type I disease.

[0158] F. Progressive (crescentic) glomerulonephritis Progressive (crescentic) glomerulonephritis (PG) is a renal syndrome that, if left untreated, rapidly progresses to acute renal failure and death within several months. In 50% of cases, PG is associated with an underlying condition such as Goodpasture's syndrome, systemic lupus erythematosus, or Wegener's granulomatosis; the remaining cases are idiopathic. Regardless of the underlying condition, PG involves severe damage to the renal glomeruli, many of which contain characteristic crescent-shaped scars. Patients with PG have hematuria, proteinuria, and may also have hypertension and edema. While the clinical picture is consistent with nephritic syndrome, the degree of proteinuria may exceed 3 g / 24 hours, a range associated with nephrotic syndrome. Untreated, the disease can progress to decreased urine output (oliguria), which is associated with decreased renal function.

[0159] G. Membranous glomerulonephritis Membranous glomerulonephritis (MGN) is a slowly progressive kidney disease that primarily affects patients aged 30 to 50 years, usually Caucasians. It can develop into nephrotic syndrome. MGN is caused by circulating immune complexes. Current research indicates that the majority of immune complexes are formed via antibody binding to antigens in situ on the glomerular basement membrane. The antigens may be endogenous to the basement membrane or deposited from the systemic circulation.

[0160] H. Obesity Experimentally induced TrpC5 deficiency in mice has been shown to cause a positive energy balance leading to excessive weight gain (Y Gao et al., Cell Rep 2017, 18(3), pp.583-92). Therefore, agonism of TrpC5 may lead to a reduction in obesity.

[0161] Measurement of urine protein levels Protein levels in urine can be measured using methods known in the art. Until recently, accurate measurement of protein required a 24-hour urine collection. In a 24-hour collection, the patient urinates and deposits the urine in a container that is kept refrigerated until the patient needs to go to the toilet. The patient is instructed to begin collecting urine after the first toilet visit of the morning. All remaining urine drops for the day are collected in the container. The next morning, the patient completes the collection by adding the first urination after waking up.

[0162] Recently, researchers have discovered that a single urine sample can provide the necessary information. The new technique compares the amount of albumin in a urine sample with the amount of creatinine, a waste product of normal muscle breakdown. This measurement is called the urinary albumin-to-creatinine ratio (UACR). A urine sample containing more than 30 milligrams of albumin per gram of creatinine (30 mg / g) signals a potential problem. If the lab test shows a value above 30 mg / g, another UACR test should be performed one to two weeks later. If the second test still shows high levels of protein, the person has persistent proteinuria, a sign of declining kidney function, and additional testing to assess kidney function is warranted.

[0163] Tests that measure the amount of creatinine in the blood also indicate whether a subject's kidneys are efficiently removing waste products. Too much creatinine in the blood is a sign of kidney damage in humans. Doctors can use creatinine measurements to estimate how efficiently the kidneys filter blood. This calculation is called the estimated glomerular filtration rate, or eGFR. Chronic kidney disease exists when the eGFR is below 60 milliliters per minute (mL / min).

[0164] TRPC5 TRPCs are a family of transient receptor potential cation channels in animals. TRPC5 is a subtype of the TRPC family of mammalian transient receptor potential ion channels. Three examples of TRPC5 are highlighted in Table 1 below.

[0165] [Table 1]

[0166] Thus, in certain embodiments, the present invention provides a method for treating or reducing the risk of developing kidney disease, comprising administering to a subject in need thereof a therapeutically effective amount of a TRPC5 inhibitory compound of the present invention (e.g., a TRPC5 inhibitory compound of Formula I or II), or a pharmaceutical composition comprising the compound.

[0167] In some embodiments, the renal disease is selected from the group consisting of focal segmental glomerulosclerosis (FSGS), diabetic nephropathy, Alport syndrome, hypertensive renal disease, nephrotic syndrome, steroid-resistant nephrotic syndrome, minimal change disease, membranous nephropathy, idiopathic membranous nephropathy, membranoproliferative glomerulonephritis (MPGN), immune complex-mediated MPGN, complement-mediated MPGN, lupus nephritis, post-infectious glomerulonephritis, thin basement membrane disease, mesangial proliferative glomerulonephritis, amyloidosis (primary), c1q nephropathy, rapidly progressive GN, anti-GBM disease, C3 glomerulonephritis, hypertensive nephrosclerosis, and IgA nephropathy. In some embodiments, the renal disease is proteinuria. In some embodiments, the renal disease is microalbuminuria or macroalbuminuria.

[0168] The present invention also provides a method for treating or reducing the risk of developing anxiety, depression, or cancer, comprising administering to a subject in need thereof a therapeutically effective amount of a TRPC5 inhibitory compound of the present invention (e.g., a TRPC5 inhibitory compound of Formula I or II), or a pharmaceutical composition containing the compound.

[0169] In certain embodiments, the present invention provides a method for treating or reducing the risk of developing obesity, comprising administering to a subject in need thereof a therapeutically effective amount of a TRPC5 agonist compound of the present invention (e.g., a TRPC5 agonist compound of Formula I or II), or a pharmaceutical composition comprising the compound.

[0170] Treatment target In one aspect of the invention, the subject is selected on the basis of having or being at risk of developing kidney disease, anxiety, depression, or cancer.

[0171] Subjects at risk for developing or having proteinuria include those with diabetes, hypertension, or certain family histories. Diabetes is the leading cause of end-stage renal disease (ESRD) in the United States. In both type 1 and type 2 diabetes, albumin in the urine is one of the first signs of deteriorating kidney function. As kidney function declines, the amount of albumin in the urine increases. Another risk factor for developing proteinuria is high blood pressure. Proteinuria in people with high blood pressure is an indicator of declining kidney function. If high blood pressure is not controlled, it can progress to complete kidney failure. African Americans have higher blood pressure than Caucasians, even when their blood pressure is only slightly elevated, making them more likely to develop kidney problems. Other groups at risk for proteinuria include American Indians, Hispanics / Latinos, Pacific Islanders, the elderly, and overweight subjects.

[0172] In one aspect of the present invention, a subject is selected based on whether or not they have proteinuria or are at risk for developing proteinuria. A subject with or at risk for developing proteinuria is a subject with one or more symptoms of the condition. Symptoms of proteinuria are known to those skilled in the art and include, but are not limited to, a large amount of protein in the urine, which may appear foamy in the toilet. A large amount of protein loss can cause edema, resulting in swelling of the hands, feet, abdomen, or face. These are signs of significant protein loss and indicate the progression of kidney disease. Laboratory testing is the only way to determine whether protein is present in a subject's urine before extensive kidney damage occurs.

[0173] The method is effective in a variety of subjects, including mammals, e.g., humans, and other animals, e.g., laboratory animals, e.g., mice, rats, rabbits, or monkeys, or domestic and farm animals, e.g., cats, dogs, goats, sheep, pigs, cows, or horses. In some embodiments, the subject is a mammal. In some embodiments, the subject is a human. [Example]

[0174] The present invention is further described in the following examples, which do not limit the scope of the invention described in the claims.

[0175] Example 1: Synthesis Method The following provide synthetic routes to exemplary compounds of the invention.

[0176] Preparation of Intermediate A

[0177] [ka]

[0178] 2,4,5-Tribromo-1-[(4-chlorophenyl)methyl]-1H-imidazole A mixture of 2,4,5-tribromo-1H-imidazole (120 g, 393.75 mmol, 1 equiv.), 1-(bromomethyl)-4-chlorobenzene (100 g, 486.67 mmol, 1.236 equiv.), and CsCO (200 g, 613.84 mmol, 1.559 equiv.) in DMF (1000 mL) was stirred at room temperature for 16 h. To the reaction mixture, EtOAc (500 mL) and HO (300 mL) were added. The organic layer was washed with HO (2×300 mL) and brine (300 mL), dried over anhydrous NaSO, and filtered. The filtrate was concentrated to give 2,4,5-tribromo-1-[(4-chlorophenyl)methyl]-1H-imidazole (170 g, crude) as a pale yellow solid.

[0179] 4,5-Dibromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole A mixture of 2,4,5-tribromo-1-[(4-chlorophenyl)methyl]-1H-imidazole (175 g, 407.61 mmol, 1 equiv.), 3-(trifluoromethoxy)phenol (87.5 g, 491.27 mmol, 1.205 equiv.), and KCO (175 g, 1266.23 mmol, 3.106 equiv.) in DMF (1000 mL) was stirred at 100 °C for 16 h. The reaction mixture was cooled to room temperature, and EtOAc (750 mL) and HO (500 mL) were added. The organic layer was washed with HO (2 × 300 mL) and brine (150 mL), then dried over anhydrous NaSO and filtered. The filtrate was concentrated to give the crude product, which was purified by silica gel column chromatography eluting with PE:EtOAc (20:1 to 10:1) to give 4,5-dibromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole (200 g, 93.19%) as a pale yellow solid.

[0180] Methyl 4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxylate (Intermediate A) To a stirred solution of 4,5-dibromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole (10.52 g, 19.98 mmol, 1 equiv.) in THF (100 mL) was added n-BuLi (25.8 mL, 64.57 mmol, 1 equiv.) dropwise at −78° C. The resulting mixture was stirred at −78° C. for 30 minutes, and then CO (g) was bubbled through the mixture at −78° C. for 50 minutes. The reaction mixture was stirred at −78° C. for 30 minutes. HATU (36.8 g, 96.86 mmol, 1.5 equiv.), MeOH (180 mL), and TEA (70 mL) were added to the above solution, and then the resulting mixture was stirred at room temperature for 12 hours. The reaction mixture was filtered, and the filtrate was concentrated to give the crude product, which was purified by silica gel column chromatography eluting with PE:EA (20:1 to 5:1) to give methyl 4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxylate (26 g, 79.63%) as a white solid. 1 H NMR (400 MHz, chloroform-d) δ 7.46-7.40 (m, 1H), 7.36-7.30 (m, 2H), 7.22 (dd, J = 8.4, 2.4 Hz, 3H), 7.13 (dtd, J = 9.9, 2.1, 1.1 Hz, 2H), 5.52 (s, 2H), 3.89 (s, 3H).

[0181] The preparation of intermediates C, E, G, I, J, K, L, M, and N shown in the table below follows the methods and protocols described for the synthesis of intermediate A, starting with the appropriate halide and phenol.

[0182] [Table 2-1]

[0183] [Table 2-2]

[0184] [Table 2-3]

[0185] 4-Bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxylic acid (Intermediate B) To a mixture of methyl 4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxylate (8 g, 15.82 mmol, 1 equiv.) in THF (50 mL) and HO (50 mL) was added LiOH (3.8 g, 158.21 mmol, 10 equiv.) and stirred at room temperature for 10 h. The resulting mixture was extracted with EA (4 × 200 mL). The combined organic layers were washed with water (1 × 100 mL) and dried over anhydrous NaSO. After filtration, the filtrate was concentrated under reduced pressure. The residue / crude product was purified by reverse-phase flash using the following conditions (Column: spherical C18 column, 20-40 μm, 120 g, Mobile phase A: water (0.1% AcOH), Mobile phase B: ACN, Flow rate: 60 mL / min, Gradient: 80-90% B in 15 min, 254 nm) to give 4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxylic acid (7.5 g, 96.42%) as a white solid. 1 H NMR(400MHz, DMSO-d6)δ13.41(s, 1H), 7.59(t, J = 8.3Hz, 1H), 7.42(dd, J = 8.7, 2.1Hz, 3H), 7.37(dt, J = 8.2, 1.5Hz, 1H), 7.33-7.29(m, 1H), 7.29-7.24(m, 2H), 5.51(s, 2H)

[0186] The preparation of intermediates D, F and H shown in the table below follows the methods and protocols described for the synthesis of intermediate B, starting with the appropriate intermediate.

[0187] [Table 3]

[0188] Preparation of Compound 1

[0189] [ka]

[0190] Methyl 4-[(2-[[(tert-butoxy)carbonyl]amino]ethyl)(methyl)amino]-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxylate A mixture of methyl 4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxylate (2 g, 3.96 mmol, 1 equiv.), tert-butyl N-[2-(methylamino)ethyl]carbamate (1.4 g, 7.91 mmol, 2.00 equiv.), Xantphos (686.6 mg, 1.19 mmol, 0.3 equiv.), Pd(dba) (362.2 mg, 0.40 mmol, 0.1 equiv.), and CsCO (3.9 g, 11.87 mmol, 3 equiv.) in dioxane (30 mL, 89.53 equiv.) was stirred at 100 °C for 14 h. The reaction mixture was filtered, and the filtrate was concentrated to give a crude product, which was purified by silica gel column chromatography eluting with PE:EA (10:1 to 3:2) to give methyl 4-[(2-[[(tert-butoxy)carbonyl]amino]ethyl)(methyl)amino]-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxylate (630 mg, 26.59%) as a pale yellow oil. 1H NMR(400MHz, chloroform-d)δ7.41(t, J = 8.2 Hz, 1H), 7.34-7.26(m, 3H), 7.18(t, J = 8.7Hz, 2H), 7.09 (d,J = 8.4Hz, 1H), 5.39(s, 2H), 5.28(s, 1H), 3.78(s, 3H), 3.38(dd, J = 18.5, 5.7Hz, 4H), 2.94(s, 3H), 1.43(s, 9H)

[0191] 4-[(2-aminoethyl)(methyl)amino]-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxylate To a stirred solution of methyl 4-[(2-[[(tert-butoxy)carbonyl]amino]ethyl)(methyl)amino]-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxylate (700 mg, 1.17 mmol, 1 equiv.) in DCM (30 mL) was added TFA (10 mL) dropwise at room temperature. The resulting mixture was then stirred at room temperature for 2 h. The reaction mixture was basified to pH 10 with KCO and extracted with ethyl acetate (5 x 50 mL), and the organic layer was then washed with brine (2 x 50 mL), dried over anhydrous NaSO, and filtered. The filtrate was concentrated to give methyl 4-[(2-aminoethyl)(methyl)amino]-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxylate (500 mg, crude) as a pale yellow oil.

