Pyrrolo[2,3-b]pyridine-3-carboxamide compositions and methods for ameliorating hearing loss

Abstract

Disclosed are N-(3-substituted thiazol-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamides (I) and (II), which are N-(3-substituted oxazol-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamides. The compounds activate Yap and inhibit Lats kinase. Therefore, the compounds are useful for the treatment of hearing loss.

Description

Detailed description of the invention

[0001] <Notice Regarding Government Rights> This invention was made with government support granted by the National Institutes of Health (NIH) under grant number T32GM007739. The U.S. Government has certain rights to this invention.

[0002] [Background of the present invention] <Technical field> This application relates in general to N-(3-substituted thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide and N-(3-substituted oxazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide, which inhibit Lats kinase and consequently increase Yap activity. These compounds are useful for inducing the proliferation of supporting cells in the inner ear and therefore potentially for treating hearing loss.

[0003] <Background information> Regeneration, initiated in response to injury, is a complex process that can restore the structure and function of damaged tissue. Some tissues in mature mammals gradually lose their regenerative capacity after development is complete. Regeneration occurs either through the activation and proliferation of resident stem cells, as in the epithelium of the skin and intestines, or through the dedifferentiation and proliferation of cells, as in the liver. In other examples, such as central nervous system tissue and cardiomyocytes, cells show little to no potential for regeneration after injury.

[0004] Given its fundamental role in development, proliferation, stem cell maintenance, and dedifferentiation, Hippo signaling is an intriguing target for driving regeneration. The potential of the Hippo pathway for regeneration has been well elucidated in numerous organs, including the heart, retina, liver, and intestines. Hippo signaling limits the size of the utricle, a sensory organ in the vestibular portion of the inner ear, in developing murids, and the Yap-Tead complex is active and required for proliferation and regeneration in the neonatal utricle. These observations suggest that chemical activation of Yap signaling may lead to the proliferation of supporting cells in adult tissue, a crucial, yet undiscovered, step in mammalian inner ear regeneration.

[0005] In our efforts to identify Yap activators, we conducted a small molecule screening of cultured cells. We identified a compound that we found to function as a Lats kinase inhibitor. To test our own hypothesis, we treated utricles cultured in vitro from adult mice with the substance and found that several days of exposure triggered a crucial step toward the re-entry of supporting cells into the cell cycle, i.e., steady hair cell regeneration.

[0006] [Overview of the prefecture] The present invention relates to N-(3-substituted azole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide, pharmaceutical compositions, and methods for inhibiting Lats or activating Yap, thereby stimulating the regeneration of target cells, particularly hair cells.

[0007] In a first aspect of this invention, formula I: [ka] (In the formula, R 1 (C1-C6) alkyl, carboxy, (C3-C7) carbomonocyclyl, (C9-C 11)Selected from the group consisting of carbobicyclic, heteromonocyclic, and heterobicyclic, wherein the (C1-C6) alkyl, (C3-C7) carbomonocyclic, (C9-C 11 )The carbobicyclic, heteromonocyclic, and heterobicyclic may optionally be substituted with one to three substituents independently selected from the group consisting of halogen, cyano, hydroxy, nitro, amino, acetoxy, carboxy, (C1-C7) hydrocarbyl, halo(C1-C6) alkyl, (C1-C3) alkoxy, halo(C1-C3) alkoxy, (C1-C6) acyl, (C1-C3) alkoxy(C1-C3) alkyl, hydroxy(C1-C3) alkyl, heteroaryl, benzenesulfonyl, (C1-C3) alkoxycarbonyl, aminocarbonyl, (C1-C3) alkylaminocarbonyl, di(C1-C3) alkylaminocarbonyl, (C1-C3) alkylamino, di(C1-C3) alkylamino, amino(C1-C3) alkyl, (C1-C3) alkylamino(C1-C3) alkyl, (C1-C3) dialkylamino(C1-C3) alkyl, (C1-C3) alkylthio, (C1-C3) alkylsulfonylamino, (C1-C3) alkylsulfinyl, (C1-C3) alkylsulfonyl, phenoxy, and benzyloxy, R 2 is selected from the group consisting of hydrogen, halogen, (C1-C7) hydrocarbyl, halo(C1-C6) alkyl, (C1-C6) acyl, hydroxy(C1-C3) alkyl, -C(=O)O(C1-C6) alkyl, -C(=O)NR 20 R 21 , and (C1-C6) oxaalkyl, R 3 is selected from the group consisting of hydrogen, halogen, (C1-C6) hydrocarbyl, halo(C1-C6) alkyl, (C1-C6) acyl, and (C1-C3) alkoxy, R 4 is selected from the group consisting of hydrogen, halogen, (C1-C6) hydrocarbyl, halo(C1-C6) alkyl, (C1-C6) acyl, and (C1-C3) alkoxy, R 10In each case, it is independently selected from the group consisting of hydrogen and methyl, R 20 It is selected from the group consisting of hydrogen and (C1-C6) hydrocarbyl, R 21 These include hydrogen, (C1-C6)hydrocarbyl, (C1-C6)oxaalkyl, amino(C1-C6)alkyl, (C1-C3)alkylamino(C1-C6)alkyl, di(C1-C3)alkylamino(C1-C6)alkyl, and -(CH2) m Selected from the group consisting of -Het, where Het is hydroxy, amino, acetoxy, carboxy, (C1-C7)hydrocarbyl, halo(C1-C6)alkyl, (C 1‐ It is an aliphatic monocyclic or bicyclic heterocycle optionally substituted with substituents selected from the group consisting of C3)alkoxy, halo(C1-C3)alkoxy, (C1-C6)acyl, (C1-C3)alkoxy(C1-C3)alkyl, hydroxy(C1-C3)alkyl, aminocarbonyl, (C1-C3)alkylaminocarbonyl, di(C1-C3)alkylaminocarbonyl, (C1-C3)alkylamino, and di(C1-C3)alkylamino. Or, R 20 and R 21 R 20 and R 21 Together with the nitrogen to which it is bonded, it forms an aliphatic heterocycle. n is 0, 1, or 2. m is 0, 1, or 2, and X is S, or n is 1 and R 1 If is a substituted phenyl, then X may also be O. However, R 1 If is phenyl, then X is sulfur and n is 1, and R 2 , R 3 , R 4 and R 10 (At least one of them is not hydrogen.) This relates to compounds represented by [the specified formula / method].

[0008] In another aspect, the present invention relates to a pharmaceutical composition comprising a pharmaceutically acceptable carrier and a compound as described herein.

[0009] In another aspect, the present invention relates to a method for activating YAP in cells expressing YAP, comprising exposing the cells to compounds as described herein.

[0010] In another aspect, the present invention relates to a method for inhibiting LATS in a cell population expressing LATS, comprising exposing the cell population to a compound as described herein.

[0011] In another aspect, the present invention relates to a method for LATS inhibition in a cell population, comprising exposing the cell population to a compound as described herein.

[0012] In another aspect, the present invention relates to a method for stimulating hair cell regeneration, comprising exposing a population of supporting cells to a compound as described herein.

[0013] [Detailed description of the invention] Equation I [ka] Compounds represented by were found to be useful in inhibiting Lats or activating Yap, and therefore potential therapeutic agents for stimulating the regeneration of target cells, particularly hair cells. Such compounds may be useful in treating hearing loss. The group of compounds represented by I can be divided into two subgroups.

[0014] In the first subgroup, X is sulfur, and the compound is given by formula II: [ka] This is thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide, represented as [formula].

[0015] In the second subgroup, X is oxygen, and the compound is given by formula III: [ka] (In the formula, n is 1 and R 1 (This is a substituted phenyl in some cases.) This is oxazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide, represented as [formula].

[0016] In some embodiments of formulas I and II, n may be 0. In other embodiments of formulas I to III, n may be 1. In these embodiments, R 10 It can be hydrogen.

[0017] In some embodiments, R 1 These are optionally substituted (C1-C4) alkyl, carboxy, phenyl, cyclohexyl, 5-membered heterocyclyl, 6-membered heterocyclyl, or heterobicyclyl. In particular, R 1 R may be methyl, ethyl, aminobutyl, and carboxyethyl. In other embodiments, R 1 is in some cases a substituted cyclohexyl, or R 1 is in some cases a substituted phenyl, or R 1 R is a heterocycline which may be substituted in some cases, such as pyridinyl, pyrazolyl, piperidinyl, tetrahydropyranil, tetrahydrofuranil, or tetrahydroisoquinolinil. 1 If is optionally a substituted phenyl, the phenyl may have one or two substituents independently selected from halogen, cyano, hydroxy, amino, carboxy, (C1-C6)hydrocarbyl, trifluoromethyl, methoxy, acetyl, formyl, hydroxy(C1-C3)alkyl, methoxycarbonyl[-C(=O)OCH3], carboxamide[-C(=O)NH2], methanesulfonylamino, and amino(C1-C3)alkyl. 1If is pyridinyl, pyrazolyl, piperidinyl, tetrahydropyranyl, or tetrahydroisoquinolinyl, each heterocycle may optionally be substituted with one or two substituents independently selected from amino, hydroxy, and (C1-C6) hydrocarbyl.

[0018] In some embodiments, R 1 R is selected from the group consisting of carboxyl and optionally substituted (C1-C4) alkyl, phenyl, cyclohexyl, 5-membered heterocyclyl, 6-membered heterocyclyl, and heterobicyclyl. In some of the above, R 1 It is selected from the group consisting of methyl, ethyl, aminobutyl, and carboxyethyl. In addition, R 1 R is a substituted cyclohexyl in some cases. In some of the above, R 1 The compound is optionally substituted heterocyclyl. The heterocycle may be optionally substituted pyridinyl, pyrazolyl, piperidinyl, tetrahydropyranyl, tetrahydrofuranyl, and tetrahydroisoquinolinyl. Optional substituents include one or two substituents independently selected from the group consisting of amino, hydroxy, and (C1-C6) hydrocarbyl compounds.

[0019] In some embodiments, R 1 R is optionally substituted with a phenyl molecule. This phenyl molecule may be substituted with one or two substituents independently selected from the group consisting of halogens, cyano, hydroxy, amino, carboxy, (C1-C6)hydrocarbyl, trifluoromethyl, methoxy, acetyl, formyl, hydroxy(C1-C3)alkyl, methoxycarbonyl, carboxamide, methanesulfonylamino, and amino(C1-C3)alkyl molecules. In some of the embodiments described above, R 1 is a phenyl compound substituted at the ortho position, and n is 0; in other embodiments, R 1 n is a substituted phenyl compound, and n is 1.

[0020] In some embodiments, R 2 These are -C(=O)O(C1-C6) alkyl and -C(=O)NR 20 R 21 , and selected from the group consisting of (C1-C6) oxaalkyl groups. In some of these embodiments, R 20 is selected from hydrogen and methyl, and R 21 These are hydrogen, methyl, (C1-C6)oxaalkyl, dimethylamino(C1-C6)alkyl, and -(CH2) m - Selected from Het. Otherwise, R 20 and R 21 R 20 and R 21 It forms a 4-7 membered aliphatic heterocycle with the nitrogen atom to which it is bonded. Typical aliphatic heterocycles include piperidine, piperazine, morpholine, pyrrolidine, azetidine, and azepine.

[0021] In some embodiments, R 2 The element is selected from hydrogen, methyl, ethyl, propyl, cyclopropyl, hydroxymethyl, and trifluoromethyl.

[0022] In some embodiments, R 3 and R 4 This is selected from hydrogen, chloro, and methyl.

[0023] Naturally, in various embodiments, the pharmaceutical composition of the present invention includes one or more pharmaceutically acceptable additives, for example, but not limited to, one or more binders, bulking agents, buffers, stabilizers, surfactants, wetting agents, smoothing agents, excipients, disintegrants, thickeners or devisers, emulsifiers, suspending agents, preservatives, antioxidants, opacifiers, fluidizers, processing aids, colorants, sweeteners, flavoring agents, fragrances, flavoring agents, excipients, glossing agents, polymer matrix systems, plasticizers, and other known additives that give the appearance of a drug or are useful in the manufacture of a pharmaceutical or drug product containing the composition of the present invention. Examples of carriers and additives well known to those skilled in the art are described in detail, for example, Ansel, Howard C. et al., “Ansel's Pharmaceutical Dosage Forms and Drug Delivery Systems,” (Philadelphia, USA), Lippincott, Williams and Wilkins, 2004; Gennaro, Alfonso R. et al., “Remington: The Science and Practice of Pharmacy,” (Philadelphia, USA), Lippincott, Williams and Wilkins, 2000; and Rowe, Raymond C., “Handbook of Pharmaceutical Excipients,” (Chicago, USA), Pharmaceutical Press, 2005.

[0024] In various embodiments, non-limiting examples of additives include, but are not limited to, corn starch, potato starch, or other starches, gelatin, natural and synthetic rubber, e.g., gum arabic, sodium alginate, alginic acid, other alginates, tragacanth powder, guar gum, cellulose and its derivatives (e.g., ethylcellulose, cellulose acetate, calcium carboxymethylcellulose, sodium carboxymethylcellulose), polyvinylpyrrolidone, methylcellulose, pregelatinized starch, hydroxypropyl methylcellulose (e.g., 2208, 2906, 2910), hydroxypropylcellulose, titanium dioxide, talc, calcium carbonate (e.g., granules or powder), microcrystalline cellulose, powdered cellulose, dextrate, kaolin, silicic acid, sorbitol, starch, pregelatinized starch. Agar, alginic acid, calcium carbonate, microcrystalline cellulose, croscarmellose sodium, crospovidone, potassium polaritrin, sodium starch glycolate, potato starch or tapioca starch, other starches, pregelatinized starch, other starches, clay, other algins, other celluloses, rubber, calcium stearate, magnesium stearate, mineral oil, diesel fuel, glycerin, sorbitol, mannitol, polyethylene glycol, other glycols, stearic acid, sodium lauryl sulfate, talc, hydrogenated vegetable oils (e.g., peanut oil, cottonseed oil, sunflower oil, sesame oil, olive oil, corn oil, and soybean oil), zinc stearate, ethyl oleate, ethyl laurate, agar, syloid® silica gel (AEROSIL200), W.R. Grace Company, Baltimore, Maryland, USA (WR Examples include synthetic silica agglomerated aerosols (manufactured by Grace Co.), sold by Degussa Co. of Plano, Texas, USA, CAB-O-SIL (registered trademark) (a calcined silicon dioxide product sold by Cabot Co. of Boston, Massachusetts, USA), colorants, and mixtures thereof.

[0025] The terms “subject” and “subject in need” are used interchangeably herein. These terms refer to patients diagnosed with an underlying disease requiring treatment. Typically, patients will be human. Subjects may be currently experiencing symptoms associated with the disease, or have experienced symptoms in the past. In addition, “subject in need” may be patients who are at risk of developing a particular disease, or who are reporting one or more of the physiological systems of the disease, even if they have not yet been diagnosed with the disease.

[0026] As used herein, the terms “treatment” and “to treat” are interchangeable. These terms refer to methods for obtaining beneficial or desired outcomes, such as therapeutic utility, without limitation. Therapeutic utility includes the eradication or improvement of the underlying disease being treated; and further includes the eradication or improvement of one or more symptoms associated with the underlying disease, even if the patient still has the underlying disease, such that improvement is observed in the patient.

[0027] As used herein, the term "optionally substituted" may be used interchangeably with "unsubstituted or substituted." The term "substituted" refers to the replacement of one or more hydrogen atoms in a given group with a given radical. For example, substituted aryls, heterocyclyls, etc., are those in which one or more H atoms in each residue are replaced with halogens, haloalkyls, alkyls, (C) 1‐8) Hydrocarbyl, acyl, alkoxyalkyl, hydroxy lower alkyl, carbonyl, phenyl, heteroaryl, benzenesulfonyl, hydroxy, lower alkoxy, haloalkoxy, oxaalkyl, carboxy, alkoxycarbonyl [i.e. -C(=O)O-alkyl], carboxamide [i.e. -C(=O)NH2], alkylaminocarbonyl [i.e. -C(=O)NH-alkyl], cyano, acetoxy, nitro, amino, alkylamino, dialkylamino, dialkylaminoalkyl, dialkylaminoalkoxy, heterocyclylalkoxy, arylalkyl, (cycloalkyl) Refers to alkyl, heterocyclyl, heterocyclylalkyl, alkylaminoalkyl, heterocyclylaminoalkyl, heterocyclylalkylaminoalkyl, cycloalkylaminoalkyl, cycloalkylalkylaminoalkyl, arylaminoalkyl, and arylalkylaminoalkyl, mercapto, alkylthio, alkylsulfinyl, benzyl, heterocyclyl, phenoxy, benzyloxy, heteroaryloxy, aminosulfonyl, amidino, guanidino, ureido, -SO2alkyl, -SO2NH2, or -SO2NHalkyl, or aryl or heterocyclyl.Preferred substituents include halogens, cyano, hydroxy, nitro, amino, acetoxy, carboxy, (C1-C7)hydrocarbyl, halo(C1-C6)alkyl, (C1-C3)alkoxy, halo(C1-C3)alkoxy, (C1-C6)acyl, (C1-C3)alkoxy(C1-C3)alkyl, hydroxy(C1-C3)alkyl, heteroaryl, benzenesulfonyl, (C1-C3)alkoxycarbonyl [i.e., -C(=O)O(C1-C3)alkyl], and carboxamide [i.e., -C These include (=O)NH2], (C1-C3)alkylaminocarbonyl [i.e., -C(=O)NH-(C1-C3)alkyl], (C1-C3)alkylamino, di(C1-C3)alkylamino, amino(C1-C3)alkyl, (C1-C3)alkylamino(C1-C3)alkyl, (C1-C3)dialkylamino(C1-C3)alkyl, (C1-C3)alkylthio, (C1-C3)alkylsulfonylamino, (C1-C3)alkylsulfinyl, (C1-C3)alkylsulfonyl, phenoxy, and benzyloxy.

[0028] Unless otherwise specified, alkyl groups are linear or branched hydrocarbyl groups. Unless otherwise specified, unsubstituted alkyl groups have 1 to 20 carbon atoms (e.g., 1 to 6 carbon atoms). Examples of alkyl groups include methyl, ethyl, propyl, isopropyl, butyl, s-, and t-butyl.

[0029] Hydrocarbons or hydrocarbyls (as substituents) include alkyl, cycloalkyl, polycycloalkyl, alkenyl, alkynyl, aryl, and combinations thereof. Examples include cyclopropylmethyl, benzyl, phenethyl, cyclohexylmethyl, camphoryl, and naphthylethyl. A hydrocarbon refers to any substituent that consists only of hydrogen and carbon as elemental components. Cycloalkyls are a subgroup of hydrocarbyls and include cyclic hydrocarbon groups with 3 to 8 carbon atoms. Examples of cycloalkyl groups include c-propyl, c-butyl, c-pentyl, and norbornyl.

[0030] Unless otherwise specified, the term "carbocycle" refers to a ring system in which all ring atoms are carbon atoms in any oxidation state. Therefore, a (C3-C8) carbocycle refers to both non-aromatic and aromatic systems, such as cyclopropane, benzene, and cyclohexene; (C8-C 12 A polycyclic carbon molecule refers to systems such as norbornane, decalin, indanes, and naphthalenes. Unless otherwise specified, a carbon ring refers to monocyclic, dicyclic, and polycyclic compounds.

[0031] Oxaalkyl refers to an alkyl residue in which one or more carbon atoms (and their associated hydrogens) are replaced by oxygen. Examples include methoxypropoxy and 3,6,9-trioxadecyl. The term oxaalkyl is used in the sense understood in this art [see "Naming and Indexing of Chemical Substances for Chemical Abstracts", 196, published by the American Chemical Society, except for the provision in 127(a)], that is, it refers to a compound in which oxygen is bonded to its adjacent atom by a single bond (forming an ether bond); it does not refer to double-bonded oxygen as seen in carbonyl groups. Alkoxy or alkoxyl is a subgroup of oxaalkyl that refers to a linear or branched group of 1 to 8 carbon atoms bonded to the parent structure via oxygen. Examples include methoxy, ethoxy, propoxy, isopropoxy, cyclopropyloxy, and cyclohexyloxy. Lower alkoxy refers to a group containing 1 to 4 carbon atoms. Regarding the purpose of this application, examples of alkoxys and lower alkoxys include methylenedioxy and ethylenedioxy.

[0032] Unless otherwise specified, acyl refers to a group of 1, 2, 3, 4, 5, 6, 7, and 8 carbon atoms in a linear, branched, and cyclic structure, which is saturated, unsaturated, aromatic, or a combination thereof, bonded to the parent structure by a carbonyl functional group, along with formyl. One or more carbon atoms in an acyl residue may be replaced with nitrogen, oxygen, or sulfur, as long as the bond site to the parent is carbonyl. Examples include formyl, acetyl, benzoyl, propionyl, isobutyryl, t-butoxycarbonyl, and benzyloxycarbonyl. Lower acyls refer to groups containing 1 to 4 carbon atoms. A double-bonded oxygen atom is called "oxo" when it is referred to as a substituent itself.

[0033] Aryl and heteroaryl mean (i) a monocyclic 5- or 6-membered heteroaromatic ring containing a phenyl group (or benzene) or 1 to 4 heteroatoms selected from O, N, or S; (ii) a bicyclic 9- or 10-membered aromatic ring or heteroaromatic ring structure containing 0 to 4 heteroatoms selected from O, N, or S; or (iii) a tricyclic 13- or 14-membered aromatic ring or heteroaromatic ring structure containing 0 to 5 heteroatoms selected from O, N, or S. Examples of 6- to 14-membered aromatic carbocyclic rings include benzene, naphthalene, indan, tetraline, and fluorene, while examples of 5- to 10-membered aromatic heteroaromatic rings include imidazole, pyridine, indole, thiophene, benzopyranone, thiazole, furan, benzimidazole, quinoline, isoquinoline, quinoxaline, pyrimidine, pyrazine, tetrazole, and pyrazole. As used herein, aryl and heteroaryl refer to residues in which one or more rings are aromatic rings, but not all rings are necessarily aromatic rings.

[0034] An arylalkyl group refers to a substituent in which an aryl residue is bonded to the parent structure by an alkyl group. Examples include benzyl and phenethyl. A heteroarylalkyl group refers to a substituent in which a heteroaryl residue is bonded to the parent structure by an alkyl group. In one embodiment, the alkyl group of an arylalkyl group or heteroarylalkyl group is an alkyl group with 1 to 6 carbon atoms. Examples include pyridinylmethyl and pyrimidinylethyl.

[0035] A heterocycle refers to a cycloalkyl or aryl carbocyclic residue in which 1 to 4 carbon atoms are replaced by heteroatoms selected from the group consisting of N, O, and S. The nitrogen and sulfur heteroatoms may be oxidized, and the nitrogen heteroatom may be quaternized. Unless otherwise specified, heterocycles may be non-aromatic (i.e., aliphatic) or aromatic. Examples of heterocycles include pyrrolidine, pyrazole, pyrrole, indole, quinoline, isoquinoline, tetrahydroisoquinoline, benzofuran, benzodioxane, benzodioxol (commonly called methylenedioxyphenyl when present as a substituent), tetrazole, morpholine, thiazole, pyridine, pyridazine, pyrimidine, thiophene, furan, oxazole, oxazoline, isoxazole, dioxane, and tetrahydrofuran. Note that heteroaryl is a subgroup of heterocycles in which the heterocycle is aromatic. Examples of heteroaromatic rings include furan, benzofuran, isobenzofuran, pyrrole, indole, isoindole, thiophene, benzothiophene, imidazole, benzimidazole, purine, pyrazole, indazole, oxazole, benzoxazole, isoxazole, benzoisoxazole, thiazole, benzothiazole, triazole, tetrazole, pyridine, quinoline, isoquinoline, pyrazine, quinoxaline, acridine, pyrimidine, quinazoline, pyridazine, cinnoline, phthalazine, and triazine.Further examples of heterocyclyl residues include piperazinyl, 2-oxopiperazinyl, 2-oxopiperidinyl, 2-oxo-pyrrolidinyl, 2-oxoazepinyl, azepinyl, 4-piperidinyl, pyrazolidinyl, imidazolyl, imidazolinyl, imidazolidinyl, pyrazinyl, oxazolidinyl, isoxazolidinyl, thiazolidinyl, isothiazolyl, quinuclidinyl, isothiazolidinyl, benzimidazolyl, thiadiazolyl, benzopyranil, benzothiazolyl, tetrahydrofuryl, tetrahydropyranil, thienyl, benzothienyl, thiamorpholinyl, thiamorpholinylsulfoxide, thiamorpholinylsulfone, oxadiazolyl, triazolyl, and tetrahydroquinolinyl.

[0036] An oxygen heterocycle is a heterocycle containing at least one oxygen atom; it may also contain additional oxygen and other heteroatoms. A sulfur heterocycle is a heterocycle containing at least one sulfur atom; it may also contain additional sulfur and other heteroatoms. Oxygen heteroaryls are a subgroup of oxygen heterocycles; examples include furan and oxazole. Sulfur heteroaryls are a subgroup of sulfur heterocycles; examples include thiophene and thiazine. A nitrogen heterocycle is a heterocycle containing at least one nitrogen atom; it may also contain additional nitrogen and other heteroatoms. Aliphatic nitrogen heterocycles include piperidine, piperazine, morpholine, pyrrolidine, thiomorpholine, azetidine, azepine, and oxazepine. Nitrogen heteroaryls are a subgroup of nitrogen heterocycles; examples include pyridine, pyrrole, and thiazole.

[0037] As used herein and as will be understood by those skilled in the art, when the term “compound” is used, it is intended to include salts of that compound unless it is clearly stated that it is further limited. Thus, for example, the statement “compound of formula I” as shown above means T32 as a free base and as a salt thereof. [ka] The compound will include (wherein Z is any counterion). In certain embodiments, the term “compound of formula I” refers to the compound or a pharmaceutically acceptable salt thereof. The term “pharmaceutically acceptable salt” refers to a salt prepared from a pharmaceutically acceptable, non-toxic acid or base, such as inorganic acids and inorganic bases, and organic acids and organic bases. If the compound of the present invention is basic, the salt can be prepared from a pharmaceutically acceptable, non-toxic acid, including inorganic acids and organic acids, as shown above in this paragraph. pharmaceutically acceptable acid addition salts suitable for the compounds of the present invention include acetic acid, adipic acid, alginic acid, ascorbic acid, aspartic acid, benzenesulfonic acid (besylic acid), benzoic acid, boric acid, butyric acid, camphoric acid, camphorsulfonic acid, carbonic acid, citric acid, ethanedisulfonic acid, ethanesulfonic acid, ethylenediaminetetraacetic acid, formic acid, fumaric acid, glucoheptonic acid, gluconic acid, glutamic acid, hydrobromic acid, hydrochloric acid, hydroiodic acid, and hydroxypropyl alcohol. Examples of addition salts include roxynaphthoic acid, isethionic acid, lactic acid, lactobionic acid, lauryl sulfonic acid, maleic acid, malic acid, mandelic acid, methanesulfonic acid, mucinic acid, naphthylenesulfonic acid, nitric acid, oleic acid, pamoic acid, pantothenic acid, phosphoric acid, pivalic acid, polygalacturonic acid, salicylic acid, stearic acid, succinic acid, sulfuric acid, tannic acid, tartaric acid, teoclatic acid, and p-toluenesulfonic acid. When a compound contains an acidic side chain, for example, R as in T34. 1When is COOH, pharmaceutically acceptable base addition salts suitable for the compounds of the present invention include, but are not limited to, metal salts made from aluminum, calcium, lithium, magnesium, potassium, sodium, and zinc, or organic salts made from lysine, arginine, N,N'-dibenzylethylenediamine, chloroprocaine, choline, diethanolamine, ethylenediamine, meglumine (N-methylglucamine), and procaine. Further pharmaceutically acceptable salts include, where appropriate, non-toxic ammonium cations bonded to alkyl groups having 1 to 20 carbon atoms, as well as carboxylic acids, sulfonic acids, and phosphonate anions.

[0038] <Treatment method> The compositions described herein may be administered to subjects with hearing loss (e.g., sensorineural hearing loss) and / or vestibular dysfunction, or at risk of developing such dysfunction, by various routes, such as local administration to the middle or inner ear (e.g., to or via the oval window, round window, or semicircular canals (e.g., horizontal semicircular canals), or by transtympanic or intramural injection), intravenous, parenteral, intradermal, percutaneous, intramuscular, intranasal, subcutaneous, percutaneous, tracheal, intraperitoneal, intra-arterial, intravascular, inhalation, perfusion, irrigation, and oral administration. The most suitable route of administration in any given case will vary depending on the specific composition being administered, the patient, the formulation method of the drug, the method of administration (e.g., time of administration and route of administration), the patient's age, weight, sex, severity of the disease being treated, the patient's diet, and the patient's excretion rate. The composition can be administered once or more times (for example, once a year, twice a year, three times a year, every other month, once a month, or every other week).

[0039] Subjects who can be treated as described herein are subjects who have hearing loss and / or vestibular dysfunction or are at risk of developing such (e.g., subjects who have hearing loss, vestibular dysfunction, or both, or are at risk of developing such). The compositions and methods described herein are for subjects who have or are at risk of developing cochlear hair cell damage (e.g., acoustic trauma, disease or infection, head injury, ototoxic drugs, or age-related damage), subjects who have or are at risk of developing vestibular hair cell damage (e.g., disease or infection, head injury, ototoxic drugs, or age-related damage), subjects who have or are at risk of developing sensorineural hearing loss, deafness, or auditory neuropathy, and vestibular It can be used to treat subjects who have or are at risk of developing functional impairments (e.g., vertigo, vertigo, loss of balance, bilateral vestibular dysfunction, oscilloscope, or balance disorders), subjects who have tinnitus (e.g., tinnitus alone, or tinnitus associated with sensorineural hearing loss or vestibular dysfunction), subjects who have gene mutations associated with hearing loss and / or vestibular dysfunction, or subjects who have a family history of hereditary hearing loss, deafness, auditory neuropathy, tinnitus, or vestibular dysfunction. In some embodiments, subjects have or are at risk of developing hearing loss and / or vestibular dysfunction associated with or resulting from loss of hair cells (e.g., hair cells in the cochlea or vestibule). The methods described herein may include a step of screening subjects for one or more mutations in genes known to be associated with hearing loss and / or vestibular dysfunction prior to treatment with or administration of the compositions described herein. Subjects can be screened for genetic mutations using standard methods known to those skilled in the art (e.g., genetic testing). The methods described herein may further include a step of evaluating the subject's hearing and / or vestibular function prior to treatment with or administration of the compositions described herein. Hearing can be evaluated using standard tests, such as audiometry, auditory brainstem response (ABR), electrocochleography (ECOG), and otoacoustic emissions.Vestibular function can be assessed using standard tests, such as eye movement tests (e.g., electrooculography (ENG) or video nystagmography (VNG)), vestibular-ocular reflex (VOR) tests (e.g., head impulse tests (Halmagyi-Curthoys test) or calorie reflex tests, which can be performed at the bedside or using a video head impulse test (VHIT)), postural tests, swivel chair tests, ECOG, vestibular evoked myoelectric potential (VEMP), and specialized clinical equilibrium function tests, such as those described in Mancini and Horak, Eur J Phys Rehabil Med, (Italy), 2010, Vol. 46, p. 239. These tests can also be used to assess the hearing and / or vestibular function of a subject after treatment with or administration of the compositions described herein. The compositions and methods described herein may be used as prophylactic measures for patients at risk of developing hearing loss and / or vestibular dysfunction, for example, patients with a family history of hearing loss or vestibular dysfunction (e.g., hereditary hearing loss or vestibular dysfunction), patients who have a gene mutation associated with hearing loss or vestibular dysfunction but have not yet shown hearing impairment or vestibular dysfunction, or patients exposed to risk factors for acquired hearing loss (e.g., acoustic trauma, disease or infection, head injury, ototoxic drugs, or aging) or risk factors for vestibular dysfunction (e.g., disease or infection, head injury, ototoxic drugs, or aging).

[0040] The compositions and methods described herein can be used to induce or increase hair cell regeneration (e.g., cochlear and / or vestibular hair cell regeneration) in subjects. Subjects who may benefit from compositions that induce or increase hair cell regeneration include subjects who suffer hearing loss or vestibular dysfunction as a result of hair cell loss (e.g., hair cell loss associated with trauma (e.g., acoustic trauma or head injury), disease or infection, ototoxic drugs, or aging), and subjects who have abnormal hair cells (e.g., hair cells that do not function properly compared to normal hair cells), damaged hair cells (e.g., hair cell damage associated with trauma (e.g., acoustic trauma or head injury), disease or infection, ototoxic drugs, or aging), or a reduced number of hair cells due to gene mutations or congenital abnormalities.

