Synthesis of pyruvate kinase activators

Abstract

Provided herein are methods for preparing activators of Pyruvate Kinase (PK) or a salt or a hydrate thereof.

Description

SYNTHESIS OF PYRUVATE KINASE ACTIVATORS RELATED APPLICATIONS

[0001] This application claims priority to U.S. Provisional Application No.63 / 525,079, filed July 05, 2023, the entirety of which is incorporated herein by reference. BACKGROUND

[0002] Pyruvate kinase (PK) is a metabolic enzyme that converts phosphoenolpyruvate to pyruvate during glycolysis. Four PK isoforms exist in mammals: the L and R isoforms are expressed in liver and red blood cells, the M1 isoform is expressed in most adult tissues, and the M2 isoform is a splice variant of M1 expressed during embryonic development. A well- known difference between the M1 and M2 isoforms of PK is that M2 is a low-activity enzyme that relies on allosteric activation by the upstream glycowlytic intermediate, fructose- 1,6-bisphosphate (FBP), whereas M1 is a constitutively active enzyme. PK activators can be used to treat a number of different disorders including PKD (Pyruvate Kinase Deficiency), thalassemia (e.g., alpha and beta-thalassemia), hereditary elliptocytosis, abetalipoproteinemia or Bassen-Kornzweig syndrome, sickle cell disease, paroxysmal nocturnal hemoglobinuria, and various anemias which include congenital anemias (e.g., enzymopathies) and hemolytic anemias (e.g. hereditary and / or congenital hemolytic anemia, acquired hemolytic anemia, chronic hemolytic anemia caused by phosphoglycerate kinase deficiency, anemia due to MDS (myelodysplastic syndromes), non-spherocytic hemolytic anemia and hereditary spherocytosis). SUMMARY

[0003] Provided herein are synthetic methods for preparing pyruvate kinase (PK) activators, or a salt or a hydrate thereof. Such activators may be prepared using compounds having the Formula (A-1) and / or Formula (B-1):wherein X, R1, R4, and Q are as defined herein.

[0004] Specific PK activators and various intermediates prepared by and / or used in the synthetic methods described herein are also disclosed. BRIEF DESCRIPTION OF THE FIGURES

[0005] FIG.1 depicts a Nuclear Magnetic Resonance (NMR) spectrum for tert-butyl 2- (6-((6-aminopyridin-2-yl)methyl)-4-methyl-5-oxo-5,6-dihydro-4H- thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-2-yl)-2-(1-(tetrahydro-2H-pyran-2-yl)-1H-pyrazol- 3-yl)acetate.

[0006] FIG.2 depicts a Nuclear Magnetic Resonance (NMR) spectrum for 2-chloro-6- formyl-4-methyl-4H-pyrrolo[2,3-d]thiazole-5-carboxylic acid. DETAILED DESCRIPTION

[0007] In one embodiment, provided is a method of preparing a compound of Formula (A-1):or a salt thereof, wherein R1is hydrogen, C1-C6alkyl, C1-C6haloalkyl, C1-C6hydroxyalkyl, C1-C6aminoalkyl, C1-C6cyanoalkyl, C2-C6alkenyl, C2-C6alkynyl, or C3-C6cycloalkyl, wherein said C3-C6cycloalkyl is optionally substituted with 1 to 3 groups selected from halo, hydroxyl, NH2, and CN; X is halo; L is a bond or C1-C6alkylene; and Q is C3-C6 cycloalkyl, 5-14 membered heterocyclyl, 6-12 membered aryl, or 5-14 membered heteroaryl, each of which is optionally substituted with 1-3 groups selected from halo, C1-C6alkyl, C1-C6haloalkyl, C1-C6hydroxyalkyl, hydroxyl, C1-C6aminoalkyl, -NH2, - NH(C1-C6alkyl), -N(C1-C6alkyl)2, CN, and NO2; the method comprising reacting a compound of Formula (A-2):with a compound of Formula (A-3):to form the compound of Formula (A-1), or a salt thereof.

[0008] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure pertains. The terminology used in the description is for describing particular embodiments only and is not intended to be limiting of the disclosure.

[0009] The use of any and all examples or exemplary language (e.g., “such as” and “e.g.”) as provided herein, is intended to better illustrate the disclosure and is not a limitation on the scope of the disclosure unless expressly claimed otherwise. Phrases such as “in one aspect” or “in one embodiment” or “in some embodiments” or “in certain embodiments” or “in further embodiments” and the like should not be construed as indicating that such elements occur or exist in isolation or that such elements are not shared by other aspects or embodiments of the disclosure. Rather, it should be understood that all aspects and embodiments may be freely combined with any and all other aspects and embodiments of the disclosure as described herein. No language in the specification should be construed as indicating that any non-claimed element is essential to the practice of the disclosure.

[0010] The terms “C1-C6alkyl” refers to a straight- or branched-chain hydrocarbon group having from 1 to 6 carbon atoms. Examples of C1-C6alkyl groups include methyl (C1), ethyl (C2), propyl (C3) (e.g., n-propyl, isopropyl), butyl (C4) (e.g., n-butyl, tert-butyl, sec-butyl, iso- butyl), pentyl (C5) (e.g., n-pentyl, 3-pentyl, amyl, neopentyl, 3-methyl-2-butanyl, tertiary amyl), and hexyl (C6) (e.g., n-hexyl).

[0011] The term “C1-C6alkylene” refers to a bivalent radical of a saturated, straight or branched chain hydrocarbon. Exemplary alkylene groups include, but are not limited to, C1- C4alkylene, C1-C3alkylene, C1-C2alkylene, and the corresponding alkylene to any of the other exemplary alkyl groups described above.

[0012] When a range of carbon atoms is used herein, for example, C1-C6, all ranges, as well as individual numbers of carbon atoms are encompassed. For example, “C1-C3” includes C1-C3, C1-C2, C2-C3, C1, C2, and C3.

[0013] As used herein, the term “C2-C6alkenyl” refers to a straight- or branched-chain group having from 2 to 6 carbon atoms in the group, wherein the group includes at least one carbon-carbon double bond. Examples of C2-C6alkenyl groups include vinyl (-CH=CH2; C2alkenyl), allyl (-CH2-CH=CH2; C3alkenyl), propenyl (-CH=CHCH3; C3alkenyl), isopropenyl (-C(CH3)=CH2; C3alkenyl), butenyl (-CH=CHCH2CH3; C4alkenyl), sec-butenyl (-C(CH3)=CHCH3; C4alkenyl), iso-butenyl (-CH=C(CH3)2; C4alkenyl), and 2-butenyl (- CH2CH=CHCH3; C4alkyl).

[0014] As used herein, the term “C2-C6alkynyl” refers to a straight- or branched-chain group having from 2 to 6 carbon atoms in the group, and wherein the group includes at least one carbon-carbon triple bond. Examples of alkynyl groups include ethynyl (-C≡CH; C2alkynyl), propargyl (-CH2-C≡CH; C3alkynyl), propynyl (-C≡CCH3; C3alkynyl), butynyl (- C≡CCH2CH3; C4alkynyl), and pentynyl (-C≡CCH2CH2CH3; C5alkynyl).

[0015] The term “C3-C6cycloalkyl” refers to cyclic-containing, non-aromatic hydrocarbon groups having from 3 to 6 carbon atoms. Examples of C3-C6cycloalkyl groups include cyclopropyl (C3), cyclobutyl (C4), cyclopentyl (C5), and cyclohexyl (C6).

[0016] The term “halo” or “halogen,” as used by itself or as part of another group refers to a fluorine, chlorine, bromine, or iodine atom.

[0017] As used herein, the term “C1-C6haloalkyl” refers to an alkyl group wherein one or more of the hydrogen atoms has been replaced with one or more halogen atoms which may be the same or different. In some embodiments, the alkyl is substituted by at least one halogen. In other embodiments, the alkyl is substituted by one, two, or three F and / or Cl. Examples of haloalkyl groups include fluoromethyl (CH2F), 1-fluoroethyl (CH(CH3)F), 2- fluoroethyl, difluoromethyl (CHF2), trifluoromethyl (CF3), pentafluoroethyl, 1,1- difluoroethyl (C(CH3)F2), 2,2-difluoroethyl (CH2CHF2), 2,2,2-trifluoroethyl (CH2CF3), 2-fluoropropan-2-yl (C(CH3)2F), 3,3,3-trifluoropropyl, 4,4,4-trifluorobutyl, trichloromethyl andthe like.