[0192] 1-[(4-chlorophenyl)methyl]-4-methyl-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepin-8-one To a stirred mixture of methyl 4-[(2-aminoethyl)(methyl)amino]-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxylate (370 mg, 0.74 mmol, 1 equiv.) in dioxane (15 mL), NaH (59.3 mg, 1.48 mmol, 2.00 equiv., 60%) was added under a nitrogen atmosphere at 0° C. for 0.5 h. The resulting mixture was stirred at 100° C. for an additional 4 h. Ethyl acetate (100 mL) and brine (50 mL) were added to the resulting mixture, and then the aqueous layer was extracted with ethyl acetate (100 mL). The combined organic layers were dried over anhydrous NaSO and filtered. The filtrate was concentrated to give the crude product, which was purified by reverse-phase flash chromatography under the following conditions (column: spherical C18 column, 20–40 μm, 120 g; mobile phase A: water (0.1% HOAc), mobile phase B: ACN; flow rate: 60 mL / min; gradient: 40% B to 70% B in 30 min, 254 nm) to give 1-[(4-chlorophenyl)methyl]-4-methyl-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepin-8-one (23 mg, 6.64%) as a pale yellow oil. 1 H NMR (400 MHz, chloroform-d) δ 7.37 (q, J = 10.5, 9.4 Hz, 1H), 7.27 (d, J = 11.1 Hz, 5H), 7.19 (d, J = 8.5 Hz, 1H), 7.07 (d, J = 8.3 Hz, 1H), 5.80 (s, 1H), 5.53 (s, 2H), 3.54-3.33 (m, 4H), 3.03 (s, 3H).

[0193] 1-[(4-chlorophenyl)methyl]-4-methyl-7-[3-(oxan-2-yloxy)propyl]-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepin-8-one A mixture of 1-[(4-chlorophenyl)methyl]-4-methyl-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepin-8-one (23 mg, 0.05 mmol, 1 equiv.), 2-(3-bromopropoxy)oxane (22.0 mg, 0.10 mmol, 2 equiv.), and KCO (20.4 mg, 0.15 mmol, 3 equiv.) in DMF (5 mL) was stirred at room temperature for 8 h. To the reaction was added EtOAc (50 mL) and HO (50 mL). The organic layer was washed with brine (2x30 mL) and concentrated to give a residue that was purified by reverse-phase flash chromatography under the following conditions (Column: spherical C18 column, 20-40 μm, 40 g; Mobile phase A: water (0.1% HOAc), Mobile phase B: ACN; Flow rate: 40 mL / min; Gradient: 50% B to 70% B in 30 min, 254 nm) to give 1-[(4-chlorophenyl)methyl]-4-methyl-7-[3-(oxan-2-yloxy)propyl]-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepin-8-one (10 mg, 33.33%) as a pale yellow oil.

[0194] 1-[(4-chlorophenyl)methyl]-7-(3-hydroxypropyl)-4-methyl-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepin-8-one (Compound 1) To a stirred solution of 1-[(4-chlorophenyl)methyl]-4-methyl-7-[3-(oxan-2-yloxy)propyl]-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepin-8-one (10 mg) in THF (5 mL) was added dropwise at room temperature. The resulting mixture was then stirred at room temperature for 1 hour. The reaction mixture was basified to pH 10 with KCO and extracted with ethyl acetate (3 x 50 mL). The organic layer was then washed with brine (2 x 20 mL), dried over anhydrous NaSO, and filtered. The filtrate was concentrated to give the crude product, which was purified by preparative chiral HPLC (Column: XBridge Preparative C18 OBD Column, 5 μm, 19*150 mm; Mobile Phase A: Water (0.1% FA), Mobile Phase B: ACN; Flow Rate: 20 mL / min; Gradient: 50% B to 85% B in 7 min; 254 nm; RT: 6.5 min) to give 1-[(4-chlorophenyl)methyl]-7-(3-hydroxypropyl)-4-methyl-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepin-8-one (2.2 mg, 25.53%) as a pale yellow oil. 1 H NMR(400MHz, methanol-d4)δ7.50(t, J = 8.2Hz, 1H), 7.33-7.26(m, 2H), 7.23-7.15(m, 5H), 5.47(s, 2H), 3.59-3.54(m, 2H), 3.53-3.41(m, 6H), 3.01(s, 3H), 1.79-1.70(m, 2H). Molecular formula C 24 H 24 Calculated [M+H] for ClF3N4O4 + :525, Observed:525.

[0195] Preparation of Compound 2

[0196] [ka]

[0197] 4-Bromo-1-[(4-chlorophenyl)methyl]-N-(3-hydroxypropyl)-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxamide A mixture of 3-aminopropan-1-ol (77.3 mg, 1.03 mmol, 1.5 equiv.) and 4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carbonyl chloride (350 mg, 0.69 mmol, 1 equiv.) in DCM (20 mL) and TEA (0.5 mL, 4.71 mmol, 5.0 equiv.) was stirred at room temperature for 16 h. The reaction mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography eluting with PE:EtOAc (1:1 to 1:2) to give 4-bromo-1-[(4-chlorophenyl)methyl]-N-(3-hydroxypropyl)-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxamide (220 mg, 58.43%) as a pale yellow solid. 1 H NMR(300MHz,chloroform-d)δ7.43(td, J = 8.0, 7.5, 1.0Hz, 1H), 7.34-7.28(m, 4H), 7.26(d, J = 2.4Hz, 2H), 7.24(s, 1H), 7.22-7.18(m, 1H), 7.16-7.08(m, 2H), 5.58(s, 2H), 3.61(dt, J = 9.8, 6.0Hz, 4H), 1.78(p, J = 5.8Hz, 2H).

[0198] 4-Bromo-1-[(4-chlorophenyl)methyl]-N-[3-(oxan-2-yloxy)propyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxamide To a mixture of 4-bromo-1-[(4-chlorophenyl)methyl]-N-(3-hydroxypropyl)-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxamide (1.00 g, 1.82 mol, 1 equiv.) in DCM (30 mL) was added 3,4-dihydro-2H-pyran (306.6 mg, 3.64 mol, 2.0 equiv.) and p-toluenesulfonic acid (15.7 mg, 0.09 mmol, 0.05 equiv.). The resulting mixture was stirred at room temperature for 16 h. The reaction mixture was concentrated to give the crude product, which was purified by silica gel column chromatography eluting with PE:EtOAc (5:1 to 3:1) to give 4-bromo-1-[(4-chlorophenyl)methyl]-N-[3-(oxan-2-yloxy)propyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxamide (1.00 g, 86.71%) as a pale yellow oil. 1 H NMR (300 MHz, chloroform-d) δ 7.52-7.05 (m, 8H), 6.92 (s, 1H), 5.56 (s, 2H), 4.58 (dd, J = 4.7, 2.6 Hz, 1H), 4.12-3.30 (m, 9H), 2.01-1.33 (m, 5H).

[0199] 2-(1-[4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazol-5-yl]-N-[3-(oxan-2-yloxy)propyl]formamido)acetic acid ethyl ester To a mixture of 4-bromo-1-[(4-chlorophenyl)methyl]-N-[3-(oxan-2-yloxy)propyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxamide (1000 mg, 1.58 mmol, 1 equiv.) in DMF (20 mL) and NaH (126.4 mg, 3.16 mmol, 2.0 equiv., 60%), 2-bromoethyl acetate (527.8 mg, 3.16 mmol, 2.0 equiv.) was added dropwise at 0 °C. The resulting mixture was warmed to room temperature and stirred at room temperature for 16 h. HO (100 mL) was added to the reaction mixture, and the resulting mixture was extracted with ethyl acetate (3 × 100 mL). The organic layer was washed with brine (100 mL), dried over anhydrous water, and filtered. The filtrate was concentrated to give a residue, which was purified by silica gel column chromatography eluting with PE:EtOAc (5:1 to 3:1) to give ethyl 2-(1-[4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazol-5-yl]-N-[3-(oxan-2-yloxy)propyl]formamido)acetate (970 mg, 87.08%) as a pale yellow oil. 1 H NMR(300MHz, chloroform-d)δ7.45-7.35(m, 4H), 7.34-7.30(m, 8H), 7.09(d, J = 15.8Hz, 8H), 6.93(s, 1H), 5.57(d, J = 5.3Hz, 2H), 5.17(s, 6H), 4.53(d, J = 27.3Hz, 1H), 4.25(q, J = 7.7, 6.9Hz, 2H), 3.96-3.18(m, 9H), 1.96-1.70(m, 3H), 1.31(d, J = 3.1Hz, 4H).

[0200] 2-(1-[4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazol-5-yl]-N-[3-(oxan-2-yloxy)propyl]formamido)-N-methylacetamide A mixture of ethyl 2-(1-[4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazol-5-yl]-N-[3-(oxan-2-yloxy)propyl]formamido)acetate (970 mg, 1.35 mmol, 1 equiv) in 2 M methylamine in methanol (3.00 mL) was irradiated with microwave radiation at 60° C. for 1 hour. The reaction mixture was cooled to room temperature and concentrated under reduced pressure. The residue was purified by silica gel column chromatography eluting with PE:EtOAc (1:6 to 1:9) to give 2-(1-[4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazol-5-yl]-N-[3-(oxan-2-yloxy)propyl]formamide)-N-methylacetamide (600 mg, 63.17%) as a colorless oil. 1 H NMR(300MHz, DMSO-d6)δ8.07-7.88(m, 1H), 7.61-7.47(m, 1H), 7.38(d, J = 1.6Hz, 6H), 7.24(dd, J = 27.9, 9.7Hz, 5H), 5.12(s, 2H), 4.49(d, J = 30.7Hz, 2H), 4.15(s, 1H), 3.79-3.55(m, 1H), 3.39(s, 6H), 3.27(d, J = 4.1Hz, 0H), 2.60(dd, J = 13.1, 4.4Hz, 5H), 1.87-1.26(m, 14H).

[0201] 1-[(4-chlorophenyl)methyl]-4-methyl-7-[3-(oxan-2-yloxy)propyl]-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-5,8-dione 2-(1-[4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazol-5-yl]-N-[3-(oxan-2-yloxy)propyl]formamide)-N-methylacetamide (500 mg, 0.71 mmol, 1 equiv.), Pd(dba) CHCl (73.5 mg, 0.07 mmol, 0.1 equiv.), P(o-Tol) (43.2 mg, 0.14 mmol, 0.2 equiv.), and CsC in toluene (10.0 mL). O3 A mixture of (462.9 mg, 1.42 mmol, 2.0 equiv.) was irradiated with microwave radiation at 120 °C for 4 h. The reaction mixture was cooled to room temperature and concentrated under reduced pressure. The residue was purified by silica gel column chromatography eluting with PE:EtOAc (3:1 to 1:1) to give 1-[(4-chlorophenyl)methyl]-4-methyl-7-[3-(oxan-2]-yloxy)propyl]-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-5,8-dione (300 mg, 67.79%) as a pale yellow oil.

[0202] 1-[(4-chlorophenyl)methyl]-7-(3-hydroxypropyl)-4-methyl-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-5,8-dione (Compound 2) A mixture of 1-[(4-chlorophenyl)methyl]-4-methyl-7-[3-(oxan-2-yloxy)propyl]-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-5,8-dione (300 mg, 0.48 mmol, 1 equiv.) in THF (10 mL) and HCl (6 M) (20 mL) was stirred at room temperature for 2 h. The reaction mixture was concentrated to give a residue. The residue was basified to pH 9 with saturated KCO(aq), and the mixture was then extracted with ethyl acetate (3 x 100 mL). The organic layer was washed with brine (50 mL) and concentrated to give the crude product, which was purified by reverse-phase flash chromatography under the following conditions (Column: spherical C18 column, 20–40 μm, 120 g; Mobile phase A: water (0.1% HOAc), Mobile phase B: ACN; Flow rate: 60 mL / min; Gradient: 50% B to 60% B in 15 min, 254 nm) to give 1-[(4-chlorophenyl)methyl]-7-(3-hydroxypropyl)-4-methyl-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-5,8-dione (220 mg, 84.78%) as a pale yellow oil. 1 H NMR(300MHz, DMSO-d6)δ7.59-7.54(m, 1H), 7.42-7.26(m, 7H), 5.44(s, 2H), 4.48(t, J = 5.1Hz, 1H), 4.05(s, 2H), 3.51(t, J = 7.1Hz, 2H), 3.39-3.33(m, 2H), 3.20(s, 3H), 1.69-1.60(m, 2H). Molecular formula C 24 H 22 Calculated [M+H] for ClF3N4O5 + :539, Observed:539.

[0203] The preparation of compounds 3-5 shown in the table below follows the methods and protocols described for the synthesis of compound 2, starting with the appropriate intermediate E.

[0204] [Table 4]

[0205] Preparation of Compounds 6 and 7

[0206] [ka]

[0207] 1-[(1R)-1-(4-chlorophenyl)ethyl]-2-(3-fluorophenoxy)-7-(3-hydroxypropyl)-4-methyl-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-5,8-dione and 1-[(1S)-1-(4-chlorophenyl)ethyl]-2-(3-fluorophenoxy)-7-(3-hydroxypropyl)-4-methyl-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-5,8-dione The crude product, 1-(1-(4-chlorophenyl)ethyl)-2-(3-fluorophenoxy)-7-(3-hydroxypropyl)-4-methyl-1,4,6,7-tetrahydroimidazo[4,5-e][1,4]diazepine-5,8-dione (200 mg), was purified by chiral HPLC under the following conditions (column: CHIRALPAK IE, 2*25 cm, 5 μm; mobile phase A: hexane 0.1% DEA - HPLC; mobile phase B: EtOH - HPLC; flow rate: 17 mL / min; gradient: 50B to 50B in 11 min; 220 / 254 nm; RT1: 7.423; RT2: 9.034) to give the separated enantiomers, compound 6 (RT 7.423 min, 73 mg, 23.14%) and compound 7 (RT 9.034 min, 77 mg, 24.41%).

[0208] Compound Characterization 6: 1H NMR(400MHz, methanol-d4)δ7.44-7.41(m, 2H), 7.38-7.31(m, 3H), 6.98-6.93(m, 1H), 6.87-6.82(m, 2H), 6.25(q, J = 7.2Hz, 1H), 4.16(s, 2H), 3.73-3.63(m, 2H), 3.56(t, J = 6.1Hz, 2H), 3.32(s, 3H), 2.00(d, J = 7.2Hz, 3H), 1.85(p, J = 6.6Hz, 2H). Molecular formula C 24 H 24 Calculated [M+H] for ClFN4O4 + :487, Observed:487.