[0041] The compositions and methods described herein may also be used to prevent or reduce hearing loss and / or vestibular dysfunction caused by damage or death of hair cells (e.g., cochlear hair cells and / or vestibular hair cells) induced by ototoxic drugs in subjects who have been receiving, are currently receiving, or are scheduled to begin treatment with ototoxic drugs. Ototoxic drugs are toxic to cells of the inner ear and may cause sensorineural hearing loss, vestibular dysfunction (e.g., rotational vertigo, dizziness, ataxia, bilateral vestibular dysfunction, oscilloscope, or balance disorders), tinnitus, or a combination of these conditions. Drugs found to be ototoxic include aminoglycoside antibiotics (e.g., gentamicin, neomycin, streptomycin, tobramycin, kanamycin, vancomycin, and amikacin), biomycin, antineoplastic agents (e.g., platinum-containing chemotherapeutic agents such as cisplatin, carboplatin, and oxaliplatin), loop diuretics (e.g., ethacric acid and furosemide), salicylates (e.g., particularly high doses of aspirin), and quinine. In some embodiments, the methods and compositions described herein can be used to treat bilateral vestibular hypodysfunction or oscillopsia. Bilateral vestibular hypodysfunction and oscillopsia may be caused by aminoglycosides (for example, the methods and compositions described herein can be used to promote or increase hair cell regeneration in subjects who have or are at risk of developing aminoglycoside-induced bilateral vestibular hypodysfunction or oscillopsia).

[0042] Treatment may include administration of compositions containing the compounds described herein in various unit doses. Each unit dose will typically contain a predetermined amount of the therapeutic composition. The amount administered, as well as the specific route of administration and dosage form, is within the scope of the skills of those skilled in the field of clinical treatment. The unit dose does not need to be administered as a single injection and may include continuous infusion over a set period. Dosage may be carried out using a syringe pump with controlled infusion rate to minimize damage to the inner ear (e.g., the cochlea and / or vestibular system).

[0043] The compositions described herein are administered in amounts sufficient to improve hearing, improve vestibular function (e.g., improve balance or reduce dizziness or vertigo), reduce tinnitus, treat bilateral vestibular dysfunction, treat oscilloscopes, treat balance disorders, increase or induce hair cell regeneration (e.g., cochlear and / or vestibular hair cell regeneration), increase hair cell count, activate YAP, and / or inhibit LATS. Hearing can be assessed using standard audiometric tests (e.g., audiometry, ABR, electrocochleography (ECOG), and otoacoustic emissions) and may be improved compared to audiometric results obtained before treatment. Vestibular function can be assessed using standard tests for balance and rotational vertigo (e.g., eye movement tests (e.g., ENG or VNG), postural tests, VOR tests (e.g., head impulse tests (Halmagyi-Curthoys test, e.g., VHIT), or calorie reflex tests), swivel chair tests, ECOG, VEMP, and specialized clinical balance function tests), and may improve compared to measurements obtained before treatment. In some embodiments, the composition is administered in an amount sufficient to improve the subject's ability to understand speech sounds. The compositions described herein may also be administered in amounts sufficient to delay or prevent the onset or progression of sensorineural hearing loss and / or vestibular dysfunction (for example, in subjects who have a gene mutation associated with hearing loss or vestibular dysfunction, a family history of hearing loss or vestibular dysfunction (e.g., hereditary hearing loss or vestibular dysfunction), or who have been exposed to risk factors associated with hearing loss or vestibular dysfunction (e.g., ototoxic drugs, head trauma, disease or infection, or acoustic trauma) but have not shown hearing loss or vestibular dysfunction (e.g., vertigo, dizziness, or ataxia), or in subjects who show mild to moderate hearing loss or vestibular dysfunction). The above effects may occur within, for example, one, two, three, four, five, six, seven, eight, nine, ten, fifteen, twenty, or twenty-five weeks, or longer, after administration of the compositions described herein.Patients may be evaluated one, two, three, four, five, six months, or more after administration of the composition, depending on the dose and route of administration used in treatment. Depending on the results of the evaluation, patients may receive additional treatment.

[0044] <Preparation of Compounds> The following abbreviations: THF (tetrahydrofuran), MeOH (methanol), DCM (dichloromethane), DMF (N,N-dimethylformamide), ACN (acetonitrile), EtOH (ethanol), SiO (ethyl acetate), IPA (2-propanol), DMSO (dimethyl sulfoxide), MTBE (methyl tert-butyl ether), TEA (triethylamine), DIPEA (N,N-diisopropylethylamine), TMEDA (tetramethylethylenediamine), DMAP (N,N-dimethylpyridine-4-amine), EDCI (N-(3-di Methylaminopropyl)-N'-ethylcarbodiimide hydrochloride, HOBt (1-hydroxybenzotriazole hydrate), HBTU (2-(1H-benzotriazole-1-yl)-1,1,3,3-tetramethyluronium hexafluorophosphate), T3P (propanephosphonic anhydride), TBAI (tetrabutylammonium iodide), LAH (lithium aluminum hydride), XPhos (2-dicyclohexylphosphin-2',4',6'-triisopropylbiphenyl), and TFA (trifluoroacetic acid) are used in the synthesis pathway.

[0045] Preparative HPLC purification refers to the use of a water / acetonitrile gradient with a suitable hydrophobic stationary phase and, in some cases, additives such as HCl, formic acid, TFA, or NH4HCO3.

[0046] The compounds of the present invention can be prepared as shown in the following general schemes I-IV and more specifically in schemes 1-69. A detailed description of the synthesis of intermediates and exemplary compounds is also disclosed below.

[0047] [ka] As shown in Scheme I, when a compound represented by formula (Ia) containing an imine group is treated with a compound represented by formula (Ib) (wherein X is chloro, bromo, or -OH) under coupling conditions well known to those skilled in the art, a compound represented by formula I is obtained. Typical conditions for the reaction of the compound of formula (Ib) (wherein X is chloro) with the compound of formula (Ia) include, but are not limited to, stirring an equimolar mixture of the compounds in a solvent such as chloroform or dichloromethane in the presence of a base, such as, but not limited to, triethylamine and N,N-dimethylpyridine-4-amine (DMAP), at 5 to 25°C for 1 to 12 hours. The acid coupling conditions for the compound of formula (Ib) (wherein X is -OH) and the compound of formula (Ia) include, but are not limited to, stirring an equimolar mixture of the compounds with a coupling reagent, such as, but not limited to, N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (EDCI), 1-hydroxybenzotriazole hydrate (HOBt), ((2-(1H-benzotriazole-1-yl)-1,1,3,3-tetramethyluronium hexafluorophosphate (HBTU), or propanephosphonic anhydride (T3P), in the presence of a base, such as, but not limited to, N,N-diisopropylethylamine (DIPEA), in a solvent such as, but not limited to, DMF, RINKAN, RINKAN, and pyridine. A typical reaction can be carried out at 25-110°C for 1-12 hours.

[0048] [ka] As shown in Scheme II, the compound represented by formula (IIa) may be converted to the compound of formula I, which is the representative compound of the present invention. Typical conditions include, but are not limited to, treatment of the compound of formula (IIa) with potassium carbonate (K2CO3) in DMF at 25°C, followed by the addition of a reagent such as R1-Y (where R1 is defined in formula I, and Y is chloro, bromo, iodine, mesyl, or tosylate). A typical reaction can be carried out in a microwave reactor at 100°C to promote alkylation.

[0049] [ka] The compound represented by formula (Ia) in Scheme I can be prepared according to the method outlined in Scheme III. The compound represented by formula (IIIa) is treated with potassium carbonate (K2CO3) in DMF at 25°C, followed by the addition of a reagent such as R1-Y (where R1 is defined in formula I and Y is chloro, bromo, iodine, mesyl, or tosylate) to obtain the compound represented by formula (IIIb / Ia). A typical reaction can be carried out in a microwave reactor at 100°C to promote alkylation.

[0050] [ka] Scheme IV describes an alternative preparation method for the compound represented by formula (Ia) in Scheme I. When the compound represented by formula (IVa) is treated with ethyl N-(thioxomethylene) carbamate and tetramethylethylenediamine (TMEDA), the compound represented by formula (IVb) is obtained. When the compound represented by formula (IVb) is treated with the compound represented by formula (IVc) in acetonitrile in the presence of cesium carbonate (Cs2CO3), the compound represented by formula (IVd) is obtained. When the compound represented by formula (IVd) is treated with sulfonyl chloride (SOCl2) in the presence of N,N-diisopropylethylamine (DIPEA), the compound represented by formula (IVe) is obtained. When the compound represented by formula (IVe) is treated with sodium hydroxide (NaOH), the compound represented by formula (IVf / Ia) is obtained.

[0051] The acid Ib used in Scheme I is shown in Table A. For known acids, the CAS number is indicated in the reference column. Synthesis schemes for all other acids are shown in this application. [Table 1]

[0052] Intermediate Ia / IIIb, used in Scheme I and prepared by the method outlined in Scheme III, are shown in Table B. [Table 2] TIFF2023512823000013.tif204131TIFF2023512823000014.tif91135

[0053] Intermediate Ia / IVf, used in Scheme I and prepared by the method outlined in Scheme IV, is shown in Table C. [Table 3] TIFF2023512823000016.tif210131TIFF2023512823000017.tif205131TIFF2023512823000018.tif215131

[0054] The preparation of the intermediates in Table D is shown in Schemes 51-52. [Table 4]

[0055] <Preparation of Intermediate B1> [ka] Chloromethylbenzene (18.96 g, 149.8 mmol, 1.5 equivalents) was added to a solution of 2-thiazoleamine (10.00 g, 99.86 mmol, 1.0 equivalent) in DMF (200 mL). The reaction mixture was heated to 80°C and stirred for 12 hours. The reaction mixture was concentrated under reduced pressure. The resulting residue was diluted with HCl (200 mL). An aqueous solution of NaOH (100 mL, 40% (wt%)) was added at 0°C and stirred at 0°C for 1 hour. The mixture was extracted with HCl (3 × 100 mL). The organic layers were washed together with brine (3 × 100 mL), dehydrated with Na₂SO₄, filtered, and concentrated under reduced pressure to obtain a residue. This residue was purified by silica gel column chromatography (petroleum ether / HCl = 5:1 → 1:1) to obtain 3-benzylthiazole-2(3H)-imine B1 (13.00 g, crude) as a yellow oil.

[0056] <Alternative method for preparing intermediate B1> [ka] <Ethyl N-(benzylcarbamoti oil)carbamate> Phenylmethaneamine (5.00 g, 46.7 mmol, 1.0 equivalent) and O-ethyl carbonisothiocyanatidate (6.43 g, 49.0 mmol, 1.05 equivalent) were dissolved in ethyl acetate (40 mL), to which TMEDA (0.54 g, 4.67 mmol, 0.1 equivalent) was added under a nitrogen atmosphere at 25°C. The mixture was stirred at 25°C for 6 hours and then concentrated under reduced pressure. The residue was triturated twice with a solution of petroleum ether and ethyl acetate (v / v = 10 / 1, 55 mL) to obtain 10.0 g of ethyl N-(benzylcarbamotioil)carbamate as a yellow solid. 1 H NMR (400 MHz, DMSO-d6) δ 11.04 (br. s, 1H), 10.22 (br. t, J = 5.6 Hz, 1H), 7.39 - 7.23 (m, 5H), 4.81 (d, J = 5.6 Hz, 2H), 4.14 (q, J = 7.2 Hz, 2H), 1.21 (t, J = 7.2 Hz, 3H).

[0057] <(Z)-Ethyl((benzylamino)((2-oxoethyl)thio)methylene)carbamate> A mixture of ethyl N-(benzylcarbamotioil)carbamate (4.00 g, 16.8 mmol, 1.0 equivalent) and cesium carbonate (13.7 g, 42.0 mmol, 2.5 equivalents) in acetonitrile (50 mL) was mixed with 2-chloroacetaldehyde (13.2 g, 67.2 mmol, 4.0 equivalents). The reaction mixture was stirred at 25 °C for 12 hours, poured into ice water (20 mL), and then extracted with MTBE (2 × 200 mL). The organic phases were combined and washed with brine (2 × 20 mL), dehydrated with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 100:1 → 3:1) to obtain 5.00 g of (Z)-ethyl((benzylamino)((2-oxoethyl)thio)methylene)carbamate as a yellow solid. LCMS (m / z [M+H] + ): 280.9.

[0058] <(Z)-Ethyl(3-benzylthiazole-2(3H)-ylidene)carbamate> A mixture of (Z)-ethyl((benzylamino)((2-oxoethyl)thio)methylene)carbamate (5.00 g, 12.4 mmol, 1.0 equivalent) and diisopropylethylamine (4.81 g, 37.2 mmol, 3.0 equivalent) in tetrahydrofuran (50 mL) was mixed with thionyl chloride (1.48 g, 12.4 mmol, 1.0 equivalent). The mixture was stirred at 25°C for 16 hours, then poured into ice water (20 mL) and extracted with ethyl acetate (3 × 100 mL). The organic phases were combined and washed with brine (20 mL), dehydrated with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 100:1 → 4:1) to obtain 2.80 g of (Z)-ethyl(3-benzylthiazole-2(3H)-ylidene)carbamate as a yellow solid. 1 H NMR (400 MHz, CD3OD) δ 7.36 - 7.28 (m, 5H), 7.22 (d, J = 4.8 Hz, 1H), 6.84 (d, J = 4.8 Hz, 1H), 5.31 (s, 2H), 4.20 (q, J = 7.2 Hz, 2H), 1.30 (t, J = 7.2Hz, 3H). LCMS (m / z [M+H] + ): 262.9.

[0059] <Intermediate B1> A mixture of (Z)-ethyl(3-benzylthiazole-2(3H)-ylidene) carbamate (2.80 g, 9.06 mmol, 1.0 equivalent) and sodium hydroxide (7.25 g, 181 mmol, 20 equivalents) in ethanol (20 mL) was stirred at 50°C for 1.5 hours. The mixture was poured into ice water (20 mL) and extracted with ethyl acetate (5 × 100 mL). The organic phases were combined and washed with brine (20 mL), dehydrated with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain 1.60 g of 3-benzylthiazole-2(3H)-imine B1 as a yellow rubbery substance.

[0060] <Preparation of Intermediate B2> [ka] To a solution of 2-thiazoleamine (1.00 g, 9.99 mmol, 1.0 equivalent) in EtOH (10 mL), MeI (1.84 g, 13.0 mmol, 808 μL, 1.3 equivalents) was added. The reaction mixture was stirred at 90°C for 12 hours. The suspension was filtered, the solid was washed with EtOH (60 mL), and dried under reduced pressure. The solid was collected, diluted with 40% NaOH aqueous solution (10 mL), and extracted with SiO (3 × 10 mL). The organic layers were washed together with 40% NaOH aqueous solution (3 × 10 mL) and brine (3 × 10 mL), dehydrated with anhydrous Na2SO4, filtered, and concentrated under reduced pressure to obtain a residue of 160 mg of 3-methyl-thiazole-2-amine B2.

[0061] <Preparation of Intermediate B3> [ka] A mixture of 2-thiazoleamine (3.00 g, 30.0 mmol, 1.0 equivalent) and ethyl iodide (EtI) (6.07 g, 38.9 mmol, 1.3 equivalents) in EtOH (30 mL) was degassed, purged three times with N2, and stirred at 90°C for 12 hours under an N2 atmosphere. The reaction mixture was diluted with 40% NaOH aqueous solution (50 mL) and extracted with SiO2 (2 × 30 mL). The organic layers were washed together with 40% NaOH aqueous solution (3 × 20 mL) and brine (3 × 20 mL), dehydrated with anhydrous Na2SO4, filtered, and concentrated under reduced pressure to obtain 2.80 g of 3-ethylthiazole-2(3H)-imine B3 as a dark red oil.

[0062] <Preparation of Intermediate B4> [ka] A mixture of 2-thiazoleamine (120 mg, 1.20 mmol, 1.0 equivalent), tert-butyl(4-bromobutan-2-yl)carbamate (302 mg, 1.20 mmol, 1.0 equivalent), and DIPEA (170 mg, 1.32 mmol, 230 L, 1.1 equivalents) was stirred in a 5 mL sealed tube at 90°C for 4 hours. The reaction mixture was diluted with MeOH (1 mL), and the mixture was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 1 / 1 → 0 / 1, then ethyl acetate / ammonium hydroxide (25%) = 50 / 1) to obtain 30 mg of tert-butyl(4-(2-iminothiazole-3(2H)-yl)butan-2-yl)carbamate B4 as a yellow oil.

[0063] <Preparation of Intermediate B5> [ka] A mixture of 2-thiazoleamine (1.00 g, 10.0 mmol, 1.0 equivalent) and 4-(bromomethyl)benzonitrile (1.96 g, 10.0 mmol, 1.0 equivalent) in i-PrOH (20 mL) was stirred at 60°C for 12 hours. The reaction mixture was filtered to collect the solid, which was washed with 20 mL of siRNA and vacuum-dried to obtain 2.1 g of 2-amino-3-(4-cyanobenzyl)thiazole-3-ium B5 as a yellow solid.

[0064] <Preparation of Intermediate B6> [ka] To a solution of 2-thiazoleamine (1.00 g, 9.99 mmol, 1.0 equivalent) and (bromomethyl)cyclohexane (1.77 g, 9.99 mmol, 1.39 mL, 1.0 equivalent) in DMF (10 mL), TBAI (1.84 g, 4.99 mmol, 0.5 equivalent) was added at room temperature. The reaction mixture was then stirred under N2 at 85°C for 12 hours. An additional 1.0 equivalent of (bromomethyl)cyclohexane was added to the mixture, and the reaction mixture was stirred under N2 at 100°C for a further 24 hours. The reaction mixture was diluted with saturated NaHCO3 aqueous solution (100 mL) and extracted with SiO2 (3 × 50 mL). The organic layers were washed together with brine (3 × 50 mL), dehydrated with Na2SO4, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate / MeOH = 0:1:0 → 0:1:1) to obtain 428 mg of 3-(cyclohexylmethyl)thiazole-2(3H)-imine B6 as a yellow solid.

[0065] <Preparation of Intermediate B7> [ka] <5-(chloromethyl)-1-methyl-1H-pyrazole> (1-methyl-1H-pyrazole-5-yl)methanol (200 mg, 1.78 mmol, 1.0 equivalent) was added to a solution in DCM (4 mL) and stirred. SOCl2 (0.32 mL) was added dropwise at 25°C. The reaction mixture was then stirred under N2 at 25°C for 2 hours. The reaction mixture was diluted with water (20 mL) and adjusted to pH 8 with saturated NaHCO3 aqueous solution. The solution was extracted with ELISA (3 × 20 mL), the organic layers were washed together with brine (3 × 10 mL), dehydrated with Na2SO4, and filtered. The filtrate was concentrated under reduced pressure to obtain 200 mg of crude 5-(chloromethyl)-1-methyl-1H-pyrazole as a yellow oil. 1 H NMR: (400MHz, CHLOROFORM-d) δ 7.41 (d, J = 1.8 Hz, 1H), 6.28 (d, J = 1.8 Hz, 1H), 4.62 (s, 2H), 3.93 (s, 3H).

[0066] <Intermediate B7> To a solution of 5-(chloromethyl)-1-methyl-1H-pyrazole (120 mg, 919 μmol, 1.0 equivalent) in i-PrOH (1 mL), 2-thiazoleamine (74 mg, 735 μmol, 0.8 equivalents) was added at room temperature, and the mixture was stirred at 60°C for 12 hours under N2. The reaction mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate / MeOH = 1:1:0 → 0:1:0 → 0:1:9) to obtain 76 mg of 3-((1-methyl-1H-pyrazole-5-yl)methyl)thiazole-2(3H)-imine B7 as a yellow solid.

[0067] <Preparation of Intermediate B8> [ka] To a solution of ethyl 2-aminothiazole-4-carboxylate (2.00 g, 11.6 mmol, 1.0 equivalent) in i-PrOH (20 mL), (bromomethyl)benzene (1.38 mL, 11.6 mmol, 1.0 equivalent) was added at 25°C. The mixture was heated to 80°C and stirred for 6 hours. The reaction mixture was cooled to room temperature and left for 3 days. The precipitate was collected by filtration, and the filtered cake was dried under high vacuum to obtain 0.80 g of ethyl 3-benzyl-2-imino-2,3-dihydrothiazole-4-carboxylate B8 as a white solid.

[0068] <Preparation of Intermediate B9> [ka] To a solution of 5-methyl-2-thiazoleamine (1.00 g, 8.76 mmol, 1.0 equivalent) in acetone (15 mL), bromomethylbenzene (1.65 g, 9.63 mmol, 1.1 equivalent) was added at 25°C. The reaction mixture was heated to 70°C and stirred for 5 hours. The reaction mixture was cooled to 25°C, and pH > 7.0 was adjusted by adding aqueous NaOH solution (2N). The reaction mixture was filtered, and the filtered cake was washed with acetone (5 mL) and MTBE (10 mL). It was dried under reduced pressure to obtain 1.1 g of crude 3-benzyl-5-methylthiazole-2(3H)-imine B9 as a brown solid. The structure was confirmed by NOE.

[0069] <Preparation of Intermediate B10> [ka] A mixture of 2-thiazoleamine (100 mg, 998 μmol, 1.0 equivalent), 2-(bromomethyl)pyridine (278 mg, 1.10 mmol, 1.1 equivalent, HBr), and K2CO3 (152 mg, 1.10 mmol, 1.1 equivalent) in acetone (5 mL) was degassed and purged three times with N2. The reaction mixture was then stirred at 60°C for 5 hours under an N2 atmosphere. The resulting mixture was concentrated under reduced pressure. The residue was purified by preparative HPLC to obtain 100 mg of compound 3-(pyridine-2-ylmethyl)thiazole-2(3H)-imine B10 as a white solid.

[0070] <Preparation of Intermediate B11> [ka] 2-(bromomethyl)benzonitrile (587 mg, 3.00 mmol, 1.0 equivalent) was added to a solution of 2-thiazoleamine (0.30 g, 3.00 mmol, 1.0 equivalent) in i-PrOH (5 mL). The reaction mixture was stirred at 25°C for 12 hours, then heated to 60°C and stirred for a further 4 hours. The reaction mixture was directly concentrated under reduced pressure to obtain a residue, which was purified by silica gel flash column chromatography (ethyl acetate / MeOH) to obtain 0.41 g of 2-((2-iminothiazole-3(2H)-yl)methyl)benzonitrile B11 as a yellow solid.

[0071] <Preparation of Intermediate B12> [ka] 2-Bromoethiethylbenzene (3.70 g, 20.0 mmol, 1.0 equivalent) was dissolved in i-PrOH (35 mL), to which 2-thiazoleamine (2.00 g, 20.0 mmol, 1.0 equivalent) was added. The reaction mixture was stirred at 80°C for 12 hours. The reaction mixture was cooled to room temperature, then concentrated directly to obtain the residue, which was purified by reverse-phase column chromatography to obtain 2.00 g of compound 3-phenethylthiazole-2(3H)-imine B12 as a pale yellow solid.

[0072] <Preparation of Intermediate B13> [ka] <(Tetrahydro-2H-pyran-4-yl)methyl-4-methylbenzenesulfonate> To a solution of (tetrahydro-2H-pyran-4-yl)methanol (2.00 g, 17.2 mmol, 1.0 equivalent) in 2-methyltetrahydrofuran (20 mL), NaOH (4.13 g, 51.6 mmol, 50% purity, 3.0 equivalents) was added, followed by the dropwise addition of a solution of 4-methylbenzenesulfonyl chloride (5.91 g, 31.0 mmol, 1.8 equivalents) in 2-methyltetrahydrofuran (6 mL) at 0°C. The reaction mixture was then stirred at 25°C for 12 hours. The reaction mixture was diluted with HCl aqueous solution (6 M), and then 3.0 mL of cyclohexane was added. The solution was stirred at 0°C for 0.5 hours, but no solid precipitate formed. Next, the mixture was extracted with SiO2 (3 × 60 mL), the organic layers were washed together with brine (100 mL), dehydrated with Na2SO4, filtered, and the filtrate was concentrated under reduced pressure to obtain 4.2 g of crude (tetrahydro-2H-pyran-4-yl)methyl-4-methylbenzenesulfonate as a white solid. LCMS (m / z [M+H] + ): 271.1.

[0073] <Intermediate B13> To a solution of (tetrahydro-2H-pyran-4-yl)methyl-4-methylbenzenesulfonate (2.70 g, 9.99 mmol, 1.0 equivalent) and 2-thiazoleamine (1.00 g, 9.99 mmol, 1.0 equivalent) in DMF (10 mL), TBAI (1.84 g, 4.99 mmol, 0.5 equivalent) was added at room temperature. The reaction mixture was stirred at 80°C for 12 hours under N2. The reaction mixture was diluted with water (80 mL) and extracted with SiO (3 × 60 mL). The organic layer was discarded, and the aqueous phase was adjusted to pH=9 with NaOH (50% purity). This aqueous solution was extracted with SiO (3 × 50 mL), the organic layer was washed with brine (100 mL), dehydrated with Na₂SO₄, and filtered. The filtrate was concentrated under reduced pressure to obtain 500 mg of 3-((tetrahydro-2H-pyran-4-yl)methyl)thiazole-2(3H)-imine B13 as a yellow solid.

[0074] <Preparation of Intermediate B14> [ka] To a solution of 2-thiazoleamine (0.60 g, 5.99 mmol, 1.0 equivalent) in i-PrOH (10 mL), methyl 3-(bromomethyl)benzoate (1.37 g, 5.99 mmol, 1.0 equivalent) was added. The reaction mixture was stirred at 60°C for 12 hours. The reaction mixture was cooled to 25°C and diluted with MTBE. This suspension was decanted and dried under reduced pressure to obtain 1.5 g of the crude product methyl 3-((2-iminothiazole-3(2H)-yl)methyl)benzoate B14 as a brown solid.

[0075] <Preparation of Intermediate B15> [ka] A mixture of 2-thiazoleamine (100 mg, 1.00 mmol, 1.0 equivalent), 2-(bromomethyl)pyridine (278 mg, 1.10 mmol, 1.1 equivalent, HBr), and K2CO3 (152 mg, 1.10 mmol, 1.1 equivalent) in acetone (5 mL) was degassed and purged three times with N2. The reaction mixture was stirred at 60°C for 5 hours under an N2 atmosphere. The resulting mixture was concentrated under reduced pressure. The residue was purified by preparative HPLC to obtain 100 mg of 3-(pyridine-2-ylmethyl)thiazole-2(3H)-imine B15 as a white solid.

[0076] <Preparation of Intermediate B16> [ka] To a solution of 2-thiazoleamine (1.00 g, 9.99 mmol, 1.0 equivalent) in ethyl acetate (20 mL), 3-(bromomethyl)benzonitrile (1.96 g, 9.99 mmol, 1.0 equivalent) was added at 25°C. The reaction mixture was then stirred at 25°C for 12 hours. The reaction mixture was diluted with ethyl acetate (100 mL), and then NaOH (100 mL, 40%) was added at 0°C. The resulting mixture was stirred at 0°C for 1 hour. This solution was extracted with ethyl acetate (3 × 100 mL). The organic layers were washed together with brine (3 × 100 mL), dehydrated with Na₂SO₄, filtered, and concentrated under vacuum. The residue was purified by silica gel column chromatography (petroleum ether / Â=5:1 → Â:NH3·H2O(25%)=100:1) to obtain 500 mg of 3-((2-iminothiazole-3(2H)-yl)methyl)benzonitrile B16 as a yellow oil.

[0077] <Preparation of Intermediate B17> [ka] <4-(chloromethyl)-1-methyl-1H-pyrazole> (1-methyl-1H-pyrazole-4-yl)methanol (500 mg, 4.46 mmol, 1.0 equivalent) was added to a solution containing DCM (1.6 mL) and stirred. SOCl2 (2.39 g, 20.1 mmol, 1.46 mL, 4.5 equivalents) was added dropwise at 25°C. The reaction mixture was stirred under N2 at 25°C for 2 hours. The reaction mixture was concentrated under reduced pressure to obtain 500 mg of 4-(chloromethyl)-1-methyl-1H-pyrazole as a white solid. 1 H NMR (400MHz, DMSO-d6) δ 7.79 (s, 1H), 7.48 (s, 1H), 4.68 (s, 2H), 3.81 (s, 3H).

[0078] <Intermediate B17> To a solution of 4-(chloromethyl)-1-methyl-1H-pyrazole (450 mg, 3.45 mmol, 1.0 equivalent) in DMF (4.5 mL), 2-thiazoleamine (345 mg, 3.45 mmol, 1.0 equivalent) was added, followed by the addition of K2CO3 (953 mg, 6.89 mmol, 2.0 equivalent) at room temperature. The mixture was then stirred under N2 at 80°C for 12 hours. The reaction mixture was diluted with water (30 mL) and extracted with RINKAN (3 × 20 mL). The organic layer was washed with brine (30 mL), dehydrated with Na2SO4, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate / MeOH = 1:1:0 → 0:1:1) to obtain 50 mg of 3-((1-methyl-1H-pyrazole-4-yl)methyl)thiazole-2(3H)-imine B17 as a yellow solid.

[0079] <Preparation of Intermediate B18> [ka] To a solution of 2-oxazoleamine (0.50 g, 5.95 mmol, 1.0 equivalent) in acetone (10 mL), bromomethylbenzene (1.12 g, 6.54 mmol, 777 μL, 1.1 equivalent) was added, and the mixture was stirred at 60°C for 5 hours. The reaction mixture was concentrated under reduced pressure to obtain 0.70 g of 3-benzyloxazole-2(3H)-imine B18 as a yellow jelly-like oil.

[0080] <Preparation of Intermediate B19> [ka] To a solution of 4-methyl-2-oxazoleamine (0.400 g, 3.26 mmol, 1.0 equivalent) in i-PrOH (2 mL), (bromomethyl)benzene (0.387 mL, 3.26 mmol, 1.0 equivalent) was added dropwise at 20°C. The reaction mixture was heated to 80°C and stirred for 3 hours. The mixture was concentrated under reduced pressure, and the residue was purified by preparative HPLC to obtain 0.20 g of 3-benzyl-4-methyloxazole-2(3H)-imine 2,2,2-trifluoroacetate B19 as a yellow oil.

[0081] <Preparation of Intermediate B20> [ka] To a solution of ethyl 2-imino-3H-oxazole-4-carboxylate (1.35 g, 8.65 mmol) in isopropyl alcohol (14 mL), bromomethylbenzene (1.03 mL, 8.65 mmol) was added at 25 °C, and the mixture was stirred at 80 °C for 12 hours. The reaction mixture was cooled to room temperature, and the precipitate was collected by filtration. The filtered cake was dried under reduced pressure to obtain 1.20 g of ethyl 3-benzyl-2-imino-2,3-dihydroxazole-4-carboxylate B20 as a white solid.

[0082] <Preparation of Intermediate C1> [ka] <Ethyl N-(phenylcarbamate oil)carbamate> To a solution of aniline (2.00 g, 21.48 mmol, 1.0 equivalent) in SiO (20 mL), ethyl N-(thioxomethylene) carbamate (2.82 g, 21.48 mmol, 1.0 equivalent) and TMEDA (250 mg, 2.15 mmol, 0.1 equivalent) were added. The reaction mixture was stirred at 25°C for 5 hours. The reaction mixture was directly concentrated under reduced pressure to obtain a yellow solid. The solid was redissolved in EtOH (10 mL), stirred for 0.5 hours, and then filtered. The filtered cake was washed with EtOH (2 × 5 mL) and dried under vacuum to obtain 3.70 g of ethyl N-(phenylcarbamotioil) carbamate as a white solid.

[0083] <(Z)-Ethyl(((2-oxoethyl)thio)(phenylamino)methylene)carbamate> To a solution of ethyl N-(phenylcarbamotioil)carbamate (2.70 g, 12.0 mmol, 1.0 equivalent) and Cs2CO3 (6.67 g, 20.5 mmol, 1.7 equivalents) in CH3CN (40 mL), 2-chloroacetaldehyde (2.95 g, 15.0 mmol, 1.3 equivalents) was added while maintaining the temperature below 25°C. After the addition, the reaction mixture was stirred at 25°C for 12 hours. The reaction mixture was diluted with MTBE (80 mL) at 20°C, washed with saturated NaHCO3 aqueous solution (75 mL) and brine (75 mL), dehydrated with Na2SO4, and filtered. The filtrate was concentrated under reduced pressure to obtain 0.32 g of the crude product (Z)-ethyl(((2-oxoethyl)thio)(phenylamino)methylene)carbamate as a brown oil.

[0084] <(Z)-Ethyl(3-phenylthiazole-2(3H)-ylidene)carbamate> To a solution of ethyl(Z)-ethyl(((2-oxoethyl)thio)(phenylamino)methylene)carbamate (3.20 g, 12.0 mmol, 1.0 equivalent) in THF (40 mL), DIPEA (4.66 g, 36.0 mmol, 3.0 equivalent) and SOCl2 (1.43 g, 12.0 mmol, 1.0 equivalent) were added at 0°C. The reaction mixture was stirred at below 15°C for 4 hours. The reaction mixture was concentrated under reduced pressure. The residue was redissolved in MTBE (100 mL), and the pH was adjusted to 8.0 by adding saturated NaHCO3 aqueous solution. The organic layer was washed with brine, dehydrated with Na2SO4, and concentrated under vacuum. The residue was purified by flash silica gel chromatography (ethyl acetate / petroleum ether) to obtain 1.40 g of (Z)-ethyl(3-phenylthiazole-2(3H)-ylidene)carbamate as a yellow solid.