[0018] The term “C1-C6cyanoalkyl” as used by itself or as part of another group refers to an alkyl as defined herein that is substituted by one or more CN. In some embodiments, the alkyl is substituted by at least one CN. In other embodiments, the alkyl is substituted by one, two, or three CN. Examples of cyanoalkyl groups include CH2CN, CH2CH2CN, CH(CN)CH3, CH2CH2CH2CN, C(CH3)2CN, CH2CH(CN)CH3, CH(CN)CH2CH3, and the like.

[0019] The term “C1-C6hydroxyalkyl” as used by itself or as part of another group refers to an alkyl group as defined herein wherein one or more of the hydrogen atoms has been replaced with one or more hydroxyl (i.e., -OH). In some embodiments, the hydroxyalkyl contains one OH. In other embodiments, the hydroxyalkyl contains two OH. In further embodiments, the hydroxyalkyl contains three OH. Examples of hydroxyalkyl groups include hydroxymethyl, hydroxyethyl (e.g., 1-hydroxyethyl, 2-hydroxyethyl), 1,2- dihydroxyethyl, hydroxypropyl (e.g., 2-hydroxypropyl, 3-hydroxypropyl), hydroxybutyl (e.g., 3-hydroxybutyl, 4-hydroxybutyl), 2-hydroxy-1-methylpropyl, 1,3-dihydroxyprop-2-yl, and the like.

[0020] The term “C1-C6aminoalkyl” as used by itself or as part of another group refers to an alkyl as defined herein that is substituted by one or more NH2. In some embodiments, the alkyl is substituted by at least one NH2. In other embodiments, the alkyl is substituted by one, two, or three NH2. Examples of cyanoalkyl groups include CH2NH2, CH2CH2NH2, CH2CH2CH2NH2, and C(CH3)2NH2.

[0021] The term “6-12 membered aryl” refers to a mono- or bicyclic- aromatic hydrocarbon ring structure having 6 or 12 carbon atoms in the ring. Examples of aryl groups include phenyl, indenyl, naphthyl, and 1,2,3,4-tetrahydronaphthyl.

[0022] The term “5-14 membered heteroaryl” refers to a mono- or bicyclic- aromatic ring structure having 5 to 14 ring atoms that include carbon atoms as well as up to four heteroatoms that are each independently selected from nitrogen, oxygen, or sulfur. Examples of heteroaryl groups include thienyl, benzo[b]thienyl, furanyl, benzofuryl, pyranyl, thiophenyl, isobenzofuranyl, benzooxazonyl, chromenyl, xanthenyl, 2H pyrrolyl, pyrrolyl, imidazolyl, pyrazolyl, pyridyl, pyrazinyl, pyrimidinyl, pyridazinyl, isoindolyl, 3H-indolyl, indolyl, indazolyl, purinyl, isoquinolyl, quinolyl, quinoxalyl, phthalazinyl, naphthyridinyl, cinnolinyl, triazolyl, tetrazolyl, thiadiazolyl, oxadiazolyl, quinazolinyl, pteridinyl, pyrimidinyl, thiazolyl, isothiazolyl, oxazolyl, isoxazolyl, furazanyl, pyrazolo[1,5-a]pyridinyl, pyrazolo[1,5-a]pyridinyl, benzoisothiazolyl, imidazol[1,5-a]pyridinyl, pyrrolo[1,2]pyridazinyl, benzo[d]thiazolyl, benzo[d]imidazolyl, benzo[d]oxazolyl, benzoisoxazolyl, isothiazolyl, and tetrahydropyrazolo[1,5-a]pyridinyl.

[0023] The term “5-14 membered heterocyclyl” as used by itself or as part of another group refers to non-aromatic, saturated or partially unsaturated, e.g., containing one or two double bonds, cyclic groups containing one, two, or three rings having from five to fourteen ring members, wherein at least one carbon atom of one of the rings is replaced with a heteroatom. Each heteroatom is independently selected from oxygen, sulfur, includingsulfoxide and sulfone, and / or nitrogen atoms, which can be oxidized or quaternized. Examples of heterocyclyl groups include azetidinyl, dioxanyl, tetrahydropyranyl, pyrrolidinyl, piperidinyl, morpholinyl, piperazinyl, pyrrolidinyl, indolinyl, oxetanyl, tetrahydrofuranyl, tetrahydrothiophenyl, azepanyl, aziridinyl, dioxolanyl, imidazolidinyl, pyrazolidinyl, thianyl, dithianyl, thiomorpholinyl, oxazepanyl, oxiranyl, tetrahydropyranyl, and the like. In some embodiments, the heterocyclyl group includes azetidinyl, pyrrolidinyl, piperidinyl, piperazinyl, morpholinyl, and 6-azaspiro[2.5]octanyl.

[0024] As used herein, the terms “salt”, “salts” or “salt form” refer to an acid addition or base addition salt of a described compound. Acid addition salts can be formed with inorganic acids and organic acids. Inorganic acids from which salts can be derived include, for example, hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, and the like. Organic acids from which salts can be derived include, for example, acetic acid, propionic acid, glycolic acid, oxalic acid, maleic acid, malonic acid, succinic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, toluenesulfonic acid, sulfosalicylic acid, and the like. It should be understood that the terms “salt”, “salts” or “salt form” refer to all stoichiometries of salts of a described compound, including, for example, bis salts of a described compound. Such bis salts would include, for example, bis acid salts such as a bis hydrochloride salt or bis hydrobromide salt as well as bis basic salts such as a bis sodium salt.

[0025] In some instances, compounds are repeated throughout the description, but may be given different reference numbers simply for ease of reading. For example, compounds 10A and 10B have the same structure but have different reference numbers due to the scheme in which they appear. To the extent that there are any discrepancies, the depicted structure takes priority over the reference number.

[0026] In some embodiments, R1in the method of preparing Formula (A-1) is C1-C6alkyl or C1-C6haloalkyl. In some embodiments, R1in the method of preparing Formula (A-1) is methyl.

[0027] In some embodiments, X in the method of preparing Formula (A-1) is Cl.

[0028] In some embodiments, L in the method of preparing Formula (A-1) is C1-C6alkylene. In some embodiments, L in the method of preparing Formula (A-1) is –CH2–.

[0029] In some embodiments, Q in the method of preparing Formula (A-1) is a 6-12 membered aryl, or 5-14 membered heteroaryl, each of which is optionally substituted with 1 to 3 groups selected from halo, C1-C6alkyl, C1-C6haloalkyl, C1-C6hydroxyalkyl, hydroxyl, C1-C6aminoalkyl, NH2, -NH(C1-C6alkyl), N(C1-C6alkyl)2, CN, and NO2. In someembodiments, Q in the method of preparing Formula (A-1) is a 5-7 membered heteroaryl substituted with -NH2.

[0030] In some embodiments, the compound of Formula (A-1) in the disclosed methods is: r a salt thereof.

[0031] In some embodiments, the compound of Formula (A-2) in the disclosed methods salt thereof.In some embodiments the compound of Formula (A-2) is a salt. In some embodiments the compound of Formula (A-2) is a bis-salt. In other embodiments the compound of Formula (A-2) is a bis-acid salt. In other embodiments the compound of Formula (A-2) is a bis-HCl salt.

[0032] Similarly, in some embodiments the compound of Formula (4) is a salt. In some embodiments the compound of Formula (4) is a bis-salt. In other embodiments the compound of Formula (4) is a bis-acid salt. In other embodiments the compound of Formula (4) is a bis- HCl salt.

[0033] In some embodiments, the compound of Formula (A-3) in the disclosed methods is: a salt thereof.