[0209] Compound Characterization 7: 1 H NMR(400MHz, methanol-d4)δ7.43-7.41(m, 2H), 7.40-7.30(m, 3H), 6.98-6.95(m, 1H), 6.87-6.82(m, 2H), 6.25(q, J = 7.2Hz, 1H), 4.17(s, 2H), 3.68(t, J = 6.8, 2H), 3.56(t, J = 6.1Hz, 2H), 3.32(s, 3H), 2.00(d, J = 7.2Hz, 3H), 1.85(p, J = 6.7Hz, 2H). Molecular formula C 24 H 24 Calculated [M+H] for ClFN4O4 + :487, Observed:487.

[0210] Preparation of Compound 8

[0211] [ka]

[0212] 1-[(4-chlorophenyl)methyl]-7-(3-methoxypropyl)-4-methyl-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-5,8-dione (Compound 8) To a mixture of 1-[(4-chlorophenyl)methyl]-7-(3-hydroxypropyl)-4-methyl-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-5,8-dione (120 mg, 0.22 mmol, 1 equiv.) in DMF (20 mL) was added NaH (44.5 mg, 1.11 mmol, 5 equiv., 60 wt%) under a nitrogen atmosphere at 0 °C for 0.5 h. To the above mixture was added CHI (94.8 mg, 0.67 mmol, 3 equiv.) at 0 °C. The resulting mixture was stirred at 80 °C for an additional 16 h. The mixture was basified to pH 10 with KCO(aq) and extracted with ethyl acetate (5 × 50 mL). The organic layer was then washed with brine (2 × 50 mL) and concentrated under reduced pressure. The residue was purified by reverse-phase flash chromatography under the following conditions (Column: XBridge Preparative OBD C18 column 30*150 mm 5 μm; Mobile phase A: water (0.05% TFA), Mobile phase B: ACN; Flow rate: 60 mL / min; Gradient: 45% B to 80% B in 7 min; 220 nm; RT: 6.55 min) to give 1-[(4-chlorophenyl)methyl]-7-(3-methoxypropyl)-4-methyl-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-5,8-dione (34.1 mg, 27.70%) as a pale yellow oil. 1 H NMR(400MHz, methanol-d4)δ7.52(t, J = 8.2Hz, 1H), 7.36-7.28(m, 6H), 7.20(d, J = 8.1Hz, 1H), 5.50(s, 2H), 4.05(s, 2H), 3.63(t, J = 6.9Hz, 2H), 3.37-3.32(m, 5H), 3.29(s, 3H), 1.89-1.82(m, 2H). Molecular formula C 25 H 24 Calculated [M+H] for ClF3N4O5 + :553, Observed:553.

[0213] Preparation of Compound 9

[0214] [ka]

[0215] 4-Bromo-1-[(4-chlorophenyl)methyl]-N-[(2,2-dimethyl-1,3-dioxolan-4-yl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxamide To a mixture of (2,2-dimethyl-1,3-dioxolan-4-yl)methanamine (617.2 mg, 4.71 mmol, 2.0 equiv) in DCM (30 mL) and TEA (1.0 mL, 9.69 mmol, 3.0 equiv) was added 4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carbonyl chloride (1.2 g, 2.35 mmol, 1 equiv) in DCM (20 mL) dropwise at 0° C. The resulting mixture was allowed to warm to room temperature and stirred at room temperature for 16 h. The reaction mixture was concentrated under reduced pressure and purified by silica gel column chromatography eluting with PE:EtOAc (3:1 to 2:1) to give 4-bromo-1-[(4-chlorophenyl)methyl]-N-[(2,2-dimethyl-1,3-dioxolan-4-yl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxamide (1.30 g, 91.37%) as a pale yellow oil. 1 H NMR(300MHz, chloroform-d)δ7.46-7.38(m, 1H), 7.35-7.29(m, 2H), 7.27-7.16(m, 3H), 7.15-7.08(m, 2H), 7.01(t, J = 5.6Hz, 1H), 5.59(s, 2H), 4.32(qd, J = 6.2, 3.8Hz, 1H), 4.07(dd, J = 8.4, 6.4Hz, 1H), 3.71-3.62(m, 2H), 3.57(dt, J = 14.1, 5.8Hz, 1H), 1.47(s, 3H), 1.38(s, 3H).

[0216] 2-(1-[4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazol-5-yl]-N-[(2,2-dimethyl-1,3-dioxolan-4-yl)methyl]formamido)acetic acid ethyl ester To a mixture of 4-bromo-1-[(4-chlorophenyl)methyl]-N-[(2,2-dimethyl-1,3-dioxolan-4-yl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxamide (1 g, 1.65 mol, 1 equiv.) in DMF (20 mL) was added NaH (99.2 mg, 2.48 mmol, 1.5 equiv., 60 wt%) under a nitrogen atmosphere at 0° C. for 0.5 h. To the above mixture was added 2-bromoethyl acetate (0.3 mL, 1.80 mmol, 1.636 equiv.) at 0° C. The resulting mixture was stirred at room temperature for an additional 16 h. Ethyl acetate (300 mL) and brine (100 mL) were added to the resulting mixture, and then the aqueous layer was extracted with ethyl acetate (100 mL). The combined organic layers were dried over anhydrous NaSO and filtered. The filtrate was concentrated to give the crude product, which was purified by silica gel column chromatography eluting with PE:EA (20:1 to 4:1) to give ethyl 2-(1-[4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazol-5-yl]-N-[(2,2-dimethyl-1,3-dioxolan-4-yl)methyl]formamido)acetate (1.1 g, 96.3%) as a pale yellow oil.

[0217] 2-(1-[4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazol-5-yl]-N-[(2,2-dimethyl-1,3-dioxolan-4-yl)methyl]formamide)-N-methylacetamide A mixture of ethyl 2-(1-[4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazol-5-yl]-N-[(2,2-dimethyl-1,3-dioxolan-4-yl)methyl]formamido)acetate (1.2 g, 1.74 mmol, 1 equiv) in 2 M methylamine in methanol (5 mL) was irradiated with microwave radiation at 60° C. for 1 hour. The mixture was cooled to room temperature. The reaction mixture was concentrated to give the crude product, which was purified by silica gel column chromatography eluting with PE:EA (1:1 to 1:8) to give 2-(1-[4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazol-5-yl]-N-[(2,2-dimethyl-1,3-dioxolan-4-yl)methyl]formamide)-N-methylacetamide (0.92 g, 78.37%) as a white solid.

[0218] 1-[(4-chlorophenyl)methyl]-7-[(2,2-dimethyl-1,3-dioxolan-4-yl)methyl]-4-methyl-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-5,8-dione A mixture of 2-(1-[4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazol-5-yl]-N-[(2,2-dimethyl-1,3-dioxolan-4-yl)methyl]formamide)-N-methylacetamide (1 g, 1.48 mmol, 1 equiv.), Pd(dba) CHCl (150 mg, 0.14 mmol, 0.098 equiv.), P(o-Tol) (90 mg, 0.30 mmol, 0.200 equiv.), and CsCO (1 g, 3.07 mmol, 2.074 equiv.) in toluene (15 mL) was irradiated with microwave radiation at 120 °C for 4 h. The mixture was cooled to room temperature. The reaction mixture was filtered, and the filtrate was concentrated to give the crude product, which was purified by silica gel column chromatography eluting with PE:EA (4:1 to 2:3) to give 1-[(4-chlorophenyl)methyl]-7-[(2,2-dimethyl-1,3-dioxolan-4-yl)methyl]-4-methyl-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-5,8-dione (470 mg, 53.39%) as a pale yellow oil.

[0219] 1-[(4-chlorophenyl)methyl]-7-(2,3-dihydroxypropyl)-4-methyl-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-5,8-dione (Compound 9) To a stirred solution of 1-[(4-chlorophenyl)methyl]-7-[(2,2-dimethyl-1,3-dioxolan-4-yl)methyl]-4-methyl-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-5,8-dione (600 mg, 1.01 mol, 1 equiv.) in THF (15 mL) was added HCl (15 mL) in HO (15.0 mL) dropwise at room temperature. The resulting mixture was then stirred at room temperature for 2 h. The mixture was basified to pH 10 with KCO(aq) and extracted with ethyl acetate (5 x 50 mL). The organic layer was then washed with brine (2 x 50 mL) and concentrated under reduced pressure. The residue was purified by reverse-phase flash chromatography under the following conditions (Column: spherical C18 column, 20–40 μm, 330 g; Mobile phase A: water (0.1% HOAc), Mobile phase B: ACN; Flow rate: 80 mL / min; Gradient: 60% B to 95% B in 35 min, 254 nm) to give 1-[(4-chlorophenyl)methyl]-7-(2,3-dihydroxypropyl)-4-methyl-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-5,8-dione (520 mg, 92.92%) as a white solid. 1 H NMR(300MHz, DMSO-d6)δ7.57(t, J = 8.3Hz, 1H), 7.41-7.35(m, 7H), 5.44(d, J = 1.8Hz, 2H), 4.81(d, J = 5.3Hz, 1H), 4.63(t, J = 5.7Hz, 1H), 4.25-4.01(m, 2H), 3.83-3.62(m, 1H), 3.55-3.48(m, 1H), 3.43-3.40(m, 1H), 3.32-3.24(m, 2H), 3.19(s, 3H). Molecular formula C 24 H 22 Calculated [M+H] for ClF3N4O6 + :555, Observed:555.

[0220] Preparation of Compound 10

[0221] [ka]

[0222] 4-Bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxylic acid A mixture of methyl 4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxylate (2.5 g, 4.94 mmol, 1 equiv.) and LiOH (1.2 g, 49.44 mmol, 10 equiv.) in THF (50 mL) and HO (50 mL) was stirred at room temperature for 10 h. The resulting mixture was extracted with EA (4 × 200 mL). The combined organic layers were washed with water (1 × 100 mL) and dried over anhydrous NaSO. After filtration, the filtrate was concentrated under reduced pressure. The residue / crude product was purified by reverse-phase flash using the following conditions (Column: spherical C18 column, 20-40 μm, 120 g Mobile phase A: water (0.1% AcOH), Mobile phase B: ACN, Flow rate: 60 mL / min, Gradient: 80-90% B in 15 min, 254 nm) to give 4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxylic acid (2.1 g, 86.40%) as a white solid. 1 H NMR(400MHz, DMSO-d6)δ13.41(s, 1H), 7.59(t, J = 8.3Hz, 1H), 7.42(dd, J = 8.7, 2.1Hz, 3H), 7.37(dt, J = 8.2, 1.5Hz, 1H), 7.33-7.29(m, 1H), 7.29-7.24(m, 2H), 5.51(s, 2H).

[0223] 4-Bromo-1-[(4-chlorophenyl)methyl]-N-(2-hydroxyethyl)-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxamide To a mixture of 4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxylic acid (1.1 g, 2.24 mmol, 1 equiv.) and 1 drop of DMF in DCM (20 mL) was added oxalic dichloride (0.9 g, 6.71 mmol, 3 equiv.) dropwise at 0° C. The mixture was stirred at room temperature for 3 hours. The resulting mixture was concentrated under vacuum. The solid was dissolved in DCM (30 mL) to give solution A for the next reaction. A mixture of 2-aminoethan-1-ol (0.4 g, 6.71 mmol, 3 equiv.) and triethylamine (0.7 g, 6.71 mmol, 3 equiv.) in DCM (10 mL) was cooled to 0° C., and solution A was added dropwise at 0° C. under a nitrogen atmosphere. The mixture was stirred at room temperature for 3 hours. The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography eluting with PE / EA (2 / 1 to 1 / 1) to give 4-bromo-1-[(4-chlorophenyl)methyl]-N-(2-hydroxyethyl)-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxamide (0.91 g, 76.06%) as a white solid. 1 H NMR(400MHz, DMSO-d6)δ7.99(t, J = 5.6Hz, 1H), 7.58(t, J = 8.3Hz, 1H), 7.45-7.37(m, 2H), 7.38-7.26(m, 5H), 5.39(s, 2H), 4.77(t, J = 5.4Hz, 1H), 3.47(q, J = 6.1Hz, 2H), 3.35-3.27(m, 2H).

[0224] 4-Bromo-1-[(4-chlorophenyl)methyl]-N-[2-(oxan-2-yloxy)ethyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxamide To a stirred mixture of 4-bromo-1-[(4-chlorophenyl)methyl]-N-(2-hydroxyethyl)-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxamide (0.9 g, 1.68 mmol, 1 equiv.) and 4-methylbenzene-1-sulfonic acid (0.0 g, 0.17 mmol, 0.1 equiv.) in DCM (50 mL) was added 3,4-dihydro-2H-pyran (0.0 g, 0.34 mmol, 0.2 equiv.) at room temperature under a nitrogen atmosphere. The mixture was reacted at room temperature for 10 hours. The resulting mixture was quenched with water (100 mL) and extracted with DCM (4 × 100 mL). The combined organic layers were washed with water (1 × 100 mL) and dried over anhydrous NaSO. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography eluting with PE / EA (4 / 1) to give 4-bromo-1-[(4-chlorophenyl)methyl]-N-[2-(oxan-2-yloxy)ethyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxamide (0.8 g, 76.81%) as a yellow oil. 1 H NMR(400MHz, DMSO-d6)δ8.09(d, J = 5.6Hz, 1H), 7.60-7.55(m, 1H), 7.43-7.38(m, 2H), 7.36-7.25(m, 5H), 5.38(s, 2H), 4.59(d, J = 4.0Hz, 1H), 3.76-3.66(m, 2H), 3.42(dq, J = 10.2, 5.9, 5.4Hz, 4H), 1.50-1.41(m, 6H).

[0225] 2-(1-[4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazol-5-yl]-N-[2-(oxan-2-yloxy)ethyl]formamido)acetic acid ethyl ester To a stirred solution of 4-bromo-1-[(4-chlorophenyl)methyl]-N-[2-(oxan-2-yloxy)ethyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxamide (0.64 g, 1.03 mmol, 1 equiv.) in DMF (15 mL) was added NaH (0.1 g, 4.14 mmol, 4 equiv.) portionwise under a nitrogen atmosphere at 0°C. The mixture was stirred at 0°C for 30 min, and ethyl 2-bromoacetate (0.7 g, 4.14 mmol, 4 equiv.) was added. The mixture was stirred at room temperature for 16 h. The resulting mixture was quenched with water (100 ml) and extracted with EA (3 x 100 ml). The combined organic layers were washed with water (1 x 100 mL) and dried over anhydrous NaSO. After filtration, the filtrate was concentrated under reduced pressure to give the crude compound. The crude product was purified by reverse-phase flash chromatography under the following conditions (column: spherical C18 column, 20–40 μm, 120 g mobile phase A: water (0.1% AcOH), mobile phase B: ACN, flow rate: 60 mL / min, gradient: 60–80% B in 35 min, 254 nm) to give ethyl 2-(1-[4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazol-5-yl]-N-[2-(oxan-2-yloxy)ethyl]formamido)acetate (400 mg, 54.87%) as a yellow oil.