[0085] <Intermediate C1> A solution of (Z)-ethyl(3-phenylthiazole-2(3H)-ylidene) carbamate (0.80 g, 3.22 mmol, 1.0 equivalent) in EtOH (10 mL) was mixed with 6N NaOH (10 mL, 60.0 mmol, 19 equivalents). The reaction mixture was stirred at 90°C for 4 hours. The reaction mixture was diluted with water (25 mL) and extracted with ELISA (2 × 25 mL). The organic layers were combined and dehydrated with Na₂SO₄, and concentrated to obtain 0.60 g of crude 3-phenylthiazole-2-imine C1 as a brown solid. This crude product was purified by reverse-phase MPLC to obtain 0.23 g of pure 3-phenylthiazole-2(3H)-imine C1 as a brown oil.

[0086] <Preparation of intermediate C2> [ka] <Ethyl N-[(2-bromo-4-pyridyl)carbamate oil]carbamate> A mixture of 2-bromopyridine-4-amine (5.00 g, 28.9 mmol, 1.0 equivalent), ethyl N-(thioxomethylene)carbamate (3.79 g, 28.9 mmol, 3.41 mL, 1.0 equivalent), and TMEDA (336 mg, 2.89 mmol, 436 μL, 0.1 equivalent) in SiO2 (50 mL) was degassed, purged three times with N2 at room temperature, and then the reaction mixture was stirred at 25°C under an N2 atmosphere for 12 hours. The mixture was directly concentrated under reduced pressure to obtain a yellow solid. The solid was redissolved in EtOH (30 mL), stirred for 10 minutes, and then filtered. The filtered cake was washed with EtOH (2 × 5 mL) and then concentrated under reduced pressure to obtain 4.2 g of ethyl N-[(2-bromo-4-pyridyl)carbamotioil]carbamate as a pale yellow solid.

[0087] <(Z)-Ethyl(((2-bromopyridine-4-yl)amino)((2-oxoethyl)thio)methylene)carbamate> To a solution of ethyl N-[(2-bromo-4-pyridyl)carbamotioyl]carbamate (1.50 g, 4.93 mmol, 1.0 equivalent) and Cs2CO3 (2.73 g, 8.38 mmol, 1.7 equivalents) in MeCN (30 mL), 2-chloroacetaldehyde (484 mg, 6.16 mmol, 397 L, 1.2 equivalents) was added at room temperature. After addition, the reaction mixture was stirred at 25 °C for 12 hours. The mixture was diluted with water (50 mL) and extracted with ethyl acetate (3 × 30 mL). The organic phases were combined and washed with brine (30 mL), dehydrated with anhydrous Na2SO4, filtered, and concentrated under vacuum. (Z)-ethyl(((2-bromopyridine-4-yl)amino)((2-oxoethyl)thio)methylene)carbamate (1.60 g, crude) was obtained as a brown oil.

[0088] <(Z)-Ethyl(3-(2-bromopyridine-4-yl)thiazole-2(3H)-ylidene)carbamate> (Z)-ethyl(((2-bromopyridine-4-yl)amino)((2-oxoethyl)thio)methylene)carbamate (1.60 g, 4.62 mmol, 1.0 equivalent) was dissolved in THF (20 mL) and DIPEA (1.79 g, 13.9 mmol, 2.42 mL, 3.0 equivalent) and SOCl2 (550 mg, 4.62 mmol, 335 L, 1.0 equivalent) were added at 0°C. The reaction mixture was stirred at below 20°C for 12 hours. The mixture was quenched with saturated NaHCO3 aqueous solution (50 mL) and extracted with dichloromethane (3 × 20 mL). The organic layers were washed together with brine, dehydrated with Na2SO4, and concentrated to obtain 1.40 g of (Z)-ethyl(3-(2-bromopyridine-4-yl)thiazole-2(3H)-ylidene)carbamate as a brown oil.

[0089] <(Z)-Ethyl(3-(2-((tert-butoxycarbonyl)amino)pyridine-4-yl)thiazole-2(3H)-ylidene)carbamate> A mixture of (Z)-ethyl(3-(2-bromopyridine-4-yl)thiazole-2(3H)-ylidene) carbamate (700 mg, 2.13 mmol, 1.0 equivalent), NH2Boc (750 mg, 6.40 mmol, 3.0 equivalent), K3PO4 (1.81 g, 8.53 mmol, 4.0 equivalent), Pd2(dba)3 (195 mg, 213 μmol, 0.1 equivalent), and xanthophos (123 mg, 213 □ mol, 0.1 equivalent) in dioxane (10 mL) was degassed, purged three times with N2, and then the reaction mixture was stirred at 110°C under an N2 atmosphere for 12 hours. The mixture was poured into water (30 mL) and extracted with ethyl acetate (3 × 20 mL). The organic layers were washed together with brine (30 mL), dehydrated with anhydrous Na2SO4, filtered, and concentrated under vacuum. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 50 / 1 → 1 / 1) to obtain 140 mg of (Z)-ethyl(3-(2-((tert-butoxycarbonyl)amino)pyridine-4-yl)thiazole-2(3H-ylidene)carbamate as a yellow solid. 1H NMR (400 MHz, METHANOL-d4) δ 8.37 (d, J= 5.50 Hz, 1H), 8.09 (s, 1H), 7.43 (d, J= 4.75 Hz, 1H), 7.29 (dd, J = 5.50, 1.75 Hz, 1H), 7.01 (d, J = 4.88 Hz, 1H), 4.18 (q, J = 7.13 Hz, 2H), 1.54 (s, 9H), 1.26 (t, J = 7.07 Hz, 3H).

[0090] <Intermediate C2> (Z)-ethyl(3-(2-((tert-butoxycarbonyl)amino)pyridine-4-yl)thiazole-2(3H)-ylidene) carbamate (110 mg, 302 μmol, 1.0 equivalent) was dissolved in EtOH (4 mL) and NaOH (6 M, 4.0 mL, 80 equivalents) was added. The reaction mixture was stirred at 60°C for 2 hours. The mixture was diluted with water (30 mL) and extracted with THF (2 × 20 mL). The organic layers were washed together with brine (30 mL), dehydrated with Na2SO4, filtered, and concentrated under reduced pressure to obtain intermediate C2, tert-butyl(4-(2-iminothiazole-3(2H)-yl)pyridine-2-yl) carbamate (100 mg, crude), as a brown solid.

[0091] <Preparation of intermediate C3> [ka] <Ethyl N-(o-tolylcarbamoteoyl)carbamate> To a solution of o-toluidine (2.00 g, 18.7 mmol, 1.0 equivalent) in ethyl acetate (20 mL), TMEDA (0.217 g, 1.87 mmol, 0.1 equivalent) and O-ethylcarbon isothiocyanate (2.45 g, 18.7 mmol, 1.0 equivalent) were added at 25°C. The mixture was stirred at 25°C for 2 hours and then evaporated under reduced pressure. The residue was triturated in ethanol (10 mL) to obtain 3.90 g of ethyl N-(o-tolylcarbamotioil)carbamate as a white solid. 1H NMR (400 MHz, CDCl3) δ 11.18 (br. s, 1H), 8.36 (br. s, 1H), 7.71 - 7.58 (m, 1H), 7.38 - 7.20 (m, 3H), 4.31 (q, J = 7.2 Hz, 2H), 2.34 (s, 3H), 1.37 (t, J = 7.2 Hz, 3H). LCMS (m / z [M+H] + ):239.1.

[0092] <(Z)-Ethyl(((2-oxoethyl)thio)(o-tolylamino)methylene)carbamate> Ethyl N-(o-tolylcarbamotioil)carbamate (3.90 g, 16.3 mmol, 1.0 equivalent) and Cs2CO3 (9.05 g, 27.8 mmol, 1.7 equivalents) were dissolved in acetonitrile (40 mL), to which 2-chloroacetaldehyde (3.85 g, 19.6 mmol, 40% aqueous solution, 1.2 equivalents) was added while maintaining the temperature below 25°C. After addition, the mixture was stirred at 25°C for 12 hours, then diluted with MTBE (100 mL), washed with saturated NaHCO3 aqueous solution (100 mL), followed by brine (50 mL), dehydrated with Na2SO4, and filtered. The filtrate was concentrated under reduced pressure to obtain 5.80 g of crude (Z)-ethyl(((2-oxoethyl)thio)(o-tolylamino)methylene)carbamate as a brown, rubbery substance. 1 H NMR (400 MHz, CDCl3) δ 7.40 - 7.27 (m, 5H), 5.39 (d, J= 5.2 Hz, 1H), 4.16 (q, J = 7.2 Hz, 2H), 3.65 (dd, J = 6.0, 12.4 Hz, 1H), 3.28 (d, J = 12.4 Hz, 1H), 2.20 (s, 3H), 1.26 (t, J= 7.2 Hz, 3H). LCMS (m / z [M+H] + ): 281.1.

[0093] <(Z)-Ethyl(3-(o-tolyl)thiazole-2(3H)-ylidene)carbamate (Z)-ethyl(((2-oxoethyl)thio)(o-tolylamino)methylene)carbamate (5.80 g, 12.1 mmol, 1.0 equivalent) and DIEA (6.32 mL, 36.3 mmol, 3.0 equivalent) were dissolved in THF (60 mL), to which SOCl2 (0.878 mL, 12.1 mmol, 1.0 equivalent) was added dropwise at 0°C. The mixture was stirred at 0°C for 4 hours, then quenched with water (100 mL), and extracted with ethyl acetate (30 mL x 3). The organic layers were combined and dehydrated with sodium sulfate, filtered, and concentrated. The residue was purified by silica gel column chromatography (petroleum ether:ethyl acetate = 10:1 → 3:1) to obtain 2.70 g of (Z)-ethyl(3-(o-tolyl)thiazole-2(3H)-ylidene)carbamate as a yellow oil. 1 H NMR (400 MHz, CDCl3) δ 7.41 - 7.28 (m, 3H), 7.25 - 7.19 (m, 1H), 6.85 (d, J = 4.8 Hz, 1H), 6.69 (d, J = 4.8 Hz, 1H), 4.21 (q, J = 7.2 Hz, 2H), 2.14 (s, 3H), 1.29 (t, J = 7.2 Hz, 3H). LCMS (m / z[M+H] + ): 263.1.

[0094] <Intermediate C3> A solution of (Z)-ethyl(3-(o-tolyl)thiazole-2(3H)-ylidene) carbamate (0.500 g, 1.79 mmol, 1.0 equivalent) and NaOH (1.43 g, 35.8 mmol, 20 equivalents) in ethanol (6 mL) was stirred at 50°C for 1 hour. The mixture was cooled to room temperature, diluted with ethyl acetate (20 mL), washed with water (20 mL), followed by brine (20 mL), dehydrated with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (dichloromethane:methanol = 20:1) to obtain 0.23 g of 3-(o-tolyl)thiazole-2(3H)-imine C3 as a yellow oil.

[0095] <Preparation of intermediate C4> [ka] <Ethyl N-[(2-methoxyphenyl)carbamate]carbamate> To a solution of 2-methoxyaniline (2.00 g, 16.2 mmol, 1.0 equivalent) in ethyl acetate (20 mL), 1.92 mL, 16.2 mmol, 1.0 equivalent and tetramethylethylenediamine (0.245 mL, 1.62 mmol, 0.1 equivalent) were added at 25°C. The mixture was stirred at this temperature for 5 hours. The mixture was concentrated under reduced pressure to remove the ethyl acetate. The residue was triturated twice with ethanol (5 mL) to obtain 3.40 g of ethyl N-[(2-methoxyphenyl)carbamoteoyl]carbamate as an off-white solid. 1 H NMR (400 MHz, DMSO-d6) δ 11.90 (br. s, 1H), 11.25 (br. s, 1H), 8.48 (dd, J = 8.0, 1.2 Hz, 1H), 7.19 (dt, J = 8.4, 1.6 Hz, 1H), 7.10 (dd, J = 8.0, 1.2 Hz, 1H), 6.96 (dt, J = 8.0, 1.2 Hz, 1H), 4.22 (q, J = 7.2 Hz, 2H), 3.85 (s, 3H), 1.25 (t, J = 7.2 Hz, 3H).

[0096] <(Z)-Ethyl(((2-Methoxyphenyl)amino)((2-Oxoethyl)thio)methylene)carbamate> To a suspension of ethyl N-[(2-methoxyphenyl)carbamate]carbamate (3.40 g, 13.4 mmol, 1.0 equivalent) and cesium carbonate (7.41 g, 22.7 mmol, 1.7 equivalents) in acetonitrile (40 mL), 2-chloroacetaldehyde (2.69 mL, 16.7 mmol, 1.2 equivalents) was added dropwise at 25°C. The resulting mixture was stirred at this temperature for 12 hours. Additional 2-chloroacetaldehyde (1.08 mL, 6.68 mmol, 0.5 equivalents) was added dropwise to the mixture at 25°C, and the mixture was stirred for a further 2 hours. The mixture was poured into water (150 mL), extracted with MTBE (100 mL x 3), the organic layers were washed together with brine (75 mL), dehydrated with anhydrous sodium sulfate, and filtered. The filtrate was concentrated under reduced pressure to obtain 4.60 g of (Z)-ethyl(((2-methoxyphenyl)amino)((2-oxoethyl)thio)methylene)carbamate as a brown oil. 1 H NMR (400 MHz, DMSO-d6): δ 7.37 (dt, J = 8.4, 1.6 Hz, 1H), 7.19 (dd, J= 7.6, 1.6 Hz, 1H), 7.13 (dd, J = 8.4, 0.8 Hz, 1H), 7.00 (dt, J = 7.6, 1.2 Hz, 1H), 6.75 (d, J = 7.2 Hz, 1H), 5.46 (dt, J =7.6, 2.4 Hz, 1H), 3.93 (q, J = 7.2 Hz, 2H), 3.77 (s, 3H), 3.59 (dd, J = 12.4, 6.4Hz, 1H), 1.12 - 1.10 (m, 3H). LCMS (m / z [M+H] + ): 296.8.

[0097] <(Z)-Ethyl(3-(2-methoxyphenyl)thiazole-2(3H)-ylidene)carbamate> (Z)-ethyl(((2-methoxyphenyl)amino)((2-oxoethyl)thio)methylene)carbamate (2.60 g, 6.10 mmol, 1.0 equivalent) was dissolved in THF (20 mL) and then diisopropylethylamine (3.19 mL, 18.3 mmol, 3.0 equivalent), followed by SOCl2 (0.442 mL, 6.10 mmol, 1.0 equivalent) was added at 0°C. The mixture was stirred at below 15°C for 4 hours. The mixture was added to saturated sodium bicarbonate aqueous solution (20 mL), stirred for 5 minutes, and extracted with ethyl acetate (100 mL x 3). The organic layers were washed together with brine (50 mL), dehydrated with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 10 / 1 → 5 / 1) to obtain 1.00 g of (Z)-ethyl(3-(2-methoxyphenyl)thiazole-2(3H)-ylidene)carbamate as an off-white solid. 1 H NMR (400 MHz, DMSO-d6): δ 7.51 - 7.47 (m, 1H), 7.35 (dd, J = 6.8, 1.6 Hz, 1H), 7.32 (d, J= 4.8 Hz, 1H), 7.25 (dd, J = 8.4, 0.8 Hz, 1H), 7.10 - 7.06 (m, 1H), 6.99 (d, J= 4.8 Hz, 1H), 3.98 (q, J = 7.2 Hz, 2H), 3.76 (s, 3H), 1.13 (t, J = 7.2 Hz, 3H). LCMS (m / z [M+H] + ): 278.8.

[0098] <Intermediate C4> To a solution of ethyl (3-(2-methoxyphenyl)thiazole-2(3H)-ylidene) carbamate (0.500 g, 1.80 mmol, 1.0 equivalent) in ethanol (5 mL), sodium hydroxide (1.44 g, 35.9 mmol, 20 equivalents) was added all at once at 25°C, and the mixture was stirred for 1 hour. The mixture was poured into water (20 mL) and extracted with ethyl acetate (50 mL x 3). The organic layers were washed together with brine (25 mL), dehydrated with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain 0.350 g of 3-(2-methoxyphenyl)thiazole-2(3H)-imine C4 as a brown oil.

[0099] <Preparation of intermediate C5> [ka] <Ethyl N-[(2-chlorophenyl)carbamate]carbamate> To a solution of 2-chloroaniline (2.20 g, 17.3 mmol, 1.0 equivalent) and O-ethylcarbon isothiocyanate (2.26 g, 17.3 mmol, 1.0 equivalent) in ethyl acetate (20 mL), tetramethylethylenediamine (0.200 g, 1.72 mmol, 0.1 equivalent) was added at 0°C. The resulting mixture was stirred at 25°C for 2 hours and then concentrated under reduced pressure. The residue was triturated with methanol (30 mL) to obtain 3.00 g of ethyl N-[(2-chlorophenyl)carbamoteoyl]carbamate as a white solid. 1 H NMR (400 MHz, DMSO-d6) δ 11.60 (s, 1H), 11.48 (s, 1H), 7.94 (dd, J = 8.0, 1.6 Hz, 1H), 7.56 (dd, J = 8.0, 1.6 Hz, 1H), 7.40 - 7.36 (m, 1H), 7.33 - 7.29 (m, 1H), 4.23 (q, J = 7.2 Hz, 2H), 1.26 (t, J =7.2 Hz, 3H).

[0100] <(Z)-Ethyl(((2-chlorophenyl)amino)((2-oxoethyl)thio)methylene)carbamate> Ethyl N-[(2-chlorophenyl)carbamate]carbamate (3.00 g, 11.6 mmol, 1.0 equivalent) and Cs2CO3 (6.42 g, 19.7 mmol, 12 equivalents) were suspended in acetonitrile (40 mL), to which 2-chloroacetaldehyde (2.84 g, 14.5 mmol, 1.2 equivalents) was added while maintaining the temperature below 25°C. The reaction mixture was stirred at 25°C for 12 hours. The mixture was poured into water (100 mL), extracted with methyl TBME (150 mL x 2), the organic layers were washed together with brine (200 mL), dehydrated with anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to obtain 3.80 g of crude (Z)-ethyl(((2-chlorophenyl)amino)((2-oxoethyl)thio)methylene)carbamate as a brown, rubbery substance. 1 H NMR (400 MHz, DMSO-d6) δ 7.66 - 7.58 (m, 1H), 7.49 - 7.35 (m, 3H), 6.99 (d, J = 6.4 Hz, 1H), 5.53- 5.42 (m, 1H), 4.00 - 3.90 (m, 2H), 3.70 - 3.65 (m, 1H), 3.23 - 3.15 (m, 1H), 1.11 (t, J = 7.2 Hz, 3H).

[0101] <(Z)-Ethyl(3-(2-chlorophenyl)thiazole-2(3H)-ylidene)carbamate> (Z)-ethyl(((2-chlorophenyl)amino)((2-oxoethyl)thio)methylene)carbamate (1.80 g, 5.98 mmol, 1.0 equivalent) was dissolved in THF (20 mL) and diisopropylethylamine (2.32 g, 17.9 mmol, 3.0 equivalent) and thionyl chloride (0.710 g, 5.98 mmol, 1.0 equivalent) were added at 0°C. The mixture was stirred at 15°C for 4 hours, then poured into water (100 mL) and extracted with ethyl acetate (150 mL x 4). The organic phases were combined and washed with brine (100 mL), dehydrated with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 1 / 0 → 10 / 1) to obtain 1.101 g of (Z)-ethyl(3-(2-chlorophenyl)thiazole-2(3H)-ylidene)carbamate as a yellow solid. 1 H NMR (400 MHz, DMSO-d6) δ 7.73 - 7.71 (m, 1H), 7.60 - 7.53 (m, 3H), 7.45 (d, J = 4.8 Hz, 1H), 7.09 (d, J= 4.8 Hz, 1H), 4.00 (q, J = 6.8 Hz, 2H), 1.14 (t, J = 7.2 Hz, 3H). LCMS (m / z [M+H] + ): 282.9.

[0102] <Intermediate C5> (Z)-ethyl(3-(2-chlorophenyl)thiazole-2(3H)-ylidene)carbamate (0.80 g, 2.75 mmol, 1.0 equivalent) was dissolved in ethanol (10 mL) and sodium hydroxide (2.34 g, 58.4 mmol, 21 equivalents) was added at 25 °C. The mixture was heated to 50 °C and stirred for 1 hour, then poured into water (80 mL) and extracted with ethyl acetate (120 mL x 4). The organic layers were washed together with brine (100 mL), dehydrated with anhydrous sodium sulfate, and filtered. The filtrate was concentrated under reduced pressure to obtain 0.54 g of 3-(2-chlorophenyl)thiazole-2(3H)-imine C5 as a yellow solid.

[0103] <Preparation of intermediate C6> [ka] <Ethyl N-(m-tolylcarbamoteoyl)carbamate> To a solution of m-toluidine (2.00 g, 18.6 mmol, 1.0 equivalent) in ethyl acetate (20 mL), o-ethylcarbon isothiocyanate (2.21 mL, 18.6 mmol, 1.0 equivalent) and tetramethylethylenediamine (0.282 mL, 1.87 mmol, 0.1 equivalent) were added. The mixture was stirred at 25°C for 5 hours. The mixture was concentrated under reduced pressure to remove the ethyl acetate. The residue was triturated twice with ethanol (5 mL) to obtain 3.00 g of ethyl N-(m-tolylcarbamotioil)carbamate as a white solid. 1 H NMR (400 MHz, DMSO-d6) δ 11.51 (br. s, 1H), 11.23 (br. s, 1H), 7.43 - 7.38 (m, 2H), 7.26 (t, J = 7.6 Hz, 1H), 7.05 (d, J = 7.6 Hz, 1H), 4.20 (q, J = 8.0, 7.2 Hz, 2H), 2.30 (s, 3H), 1.25 (t, J = 6.8 Hz, 3H).

[0104] <(Z)-Ethyl(((2-oxoethyl)thio)(m-tolylamino)methylene)carbamate> Ethyl N-(m-tolylcarbamotioil)carbamate (3.00 g, 12.6 mmol, 1.0 equivalent) and cesium carbonate (6.97 g, 21.4 mmol, 1.7 equivalents) were suspended in acetonitrile (40 mL). 2-chloroacetaldehyde (2.53 mL, 15.7 mmol, 1.25 equivalents) was added dropwise at 25°C, and the mixture was stirred for 12 hours. The mixture was poured into water (150 mL), extracted with methyl tert-butyl ether (100 mL x 3), and the organic layers were washed together with brine (75 mL), dehydrated with sodium sulfate, and filtered. The filtrate was concentrated under reduced pressure to obtain 4.00 g of (Z)-ethyl(((2-oxoethyl)thio)(m-tolylamino)methylene)carbamate as a brown oil. 1 H NMR (400 MHz, DMSO-d6) δ 7.34 - 7.30 (m, 1H), 7.16 - 7.13 (m, 3H), 6.85 (d, J = 7.2 Hz, 1H), 5.60 (dt, J=7.2, 1.6 Hz, 1H), 3.96 (q, J = 7.2 Hz, 2H), 2.32 (s, 3H), 1.14 - 1.10 (m, 3H). LCMS (m / z [M+H] + ): 280.9.

[0105] <(Z)-Ethyl(3-(m-tolyl)thiazole-2(3H)-ylidene)carbamate> (Z)-ethyl(((2-oxoethyl)thio)(m-tolylamino)methylene)carbamate (2.00 g, 5.58 mmol, 1.0 equivalent) was dissolved in THF (20 mL), to which diisopropylethylamine (2.92 mL, 16.8 mmol, 3.0 equivalent), followed by SOCl2 (0.405 mL, 5.58 mmol, 1.0 equivalent), was added at 0°C under a nitrogen atmosphere. The mixture was stirred at below 15°C for 4 hours. The mixture was poured into saturated sodium bicarbonate aqueous solution to adjust the pH to 8, and then extracted with ethyl acetate (100 mL x 3). The organic layers were washed together with brine (100 mL), dehydrated with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 10 / 1) to obtain 1.00 g of (Z)-ethyl(3-(m-tolyl)thiazole-2(3H)-ylidene)carbamate as a yellow oil. 1 H NMR (400 MHz, DMSO-d6) δ 7.49 (d, J = 4.8 Hz, 1H), 7.42 (t, J = 8.0 Hz,1H), 7.30 - 7.28 (m, 3H), 7.05 (d, J = 4.8 Hz, 1H), 4.04 - 3.99 (m, 2H), 2.37 (s, 3H), 1.15 (t, J= 7.2 Hz, 3H). LCMS (m / z [M+H] + ): 262.8.

[0106] <Intermediate C6> (Z)-ethyl(3-(m-tolyl)thiazole-2(3H)-ylidene)carbamate (0.200 g, 0.762 mmol, 1.0 equivalent) was dissolved in ethanol (2 mL) and sodium hydroxide (0.610 g, 15.2 mmol, 20 equivalents) was added all at once at 25 °C. The mixture was then heated to 50 °C and stirred for 1 hour. The mixture was placed in water (50 mL) and extracted with ethyl acetate (50 mL x 4). The organic layers were washed together with brine (20 mL), dehydrated with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain 140 mg of 3-(m-tolyl)thiazole-2(3H)-imine C6 as a brown oil.

[0107] <Preparation of intermediate C7> [ka] <Ethyl N-[(3-methoxyphenyl)carbamate oil]carbamate> To a solution of 3-methoxyaniline (2.00 g, 16.2 mmol, 1.0 equivalent) in ethyl acetate (10 mL), TMEDA (0.189 g, 1.62 mmol, 0.1 equivalent) and O-ethylcarbon isothiocyanate (2.13 g, 16.2 mmol, 1.0 equivalent) were added at 25°C. The mixture was stirred at 25°C for 2 hours and then evaporated under reduced pressure. The residue was crystallized from petroleum ether / ethyl acetate (v / v=5:1, 10 mL) to obtain 3.70 g of ethyl N-[(3-methoxyphenyl)carbamoteoyl]carbamate as a yellow solid. 1 H NMR (400 MHz, CDCl3) δ 11.48 (br. s, 1H), 8.11 (br. s, 1H), 7.41 (t, J = 2.0 Hz, 1H), 7.30 (t, J = 8.0 Hz, 1H), 7.14 (dd, J = 1.6, 8.0 Hz, 1H), 6.82 (ddd, J = 0.8, 2.4, 8.4 Hz, 1H), 4.30 (q, J = 7.2 Hz, 2H), 3.83 (s, 3H), 1.36 (t, J = 7.2 Hz, 3H). LCMS (m / z [M+H] + ): 255.1.

[0108] <(Z)-Ethyl(((3-Methoxyphenyl)amino)((2-Oxoethyl)thio)methylene)carbamate> To a solution of ethyl N-[(3-methoxyphenyl)carbamate]carbamate (3.70 g, 14.5 mmol, 1.0 equivalent) and Cs2CO3 (8.01 g, 24.6 mmol, 1.7 equivalents) in acetonitrile (40 mL), 2-chloroacetaldehyde (3.40 g, 17.3 mmol, 40% aqueous solution, 1.2 equivalents) was added while maintaining the temperature below 25°C. The mixture was stirred at 25°C for 12 hours, then diluted with MTBE (100 mL), washed with saturated NaHCO3 aqueous solution (100 mL), followed by brine (50 mL), dehydrated with Na2SO4, and filtered. The filtrate was concentrated under reduced pressure to obtain 5.00 g of (Z)-ethyl(((3-methoxyphenyl)amino)((2-oxoethyl)thio)methylene)carbamate as a yellow oil. 1 H NMR (400 MHz, CDCl3) δ 7.33 (t, J = 8.0 Hz, 1H), 7.04 - 6.95 (m, 2H), 6.87 (ddd, J = 0.8, 2.4, 8.4 Hz, 1H), 5.61 (d, J = 5.2 Hz, 1H), 4.18 (q, J= 7.2 Hz, 2H), 3.81 (s, 3H), 3.60 (dd, J= 5.6, 12.0 Hz, 1H), 3.22 (dd, J = 1.2, 12.4 Hz, 1H), 1.28 (t, J = 7.2 Hz, 3H). 297.1 [M+H] + ): 297.1.

[0109] <(Z)-Ethyl(3-(3-methoxyphenyl)thiazole-2(3H)-ylidene)carbamate> (Z)-ethyl(((3-methoxyphenyl)amino)((2-oxoethyl)thio)methylene)carbamate (5.00 g, 11.3 mmol, 1.0 equivalent) and DIEA (4.37 g, 33.8 mmol, 3.0 equivalent) were dissolved in THF (60 mL), to which SOCl2 (0.818 mL, 11.3 mmol, 1.0 equivalent) was added dropwise at 0°C. The mixture was stirred at 0°C for 4 hours, then quenched with water (100 mL), and extracted with ethyl acetate (30 mL x 3). The organic layers were combined and dehydrated with sodium sulfate, filtered, and concentrated. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 10:1 → 3:1) to obtain 2.70 g of (Z)-ethyl(3-(3-methoxyphenyl)thiazole-2(3H)-ylidene)carbamate as a yellow oil. 1 H NMR (400 MHz, CDCl3) δ 7.38 (t, J = 8.0 Hz, 1H), 7.07 - 6.98 (m, 3H), 6.94 (dd, J= 2.0, 8.0 Hz, 1H), 6.66 (d, J = 4.8 Hz, 1H), 4.22 (q, J = 7.2 Hz, 2H), 3.83 (s, 3H), 1.30 (t, J = 7.2 Hz, 3H). LCMS (m / z [M+H] + ): 279.1.

[0110] <Intermediate C7> A solution of (Z)-ethyl(3-(3-methoxyphenyl)thiazole-2(3H)-ylidene) carbamate (0.500 g, 1.70 mmol, 1.0 equivalent) and NaOH (1.36 g, 34.1 mmol, 20 equivalents) in ethanol (6 mL) was stirred at 50°C for 1 hour. The mixture was cooled to room temperature, diluted with ethyl acetate (20 mL), washed with water (20 mL), followed by brine (20 mL), dehydrated with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (dichloromethane:methanol = 20:1) to obtain 230 mg of 3-(3-methoxyphenyl)thiazole-2(3H)-imine C7 as a yellow oil.

[0111] <Preparation of Intermediate C8>

Chem.

[0112] <(Z)-Ethyl (((3-chlorophenyl)amino)((2-oxoethyl)thio)methylene)carbamate> Ethyl N-[(3-chlorophenyl)carbamate]carbamate (3.20 g, 12.4 mmol, 1.0 equivalent) and cesium carbonate (6.85 g, 21.0 mmol, 1.7 equivalents) were suspended in acetonitrile (40 mL), to which 2-chloroacetaldehyde (2.49 mL, 15.4 mmol, 1.25 equivalents) was added dropwise at 25°C. The resulting mixture was stirred at this temperature for 12 hours. The mixture was poured into water (150 mL) and extracted at 25°C using MTBE (100 mL x 3). The organic layers were washed together with brine (75 mL), dehydrated with anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to obtain 3.80 g of (Z)-ethyl(((3-chlorophenyl)amino)((2-oxoethyl)thio)methylene)carbamate as a brown oil. 1 H NMR (400 MHz, DMSO-d6): δ 7.50 - 7.46 (m, 2H), 7.43 - 7.40 (m, 1H), 7.38 - 7.36 (m, 1H), 6.95 (br. s, 1H), 5.69 - 5.68 (m, 1H), 3.98 (t, J= 7.6Hz, 2H), 3.68 - 3.55 (m, 1 H), 3.10 - 3.05 (m, 1 H), 1.14 (t, J = 7.6Hz, 3H). LCMS (m / z [M+H] + ): 300.8.

[0113] <(Z)-Ethyl(3-(3-chlorophenyl)thiazole-2(3H)-ylidene)carbamate> To a solution of (Z)-ethyl(((3-chlorophenyl)amino)((2-oxoethyl)thio)methylene) carbamate (1.80 g, 3.68 mmol, 1.0 equivalent) in THF (20 mL), diisopropylethylamine (1.92 mL, 11.0 mmol, 3.0 equivalent), followed by SOCl2 (0.267 mL, 3.68 mmol, 1.0 equivalent), was added at 0°C under a nitrogen atmosphere. The mixture was stirred at below 15°C for 4 hours. SOCl2 (0.134 mL, 1.84 mmol, 0.5 equivalent) was added dropwise to the mixture at 0°C, and the reaction mixture was stirred at 15°C for 2 hours. The mixture was added to saturated sodium bicarbonate aqueous solution to adjust the pH to 8, and extracted with ethyl acetate (100 mL x 3). The organic layers were washed together with brine (100 mL), dehydrated with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 10 / 1 → 5 / 1) to obtain 0.95 g of (Z)-ethyl(3-(3-chlorophenyl)thiazole-2(3H)-ylidene) carbamate as an off-white solid. 1 H NMR (400 MHz, DMSO-d6): δ 7.70 (s, 1H), 7.58 - 7.54 (m, 3H), 7.53 - 7.51 (m, 1H), 7.07 (d, J = 4.8 Hz, 1H), 4.03 (q, J = 7.2 Hz, 2H), 1.16 (t, J = 7.2 Hz, 3H). LCMS (m / z [M+H] + ): 282.8.