[0034] In some embodiments, the compound of Formula (A-2), or a salt thereof, is reacted with the compound of Formula (A-3) in the presence of an acid (e.g., hydrochloric acid, acetic acid, phosphoric acid, sulfuric acid, citric acid, formic acid, methanesulfonic acid, or p-toluenesulfonic acid ) and one or more solvents (e.g., a polar protic solvent such as water, methanol, ethanol, n-propanol, i-propanol, or t-butanol, or any combination thereof).In some embodiments, the compound of Formula (A-2), or a salt thereof, is reacted with the compound of Formula (A-3) in the presence of citric acid and one or more solvents (e.g., a polar protic solvent such as water, methanol, ethanol, n-propanol, i-propanol, or t-butanol, or any combination thereof). In some embodiments one or more solvents is an alcohol. In some embodiments, the compound of Formula (A-2), or a salt thereof, is reacted with the compound of Formula (A-3) in the presence of an acid (e.g., hydrochloric acid, acetic acid, phosphoric acid, sulfuric acid, citric acid, formic acid, methanesulfonic acid, or p- toluenesulfonic acid) and ethanol. In some embodiments, the compound of Formula (A-2), or a salt thereof, is reacted with the compound of Formula (A-3) in the presence of an acid (e.g., hydrochloric acid, acetic acid, phosphoric acid, sulfuric acid, citric acid, formic acid, methanesulfonic acid, or p-toluenesulfonic acid) and ethanol and at least one other solvent. In some embodiments, the compound of Formula (A-2), or a salt thereof, is reacted with the compound of Formula (A-3) in the presence of citric acid and ethanol. In other embodiments, the compound of Formula (A-2), or a salt thereof, is reacted with the compound of Formula (A-3) in the presence of citric acid and a mixture of ethanol and i-propanol (IPA) . In other embodiments, the compound of Formula (A-2), or a salt thereof, is reacted with the compound of Formula (A-3) in the presence of citric acid and n-propanol. In some embodiments, the compound of Formula (4), or a salt thereof, is reacted with the compound of Formula (8A) in the presence of an acid and one or more solvents. In some embodiments, the bis-acid salt of the compound of Formula (4) is reacted with the compound of Formula (8A) in the presence of an acid and one or more solvents. In some embodiments, the bis-acid salt of the compound of Formula (4) is reacted with the compound of Formula (8A) in the presence of a citric acid or hydrochloric acid and one or more alcohol solvents. In some embodiments, the bis-HCl salt of the compound of Formula (4) is reacted with the compound of Formula (8A) in the presence of a citric acid or hydrochloric acid and one or more of ethanol, isopropyl alcohol and n-propanol. In other embodiments the bis acid salt of the compound of Formula (4) is reacted with the compound of Formula (8A) in the presence of one or more solvents, without the presence of an additional acid.

[0035] In one embodiment, provided herein is a method of preparing a compound of Formula (A-3):(A-3), wherein R1is hydrogen, C1-C6alkyl, C1-C6haloalkyl, C1-C6hydroxyalkyl, C1-C6aminoalkyl, C1-C6cyanoalkyl C2-C6alkenyl, C2-C6alkynyl, or C3-C6cycloalkyl, wherein said C3-C6cycloalkyl is optionally substituted with 1-3 groups selected from halo, hydroxyl, NH2, and CN; and X is halo; the method comprising: hydrolyzing a compound of Formula (A-6):(A-6) wherein R2is C1-C6alkyl, C1-C6haloalkyl, C1-C6hydroxyalkyl, C1-C6aminoalkyl, C1- C6cyanoalkyl C2-C6alkenyl, C2-C6alkynyl, or C3-C6cycloalkyl, wherein said C3-C6cycloalkyl is optionally substituted with 1 to 3 groups selected from halo, hydroxyl, NH2, and CN to form compound of Formula (A-3). In an embodiment, R2is C1-C6alkyl.

[0036] In some embodiments, R1in the method of preparing a compound of Formula (A- 3) is C1-C6alkyl or C1-C6haloalkyl. In some embodiments, R1in the compound of Formula (A-3) is methyl.

[0037] In some embodiments, X in the method of preparing a compound of Formula (A- 3) is Cl or Br. In an embodiment, X in the method of preparing a compound of Formula (A-3) is Cl.

[0038] In some embodiments, R2in the method of preparing a compound of Formula (A- 3) is ethyl.

[0039] In some embodiments, the compound of Formula (A-3) in the disclosed methods is:

[0040] In some embodiments, the compound of Formula (A-6) in the disclosed methods is:

[0041] In some embodiments, the compound of Formula (A-4) in the disclosed methods is:

[0042] In some embodiments, the compound of Formula (A-6) is hydrolyzed in the presence of a base (e.g., lithium hydroxide (LiOH), calcium hydroxide (Ca(OH)2), potassium hydroxide (KOH), sodium hydroxide (NaOH), or barium hydroxide (Ba(OH)2)) and one or more solvents (e.g., a mixture of water and at least one polar aprotic solvent). In some embodiments, the compound of Formula (A-6) is hydrolyzed in the presence of a base (e.g., lithium hydroxide (LiOH), calcium hydroxide (Ca(OH)2), potassium hydroxide (KOH), sodium hydroxide (NaOH), or barium hydroxide (Ba(OH)2)), water, and at least one polar aprotic solvent. In some embodiments, the compound of Formula (A-6) is hydrolyzed in the presence of a base (e.g., lithium hydroxide (LiOH), calcium hydroxide (Ca(OH)2), potassium hydroxide (KOH), sodium hydroxide (NaOH), or barium hydroxide (Ba(OH)2)), water, and at least one polar aprotic solvent selected from dimethylacetamide (DMAC), 2-methyl tetrahydrofuran (2-Me THF), tetrahydrofuran (THF), acetonitrile (ACN), or dimethylsulfoxide (DMSO), or any combination thereof. In some embodiments, the compound of Formula (A-6) is hydrolyzed in the presence of sodium hydroxide (NaOH), water, and at least one polar aprotic solvent. In some embodiments, the compound of Formula (A-6) is hydrolyzed in the presence of sodium hydroxide (NaOH), water, and at least one polar aprotic solvent selected from dimethylacetamide (DMAC), 2-methyl tetrahydrofuran (2-Me THF), tetrahydrofuran (THF), acetonitrile (ACN), or dimethylsulfoxide (DMSO), or any combination thereof. In some embodiments, the compound of Formula (A-6) is hydrolyzed in the presence of sodium hydroxide (NaOH), water, and tetrahydrofuran.

[0043] In some embodiments, the compound of Formula (A-6) is prepared by reacting a compound of Formula (A-5):with phosphorus oxychloride (POCl3), dimethylformamide (DMF), and one or more additional solvents (e.g., a non-polar solvent such as dioxane, dichloroethane, benzene, toluene, chlorobenzene, dichlorobenzene, or xylene) to form the compound of Formula (A-6). In some embodiments, the compound of Formula (A-6) is prepared by reacting a compound of Formula (A-5) with phosphorus oxychloride (POCl3), dimethylformamide (DMF), in chlorobenzene.

[0044] In some embodiments, the compound of Formula (A-5) in the disclosed methods is:

[0045] In some embodiments, the compound of Formula (A-5) is prepared by reacting a compound of Formula (A-4):with a halogenating agent (e.g., elemental halogen, carbon tetrachloride, hexachloroethane, carbon tetrabromide, n-bromosuccinmide (NBS), or n-chlorosuccinimide (NCS)), a base (e.g., lithium diisopropylamide (LDA), sodium bis(trimethylsilyl)amide (NaHMDS), lithium bis(trimethylsilyl)amide (LiHMDS), potassium bis(trimethylsilyl)amide (KHMDS), and sodium hydride (NaH)) and at least one solvent (e.g., a polar aprotic solvent such as dimethylacetamide (DMCA), 2-methyl tetrahydrofuran (2-Me THF), tetrahydrofuran (THF), acetone, dimethylformamide (DMF), acetonitrile (ACN), and dimethylsulfoxide (DMSO), or any combination thereof) to form the compound of Formula (A-5).

[0046] In some embodiments, the compound of Formula (A-5) is prepared by reacting a compound of Formula (A-4) with hexachloroethane, a base (e.g., lithium diisopropylamide (LDA), sodium bis(trimethylsilyl)amide (NaHMDS), lithium bis(trimethylsilyl)amide (LiHMDS), potassium bis(trimethylsilyl)amide (KHMDS), and sodium hydride (NaH),) andat least one solvent (e.g., a polar aprotic solvent) to form the compound of Formula (A-5). In some embodiments, the compound of Formula (A-5) is prepared by reacting a compound of Formula (A-4) with hexachloroethane, lithium bis(trimethylsilyl)amide (LiHMDS), and at least one solvent (e.g., a polar aprotic solvent) to form the compound of Formula (A-5). In some embodiments, the compound of Formula (A-5) is prepared by reacting a compound of Formula (A-4) with hexachloroethane, lithium bis(trimethylsilyl)amide (LiHMDS), and at least one solvent (e.g., a polar aprotic solvent) to form the compound of Formula (A-5). In some embodiments, the compound of Formula (A-5) is prepared by reacting a compound of Formula (A-4) with hexachloroethane, lithium bis(trimethylsilyl)amide (LiHMDS), in THF to form the compound of Formula (A-5).

[0047] In some embodiments, the compound of Formula (A-4) in the disclosed methods is:(5A) .