[0226] 2-(1-[4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazol-5-yl]-N-[2-(oxan-2-yloxy)ethyl]formamido)-N-methylacetamide To a 20 mL vessel, ethyl 2-(1-[4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazol-5-yl]-N-[2-(oxan-2-yloxy)ethyl]formamido)acetate (0.4 g, 0.57 mmol, 1 equiv.) and CH3NH2·MeOH (6 mL, 30%) were added at room temperature. The mixture was reacted under microwave irradiation at 60 °C for 1 hour. The resulting mixture was cooled to room temperature and concentrated in vacuo. The residue was purified by silica gel column chromatography eluting with PE / EA (1 / 1) to give 2-(1-[4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazol-5-yl]-N-[2-(oxan-2-yloxy)ethyl]formamide)-N-methylacetamide (0.28 g, 71.52%) as a yellow oil. 1 H NMR(400MHz, chloroform-d)δ7.42(d, J = 8.5Hz, 1H), 7.33(d, J = 8.6Hz, 3H), 7.14(dd, J = 31.1, 7.5Hz, 4H), 5.17(s, 2H), 4.41(s, 1H), 4.17(s, 2H), 3.75(d, J = 53.6Hz, 4H), 3.51(s, 2H), 2.82(s, 3H), 1.73(d, J = 36.0Hz, 6H).

[0227] 1-[(4-chlorophenyl)methyl]-4-methyl-7-[2-(oxan-2-yloxy)ethyl]-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-5,8-dione To a 20 mL vessel, 2-(1-[4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazol-5-yl]-N-[2-(oxan-2-yloxy)ethyl]formamide)-N-methylacetamide (280 mg, 0.41 mmol, 1 equiv.), Pd(dba) (55.9 mg, 0.06 mmol, 0.150 equiv.), CsCO (397.7 mg, 1.22 mmol, 3.008 equiv.), and tris(2-methylphenyl)phosphane (37.23 mg, 0.12 mmol, 0.301 equiv.) were added at room temperature. The mixture was heated at 120 °C under microwave conditions for 4 h. The mixture was cooled to room temperature. The residue was purified by silica gel column chromatography eluting with PE / EA (1 / 1) to give 1-[(4-chlorophenyl)methyl]-4-methyl-7-[2-(oxan-2-yloxy)ethyl]-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-5,8-dione (110 mg, 44.51%) as a yellow oil.

[0228] 1-[(4-chlorophenyl)methyl]-7-(2-hydroxyethyl)-4-methyl-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-5,8-dione (Compound 10) A mixture of 1-[(4-chlorophenyl)methyl]-4-methyl-7-[2-(oxan-2-yloxy)ethyl]-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-5,8-dione (120 mg, 200 mmol, 1 equiv.) and 2 M HCl (20 mL) in THF (20 mL) was stirred at room temperature for 1 h. The reaction was monitored by LCMS. The resulting mixture was extracted with EA (3 × 200 mL). The combined organic layers were washed with water (1 × 200 mL) and dried over anhydrous NaSO. After filtration, the filtrate was concentrated under reduced pressure. The crude product (88 mg) was purified by preparative HPLC under the following conditions (Column: XBridge preparative C18 OBD column, 5 μm, 19*150 mm; Mobile phase A: water (0.05% TFA), Mobile phase B: ACN; Flow rate: 20 mL / min; Gradient: 35% B to 60% B in 12 min; 254 nm; RT: 11.70 min) to give 1-[(4-chlorophenyl)methyl]-7-(2-hydroxyethyl)-4-methyl-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-5,8-dione (60 mg, 73.28%) as a yellow semi-solid. 1 H NMR(400MHz, methanol-d4)δ7.52(t, J = 8.3Hz, 1H), 7.34-7.27(m, 6H), 7.20(d, J = 8.3Hz, 1H), 5.51(s, 2H), 4.14(s, 2H), 3.74(t, J = 5.5Hz, 2H), 3.67(t, J = 5.6Hz, 2H), 3.33(s, 3H). Molecular formula C 23 H 20 Calculated [M+H] for ClF3N4O5 + :525, Observed:525.

[0229] The preparation of compounds 11-14 shown in the table below follows the methods and protocols described for the synthesis of compound 10, starting with the appropriate amine.

[0230] [Table 5]

[0231] Preparation of compounds 15 and 16:

[0232] [ka]

[0233] 1-[(4-chlorophenyl)methyl]-7-[(1R,3R)-3-hydroxycyclopentyl]-4-methyl-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-5,8-dione and 1-[(4-chlorophenyl)methyl]-7-[(1S,3S)-3-hydroxycyclopentyl]-4-methyl-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-5,8-dione The crude product 1-(4-chlorobenzyl)-7-((trans-3-hydroxycyclopentyl)-4-methyl-2-(3-(trifluoromethoxy)phenoxy)-1,4,6,7-tetrahydroimidazo[4,5-e][1,4]diazepine-5,8-dione was purified by chiral preparative HPLC under the following conditions (column: CHIRALPAK IC, 2*25 cm, 5 μm; mobile phase A: hexane 0.1% DEA--HPLC, mobile phase B: EtOH--HPLC; flow rate: 20 mL / min; gradient: 20B to 20B in 18 min; 220 / 254 nm; RT1: 12.901; RT2: 15.068) to give compound 15 (RT 12.901 min, 23.8 mg, 16.08%) and compound 16 (RT The yield was 15.068 min, 22.0 mg, 14.87%).

[0234] Characterization of compound 15: 1H NMR(400MHz, methanol-d4)δ7.52(t, J = 8.3Hz, 1H), 7.43-7.24(m, 6H), 7.20(d, J = 8.4Hz, 1H), 5.51(s, 2H), 5.17(p, J = 8.5Hz, 1H), 4.40(s, 1H), 3.93(s, 2H), 3.33(s, 3H), 2.10-2.09(m, 2H), 1.99-1.97(m, 2H), 1.68-1.59(m, 2H). Molecular formula C 26 H 24 Calculated [M+H] for ClF3N4O5 + :565, Observed:565.

[0235] Characterization of compound 16: 1 H NMR(400MHz, methanol-d4)δ7.52(t, J = 8.3Hz, 1H), 7.36-7.27(m, 6H), 7.20(d, J = 8.4Hz, 1H), 5.51(s, 2H), 5.18(p, J = 8.5Hz, 1H), 4.40(s, 1H), 3.93(s, 2H), 3.33(s, 3H), 2.10-2.08(m, 2H), 1.87-1.77(m, 2H), 1.68-1.59(m, 2H). Molecular formula C 26 H 24 Calculated [M+H] for ClF3N4O5 + :565, Observed:565.

[0236] Preparation of compounds 17 and 18:

[0237] [ka]

[0238] 2-Amino-4-[(tert-butyldimethylsilyl)oxy]butanoic acid To a stirred solution of 2-amino-4-hydroxybutanoic acid (5.0 g, 41.97 mmol, 1 equiv.) in MeCN (150 mL) and DBU (6.6 mL, 43.25 mmol, 1.05 equiv.) at 0 °C was slowly added TBSCl (6.6 g, 44.07 mmol, 1.05 equiv.). The resulting mixture was warmed to room temperature and stirred at room temperature for 16 h. The reaction mixture was filtered, and the filter cake was washed with MeCN (3 × 50 mL). The combined filtrates were concentrated to give 2-amino-4-[(tert-butyldimethylsilyl)oxy]butanoic acid (8.5 g, crude). 1 H NMR (300 MHz, methanol-d₄) δ 3.89 (t, J = 6.1 Hz, 2H), 3.79 (t, J = 5.9 Hz, 1H), 3.74–3.55 (m, 2H), 0.95 (s, 9H), 0.14 (s, 6H).

[0239] 2-Amino-4-[(tert-butyldimethylsilyl)oxy]butanoic acid methyl ester To a mixture of 2-amino-4-[(tert-butyldimethylsilyl)oxy]butanoic acid (5 g, 21.42 mmol, 1 equiv.) in toluene (200 mL) and MeOH (50 mL) was added trimethylsilyldiazomethane (60 mL, 0.53 mmol, 0.025 equiv.) at room temperature. The resulting mixture was stirred at room temperature for 16 hours. The reaction mixture was concentrated to give methyl 2-amino-4-[(tert-butyldimethylsilyl)oxy]butanoate (5 g, crude) as a pale yellow oil.

[0240] 2-([4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazol-5-yl]formamido)-4-[(tert-butyldimethylsilyl)oxy]butanoic acid methyl ester A mixture of 4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxylic acid (1.1 g, 2.24 mmol, 1 equiv.) and (COCl) (0.6 mL, 4.73 mmol, 3.148 equiv.) in DCM (20.0 mL) and DMF (5 drops) was stirred at room temperature for 1 hour. The resulting mixture was concentrated to give the crude product. To the above crude product, methyl 2-amino-4-[(tert-butyldimethylsilyl)oxy]butanoate (1.1 g, 4.45 mmol, 1.987 equiv.), TEA (1.6 mL, 15.37 mmol, 5 equiv.), and DCM (25.0 mL) were added, and the resulting mixture was then stirred at room temperature for 4 hours. The reaction mixture was purified by silica gel column chromatography eluting with PE:EA (20:1 to 5:1) to give methyl 2-([4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazol-5-yl]formamido)-4-[(tert-butyldimethylsilyl)oxy]butanoate (1.2 g, 74.38%) as a pale yellow oil. 1 H NMR(400MHz, DMSO-d6)δ8.51(d, J = 7.6Hz, 1H), 7.68-7.53(m, 1H), 7.44-7.23(m, 7H), 5.47-5.26(m, 2H), 4.55(ddd, J = 9.5, 7.5, 4.3Hz, 1H), 3.70(dd, J = 7.1, 4.7Hz, 2H), 3.34(s, 3H), 2.08-1.89(m, 2H), 0.85(s, 9H), 0.01(d, J = 5.2Hz, 7H).

[0241] 2-(1-[4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazol-5-yl]-N-methylformamido)-4-[(tert-butyldimethylsilyl)oxy]butanoic acid methyl ester To a mixture of methyl 2-([4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazol-5-yl]formamido)-4-[(tert-butyldimethylsilyl)oxy]butanoate (1.2 g, 1.66 mmol, 1 equiv.) in DMF (20 mL) was added NaH (100 mg, 2.50 mmol, 1.502 equiv., 60 wt%) at 0 °C under a nitrogen atmosphere for 0.5 h. To the above mixture was added CHI (0.2 mL, 3.21 mmol, 1.930 equiv.) at 0 °C. The resulting mixture was stirred at room temperature for an additional 16 h. Ethyl acetate (300 mL) and brine (300 mL) were added to the resulting mixture, and then the aqueous layer was extracted with ethyl acetate (200 mL). The combined organic layers were dried over anhydrous NaSO and filtered. The filtrate was purified by silica gel column chromatography eluting with PE:EA (20:1 to 6:1) to give methyl 2-(1-[4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazol-5-yl]-N-methylformamido)-4-[(tert-butyldimethylsilyl)oxy]butanoate (880 mg, 71.93%) as a pale yellow oil.

[0242] 2-(1-[4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazol-5-yl]-N-methylformamido)-4-[(tert-butyldimethylsilyl)oxy]-N-methylbutanamide A mixture of methyl 2-(1-[4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazol-5-yl]-N-methylformamido)-4-[(tert-butyldimethylsilyl)oxy]butanoate (1.1 g, 1.50 mmol, 1 equiv) in 2 M methylamine in methanol (5 mL) was irradiated with microwave radiation at 60° C. for 1 hour. The reaction mixture was purified by silica gel column chromatography eluting with PE:EA (1:1 to 1:8) to give 2-(1-[4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazol-5-yl]-N-methylformamido)-4-[(tert-butyldimethylsilyl)oxy]-N-methylbutanamide (550 mg, 50.07%) as a pale yellow oil. 1 H NMR(300MHz, DMSO-d6)δ7.70(s, 1H), 7.58(t, J = 8.2Hz, 1H), 7.46-7.21(m, 6H), 5.16(s, 2H), 4.94(s, 1H), 3.60(s, 3H), 2.88(s, 3H), 2.60(d, J = 4.5Hz, 3H), 2.03(d, J = 8.7Hz, 1H), 1.90(d, J = 12.3Hz, 1H), 0.86(s, 9H), 0.02(d, J = 2.7Hz, 6H).

[0243] 6-[2-[(tert-butyldimethylsilyl)oxy]ethyl]-1-[(4-chlorophenyl)methyl]-4,7-dimethyl-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-5,8-dione A mixture of 2-(1-[4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazol-5-yl]-N-methylformamido)-4-[(tert-butyldimethylsilyl)oxy]-N-methylbutanamide (450 mg, 610 mmol, 1 equiv.), Pd(dba) CHCl (63.45 mg, 0.06 mmol, 0.100 equiv.), P(o-Tol) (37.35 mg, 0.12 mmol, 0.200 equiv.), and CsCO (400 mg, 1.23 mmol, 2.003 equiv.) in toluene (8 mL) was irradiated with microwave radiation at 120 °C for 4 h. The mixture was cooled to room temperature. The reaction mixture was filtered, and the filtrate was concentrated to give a crude product, which was purified by silica gel column chromatography eluting with PE:EA (2:1 to 2:3) to give 6-[2-[(tert-butyldimethylsilyl)oxy]ethyl]-1-[(4-chlorophenyl)methyl]-4,7-dimethyl-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-5,8-dione (220 mg, 54.95%) as a pale yellow oil.