[0114] <Intermediate C8> (Z)-ethyl(3-(3-chlorophenyl)thiazole-2(3H)-ylidene)carbamate (0.200 g, 0.707 mmol, 1.0 equivalent) was dissolved in ethanol (2 mL) and sodium hydroxide (0.566 g, 14.2 mmol, 20 equivalents) was added all at once at 25°C. The mixture was then stirred at this temperature for 1 hour. The mixture was added to water (50 mL) and extracted with ethyl acetate (50 mL x 4). The organic layers were washed together with brine (20 mL), dehydrated with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain 130 mg of 3-(3-chlorophenyl)thiazole-2(3H)-imine C8 as a brown solid.

[0115] <Preparation of intermediate C9> [ka] To a solution of p-toluidine (2.00 g, 18.7 mmol, 1.0 equivalent) in ethyl acetate (10 mL), TMEDA (0.217 g, 1.87 mmol, 0.1 equivalent) and O-ethylcarbon isothiocyanate (2.45 g, 18.7 mmol, 1.0 equivalent) were added at 25°C. The mixture was stirred at 25°C for 2 hours and then evaporated under reduced pressure. The residue was triturated with ethanol (10 mL) to obtain 3.70 g of ethyl N-(p-tolylcarbamotioil)carbamate as a yellow solid. 1 H NMR (400 MHz, CDCl3) δ 11.35 (br s, 1H), 8.21 (br s, 1H), 7.48 (d, J = 8.4 Hz, 2H), 7.21 (d, J = 8.0 Hz, 2H), 4.29 (q, J = 7.2 Hz, 2H), 2.37 (s, 3H), 1.35 (t, J = 7.2 Hz, 3H). LCMS (m / z [M+H] + ): 239.1.

[0116] <(Z)-Ethyl(((2-oxoethyl)thio)(p-tolylamino)methylene)carbamate> A solution of ethyl N-(p-tolylcarbamothioyl)carbamate (3.70 g, 15.5 mmol, 10 eq) and Cs2CO3 (8.59 g, 26.4 mmol, 1.7 eq) in acetonitrile (40 mL) was added 2-chloroacetaldehyde (3.65 g, 18.6 mmol, 40% aqueous solution, 1.2 eq) while maintaining the temperature below 25 °C. After the addition, the mixture was stirred at 25 °C for 12 h, then diluted with MTBE (100 mL), washed with saturated aqueous NaHCO3 (100 mL), followed by brine (50 mL), and dried over Na2SO4 and filtered. The filtrate was concentrated under reduced pressure to give 5.30 g of crude (Z)-ethyl (((2-oxoethyl)thio)(p-tolylamino)methylene)carbamate as a yellow oil. 1 H NMR (400 MHz, CDCl3) δ 7.39 - 7.18 (m, 5H), 5.56 (dd, J = 0.5, 5.6 Hz, 1H), 4.16 (q, J = 7.2 Hz, 2H), 3.57 (dd, J = 5.6, 12.4 Hz, 1H), 3.19 (dd, J = 1., 12.0 Hz, 1H), 2.35 (s, 3H), 1.26(t, J = 7.2 Hz, 3H). LCMS (m / z [M+H] + ): 281.1.

[0117] <(Z)-ethyl (3-(p-tolyl)thiazol-2(3H)-ylidene)carbamate> It should be noted that there is a small error in the original text where "J = 1." in the NMR data should probably be "J = 1.2". This has been corrected in the translation for the sake of correct chemical data representation.(Z)-ethyl(((2-oxoethyl)thio)(p-tolylamino)methylene)carbamate (5.30 g, 10.8 mmol, 1.0 equivalent) and DIEA (4.19 g, 32.4 mmol, 3.0 equivalent) were dissolved in THF (60 mL), to which SOCl2 (0.783 mL, 10.8 mmol, 1.0 equivalent) was added dropwise at 0°C. The mixture was stirred at 0°C for 4 hours, then quenched with water (100 mL), and extracted with ethyl acetate (30 mL x 3). The organic layers were combined and dehydrated with sodium sulfate, filtered, and concentrated. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 10:1 → 3:1) to obtain 2.20 g of (Z)-ethyl(3-(p-tolyl)thiazole-2(3H)-ylidene)carbamate as a yellow oil. 1 H NMR (400 MHz, CDCl3) δ 7.26 (d, J = 8.4 Hz, 2H), 7.21 (d, J = 8.4 Hz, 2H), 6.93 (d, J = 4.8 Hz, 1H), 6.58 (d, J = 4.8 Hz, 1H), 4.14 (q, J = 7.2 Hz, 2H), 2.32 (s, 3H), 1.22 (t, J = 7.2 Hz, 3H). LCMS o(m / z [M+H] + ): 263.1.

[0118] <Intermediate C9> A solution of (Z)-ethyl(3-(p-tolyl)thiazole-2(3H)-ylidene) carbamate (0.500 g, 1.86 mmol, 1.0 equivalent) and NaOH (1.49 g, 37.2 mmol, 20 equivalents) in ethanol (6 mL) was stirred at 50°C for 1 hour. The mixture was cooled to room temperature, diluted with ethyl acetate (20 mL), washed with water (20 mL), followed by brine (20 mL), dehydrated with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (dichloromethane / methanol = 20:1) to obtain 230 mg of 3-(p-tolyl)thiazole-2(3H)-imine C9 as a yellow oil.

[0119] <Preparation of intermediate C10> [ka] <Ethyl N-[(4-methoxyphenyl)carbamate]carbamate> To a solution of 4-methoxyaniline (2.00 g, 16.2 mmol, 1.0 equivalent) and O-ethylcarbon isothiocyanate (2.13 g, 16.2 mmol, 1.0 equivalent) in ethyl acetate (20 mL), tetramethylethylenediamine (0.189 g, 1.62 mmol, 0.1 equivalent) was added at 25°C, and the mixture was stirred at 25°C for 2 hours. The mixture was concentrated under vacuum, the residue was triturated with ethanol (8 mL), and the precipitate was dried under vacuum to obtain 3.50 g of ethyl N-[(4-methoxyphenyl)carbamoteoyl]carbamate as a white solid. 1 H NMR (400 MHz, CDCl3): δ 11.37 (br. s, 1H), 11.18 (br. s, 1H), 7.44 (d, J =8.8 Hz, 2H), 6.94 (d, J =8.8 Hz, 2H), 4.20 (q, J = 7.2 Hz, 2H), 3.76 (s, 1H), 1.26 (t, J = 6.8 Hz, 3H).

[0120] <(Z)-Ethyl(((4-Methoxyphenyl)amino)((2-Oxoethyl)thio)methylene)carbamate> Ethyl N-[(4-methoxyphenyl)carbamate]carbamate (3.50 g, 13.7 mmol, 1.0 equivalent) and cesium carbonate (7.62 g, 23.4 mmol, 1.7 equivalents) were dissolved in acetonitrile (40 mL), to which 2-chloroacetaldehyde (3.38 g, 17.2 mmol, 1.25 equivalents) was added while maintaining the temperature below 25°C. After addition, the mixture was stirred at 25°C for 12 hours. The mixture was poured into water (150 mL) and extracted with TBME (150 mL x 2). The organic layers were washed together with brine (200 mL), dehydrated with sodium sulfate, and filtered. The filtrate was concentrated under reduced pressure to obtain 3.80 g of (Z)-ethyl(((4-methoxyphenyl)amino)((2-oxoethyl)thio)methylene)carbamate as a brown, rubbery substance. 1H NMR (400 MHz, DMSO-d6): δ 7.28 - 7.20 (m, 2H), 7.02 - 6.95 (m, 2H), 6.92 (d, J= 7.2 Hz, 1H), 5.59 - 5.54 (m, 1H), 3.95 (q, J = 7.2 Hz, 2H), 3.78 (s, 3H), 1.12 (t, J = 7.2 Hz, 3H). LCMS (m / z [M +H] + ): 297.0.

[0121] <(Z)-Ethyl(3-(4-methoxyphenyl)thiazole-2(3H)-ylidene)carbamate> (Z)-ethyl(((4-methoxyphenyl)amino)((2-oxoethyl)thio)methylene)carbamate (1.80 g, 6.07 mmol, 1.0 equivalent) was dissolved in tetrahydrofuran (20 mL) to which DIEA (2.36 g, 18.2 mmol, 3.0 equivalents) and then thionyl chloride (0.723 g, 6.07 mmol, 1.0 equivalent) were added at 0°C. After addition, the mixture was heated to 15°C and stirred for 4 hours. The mixture was directly concentrated under reduced pressure. The residue was dissolved in ethyl acetate (100 mL) and the pH was adjusted to 8.0 by adding saturated sodium bicarbonate aqueous solution. The organic layer was separated, washed with brine (20 mL), dehydrated with sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 10 / 1 → 5 / 1) to obtain 1.20 g of (Z)-ethyl(3-(4-methoxyphenyl)thiazole-2(3H)-ylidene)carbamate as a yellow solid. 1 H NMR (400 MHz, CDCl3) δ 7.47 (d, J= 4.8 Hz, 1H), 7.41 (d, J = 5.2 Hz, 2H), 7.07 (d, J = 4.8 Hz, 2H), 7.04 (d, J = 4.8 Hz, 1H), 4.01 (q, J = 6.8 Hz, 2H), 3.82 (s, 3H), 1.16 (t, J = 7.2 Hz, 3H).

[0122] <Intermediate C10> A solution of ethyl (3-(4-methoxyphenyl)thiazole-2(3H)-ylidene) carbamate (0.200 g, 0.718 mmol, 1.0 equivalent) in ethanol (2 mL) was mixed with sodium hydroxide (0.575 g, 14.4 mmol, 20 equivalents). The mixture was stirred at 50°C for 2 hours. The mixture was diluted with water (30 mL) and extracted with ethyl acetate (15 mL x 2). The organic layers were dehydrated together with sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography (dichloromethane / methanol = 10 / 1) to obtain 100 mg of 3-(4-methoxyphenyl)thiazole-2(3H)-imine C10 as a yellow oil.

[0123] <Preparation of intermediate C11> [ka] <Ethyl N-[(4-chlorophenyl)carbamate oil]carbamate> To a solution of 4-chloroaniline (2.00 g, 15.7 mmol, 1.0 equivalent) and O-ethylcarbon isothiocyanate (2.06 g, 15.7 mmol, 1.0 equivalent) in ethyl acetate (20 mL), tetramethylethylenediamine (0.182 g, 1.57 mmol, 0.1 equivalent) was added at 25°C, and the resulting mixture was stirred at 25°C for 2 hours. The mixture was concentrated under reduced pressure. The residue was triturated with ethanol (8 mL) to obtain 3.00 g of ethyl N-[(4-chlorophenyl)carbamoteoyl]carbamate as a white solid. 1 H NMR (400 MHz, CDCl3-d): δ 11.51 (br. s, 1H), 11.31 (br. s, 1H), 7.62 (d, J =8.8 Hz, 2H), 7.44 (d, J =8.8 Hz, 2H), 4.21 (q, J = 7.2 Hz, 2H), 1.26 (t, J = 6.8 Hz, 3H).

[0124] <(Z)-Ethyl(((4-chlorophenyl)amino)((2-oxoethyl)thio)methylene)carbamate> Ethyl ethyl N-[(4-chlorophenyl)carbamate]carbamate (3.00 g, 11.60 mmol, 1.0 equivalent) and cesium carbonate (6.42 g, 19.7 mmol, 1.7 equivalents) were suspended in acetonitrile (40 mL). 2-chloroacetaldehyde (2.84 g, 14.5 mmol, 1.25 equivalents) was added dropwise to this suspension while maintaining the temperature below 25°C. After the addition, the mixture was stirred at 25°C for 12 hours. The mixture was diluted with tert-butyl methyl ether (150 mL x 2), washed with brine (50 mL), dehydrated with sodium sulfate, and filtered. The filtrate was concentrated under reduced pressure to obtain 3.50 g of (Z)-ethyl(((4-chlorophenyl)amino)((2-oxoethyl)thio)methylene)carbamate as a brown, rubbery substance. 1 H NMR (400 MHz, DMSO-d6) δ 7.57 - 7.46 (m, 2H), 7.43 - 7.33 (m, 2H), 6.92(d, J = 7.2 Hz, 1H), 5.70 - 5.61 (m, 1H), 3.98 (q, J=6.8 Hz, 2H), 1.13 (t, J = 7.2 Hz, 3H). LCMS (m / z [M +H] + ): 301.0.

[0125] <(Z)-Ethyl(3-(4-chlorophenyl)thiazole-2(3H)-ylidene)carbamate> To a solution of (Z)-ethyl(((4-chlorophenyl)amino)((2-oxoethyl)thio)methylene) carbamate (1.80 g, 5.78 mmol, 1.0 equivalent) in tetrahydrofuran (20 mL), N-ethyl-N-isopropylpropan-2-amine (2.24 g, 17.3 mmol, 3.0 equivalent) and thionyl chloride (0.687 g, 5.78 mmol, 1.0 equivalent) were added at 0°C. The mixture was stirred at 25°C for 4 hours. The mixture was directly concentrated under reduced pressure. The residue was dissolved in ethyl acetate (100 mL) and the pH was adjusted to 8.0 by adding saturated sodium bicarbonate aqueous solution. The organic layer was separated, washed with brine (20 mL), dehydrated with sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 10 / 1 → 5 / 1) to obtain 1.10 g of (Z)-ethyl(3-(4-chlorophenyl)thiazole-2(3H)-ylidene)carbamate as a yellow solid. 1 H NMR (400 MHz, CDCl3-d): δ 7.65 - 7.55 (m, 4H), 7.54 (d, J = 4.4 Hz, 1H), 7.07 (d, J = 4.4 Hz, 1H), 4.03 (q, J = 7.2 Hz, 2H), 1.16 (t, J = 7.2Hz, 3H).

[0126] <Intermediate C11> A solution of (Z)-ethyl(3-(4-chlorophenyl)thiazole-2(3H)-ylidene) carbamate (0.200 g, 0.707 mmol, 1.0 equivalent) in ethanol (2 mL) was mixed with sodium hydroxide (0.569 g, 14.2 mmol, 20 equivalents) at 25°C. The reaction mixture was then heated to 50°C and stirred for 1.5 hours. The mixture was diluted with water (80 mL) and extracted with ethyl acetate (50 mL x 2). The organic layers were combined and dehydrated with sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to obtain 120 mg of 3-(4-chlorophenyl)thiazole-2(3H)-imine C11 as a yellow oil.

[0127] <Preparation of intermediate C12> [ka] <Ethyl N-(3-pyridylcarbamoteoyl)carbamate> To a solution of pyridine-3-amine (1.00 g, 10.6 mmol, 1.0 equivalent) and O-ethylcarbon isothiocyanate (1.39 g, 10.6 mmol, 1.0 equivalent) in ethyl acetate (10 mL), tetramethylethylenediamine (0.123 g, 1.06 mmol, 0.1 equivalent) was added at 25°C, and the mixture was then stirred at this temperature for 1 hour. The mixture was concentrated, the residue was triturated with ethanol (10 mL), the precipitate was collected by filtration, and dried under reduced pressure to obtain 1.80 g of ethyl N-(3-pyridylcarbamotioil)carbamate as a white solid. 1 H NMR (400MHz, CDCl3) δ 11.56 (br. s, 1H), 8.70 (d, J=2.4 Hz, 1H), 8.52 (dd, J = 4.8 Hz, 1.6 Hz, 1H), 8.30 (br. s, 1H), 8.28 - 8.23 ​​(m, 1H), 7.39 - 7.33 (m, 1H), 4.33 (q, J = 7.2 Hz, 2H), 1.38 (t, J = 7.2 Hz, 3H).

[0128] <(Z)-Ethyl(((2-oxoethyl)thio)(pyridine-3-ylamino)methylene)carbamate> 0.60 g, 2.66 mmol, 1.0 equivalent) of ethyl N-(3-pyridylcarbamotioyl)carbamate and cesium carbonate (1.48 g, 4.53 mmol, 1.7 equivalents) were suspended in acetonitrile (10 mL), to which 2-chloroacetaldehyde (1.05 g, 5.33 mmol, 2.0 equivalents) was added dropwise while maintaining the temperature at 25 °C. After the addition, the mixture was stirred at this temperature for 12 hours. The mixture was poured into water (100 mL) and extracted with ethyl acetate (50 mL x 3). The organic layers were washed together with brine (100 mL), dehydrated with anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to obtain 0.40 g of (Z)-ethyl(((2-oxoethyl)thio)(pyridine-3-ylamino)methylene)carbamate as a white solid. 1 H NMR (400 MHz, DMSO-d6) δ 8.58 (d, J = 2.0 Hz , 1H), 8.52 (dd, J = 1.6 Hz , 4.8Hz, 1H), 7.83-7.80 (m, 1H), 7.50 (q, J = 4.8 Hz , 1H),7.00 (d, J = 7.2 Hz , 1H), 5.72 (t, J = 6.0 Hz , 1H), 4.04-3.96 (m, 2H), 3.66 - 3.61 (m, 1H), 3.15 (dd, J= 12.4 Hz , 1.2 Hz, 1H), 1.13 (t, J = 6.8 Hz, 3H).LCMS (m / z [M +H] + ): 268.1.

[0129] <(Z)-Ethyl(3-(pyridine-3-yl)thiazole-2(3H)-ylidene)carbamate> (Z)-ethyl(((2-oxoethyl)thio)(pyridine-3-ylamino)methylene)carbamate (0.570 g, 2.13 mmol, 1.0 equivalent) was dissolved in tetrahydrofuran (6 mL), to which diisopropylethylamine (0.830 g, 6.40 mmol, 3.0 equivalent) and thionyl chloride (0.250 g, 2.13 mmol, 1.0 equivalent) were added at 0°C. The mixture was stirred at 25°C for 4 hours. The mixture was directly concentrated under reduced pressure. The residue was dissolved in ethyl acetate (100 mL) and the pH was adjusted to 8.0 with saturated sodium bicarbonate aqueous solution. The organic layer was separated, washed with brine (20 mL), dehydrated with sodium sulfate, filtered, and concentrated under reduced pressure to obtain 0.40 g of (Z)-ethyl(3-(pyridine-3-yl)thiazole-2(3H)-ylidene)carbamate as a yellow solid. 1 H NMR (400 MHz, DMSO-d6) δ 8.77 (d, J = 2.4 Hz, 1H), 8.65 (dd, J = 4.8 Hz, 1.6 Hz, 1H), 8.07 - 8.00 (m, 1H), 7.63 (d, J = 4.8 Hz, 1H), 7.63 - 7.59 (m, 1H), 7.12 (d, J = 4.8 Hz, 1H), 4.03 (q, J = 7.2 Hz, 2H), 1.15 (t, J = 7.2 Hz, 3H).

[0130] <Intermediate C12> To a solution of ethyl (3-(pyridine-3-yl)thiazole-2(3H)-ylidene) carbamate (0.200 g, 0.800 mmol, 1.0 equivalent) in ethanol (2 mL), sodium hydroxide (0.640 g, 16.1 mmol, 20 equivalents) was added at 25°C, and the mixture was then heated to 50°C and stirred for 1 hour. This mixture was diluted with water (50 mL) and extracted with ethyl acetate (50 mL x 2). The organic layers were dehydrated with sodium sulfate, filtered, and concentrated to obtain 140 mg of 3-(pyridine-3-yl)thiazole-2(3H)-imine C12 as a brown oil. 11H NMR (400 MHz, DMSO-d6, 400 MHz) δ 8.80 (d, J = 2.0 Hz, 1H), 8.45 (d, J = 4.0 Hz, 1H), 8.28 (br.s, 1H), 8.06 (d, J = 7.6 Hz, 1H), 7.51 - 7.43 (m, 1H), 7.12 (d, J = 5.2 Hz, 1H), 6.25 (d, J = 5.2 Hz, 1H).

[0131] <Preparation of Intermediate C13>

Chem.

[0132] <(Z)-Ethyl (((2-oxoethyl)thio)(pyridin-2-ylamino)methylene)carbamate> To a suspension of ethyl N-(2-pyridylcarbamotioyl)carbamate (0.800 g, 3.55 mmol, 1.0 equivalent) and cesium carbonate (1.97 g, 6.04 mmol, 1.7 equivalents) in acetonitrile (8 mL), 2-chloroacetaldehyde (0.714 mL, 4.44 mmol, 1.25 equivalents) was added dropwise at 25°C. After the addition, the mixture was stirred for 5 hours. Next, 2-chloroacetaldehyde (0.285 mL, 1.78 mmol, 0.5 equivalents) was added to the mixture and stirred for a further 2 hours. The mixture was poured into water (20 mL) and extracted with ethyl acetate (70 mL x 3). The organic layers were washed with brine (50 mL), dehydrated with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain (Z)-ethyl (((2-oxoethyl)thio)(pyridine-2-ylamino)methylene)carbamate (1.00 g) as a brown oil. 1 H NMR (400 MHz, CDCl3): δ 8.34 - 8.32 (m, 1H), 8.06 (d, J = 8.4 Hz, 1H), 7.93 - 7.79 (m, 1H), 7.17 - 7.14 (m, 1H), 6.19 (d, J = 6.4 Hz, 1H), 5.44 (s, 1H), 4.28 - 4.22 (m, 2H), 3.51 - 3.47 (m, 1H), 3.28 - 3.24 (m, 1H), 1.33 (t, J= 7.2 Hz, 3H).

[0133] <(Z)-Ethyl(3-(pyridine-2-yl)thiazole-2(3H)-ylidene)carbamate> A solution of (Z)-ethyl(((2-oxoethyl)thio)(pyridin-2-ylamino)methylene)carbamate (0.500 g, 1.87 mmol, 1.0 eq) in THF (5 mL) was added with N-ethyl-N-isopropylpropan-2-amine (0.977 mL, 5.61 mmol, 3.0 eq), followed by SOCl2 (136 μL, 1.87 mmol, 1.0 eq) at 0 °C. The mixture was stirred at below 15 °C for 4 h. The mixture was poured into water (10 mL) and extracted with ethyl acetate (40 mL×3). The combined organic layers were washed with brine (25 mL), dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 20 / 1→10 / 1) to obtain 210 mg of (Z)-ethyl(3-(pyridin-2-yl)thiazol-2(3H)-ylidene)carbamate as a brown solid. 1 H NMR (400 MHz, CDCl3) δ 8.51 - 8.49 (m, 1H), 8.42 (d, J = 8.4 Hz, 1H), 7.92 - 7.88 (m, 1H), 7.80 (d, J = 5.2 Hz, 1H), 7.31 - 7.28 (m, 1H), 0.65 (d, J = 5.2Hz, 1H), 4.28 (q, J = 7.2 Hz, 2H), 1.36 (t, J = 7.2 Hz, 3H). LCMS (m / z[M+H] + ): 249.8.

[0134] <Intermediate C13> (Z)-ethyl(3-(pyridine-2-yl)thiazole-2(3H)-ylidene)carbamate (0.210 g, 0.842 mmol, 1.0 equivalent) was added at 25°C to a solution of sodium hydroxide (0.674 g, 16.8 mmol, 20.0 equivalents) in ethanol (3 mL). The mixture was then heated to 50°C and stirred for 1 hour. The mixture was poured into water (15 mL) and stirred for 3 minutes, and extracted with ethyl acetate (40 mL x 3). The organic phases were combined and washed with brine (15 mL x 2), dehydrated with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain 130 mg of 3-(pyridine-2-yl)thiazole-2(3H)-imine C13 as a brown oil. 1 H NMR (400 MHz, DMSO-d6) δ 8.82 (s, 1H), 8.62 - 8.60 (m, 1H), 8.41 (d, J= 4.0 Hz, 1H), 7.89 - 7.86 (m, 1H), 7.60 (s, 1H), 7.24 - 7.21 (m, 1H), 7.26 (d, J = 5.2 Hz, 1H). LCMS (m / z [M+H] + ): 177.9.

[0135] <Preparation of intermediate C14> [ka] <Ethyl N-(tetrahydropyran-4-yl carbamoti oil) carbamate> A mixture of tetrahydro-2H-pyran-4-amine (0.800 g, 7.91 mmol, 1.0 equivalent) and O-ethylcarbon isothiocyanate (1.09 g, 8.30 mmol, 1.05 equivalent) in ethyl acetate (10 mL) was mixed with TMEDA (92 mg, 0.791 mmol, 0.1 equivalent). The mixture was stirred at 25°C for 6 hours and then concentrated under reduced pressure. The residue was triturated twice with a petroleum ether / ethanol solution (v / v=20 / 1, 21 mL) to obtain 1.80 g of ethyl N-(tetrahydropyran-4-yl carbamotioil)carbamate as a yellow solid. 1H NMR (400 MHz, DMSO-d6) δ 10.99 (br. s, 1H), 9.83 (d, J = 7.2 Hz, 1H), 4.40 - 4.26 (m, 1H), 4.12 (q, J = 7.2 Hz, 2H), 3.87 - 3.73 (m, 2H), 3.48 - 3.36 (m, 2H), 1.97 - 1.86 (m, 2H), 1.59 - 1.45 (m, 2H), 1.21(t, J= 7.2 Hz, 3H).

[0136] <(Z)-Ethyl(((2-oxoethyl)thio)((tetrahydro-2H-pyran-4-yl)amino)methylene)carbamate> A mixture of ethyl N-(tetrahydropyran-4-yl carbamotioil) carbamate (0.800 g, 3.44 mmol, 1.0 equivalent) and cesium carbonate (1.91 g, 5.85 mmol, 1.7 equivalents) in acetonitrile (10 mL) was mixed with 2-chloroacetaldehyde (2.09 g, 10.66 mmol, 3.1 equivalents). The resulting mixture was stirred at 25°C for 9 hours, then poured into ice water (10 mL) and extracted with ethyl acetate (100 mL x 2). The organic phases were combined and washed with brine (20 mL x 2), dehydrated with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 100 / 1 → 0 / 1) to obtain 0.30 g of (Z)-ethyl(((2-oxoethyl)thio)((tetrahydro-2H-pyran-4-yl)amino)methylene)carbamate as a yellow solid. LCMS (m / z [M+H] + ): 274.9.

[0137] <(Z)-Ethyl(3-(tetrahydro-2H-pyran-4-yl)thiazole-2(3H)-ylidene)carbamate> A mixture of (Z)-ethyl(((2-oxoethyl)thio)((tetrahydro-2H-pyran-4-yl)amino)methylene)carbamate (0.30 g, 0.52 mmol, 1.0 equivalent) and diisopropylethylamine (133 mg, 1.03 mmol, 2.0 equivalents) in tetrahydrofuran (10 mL) was mixed with thionyl chloride (61 mg, 0.52 mmol, 1.0 equivalent). The mixture was stirred at 25°C for 2 hours, then poured into ice water (20 mL) and extracted with ethyl acetate (100 mL x 3). The organic layers were washed together with brine (20 mL), dehydrated with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 100 / 1 → 0 / 1) to obtain 120 mg of (Z)-ethyl(3-(tetrahydro-2H-pyran-4-yl)thiazole-2(3H)-ylidene)carbamate as a yellow solid. 1 H NMR (400 MHz, CD3OD) δ 7.41 (d, J =4.8Hz, 1H), 6.87 (d, J = 4.8 Hz, 1H), 5.02 - 4.91 (m, 1H), 4.20 (q, J = 7.2 Hz, 2H), 4.09 - 4.04 (m, 2H), 3.62 - 3.58 (m, 2H), 1.99 - 1.89 (m, 4H), 1.31 (t, J = 7.2 Hz, 3H). LCMS (m / z [M+H] + ): 256.9.

[0138] <Intermediate C14> A mixture of (Z)-ethyl(3-(tetrahydro-2H-pyran-4-yl)thiazole-2(3H)-ylidene) carbamate (0.120 g, 0.468 mmol, 1.0 equivalent) and sodium hydroxide (0.374 g, 9.36 mmol, 20 equivalents) in ethanol (1 mL) was stirred at 50°C for 1 hour. The mixture was concentrated under reduced pressure, and the residue was purified by silica gel column chromatography (dichloromethane / methanol = 100 / 1 → 10 / 1) to obtain 80 mg of 3-(tetrahydro-2H-pyran-4-yl)thiazole-2(3H)-imine C14 as a yellow solid.

[0139] <Preparation of intermediate C15> [ka] <Ethyl N-(phenylcarbamate oil)carbamate> To a solution of aniline (1.96 mL, 21.5 mmol, 1.0 equivalent) in ethyl acetate (20 mL), O-ethylcarbon isothiocyanate (2.54 mL, 21.5 mmol, 1.0 equivalent) and tetramethylethylenediamine (0.324 mL, 2.15 mmol, 0.1 equivalent) were added. The mixture was stirred at 25°C for 4 hours and then concentrated directly under reduced pressure. The residue was triturated with petroleum ether (8 ml) to obtain 4.20 g of ethyl N-(phenylcarbamotioil)carbamate as a yellow oil. 1 H NMR (400 MHz, DMSO-d6) δ 11.55 (br. s, 1H), 11.25 (br. s, 1H), 7.61 -7.59 (m, 2H), 7.41 - 7.37 (m, 2H), 7.26 -7.22 (m, 1H), 4.21 (q, J = 7.2 Hz, 2H), 1.26 (t, J = 7.2 Hz, 3H).

[0140] <(Z)-Ethyl(((2-oxopropyl)thio)(phenylamino)methylene)carbamate> To a solution of N-(phenylcarbamotioil)carbamate (4.20 g, 18.7 mmol, 1.0 equivalent) and cesium carbonate (10.4 g, 31.8 mmol, 1.7 equivalents) in acetonitrile (50 mL), 1-chloropropan-2-one (2.17 g, 23.4 mmol, 1.25 equivalents) was added while maintaining the temperature below 25°C. After addition, the mixture was stirred at 25°C for 2 hours, then diluted with MTBE (120 mL), washed with saturated sodium bicarbonate aqueous solution (100 mL), followed by brine (100 mL), dehydrated with anhydrous sodium sulfate, and filtered. The filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 50:1 → 4:1) to obtain 1.80 g of (Z)-ethyl(((2-oxopropyl)thio)(phenylamino)methylene)carbamate as a brown oil. 1 H NMR (400 MHz, CDCl3) δ 7.46 -7.42 (m, 2H), 7.40 -7.36 (m, 1H), 7.28 - 7.27 (m, 1H), 7.27 - 7.26(m, 1H), 4.14 (q, J = 7.2.0 Hz, 2H), 3.47(d, J = 12.0 Hz, 1H), 3.65 (d, J = 12.0 Hz, 1H), 1.47 (s, 3H), 1.26 (t, J = 7.6 Hz, 4H). LCMS (m / z [M+H] + ): 280.9.

[0141] <(Z)-Ethyl(4-methyl-3-phenylthiazole-2(3H)-ylidene)carbamate> (Z)-ethyl(((2-oxopropyl)thio)(phenylamino)methylene)carbamate (0.600 g, 2.09 mmol, 1.0 equivalent) was dissolved in THF (6 mL) and diisopropylethylamine (0.810 g, 6.27 mmol, 3.0 equivalent) and thionyl chloride (0.167 mL, 2.30 mmol, 1.1 equivalent) were added at 0°C. The mixture was stirred at 20°C for 4 hours and then concentrated under reduced pressure. The residue was diluted with saturated sodium bicarbonate aqueous solution (40 mL) and extracted with ethyl acetate (30 mL x 3). The organic phases were combined and washed with brine (60 mL), dehydrated with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by reverse-phase HPLC to obtain 0.32 g of (Z)-ethyl(4-methyl-3-phenylthiazole-2(3H)-ylidene)carbamate as a yellow oil. 1 H NMR (400 MHz, CDCl3) δ 7.68(s, 3H), 7.45 (s, 2H), 6.92(s, 1H), 4.34(q, J = 7.2 Hz, 2H), 2.08(s, 3H), 1.34 (t, J = 7.2 Hz, 3H). LCMS (m / z [M+H] + ): 263.3.

[0142] <Intermediate C15> (Z)-ethyl(4-methyl-3-phenylthiazole-2(3H)-ylidene) carbamate (0.240 g, 0.891 mmol, 1.0 equivalent) was dissolved in ethanol (4 mL) and sodium hydroxide (0.713 g, 17.8 mmol, 20 equivalents) was added at 25°C. The mixture was heated to 60°C and stirred for 1 hour, then diluted with water (40 mL) and extracted with ethyl acetate (30 mL x 3). The organic layers were washed together with brine (50 mL), dehydrated with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain 180 mg of 4-methyl-3-phenylthiazole-2(3H)-imine C15 as a yellow solid.