[0048] In one embodiment, provided herein is a method of preparing compound of Formula (B-1):wherein R1is hydrogen, C1-C6alkyl, C1-C6haloalkyl, C1-C6hydroxyalkyl, C1-C6aminoalkyl, C1-C6cyanoalkyl C2-C6alkenyl, C2-C6alkynyl, or C3-C6cycloalkyl, wherein said C3-C6cycloalkyl is optionally substituted with 1-3 groups selected from halo, hydroxyl, NH2, and CN; R4is C1-C6alkyl, C1-C6haloalkyl, 6-12 membered aryl, or 5-14 membered heteroaryl, wherein said 6-12 membered aryl or 5-14 membered heteroaryl is optionally substituted with 1 to 3 groups selected from halo or C1-C6alkyl; L is a bond or C1-C6alkylene; and Q is C3-C6cycloalkyl, 5-14 membered heterocyclyl, 6-12 membered aryl, or 5-14 membered heteroaryl, each of which is optionally substituted with 1-3 groups selected fromhalo, C1-C6alkyl, C1-C6haloalkyl, C1-C6hydroxyalkyl, hydroxyl, C1-C6aminoalkyl, -NH2, - NH(C1-C6alkyl), -N(C1-C6alkyl)2, CN, and NO2; the method comprising: reacting a compound of Formula (A-2):(A-2) or a salt thereof, with a compound of Formula (B-5):to form the compound of Formula (B-1).

[0049] In some embodiments, the compound of Formula (A-2), or a salt thereof, is reacted with the compound of Formula (B-5) in the presence of an acid (e.g., hydrochloric acid, acetic acid, trifluoroacetic acid, phosphoric acid, sulfuric acid, citric acid, formic acid) and at least one solvent (e.g., a polar protic solvent such as methanol, ethanol, n-propanol, i- propanol, sec-butanol, or t-butanol, or any combination thereof). In some embodiments, the compound of Formula (A-2), or a salt thereof, is reacted with the compound of Formula (B-5) in the presence of an acid (e.g., hydrochloric acid, trifluoroacetic acid, acetic acid, phosphoric acid, sulfuric acid, citric acid, formic acid, or carbonic acid) in ethanol or n-propanol or any combination thereof. In some embodiments the compound of Formula (A-2) is a bis salt. In some embodiments the compound of Formula (A-2) is a bis acid salt. In some embodiments the compound of Formula (A-2) is a bis HCl salt.

[0050] In some embodiments, the compound of Formula (B-5) in the disclosed methods is prepared by hydrolyzing a compound of the Formula (B-4):wherein R2is C1-C6alkyl, C1-C6haloalkyl, C1-C6hydroxyalkyl, C1-C6aminoalkyl, C1- C6cyanoalkyl C2-C6alkenyl, C2-C6alkynyl, or C3-C6cycloalkyl, wherein said C3-C6cycloalkyl is optionally substituted with 1-3 groups selected from halo, hydroxyl, NH2, and CN. In another embodiment, R2is C1-C6alkyl.

[0051] In some embodiments, the compound of Formula (B-4) is hydrolyzed in the presence of a base (e.g., lithium hydroxide (LiOH), calcium hydroxide (Ca(OH)2), potassium hydroxide (KOH), sodium hydroxide (NaOH), or barium hydroxide (Ba(OH)2)) and one or more solvents (e.g., a mixture of water and at least one polar aprotic solvent). In some embodiments, the compound of Formula (B-4) is hydrolyzed in the presence of a base (e.g., lithium hydroxide (LiOH), calcium hydroxide (Ca(OH)2), potassium hydroxide (KOH), sodium hydroxide (NaOH), or barium hydroxide (Ba(OH)2)), water, and at least one polar aprotic solvent. In some embodiments, the compound of Formula (B-4) is hydrolyzed in the presence of a base (e.g., lithium hydroxide (LiOH), calcium hydroxide (Ca(OH)2), potassium hydroxide (KOH), sodium hydroxide (NaOH), or barium hydroxide (Ba(OH)2)), water, and at least one polar aprotic solvent selected from dimethylacetamide (DMAC), 2-methyl tetrahydrofuran (2-Me THF), tetrahydrofuran (THF), dimethylformamide (DMF), acetonitrile (ACN), or dimethylsulfoxide (DMSO), or any combination thereof. In some embodiments, the compound of Formula (B-4) is hydrolyzed in the presence of sodium hydroxide (NaOH), water, and at least one polar aprotic solvent. In some embodiments, the compound of Formula (B-4) is hydrolyzed in the presence of sodium hydroxide (NaOH), water, and at least one polar aprotic solvent selected from dimethylacetamide (DMAC), 2-methyl tetrahydrofuran (2-Me THF), tetrahydrofuran (THF), dimethylformamide (DMF), acetonitrile (ACN), or dimethylsulfoxide (DMSO), or any combination thereof. In some embodiments, the compound of Formula (B-4) is hydrolyzed in the presence of sodium hydroxide (NaOH), water, and tetrahydrofuran.

[0052] In some embodiments, the compound of Formula (B-4) is prepared by reacting a compound of Formula (B-3):with an oxidizing agent (e.g., hydrogen peroxide and sodium tungstate (Na2WO4), 39% peracetic acid, trichloroisocyanuric acid (TCCA) and (sodium hypochlorite) NaOCl, oxone, hydrogen peroxide and SeO2, or hydrogen peroxide and MoCl2O2) in at least one solvent (e.g., a non-polar solvent such as dichloromethane, toluene, chlorobenzene, dichlorobenzene,or xylene, or any combination thereof) to form a compound of Formula (B-4). In some embodiments, the compound of Formula (B-4) is prepared by reacting a compound of Formula (B-3) with hydrogen peroxide and sodium tungstate (Na2WO4)) in chlorobenzene to form a compound of Formula (B-4). In some embodiments, the compound of Formula (B-4) is prepared by reacting a compound of Formula (B-3) with an oxidizing agent selected from 39% peracetic acid; trichloroisocyanuric acid (TCCA) and (sodium hypochlorite) NaOCl; oxone; hydrogen peroxide and SeO2; or hydrogen peroxide and MoCl2O2; in at least one solvent (e.g., a polar solvent such as ethanol, tetrahydrofuran (THF), water, acetic acid, ethyl acetate, or any mixtures thereof) to form a compound of Formula (B-4).

[0053] In some embodiments, the method of forming the compound of Formula (B-4) further comprises at least one phase transfer catalyst such as methyl tri-octylammonium hydrogen sulfate or tetrabutylammonium hydrogen sulfate. In some embodiments, the phase transfer catalyst is methyl tri-octylammonium hydrogen sulfate.

[0054] In some embodiments, the method of forming the compound of Formula (B-4) further comprises at least one acid such as phenyl phosphonic acid and acetic acid. In some embodiments, the acid is phenyl phosphonic acid.

[0055] In some embodiments, the compound of Formula (B-3) is prepared by reacting a compound of Formula (B-2):with phosphorus oxychloride (POCl3), dimethylformamide (DMF), and one or more solvent(s) such as a non-polar solvent (e.g., toluene, chlorobenzene, dichlorobenzene, or xylene, or any combination thereof) to form the compound of Formula (B-3). In some embodiments, the compound of Formula (B-3) is prepared by reacting a compound of Formula (B-2) with phosphorus oxychloride (POCl3) and dimethylformamide (DMF) in chlorobenzene.

[0056] In some embodiments, R1in the methods of preparing the compound of Formula (B-1), Formula (B-5), Formula (B-4), Formula (B-3), and or Formula (B-2) is C1-C6alkyl or C1-C6haloalkyl. In some embodiments, R1in the methods of preparing the compound of Formula (B-1), Formula (B-5), Formula (B-4), Formula (B-3), and or Formula (B-2) is methyl.

[0057] In some embodiments, R2in the methods of preparing the compound of Formula (B-4), Formula (B-3), and or Formula (B-2) is ethyl.

[0058] In some embodiments, L in the methods of preparing the compound of Formula (B-1) is C1-C6alkylene. In some embodiments, L in the methods of preparing the compound of Formula (B-1) is –CH2–.

[0059] In some embodiments, Q in the methods of preparing the compound of Formula (B-1) is a 6-12 membered aryl, or 5-14 membered heteroaryl, each of which is optionally substituted with 1-3 groups selected from halo, C1-C6alkyl, C1-C6haloalkyl, C1-C6hydroxyalkyl, hydroxyl, C1-C6aminoalkyl, -NH2, -NH(C1-C6alkyl), N(C1-C6alkyl)2, -CN, or –NO2.In some embodiments, Q in the methods of preparing the compound of Formula (B-1) is a 5-7 membered heteroaryl substituted with -NH2.