[0244] (6S)-1-[(4-chlorophenyl)methyl]-6-(2-hydroxyethyl)-4,7-dimethyl-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-5,8-dione and (6R)-1-[(4-chlorophenyl)methyl]-6-(2-hydroxyethyl)-4,7-dimethyl-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-5,8-dione To a stirred solution of 6-[2-[(tert-butyldimethylsilyl)oxy]ethyl]-1-[(4-chlorophenyl)methyl]-4,7-dimethyl-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-5,8-dione (200 mg, 310 mmol, 1 equiv.) in THF (10 mL) was added 6 M HCl (10 mL) dropwise at room temperature. The resulting mixture was stirred at room temperature for 2 h. The reaction mixture was basified to pH 10 with KCO and extracted with ethyl acetate (3 × 100 mL). The organic layer was then washed with brine (2 × 50 mL), dried over anhydrous NaSO, and filtered. The filtrate was concentrated to give the crude product, which was purified by preparative chiral HPLC (column: CHIRALPAK IG, 20*250 mm, 5 μm; mobile phase A: hexane 0.1% DEA-HPLC, mobile phase B: EtOH-HPLC; flow rate: 20 mL / min; gradient: 20B to 20B in 18 min; 220 / 254 nm; RT1: 10.297; RT2: 13.612) to give compound 17 (RT 10.297 min, 20.2 mg, 12.24%) and compound 18 (RT 13.612 min, 18.2 mg, 11.03%) as pale yellow oils.

[0245] Characterization of compound 17: 1 H NMR(300MHz, DMSO-d6)δ7.63-7.50(m, 1H), 7.46-7.24(m, 7H), 5.65-5.32(m, 2H), 4.72-4.59(m, 1H), 4.41-4.30(m, 1H), 3.55-3.43(m, 0.8H), 3.42-3.36(m, 0.7H), 3.23(s, 3H), 3.08-3.01(m, 1H), 3.00-2.90(m, 0.3H), 2.84(s, 2.2H), 2.25-2.14(m, 0.7H), 2.07-1.93(m, 0.7H), 1.61-1.49(m, 0.7H). Molecular formula C 24 H 22 Calculated [M+H] for ClF3N4O5 + :539, Observed:539.

[0246] Characterization of compound 18:1 H NMR(300MHz, DMSO-d6)δ7.63-7.53(m, 1H), 7.47-7.23(m, 7H), 5.64-5.32(m, 2H), 4.72-4.59(m, 1H), 4.39-4.27(m, 1H), 3.55-3.42(m, 0.8H), 3.42-3.36(m, 0.8H), 3.23(s, 3H), 3.08-3.01(m, 1H), 3.00-2.89(m, 0.5H), 2.84(s, 2.2H), 2.27-2.13(m, 0.7H), 2.08-1.94(m, 0.7H), 1.62-1.49(m, 0.5H). Molecular formula C 24 H 22 Calculated [M+H] for ClF3N4O5 + :539, Observed:539.

[0247] Preparation of compounds 19 and 20:

[0248] [ka]

[0249] 4-Bromo-1-[(4-chlorophenyl)methyl]-2-(3-fluorophenoxy)-N-(3-hydroxypropyl)-1H-imidazole-5-carboxamide To a mixture of 4-bromo-1-[(4-chlorophenyl)methyl]-2-(3-fluorophenoxy)-1H-imidazole-5-carboxylic acid (2 g, 4.70 mmol, 1 eq.) and 1 drop of DMF in DCM (30 mL) was added oxalyl dichloride (1.8 g, 14.10 mmol, 3 eq.) dropwise at 0° C. This mixture was stirred at room temperature for 3 hours. The resulting mixture was concentrated under vacuum. The solid was dissolved in DCM (30 mL) to give solution A. A mixture of 3-aminopropan-1-ol (1.8 g, 23.49 mmol, 5 eq.) and triethylamine (2.4 g, 23.49 mol, 5 eq.) in DCM (5 mL) was cooled to 0° C., and solution A was added dropwise at 0° C. under a nitrogen atmosphere. The mixture was stirred at room temperature for 3 hours. The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography eluting with PE / EA (2 / 1 to 1 / 1) to give 4-bromo-1-[(4-chlorophenyl)methyl]-2-(3-fluorophenoxy)-N-(3-hydroxypropyl)-1H-imidazole-5-carboxamide (1.65 g, 72.74%) as a white solid.

[0250] 4-Bromo-1-[(4-chlorophenyl)methyl]-2-(3-fluorophenoxy)-N-[3-(oxan-2-yloxy)propyl]-1H-imidazole-5-carboxamide To a stirred mixture of 4-bromo-1-[(4-chlorophenyl)methyl]-2-(3-fluorophenoxy)-N-(3-hydroxypropyl)-1H-imidazole-5-carboxamide (1.65 g, 3.42 mmol, 1 equiv.) and 3,4-dihydro-2H-pyran (1.4427 g, 17.15 mmol, 5.018 equiv.) in DCM (50 mL) was added 4-methylbenzene-1-sulfonic acid (0.059 g, 0.34 mmol, 0.100 equiv.) at room temperature. The mixture was stirred at room temperature for 16 hours. The resulting mixture was washed with saturated aqueous NaHCO3. The DCM layer was concentrated under reduced pressure. The residue was purified by silica gel column chromatography eluting with PE / EA (6 / 1 to 3 / 1) to give 4-bromo-1-[(4-chlorophenyl)methyl]-2-(3-fluorophenoxy)-N-[3-(oxan-2-yloxy)propyl]-1H-imidazole-5-carboxamide (1.65 g, 85.16%) as a white solid.

[0251] Ethyl 2-(1-[4-bromo-1-[(4-chlorophenyl)methyl]-2-(3-fluorophenoxy)-1H-imidazol-5-yl]-N-[3-(oxan-2-yloxy)propyl]formamido)propanoate To a stirred solution of 4-bromo-1-[(4-chlorophenyl)methyl]-2-(3-fluorophenoxy)-N-[3-(oxan-2-yloxy)propyl]-1H-imidazole-5-carboxamide (2.5 g, 4.41 mmol, 1 equiv) in DMF (15 mL) was added NaH (0.7 g, 17.64 mmol, 4 equiv, 60%) portionwise under a nitrogen atmosphere at 0 °C. The mixture was stirred at 0 °C for 30 min, and ethyl 2-bromopropanoate (3.1936 g, 17.64 mmol, 4.000 equiv) was added. The mixture was stirred at room temperature for 16 h. The resulting mixture was quenched with water (100 mL) and extracted with EA (3 × 100 mL). The combined organic layers were washed with water (1 × 100 mL) and dried over anhydrous NaSO. After filtration, the filtrate was concentrated under reduced pressure to give the crude compound. The crude product was purified by reverse-phase flash chromatography using the following conditions: column: spherical C18 column, 20-40 μm, 120 g, mobile phase A: water (0.1% AcOH), mobile phase B: ACN, flow rate: 60 mL / min, gradient: 60-80% B in 35 min, 254 nm) to give ethyl 2-(1-[4-bromo-1-[(4-chlorophenyl)methyl]-2-(3-fluorophenoxy)-1H-imidazol-5-yl]-N-[3-(oxan-2-yloxy)propyl]formamido)propanoate (1.38 g, 46.91%) as a yellow oil.

[0252] 2-(1-[4-bromo-1-[(4-chlorophenyl)methyl]-2-(3-fluorophenoxy)-1H-imidazol-5-yl]-N-[3-(oxan-2-yloxy)propyl]formamido)-N-methylpropanamide To a 20 mL vessel, ethyl 2-(1-[4-bromo-1-[(4-chlorophenyl)methyl]-2-(3-fluorophenoxy)-1H-imidazol-5-yl]-N-[3-(oxan-2-yloxy)propyl]formamido)propanoate (1.3 g, 1 equivalent) and methylamine in methanol (6 mL, 30%) were added at room temperature. The mixture was reacted under microwave conditions at 60° C. for 1 hour. The resulting mixture was cooled to room temperature and concentrated in vacuo. The residue was purified by silica gel column chromatography eluting with PE / EA (1 / 1) to give 2-(1-[4-bromo-1-[(4-chlorophenyl)methyl]-2-(3-fluorophenoxy)-1H-imidazol-5-yl]-N-[3-(oxan-2-yloxy)propyl]formamide)-N-methylpropanamide (1.2 g, 94.43%) as a yellow oil.

[0253] 1-[(4-chlorophenyl)methyl]-2-(3-fluorophenoxy)-4,6-dimethyl-7-[3-[(2R)-oxan-2-yloxy]propyl]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-5,8-dione To a 20 mL vessel, 2-(1-[4-bromo-1-[(4-chlorophenyl)methyl]-2-(3-fluorophenoxy)-1H-imidazol-5-yl]-N-[3-(oxan-2-yloxy)propyl]formamido)-N-methylpropanamide (1.1936 g, 1.83 mol, 1 equiv.), Pd(dba) (0.2 g, 0.18 mmol, 0.1 equiv.), CsCO (1.1966 g, 3.67 mmol, 2.006 equiv.), and tris(2-methylphenyl)phosphane (0.1 g, 0.37 mmol, 0.2 equiv.) were added at room temperature. The mixture was heated at 120 °C under microwave conditions for 5 h. The mixture was cooled to room temperature. The residue was purified by silica gel column chromatography eluting with PE / EA (1 / 1) to give 1-[(4-chlorophenyl)methyl]-2-(3-fluorophenoxy)-4,6-dimethyl-7-[3-[(2R)-oxan-2-yloxy]propyl]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-5,8-dione (340 mg, 32.52%) as a yellow oil.

[0254] (6S)-1-[(4-chlorophenyl)methyl]-2-(3-fluorophenoxy)-7-(3-hydroxypropyl)-4,6-dimethyl-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-5,8-dione and (6R)-1-[(4-chlorophenyl)methyl]-2-(3-fluorophenoxy)-7-(3-hydroxypropyl)-4,6-dimethyl-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-5,8-dione A mixture of 1-[(4-chlorophenyl)methyl]-2-(3-fluorophenoxy)-4,6-dimethyl-7-[3-[(2R)-oxan-2-yloxy]propyl]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-5,8-dione (200 mg, 0.35 mmol, 1 equiv.) and HCl (2 M; 20 mL) in THF (20 mL) was stirred at room temperature for 1 h. The resulting mixture was extracted with EA (3 × 200 mL). The combined organic layers were washed with water (1 × 200 mL) and dried over anhydrous NaSO. After filtration, the filtrate was concentrated under reduced pressure. The crude product (170 mg) was purified by preparative HPLC under the following conditions (Column: XBridge preparative C18 OBD column, 5 μm, 19*150 mm; Mobile phase A: water (0.05% TFA), Mobile phase B: ACN; Flow rate: 20 mL / min; Gradient: 40% B to 70% B in 7 min; 254 nm; RT: 6.32 min) to give 1-[(4-chlorophenyl)methyl]-2-(3-fluorophenoxy)-7-(3-hydroxypropyl)-4,6-dimethyl-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-5,8-dione (140 mg). The racemate was separated by chiral HPLC under the following conditions (column: CHIRALPAK IF, 2*25 cm, 5 μm; mobile phase A: hexane 0.1% DEA--HPLC, mobile phase B: EtOH--HPLC; flow rate: 15 mL / min; gradient: 50B to 50B in 13 min; 220 / 254 nm; RT1: 8.349; RT2: 9.504) to give compound 19 (RT 8.349 min, 20 mg, 11.73%) and compound 20 (RT 9.504 min, 20 mg, 11.73%).

[0255] Characterization of compound 19: 1H NMR(400MHz, methanol-d4)δ7.45(dd, J = 13.8, 6.2Hz, 1H), 7.41-7.29(m, 4H), 7.23-6.99(m, 3H), 5.63-5.53(m, 1H), 5.45-5.38(m, 1H), 4.38(dd, J = 16.4, 8.3Hz, 1H), 3.96(dt, J = 14.8, 7.7Hz, 1H), 3.64(t, J = 7.0Hz, 1H), 3.49(t, J = 6.3Hz, 2H), 3.35(s, 3H), 1.85-1.73(m, 1H), 1.69-1.62(dt, J = 13.5, 6.9Hz, 1H), 1.56(d, J = 7.0Hz, 2H), 1.10(d, J = 7.5Hz, 1H). Molecular formula C 24 H 24 Calculated [M+H] for ClFN4O4 + :487, Observed:487.

[0256] Characterization of compound 20: 1 H NMR(400MHz, methanol-d4)δ7.49-7.40(m, 1H), 7.37-7.29(m, 4H), 7.22-7.00(m, 3H), 5.62-5.53(m, 1H), 5.42-5.39(m, 1H), 4.43-4.33(m, 1H), 3.99-3.92(dt, J = 14.9, 7.7Hz, 1H), 3.64(t, J = 8.0Hz, 1H), 3.49(t, J = 8.0Hz, 2H), 3.35(s, 3H), 1.82-1.75(m, 1H), 1.69-1.62(dt, J = 13.7, 6.9Hz, 1H), 1.56(d, J = 8.0Hz, 2H), 1.10(d, J = 8.0Hz, 1H). Molecular formula C 24 H 24 Calculated [M+H] for ClFN4O4 + :487, Observed:487.

[0257] Preparation of Compound 21:

[0258] [ka]

[0259] 1-[(4-chlorophenyl)methyl]-4-[(2-methoxy-2-oxoethyl)(methyl)amino]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxylate methyl ester A mixture of methyl 4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxylate (1.00 g, 1.98 mol, 1 equiv.), methyl 2-(methylamino)acetate (1019.6 mg, 9.89 mol, 5.0 equiv.), Pd(dba) (452.7 mg, 0.49 mmol, 0.25 equiv.), Xantphos (572.1 mg, 0.99 mmol, 0.50 equiv.), and CsCO (6.4 g, 19.78 mmol, 10.0 equiv.) in dioxane (100 mL) was stirred under a nitrogen atmosphere at 100 °C for 16 h. The reaction mixture was cooled to room temperature, and ethyl acetate (200 mL) and HO (200 mL) were added. The organic layer was washed with brine (3 × 50 mL), dried over anhydrous NaSO, and filtered. The filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography eluting with PE:EtOAc (6:1 to 4:1) to give methyl 1-[(4-chlorophenyl)methyl]-4-[(2-methoxy-2-oxoethyl)(methyl)amino]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxylate (150 mg, 14.37%) as a pale yellow oil.

[0260] 4-[(carbamoylmethyl)(methyl)amino]-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxylate A mixture of methyl 1-[(4-chlorophenyl)methyl]-4-[(2-methoxy-2-oxoethyl)(methyl)amino]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxylate (300 mg, 570 mmol, 1 equiv.) in ammonia solution (10 mL) was irradiated with microwave radiation for 4 hours at 120° C. The reaction mixture was concentrated under reduced pressure. The residue was purified by reverse-phase flash chromatography under the following conditions (Column: spherical C18 column, 20–40 μm, 120 g; Mobile phase A: water (0.1% HOAc), Mobile phase B: ACN; Flow rate: 60 mL / min; Gradient: 60% B to 70% B in 10 min, 254 nm) to give methyl 4-[(carbamoylmethyl)(methyl)amino]-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxylate (230 mg, 39.46%) as an off-white solid.