[0143] <Preparation of intermediate C16> [ka] <Ethyl N-(benzylcarbamoti oil)carbamate> Phenylmethaneamine (5.00 g, 46.7 mmol, 1.0 equivalent) and O-ethylcarbon isothiocyanate (6.43 g, 49.0 mmol, 1.05 equivalent) were dissolved in ethyl acetate (40 mL), to which TMEDA (0.542 g, 4.67 mmol, 1.0 equivalent) was added under a nitrogen atmosphere at 25°C. The mixture was stirred at 25°C for 6 hours and then concentrated under reduced pressure. The residue was triturated twice with a solution of petroleum and ethyl acetate (v / v = 10 / 1, 55 mL) to obtain 10.0 g of ethyl N-(benzylcarbamotioil)carbamate as a yellow solid. 1 H NMR (400 MHz, DMSO-d6) δ 11.04 (br. s, 1H), 10.22 (br. t, J = 5.6 Hz, 1H), 7.39 - 7.23 (m, 5H), 4.81 (d, J = 5.6 Hz, 2H), 4.14 (q, J = 7.2 Hz, 2H), 1.21 (t, J = 7.2 Hz, 3H).

[0144] <(Z)-Ethyl((benzylamino)((2-oxobutyl)thio)methylene)carbamate> To a suspension of ethyl N-(benzyl carbamotioil) carbamate (3.00 g, 12.6 mmol, 1.0 equivalent) and Cs2CO3 (6.97 g, 21.4 mmol, 1.7 equivalents) in acetonitrile (45 mL), 1-bromobutan-2-one (2.38 g, 15.7 mmol, 1.25 equivalents) was added while maintaining the temperature below 25°C. After addition, the mixture was stirred at 25°C for 5 hours, then poured into water (80 mL) and extracted with ethyl acetate (120 mL x 3). The organic layers were washed together with brine (100 mL x 2), dehydrated with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 1:0 → 5:1) to obtain 2.50 g of (Z)-ethyl((benzylamino)((2-oxobutyl)thio)methylene)carbamate as a yellow oil. 1H NMR (400 MHz, CDCl3) δ 7.32 - 7.27 (m, 3H), 7.25 - 7.22 (m, 2H), 4.96 (d, J = 15.6 Hz, 1H), 4.71 (d, J= 15.6 Hz, 1H), 4.18 (q, J = 7.2 Hz, 2H), 3.28 (d, J = 12.0 Hz, 1H), 3.11 (d, J = 12.0 Hz, 1H), 1.97 - 1.92(m, 1H) ,1.67 - 1.62 (m, 1H), 1.31 - 1.28 (m, 3H), 0.88 (t, J = 7.6 Hz, 3H ).LCMS (m / z [M+H] + ): 308.9.

[0145] <(Z)-Ethyl(3-benzyl-4-ethylthiazole-2(3H)-ylidene)carbamate> To a solution of ethyl(Z)-ethyl((benzylamino)((2-oxobutyl)thio)methylene)carbamate (2.50 g, 8.11 mmol, 1.0 equivalent) in THF (38 mL), diisopropylethylamine (3.14 g, 24.3 mmol, 3.0 equivalent) and SOCl2 (0.964 g, 8.11 mmol, 1.0 equivalent) were added at 0°C. The mixture was stirred at 25°C for 16 hours, then poured into water (80 mL) and extracted with ethyl acetate (100 mL x 3). The organic layers were washed together with brine (100 mL x 2), dehydrated with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain 2.20 g of (Z)-ethyl(3-benzyl-4-ethylthiazole-2(3H)-ylidene)carbamate as a dark brown oil. 11H NMR (400 MHz, DMSO-d6) δ 7.37 - 7.33 (m, 2H), 7.29 - 7.27 (m, 1H), 7.09 (d, J = 7.2 Hz, 2H), 6.61 (s, 1H), 5.36 - 5.34 (m, 2H), 4.07 - 4.03 (m, 2H), 2.46 (dd, J = 2.0 Hz, 7.6 Hz, 2H), 1.17 (t, J = 7.2 Hz, 3H), 1.11 (t, J = 7.2 Hz, 3H). LCMS (m / z [M+H] + ): 291.1.

[0146] <Intermediate C16> (Z)-Ethyl (3-benzyl-4-ethylthiazol-2(3H)-ylidene)carbamate (1.00 g, 3.44 mmol, 1.0 eq) in ethanol (10 mL) was added 6N NaOH (2.75 g, 68.9 mmol, 20 eq) at 25 °C. The mixture was heated to 50 °C and stirred for 1 h, then poured into water (80 mL) and extracted with ethyl acetate (100 mL × 3). The combined organic layers were washed with brine (100 mL × 2), dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 1:0 → 5:1) to give 0.49 g of 3-benzyl-4-ethylthiazol-2(3H)-imine C16 as a dark brown oil.

[0147] <Preparation of Intermediate C17>

Chemical Structure

[0148] <Ethyl(Z)-ethyl(3-benzyl-4-isopropylthiazole-2(3H)-ylidene)carbamate> (Z)-ethyl((benzylamino)((3-methyl-2-oxobutyl)thio)methylene)carbamate (0.670 g, 2.08 mmol, 1.0 equivalent) and diisopropylethylamine (1.09 mL, 6.23 mmol, 3.0 equivalents) were dissolved in THF (10 mL) to which thionyl chloride (0.247 g, 2.08 mmol, 1.0 equivalent) was added. The mixture was stirred at 25°C for 16 hours, then poured into water (20 mL) and extracted with ethyl acetate (100 mL x 3). The organic layers were washed together with brine (20 mL), dehydrated with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 100 / 1 → 4 / 1) to obtain 0.53 g of ethyl(Z)-ethyl(3-benzyl-4-isopropylthiazole-2(3H)-ylidene)carbamate as a yellow solid. 1H NMR (400 MHz, DMSO-d6) δ 7.38 - 7.24 (m, 3H), 7.04(d, J=7.2 Hz, 2H), 6.70 (s, 1H), 5.39 (s, 2H), 4.04 (q, J = 7.2 Hz, 2H), 2.87 - 2.77 (m, 1H), 1.16 (t, J =7.2 Hz, 3H), 1.08 (d, J= 6.8 Hz, 6H). LCMS (m / z: 305.1[M+H] + ): 305.1.

[0149] <Intermediate C17> A mixture of ethyl(Z)-ethyl(3-benzyl-4-isopropylthiazole-2(3H)-ylidene) carbamate (0.300 g, 0.830 mmol) and sodium hydroxide (0.664 g, 16.61 mmol) in ethanol (5 mL) was stirred at 50°C for 1 hour. The mixture was poured into water (10 mL) and extracted with ethyl acetate (100 mL x 3). The organic layers were washed together with brine (10 mL), dehydrated with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (dichloromethane / methanol = 100 / 1 → 10 / 1) to obtain 150 mg of 3-benzyl-4-isopropylthiazole-2(3H)-imine C17 as a yellow solid.

[0150] <Preparation of intermediate C18> [ka] <(Z)-Ethyl((benzylamino)((2-cyclopropyl-2-oxoethyl)thio)methylene)carbamate> To a mixture of ethyl N-(benzylcarbamotioyl)carbamate (0.500 g, 2.10 mmol, 1.0 equivalent) and cesium carbonate (1.16 g, 3.57 mmol, 1.7 equivalents) in acetonitrile (5 mL), 2-bromo-1-cyclopropyl-ethanone (0.513 g, 3.15 mmol, 1.5 equivalents) was added. The resulting mixture was stirred at 25°C for 5 hours, then poured into ice water (10 mL) and extracted with MTBE (100 mL x 2). The organic layers were washed together with brine (20 mL x 2), dehydrated with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain (Z)-ethyl((benzylamino)((2-cyclopropyl-2-oxoethyl)thio)methylene)carbamate (0.670 g, crude) as a yellow solid. LCMS (m / z [M+H] + ): 321.1.

[0151] <Ethyl(Z)-ethyl(3-benzyl-4-cyclopropylthiazole-2(3H)-ylidene)carbamate> A mixture of (Z)-ethyl((benzylamino)((2-cyclopropyl-2-oxoethyl)thio)methylene) carbamate (0.670 g, 2.08 mmol, 1.0 equivalent) and diisopropylethylamine (0.810 g, 6.27 mmol, 3.0 equivalents) in tetrahydrofuran (10 mL) was mixed with thionyl chloride (0.248 g, 2.09 mmol, 1.2 equivalents). The mixture was stirred at 25°C for 16 hours, then poured into ice water (20 mL) and extracted with ethyl acetate (100 mL x 3). The organic layers were washed together with brine (20 mL), dehydrated with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 100 / 1 → 4 / 1) to obtain 0.25 g of ethyl(Z)-ethyl(3-benzyl-4-cyclopropylthiazole-2(3H)-ylidene)carbamate as a yellow, rubbery substance. 1H NMR (400 MHz, DMSO-d6): δ 7.40 - 7.24 (m, 3H), 7.14 (d, J = 7.2Hz, 2H), 6.59 (s, 1H), 5.48(s, 2H), 4.03 (q, J = 7.2Hz, 2H), 1.69 - 1.60 (m, 1H), 1.17 (t, J = 7.2 Hz, 3H), 0.82 - 0.75 (m, 2H), 0.60 - 0.54 (m, 2H). LCMS (m / z [M+H] + ): 305.1.

[0152] <Intermediate C18> A mixture of ethyl(Z)-ethyl(3-benzyl-4-cyclopropylthiazole-2(3H)-ylidene) carbamate (0.200 g, 0.661 mmol, 1.0 equivalent) and 6N sodium hydroxide (0.529 g, 13.23 mmol, 20 equivalents) in ethanol (4 mL) was stirred at 50°C for 1 hour. The mixture was poured into ice water (10 mL) and extracted with ethyl acetate (100 mL x 3). The organic layers were washed together with brine (10 mL), dehydrated with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain 160 mg of 3-benzyl-4-cyclopropylthiazole-2(3H)-imine C18 as a yellow solid. 1 H NMR (400 MHz, DMSO-d6) δ 7.57 (br. s, 1H), 7.36 - 7.30 (m, 2H), 7.27 - 7.20 (m, 3H), 5.64 (s, 1H), 5.04 (s, 2H), 1.48 - 1.36 (m, 1H), 0.72 - 0.64 (m, 2H), 0.52 - 0.42 (m, 2H). LCMS (m / z [M+H] + ): 233.2.

[0153] <Preparation of intermediate C19> [ka] <(Z)-Ethyl((benzylamino)((3,3,3-trifluoro-2-oxopropyl)thio)methylene)carbamate> To a suspension of ethyl N-(benzyl carbamotioil) carbamate (0.500 g, 2.10 mmol, 1.0 equivalent) and cesium carbonate (1.16 g, 3.57 mmol, 1.7 equivalents) in acetonitrile (8 mL), 3-bromo-1,1,1-trifluoropropan-2-one (0.654 mL, 6.30 mmol, 3.0 equivalents) was added while maintaining the temperature below 25°C. After addition, the mixture was stirred at 25°C for 12 hours, then poured into water (40 mL) and extracted with MTBE (30 mL x 3). The organic layers were washed together with brine (50 mL), dehydrated with anhydrous sodium sulfate, and filtered. The filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 50:1 → 10:1) to obtain 0.75 g of (Z)-ethyl((benzylamino)((3,3,3-trifluoro-2-oxopropyl)thio)methylene)carbamate as a yellow oil. 1 H NMR (400 MHz, CDCl3) δ 7.35 - 7.28 (m, 5H), 5.37 (d, J = 15.6 Hz,1H), 4.62 (d, J = 16.0 Hz, 1H), 4.27 - 4.18 (m, 2H), 3.79 (br. s, 1H), 3.69 (d, J = 12.8 Hz, 1H), 3.25 (dd, J = 12.0 Hz, 0.8 Hz, 1H), 1.35 - 1.31 (m, 3H). 19 F NMR (400 MHz, CDCl3) δ-79.04 (s, 3F). LCMS (m / z [M+H] + ): 348.9.

[0154] <(Z)-Ethyl(3-benzyl-4-(trifluoromethyl)thiazole-2(3H)-ylidene)carbamate> (Z)-ethyl((benzylamino)((3,3,3-trifluoro-2-oxopropyl)thio)methylene)carbamate (0.550 g, 1.44 mmol, 1.0 equivalent) was dissolved in THF (20 mL) and diisopropylethylamine (0.754 mL, 4.33 mmol, 3.0 equivalent) and thionyl chloride (0.115 mL, 1.59 mmol, 1.1 equivalent) were added at 0°C. The mixture was heated to 60°C and stirred for 2 hours, then added to saturated sodium bicarbonate aqueous solution (40 mL) and extracted with ethyl acetate (30 mL x 3). The organic layers were washed together with brine (60 mL), dehydrated with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 50:1 → 10:1) to obtain 0.40 g of (Z)-ethyl(3-benzyl-4-(trifluoromethyl)thiazole-2(3H)-ylidene)carbamate as a brown oil. 1 H NMR (400 MHz, CDCl3): δ 7.34 - 7.28 (m, 3H), 7.18 - 7.13 (m, 3H), 5.48 (s, 2H), 4.26 (q, J= 7.2 Hz, 2H), 1.34 (t, J = 6.8 Hz, 3H). LCMS (m / z [M+H] + ): 330.9.

[0155] <Intermediate C19> A solution of (Z)-ethyl(3-benzyl-4-(trifluoromethyl)thiazole-2(3H)-ylidene) carbamate (0.300 g, 0.884 mmol, 1.0 equivalent) in ethanol (5 mL) was prepared by adding sodium hydroxide (0.707 g, 17.7 mmol, 20 equivalents) at 25°C. The mixture was heated to 60°C and stirred for 1 hour, then poured into water (40 mL) and extracted with ethyl acetate (30 mL x 3). The organic layers were washed together with brine (50 mL), dehydrated with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 20:1) to obtain 150 mg of 3-benzyl-4-(trifluoromethyl)thiazole-2(3H)-imine C19 as a brown oil.

[0156] <Preparation of intermediate C20> [ka] <Ethyl N-[(3-cyanophenyl)carbamate oil]carbamate> To a stirred mixture containing 3-aminobenzonitrile (2.50 g, 21.16 mmol, 1.0 equivalent) in SiO2 (25 mL), O-ethylcarbon isothiocyanate (2.78 g, 21.16 mmol, 1.0 equivalent) and TMEDA (246 mg, 2.12 mmol, 319 μL, 0.1 equivalent) were added at 25°C. The reaction mixture was then stirred under N2 at 25°C for 5 hours. The reaction mixture was concentrated under reduced pressure. EtOH (50 mL) was added to the mixture, and the resulting mixture was stirred at 25°C for 0.5 hours. The mixture was filtered, and the filtered cake was dried under reduced pressure to obtain 2.80 g of ethyl N-[(3-cyanophenyl)carbamate]carbamate as a yellow solid.

[0157] <(Z)-Ethyl(((3-Cyanophenyl)amino)((2-Oxoethyl)thio)methylene)carbamate> A mixture of ethyl N-[(3-cyanophenyl)carbamate]carbamate (2.80 g, 11.2 mmol, 1.0 equivalent) and Cs2CO3 (6.22 g, 19.1 mmol, 1.7 equivalents) in MeCN (40 mL) was mixed with 2-chloroacetaldehyde (2.76 g, 14.0 mmol, 2.26 mL, 1.2 equivalents) at 25°C. The reaction mixture was stirred under N2 at 25°C for 12 hours. An additional 1.0 equivalent of 2-chloroacetaldehyde was added, and the reaction mixture was stirred at 25°C for another 12 hours. The reaction mixture was diluted with MTBE (100 mL) and washed with NaHCO3 solution (2 × 50 mL). The organic layer was washed with brine (50 mL), dehydrated with Na2SO4, and filtered. The filtrate was concentrated under reduced pressure to obtain 2.80 g of (Z)-ethyl(((3-cyanophenyl)amino)((2-oxoethyl)thio)methylene)carbamate as a brown solid.

[0158] <(Z)-Ethyl(3-(3-cyanophenyl)thiazole-2(3H)-ylidene)carbamate> A mixture of (Z)-ethyl(((3-cyanophenyl)amino)((2-oxoethyl)thio)methylene) carbamate (2.80 g, 9.61 mmol, 1.0 equivalent) in THF (30 mL) was mixed with DIPEA (3.73 g, 28.8 mmol, 5.02 mL, 3.0 equivalent) and SOCl2 (1.14 g, 9.61 mmol, 697 μL, 1.0 equivalent) at 0°C. The reaction mixture was then stirred under N2 at 25°C for 15 hours. The reaction mixture was concentrated under reduced pressure. The residue was dissolved in MTBE (200 mL), the mixture was adjusted to pH=8 with saturated NaHCO3 aqueous solution, and the organic layer was discarded. The aqueous phase was extracted with SiO2 (3 × 100 mL), the organic layer was washed with brine (100 mL), dehydrated with Na2SO4, filtered, and the filtrate was concentrated under reduced pressure. MTBE (10 mL) was added to the mixture, and the mixture was stirred at 25°C for 0.5 hours. The mixture was then filtered, and the filtered cake was dried under reduced pressure to obtain 1.30 g of ethyl(Z)-ethyl(3-(3-cyanophenyl)thiazole-2(3H)-ylidene)carbamate as a brown solid. 1H NMR: (400 MHz, DMSO-d6) δ 8.12 (s, 1H), 7.99-7.91 (m, 2H), 7.79-7.74 (m, 1H), 7.61 (d, J = 4.8 Hz, 1H), 7.11 (d, J = 4.8 Hz, 1H), 4.09-3.98 (m, 2H), 1.17 (t, J = 7.1 Hz, 3H).

[0159] <Intermediate C20> A mixture of (Z)-ethyl(3-(3-cyanophenyl)thiazole-2(3H)-ylidene) carbamate (400 mg, 1.46 mmol, 1.0 equivalent) in EtOH (4 mL) was mixed with NaOH (6 M, 4.00 mL, 16.4 equivalents) at 25°C. The reaction mixture was then stirred under N2 at 25°C for 12 hours and at 60°C for 2 hours. The reaction mixture was adjusted to pH=7 with concentrated HCl at 0°C, and the resulting solution was filtered. The filtrate was concentrated under reduced pressure. The residue was purified by preparative HPLC to obtain 130 mg of 3-(2-iminothiazole-3(2H)-yl)benzamide C20 as a brown solid.

[0160] <Preparation of intermediate C21> [ka] <tert‐ブチル7‐ニトロ‐3,4‐ジヒドロイソキノリン‐2(1H)‐カルボキシラート> To a solution of 7-nitro-1,2,3,4-tetrahydroisoquinoline (4.00 g, 22.4 mmol, 1.0 equivalent) in DCM (40 mL), TEA (4.54 g, 44.9 mmol, 6.25 mL, 2.0 equivalents) and di-tert-butyldicarbonate (5.39 g, 24.7 mmol, 5.67 mL, 1.1 equivalents) were added at 25°C. The reaction mixture was stirred at 25°C for 2 hours. The reaction mixture was diluted with water (100 mL) and extracted with SiO (4 × 200 mL). The organic layers were washed together with brine (100 mL), dehydrated with Na₂SO₄, filtered, and concentrated under reduced pressure to obtain the residue. The residue was purified by flash silica gel chromatography (eluent with a 0-17% ethyl acetate / petroleum ether gradient, 100 mL / min) to obtain 5.30 g of tert-butyl 7-nitro-3,4-dihydroisoquinoline-2(1H)-carboxylate as a yellow oil. 1 H NMR (400 MHz, DMSO-d6) δ 8.12 (s, 1H), 8.02 (dd, J = 1.9, 8.4 Hz, 1H), 7.45 (d, J = 8.4 Hz, 1H), 4.63 (br s, 2H), 3.64-3.52 (m, 2H), 2.97-2.80 (m, 2H), 1.43 (s, 9H).

[0161] <tert‐ブチル7‐アミノ‐3,4‐ジヒドロイソキノリン‐2(1H)‐カルボキシラート> To a solution of tert-butyl 7-nitro-3,4-dihydroisoquinoline-2(1H)-carboxylate (5.9 g, 21.2 mmol, 1.0 equivalent) in MeOH (50 mL), Pd / C (2.5 g, 10% purity) was added under N2 at 25°C. The suspension was degassed under vacuum and purged three times with H2. The reaction mixture was stirred under H2 (15 psi) at 25°C for 12 hours. The reaction mixture was filtered, and the filtrate was concentrated under reduced pressure to obtain tert-butyl 7-amino-3,4-dihydroisoquinoline-2(1H)-carboxylate (4.5 g, crude) as a red oil.

[0162] <tert‐ブチル7‐(3‐(エトキシカルボニル)チオウレイド)‐3,4‐ジヒドロイソキノリン‐2(1H)‐カルボキシラート> To a mixture of tert-butyl 7-amino-3,4-dihydroisoquinoline-2(1H)-carboxylate (1.00 g, 4.03 mmol, 1.0 equivalent) in SiO2 (10 mL), O-ethylcarbon isothiocyanate (528 mg, 4.03 mmol, 1.0 equivalent) and TMEDA (47 mg, 0.40 mmol, 61 μL, 0.1 equivalent) were added at 25°C. The reaction mixture was then stirred under N2 at 25°C for 5 hours. The reaction mixture was concentrated under reduced pressure to remove the solvent. The residue was dissolved in EtOH (10 mL), and the mixture was stirred at 25°C for 0.5 hours. The mixture was filtered, and the filtered cake was dried under reduced pressure to obtain 1.20 g of tert-butyl 7-(3-(ethoxycarbonyl)thioureid)-3,4-dihydroisoquinoline-2(1H)-carboxylate as a white solid. LCMS (m / z [M+Na] + ): 402.2.

[0163] <(Z)-tert-butyl7-((((ethoxycarbonyl)imino)((2-oxoethyl)thio)methyl)amino)-3,4-dihydroisoquinoline-2(1H)-carboxylate> To a mixture of tert-butyl 7-(3-(ethoxycarbonyl)thioureid)-3,4-dihydroisoquinoline-2(1H)-carboxylate (1.80 g, 4.74 mmol, 1.0 equivalent) in MeCN (20 mL), Cs2CO3 (2.63 g, 8.06 mmol, 1.7 equivalents) and 2-chloroacetaldehyde (1.16 g, 5.93 mmol, 954 μL, 1.25 equivalents) were added at 25°C. The reaction mixture was then stirred under N2 at 25°C for 12 hours. The reaction mixture was diluted with MTBE (100 mL) and then washed with saturated NaHCO3 aqueous solution (2 × 50 mL). The organic layer was washed with brine (50 mL), dehydrated with Na2SO4, filtered, and the filtrate was concentrated under reduced pressure to obtain (Z)-tert-butyl 7-((((ethoxycarbonyl)imino)((2-oxoethyl)thio)methyl)amino)-3,4-dihydroisoquinoline-2(1H)-carboxylate (2.4 g, crude) as a brown oil.

[0164] <tert‐ブチル7‐[(2Z)‐2‐エトキシカルボニルイミノチアゾール‐3‐イル]‐3,4‐ジヒドロ‐1H‐イソキノリン‐2‐カルボキシラート> A mixture of (Z)-tert-butyl 7-((((ethoxycarbonyl)imino)((2-oxoethyl)thio)methyl)amino)-3,4-dihydroisoquinoline-2(1H)-carboxylate (2.40 g, 5.69 mmol, 1.0 equivalent) in THF (30 mL) was mixed with DIPEA (2.21 g, 17.1 mmol, 2.98 mL, 3.0 equivalent) and SOCl2 (677 mg, 5.69 mmol, 413 μL, 1.0 equivalent) at 0°C. The reaction mixture was then stirred under N2 at 25°C for 12 hours. The reaction mixture was diluted with water (100 mL) and extracted with SiO2 (3 × 50 mL). The organic layer was washed with brine (100 mL), dehydrated with Na2SO4, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by flash silica gel chromatography (ISCO®; 20 g SepaFlash® silica flash column, eluent with a 0-40% ethyl acetate / petroleum ether gradient, 75 mL / min) to obtain 1.0 go of (Z)-tert-butyl 7-((((ethoxycarbonyl)imino)((2-oxoethyl)thio)methyl)amino)-3,4-dihydroisoquinoline-2(1H)-carboxylate as a brown oil.

[0165] <Intermediate C21> (Z)-tert-butyl7-((((ethoxycarbonyl)imino)((2-oxoethyl)thio)methyl)amino)-3,4-dihydroisoquinoline-2(1H)-carboxylate (800 mg, 1.98 mmol, 1.0 equivalent) in EtOH (8 mL) was mixed with NaOH (6 M, 8.00 mL, 24 equivalents) at room temperature. The reaction mixture was then stirred under N2 at 60°C for 2 hours. The reaction mixture was filtered, the filtrate was washed with ELISA (3 × 20 mL), and the filtrate was concentrated under reduced pressure. The residue was purified by reverse-phase column chromatography to obtain 170 mg of tert-butyl7-(2-iminothiazole-3(2H)-yl)-3,4-dihydroisoquinoline-2(1H)-carboxylate C21 as a yellow solid.

[0166] <Preparation of intermediate C22> [ka] <Ethyl N-[(2-bromophenyl)carbamate]carbamate> To a solution of 2-bromoaniline (10.00 g, 58.1 mmol, 1.0 equivalent) in SiO2 (100 mL), ethyl N-(thioxomethylene)carbamate (7.62 g, 58.1 mmol, 1.0 equivalent) and TMEDA (675 mg, 5.81 mmol, 877 μL, 0.1 equivalent) were added at room temperature. The mixture was stirred at 20°C for 12 hours. The reaction mixture was concentrated under reduced pressure. The residue was triturated with EtOH (50 mL) for 0.5 hours. The mixture was filtered, and the filtered cake was washed with EtOH (2 × 30 mL). The filtered cake was collected, and 16.0 g of ethyl N-[(2-bromophenyl)carbamotioil]carbamate was obtained as a white solid.

[0167] <(Z)-Ethyl(((2-bromophenyl)amino)((2-oxoethyl)thio)methylene)carbamate> To a solution of ethyl N-[(2-bromophenyl)carbamotioil]carbamate (16.0 g, 52.8 mmol, 1.0 equivalent) in ACN (200 mL), Cs2CO3 (31.0 g, 95.0 mmol, 1.8 equivalents) and 2-chloroacetaldehyde (20.7 g, 106 mmol, 17.0 mL, 2.0 equivalents) were added at room temperature. The mixture was stirred at 20°C for 12 hours. The reaction mixture was quenched by adding water (500 mL) and extracted with ethyl acetate (3 × 500 mL). The organic layers were dehydrated with Na2SO4, filtered, and concentrated under reduced pressure to obtain 24.0 g of (Z)-ethyl(((2-bromophenyl)amino)((2-oxoethyl)thio)methylene)carbamate as a pale yellow solid.

[0168] <(Z)-Ethyl(3-(2-bromophenyl)thiazole-2(3H)-ylidene)carbamate> (Z)-ethyl(((2-bromophenyl)amino)((2-oxoethyl)thio)methylene) carbamate (24.0 g, 69.5 mmol, 1.0 equivalent) was dissolved in THF (240 mL) and DIPEA (27.0 g, 209 mmol, 36.3 mL, 3.0 equivalent) and SOCl2 (8.27 g, 69.5 mmol, 5.0 mL, 1.0 equivalent) were added at 0°C. The mixture was stirred at 15°C for 4 hours. The reaction mixture was quenched by adding saturated NaHCO3 solution (400 mL) and extracted with ethyl acetate (3 × 400 mL). The organic layers were dehydrated together with Na2SO4, filtered, and concentrated under reduced pressure to obtain the residue. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 3 / 1 → 1 / 1) to obtain 13.43 g of (Z)-ethyl(3-(2-bromophenyl)thiazole-2(3H)-ylidene)carbamate. LCMS: (m / z [M+H + ]): 329.0.

[0169] <Intermediate C22> (Z)-ethyl(3-(2-bromophenyl)thiazole-2(3H)-ylidene)carbamate (2.00 g, 6.11 mmol, 1.0 equivalent) was dissolved in EtOH (20 mL), to which 6N NaOH (4.89 g, 122 mmol, 20 equivalents) was added at room temperature. The mixture was stirred under N2 at 50°C for 1.5 hours. The reaction mixture was quenched by adding water (40 mL) and extracted with ethyl acetate (3 × 40 mL). The organic layers were dehydrated together with Na2SO4, filtered, and concentrated under reduced pressure to obtain 1.56 g of 3-(2-bromophenyl)thiazole-2(3H)-imine C22 as a yellow oil. LCMS (m / z [M+H] + ): 255.1.

[0170] <Preparation of intermediate C23> [ka] <tert‐ブチル2‐アミノベンジルカルバマート> A solution of 2-(aminomethyl)aniline (5.00 g, 40.9 mmol, 1.0 equivalent) in DCM (50 mL) was added to a solution of BOC2O (8.93 g, 40.9 mmol, 9.40 mL, 1.0 equivalent) in DCM (20 mL). The reaction mixture was stirred at 25°C for 12 hours under an N2 atmosphere. The reaction mixture was extracted with H2O (25 mL). The organic layers were washed together with brine (25 mL), dehydrated with Na2SO4, filtered, and concentrated under reduced pressure to obtain 8.36 g of tert-butyl2-aminobenzylcarbamate as a pale yellow solid. 1 H NMR (400 MHz, DMSO-d6) δ 7.18 (s, 1H), 7.19-7.18 (m, 1H), 7.05-7.00 (m, 1H), 7.02 (dt, J = 1.3, 7.6 Hz, 1H), 6.95 (d, J = 7.3 Hz, 1H), 6.65-6.53 (m, 2H), 4.19-4.14 (m, 1H), 4.19-4.11 (m, 1H), 4.16 (d, J = 6.4 Hz, 2H), 1.36-1.36 (m, 1H), 1.37 (s, 9H). LCMS(m / z [M+H] + ): 166.9.

[0171] <Ethyl N-[[2-[(tert-butoxycarbonylamino)methyl]phenyl]carbamate oil> To a solution of tert-butyl-2-aminobenzylcarbamatenyl)methyl carbamate (5.00 g, 22.5 mmol, 1.0 equivalent) in SiO2 (50 mL), TMEDA (2.61 g, 22.5 mmol, 3.39 mL, 1.0 equivalent) and ethyl N-(thioxomethylene) carbamate (4.43 g, 33.7 mmol, 1.5 equivalent) were added. The reaction mixture was stirred at 25°C for 5 hours. The mixture was directly concentrated under reduced pressure to obtain a yellow oil. This oil was dissolved in MTBE (20 mL) and stirred for 0.5 hours, then filtered. The filtered cake was washed with MTBE (2 × 10 mL) and dried under vacuum to obtain 7.75 g of ethyl N-[[2-[(tert-butoxycarbonylamino)methyl]phenyl]carbamotioil]carbamate as a yellow solid. 1 H NMR (400MHz, DMSO-d6) δ 11.18 (br s, 1H), 11.21-11.15 (m, 1H), 11.24-11.09 (m, 1H), 11.24-11.09 (m, 1H), 7.35-7.31 (m, 1H), 7.29-7.19 (m, 3H), 7.12-7.03 (m, 1H), 7.12-7.02 (m, 1H), 7.13-7.00 (m, 1H), 4.62 (s, 1H), 4.65-4.55 (m, 1H), 4.29-4.12 (m, 2H), 4.10-4.02 (m, 2H), 1.37 (s, 9H), 1.28-1.19 (m, 1H), 1.23 (td, J = 7.2, 16.0 Hz, 3H). LCMS (m / z [M+H] + ): 298.1.

[0172] <(Z)-Ethyl(((2-(((tert-butoxycarbonyl)amino)methyl)phenyl)amino)((2-oxoethyl)thio)methylene)carbamate> To a solution of ethyl N-[[2-[(tert-butoxycarbonylamino)methyl]phenyl]carbamate]carbamate (3.70 g, 10.5 mmol, 1.0 equivalent) in MeCN (50 mL), Cs2CO3 (5.80 g, 17.8 mmol, 1.7 equivalents) and then 2-chloroacetaldehyde (2.57 g, 13.1 mmol, 2.10 mL, 1.25 equivalents) were added at 10°C. The reaction mixture was stirred at 20°C for 12 hours. The reaction mixture was filtered and concentrated under reduced pressure. The residue was diluted with water (50 mL) and extracted with SiO (3 × 20 mL). The organic layers were washed together with brine (50 mL), dehydrated with Na2SO4, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 1 / 0 → 1 / 1) to obtain 3.0 g of (Z)-ethyl(((2-(((tert-butoxycarbonyl)amino)methyl)phenyl)amino)((2-oxoethyl)thio)methylene)carbamate as a yellow solid. LCMS (m / z [M+H] + ):396.1.