[0060] In some embodiments R4in the methods of preparing the compound of (B-1) is C1-C6alkyl, C1-C6haloalkyl, 6-12 membered aryl, or 5-14 membered heteroaryl, wherein said 6-12 membered aryl or 5-14 membered heteroaryl is optionally substituted with 1 to 3 groups selected from halo and C1-C6alkyl. In some embodiments R4is 6-12 membered aryl substituted with C1-C6alkyl.

[0061] In some embodiments, the compound of Formula (B-1) in the disclosed methods is:

[0062] In some embodiments, the compound of Formula (B-2) in the disclosed methods is:

[0063] In some embodiments, the compound of Formula (B-3) in the disclosed methods is:

[0064] In some embodiments, the compound of Formula (B-4) in the disclosed methods is:

[0065] In some embodiments, the compound of Formula (B-5) in the disclosed methods is:(8B).

[0066] In some embodiments, the compound of Formula (A-2) in the disclosed methods is: salt thereofIn some embodiments compound 4 is a bis salt. In some embodiments compound 4 is a bis acid salt. In some embodiments compound 4 is a bis HCl salt.

[0067] In one embodiment, provided is a Method (C) of preparing the compound of Formula (C-1)or a salt or hydrate thereof, whereinR1is hydrogen, C1-C6alkyl, C1-C6haloalkyl, C1-C6hydroxyalkyl, C1-C6aminoalkyl, C1-C6cyanoalkyl, C2-C6alkenyl, C2-C6alkynyl, or C3-C6cycloalkyl, wherein said C3-C6cycloalkyl is optionally substituted with 1 to 3 groups selected from halo, hydroxyl, NH2, and CN; L is a bond or C1-C6alkylene; and Q is C3-C6cycloalkyl, 5-14 membered heterocyclyl, 6-12 membered aryl, or 5-14 membered heteroaryl, each of which is optionally substituted with 1-3 groups selected from halo, C1-C6alkyl, C1-C6haloalkyl, C1-C6hydroxyalkyl, hydroxyl, C1-C6aminoalkyl, -NH2, - NH(C1-C6alkyl), -N(C1-C6alkyl)2, CN, and NO2; the method comprising the steps of: a) reacting a compound of Formula (C-2):wherein P1is a protecting group; R3is C1-C6alkyl; with a compound of Formula (C-3):wherein X is halo or S(O)2R4; R4is C1-C6alkyl, C1-C6haloalkyl, 6-12 membered aryl, or 5-14 membered heteroaryl, wherein said 6-12 membered aryl or 5-14 membered heteroaryl is optionally substituted with 1 to 3 groups selected from halo or C1-C6alkyl; to form a compound of Formula (C-4):(C-4); and b) deprotecting the compound to form a compound of Formula (C-1).

[0068] In some embodiments, L in the method of preparing a compound of Formula (C- 1) is C1-C6alkylene. In some embodiments, L in the method of preparing a compound of Formula (C-1) is –CH2–.

[0069] In some embodiments, Q in the method of preparing a compound of Formula (C- 1) is a 6-12 membered aryl, or 5-14 membered heteroaryl, each of which is optionally substituted with 1 to 3 groups selected from halo, C1-C6alkyl, C1-C6haloalkyl, C1-C6hydroxyalkyl, hydroxyl, C1-C6aminoalkyl, NH2, -NH(C1-C6alkyl), N(C1-C6alkyl)2, CN, and NO2.In some embodiments, Q in the method of preparing a compound of Formula (C-1) is a 5-7 membered heteroaryl substituted with -NH2.

[0070] In some embodiments, the compound of Formula (C-1) that is prepared according to the methods described above is:

[0071] In some embodiments, the compound of Formula (C-1) can be isolated as a free- base form or a salt form. In some embodiments, the compound of Formula (C-1) can be isolated as a free base or a salt form of the compound:

[0072] In some embodiments, X in the method of preparing a compound of Formula (C- 1) is Cl. In some embodiments, the compound of Formula (C-3) is a compound of Formula (A-1). In other embodiments, the compound of Formula (C-3) is a compound of Formula (A- 1), wherein the compound of Formula (A-1) was prepared by the disclosed methods.

[0073] In some embodiments, X in the method of preparing a compound of Formula (C- 1) is S(O)2R4, and R4is 6-12 membered aryl, wherein said 6-12 membered aryl is optionally substituted with 1 to 3 groups selected from halo and C1-C6alkyl. In some embodiments, X in the method of preparing a compound of Formula (C-1) is S(O)2R4, and R4is 4-methyl benzene. In some embodiments, the compound of Formula (C-3) is a compound of Formula(B-1). In other embodiments, the compound of Formula (C-3) is a compound of Formula (B- 1), wherein the compound of Formula (B-1) was prepared by the disclosed methods.

[0074] In some embodiments, the compound of Formula (C-3) in the disclosed methods is:

[0075] In some embodiments, the compound of Formula (C-3) in the disclosed methods is:

[0076] In some embodiments, P1in the method of preparing a compound of Formula (C- 1) is t-butyloxycarbonyl (BOC), fluorenylmethyloxycarbonyl (FMOC), acetate, benzyl, trityl, tetrahydropyranyl ether (THP), pivaloyl, or tosylate. In some embodiments, P1is tetrahydropyranyl ether (THP).

[0077] In some embodiments, R3in the method of preparing a compound of Formula (C- 1) is t-butyl.

[0078] In some embodiments, the compound of Formula (C-2) is:

[0079] In some embodiments, the compound of Formula (C-4) is:

[0080] In some embodiments, the compound of Formula (C-2) is reacted with the compound of Formula (C-3) in the presence of a base (e.g., lithium bis(trimethylsilyl)amide (LiHMDS), , lithium t-butoxide (LiOtBu), sodium t-butoxide (NaOtBu), potassium t- butoxide (KOtBu), or sodium hydride (NaH)) and one or more solvents (e.g., polar aprotic solvent such as tetrahydrofuran (THF), 2-methyl tetrahydrofuran (2-MeTHF), acetone, dimethylformamide (DMF), dimethylacetamide (DMA), acetonitrile (ACN), or dimethylsulfoxide (DMSO), or any combination thereof). In some embodiments, the compound of Formula (C-2) is reacted with the compound of Formula (C-3) in the presence of lithium bis(trimethylsilyl)amide (LiHMDS) and one or more solvents (e.g., polar aprotic solvent such as tetrahydrofuran (THF), 2-methyl tetrahydrofuran (2-MeTHF), acetone, dimethylformamide (DMF), dimethylacetamide (DMA), acetonitrile (ACN), or dimethylsulfoxide (DMSO), or any combination thereof). In some embodiments, the compound of Formula (C-2) is reacted with the compound of Formula (C-3) in the presence of lithium t-butoxide (LiOtBu) and one or more solvents (e.g., polar aprotic solvent such as tetrahydrofuran (THF), 2-methyl tetrahydrofuran (2-MeTHF), acetone, dimethylformamide (DMF), dimethylacetamide (DMA), acetonitrile (ACN), or dimethylsulfoxide (DMSO), or any combination thereof). In some embodiments, the compound of Formula (C-2) is reacted with the compound of Formula (C-3) in the presence of lithium bis(trimethylsilyl)amide (LiHMDS) and tetrahydrofuran (THF). In some embodiments, the compound of Formula (C- 2) is reacted with the compound of Formula (C-3) in the presence of lithium t-butoxide (LiOtBu) and 2-methyl tetrahydrofuran (2-MeTHF) and dimethylacetamide (DMA), acetonitrile (ACN).

[0081] In some embodiments, the compound of Formula (C-4) is deprotected in the presence of an acid (e.g., hydrochloric acid, acetic acid, phosphoric acid, sulfuric acid, citric acid, trifluoroacetic acid, formic acid) and one or more solvents (e.g., a polar protic solvent such as water, methanol, ethanol, n-propanol, i-propanol, or t-butanol, or any combination thereof) to provide a compound of Formula (C-1). In some embodiments, the compound of Formula (C-4) is deprotected in the presence of hydrochloric acid and one or more solvents (e.g., a polar protic solvent such as water, methanol, ethanol, n-propanol, i-propanol, or t- butanol, or any combination thereof). In some embodiments, the compound of Formula (C-4) is deprotected in the presence of an acid (e.g., hydrochloric acid, acetic acid, phosphoric acid, sulfuric acid, citric acid, formic acid) and n-propanol. In some embodiments, the compound of Formula (C-4) is deprotected in the presence of hydrochloric acid and n-propanol.

[0082] In some embodiments, the compound of Formula (C-1) that is prepared according to the methods disclosed above is:or salt, tautomer, or hydrate thereof.

[0083] In one embodiment, provided herein is a compound having the structure:or a salt, tautomer or hydrate thereof.