[0261] 1-[(4-chlorophenyl)methyl]-4-methyl-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-6,8-dione A mixture of methyl 4-[(carbamoylmethyl)(methyl)amino]-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxylate (120 mg, 0.23 mmol, 1 equiv.) in dioxane (20 mL) and NaH (18.7 mg, 0.47 mmol, 2.0 equiv., 60 wt%) was refluxed for 15 min. HOAc (0.5 mL) was added to the reaction mixture, and the resulting mixture was stirred at room temperature for 16 h. The reaction mixture was concentrated under reduced pressure. The residue was purified by reverse-phase flash chromatography under the following conditions (Column: spherical C18 column, 20–40 μm, 120 g; Mobile phase A: water (0.1% HOAc), Mobile phase B: ACN; Flow rate: 60 mL / min; Gradient: 60% B to 70% B in 15 min, 254 nm) to give 1-[(4-chlorophenyl)methyl]-4-methyl-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-6,8-dione (45 mg, 40.00%) as an off-white solid.

[0262] 1-[(4-chlorophenyl)methyl]-4-methyl-7-[3-(oxan-2-yloxy)propyl]-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-6,8-dione A mixture of 1-[(4-chlorophenyl)methyl]-4-methyl-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-6,8-dione (100 mg, 0.21 mmol, 1 equiv.), 2-(3-bromopropoxy)oxane (139.2 mg, 0.62 mmol, 3.0 equiv.), and K2CO3 (86.2 mg, 0.62 mmol, 3.0 equiv.) in DMF (15.0 mL) was stirred at 50 °C for 16 h. The reaction mixture was cooled to room temperature, and HO (100 mL) was added. The resulting mixture was extracted with EtOAc (3 × 50 mL). The combined organic layers were washed with brine (2 × 50 mL) and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluting with PE:EA (6:1 to 3:1) to give 1-[(4-chlorophenyl)methyl]-4-methyl-7-[3-(oxan-2-yloxy)propyl]-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-6,8-dione (120 mg, 92.6:1%) as a pale yellow oil. 1 H NMR(400MHz, chloroform-d)δ7.43(t, J = 8.3Hz, 1H), 7.27(d, J = 8.7Hz, 5H), 7.17(dd, J = 33.5, 8.4Hz, 2H), 5.58(s, 2H), 4.60(d, J = 14.8Hz, 2H), 3.88(s, 2H), 3.85-3.72(m, 2H), 3.63(dt, J = 10.4, 5.6Hz, 1H), 3.58-3.47(m, 2H), 3.13(s, 3H), 2.04-1.58(m, 8H).

[0263] 1-[(4-chlorophenyl)methyl]-7-(3-hydroxypropyl)-4-methyl-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]]diazepine-6,8-dione A mixture of 1-[(4-chlorophenyl)methyl]-4-methyl-7-[3-(oxan-2-yloxy)propyl]-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-6,8-dione (45 mg, 0.07 mmol, 1 equiv.) in THF (10 mL) and 6 M HCl (10 mL) was stirred at room temperature for 2 h. The reaction mixture was concentrated under reduced pressure. The residue was basified to pH 9 with saturated KCO(aq). The resulting mixture was extracted with ethyl acetate (3 x 50 mL), and the combined organic layers were washed with brine (50 mL) and concentrated to give the crude product, which was purified by preparative HPLC under the following conditions (Column: XSelect CSH Preparative C18 OBD Column, 5 μm, 19*150 mm; Mobile phase A: water (0.1% FA), Mobile phase B: ACN; Flow rate: 20 mL / min; Gradient: 50% B to 87% B in 7 min; 254 nm; RT: 6.58 min) to give 1-[(4-chlorophenyl)methyl]-7-(3-hydroxypropyl)-4-methyl-2-[3-(trifluoromethoxy)phenoxy]-1H,4H,5H,6H,7H,8H-imidazo[4,5-e][1,4]diazepine-6,8-dione (3.9 mg, 9.52%) as an off-white solid. 1 H NMR(400MHz, chloroform-d)δ7.54-7.45(m, 1H), 7.43-7.28(m, 2H), 7.24-7.02(m, 5H), 5.57(s, 2H), 4.04(t, J = 6.0Hz, 2H), 3.90(s, 2H), 3.50(t, J = 5.5Hz, 2H), 3.15(s, 3H), 1.80(p, J = 5.7Hz, 2H). Molecular formula C 24 H 22 Calculated [M+H] for ClF3N4O5 + :539, Observed:539.

[0264] Preparation of compounds 22 and 23:

[0265] [ka]

[0266] 1-[(4-chlorophenyl)methyl]-4-[2-(methoxycarbonyl)pyrrolidin-1-yl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxylate methyl ester A mixture of methyl 4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxylate (1.5 g, 2.97 mmol, 1 equiv.), Pd(dba) (280 mg, 0.31 mmol, 0.103 equiv.), Xantphos (520 mg, 0.90 mmol, 0.303 equiv.), methyl pyrrolidine-2-carboxylate (800.0 mg, 6.19 mmol, 2.088 equiv.), and CsCO (4.9 g, 15.04 mmol, 5.070 equiv.) in dioxane (40 mL) was stirred at 100 °C for 14 h. The reaction mixture was filtered, and the filtrate was concentrated to give the crude product, which was purified by silica gel column chromatography eluting with PE:EA (20:1 to 4:1) to give methyl 1-[(4-chlorophenyl)methyl]-4-[2-(methoxycarbonyl)pyrrolidin-1-yl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxylate (830 mg, 50.51%) as a pale yellow oil.

[0267] 4-(2-carbamoylpyrrolidin-1-yl)-1-(4-chlorobenzyl)-2-(3-(trifluoromethoxy)phenoxy)-1H-imidazole-5-carboxylate methyl ester A mixture of methyl 1-[(4-chlorophenyl)methyl]-4-[2-(methoxycarbonyl)pyrrolidin-1-yl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxylate (1.65 g, 2.98 mmol, 1 equiv) in ammonia solution (7.0 M in methanol) (10 mL) was irradiated with microwave radiation for 10 hours at 140° C. The reaction mixture was cooled to room temperature and concentrated to give a residue which was purified by flash chromatography on a silica gel column eluted with 70% ethyl acetate in petroleum ether to give methyl 4-(2-carbamoylpyrrolidin-1-yl)-1-(4-chlorobenzyl)-2-(3-(trifluoromethoxy)phenoxy)-1H-imidazole-5-carboxylate (300 mg, 24% yield) as a pale yellow semi-solid. 1 H NMR(400MHz, chloroform-d)δ7.41(t, J = 8.3Hz, 1H), 7.34-7.24(m, 4H), 7.23-7.18(m, 3H), 7.12-7.07(m, 1H), 6.52(s, 1H), 5.41(q, J = 15.4Hz, 2H), 4.53(dd, J = 8.2, 5.2Hz, 1H), 3.87-3.69(m, 4H), 3.32(dt, J = 10.5, 7.1Hz, 1H), 2.28-2.09(m, 2H), 2.05-1.83(m, 2H).

[0268] 5-[(4-chlorophenyl)methyl]-4-[3-(trifluoromethoxy)phenoxy]-1,3,5,8-tetraazatricyclo[8.3.0.0^[2,6]]trideca-2(6),3-diene-7,9-dione To a mixture of methyl 4-(2-carbamoylpyrrolidin-1-yl)-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxylate (185 mg, 340 mmol, 1 equiv.) in dioxane (40 mL) was added NaH (96.1 mg, 2.40 mmol, 7.0 equiv., 60 wt%) at 0° C., and the reaction mixture was then refluxed for 15 minutes. The reaction mixture was cooled to room temperature and concentrated under reduced pressure. The residual product was purified by reverse-phase flash chromatography (C18 spherical column, 20-40 μm, 120 g; mobile phase A: water (0.1% HOAc), mobile phase B: ACN; flow rate: 60 mL / min; gradient: 65% B to 85% B in 15 min at 254 nm) to give 5-[(4-chlorophenyl)methyl]-4-[3-(trifluoromethoxy)phenoxy]-1,3,5,8-tetraazatricyclo[8.3.0.0^[2,6]]trideca-2(6),3-diene-7,9-dione (115 mg, 66.09%) as an off-white solid. 1 H NMR(400MHz, DMSO-d6)δ10.52(s, 1H), 7.58(t, J = 8.3Hz, 1H), 7.47(d, J = 2.7Hz, 1H), 7.44-7.36(m, 3H), 7.36-7.27(m, 3H), 5.52(dd, J = 91.6, 15.5Hz, 2H), 3.81(dd, J = 8.3, 5.2Hz, 1H), 3.51(dt, J = 10.1, 7.3Hz, 1H), 3.32(d, J = 7.1Hz, 1H), 2.50(s, 1H), 2.12-1.96(m, 1H), 1.81(qt, J = 12.3, 6.6Hz, 2H).

[0269] 5-[(4-chlorophenyl)methyl]-8-[3-(oxan-2-yloxy)propyl]-4-[3-(trifluoromethoxy)phenoxy]-1,3,5,8-tetraazatricyclo[8.3.0.0^[2,6]]trideca-2(6),3-diene-7,9-dione A mixture of 5-[(4-chlorophenyl)methyl]-4-[3-(trifluoromethoxy)phenoxy]-1,3,5,8-tetraazatricyclo[8.3.0.0^[2,6]]trideca-2(6),3-diene-7,9-dione (150 mg, 300 mmol, 1 equiv), 2-(3-bromopropoxy)oxane (198.1 mg, 890 mmol, 3.000 equiv), and KCO (122.7 mg, 0.89 mmol, 3.0 equiv) in DMF (15.0 mL) was stirred at 50° C. for 16 h. The reaction was cooled to room temperature, and EtOAc (100 mL) and HO (100 mL) were added. The organic layer was washed with brine (2 × 30 mL), concentrated under reduced pressure, and purified by reverse-phase flash chromatography (120 g, 20–40 μm spherical C18 column; mobile phase A: water (0.1% HOAc), mobile phase B: ACN; flow rate: 60 mL / min; gradient: 90% B to 98% B in 10 min at 254 nm) to give 5-[(4-chlorophenyl)methyl]-8-[3-(oxan-2-yloxy)propyl]-4-[3-(trifluoromethoxy)phenoxy]-1,3,5,8-tetraazatricyclo[8.3.0.0^[2,6]]trideca-2(6),3-diene-7,9-dione (185 mg, 96.31%) as a pale yellow oil. 1 H NMR(400MHz, chloroform-d)δ7.43(t, J = 8.3Hz, 1H), 7.30(s, 4H), 7.27-7.19(m, 2H), 7.13(d, J = 8.3Hz, 1H), 5.73(d, J = 14.9Hz, 1H), 5.43(d, J = 15.1Hz, 1H), 4.65-4.49(m, 1H), 4.15(ddt, J = 19.0, 13.7, 7.1Hz, 1H), 3.98-3.61(m, 4H), 3.46(tt, J = 16.5, 7.6Hz, 4H), 2.85(dt, J = 12.2, 6.3Hz, 1H), 2.18-2.02(m, 2H), 1.99-1.77(m, 3H), 1.76-1.48(m, 6H).

[0270] (10R)-5-[(4-chlorophenyl)methyl]-8-(3-hydroxypropyl)-4-[3-(trifluoromethoxy)phenoxy]-1,3,5,8-tetraazatricyclo[8.3.0.0^[2,6]]trideca-2(6),3-diene-7,9-dione and (10S)-5-[(4-chlorophenyl)methyl]-8-(3-hydroxypropyl)-4-[3-(trifluoromethoxy)phenoxy]-1,3,5,8-tetraazatricyclo[8.3.0.0^[2,6]]trideca-2(6),3-diene-7,9-dione A mixture of 5-[(4-chlorophenyl)methyl]-8-[3-(oxan-2-yloxy)propyl]-4-[3-(trifluoromethoxy)phenoxy]-1,3,5,8-tetraazatricyclo[8.3.0.0^[2,6]]trideca-2(6),3-diene-7,9-dione (160 mg, 250 mmol, 1 equiv.) in THF (5 mL) and 2 M HCl (5 mL) was stirred at room temperature for 2 h. The pH of the reaction mixture was adjusted to 9 with saturated NaHCO3 (aq.), EtOAc (100 mL) was added, and the organic layer was then washed with brine (2 × 50 mL) and concentrated under reduced pressure. The residue was purified by preparative HPLC under the following conditions (Column: XBridge Shield RP18 OBD column 30*150 mm, 5 μm, Mobile phase A: water (0.1% FA), Mobile phase B: ACN; Flow rate: 60 mL / min; Gradient: 50% B to 80% B in 7 min; 254 nm; RT: 6.45 min) to give racemic 5-[(4-chlorophenyl)methyl]-8-(3-hydroxypropyl)-4-[3-(trifluoromethoxy)phenoxy]-1,3,5,8-tetraazatricyclo[8.3.0.0^[2,6]]trideca-2(6),3-diene-7,9-dione (110 mg, 78.99%) as a colorless oil. 90 mg of this material was subjected to chiral preparative HPLC (column: CHIRALPAK AD-H, 2.0 cm ID*25 cm L; mobile phase A: hexane 0.1% DEA-HPLC, mobile phase B: EtOH-HPLC; flow rate: 20 mL / min; gradient: 15B to 15B in 15 min; 220 / 254 nm; RT1: 9.279; RT2: 10.54) to give compound 22 (RT 9.279 min, 17.7 mg, 19.67%) and compound 23 (RT 10.54 min, 19.11%).

[0271] Characterization of compound 22: 1H NMR(400MHz, methanol-d4)δ7.55-7.51(m, 1H), 7.36-7.27(m, 6H), 7.23-7.20(m, 1H), 5.70(d, J = 16.0Hz, 1H), 5.52(d, J = 16.0Hz, 1H), 4.13-4.05(m, 1H), 3.90-3.83(m, 1H), 3.75-3.73(m, 1H), 3.69-3.62(m, 1H), 3.58-3.51(m, 2H), 3.45-3.41(m, 1H), 2.80-2.72(m, 1H), 2.19-2.10(m, 1H), 2.05-1.92(m, 2H), 1.91-1.68(m, 2H). Molecular formula C 26 H 24 Calculated [M+H] for ClF3N4O5 + :565, Observed:565.