[0173] <(Z)-Ethyl(3-(2-(((tert-butoxycarbonyl)amino)methyl)phenyl)thiazole-2(3H)-ylidenecarbamate> A solution of (Z)-ethyl(((2-(((tert-butoxycarbonyl)amino)methyl)phenyl)amino)((2-oxoethyl)thio)methylene)carbamate (2.00 g, 5.06 mmol, 1.0 equivalent) in THF (40 mL) was mixed with DIPEA (1.96 g, 15.2 mmol, 2.64 mL, 3.0 equivalent) and SOCl2 (602 mg, 5.06 mmol, 367 μL, 1.0 equivalent) at 0°C. The reaction mixture was stirred at 15°C for 1 hour under an N2 atmosphere. The reaction mixture was concentrated under reduced pressure to remove the solvent. The residue was diluted with saturated NaHCO3 aqueous solution (25 mL) and extracted with MTBE (3 × 25 mL). The organic layers were washed together with brine (25 mL), dehydrated with Na2SO4, filtered, and concentrated under reduced pressure to obtain the residue. This residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 1 / 0 → 2 / 1) to obtain 1.06 g of (Z)-ethyl(3-(2-(((tert-butoxycarbonyl)amino)methyl)phenyl)thiazole-2(3H)-ylidene)carbamate as a yellow solid. 1 H NMR (400MHz, DMSO-d6) δ 7.55-7.47 (m, 1H), 7.42 (br s, 1H), 7.46-7.39 (m, 1H), 7.37 (d, J = 4.6 Hz, 1H), 7.28 (d, J = 7.7 Hz, 1H), 7.24 (br t, J = 5.6 Hz, 1H), 7.08 (d, J = 4.9 Hz, 1H), 4.07-3.93 (m, 2H), 3.88 (br d, J = 6.0 Hz, 2H), 1.40-1.21 (m, 9H), 1.20-1.09 (m, 3H).LCMS (m / z [M+H] + ): 378.3.

[0174] <Intermediate C23> (Z)-ethyl(3-(2-(((tert-butoxycarbonyl)amino)methyl)phenyl)thiazole-2(3H)-ylidene)carbamate (340 mg, 901 μmol, 1.0 equivalent) was dissolved in EtOH (3.4 mL) and NaOH (831 mg, 20.8 mmol, 23 equivalents) was added. The reaction mixture was stirred at 50°C for 1.5 hours under an N2 atmosphere. The reaction mixture was diluted with water (20 mL) and extracted with RINKAN (3 × 20 mL). The organic layers were washed together with brine (20 mL), dehydrated with Na2SO4, filtered, and concentrated under reduced pressure. The residue was purified by preparative HPLC to obtain 133 mg of tert-butyl 2-(2-iminothiazole-3(2H)-yl)benzylcarbamate C23 as a pale yellow solid.

[0175] <Preparation of intermediate C24> [ka] <tert‐ブチル3‐アミノベンジルカルバマート> A solution of 3-(aminomethyl)aniline (5.00 g, 40.9 mmol, 1.0 equivalent) in MeCN (50 mL) was added dropwise to a solution of Boc2O (8.13 g, 37.2 mmol, 8.56 mL, 0.9 equivalents) in MeCN (50 mL) at 25°C. The reaction mixture was then stirred under N2 at 25°C for 12 hours. The reaction mixture was diluted with ethyl acetate (100 mL), filtered through a silica gel layer, and then washed with ethyl acetate (500 mL). The filtrate was concentrated under reduced pressure to obtain 9.4 g of tert-butyl 3-aminobenzylcarbamate as a yellow oil. 1 H NMR (400MHz, DMSO-d6) δ 7.23 (br t, J = 5.9 Hz, 1H), 6.95 (t, J = 7.6 Hz, 1H), 6.48-6.38(m, 3H), 4.99 (s, 2H), 4.00 (br d, J= 6.1 Hz, 2H), 1.42 (s, 9H).

[0176] <Ethyl N-[[3-[(tert-butoxycarbonylamino)methyl]phenyl]carbamate oil> To a solution of tert-butyl 3-aminobenzyl carbamate (5.00 g, 22.5 mmol, 1.0 equivalent) in SiO2 (50 mL), ethyl O-ethylcarbon isothiocyanate (2.95 g, 22.5 mmol, 1.0 equivalent) and TMEDA (261 mg, 2.25 mmol, 339 μL, 0.1 equivalent) were added at 25°C. The reaction mixture was stirred under N2 at 25°C for 12 hours. The reaction mixture was concentrated under reduced pressure. The residue was diluted with MTBE (50 mL) and stirred at 25°C for 0.5 hours. This was filtered to recover 4.3 g of ethyl N-[[3-[(tert-butoxycarbonylamino)methyl]phenyl]carbamote oil]carbamate as a white solid. 1 H NMR (400MHz, DMSO-d6) δ 11.55 (s, 1H), 11.22 (br s, 1H), 7.53 (br d, J = 7.8 Hz, 1H), 7.46-7.38 (m, 2H), 7.33 (t, J = 7.8 Hz, 1H), 7.11 (d, J= 7.6 Hz, 1H), 4.22 (q, J = 7.1 Hz, 2H), 4.13 (br d, J = 6.1 Hz, 2H), 1.40 (s, 9H), 1.27 (t, J = 7.1 Hz, 3H).

[0177] <(Z)-Ethyl(((3-(((tert-butoxycarbonyl)amino)methyl)phenyl)amino)((2-oxoethyl)thio)methylene)carbamate> To a solution of ethyl N-[[3-[(tert-butoxycarbonylamino)methyl]phenyl]carbamate]carbamate (4.30 g, 12.2 mmol, 1.0 equivalent) and Cs2CO3 (6.74 g, 20.7 mmol, 1.7 equivalents) in MeCN (44 mL), 2-chloroacetaldehyde (2.98 g, 15.2 mmol, 2.45 mL, 1.25 equivalents) was added while maintaining the temperature below 25°C. After addition, the reaction mixture was stirred under N2 at 25°C for 12 hours. The reaction mixture was diluted with MTBE (80 mL), washed with saturated NaHCO3 aqueous solution (75 mL) and brine (75 mL), dehydrated with Na2SO4, and filtered. The filtrate was concentrated under reduced pressure to obtain 4.7 g of (Z)-ethyl(((3-(((tert-butoxycarbonyl)amino)methyl)phenyl)amino)((2-oxoethyl)thio)methylene)carbamate as a brown solid.

[0178] <(Z)-Ethyl(3-(3-(((tert-butoxycarbonyl)amino)methyl)phenyl)thiazole-2(3H)-ylidene)carbamate> (Z)-ethyl(((3-(((tert-butoxycarbonyl)amino)methyl)phenyl)amino)((2-oxoethyl)thio)methylene)carbamate (4.70 g, 11.9 mmol, 1.0 equivalent) was dissolved in THF (47 mL) and DIPEA (4.61 g, 35.6 mmol, 6.21 mL, 3.0 equivalent) and SOCl2 (1.41 g, 11.9 mmol, 862 μL, 1.0 equivalent) were added at 0°C. The reaction mixture was stirred under N2 at 25°C for 1 hour. The reaction mixture was concentrated under reduced pressure. The residue was dissolved in SiO (20 mL), diluted with saturated NaHCO3 aqueous solution (60 mL), and extracted with SiO (3 × 80 mL). The organic layers were washed together with brine (80 mL), dehydrated with Na2SO4, and filtered. The filtrate was concentrated under reduced pressure. The residue was purified by flash silica gel chromatography (eluting at 75 mL / min using a 0-40% ethyl acetate / petroleum ether gradient) to obtain 2.4 g of (Z)-ethyl(3-(3-(((tert-butoxycarbonyl)amino)methyl)phenyl)thiazole-2(3H)-ylidene)carbamate as a white solid. 1 H NMR (400MHz, DMSO-d6) δ 7.53-7.47 (m, 2H), 7.41-7.33 (m, 3H), 7.08 (d, J = 4.8 Hz, 1H), 4.21 (br d, J = 6.1 Hz, 2H), 4.06-4.00 (m, 2H), 1.40(s, 9H), 1.16 (t, J= 7.1 Hz, 3H).

[0179] <Intermediate C24> (Z)-ethyl(3-(3-(((tert-butoxycarbonyl)amino)methyl)phenyl)thiazole-2(3H)-ylidene)carbamate (1.00 g, 2.65 mmol, 1.0 equivalent) was dissolved in EtOH (10 mL) and 6N NaOH (2.40 g, 60.0 mmol, 23.0 equivalents) was added at room temperature. The reaction mixture was stirred under N2 at 60°C for 5 hours. The reaction mixture was adjusted to pH 7 with HCl aqueous solution (6 M). The neutralized solution was then filtered, and the filtrate was washed with EtOH (200 mL). The filtrate was concentrated under reduced pressure. The crude product was purified by reverse-phase column chromatography to obtain 300 mg of tert-butyl 3-(2-iminothiazole-3(2H)-yl)benzylcarbamate C24 as a yellow solid.

[0180] <Preparation of intermediate C25> [ka] <Ethyl N-[(4-bromophenyl)carbamate]carbamate> To a solution of 4-bromoaniline (5.00 g, 29.1 mmol, 1.0 equivalent) in SiO2 (50 mL), ethyl N-(thioxomethylene)carbamate (3.81 g, 29.1 mmol, 1.0 equivalent) and TMEDA (338 mg, 2.91 mmol, 439 μL, 0.1 equivalent) were added at room temperature. The reaction mixture was stirred at 20°C for 12 hours. The reaction mixture was concentrated under reduced pressure to obtain a residue. This residue was triturated with EtOH (25 mL) for 0.5 hours. The mixture was filtered, the filtration cake was washed with EtOH (2 × 25 mL), and dried to obtain 7.5 g of ethyl N-[(4-bromophenyl)carbamotioil]carbamate as a white solid.

[0181] <(Z)-Ethyl(((4-bromophenyl)amino)((2-oxoethyl)thio)methylene)carbamate> To a solution of ethyl N-[(4-bromophenyl)carbamotioyl]carbamate (7.50 g, 24.7 mmol, 1.0 equivalent) in CH3CN (100 mL), Cs2CO3 (13.70 g, 42.0 mmol, 1.7 equivalents) and 2-chloroacetaldehyde (6.07 g, 30.9 mmol, 4.97 mL, 1.25 equivalents) were added at room temperature. The reaction mixture was stirred under N2 at 20°C for 12 hours. Cs2CO3 (8.06 g, 24.7 mmol, 1.0 equivalent) and 2-chloroacetaldehyde (3.64 g, 18.6 mmol, 2.98 mL, 0.75 equivalents) were added to the reaction solution. The reaction mixture was stirred under N2 at 20°C for 4 hours. The reaction mixture was quenched by adding water (300 mL) and extracted with ethyl acetate (3 × 300 mL). The organic layers were combined and dehydrated with Na₂SO₄, filtered, and concentrated under reduced pressure to obtain 10 g of (Z)-ethyl(((4-bromophenyl)amino)((2-oxoethyl)thio)methylene)carbamate as a pale yellow solid. LCMS (m / z[M+H] + ): 345.0.

[0182] <(Z)-Ethyl(3-(4-bromophenyl)thiazole-2(3H)-ylidene)carbamate> A solution of (Z)-ethyl(((4-bromophenyl)amino)((2-oxoethyl)thio)methylene) carbamate (10.00 g, 28.97 mmol, 1.0 equivalent) in THF (120 mL) was mixed with DIPEA (11.23 g, 86.90 mmol, 15.14 mL, 3.0 equivalent) and SOCl2 (3.45 g, 28.97 mmol, 2.10 mL, 1.0 equivalent) at 0°C. The reaction mixture was stirred at 15°C for 4 hours. The reaction mixture was quenched by adding saturated NaHCO3 solution (200 mL) and extracted with ethyl acetate (3 × 200 mL). The organic layers were combined and dehydrated with Na2SO4, filtered, and concentrated under reduced pressure to obtain the residue. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 3 / 1 → 1 / 1) to obtain 3.60 g of (Z)-ethyl (3-(4-bromophenyl)thiazole-2(3H)-ylidene) carbamate compound as a yellow solid.1 H NMR (400 MHz, DMSO-d6) δ 7.75 (d, J=8.63 Hz, 2H), 7.47-7.56 (m, 3H), 7.08 (d, J=4.88 Hz, 1H), 4.03 (q, J=7.09 Hz, 2H), 1.16 (t, J=7.07 Hz, 3H).

[0183] <Intermediate C25> (Z)-ethyl(3-(4-bromophenyl)thiazole-2(3H)-ylidene)carbamate (1.00 g, 3.06 mmol, 1.0 equivalent) was dissolved in EtOH (10 mL) and 6N NaOH (2.44 g, 61.1 mmol, 20 equivalents) was added. The reaction mixture was stirred under N2 at 50°C for 1.5 hours. This reaction was combined with a preceding batch (using 100 mg of the starting material). The reaction mixture was quenched by adding water (100 mL) and extracted with ethyl acetate (3 × 100 mL). The organic layers were combined and dehydrated with Na₂SO₄, filtered, and concentrated under reduced pressure to obtain 750 mg of 3-(4-bromophenyl)thiazole-2(3H)-imine C25 as a yellow oil.

[0184] <Preparation of intermediate C26> [ka] <Ethyl N-((cyclohexylmethyl)carbamate)carbamate> To a solution of cyclohexylmethaneamine (2.00 g, 17.7 mmol, 1.0 equivalent) in ethyl acetate (20 mL), O-ethylcarbon isothiocyanate (2.09 mL, 17.7 mmol, 1.0 equivalent) and tetramethylethylenediamine (0.267 mL, 1.77 mmol, 0.1 equivalent) were added at 25°C. The mixture was then stirred at this temperature for 5 hours. The mixture was concentrated under reduced pressure, and the residue was triturated twice with ethanol (10 mL) to obtain 3.50 g of ethyl N-((cyclohexylmethyl)carbamate)carbamate as an off-white solid. 1H NMR (400 MHz, DMSO-d6) δ 10.94 (br. s, 1H), 9.92 (br. t, J = 5.2 Hz, 1H), 4.14 (q, J = 7.2 Hz, 2H), 3.42 - 3.39 (m, 2H), 2.83 (s, 1H), 1.69 - 1.60 (m, 6H), 1.22 - 1.10 (m, 5H), 0.98 - 0.90 (m, 2H). LCMS (m / z [M+H] + ): 244.9.

[0185] <(Z)-Ethyl(3-(cyclohexylmethyl)-4-methylthiazole-2(3H)-ylidene)carbamate> To a solution of ethyl N-((cyclohexylmethyl)carbamate) carbamate (1.00 g, 4.09 mmol, 1.0 equivalent) and cesium carbonate (2.27 g, 6.96 mmol, 1.7 equivalents) in acetonitrile (10 mL), 1-chloropropan-2-one (0.568 g, 6.14 mmol, 1.5 equivalents) was added dropwise at 25°C. After addition, the mixture was stirred at this temperature for 2 hours. The mixture was poured into water (30 mL) and extracted with ethyl acetate (50 mL x 3). The organic layers were washed together with brine (25 mL x 2), dehydrated with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 10 / 1 → 5 / 1) to obtain 0.80 g of (Z)-ethyl(3-(cyclohexylmethyl)-4-methylthiazole-2(3H)-ylidene)carbamate as a yellow oil. 1 H NMR (400 MHz, DMSO-d6) δ 6.44 (s, 1H), 4.01 - 3.99 (m, 2H), 3.22 - 3.09 (m, 2H), 3.08 - 3.05 (m, 1H), 1.65 - 1.61 (m, 6H), 1.48 (s, 3H), 1.15 - 1.14 (m, 3H), 1.12 - 1.10 (m, 2H), 0.90 - 0.88 (m, 2H). LCMS (m / z [M+H] + ): 282.9.

[0186] <Intermediate C26> A solution of (Z)-ethyl(3-(cyclohexylmethyl)-4-methylthiazole-2(3H)-ylidene) carbamate (0.350 g, 1.08 mmol, 1.0 equivalent) in ethanol (3 mL) was to be mixed with 6N sodium hydroxide (0.862 g, 21.5 mmol, 20 equivalents) at 25°C in a single addition. The reaction mixture was heated to 50°C and stirred for 1 hour. The mixture was added to water (15 mL) and extracted with ethyl acetate (30 mL x 3). The organic phases were combined and washed with brine (25 mL x 2), dehydrated with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 10 / 1 → 0 / 1) to obtain 150 mg of 3-(cyclohexylmethyl)-4-methylthiazole-2(3H)-imine C26 as a yellow oil.

[0187] <Preparation of intermediate C27> [ka] <Ethyl N-(cyclohexylcarbamate)carbamate> A mixture of cyclohexaneamine (0.800 g, 8.07 mmol, 1.0 equivalent) and O-ethylcarbon isothiocyanate (1.11 g, 8.47 mmol, 1.05 equivalent) in ethyl acetate (10 mL) was mixed with TMEDA (93 mg, 0.806 mmol, 1.0 equivalent) at 25°C. The mixture was stirred at this temperature for 6 hours and then concentrated under reduced pressure. The residue was purified by silica gel column chromatography (column height: 250 mm, diameter: 100 mm, silica gel of 100-200 mesh, petroleum ether / ethyl acetate = 100 / 1 → 3 / 1) to obtain 1.80 g of ethyl N-(cyclohexylcarbamotioil)carbamate as a yellow, rubbery substance. 1H NMR (400 MHz, CD3Cl) δ 9.64 (br. s, 1H), 7.96 (br. s, 1H), 4.29 - 4.22 (m, 2H), 2.09 - 2.01 (m, 2H), 1.77 - 1.68 (m, 2H), 1.62-1.63 (m, 1H), 1.49 - 1.33 (m, 4H), 1.33 - 1.19 (m, 5H). LCMS (m / z [M+H] + ): 231.1.

[0188] <(Z)-Ethyl((Cyclohexylamino)((2-Oxopropyl)thio)methylene)carbamate> A mixture of ethyl N-(cyclohexylcarbamotioil)carbamate (0.800 g, 3.47 mmol, 1.0 equivalent) and cesium carbonate (1.92 g, 5.90 mmol, 1.7 equivalents) in acetonitrile (10 mL) was mixed with 1-chloropropan-2-one (0.353 g, 3.82 mmol, 1.1 equivalents) at 25°C. The resulting mixture was stirred at this temperature for 4 hours, then poured into ice water (10 mL) and extracted with MTBE (100 mL x 2). The organic phases were combined and washed with brine (20 mL x 2), dehydrated with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain 900 mg of crude (Z)-ethyl((cyclohexylamino)((2-oxopropyl)thio)methylene)carbamate as a yellow solid, which was used directly in the next step without purification. LCMS (m / z [M+H] + ): 287.1.

[0189] <(Z)-Ethyl(3-cyclohexyl-4-methylthiazole-2(3H)-ylidene)carbamate> A mixture of (Z)-ethyl((cyclohexylamino)((2-oxopropyl)thio)methylene)carbamate (0.900 g, 3.14 mmol, 1.0 equivalent) and diisopropylethylamine (0.812 g, 6.29 mmol, 2.2 equivalents) in tetrahydrofuran (20 mL) was mixed with thionyl chloride (0.224 g, 1.89 mmol, 0.6 equivalents) at 0°C. The mixture was stirred at 25°C for 2 hours, then poured into ice water (20 mL) and extracted with ethyl acetate (100 mL x 3). The organic phases were combined and washed with brine (20 mL), dehydrated with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 100 / 1 → 4 / 1) to obtain 0.50 g of (Z)-ethyl(3-cyclohexyl-4-methylthiazole-2(3H)-ylidene) carbamate as a yellow solid. 1 H NMR (400 MHz, DMSO-d6) δ 6.54 (br. s, 1H), 4.09-4.03 (m, 2H), 2.29 - 2.25 (m, 3H), 1.82 - 1.62 (m, 6H), 1.36 - 1.23 (m, 3H), 1.21 - 1.19 (m, 5H).LCMS (m / z [M+H] + ): 268.9.

[0190] <Intermediate C27> A mixture of (Z)-ethyl(3-cyclohexyl-4-methylthiazole-2(3H)-ylidene) carbamate (0.200 g, 0.745 mmol, 1.0 equivalent) in ethanol (4 mL) was mixed with NaOH (0.596 g, 14.90 mmol, 20 equivalents) at 25°C. The mixture was heated to 50°C and stirred for 1 hour, then poured into ice water (50 mL) and extracted with ethyl acetate (50 mL x 5). The organic phases were combined and washed with brine (10 x 2 mL), dehydrated with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain 120 mg of 3-cyclohexyl-4-methylthiazole-2(3H)-imine C27 as a yellow solid.

[0191] <Preparation of intermediate C28> [ka] <Ethyl N-[(2-chlorophenyl)carbamate]carbamate> To a solution of 2-chloroaniline (8.26 mL, 78.4 mmol) in ethyl acetate (100 mL), 10.3 g, 78.4 mmol of o-ethylcarbon isothiocyanate and 1.18 mL, 7.84 mmol of tetramethylethylenediamine were gradually added at 0°C, and the mixture was then stirred at 50°C for 5 hours. The reaction mixture was cooled to room temperature and quenched by adding 100 mL of methanol dropwise. The precipitated solid was collected by filtration and then triturated with methanol (100 mL) at 25°C for 10 minutes to obtain 16.5 g of ethyl N-[(2-chlorophenyl)carbamotioil]carbamate as a white solid. 1 H NMR (DMSO-d6, 400 MHz) δ 11.60 (br. s, 1H), 11.47 (br. s, 1H), 7.94 (d, J = 8.0 Hz, 1H), 7.57 - 7.55 (m, 1H), 7.41 - 7.36 (m, 1H), 7.33 - 7.29 (m, 1H), 4.23 (q, J = 7.2 Hz, 2H), 1.27 (t, J = 7.2 Hz, 3H). LCMS (m / z [M+H] + ): 258.9.

[0192] <(Z)-methyl3-((N-(2-chlorophenyl)-N'-(ethoxycarbonyl)carbamimidoyl)thio)-2-oxopropanoate> To a solution of ethyl N-[(2-chlorophenyl)carbamotioil]carbamate (5.00 g, 19.3 mmol) and cesium carbonate (7.56 g, 23.2 mmol) in acetonitrile (100 mL), methyl 3-bromo-2-oxopropanoate (2.67 mL, 25.1 mmol) was added at 25°C, and the mixture was stirred at 25°C for 2 hours. The reaction mixture was quenched with water (100 mL) at 25°C and extracted with ethyl acetate (200 mL x 3). The organic layers were washed together with brine (200 mL x 2), dehydrated with anhydrous sodium 2SO4, and filtered. The filtrate was concentrated under reduced pressure. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 10 / 1 to 2 / 1) to obtain 2.80 g of (Z)-methyl3-((N-(2-chlorophenyl)-N'-(ethoxycarbonyl)carbamimidoyl)thio)-2-oxopropanoate as a brown oil. 1 H NMR (CDCl3, 400 MHz) δ 7.53 - 7.49 (m, 1H), 7.47 - 7.44 (m, 1H), 7.35 - 7.31 (m, 2H), 4.90 (br. s, 1H), 4.16 (q, J = 7.2 Hz, 2H), 4.00 (d, J = 11.6 Hz, 1H), 3.75 (s, 3H), 3.47 - 3.42 (m, 1H), 1.26 (t, J = 7.2 Hz, 3H). LCMS (m / z [M+H] + ): 358.9.

[0193] <(Z)-methyl3-(2-chlorophenyl)-2-((ethoxycarbonyl)imino)-2,3-dihydrothiazole-4-carboxylate> (Z)-methyl 3-((N-(2-chlorophenyl)-N'-(ethoxycarbonyl)carbamimidoyl)thio)-2-oxopropanoate (2.80 g, 7.80 mmol) and DIPEA (2.04 mL, 11.7 mmol) in THF (15 mL) were mixed with thionyl chloride (0.85 mL, 11.7 mmol) at 0°C. The mixture was stirred under a nitrogen atmosphere at 25°C for 2 hours. The reaction mixture was quenched with water (100 mL) at 25°C and extracted with ethyl acetate (200 mL x 3). The organic layers were washed together with brine (200 mL x 2), dehydrated with anhydrous Na2SO4, and filtered. The filtrate was concentrated under reduced pressure to obtain 2.20 g of (Z)-methyl 3-(2-chlorophenyl)-2-((ethoxycarbonyl)imino)-2,3-dihydrothiazole-4-carboxylate as a yellow oil, which was used directly in the next step without further purification. 1 H NMR (CDCl3, 400 MHz) δ 7.64 (s, 1H), 7.54 - 7.51 (m, 1H), 7.46 - 7.41 (m, 2H), 7.40 - 7.37 (m, 1H), 4.20 (q, J = 7.2 Hz, 2H), 3.74 (s, 3H), 1.29 (t, J = 7.2 Hz, 3H). LCMS (m / z [M+H] + ): 340.9.

[0194] <(Z)-Ethyl(3-(2-chlorophenyl)-4-(methylcarbamoyl)thiazole-2(3H)-ylidene)carbamate> A mixture of (Z)-methyl 3-(2-chlorophenyl)-2-((ethoxycarbonyl)imino)-2,3-dihydrothiazole-4-carboxylate (1.5 g, 4.40 mmol) in THF (15 mL) was mixed with methylamine (2 M, 22 mL) dropwise, and 3,4,6,7,8,9-hexahydro-2H-pyrimido[1,2-a]pyrimidine (0.306 g, 2.20 mmol) was added at 25 °C. The mixture was stirred under a nitrogen atmosphere at 60 °C for 2 hours, then gradually poured into ice water (50 mL), and subsequently extracted with ethyl acetate (100 mL x 3). The organic layers were washed together with brine (200 mL x 2), dehydrated with anhydrous Na2SO4, and filtered. The filtrate was concentrated under reduced pressure to obtain 1.55 g of (Z)-ethyl(3-(2-chlorophenyl)-4-(methylcarbamoyl)thiazole-2(3H)-ylidene)carbamate as a yellow solid. 1 H NMR (DMSO-d6, 400 MHz) δ8.63 (br. d, J = 4.8 Hz, 1H), 7.59 (d, J = 7.6 Hz, 1H), 7.54 (s, 1H), 7.49 - 7.43 (m, 3H), 4.02 (q, J = 7.2 Hz, 2H), 2.57 (d, J = 4.8 Hz, 3H), 1.19 - 1.04 (t, J = 7.2 Hz, 3H). LCMS (m / z [M+H] + ): 339.9.

[0195] <Intermediate C28> To a solution of (Z)-ethyl(3-(2-chlorophenyl)-4-(methylcarbamoyl)thiazole-2(3H)-ylidene) carbamate (0.360 g, 1.06 mmol) in DMF (7 mL), bromotrimethylsilane (0.687 mL, 5.30 mmol) was added at 25°C. The reaction mixture was stirred at 80°C for 1 hour, and then gradually poured into ice water (20 mL). The mixture was neutralized with saturated sodium bicarbonate aqueous solution. The mixture was directly purified by reverse-phase flush (0.1% NH3·H2O / MeCN / water) to obtain 80 mg of 3-(2-chlorophenyl)-2-imino-N-methyl-2,3-dihydrothiazole-4-carboxamide C28 as a yellow solid.

[0196] <Preparation of intermediate C29> [ka] <Preparation of intermediate C29> <Ethyl(Z)-(3-(2-chlorophenyl)-4-(dimethylcarbamoyl)thiazole-2(3H)-ylidene)carbamate> A mixture of methyl(Z)-3-(2-chlorophenyl)-2-((ethoxycarbonyl)imino)-2,3-dihydrothiazole-4-carboxylate (0.380 g, 1.12 mmol) and dimethylamine (2 M, 5.6 mL) in THF (1 mL) was to be combined with 3,4,6,7,8,9-hexahydro-2H-pyrimido[1,2-a]pyrimidine (0.078 g, 0.560 mmol) at 25°C under a nitrogen atmosphere. The reaction mixture was stirred at 60°C for 2 hours and then carefully poured into 100 mL of ice water. The aqueous phase was extracted with ethyl acetate (50 mL x 3) and then washed with brine (10 mL). The organic layer was dehydrated with anhydrous Na₂SO₄ and filtered. The filtrate was concentrated under reduced pressure to obtain a residue, which was purified by reverse-phase flushing (0.1% trifluoroacetic acid / MeCN / water) to obtain 140 mg of ethyl(Z)-[3-(2-chlorophenyl)-4-(dimethylcarbamoyl)thiazole-2-ylidene]carbamate as a yellow solid. 1H NMR (DMSO-d6, 400 MHz) δ 7.63 - 7.61 (m, 1H), 7.52 - 7.43 (m, 3H), 7.36 (s, 1H), 4.04 (m, 2H), 3.11 (s, 3H), 2.76 (s, 3H), 1.14 (t, J = 7.2 Hz, 3H). LCMS (m / z[M+H] + ): 353.9.

[0197] <Intermediate C29> To a solution of ethyl(Z)-[3-(2-chlorophenyl)-4-(dimethylcarbamoyl)thiazole-2-ylidene]carbamate (0.390 g, 1.10 mmol) in ethyl alcohol (4 mL), solid sodium hydroxide (0.960 g, 24.0 mmol) was added. The mixture was stirred at 50°C for 2 hours. The crude product was purified directly by reverse-phase flush (0.1% ammonium hydroxide / MeCN / water) without workup to obtain 90 mg of 3-(2-chlorophenyl)-2-imino-N,N-dimethylthiazole-4-carboxamide C29 as a yellow solid.

[0198] <Preparation of Intermediate D1> [ka] <2-phenylacetyl chloride> A solution of 2-phenylacetic acid (5.00 g, 36.7 mmol, 4.63 mL, 1.0 equivalent) in DCM (50 mL) was mixed with DMF (268 mg, 3.67 mmol, 283 μL, 0.1 equivalent), and then SOCl2 (8.74 g, 73.4 mmol, 5.33 mL, 2.0 equivalent) was added to the mixture at 0°C. The reaction mixture was stirred at 25°C for 1 hour. The reaction mixture was concentrated under reduced pressure to remove the solvent. Compound 2-phenylacetyl chloride (5.68 g, crude) was obtained as a colorless oil.

[0199] <1-Diazo-3-phenylpropan-2-one> To a solution of 2-phenylacetyl chloride (5.68 g, 36.7 mmol, 4.90 mL, 1.0 equivalent) in MeCN (50 mL), TMSCHN2 (2 M, 36.74 mL, 2 equivalents) was added at 0°C. The reaction mixture was then stirred at 25°C for 2 hours. The resulting yellow solution was used directly in the next step without further purification.

[0200] <1-Bromo-3-phenylpropan-2-one> To a solution of 1-diazo-3-phenylpropan-2-one in MeCN, HBr (13.5 g, 55.1 mmol, 9.06 mL, 1.5 equivalents) (33% AcOH solution) was added. The mixture was stirred at 25°C for 12 hours. The mixture was poured into water (50 mL) and extracted with ethyl acetate (3 × 30 mL). The organic phase was washed with brine (50 mL), dehydrated with Na₂SO₄, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 0 / 1 → 6 / 1) to obtain 2.30 g of 1-bromo-3-phenylpropan-2-one as a red oil.

[0201] <Intermediate D1> To a solution of thiourea (715 mg, 9.39 mmol, 1.0 equivalent) in refluxed EtOH (20 mL), 1-bromo-3-phenylpropan-2-one (2.00 g, 9.39 mmol, 1.0 equivalent) and pyridine (742 mg, 9.39 mmol, 758 μL, 1.0 equivalent) were added. The reaction mixture was stirred at 70°C for 4 hours. The reaction mixture was concentrated under reduced pressure to remove the solvent. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 50 / 1 → 1 / 1) to obtain 0.6 g of crude product. 490 mg of the crude product was purified by preparative HPLC to obtain 160 mg of 4-benzylthiazole-2(3H)-imine D1 as a white solid. 1 1H NMR: (400 MHz, DMSO-d6) δ 7.31-7.12 (m, 5H), 6.83 (s, 2H), 6.12 (s, 1H), 3.71 (s, 2H). The structure was confirmed by HMBC and HSQC.

[0202] <Preparation of intermediate D3> [ka] To a solution of benzylthiourea (1.00 g, 6.02 mmol, 1.0 equivalent) in acetone (15 mL), HCl (1.20 g, 12.0 mmol, 1.18 mL, 36% purity, 2.0 equivalents) and DMSO (939 mg, 12.0 mmol, 939 μL, 2.0 equivalents) were added at 40°C. The reaction mixture was stirred for 48 hours. The reaction mixture was directly concentrated under reduced pressure. The residue was purified by reverse-phase MPLC to obtain 0.80 g of 3-benzyl-4-methylthiazole-2(3H)-imine D3 as a white solid.

[0203] <Preparation of Acid D> [ka] <4-methyl-1H-pyrrolo[2,3-b]pyridine> To a solution of 4-chloro-1H-pyrrolo[2,3-b]pyridine (10.00 g, 65.54 mmol, 1.0 equivalent) and Pd(dppf)Cl2 (1.20 g, 1.64 mmol, 0.025 equivalents) in toluene (200 mL), MeMgBr (3 M, 109 mL, 5.0 equivalents) was added dropwise at 25°C. After addition, the reaction mixture was stirred at 80°C for 12 hours, then cooled to 25°C, quenched by adding 0°C ice water (300 mL), and extracted with ELISA (2 × 200 mL). The organic layers were washed together with saturated NH4Cl aqueous solution (400 mL) and brine (200 mL), dehydrated with Na2SO4, and filtered. The filtrate was concentrated under reduced pressure to obtain the residue. The residue was purified by silica gel flash chromatography (0-100% ethyl acetate / petroleum ether gradient, elution at 100 mL / min) to obtain 7.00 g of 4-methyl-1H-pyrrolo[2,3-b]pyridine as a white solid. LCMS (m / z [M+H] + ): 133.1.