[0084] In one embodiment, provided herein is a compound having the structure:or a salt, tautomer or hydrate thereof.

[0085] In one embodiment, provided herein is a compound having the structure:or a salt, tautomer or hydrate thereof.

[0086] In one embodiment, provided herein is a compound having the structure:or a salt, tautomer or hydrate thereof.

[0087] In one embodiment, provided herein is a compound having the structure:or a salt, tautomer or hydrate thereof.

[0088] In one embodiment, provided herein is a compound having the structure:or a salt, tautomer or hydrate thereof.

[0089] In one embodiment, provided herein is a compound having the structure:or a salt, tautomer or hydrate thereof.

[0090] In one embodiment, provided herein is a bis acid compound having the structure:or a tautomer or hydrate thereof.EXEMPLIFICATION

[0091] While a number of embodiments have been described, the scope of this disclosure is to be defined by the appended claims, and not by the specific embodiments that have been represented by way of example. The contents of all references (including literature references, issued patents, published patent applications, and co-pending patent applications) cited throughout this application are hereby expressly incorporated herein in their entireties by reference. Unless otherwise defined, all technical and scientific terms used herein are accorded the meaning commonly known to one with ordinary skill in the art.

[0092] Typical abbreviations used are outlined below.Route A:

[0093] Synthesis of tert-butyl (E)-2-((6-((tert-butoxycarbonyl)amino)pyridin-2- yl)methylene)hydrazine-1-carboxylate (2):

[0094] To a room temperature mixture of (1) (1 wt) and MeOH (2 vol) is added AcOH (0.05 vol, 0.2 eq). The mixture was warmed to 50°C and then a methanolic solution (1 vol) of Boc-hydrazine (0.61 wts) was added over 2.5 h. The mixture was stirred at 50°C until the reaction was complete. The mixture was cooled to 20°C, and water (1.5 vol) was charged over 1 h, stirred for 4 h more, and filtered. The wet cake was washed with 2:1 MeOH:water (1.5 vol), and dried under vacuum at 40°C for 20 h to recover (2) as an off-white solid.

[0095] Synthesis of tert-butyl 2-((6-((tert-butoxycarbonyl)amino)pyridin-2- yl)methyl)hydrazine-1-carboxylate (3):

[0096] A mixture of (2) (1 wt) in MeOH (8 vol) at 25°C was evacuated and flushed with nitrogen three times.10% Pd / C (0.04 wt) was charged, and the reactor was evacuated and flushed with hydrogen three times. The mixture was heated to 50°C, and the pressure was adjusted to 15-20 psi and stirred for 6-12 h. Once complete, the reaction was cooled to 20°C, and evacuated and flushed with nitrogen three times. The mixture was filtered quickly over Celite (0.6 wt), and the solid was rinsed with MeOH (1 wt). The filtrate was added to a reactor at 25°C. Water (7.5 vol) was added dropwise over 2 h to induce precipitation. The suspension was stirred for 3 h, filtered, washed with 4:3 MeOH:water (1 vol), and dried under vacuum at 50°C for 24 h to obtain (3) as an off-white solid.

[0097] Synthesis of 6-(hydrazineylmethyl)pyridin-2-amine dihydrochloride (4):

[0098] To a reactor containing HCl (6 N, 5 vol) was added (3) (1.0 wt) portion-wise while maintaining the internal temperature between 10-20°C. The mixture was heated to 50°C for 2 h (or until complete), cooled to 20°C, and treated with EtOH (10 vol) over 2 h toinduce precipitation and held for 2 h more. The suspension was filtered, washed with EtOH (2 vol), and dried under vacuum at 50°C for 24 h to obtain (4) as a white solid.

[0099] Synthesis of ethyl 4-methyl-4H-pyrrolo[2,3-d]thiazole-5-carboxylate (5A):

[0100] Commercially available 2,4-thiazolidindione is aromatized and brominated through treatment with phosphorous pentoxide and TBAB in a similar manner to that reported by Sampson (See, Grubb, M.; Schmidt, M. J.; Seed, A. J.; Sampson, P. Synthesis 2012, 44, 1026−1029), to produce 2,4-dibromothiazole. The formylation of 2,4- dibromothiazole at the 5 position to generate compound (6-1) is achieved via de-protonation with lithium-tetramethyl piperidene (TMP) and introduction of the formyl group from ethyl formate. The formyl group is reacted in an Erlenmeyer Plochl type reaction with the readily available N-benzoyl-glycine (Hippuric acid) and acetic anhydride and esterified under treatment with ethanol (See, Zhao, H. Koenig S.G. Dankwardt, J.W. Singh, S.P. OPRD 18(1):198–204, 2013). The removal of the bromine at the 2-thiazole position is achieved bytreatment with zinc metal to provide (6-2). The cyclization of (6-2) is carried out under copper catalyzed amidation and the benzoyl is removed via treatment with ethylenediamine. The final stage of N-methylation is carried out in acetone through treatment with dimethyl sulfate to afford compound (5A).

[0101] Synthesis of ethyl 2-chloro-4-methyl-4H-pyrrolo[2,3-d]thiazole-5-carboxylate (6A):

[0102] To a reactor containing (5A) (1 wt) and THF (10 vol) at 10°C was added hexachloroethane (1.35 wt). The mixture was cooled to -40°C and LiHMDS (1.0M in THF, 4.5 wt) was added dropwise over 1 h and was further stirred for 2 h while maintaining the temperature between -30 to -50°C. Once complete, AcOH (0.66 wt) was added at -40°C over 30 min and then the mixture was warmed to 25°C over 1 h. Water (5 vol) was added over 2 h at 25°C. The layers were separated. The organic layer was washed with 5% Na2SO4(5 wt), was concentrated under vacuum to 2.5 vol and DCM (6.65 wt) was added. After 20 min of agitation, the mixture was allowed to stand, and the layers were separated. The organic layer was concentrated under vacuum at 30°C to 2.5 vol. Acetone (3.85 wt) was added, and the mixture was concentrated again to 2.5 vol (2x). Acetone (2.5 vol) was added and the mixture was transferred dropwise to 5°C water (5.0 vol) over a 2 h period and the suspension was stirred at 5°C for up to 10 h, filtered and washed with a 5°C 4:3 water:acetone (4.5 vol) mixture. The wet cake was dried under vacuum at 25°C for 24 h to obtain (6A) as a yellow solid.

[0103] Synthesis of ethyl 2-chloro-6-formyl-4-methyl-4H-pyrrolo[2,3-d]thiazole-5- carboxylate (7A):

[0104] To a reactor containing (6A) (1 wt) and 1,2-dichlorobenzene (2.6 wt) at 25°C was added POCl3(2.5 wt). The mixture was heated to 100°C and then DMF (0.90 wt) was added over 12 h, and then the mixture was stirred at 100°C for 6 h or until complete. Once complete, the reaction was cooled to 50°C and DMF (5.25 wt) was added over 1 h. Thismixture was added to a cooled (10-20°C) solution of EtOH (9.5 wt) and water (6.0 wt) and the temperature was maintained below 30°C during the addition. The suspension that forms was stirred between 15-30°C for 20 h and was filtered. The wet cake was washed with 1:1 v:v EtOH:H2O (2.0 vol). The wet cake was slurried with 1:1 EtOH:H2O (5 vol) at 50°C for 2 h, cooled to 20°C, filtered, rinsed with 4:3 H2O:EtOH (3.8 vol) and dried under vacuum at 50°C for 24 h to provide (7A) as a light brown solid.

[0105] Synthesis of 2-chloro-6-formyl-4-methyl-4H-pyrrolo[2,3-d]thiazole-5- carboxylic acid (8A):

[0106] To a reactor containing (7A) (1 wt), THF (5 vol) and water (5 vol) at 15°C was added dropwise NaOH (5% w / w, 5.0 vol, 1.5 eq) over 30 min while keeping the temperature below 30°C. After the addition, the mixture is stirred at 20-30°C for 4 h or until complete. Once complete, HCl (1 N) was added at 15°C until pH 1-2, and this induced precipitation. The suspension was filtered, washed with water (8 vol), and the wet cake was dried under vacuum at 55°C for 24 h to provide (8A) as a brown solid (see, FIG.2).