[0272] Characterization of compound 23: 1 H NMR(400MHz, methanol-d4)δ7.55-7.51(m, 1H), 7.35-7.28(m, 6H), 7.23-7.21(m, 1H), 5.70(d, J = 16.0Hz, 1H), 5.52(d, J = 16.0Hz, 1H), 4.13-4.06(m, 1H), 3.90-3.82(m, 1H), 3.77-3.73(m, 1H), 3.69-3.62(m, 1H), 3.55-3.51(m, 2H), 3.46-3.41(m, 1H), 2.79-2.70(m, 1H), 2.19-2.10(m, 1H), 2.04-1.94(m, 2H), 1.79-1.70(m, 2H). Molecular formula C 26 H 24 Calculated [M+H] for ClF3N4O5 + :565, Observed:565.

[0273] Preparation of compounds 24 and 25: The preparation of the racemates of compounds 24 and 25 follows the methods and protocols described for the synthesis of compounds 22 and 23, starting from intermediate I.

[0274] [ka]

[0275] (10R)-5-Butyl-4-(3-fluorophenoxy)-8-(3-hydroxypropyl)-1,3,5,8-tetraazatricyclo[8.3.0.0^[2,6]]trideca-2(6),3-diene-7,9-dione and (10S)-5-Butyl-4-(3-fluorophenoxy)-8-(3-hydroxypropyl)-1,3,5,8-tetraazatricyclo[8.3.0.0^[2,6]]trideca-2(6),3-diene-7,9-dione The crude product, 5-butyl-4-(3-fluorophenoxy)-8-(3-hydroxypropyl)-1,3,5,8-tetraazatricyclo[8.3.0.0^[2,6]]trideca-2(6),3-diene-7,9-dione, was purified by chiral HPLC (column: XBridge Shield RP18 OBD column, 5 μm, 19 × 150 mm; mobile phase A: water (10 mmol / L NH4HCO3), mobile phase B: ACN; flow rate: 20 mL / min; gradient: 45% B to 70% B in 7 min; 220 nm; RT1: 15.00 min, RT2: 17.33 min) to give compound 24 (RT = 15.00 min, 20.3 mg, 8.09%) and compound 25 (RT = 17.33 min, 18.9 mg, 7.53%).

[0276] Characterization of compound 24: 1H NMR(400MHz, メタノール-d4)δ7.51-7.40(m, 1H), 7.19-7.08(m, 2H), 7.09-6.99(m, 1H), 4.41(dt, J = 13.5, 6.8Hz, 1H), 4.26(dt, J = 13.7, 7.4Hz, 1H), 4.13(ddd, J = 13.4, 7.7, 5.8Hz, 1H), 3.88(dt, J = 13.6, 7.3Hz, 1H), 3.75(dd, J = 8.2, 4.7Hz, 1H), 3.70-3.50(m, 3H), 3.42(dd, J = 10.7, 5.8Hz, 1H), 2.77(dq, J = 12.2, 6.0Hz, 1H), 2.16(dq, J = 12.7, 7.8Hz, 1H), 1.98(h, J = 6.1Hz, 2H), 1.79(dq, J = 14.5, 7.1Hz, 4H), 1.38(p, J = 7.4Hz, 2H), 0.98(t, J = 7.4Hz, 3H). Molecular formula C 22 H 27 Calculation of FN4O4[M+H] + :431, View value: 431.

[0277] Evaluation of properties of compound 25: 1H NMR(400MHz, methanol-d4)δ7.49-7.38(m, 1H), 7.19-7.10(m, 2H), 7.03(d, J = 7.8Hz, 1H), 4.41(dt, J = 13.6, 6.8Hz, 1H), 4.26(dt, J = 13.9, 7.4Hz, 1H), 4.17-4.08(m, 1H), 3.88(dt, J = 13.5, 7.2Hz, 1H), 3.75(dd, J = 8.2, 4.7Hz, 1H), 3.70-3.52(m, 3H), 3.43(dt, J = 11.0, 5.8Hz, 1H), 2.77(dq, J = 12.3, 6.1Hz, 1H), 2.16(dq, J = 12.6, 7.8Hz, 1H), 1.99(hept, J = 6.1Hz, 2H), 1.80(dp, J = 14.6, 7.1Hz, 4H), 1.35(dd, J = 17.2, 9.9Hz, 2H), 0.98(t, J = 7.4Hz, 3H). Molecular formula C 22 H 27 Calculated [M+H] of FN4O4 + :431, Observed:431.

[0278] The following compounds were prepared by a procedure similar to the preparation of compounds 22 and 23.

[0279] [Table 6-1]

[0280] [Table 6-2]

[0281] [Table 6-3]

[0282] [Table 6-4]

[0283] Preparation of compounds 34 and 35:

[0284] [ka]

[0285] Methyl 4-[2-([[(tert-butoxy)carbonyl]amino]methyl)pyrrolidin-1-yl]-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxylate A mixture of methyl 4-bromo-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxylate (Intermediate A, 6 g, 11.87 mmol, 1 equiv.), tert-butyl N-[(pyrrolidin-2-yl)methyl]carbamate (4.8 g, 23.73 mmol, 2.00 equiv.), Xantphos (2.1 g, 3.56 mmol, 0.3 equiv.), Pd(dba) (1.1 g, 1.19 mmol, 0.1 equiv.), and CsCO (19.3 g, 59.33 mmol, 5 equiv.) in dioxane (100 mL) was stirred at 100 °C for 16 h. The reaction mixture was filtered, and the filtrate was concentrated to give a crude product, which was purified by silica gel column chromatography eluting with PE:EA (20:1 to 1:1) to give methyl 4-[2-([[(tert-butoxy)carbonyl]amino]methyl)pyrrolidin-1-yl]-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxylate (3.4 g, 45.84%) as a pale yellow oil. 1H NMR(400MHz, chloroform-d)δ7.42(t, J = 8.2Hz, 1H), 7.31(d, J = 8.2Hz, 2H), 7.23(q, J = 9.0, 8.4Hz, 4H), 7.10(d, J = 8.3Hz, 1H), 5.45(d, J = 15.8Hz, 1H), 5.33(d, J = 15.4Hz, 1H), 5.02(s, 1H), 4.16(dt, J = 23.2, 6.8Hz, 1H), 3.76(s, 4H), 3.30(s, 2H), 3.11(s, 1H), 1.97(d, J = 41.7Hz, 2H), 1.75(d, J = 6.8Hz, 2H), 1.43(s, 9H).

[0286] Methyl 4-[2-(aminomethyl)pyrrolidin-1-yl]-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxylate A stirred solution of methyl 4-[2-([[((tert-butoxy)carbonyl]amino]methyl)pyrrolidin-1-yl]-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxylate (3.4 g, 5.44 mmol, 1 equiv.) in HCl (4 M) (30 mL, 987.36 mmol, 181.51 equiv.) was stirred at room temperature for 2 h. The reaction mixture was cooled to 100°C with KCO3. After basification to pH 10 and extraction with ethyl acetate (5 × 150 mL), the organic layer was washed with brine (2 × 150 mL), dried over anhydrous NaSO, and filtered. The filtrate was concentrated to give methyl 4-[2-(aminomethyl)pyrrolidin-1-yl]-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxylate (2.85 g, crude) as a pale yellow oil. 1H NMR(400MHz, chloroform-d)δ7.39(t, J = 8.3Hz, 1H), 7.34-7.28(m, 3H), 7.19(t, J = 7.6Hz, 3H), 7.07(d, J = 8.3Hz, 1H), 5.44(d, J = 15.5Hz, 1H), 5.32(d, J = 15.4Hz, 1H), 4.25-4.08(m, 2H), 3.92-3.66(m, 5H), 3.11(ddd, J = 10.9, 6.8, 3.2Hz, 1H), 2.81(d, J = 4.9Hz, 2H), 2.04-1.86(m, 2H), 1.75(td, J = 12.2, 7.8Hz, 2H).

[0287] 5-[(4-chlorophenyl)methyl]-4-[3-(trifluoromethoxy)phenoxy]-1,3,5,8-tetraazatricyclo[8.3.0.0^[2,6]]trideca-2(6),3-dien-7-one To a mixture of methyl 4-[2-(aminomethyl)pyrrolidin-1-yl]-1-[(4-chlorophenyl)methyl]-2-[3-(trifluoromethoxy)phenoxy]-1H-imidazole-5-carboxylate (2.75 g, 5.24 mmol, 1 equiv.) in dioxane (50 mL) was added NaH (1.5 g, 36.67 mmol, 7.00 equiv., 60%) at 0° C., and then the reaction mixture was refluxed for 15 minutes. The reaction mixture was cooled to room temperature and concentrated to give a residue. The residual product was purified by silica gel column chromatography, eluting with PE:EA (1:2 to 0:1) to give 5-[(4-chlorophenyl)methyl]-4-[3-(trifluoromethoxy)phenoxy]-1,3,5,8-tetraazatricyclo[8.3.0.0^[2,6]]trideca-2(6),3-dien-7-one (1.5 g, 58.09%) as a white solid. 1H NMR(400MHz, chloroform-d)δ7.38(t, J = 8.3Hz, 1H), 7.26(s, 5H), 7.18(dd, J = 8.4, 2.3Hz, 1H), 7.06(d, J = 8.3Hz, 1H), 5.67(d, J = 14.9Hz, 2H), 5.40(d, J = 14.9Hz, 1H), 3.68-3.38(m, 4H), 3.22-3.09(m, 1H), 2.18(dt, J = 12.8, 6.4Hz, 1H), 1.93(tdd, J = 21.4, 11.8, 6.7Hz, 2H), 1.61(qd, J = 11.5, 7.9Hz, 1H),

[0288] 5-[(4-chlorophenyl)methyl]-8-[3-(oxan-2-yloxy)propyl]-4-[3-(trifluoromethoxy)phenoxy]-1,3,5,8-tetraazatricyclo[8.3.0.0^[2,6]]trideca-2(6),3-dien-7-one To a mixture of 5-[(4-chlorophenyl)methyl]-4-[3-(trifluoromethoxy)phenoxy]-1,3,5,8-tetraazatricyclo[8.3.0.0^[2,6]]trideca-2(6),3-dien-7-one (350 mg, 0.71 mmol, 1 equiv.) in DMF (40 mL) was added NaH (85.2 mg, 2.13 mmol, 3 equiv., 60%) at 0 °C under a nitrogen atmosphere for 0.5 h. To the above mixture was added 2-(3-bromopropoxy)oxane (475.3 mg, 2.13 mmol, 3.00 equiv.) at 0 °C. The resulting mixture was stirred at 50 °C for an additional 16 h. Ethyl acetate (300 mL) and brine (100 mL) were added to the resulting mixture, and the aqueous layer was then extracted with ethyl acetate (200 mL). The combined organic layers were dried over anhydrous Na2SO4 and filtered. The filtrate was concentrated to give the crude product, which was purified by silica gel column chromatography eluting with PE:EA (10:1 to 1:1) to give 5-[(4-chlorophenyl)methyl]-8-[3-(oxan-2-yloxy)propyl]-4-[3-(trifluoromethoxy)phenoxy]-1,3,5,8-tetraazatricyclo[8.3.0.0^[2,6]]trideca-2(6),3-dien-7-one (300 mg, 66.52%) as a pale yellow oil.

[0289] (10R)-5-[(4-chlorophenyl)methyl]-8-(3-hydroxypropyl)-4-[3-(trifluoromethoxy)phenoxy]-1,3,5,8-tetraazatricyclo[8.3.0.0^[2,6]]trideca-2(6),3-dien-7-one and (10S)-5-[(4-chlorophenyl)methyl]-8-(3-hydroxypropyl)-4-[3-(trifluoromethoxy)phenoxy]-1,3,5,8-tetraazatricyclo[8.3.0.0^[2,6]]trideca-2(6),3-dien-7-one To a stirred solution of 5-[(4-chlorophenyl)methyl]-8-[3-(oxan-2-yloxy)propyl]-4-[3-(trifluoromethoxy)phenoxy]-1,3,5,8-tetraazatricyclo[8.3.0.0^[2,6]]trideca-2(6),3-dien-7-one (300 mg, 0.47 mmol, 1 equiv.) in THF (15 mL) was added dropwise HCl (2 M) (15 mL) at room temperature. The resulting mixture was then stirred at room temperature for 2 h. The reaction mixture was basified to pH 10 with KCO and extracted with ethyl acetate (3 x 100 mL). The organic layer was then washed with brine (2 x 50 mL), dried over anhydrous NaSO, and filtered. The filtrate was concentrated to give the crude product, which was purified by preparative chiral HPLC (column: (R,R) Whelk-01, 21.1*250 mm, 5 μm; mobile phase A: Hex--HPLC, mobile phase B: EtOH--HPLC; flow rate: 20 mL / min; gradient: 50B to 50B in 14 min; 220 / 254 nm; RT1: 8.638; RT2: 11.063) to give compound 34 (37.8 mg, 14.52%) and compound 35 (51.9 mg, 19.94%).

[0290] Characterization of compound 34: 1 H NMR (400 MHz, methanol-d4) chemical shifts: 7.49 (t, J = 8.2 Hz, 1H), 7.29 (d, J = 8.2 Hz, 2H), 7.26-7.18 (m, 4H), 7.15 (d, J = 8.3 Hz, 1H), 5.64 (d, J = 15.3 Hz, 1H), 5.34 (d, J = 15.3 Hz, 1H), 3.80 (dt, J = 14.3, 7.4 Hz, 1H), 3.65-3.38 (m, 6H), 3.25 (dd, J = 13.4, 6.5 Hz, 1H), 3.15 (dd, J = 14.8, 7.6 Hz, 1H), 2.24 (dt, J = 12.0, 6.0Hz, 1H), 2.02-1.85(m, 2H), 1.76(p, J = 6.5Hz, 2H), 1.69-1.56(m, 1H). Molecular formula C 26 H 26 Calculated [M+H] for ClF3N4O4 + :551, Observed:551.