[0204] <2,2,2-Trichloro-1-(4-methyl-1H-pyrrolo[2,3-b]pyridin-3-yl)ethanone> To a solution of 4-methyl-1H-pyrrolo[2,3-b]pyridine (1.00 g, 7.57 mmol, 1.0 equivalent) in DCM (20 mL), AlCl3 (2.52 g, 18.9 mmol, 2.5 equivalents) was added at 25°C. The reaction mixture was stirred at 25°C for 10 minutes, and then 2,2,2-trichloroacetyl chloride (2.06 g, 11.4 mmol, 1.5 equivalents) was added. The reaction mixture was stirred at 25°C for a further 3 hours and 50 minutes. The mixture was poured into ice water (30 mL), and the precipitate was filtered. The filtrate was extracted with DCM (2 × 30 mL), concentrated under reduced pressure, and 2.0 g of crude 2,2,2-trichloro-1-(4-methyl-1H-pyrrolo[2,3-b]pyridine-3-yl)etanone was obtained as a pale yellow solid.

[0205] <4-methyl-1H-pyrrolo[2,3-b]pyridine-3-carboxylic acid (acid D)> A mixture of 2,2,2-trichloro-1-(4-methyl-1H-pyrrolo[2,3-b]pyridine-3-yl)ethanone (2.00 g, 7.21 mmol, 1.0 equivalent) in an aqueous NaOH solution (3 M, 30 mL, 12.5 equivalents) was stirred at 25°C for 3 hours under an N2 atmosphere. The mixture was cooled to 0°C and the pH was adjusted to <6.0 by adding concentrated aqueous HCl. The precipitate was collected by filtration, the filtration cake was washed with H2O (2 × 5 mL), and dried under vacuum to obtain 1.2 g of crude 4-methyl-1H-pyrrolo[2,3-b]pyridine-3-carboxylic acid (acid D) as an off-white solid.

[0206] <Preparation of Example T01> [ka] EDCI (1.81 g, 9.46 mmol, 1.5 equivalents) was added to a mixture of 3-benzylthiazole-2(3H)-imine B1 (1.20 g, 6.31 mmol, 1.0 equivalent), 1H-pyrrolo[2,3-b]pyridine-3-carboxylic acid A (0.818 g, 5.05 mmol, 0.8 equivalent), HOBt (1.28 g, 9.46 mmol, 1.5 equivalent), and diisopropylethylamine (2.45 g, 18.9 mmol, 3.0 equivalent) in dimethylformamide (10 mL). The mixture was stirred at 25°C for 1.5 hours and then poured into ice water (200 mL). The precipitate was collected by filtration, the filtered cake was washed with acetonitrile (3 × 30 mL), then triturated with acetonitrile / methanol = 5:1 (5 × 30 mL), followed by reverse-phase HPLC and recrystallization from methanol (2 × 250 mL) to obtain 352 mg of (Z)-N-(3-benzylthiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T01 as a white solid. 1 H-NMR (DMSO-d6, 400 MHz) δ12.18 (br. s, 1H), 8.53 - 8.50 (m, 1H), 8.25 - 8.10 (m, 2H), 7.60 (d, J= 4.8 Hz, 1H), 7.42 - 7.40 (m, 5H), 7.36 (d, J = 7.2 Hz, 1H), 6.96 (d, J= 4.8 Hz, 1H), 5.52 (s, 2H). LCMS (m / z [M+H] + ) = 335.1.

[0207] The examples in Table E below were prepared using intermediate B1 and the appropriate acid in Table A, in the same manner as shown above in Scheme 54. Table E [Table 5]

[0208] The examples in Table F are prepared using appropriate intermediates from Tables B / C and Acid A from Table A, in the same manner as described above in Scheme 54. Table F [Table 6] TIFF2023512823000076.tif215141TIFF2023512823000077.tif215141TIFF20235128230 00078.tif204141TIFF2023512823000079.tif211141TIFF2023512823000080.tif184141

[0209] Further examples of the preparation of Table G are shown in Schemes 55-71. Table G [Table 7] TIFF2023512823000082.tif214141TIFF2023512823000083.tif188141TIFF2023512823000084.tif187141TIFF2023512823000085.tif45128

[0210] <Preparation of Examples T37 and T38> [ka] <Example T37> To a solution of 1H-pyrrolo[2,3-b]pyridine-3-carboxylic acid A (199 mg, 1.23 mmol, 1.0 equivalent) in DCM (10 mL), oxalyl chloride (COCl)2 (156 mg, 1.23 mmol, 107 μL, 1.0 equivalent) and DMF (1 μL, 12.3 μmol, 0.01 equivalent) were added at 0°C. The reaction mixture was stirred at 15°C for 2 hours, and then TEA (187 mg, 1.84 mmol, 1.5 equivalents) and 3-methylthiazole-2-imine B2 (140 mg, 1.23 mmol, 1.0 equivalent) were added. The resulting mixture was stirred at 15°C for 12 hours. The reaction mixture was quenched by adding H2O (20 mL) and extracted with RINKAN (3 × 15 mL). The organic layers were washed together with brine (3 × 10 mL), dehydrated with anhydrous Na2SO4, filtered, and concentrated under reduced pressure. The residue was purified by preparative HPLC to obtain 21 mg of (Z)-N-(3-methylthiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T37 as a white solid.

[0211] <Example T38> To a solution of 1H-pyrrolo[2,3-b]pyridine-3-carboxylic acid A (379 mg, 2.34 mmol, 1.0 equivalent) in DCM (10 mL), oxalyl chloride (COCl)2 (297 mg, 2.34 mmol, 1.0 equivalent) and DMF (2 μL, 23.4 μmol, 0.01 equivalent) were added at 0°C. The reaction mixture was stirred at 15°C for 2 hours, and then TEA (355 mg, 3.51 mmol, 1.5 equivalent) and 3-ethylthiazole-2-imine B3 (300 mg, 2.34 mmol, 1.0 equivalent) were added. The resulting mixture was stirred at 15°C for 12 hours. The reaction mixture was quenched by adding water (30 mL) and extracted with RINKAN (3 × 20 mL). The organic layers were washed together with brine (2 × 20 mL), dehydrated with anhydrous Na2SO4, filtered, and concentrated under reduced pressure. The residue was purified by preparative HPLC to obtain 24 mg of (Z)-N-(3-ethylthiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T38 as a white solid.

[0212] <Preparation of Example T39> [ka] <(Z)-N-(thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide> To a solution of 1H-pyrrolo[2,3-b]pyridine-3-carboxylic acid A (0.60 g, 3.70 mmol, 1.0 equivalent) in pyridine (5.88 g, 74.3 mmol, 20 equivalents), HBTU (2.81 g, 7.40 mmol, 2.0 equivalents) and thiazole-2-amine (556 mg, 5.55 mmol, 1.5 equivalents) were added. The reaction mixture was heated to 100°C and stirred at 100°C for 12 hours. The reaction mixture was cooled to 0°C and poured into ice water (15 mL). Solid matter was recovered by filtration. The filtration cake was washed with water (2 × 5 mL) and dried under reduced pressure to obtain the crude product. The crude product was recrystallized by refluxing MeOH / EtOH (4 mL / 4 mL) at 60°C for 0.5 hours, then cooled to 25°C and filtered. The solid was washed with cold EtOH (5 mL) and dried in vacuum to obtain 0.41 g of (Z)-N-(thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide as a brown solid.

[0213] <(Z)-N-(3-(1-phenylethyl)thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T39> (Z)-N-(thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide (0.20 g, 819 ml, 1.0 equivalent) was dissolved in DMF (3 mL) and K2CO3 (226 mg, 1.64 mmol, 2.0 equivalent) and 1-chloroethylbenzene (173 mg, 1.23 mmol, 1.5 equivalent) were added. The reaction mixture was stirred at 25°C for 12 hours. The reaction mixture was heated at 60°C for 18 hours. The reaction mixture was cooled to room temperature and filtered to obtain a yellow solution in DMF. The yellow solution was purified by preparative HPLC to obtain 25 mg of (Z)-N-(3-(1-phenylethyl)thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T39 as a pale yellow solid.

[0214] <Preparation of Example T41> [ka] <(Z)-tert-butyl(4-(2-((1H-pyrrolo[2,3-b]pyridine-3-carbonyl)imino)thiazole-3(2H)-yl)butan-2-yl)carbamate> To a solution of 1H-pyrrolo[2,3-b]pyridine-3-carboxylic acid A (30 mg, 184 μmol, 1.0 equivalent), HOBt (37 mg, 276 μmol, 1.5 equivalents), EDCI (53 mg, 276 μmol, 1.5 equivalents), and DIPEA (71 mg, 553 μmol, 96 μL, 3.0 equivalents) in DMF (1 mL), tert-butyl(4-(2-iminothiazole-3(2H)-yl)butan-2-yl)carbamate B4 (50 mg, 184 μmol, 1.0 equivalent) was added. The reaction mixture was stirred at 25°C for 12 hours. The reaction mixture was diluted with 20 mL of water and extracted with RINKAN (3 × 10 mL). The organic layers were washed together with 20 mL of brine, dehydrated with Na₂SO₄, filtered, and concentrated under reduced pressure. The residue was purified by preparative TLC (petroleum ether / ethyl acetate) of silica gel to obtain 40 mg of (Z)-tert-butyl(4-(2-((1H-pyrrolo[2,3-b]pyridine-3-carbonyl)imino)thiazole-3(2H)-yl)butan-2-yl)carbamate as a white solid.

[0215] <(Z)-N-(3-(3-aminobutyl)thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T41> A mixture of (Z)-tert-butyl(4-(2-((1H-pyrrolo[2,3-b]pyridine-3-carbonyl)imino)thiazole-3(2H)-yl)butan-2-yl)carbamate (40 mg, 96 μmol, 1.0 equivalent) in HCl / Â (4M, 2 mL) was stirred at 25°C for 1 hour. The reaction mixture was concentrated under reduced pressure. The residue was purified by preparative HPLC to obtain 28 mg of (Z)-N-(3-(3-aminobutyl)thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T41 as a white solid.

[0216] <Preparation of Examples T42 and T43> [ka] <(Z)-N-(3-(2-cyanobenzyl)thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T42> To a solution of 1H-pyrrolo[2,3-b]pyridine-3-carboxylic acid A (369 mg, 2.28 mmol, 1.0 equivalent) in DMF (6 mL), EDCI (524 mg, 2.73 mmol, 1.2 equivalents), HOBt (369 mg, 2.73 mmol, 1.2 equivalents), TEA (691 mg, 6.83 mmol, 3.0 equivalents), and 2-((2-iminothiazole-3(2H)-yl)methyl)benzonitrile B11 (0.49 g, 2.28 mmol, 1.0 equivalent) were added. The reaction mixture was stirred at 25°C for 12 hours. The reaction mixture was quenched by adding 15 mL of water at 0°C, the precipitate was collected by filtration, washed with 2 × 5 mL of water, and dried under vacuum to obtain 0.60 g of the crude product (Z)-N-(3-(2-cyanobenzyl)thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T42 as a gray solid. 180 mg of the crude product T42 was further purified by preparative HPLC to obtain 92 mg of pure (Z)-N-(3-(2-cyanobenzyl)thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T42 as a pale yellow solid.

[0217] <(Z)-N-(3-(2-(aminomethyl)benzyl)thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T43> To a solution of LAH (48 mg, 1.25 mmol, 3.0 equivalents) in THF (4 mL), (Z)-N-(3-(2-cyanobenzyl)thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T42 (0.15 g, 417 μmol, 1.0 equivalent) was added in small amounts at 0°C. After addition, the reaction mixture was gradually warmed to 25°C and stirred at 25°C for 4 hours. The reaction mixture was quenched by adding HCl aqueous solution (2N, 0.5 mL) at 0°C and stirred for 0.5 hours. The mixture was diluted with THF / DMF (20 mL / 4 mL) and filtered. The filtered cake was washed with MeOH (2 × 5 mL). The filtrates were combined and concentrated under reduced pressure. The residue was purified by preparative HPLC to obtain 8.3 mg of (Z)-N-(3-(2-(aminomethyl)benzyl)thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T43 as an off-white solid.

[0218] <Preparation of Examples T44 and T45> [ka] <(Z)-N-(3-(3-cyanobenzyl)thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T44> To a solution of 1H-pyrrolo[2,3-b]pyridine-3-carboxylic acid A (264 mg, 1.63 mmol, 1.0 equivalent), HOBt (330 mg, 2.44 mmol, 1.5 equivalent), EDCI (468 mg, 2.44 mmol, 1.5 equivalent), and DIPEA (630 mg, 4.88 mmol, 850 μL, 3.0 equivalent) in DMF (8 mL), 3-((2-iminothiazole-3(2H)-yl)methyl)benzonitrile B17 (350 mg, 1.63 mmol, 1.0 equivalent) was added at 25°C. The reaction mixture was then stirred at 25°C for 12 hours. The reaction mixture was diluted with 50 mL of water and extracted with RINKAN (3 × 30 mL). The organic layers were washed together with 50 mL of brine, dehydrated with Na₂SO₄, filtered, and concentrated under reduced pressure. The residue was diluted with MeOH (20 mL) and stirred for 20 minutes; the suspension was filtered to collect the solid. The collected solid was washed with MeOH (10 mL) and then dried under vacuum to obtain 500 mg of (Z)-N-(3-(3-cyanobenzyl)thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T44 as a yellow solid. 100 mg of the crude product was purified by preparative HPLC to obtain 62 mg of pure (Z)-N-(3-(3-cyanobenzyl)thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T44 as a white solid.

[0219] <(Z)-N-(3-(3-(aminomethyl)benzyl)thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T45> (Z)-N-(3-(3-cyanobenzyl)thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T44 (200 mg, 556 μmol, 1.0 equivalent) was suspended in THF (5 mL) and LAH (63 mg, 1.67 mmol, 3.0 equivalents) was added at 0°C under N2 conditions. The reaction mixture was then stirred at 25°C for 3 hours. The reaction mixture was quenched by adding water (5 mL) and acidified to pH=3 with HCl (4N), at which point the mixture became clear. The mixture was directly purified by preparative HPLC to obtain 76 mg of (Z)-N-(3-(3-(aminomethyl)benzyl)thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T45 as a white solid.

[0220] <Preparation of Examples T46, T47, and T48> [ka] <(Z)-N-(3-(4-cyanobenzyl)thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T46> To a solution of 1H-pyrrolo[2,3-b]pyridine-3-carboxylic acid A (1.09 g, 6.75 mmol, 1.0 equivalent), HOBt (1.37 g, 10.1 mmol, 1.5 equivalent), EDCI (1.94 g, 10.1 mmol, 1.5 equivalent), and DIPEA (2.62 g, 20.3 mmol, 3.5 mL, 3.0 equivalent) in DMF (20 mL), 4-((2-iminothiazole-3(2H)-yl)methyl)benzonitrile B5 (2.00 g, 6.80 mmol, 1.0 equivalent) was added at 25°C. The reaction mixture was stirred at 25°C for 12 hours. The reaction mixture was diluted with 150 mL of water and extracted with SiO (3 × 100 mL). The organic layers were washed together with 200 mL of brine, dehydrated with Na2SO4, filtered, and concentrated under reduced pressure to obtain the residue. The residue was diluted with MeOH (50 mL) and stirred for 20 minutes; the suspension was filtered to collect the solids. The filtered cake was washed with MeOH (20 mL) and dried under vacuum to obtain 1.8 g of (Z)-N-(3-(4-cyanobenzyl)thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T46 as a yellow solid. 200 mg of the crude product T46 was purified by preparative HPLC to obtain 132 mg of pure (Z)-N-(3-(4-cyanobenzyl)thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T46 as a white solid.

[0221] <(Z)-N-(3-(4-(aminomethyl)benzyl)thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T47> <(Z)-N-(3-(4-formylbenzyl)thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T48> A mixture of (Z)-N-(3-(4-cyanobenzyl)thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T46 (300 mg, 835 μmol, 1.0 equivalent) in THF (7 mL) was mixed with LAH (95 mg, 2.5 mmol, 3.0 equivalents) under N2 conditions at 0°C. The mixture was stirred at 25°C for 3 hours. The reaction mixture was quenched by adding water (5 mL) and acidified to pH=3 with 4N HCl. The mixture was purified by preparative HPLC to obtain 108 mg of (Z)-N-(3-(4-(aminomethyl)benzyl)thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T47 as a white solid, and 31 mg of (Z)-N-(3-(4-formylbenzyl)thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T48 as a white solid. <Preparation of Examples T54 and T64>

[0222] To a solution of an appropriate imine (1.0 equivalent) in pyridine, HBTU (2.0 equivalents) and 1H-pyrrolo[2,3-b]pyridine-3-carboxylic acid A (1.0 equivalent) were added. The reaction mixture was stirred at 100°C for 12 hours. The reaction mixture was concentrated under reduced pressure. The residue was purified by preparative HPLC to obtain T54 and T64.

[0223] <Preparation of Example T51> [ka] <N‐(チアゾール‐2‐イル)‐1H‐ピロロ[2,3‐b]ピリジン‐3‐カルボキサミド> To a solution of 1H-pyrrolo[2,3-b]pyridine-3-carboxylic acid A (0.60 g, 3.70 mmol, 1.0 equivalent) in pyridine (6 mL), HBTU (2.81 g, 7.40 mmol, 2.0 equivalent) and 2-thiazoleamine (556 mg, 5.55 mmol, 1.5 equivalent) were added at 0°C. The reaction mixture was gradually heated to 100°C and stirred at 100°C for 12 hours. The reaction mixture was poured into ice water (12 mL). The solid was collected by filtration, washed with H2O (5 × 2 mL), and dried under reduced pressure. The crude product was recrystallized by refluxing MeOH / EtOH (4 mL / 4 mL) at 70°C for 0.5 hours, then cooled to room temperature and filtered to obtain 0.41 g of N-(thiazole-2-yl)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide as a brown solid.

[0224] <Methyl 3-[(2Z)-2-(1H-pyrrolo[2,3-b]pyridine-3-carbonylimino)thiazole-3-yl]propanoate> To a solution of N-(thiazole-2-yl)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide (0.29 g, 1.19 mmol, 1.0 equivalent) in DMF (5 mL), K2CO3 (656 mg, 4.75 mmol, 4.0 equivalents) and methyl 3-bromopropanoate (595 mg, 3.56 mmol, 3.0 equivalents) were added. After addition, the reaction mixture was transferred to a sealed container for microwave reaction and stirred in a microwave reactor at 100°C for 2 hours. The reaction mixture was quenched by adding H2O (10 mL) at 0°C and extracted with ₹ (2 × 10 mL). The organic layers were washed together with H2O (20 mL) and brine (20 mL), dehydrated with Na2SO4, and filtered. The filtrate was concentrated under reduced pressure. The residue was purified by preparative silica gel TLC (etroleum ether / ethyl acetate = 1:1) to obtain 0.30 g of methyl 3-[(2Z)-2-(1H-pyrrolo[2,3-b]pyridine-3-carbonylimino)thiazole-3-yl]propanoate as a brown solid. A portion of the residue (0.10 g) was further purified twice by preparative silica gel TLC (petroleum ether / ethyl acetate = 2:1) to obtain 20 mg of methyl 3-[(2Z)-2-(1H-pyrrolo[2,3-b]pyridine-3-carbonylimino)thiazole-3-yl]propanoate as a white solid.

[0225] <Example T51> (Z)-methyl 3-(2-((1H-pyrrolo[2,3-b]pyridine-3-carbonyl)imino)thiazole-3(2H)-yl)propanoate (0.20 g, 605 μmol, 1.0 equivalent) was dissolved in dioxane (4 mL) / H2O (1 mL), to which LiOH.H2O (127 mg, 3.03 mmol, 5.0 equivalents) was added. The reaction mixture was stirred at 25°C for 12 hours. The mixture was concentrated. The residue was diluted with water (5 mL) and the pH was adjusted to < 5.0 by adding concentrated HCl. The suspension was filtered. The filtered cake was washed with H2O (5 mL) and dried under reduced pressure. The residue was purified by preparative HPLC to obtain 52 mg of (Z)-3-(2-((1H-pyrrolo[2,3-b]pyridine-3-carbonyl)imino)thiazole-3(2H)-yl)propanoic acid as a white solid.

[0226] <Preparation of Example T52> [ka] A mixture of N-(thiazole-2-yl)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide (100 mg, 409.38 μmol, 1.0 equivalent), 4-(bromomethyl)pyridine (104 mg, 409 μmol, 1.0 equivalent, HBr), and K2CO3 (170 mg, 1.23 mmol, 3.0 equivalents) in DMF (1 mL) was degassed and purged three times with N2. The reaction mixture was then stirred at 30°C for 12 hours under an N2 atmosphere. The reaction mixture was concentrated under reduced pressure. The residue was purified by preparative HPLC to obtain 37 mg of (Z)-N-(3-(pyridine-4-ylmethyl)thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T52 as a white solid.

[0227] <Preparation of Example T53> [ka] A mixture of N-(thiazole-2-yl)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide (100 mg, 409 μmol, 1.0 equivalent), 3-(bromomethyl)pyridine (104 mg, 409 μmol, 1.0 equivalent, HBr), and K2CO3 (170 mg, 1.23 mmol, 3.0 equivalents) in DMF (1 mL) was degassed and purged three times with N2. The reaction mixture was then stirred at 30°C for 12 hours under an N2 atmosphere. The reaction mixture was concentrated under reduced pressure. The residue was purified by preparative HPLC to obtain 32 mg of (Z)-N-(3-(pyridine-3-ylmethyl)thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T53 as a white solid.

[0228] <Preparation of Example T57> [ka] To a solution of N-(thiazole-2-yl)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide (200 mg, 819 μmol, 1.0 equivalent) in DMF (2 mL), K2CO3 (226 mg, 1.64 mmol, 2.0 equivalents) was added, followed by the addition of bromocyclohexane (2.67 g, 16.4 mmol, 2.02 mL, 20 equivalents) at room temperature. The reaction mixture was then stirred at 100°C for 12 hours. A further 10 equivalents of bromocyclohexane were added to the reaction mixture, and the mixture was then stirred under N2 at 100°C for another 12 hours. The reaction mixture was diluted with water (50 mL) and extracted with RINKAN (50 mL x 3). The organic layers were washed together with brine (100 mL), dehydrated with Na2SO4, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by preparative HPLC to obtain 7.3 mg of (Z)-N-(3-cyclohexylthiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T57 as a white solid. Its structure was confirmed by NOE.

[0229] <Preparation of Example T61> [ka] <(Z)-tert-butyl3-(2-((1H-pyrrolo[2,3-b]pyridine-3-carbonyl)imino)thiazole-3(2H)-yl)benzylcarbamate> To a solution of 1H-pyrrolo[2,3-b]pyridine-3-carboxylic acid A (80 mg, 491 μmol, 1.0 equivalent) in DMF (2 mL), EDCI (141 mg, 737 μmol, 1.5 equivalent), HOBt (100 mg, 737 μmol, 1.5 equivalent), DIPEA (190 mg, 1.47 mmol, 257 μL, 3.0 equivalent), and tert-butyl 3-(2-iminothiazole-3(2H)-yl)benzylcarbamate C24 (150 mg, 491 μmol, 1.0 equivalent) were added at 25°C. The reaction mixture was then stirred under N2 at 25°C for 2 hours. When the reaction mixture was diluted with water (5 mL), a solid precipitate formed. The solid was recovered by filtration, washed with 5 mL of water, and dried under reduced pressure to obtain 170 mg of (Z)-tert-butyl3-(2-((1H-pyrrolo[2,3-b]pyridine-3-carbonyl)imino)thiazole-3(2H)-yl)benzylcarbamate as a white solid.

[0230] <(Z)-N-(3-(3-(aminomethyl)phenyl)thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T61> (Z)-tert-butyl3-(2-((1H-pyrrolo[2,3-b]pyridine-3-carbonyl)imino)thiazole-3(2H)-yl)benzylcarbamate (170 mg, 378 μmol, 1.0 equivalent) was dissolved in 1 mL of HCl / HCl (378 μmol, 3 mL, 1.0 equivalent) at 25°C. The solution was then stirred under N2 at 25°C for 1 hour. The reaction mixture was filtered, and the filtered cake was washed with HCl (30 mL). The filtered cake was concentrated under reduced pressure and purified by preparative HPLC to obtain 52 mg of (Z)-N-(3-(3-(aminomethyl)phenyl)thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T61 as a white solid.

[0231] <Preparation of Example T62> [ka] <(Z)-tert-butyl7-(2-((1H-pyrrolo[2,3-b]pyridine-3-carbonyl)imino)thiazole-3(2H)-yl)-3,4-dihydroisoquinoline-2(1H)-carboxylate> To a mixture of 1H-pyrrolo[2,3-b]pyridine-3-carboxylic acid A (80 mg, 493 μmol, 1.0 equivalent) in DMF (3 mL), HOBt (100 mg, 740 μmol, 1.5 equivalent), EDCI (142 mg, 740 μmol, 1.5 equivalent), DIPEA (191 mg, 1.48 mmol, 258 μL, 3.0 equivalent), and tert-butyl 7-(2-iminothiazole-3(2H)-yl)-3,4-dihydroisoquinoline-2(1H)-carboxylate C21 (164 mg, 493 μmol, 1.0 equivalent) were added at 25°C. The reaction mixture was then stirred under N2 at 25°C for 12 hours. The reaction mixture was diluted with water (20 mL), filtered, and the solid was collected. The recovered solid was dried under reduced pressure to obtain 200 mg of (Z)-tert-butyl7-(2-((1H-pyrrolo[2,3-b]pyridine-3-carbonyl)imino)thiazole-3(2H)-yl)-3,4-dihydroisoquinoline-2(1H)-carboxylate as a brown solid. This crude product was used directly in the next step.

[0232] <(Z)-N-(3-(1,2,3,4-tetrahydroisoquinoline-7-yl)thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T62> A mixture of (Z)-tert-butyl7-(2-((1H-pyrrolo[2,3-b]pyridine-3-carbonyl)imino)thiazole-3(2H)-yl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (170 mg, 357 μmol, 1.0 equivalent) in 1 mL of HCl / Â (4 M, 3 mL, 33 equivalents) was added at 25°C. The reaction mixture was then stirred under N2 at 25°C for 2 hours. The solid was collected by filtration and purified by preparative HPLC to obtain 88 mg of (Z)-N-(3-(1,2,3,4-tetrahydroisoquinoline-7-yl)thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T62 as a white solid.

[0233] <Preparation of Example T72> [ka] <(Z)-Ethyl 2-((1H-pyrrolo[2,3-b]pyridine-3-carbonyl)imino)-3-benzyl-2,3-dihydrothiazole-4-carboxylate> To a solution of 1H-pyrrolo[2,3-b]pyridine-3-carboxylic acid (0.238 g, 1.47 mmol, 1.0 equivalent) in DMF (5 mL), EDCI (0.422 g, 2.20 mmol, 1.5 equivalent), HOBT (0.298 g, 2.20 mmol, 1.5 equivalent), diisopropylethylamine (0.758 mL, 4.41 mmol, 3.0 equivalent), and ethyl 3-benzyl-2-imino-2,3-dihydrothiazole-4-carboxylate (0.400 g, 1.47 mmol, 1.0 equivalent) were added at 25°C. The mixture was then stirred at 25°C for 1 hour. The reaction mixture was then poured into ice water (15 mL) to precipitate the solids, which were recovered by filtration. This filtered cake was triturated three times with ethanol (15 mL) at 25°C to obtain 0.40 g of (Z)-ethyl 2-((1H-pyrrolo[2,3-b]pyridine-3-carbonyl)imino)-3-benzyl-2,3-dihydrothiazole-4-carboxylate as a white solid. 1H NMR (400 MHz, DMSO-d6): δ 12.27 (br. s, 1H), 8.45 - 8.42 (m, 1H), 8.26 (dd, J = 4.8, 1.2 Hz, 1H), 8.21 - 8.20 (m, 1H), 7.99 (s, 1H), 7.35 - 7.31 (m, 2H), 7.27 - 7.2 (m, 3H), 7.14 (dd, J = 8.0, 4.8 Hz, 1H), 5.93 (s, 2H), 4.27 (q, J = 7.2 Hz, 2H), 1.25 (t, J = 7.2 Hz, 3H).LCMS (m / z [M+H] + ): 407.0.

[0234] <(Z)-N-(3-benzyl-4-(hydroxymethyl)thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T72> A suspension of (Z)-ethyl 2-((1H-pyrrolo[2,3-b]pyridine-3-carbonyl)imino)-3-benzyl-2,3-dihydrothiazole-4-carboxylate (0.200 g, 0.407 mmol) in THF (5 mL) was to be added dropwise to a THF solution of lithium borohydride (2 M, 0.4 mL, 0.8 mmol) under a nitrogen atmosphere at 0°C. After addition, the reaction mixture was warmed to 25°C and stirred for 1 hour. The mixture was then added dropwise to water (5 mL) at 25°C, followed by saturated sodium carbonate aqueous solution (5 mL). The aqueous phase was extracted with ethyl acetate (50 mL x 3). The organic layers were washed together with brine (15 mL), dehydrated with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by preparative HPLC to obtain 32 mg of (Z)-N-(3-benzyl-4-(hydroxymethyl)thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T72 as a white solid.

[0235] <Preparation of Example T75> [ka] <(Z)-tert-butyl4-(2-((1H-pyrrolo[2,3-b]pyridine-3-carbonyl)imino)thiazole-3(2H)-yl)piperidine-1-carboxylate> (Z)-N-(thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide (20 mg, 82 μmol, 1.0 equivalent) was dissolved in DMF (1 mL) and K2CO3 (57 mg, 246 μmol, 3.0 equivalents) and tert-butyl 4-bromopiperidine-1-carboxylate (216 mg, 819 μmol, 10.0 equivalents) were added. The reaction mixture was transferred to a microwave reaction tube. The sealed tube was heated in a microwave reactor at 100°C for 2 hours. When the reaction mixture was quenched by adding water (5 mL), a brown precipitate was formed. The solid was collected by filtration and washed with water (2 mL). The filtrate was dried under reduced pressure to obtain the crude product (Z)-tert-butyl4-(2-((1H-pyrrolo[2,3-b]pyridine-3-carbonyl)imino)thiazole-3(2H)-yl)piperidine-1-carboxylate (30 mg, crude) as a brown solid.

[0236] <(Z)-N-(3-(piperidine-4-yl)thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide hydrochloride T75> (Z)-tert-butyl4-(2-((1H-pyrrolo[2,3-b]pyridine-3-carbonyl)imino)thiazole-3(2H)-yl)piperidine-1-carboxylate (0.03 g, 70 umol, 1.0 equivalent) was dissolved in 1 mL of HCl / HCl (4 M, 1.2 mL, 68.4 equivalents) dropwise. The reaction mixture was stirred at 25°C for 2 hours. The reaction mixture was concentrated under reduced pressure. The residue was purified by preparative HPLC to obtain 5 mg of (Z)-N-(3-(piperidine-4-yl)thiazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide hydrochloride T75 as a white solid.

[0237] <Preparation of Example T77> [ka] To a solution of 3-benzyl-4-methyloxazole-2(3H)-imine 2,2,2-trifluoroacetate B19 (0.100 g, 0.285 mmol, 1.0 equivalent) and triethylamine (0.198 mL, 1.43 mmol, 5.0 equivalents) in chloroform (1 mL), N,N-dimethylpyridine-4-amine (3 mg, 0.03 mmol, 0.1 equivalent) and 1H-pyrrolo[2,3-b]pyridine-3-carbonyl chloride hydrochloride A (62 mg, 0.28 mmol, 1.0 equivalent) in chloroform (2 mL) were added at 0°C. The reaction mixture was warmed to 25°C and stirred for 1 hour. The mixture was concentrated under reduced pressure, the residue was dissolved in methanol (2 mL), and then poured into water (10 mL) to precipitate the solids. The precipitate was collected by filtration and purified by preparative HPLC to obtain 8 mg of (Z)-N-(3-benzyl-4-methyloxazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T77 as a white solid.

[0238] <Preparation of Example T78> [ka] <Ethyl(Z)-2-((1H-pyrrolo[2,3-b]pyridine-3-carbonyl)imino)-3-benzyl-2,3-dihydroxazole-4-carboxylate> To a solution of ethyl 3-benzyl-2-imino-2,3-dihydroxazole-4-carboxylate B20 (0.450 g, 1.83 mmol) and 1H-pyrrolo[2,3-b]pyridine-3-carboxylic acid (acid A) (0.356 g, 2.19 mmol) in DMF (6 mL), DIPEA (1.59 mL, 9.14 mmol), EDCI (0.525 g, 2.74 mmol), and HOBt (0.247 g, 1.83 mmol) were added at 25°C. The reaction mixture was stirred at 50°C for 2 hours. This reaction was quenched with water (50 mL) and then extracted with ethyl acetate (30 mL x 3). The organic layers were washed together with saturated sodium bicarbonate aqueous solution (50 mL x 3) and brine (50 mL x 3), dehydrated with anhydrous Na2SO4, and filtered. The filtrate was concentrated under reduced pressure. The crude product was purified by reverse-phase HPLC (under 0.1% NH3·H2O conditions) to obtain 0.220 g of (Z)-ethyl 2-((1H-pyrrolo[2,3-b]pyridine-3-carbonyl)imino)-3-benzyl-2,3-dihydroxazole-4-carboxylate as a yellow solid. 1 H NMR (DMSO-d6, 400 MHz) δ12.18 (br. s, 1H), 8.52 (s, 1H), 8.34 (dd, J = 1.6, 6.4 Hz, 1H), 8.24 (dd, J = 1.6, 3.2 Hz, 1H), 8.07 (d, J = 2.0 Hz, 1H), 7.37 - 7.36 (m, 4H), 7.33 - 7.29 (m, 1H), 7.10 (dd, J = 3.2, 4.8 Hz, 1H), 5.37 (s, 2H), 4.28 (q, J = 7.2 Hz, 2H), 1.25 (t, J = 7.2 Hz, 3H). + ): 391.0.