[0107] Synthesis of 6-((6-aminopyridin-2-yl)methyl)-2-chloro-4-methyl-4,6-dihydro- 5H-thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-5-one (9A):

[0108] To a reactor containing (4) (0.95 wt) in EtOH (20 vol) at 20°C was added sodium citrate monobasic (1.75 wt) and anhydrous citric acid (0.39 wt), and the mixture was stirred for 30 min. Compound (8A) (1.0 wt) was added and the mixture was warmed to 75°C for up to 10 h (or until complete by IPC), and then was cooled to 10°C and was stirred for 10 h. Thesuspension was filtered, and the wet cake was washed with cold EtOH (2.0 vol). [Note: the following sequence can be repeated to improve the purity if desired. The wet cake was transferred to a reactor containing water (25 vol), was mixed at 20°C, and was treated with ammonium hydroxide (25 wt%) dropwise until the solution pH reached 8-9. The basified mixture was stirred at 20°C for up to 8 h, was filtered, the wet cake was washed with water (4 vol)]. Once the desired purity was obtained, the wet cake was dried under vacuum at 50-60°C for 24 h to produce (9A) as a light brown solid.

[0109] Synthesis of 2-(1-(tetrahydro-2H-pyran-2-yl)-1H-pyrazol-3-yl)acetic acid (11):

[0110] A reactor containing (10) (1.0 wt), TsOH-H2O (0.12 wt) and 2-MeTHF (8.0 vol) at 25°C was warmed to 60°C. DHP (1.2 wt) was added dropwise at 60°C, and the mixture was stirred at 60°C for 4 h. Upon full conversion, the mixture was cooled to 25°C, MTBE (3.75 wt) was added, and the reaction was cooled to 10°C. NaOH (1 N, 8.0 wt) was added dropwise at 10°C and then the mixture was warmed to 20°C. The layers were separated. To the aqueous layer was added DCM (5.0 vol). The biphasic mixture was cooled to 10°C and was treated with HCl (2 N) over 1 h until the pH was 3-4. After mixing at 20°C for 1 h, the two layers were allowed to separate, and the organic layer was reserved. The aqueous layer was re-extracted with DCM (5.0 vol) as above two more times. The combined organic layers were washed with water (3.0 vol), concentrated under vacuum in a reactor to 2.5 vol, EtOAc (10 vol) was added, and the mixture was concentrated to 2.5 vol. More EtOAc (5.0 vol) was added and again the mixture was concentrated to 2.5 vol. The concentrate was warmed to 55°C to produce a solution, was cooled to 35°C, and seed (0.010 wt) was added. The mixture was stirred at 35°C for 3 h, was cooled to 0°C over 8 h and was held at this temperature for 8 h. The suspension was filtered, washed with cold EtOAc (0.5 vol), and was dried under vacuum at 35°C for 25 h to afford (11) as an off-white solid.

[0111] Synthesis of tert-butyl 2-(1-(tetrahydro-2H-pyran-2-yl)-1H-pyrazol-3- yl)acetate (12):

[0112] To a reactor containing (11) (1.0 wt) and t-BuOH (2.5 vol) was added DMAP (0.03 wt) and THF (2.5 vol) at 25°C. After 30 min, the mixture was warmed to 50°C and then a solution of Boc2O (1.15 wt) in THF (2.0 vol) was added dropwise over at least 5 hours and then further stirred for up to 2 h. Upon completion, the reaction was cooled to 25°C, and heptane (5.0 vol) and water (5.0 vol) were both added. The mixture was stirred at 25°C for 2h and then the layers were separated. The organic layer was concentrated to 2 vol under vacuum. Heptane (10 vol) was added and again the mixture was concentrated to 2 vol. More heptane (10 vol) was added, and the diluted mixture was cycled through a charcoal filter for 24 h. The filtrate was concentrated to 2 vol and 2-MeTHF (6.3 vol) was added; this cycle was repeated at least two more times. This solution of (12) was used without further processing in the next step.

[0113] Synthesis of tert-butyl 2-(6-((6-aminopyridin-2-yl)methyl)-4-methyl-5-oxo- 5,6-dihydro-4H-thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-2-yl)-2-(1-(tetrahydro-2H- p

[0114] To a reactor containing (9A) (1.0 wt, 1.0 eq) and THF (13 vol) was added the 2- MeTHF solution of (12) (1.1 eq) and more THF (2.8 vol). The mixture was cooled to -25°C, and LiHMDS (1.0 M in THF, 7.0 wt, 2.5 eq) was added dropwise over at least 1 h. The mixture was stirred for 3 h at -25°C, was warmed to -10°C and was stirred at -10°C for 3 h. Once complete, AcOH (0.98 wt) in 2-MeTHF (2.0 vol) was added dropwise at -10°C until pH was 7-8. The mixture was stirred at -10°C for 2 h, warmed to 10°C over 1 h, water (8 vol) was added, and then was further warmed to 20°C and maintained for 1 h. The reaction was filtered over Celite (1.2 wt) and the Celite bed was washed with THF (2.0 vol). The biphasic mixture was allowed to separate, and the organic layer was washed with 10% Na2SO4 (5 vol) and then was concentrated to 4 vol under vacuum. Toluene (5.4 vol) was added, the mixture was concentrated to 4 vol, and this sequence was repeated at least once (KF <0.5%). Toluene (7.0 vol) was added, and the mixture was warmed to 60°C to dissolve all the solids. The reaction was cooled to 43°C, seed (0.05 wt) was added, the light suspension was stirred for 8 h at 43°C, and then heptane (10 vol) was added dropwise over 8 h. The suspension was cooled to 33°C and held for 24 h, and then filtered. The wet cake was washed with heptane (2.0 vol) and then dried under vacuum at 40°C for 24 h to produce (10A) as a brown solid (see, FIG.1).

[0115] Synthesis of 2-((1H-pyrazol-3-yl)methyl)-6-((6-aminopyridin-2-yl)methyl)-4- methyl-4,6-dihydro-5H-thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-5-one hydrate (14):

[0116] To a reactor containing (10A) (1.0 wt) and n-PrOH (10 vol) at 25°C was added conc. HCl (3.0 vol, 20 eq) over at least 30 min, and then the mixture was warmed to 75°C and stirred for at least 1 h (or until reaction is complete). The reaction was then was cooled to 25°C over 2 h. Isopropyl acetate (20 vol) was added, and the suspension was stirred for 4 h at 25°C and then filtered. The wet cake was washed with IPAc (3.0 vol), transferred to another reactor, and dissolved with water (20 vol) and warmed to 35°C. Ammonium hydroxide (25 wt%) was added until the solution pH was 3-4. Seed (0.005 wt) was added and after 2 h at 35°C, more ammonium hydroxide (25 wt%) was added over 2 h until the pH was 8-9. After 4 h, the suspension was filtered, and the wet cake was washed with water (4.0 vol) and was transferred to a reactor. n-Propanol (13 vol) and water (2.5 vol) were added to the wet cake, and the suspension was warmed to 80°C. Once total dissolution was achieved, the solution was cooled to 65°C, seed (0.005 wt) was added. The light suspension was stirred at 65°C for 2 h, cooled to 20°C over at least 3 h, held for 2 h, filtered, and washed with n- propanol / heptane (0.1 vol:1 vol) and then with heptane (2 vol). The wet cake was dried under vacuum at 30°C for 10 h to provide (14).

[0117] Synthesis of 2-((1H-pyrazol-3-yl)methyl)-6-((6-aminopyridin-2-yl)methyl)-4- methyl-4,6-dihydro-5H-thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-5-one phosphate salt (15):

[0118] All solvents were degassed by bubbling with nitrogen to remove oxygen. To a reactor containing DMSO (8.0 vol) and phosphoric acid (85%, 0.31 wt) at 25°C was added (14) (1.0 wt). The mixture was warmed to 55°C, and polish-filtered to a 50°C reactor followed by a 1.0 vol DMSO rinse. To the 50°C solution was added a 1 / 7.2 / 1.8 DMSO / EtOH / H2O mixture (1.3 wt) under nitrogen. Seed (0.010 wt) was added, and the light suspension was stirred at 50°C for 5 h. More of the 1 / 7.2 / 1.8 DMSO / EtOH / H2O mixture (20wt) was added over at least 4 h, and then the suspension was cooled to 25°C over 3 h and held at 25°C for 7 h. The suspension was wet-milled at 25°C for at least 2 h, filtered, and rinsed with 2.5 / 4 / 1 DMSO / EtOH / H2O (2 vol) and then with 4 / 1 EtOH / H2O (2 vol) at least three times until DMSO <4000 ppm. The wet cake was dried under vacuum at 50°C for 20 h to provide (15) as an off-white solid. Route B:Scheme 3

[0119] Synthesis of tert-butyl (E)-2-((6-((tert-butoxycarbonyl)amino)pyridin-2- yl)methylene)hydrazine-1-carboxylate (2):

[0120] To a room temperature mixture of (1) (1 wt) and MeOH (2 vol) is added AcOH (0.05 vol, 0.2 eq). The mixture was warmed to 50°C and then a methanolic solution (1 vol) of Boc-hydrazine (0.62 wts) was added over 2.5 h. The mixture was stirred at 50°C for at least 2 h (or until an IPC showed the reaction was complete). The mixture was charged with water (3.5 vol) over 2 h, cooled to 20°C over 7 h, and filtered. The wet cake was washed with 2:1 MeOH:water (1.5 vol), and dried under vacuum at 60°C for 20 h to recover (2) as an off-white solid.