[0291] Characterization of compound 35: 1 H NMR (400 MHz, methanol-d4) chemical shifts: 7.49 (t, J = 8.2 Hz, 1H), 7.29 (d, J = 8.3 Hz, 2H), 7.26-7.18 (m, 4H), 7.15 (d, J = 8.3 Hz, 1H), 5.64 (d, J = 15.2 Hz, 1H), 5.34 (d, J = 15.3 Hz, 1H), 3.80 (dt, J = 14.3, 7.4 Hz, 1H), 3.66-3.38 (m, 6H), 3.25 (dd, J = 13.5, 6.4 Hz, 1H), 3.15 (dd, J = 14.8, 7.6 Hz, 1H), 2.24 (dt, J = 12.3, 6.2Hz, 1H), 2.11-1.85(m, 2H), 1.76(p, J = 6.4Hz, 2H), 1.70-1.58(m, 1H). Molecular formula C 26 H 26 Calculated [M+H] for ClF3N4O4 + :551, Observed:551.

[0292] Example 2: Assay Protocol

[0293] I. Human TRPC5-expressing cells. ICLN-1633 cells expressing TRPC5 (HEK-TREx hTRPC5) were generated as follows. Commercially available HekTrex-293 cells were seeded at 0.7 x 10 cells / well in 1 x 6-well plates 24 h prior to transfection using 2 mL of antibiotic-free cell growth medium (1 x DMEM / high glucose (Hyclone #SH30022.02); 10% fetal bovine serum (Sigma), 2 mM sodium pyruvate, 10 mM HEPES). The human TRPC5 coding sequence (NM_012471 containing the silent T478C mutation) was cloned into pcDNA5 / TO (Invitrogen; catalog no. V103320) using hygromycin as a resistance gene, and the plasmid (SEQ ID NO: 1) was propagated in T-Rex-293 cells (Invitrogen; catalog no. R71007) according to the manufacturer's instructions. On day 2, 2 μg of plasmid DNA and 6 μl of Xtreme-GENE HP Reagent in Optimem (total volume 200 μl) were prepared and incubated at room temperature for 15 minutes. The plasmid solution was then gently added dropwise to each well to cover it, and the plate was gently rotated to mix the complex with the medium for approximately 30 seconds. The transfected cells were incubated at 37°C in a 10% CO2 incubator for 24 hours. The transfected cells were harvested and transferred to 2 x 150 mm dishes containing cell growth medium without antibiotics at 37°C.

[0294] The next day, selection for generating stable pools was initiated by adding cell growth medium containing 150 μg / mL hygromycin and 5 μg / mL blasticidin. The medium containing the selection agent was replaced every 1–2 days as needed to remove dead cells. After 7 days, the hygromycin concentration was reduced to 75 μg / mL, and cell growth was continued.

[0295] Single clones were selected as follows: Stable pools were diluted to 10 cells / mL and seeded (100 μL / well) into 24 x 96-well plates (approximately 1 cell / well) and grown in cell growth medium for 7 days. Fresh medium (100 μL) was added, and cells were grown for an additional 1–2 weeks before being cryopreserved or used immediately.

[0296] II. Automated patch clamp assay (Qpatch) Automated electrophysiological assays were performed at room temperature. On the day of the experiment, TRPC5 cells were cultured according to standard operating procedures. Briefly, cells were harvested using TrypLE™ Express, resuspended in serum-free medium, added to the automated platform, and used within 0.5–3 h. Prior to the assay, internal and external saline solutions were freshly prepared. The external solution contained 145 mM NaCl, 4 mM KCl, 2 mM CaCl2, 1 mM MgCl2, 10 mM HEPES, 10 mM glucose, pH 7.4 with NaOH, and 310 mOsm / L. The internal solution contained 120 mM L-aspartate, 120 mM CsOH.H2O, 20 mM CsCl, 2 mM MgCl2, 8.8 mM CaCl2, 10 mM EGTA, 10 mM HEPES, and 2 mM Na2ATP; pH 7.2 and 330 mOsm / L with CsOH. Free internal Ca was measured according to the WCabuf software. 2+ The concentration was buffered to 1 μM. Compound profiling was performed using the automated electrophysiological platform QPatch 16 from Sophion (Denmark). Seal series resistance and quality were continuously monitored throughout the experiment. Data were analyzed using Sophion QPatch Assay Software 5.6 (Odense). Data were normalized using the maximum activation obtained during pre-compound agonist application as the highest value (1.0) and the maximum inhibition induced by ML-204 as the lowest value (0.0). IC50 values ​​were calculated using a least-squares regression algorithm (Hill equation).

[0297] To monitor ionic currents, voltage ramps from -100 mV to +100 mV over 300 ms were applied every 10 s from a holding potential of -60 mV. Antagonist mode: After a minimum 60 s control period of recording, an EC60 concentration of the TRPC5 agonist, rosiglitazone (30 μM), was applied to activate the channel. After steady-state conditions were reached, simultaneous application of increasing concentrations of test compound was applied, followed by application of the EC60 and saturating concentrations of the specific blocker, ML-204 (100 μM).

[0298] III. FLIPR Protocol: Compounds were typically prepared or supplied as stock solutions up to 10 mM, using DMSO as the vehicle. Ten-point dose-response curves were generated using an Echo-550 acoustic dispenser. Compound source plates were created by serially diluting compound stocks to create 10 mM, 1 mM, and 0.1 mM solutions in DMSO in an Echo-certified LDV plate. Echo then serially dispensed 100% DMSO stock solutions into the source dose-response plate, generating a 4-fold dilution scheme. 100% DMSO was added to the dispensed dose-response plate to a final volume of 5 μl. 300 nl of the dose-response stock plate was then dispensed into the preincubation and stimulation assay plate. 50 μl of preincubation buffer and 100 μl of stimulation buffer were then added to the plate to yield a final assay test concentration range of 30 μM to 0.0001 μM with a final DMSO concentration of 0.3%.

[0299] Human ICLN-1633 cells expressing TRPC5 were plated in 384-well black poly-D-lysine-coated microplates and maintained in TRPC5 growth medium the day before use in experiments. TRPC5 expression was induced by applying 1 μg / mL tetracycline at the time of plating. The medium was removed from the plates, and 10 μl of 4 μM Fluo-4 AM (mixed with an equal volume of Pluronic F-127) in Earls' buffered salt solution (EBSS) was added to the cells. The cells were incubated at room temperature, protected from light, for 60–90 minutes. After the incubation period, the dye was removed and replaced with 10 μl of EBSS. The cell, preincubation, and stimulation plates were loaded into the FLIPR-II and the assay was initiated. The FLIPR measured a 10-second baseline, followed by the addition of 10 μl of 2X compound (or control). The change in fluorescence was monitored for an additional 5 minutes. After a 5-minute preincubation, 20 μl of 2× riluzole (containing 1× compound or control) was added to the cell plate. The final riluzole stimulation concentration in the assay was 30 μM. After riluzole addition, changes in fluorescence were monitored for an additional 5 minutes. A decrease in the riluzole-activated calcium response compared to control wells was reported as inhibition. A compound-mediated increase in the riluzole response compared to the control riluzole response (no test agent present) without an enhancement of calcium influx during the preincubation phase was reported as an agonist response.

[0300] Compound inhibition of the TRPC5 calcium response was determined as follows: After addition of riluzole, fluorescence was monitored for 5 minutes. For inhibition, the maximum relative fluorescence response (minus the control response of 1 μM of an internal control compound known to maximally block the TRPC5 calcium response, "REF INHIB" in the formula below) was captured and exported from FLIPR.

[0301] Compound inhibition is calculated using the following formula:

[0302]

number

[0303] Compound activation (agonism) of the TRPC5 calcium response was determined as follows: After the first compound addition, fluorescence was monitored for 5 minutes. The maximum relative fluorescence response (minus the EBSS buffer only control response) was captured and exported from the FLIPR. Compound activation was calculated using the following formula:

[0304]

number

[0305] Example 3: Exemplary Biological Assay Data Table 2: QPatch, FLIPR inhibitor, and FLIPR agonist potency ranges for representative compounds of the present disclosure.

[0306] Effectiveness: Qpatch assay: A = 0.001-1 μM; B = 1-30 μM; C = >30 μM; ND = not tested.

[0307] FLIPR assay (inhibitor and agonist activity): A = 0.001-1 μM; B = 1-10 μM; C = >10 μM; D = Tests positive for agonism but not EC 50 not calculated; ND = not tested.

[0308] The pairs of enantiomers shown in Table 2 (see, e.g., compounds 6 and 7; compounds 12 and 13; compounds 15 and 16, etc.) have stereochemistry relative to each other. In other words, the two enantiomers were separated from a racemic mixture, but the absolute stereochemistry of each has not been determined.

[0309] [Table 7-1]

[0310] [Table 7-2]

[0311] [Table 7-3]

[0312] [Table 7-4]

[0313] SEQ ID NO: 1: TRPC5 plasmid sequence The DNA sequence of the TRPC5 plasmid used in Example 2 is shown below, where the underlined nucleic acid represents the one encoding human TRPC5.

[0314] [ka]

[0315] [ka]

[0316] [ka]

[0317] [ka]

[0318] Incorporation by Reference All US patents and US and PCT published patent applications cited herein are hereby incorporated by reference.

[0319] Doctrine of Equivalents The above specification is sufficient to enable one skilled in the art to practice the present invention. The present invention should not be limited in scope by the provided examples, as the examples are intended as a single illustration of one aspect of the present invention, and other functionally equivalent embodiments are within the scope of the present invention. Various modifications of the present invention, in addition to those shown and described herein, will be apparent to those skilled in the art from the above specification and are intended to be included within the scope of the appended claims. The advantages and objectives of the present invention are not necessarily encompassed by each embodiment of the present invention.

Claims

1. A compound of formula (II), or a tautomer or a pharmaceutically acceptable salt thereof: 【Chemistry 1】 (In the formula, A' is N and A is CR; R is L-R １ and L does not exist, CH ２ , O, SO ２ , or NR ２ and R １ is selected from optionally substituted alkyl, optionally substituted aryl, and optionally substituted heteroaryl; when L is absent, R １ is further selected from H; Each R ２ are independently H or alkyl; R ３ is an optionally substituted alkyl, an optionally substituted alkylene -OR ２ , optionally substituted cycloalkylene -OR ２ , optionally substituted alkylene-N(R ７ ) ２ , optionally substituted cycloalkylene-N(R ７ ) ２ , optionally substituted alkylene -C(O)N(R ２ ) ２ , optionally substituted cycloalkylene -C(O)N(R ２ ) ２ , optionally substituted alkylene-S(O) ２ N (R ２ ) ２ and optionally substituted cycloalkylene-S(O) ２ N (R ２ ) ２ is selected from R ４ is selected from alkyl, optionally substituted alkylene-(6-membered aryl), and optionally substituted alkylene-(5-6 membered heteroaryl); Each R ５ is H, N(R ２ ) ２ , OR ２ are independently selected from Each R ７ is H, alkyl, (alkyl)C(O)-, (aryl)C(O)-, (alkyl)S(O) ２ - and (aryl)S(O) ２ are independently selected from X' is -C(O)-, CH ２ , CHR ３’ , or C(R ３’ ) ２ and Each R ３’ is an optionally substituted alkyl, an optionally substituted alkylene -OR ２ , optionally substituted cycloalkylene -OR ２ , optionally substituted alkylene-N(R ７ ) ２ , optionally substituted cycloalkylene-N(R ７ ) ２ , optionally substituted alkylene -C(O)N(R ２ ) ２ , optionally substituted cycloalkylene -C(O)N(R ２ ) ２ , optionally substituted alkylene-S(O) ２ N (R ２ ) ２ and optionally substituted cycloalkylene-S(O) ２ N (R ２ ) ２ are independently selected from, and Z does not exist.)

2. 2. The compound of claim 1, or a tautomer or pharmaceutically acceptable salt thereof, wherein L is absent.

3. The L is CH ２ 2. The compound of claim 1, wherein:

4. 2. The compound of claim 1, wherein L is O, or a tautomer or a pharmaceutically acceptable salt thereof.

5. The R １ The compound of any one of claims 1 to 4, or a tautomer or pharmaceutically acceptable salt thereof, wherein is optionally substituted phenyl.

6. The substituted phenyl may be a halogen, —CF ３ , -C(H)F ２ , and -OCF ３ 6. The compound of claim 5, or a tautomer or pharmaceutically acceptable salt thereof, substituted with one or more substituents independently selected from:

7. The R １ The compound of any one of claims 1 to 4, or a tautomer or pharmaceutically acceptable salt thereof, wherein is optionally substituted alkyl.

8. The R ２ The compound of any one of claims 1 to 7, or a tautomer or a pharmaceutically acceptable salt thereof, wherein

9. The R ３ The compound of any one of claims 1 to 8, or a tautomer or a pharmaceutically acceptable salt thereof, wherein is alkylene-OH.

10. The R ３ but 【Chemistry 2】 9. The compound of claim 1, wherein the compound is selected from the group consisting of:

11. The R ４ The compound of any one of claims 1 to 10, or a tautomer or pharmaceutically acceptable salt thereof, wherein is alkyl.

12. The R ４ 11. The compound of any one of claims 1 to 10, or a tautomer or pharmaceutically acceptable salt thereof, wherein is optionally substituted alkylene-(6-membered aryl).

13. The aryl may be a halogen, —CF ３ , -C(H)F ２ , or -OCF ３ 13. The compound of claim 12, substituted with: or a tautomer or a pharmaceutically acceptable salt thereof.

14. 13. The compound of claim 12, or a tautomer or pharmaceutically acceptable salt thereof, wherein the aryl of said alkylene-(6-membered aryl) is phenyl and is substituted with one or more halogens.

15. The R ４ teeth 【Transformation 3】 11. The compound according to any one of claims 1 to 10, wherein:

16. Each of the R ５ The compound of any one of claims 1 to 15, or a tautomer or a pharmaceutically acceptable salt thereof, wherein

17. 17. The compound according to any one of claims 1 to 16, or a tautomer or pharmaceutically acceptable salt thereof, wherein X' is -C(O)-.

18. The X' is CH ２ 17. The compound of any one of claims 1 to 16, wherein:

19. The compound is 【Chemistry 4】 【Transformation 5】 【Transformation 6】 2. The compound of claim 1, wherein:

20. 20. A composition comprising a compound according to any one of claims 1 to 19, or a tautomer or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable excipient.

21. A pharmaceutical composition comprising a compound according to any one of claims 1 to 19, or a tautomer or a pharmaceutically acceptable salt thereof, A pharmaceutical composition for treating or reducing the risk of developing kidney disease, diabetic retinopathy, anxiety, depression, pain, obesity, or cancer.

22. 22. The pharmaceutical composition of claim 21 for treating or reducing the risk of developing anxiety, depression, or pain.

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