[0239] <(Z)-N-(3-benzyl-4-(hydroxymethyl)oxazole-2(3H)-ylidene)-1H-pyrrolo[2,3-b]pyridine-3-carboxamide T78> A solution of (Z)-ethyl 2-((1H-pyrrolo[2,3-b]pyridine-3-carbonyl)imino)-3-benzyl-2,3-dihydroxazole-4-carboxylate (0.100 g, 0.231 mmol) in THF (3.0 mL) was mixed with lithium borohydride solution (0.348 mL, dissolved in THF to 2 M) dropwise at 0°C under a nitrogen atmosphere. The reaction mixture was stirred at 0°C for 1 hour. The mixture was poured into 20 mL of water at 25°C, and then saturated ammonium chloride aqueous solution (20 mL) was added. The aqueous phase was extracted with ethyl acetate (30 mL x 3). The organic phases were combined and washed with brine (50 mL), dehydrated with anhydrous Na2SO4, and filtered. The filtrate was concentrated under reduced pressure. The mixture was subjected to preparative HPLC (column: Gemini®-NX C, Phenomenex). 18 The sample was purified using a 75*30mm*3um mobile phase (water (10mM NH4HCO3)-ACN), with a B% concentration of 17%-47% and a min of 8 minutes, to obtain 9 mg of (Z)-N-(3-benzyl-4-(hydroxymethyl)oxazole-2(3H)-ylidene)-1-pyrrolo[2,3-b]pyridine-3-carboxamide T78 as a white solid.

[0240] <Preparation of Examples T79 and T80> [ka] <Ethyl(2Z)-3-benzyl-2-(1H-pyrrolo[2,3-b]pyridine-3-carbonylimino)thiazole-4-carboxylate T79> To a solution of 1H-pyrrolo[2,3-b]pyridine-3-carboxylic acid (acid A) (4.62 g, 28.4 mmol) in DMF (100 mL), HOBt (3.08 g, 22.8 mmol), EDCI (8.20 g, 42.8 mmol), DIPEA (14.9 mL, 85.6 mmol), and ethyl 3-benzyl-2-iminothiazole-4-carboxylate hydrobromide B8 (10.0 g, 28.5 mmol) were added at 25°C. The resulting mixture was stirred at 25°C for 1 hour. The reaction was then poured into ice water (200 mL). The precipitate was collected by filtration, and the filtered cake was triturated with ethanol (150 mL) to obtain 10.0 g of ethyl(2Z)-3-benzyl-2-(1H-pyrrolo[2,3-b]pyridine-3-carbonylimino)thiazole-4-carboxylate T79 as a white solid.

[0241] <(Z)-3-benzyl-N-[2-(dimethylamino)ethyl]-2-(1H-pyrrolo[2,3-b]pyridine-3-carbonylimino)thiazole-4-carboxamide T80> Ethyl(2Z)-3-benzyl-2-(1H-pyrrolo[2,3-b]pyridine-3-carbonylimino)thiazole-4-carboxylate T79 (0.050 g, 0.123 mmol) and N',N'-dimethylethane-1,2-diamine (0.108 g, 1.23 mmol, 0.134 mL) were dissolved in THF (1 mL), to which 3,4,6,7,8,9-hexahydro-2H-pyrimido[1,2-a]pyrimidine (0.009 g, 0.062 mmol) was added. The mixture was stirred at 60°C for 1 hour. The reaction mixture was concentrated under reduced pressure. The residue was purified by preparative HPLC (Phenomenex Waters Xbridge™ 150*25mm*5um; mobile phase: [water (10mM NH4HCO3)-acetonitrile]; B%: 24%-54%, 10 min) to obtain 54 mg of (Z)-3-benzyl-N-[2-(dimethylamino)ethyl]-2-(1H-pyrrolo[2,3-b]pyridine-3-carbonylimino)thiazole-4-carboxamide T80 as a white solid.

[0242] <General method> LCMS conditions [Table 8] Method for IVKA assay at 10 μM:

[0243] An in vitro kinase assay (HTRF® KinEASE®-STK S1, CisBio 62ST1PEB) was optimized to the linear reaction range of the Lats1 enzyme (Carna 01-123). The reaction was carried out with 10 μM STK1 substrate and 10 μM ATP or 2 mM ATP, as shown below. The DMSO concentration was maintained at 0.5% for all experiments, and the compound dissolved in DMSO was added to the reaction using a Janus® 384MDT (PerkinElmer) equipped with a 50 nL pin tool (V and P Scientific, Inc.). The enzyme, substrate, ATP, and the test compound of the present invention were combined in a low-volume 384-well plate and stirred at room temperature for 50 minutes unless otherwise specified. The reaction was stopped by adding a detection reagent prepared at an 8:1 biotin:streptavidin ratio and stirring at room temperature for 60 minutes. All reactions were performed in triplicate, and the signal was detected using Synergy®NEO (Biotek).

[0244] Reagents for IVKA in 2 mM ATP, final concentration at assay: Kit: HTRFKinEASE-STKS1, Cisbio's 62ST1PEB); 1× kinase buffer (KB); 5 mM gCl2; 1 mM DTT; ATP 2 mM; STK1, 2.5 μM; 50 pg / μL (Lats1, Karna's 01-123); BSA 1 mg / mL; Biotin:Streptavidin ratio 8:1; Detection reagent Streptavidin-XL665: 156 nM for 10 μL of final reaction volume and 20 μL of final detection volume. The assay was performed in a 384-well plate. Drug dose curves were prepared as 16-point, triplicate curves, each diluted 3-fold with 100% DMSO. The Lats1 activity measured as described above is shown in the following table.

[0245] [Table 9] Compound T01 (TIFF2023512823000105.tif203145) was tested in detail to investigate its mechanism of action and usefulness in regenerating ear hair cells. The effect of the compound was tested on utricles isolated from 8-12 week old mice. The inner ear was excised from mice euthanized with fluoride and placed in ice-cold Hanks equilibrium salt solution, and cultured as previously described by Gnedeva, K. and Hudspeth [Proc. Natl. Acad. Sci., (US), 2015, Vol. 112, p. 14066-14071]. For the proliferation assay, the utricles were cultured with 10 μM of 5-ethynyl-2'-deoxyuridine (EdU), which is detected by click chemistry.

[0246] Immunohistochemical analysis revealed that T01 promotes robust Yap nuclear translocation in supporting cells after 24 hours of treatment at a concentration of 10 μM (quantified as a ratio to the constitutively expressed protein Sall2; control group = 0.6; T01 treatment group = 1.0; p<0.0001 by unpaired two-sided t-test, n=570 (control group nuclei) and 680 (treatment group nuclei)), and that T01 causes a significant reduction in Yap phosphorylation levels, as detected by Western blotting. After 5 days of treatment, T01 induced robust re-entry into the cell cycle of adult utricle supporting cells, resulting in hundreds of EdU+ daughter supporting cells (control group = 20 EdU+ supporting cells; T01 treatment group = 250 EdU+ supporting cells; p=0.021 by unpaired one-sided t-test, n=2 for the control group and n=3 for the T01 group).

Claims

1. Formula I: 【Chemistry 1】 (In the formula, R １ (C １ -C ６ ) alkyl, carboxy, (C ３ -C ７ ) carbomonocyclyl, (C ９ -C １１ ) selected from the group consisting of carbobicyclyl, heteromonocyclyl, and heterobicyclyl, １ -C ６ ) alkyl, (C ３ -C ７ ) carbomonocyclyl, (C ９ -C １１ ) Carbobicyclyl, heteromonocyclyl, and heterobicyclyl are optionally selected from halogen, cyano, hydroxy, nitro, amino, acetoxy, carboxy, (C １ -C ７ ) hydrocarbyl, halo(C １ -C ６ ) alkyl, (C １ -C ３ ) alkoxy, halo (C １ -C ３ ) alkoxy, (C １ -C ６ ) acyl, (C １ -C ３ ) Alkoxy (C １ -C ３ ) alkyl, hydroxy (C １ -C ３ ) alkyl, heteroaryl, benzenesulfonyl, (C １ -C ３ ) alkoxycarbonyl, aminocarbonyl, (C １ -C ３ ) alkylaminocarbonyl, di(C １ -C ３ ) alkylaminocarbonyl, (C １ -C ３ ) alkylamino, di(C １ -C ３ ) alkylamino, amino (C １ -C ３ ) alkyl, (C １ -C ３ ) alkylamino (C １ -C ３ ) alkyl, (C １ -C ３ ) Dialkylamino (C １ -C ３ ) alkyl, (C １ -C ３ ) alkylthio, (C １ -C ３ ) alkylsulfonylamino, (C １ -C ３ ) alkylsulfinyl, (C １ -C ３ ) optionally substituted with 1 to 3 substituents independently selected from the group consisting of alkylsulfonyl, phenoxy, and benzyloxy; R ２ is hydrogen, halogen, (C １ -C ７ ) hydrocarbyl, halo(C １ -C ６ ) alkyl, (C １ -C ６ ) acyl, hydroxy (C １ -C ３ ) alkyl, -C(=O)O(C １ -C ６ ) alkyl, -C(=O)NR ２０ R ２１ , and (C １ -C ６ ) oxaalkyl; R ３ is hydrogen, halogen, (C １ -C ６ ) hydrocarbyl, halo(C １ -C ６ ) alkyl, (C １ -C ６ ) acyl, and (C １ -C ３ ) alkoxy; R ４ is hydrogen, halogen, (C １ -C ６ ) hydrocarbyl, halo(C １ -C ６ ) alkyl, (C １ -C ６ ) acyl, and (C １ -C ３ ) alkoxy; R １０ is independently selected in each occurrence from the group consisting of hydrogen and methyl; R ２０ is hydrogen and (C １ -C ６ ) hydrocarbyl; R ２１ is hydrogen, (C １ -C ６ ) hydrocarbyl, (C １ -C ６ ) oxaalkyl, amino (C １ -C ６ ) alkyl, (C １ -C ３ ) alkylamino (C １ -C ６ ) alkyl, di(C １ -C ３ ) alkylamino (C １ -C ６ ) alkyl, and -(CH ２ ) ｍ -Het, where Het is selected from the group consisting of hydroxy, amino, acetoxy, carboxy, (C １ -C ７ ) hydrocarbyl, halo(C １ -C ６ ) alkyl, (C １‐ C ３ ) alkoxy, halo (C １ -C ３ ) alkoxy, (C １ -C ６ ) acyl, (C １ -C ３ ) Alkoxy (C １ -C ３ ) alkyl, hydroxy (C １ -C ３ ) alkyl, aminocarbonyl, (C １ -C ３ ) alkylaminocarbonyl, di(C １ -C ３ ) alkylaminocarbonyl, (C １ -C ３ ) alkylamino, and di(C １ -C ３ ) an aliphatic monocyclic or bicyclic heterocycle optionally substituted with a substituent selected from the group consisting of alkylamino; Or, R ２０ and R ２１ is R ２０ and R ２１ forms an aliphatic heterocycle together with the nitrogen to which it is attached, n is 0, 1 or 2; m is 0, 1 or 2, and X is S or n is 1 and R １ When is optionally substituted phenyl, X is S or O; However, R １ When is phenyl, X is sulfur, and n is 1, R ２ , R ３ , R ４ and R １０ At least one of is other than hydrogen. A compound represented by the formula:

2. R ２ is -C(=O)O(C １ -C ６ ) alkyl, -C(=O)NR ２０ R ２１ , and (C １ -C ６ 2. The compound of claim 1, wherein said compound is selected from the group consisting of:

3. R ２０ is selected from hydrogen and methyl, and R ２１ is hydrogen, methyl, (C １ -C ６ ) oxaalkyl, dimethylamino (C １ -C ６ ) alkyl, and -(CH ２ ) ｍ The compound according to claim 2 , wherein the group is selected from -Het.

4. R ２０ and R ２１ is R ２０ and R ２１ The compound according to claim 1, wherein, together with the nitrogen to which it is attached, forms a 4- to 7-membered aliphatic heterocycle.

5. Below formula: 【Chemistry 2】 (In the formula, R １ (C １ -C ６ ) alkyl, carboxy, (C ３ -C ７ ) carbomonocyclyl, (C ９ -C １１ ) selected from the group consisting of carbobicyclyl, heteromonocyclyl, and heterobicyclyl, １ -C ６ ) alkyl, (C ３ -C ７ ) carbomonocyclyl, (C ９ -C １１ ) Carbobicyclyl, heteromonocyclyl, and heterobicyclyl are optionally selected from halogen, cyano, hydroxy, nitro, amino, acetoxy, carboxy, (C １ -C ７ ) hydrocarbyl, halo(C １ -C ６ ) alkyl, (C １ -C ３ ) alkoxy, halo (C １ -C ３ ) alkoxy, (C １ -C ６ ) acyl, (C １ -C ３ ) Alkoxy (C １ -C ３ ) alkyl, hydroxy (C １ -C ３ ) alkyl, heteroaryl, benzenesulfonyl, (C １ -C ３ ) alkoxycarbonyl, aminocarbonyl, (C １ -C ３ ) alkylaminocarbonyl, (C １ -C ３ ) alkylamino, di(C １ -C ３ ) alkylamino, amino (C １ -C ３ ) alkyl, (C １ -C ３ ) alkylamino (C １ -C ３ ) alkyl, (C １ -C ３ ) Dialkylamino (C １ -C ３ ) alkyl, (C １ -C ３ ) alkylthio, (C １ -C ３ ) alkylsulfonylamino, (C １ -C ３ ) alkylsulfinyl, (C １ -C ３ ) optionally substituted with 1 to 3 substituents independently selected from the group consisting of alkylsulfonyl, phenoxy, and benzyloxy; R ２ is hydrogen, halogen, (C １ -C ６ ) hydrocarbyl, halo(C １ -C ６ ) alkyl, (C １ -C ６ ) acyl, hydroxy (C １ -C ３ ) alkyl, and (C １ -C ３ ) alkoxy; R ３ is hydrogen, halogen, (C １ -C ６ ) hydrocarbyl, halo(C １ -C ６ ) alkyl, (C １ -C ６ ) acyl, and (C １ -C ３ ) alkoxy; R ４ is hydrogen, halogen, (C １ -C ６ ) hydrocarbyl, halo(C １ -C ６ ) alkyl, (C １ -C ６ ) acyl, and (C １ -C ３ ) alkoxy; R １０ is independently selected in each occurrence from the group consisting of hydrogen and methyl; n is 0, 1 or 2, and X is O or S; However, R １ When is phenyl, X is sulfur, and n is 1, R ２ , R ３ , R ４ and R １０ At least one of is other than hydrogen. The compound according to claim 1 ,

6. The compound according to any one of claims 1 to 5, wherein n is 0.

7. The compound according to any one of claims 1 to 5, wherein n is 1.

8. R １０ The compound of claim 7 , wherein is hydrogen.

9. R １ is carboxy as well as optionally substituted (C １ -C ４ 6. The compound according to any one of claims 1 to 5, wherein the aryl group is selected from the group consisting of alkyl, phenyl, cyclohexyl, 5-membered heterocyclyl, 6-membered heterocyclyl, and heterobicyclyl.

10. R １ 10. The compound of claim 9, wherein is selected from the group consisting of methyl, ethyl, aminobutyl, and carboxyethyl.

11. R １ The compound of claim 9 , wherein is optionally substituted cyclohexyl.

12. R １ The compound of claim 9 , wherein is optionally substituted phenyl.

13. R １ The compound of claim 9 , wherein is optionally substituted heterocyclyl.

14. R １ 14. The compound of claim 13, wherein is selected from the group consisting of pyridinyl, pyrazolyl, piperidinyl, tetrahydropyranyl, tetrahydrofuranyl, and tetrahydroisoquinolinyl, each of which is optionally substituted.

15. R １ is phenyl or halogen, cyano, hydroxy, amino, carboxy, (C １ -C ６ ) hydrocarbyl, trifluoromethyl, methoxy, acetyl, formyl, hydroxy (C １ -C ３ ) alkyl, methoxycarbonyl, carboxamido, methanesulfonylamino, and amino (C １ -C ３ 13. The compound of claim 12, wherein said phenyl is substituted with one or two substituents independently selected from the group consisting of: alkyl, aryl, phenyl, phenyl substituted with one or two substituents independently selected from the group consisting of: alkyl, aryl, aryl substituted with one or two substituents independently selected from the group consisting of: alkyl ...

16. R １ 16. The compound of claim 15, wherein is phenyl substituted at the ortho position and n is 0.

17. R １ are each an amino group, a hydroxy group, and a (C １ -C ６ 15. The compound of claim 14, wherein the aryl group is selected from the group consisting of pyridinyl, pyrazolyl, piperidinyl, tetrahydropyranyl, and tetrahydroisoquinolinyl, optionally substituted with 1 or 2 substituents independently selected from the group consisting of hydrocarbyl.

18. 10. The compound of claim 9, wherein X is S.

19. 10. The compound of claim 9, wherein X is O.

20. R ３ and R ４ 10. The compound of claim 9, wherein is independently selected from the group consisting of hydrogen, chloro and methyl.

21. R ２ 10. The compound of claim 9, wherein is selected from the group consisting of hydrogen, methyl, ethyl, propyl, cyclopropyl, hydroxymethyl, and trifluoromethyl.

22. A compound according to claim 1 selected from the compounds in the table below. 【Chemistry 3】 【change】 【change】 【change】 【change】 23. The compound of claim 1, which is a compound of the following formula: 【Chemistry 4】

24. A pharmaceutical composition comprising a pharma- ceutically acceptable carrier and a compound according to any one of claims 1 to 23.

25. Below formula: 【Chemistry 5】 (In the formula, R １ (C １ -C ６ ) alkyl, carboxy, (C ３ -C ７ ) carbomonocyclyl, (C ９ -C １１ ) selected from the group consisting of carbobicyclyl, heteromonocyclyl, and heterobicyclyl, １ -C ６ ) alkyl, (C ３ -C ７ ) carbomonocyclyl, (C ９ -C １１ ) Carbobicyclyl, heteromonocyclyl, and heterobicyclyl are optionally selected from halogen, cyano, hydroxy, nitro, amino, acetoxy, carboxy, (C １ -C ７ ) hydrocarbyl, halo(C １ -C ６ ) alkyl, (C １ -C ３ ) alkoxy, halo (C １ -C ３ ) alkoxy, (C １ -C ６ ) acyl, (C １ -C ３ ) Alkoxy (C １ -C ３ ) alkyl, hydroxy (C １ -C ３ ) alkyl, heteroaryl, benzenesulfonyl, (C １ -C ３ ) alkoxycarbonyl, aminocarbonyl, (C １ -C ３ ) alkylaminocarbonyl, di(C １ -C ３ ) alkylaminocarbonyl, (C １ -C ３ ) alkylamino, di(C １ -C ３ ) alkylamino, amino (C １ -C ３ ) alkyl, (C １ -C ３ ) alkylamino (C １ -C ３ ) alkyl, (C １ -C ３ ) Dialkylamino (C １ -C ３ ) alkyl, (C １ -C ３ ) alkylthio, (C １ -C ３ ) alkylsulfonylamino, (C １ -C ３ ) alkylsulfinyl, (C １ -C ３ ) optionally substituted with 1 to 3 substituents independently selected from the group consisting of alkylsulfonyl, phenoxy, and benzyloxy; R ２ is hydrogen, halogen, (C １ -C ６ ) hydrocarbyl, halo(C １ -C ６ ) alkyl, (C １ -C ６ ) acyl, hydroxy (C １ -C ３ ) alkyl, -C(=O)O(C １ -C ６ ) alkyl, -C(=O)NR ２０ R ２１ , and (C １ -C ６ ) oxaalkyl; R ３ is hydrogen, halogen, (C １ -C ６ ) hydrocarbyl, halo(C １ -C ６ ) alkyl, (C １ -C ６ ) acyl, and (C １ -C ３ ) alkoxy; R ４ is hydrogen, halogen, (C １ -C ６ ) hydrocarbyl, halo(C １ -C ６ ) alkyl, (C １ -C ６ ) acyl, and (C １ -C ３ ) alkoxy; R １０ is independently selected in each occurrence from the group consisting of hydrogen and methyl; R ２０ is hydrogen and (C １ -C ６ ) hydrocarbyl; R ２１ is hydrogen, (C １ -C ６ ) hydrocarbyl, (C １ -C ６ ) oxaalkyl, amino (C １ -C ６ ) alkyl, (C １ -C ３ ) alkylamino (C １ -C ６ ) alkyl, di(C １ -C ３ ) alkylamino (C １ -C ６ ) alkyl, and -(CH ２ ) ｍ -Het, where Het is selected from the group consisting of hydroxy, amino, acetoxy, carboxy, (C １ -C ７ ) hydrocarbyl, halo(C １ -C ６ ) alkyl, (C １ -C ３ ) alkoxy, halo (C １ -C ３ ) alkoxy, (C １ -C ６ ) acyl, (C １ -C ３ ) Alkoxy (C １ -C ３ ) alkyl, hydroxy (C １ -C ３ ) alkyl, aminocarbonyl, (C １ -C ３ ) alkylaminocarbonyl, di(C １ -C ３ ) alkylaminocarbonyl, (C １ -C ３ ) alkylamino, and di(C １ -C ３ ) an aliphatic monocyclic or bicyclic heterocycle optionally substituted with a substituent selected from the group consisting of alkylamino; Or, R ２０ and R ２１ is R ２０ and R ２１ forms an aliphatic heterocycle together with the nitrogen to which it is attached, n is 0, 1 or 2; m is 0, 1 or 2, and X is S or n is 1 and R １ When is optionally substituted phenyl, X is S or O. A composition for activating YAP in a cell expressing YAP, comprising a compound represented by the formula: The composition, wherein the cell is exposed to the compound.

26. Below formula: 【Chemistry 6】 (In the formula, R １ (C １ -C ６ ) alkyl, carboxy, (C ３ -C ７ ) carbomonocyclyl, (C ９ -C １１ ) selected from the group consisting of carbobicyclyl, heteromonocyclyl, and heterobicyclyl, １ -C ６ ) alkyl, (C ３ -C ７ ) carbomonocyclyl, (C ９ -C １１ ) Carbobicyclyl, heteromonocyclyl, and heterobicyclyl are optionally selected from halogen, cyano, hydroxy, nitro, amino, acetoxy, carboxy, (C １ -C ７ ) hydrocarbyl, halo(C １ -C ６ ) alkyl, (C １ -C ３ ) alkoxy, halo (C １ -C ３ ) alkoxy, (C １ -C ６ ) acyl, (C １ -C ３ ) Alkoxy (C １ -C ３ ) alkyl, hydroxy (C １ -C ３ ) alkyl, heteroaryl, benzenesulfonyl, (C １ -C ３ ) alkoxycarbonyl, aminocarbonyl, (C １ -C ３ ) alkylaminocarbonyl, di(C １ -C ３ ) alkylaminocarbonyl, (C １ -C ３ ) alkylamino, di(C １ -C ３ ) alkylamino, amino (C １ -C ３ ) alkyl, (C １ -C ３ ) alkylamino (C １ -C ３ ) alkyl, (C １ -C ３ ) Dialkylamino (C １ -C ３ ) alkyl, (C １ -C ３ ) alkylthio, (C １ -C ３ ) alkylsulfonylamino, (C １ -C ３ ) alkylsulfinyl, (C １ -C ３ ) optionally substituted with 1 to 3 substituents independently selected from the group consisting of alkylsulfonyl, phenoxy, and benzyloxy; R ２ is hydrogen, halogen, (C １ -C ６ ) hydrocarbyl, halo(C １ -C ６ ) alkyl, (C １ -C ６ ) acyl, hydroxy (C １ -C ３ ) alkyl, -C(=O)O(C １ -C ６ ) alkyl, -C(=O)NR ２０ R ２１ , and (C １ -C ６ ) oxaalkyl; R ３ is hydrogen, halogen, (C １ -C ６ ) hydrocarbyl, halo(C １ -C ６ ) alkyl, (C １ -C ６ ) acyl, and (C １ -C ３ ) alkoxy; R ４ is hydrogen, halogen, (C １ -C ６ ) hydrocarbyl, halo(C １ -C ６ ) alkyl, (C １ -C ６ ) acyl, and (C １ -C ３ ) alkoxy; R １０ is independently selected in each occurrence from the group consisting of hydrogen and methyl; R ２０ is hydrogen and (C １ -C ６ ) hydrocarbyl; R ２１ is hydrogen, (C １ -C ６ ) hydrocarbyl, (C １ -C ６ ) oxaalkyl, amino (C １ -C ６ ) alkyl, (C １ -C ３ ) alkylamino (C １ -C ６ ) alkyl, di(C １ -C ３ ) alkylamino (C １ -C ６ ) alkyl, and -(CH ２ ) ｍ -Het, where Het is selected from the group consisting of hydroxy, amino, acetoxy, carboxy, (C １ -C ７ ) hydrocarbyl, halo(C １ -C ６ ) alkyl, (C １ -C ３ ) alkoxy, halo (C １ -C ３ ) alkoxy, (C １ -C ６ ) acyl, (C １ -C ３ ) Alkoxy (C １ -C ３ ) alkyl, hydroxy (C １ -C ３ ) alkyl, aminocarbonyl, (C １ -C ３ ) alkylaminocarbonyl, di(C １ -C ３ ) alkylaminocarbonyl, (C １ -C ３ ) alkylamino, and di(C １ -C ３ ) an aliphatic monocyclic or bicyclic heterocycle optionally substituted with a substituent selected from the group consisting of alkylamino; Or, R ２０ and R ２１ is R ２０ and R ２１ forms an aliphatic heterocycle together with the nitrogen to which it is attached, n is 0, 1 or 2; m is 0, 1 or 2, and X is S or n is 1 and R １ When is optionally substituted phenyl, X is S or O. A composition for inhibiting LATS in a cell expressing LATS, comprising a compound represented by the formula: The composition, wherein the cell is exposed to the compound.

27. Below formula: 【Chemistry 7】 (In the formula, R １ (C １ -C ６ ) alkyl, carboxy, (C ３ -C ７ ) carbomonocyclyl, (C ９ -C １１ ) selected from the group consisting of carbobicyclyl, heteromonocyclyl, and heterobicyclyl, １ -C ６ ) alkyl, (C ３ -C ７ ) carbomonocyclyl, (C ９ -C １１ ) Carbobicyclyl, heteromonocyclyl, and heterobicyclyl are optionally selected from halogen, cyano, hydroxy, nitro, amino, acetoxy, carboxy, (C １ -C ７ ) hydrocarbyl, halo(C １ -C ６ ) alkyl, (C １ -C ３ ) alkoxy, halo (C １ -C ３ ) alkoxy, (C １ -C ６ ) acyl, (C １ -C ３ ) Alkoxy (C １ -C ３ ) alkyl, hydroxy (C １ -C ３ ) alkyl, heteroaryl, benzenesulfonyl, (C １ -C ３ ) alkoxycarbonyl, aminocarbonyl, (C １ -C ３ ) alkylaminocarbonyl, di(C １ -C ３ ) alkylaminocarbonyl, (C １ -C ３ ) alkylamino, di(C １ -C ３ ) alkylamino, amino (C １ -C ３ ) alkyl, (C １ -C ３ ) alkylamino (C １ -C ３ ) alkyl, (C １ -C ３ ) Dialkylamino (C １ -C ３ ) alkyl, (C １ -C ３ ) alkylthio, (C １ -C ３ ) alkylsulfonylamino, (C １ -C ３ ) alkylsulfinyl, (C １ -C ３ ) optionally substituted with 1 to 3 substituents independently selected from the group consisting of alkylsulfonyl, phenoxy, and benzyloxy; R ２ is hydrogen, halogen, (C １ -C ６ ) hydrocarbyl, halo(C １ -C ６ ) alkyl, (C １ -C ６ ) acyl, hydroxy (C １ -C ３ ) alkyl, -C(=O)O(C １ -C ６ ) alkyl, -C(=O)NR ２０ R ２１ , and (C １ -C ６ ) oxaalkyl; R ３ is hydrogen, halogen, (C １ -C ６ ) hydrocarbyl, halo(C １ -C ６ ) alkyl, (C １ -C ６ ) acyl, and (C １ -C ３ ) alkoxy; R ４ is hydrogen, halogen, (C １ -C ６ ) hydrocarbyl, halo(C １ -C ６ ) alkyl, (C １ -C ６ ) acyl, and (C １ -C ３ ) alkoxy; R １０ is independently selected in each occurrence from the group consisting of hydrogen and methyl; R ２０ is hydrogen and (C １ -C ６ ) hydrocarbyl; R ２１ is hydrogen, (C １ -C ６ ) hydrocarbyl, (C １ -C ６ ) oxaalkyl, amino (C １ -C ６ ) alkyl, (C １ -C ３ ) alkylamino (C １ -C ６ ) alkyl, di(C １ -C ３ ) alkylamino (C １ -C ６ ) alkyl, and -(CH ２ ) ｍ -Het, where Het is selected from the group consisting of hydroxy, amino, acetoxy, carboxy, (C １ -C ７ ) hydrocarbyl, halo(C １ -C ６ ) alkyl, (C １ -C ３ ) alkoxy, halo (C １ -C ３ ) alkoxy, (C １ -C ６ ) acyl, (C １ -C ３ ) Alkoxy (C １ -C ３ ) alkyl, hydroxy (C １ -C ３ ) alkyl, aminocarbonyl, (C １ -C ３ ) alkylaminocarbonyl, di(C １ -C ３ ) alkylaminocarbonyl, (C １ -C ３ ) alkylamino, and di(C １ -C ３ ) an aliphatic monocyclic or bicyclic heterocycle optionally substituted with a substituent selected from the group consisting of alkylamino; Or, R ２０ and R ２１ is R ２０ and R ２１ forms an aliphatic heterocycle together with the nitrogen to which it is attached, n is 0, 1 or 2; m is 0, 1 or 2, and X is S or n is 1 and R １ When is optionally substituted phenyl, X is S or O. A composition for stimulating hair cell regeneration comprising a compound represented by the formula: The composition, wherein a feeder cell population is exposed to the compound.

28. The composition of any one of claims 25 to 27, wherein the cell, cell population, or support cell population is in vivo in a subject.

29. 30. The composition of claim 28, wherein the subject has or is at risk of developing hearing loss.

30. 30. The composition of claim 28, wherein the subject has or is at risk of developing vestibular dysfunction.

31. A composition for treating a subject having or at risk of developing hearing loss, comprising a compound according to any one of claims 1 to 23, wherein an effective amount of the compound is administered to the subject.

32. 32. The composition of claim 29 or 31, wherein the hearing loss is hereditary hearing loss.

33. 33. The composition of claim 32, wherein the hereditary hearing loss is autosomal dominant hearing loss, autosomal recessive hearing loss, or X-linked hearing loss.

34. 32. The composition of claim 29 or 31, wherein the hearing loss is acquired hearing loss.

35. 35. The composition of claim 34, wherein the acquired hearing loss is noise-induced hearing loss, age-related hearing loss, disease or infection-related hearing loss, head trauma-related hearing loss, or ototoxic drug-induced hearing loss.

36. A composition for treating a human subject having or at risk of developing tinnitus, comprising a compound according to any one of claims 1 to 23, wherein an effective amount of the compound is administered to the subject.

37. A composition for treating a subject having or at risk of developing a vestibular dysfunction, comprising a compound described in any one of claims 1 to 23, wherein an effective amount of the compound is administered to the subject.

38. 40. The composition of claim 30 or 37, wherein the vestibular dysfunction comprises vertigo, dizziness, disequilibrium, bilateral vestibular hypofunction, oscillopsia, or balance disorders.

39. 40. The composition of any one of claims 30, 37 or 38, wherein the vestibular dysfunction is age-related vestibular dysfunction, head trauma-related vestibular dysfunction, disease or infection-related vestibular dysfunction, or ototoxic drug-induced vestibular dysfunction.

40. 40. The composition of claim 35 or 39, wherein the ototoxic drug is an aminoglycoside, an antineoplastic agent, ethacrynic acid, furosemide, a salicylate, or quinine.

41. 40. The composition of any one of claims 30, 37 and 38, wherein the vestibular dysfunction is associated with a genetic mutation.

42. 39. The composition of any one of claims 30, 37 and 38, wherein the vestibular dysfunction is idiopathic.

43. The composition of any one of claims 28 to 42, which is administered topically.

44. 44. The composition of claim 43, administered to the inner ear or middle ear.

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