[0121] Synthesis of tert-butyl 2-((6-((tert-butoxycarbonyl)amino)pyridin-2- yl)methyl)hydrazine-1-carboxylate (3):

[0122] A mixture of (2) (1 wt) in MeOH (8 vol) at 25°C was evacuated and flushed with nitrogen three times.10% Pd / C (0.04 wt) was charged, and the reactor was evacuated and flushed with hydrogen three times. The mixture was heated to 50°C, and the pressure was adjusted to 15-20 psi and stirred for at least 6 h. Once complete, the reaction was cooled to 20°C, and evacuated and flushed with nitrogen three times. The mixture was filtered quickly over Celite (0.6 wt), and the solid was rinsed with MeOH (2 vol). The filtrate was added to a reactor at 25°C. Water (7.5 vol) was added dropwise over 2 h to induce precipitation. The suspension was stirred for 12 h, filtered, washed with 4:3 MeOH:water (1 vol), and dried under vacuum at 50°C for 24 h to obtain (3) as an off-white solid.

[0123] Synthesis of 6-(hydrazineylmethyl)pyridin-2-amine (4):

[0124] To a reactor containing HCl (6 N, 5 vol) at 15°C was added (3) (1.0 wt) portion- wise while maintaining the internal temperature between 10-20°C. The mixture was heated to 50°C for 2 h (or until complete), cooled to 20°C, and treated with EtOH (10 vol) over 2 h to induce precipitation and held for 2 h more. The suspension was filtered, washed with EtOH (2 vol), and dried under vacuum at 50°C for 20 h to obtain (4) as a white solid.

[0125] Synthesis of ethyl 6-formyl-4-methyl-2-(p-tolylthio)-4H-pyrrolo[2,3- d]thiazole-5-carboxylate (6B):

[0126] A reactor at 25°C was charged with (5B) (1 wt), chlorobenzene (3.5 vol) and POCl3(1.12 vol, 4.0 eq), and was warmed to 100°C. DMF (0.70 vol, 3.0 eq) was added dropwise over at least 4 h and the reactor was maintained at 100°C for at least another 4 h (or longer if needed) to reach completion. The temperature was lowered to 20°C, and the mixture was added over 6 h to a separate reactor containing 5% Na2SO4 (10 vol) at 5°C. EtOAc (5.0 vol) was added, the biphasic mixture was stirred for 2 h at 20°C, and the layers were allowed to separate. The organic phase was slowly neutralized to pH 7-9 by adding NaHCO3(~5 vol) at 20°C. The organic phase was separated, washed with water (5 vol), and then cycled through a charcoal filter for 15 h at 25°C. The filtrate was concentrated to 3V under vacuum, warmed to 40°C, and charged with heptane (5.0 vol) over 3 h and then seed (0.005 wt). The light suspension was held at 40°C for 3 h, and then more heptane (3.0 vol) was added over 3 h at 40°C. The suspension was cooled slowly over 5 h to 0°C, held for 9 h at 0°C, and filtered. The wet cake was washed with rinsed with heptane (4.5 vol) and dried under vacuum at 50°C for at least 18 h to provide (6B) as a light yellow solid.

[0127] Synthesis of ethyl 6-formyl-4-methyl-2-tosyl-4H-pyrrolo[2,3-d]thiazole-5- carboxylate (7B):

[0128] To a reactor was charged (6B) (1 wt), chlorobenzene (16 vol), sodium tungstate dihydrate (0.046 wt, 0.05 eq), phenylphosphonic acid (0.022 wt, 0.05 eq), and methyltri- octylammonium hydrogen sulfate (0.065 wt, 0.05 eq), and the mixture was stirred and cooled to 5°C. Hydrogen peroxide (30%, 0.79 wt, 2.5 eq) was added dropwise over 2 h at 5°C, andthen the reaction was warmed to 35°C and held for at least 4 h (or until complete). The reaction was cooled to 20°C and treated with 10% Na2SO3(10 vol) at 20°C over 2 h. The biphasic mixture was stirred for 2 h at 20°C, and the layers were allowed to separate. The organic layer was stirred with water (5 vol) for 1 h, the organic layer was warmed to 40°C, and heptane (16 vol) was added dropwise over 4 h. The mixture was held for 2 h, was cooled to 0°C over 4 h, held for 6 h, and then filtered. The wet cake was rinsed with heptane (3 vol) at 0°C and dried under vacuum at 45°C for at least 24 h to provide (7B) as an off-white solid.

[0129] Synthesis of 6-formyl-4-methyl-2-tosyl-4H-pyrrolo[2,3-d]thiazole-5- carboxylic acid (8B):

[0130] To a reactor containing (7B) (1 wt) and THF (10 vol) at 10°C was added aqueous LiOH (14%, 2.3 vol, 3.0 eq) over a 2 h period. The mixture was stirred at 20°C for at least 8 h (or until the reaction is complete). The reaction was cooled to 10°C, and aqueous HCl (2 N) was added until the pH was 1-2. The reaction was warmed to 20°C, stirred for 2 h, water (10 vol) was added, and the suspension was stirred for 7 h. The suspension was filtered, washed with water (6.0 vol), and dried under vacuum at 55°C for 24 h to provide (8B) as a yellow solid.

[0131] Synthesis of 6-((6-aminopyridin-2-yl)methyl)-4-methyl-2-tosyl-4,6-dihydro- 5H-thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-5-one (9B):

[0132] To a reactor containing (8B) (1.0 wt) in EtOH (12 vol) at 20°C was added (4) (1.1 eq), EtOH (7 vol) and i-PrOH (1 vol). Sodium citrate monobasic (1.18 wt) and citric acid (anhydrous, 0.26 wt) was added, and the mixture was warmed to 75°C and stirred for at least 40 h (or longer until the reaction was complete). The reaction was cooled to 20°C over 4 h and filtered. The wet cake was washed with ethanol (4.0 vol) and then transferred back to the reactor and dissolved in water (25 vol) at 20°C. Ammonium hydroxide (25%, ~1.2 vol) was added over 30 min to adjust the pH to 8-9, the suspension was stirred for 8 h at 20°C, andthen was filtered and washed with water (4.0 vol) and dried under vacuum at 55°C for 24 h to produce (9B) as an off-white solid.

[0133] Synthesis of tert-butyl 2-(1-(tetrahydro-2H-pyran-2-yl)-1H-pyrazol-3- yl)acetate (12):

[0134] To a reactor containing (11) (1.0 wt) and t-BuOH (2.5 vol) was added DMAP (0.03 wt) and THF (2.5 vol) at 25°C. After 30 min, the mixture was warmed to 50°C and then a solution of Boc2O (1.15 wt) in THF (2.0 vol) was added dropwise over at least 5 hours and then further stirred for up to 2 h. Upon completion, the reaction was cooled to 25°C, and heptane (5.0 vol) and water (5.0 vol) were both added. The mixture was stirred at 25°C for 2 h and then the layers were separated. The organic layer was concentrated to 2 vol under vacuum. Heptane (10 vol) was added and again the mixture was concentrated to 2 vol. More heptane (10 vol) was added, and the diluted mixture was cycled through a charcoal filter for 24 h. The filtrate was concentrated to 2 vol and 2-MeTHF (6.3 vol) was added; this cycle was repeated at least two more times. This solution of (12) was used without further processing in the next step.

[0135] Alternative Synthesis of tert-butyl 2-(6-((6-aminopyridin-2-yl)methyl)-4- methyl-5-oxo-5,6-dihydro-4H-thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-2-yl)-2-(1- (tetrahydro-2H-pyran-2-yl)-1H-pyrazol-3-yl)acetate (10B):

[0136] A reactor was charged with (9B) (1.0 wt, 1.0 eq), DMAc (5.0 vol), and the (12) solution (1.1 eq.) in 2-MeTHF. After mixing at 25°C, the reaction mixture was cooled to 0°C and charged dropwise with lithium tert-butoxide (1 M in THF, 6.5 vol, 3.0 eq) over 2 h under nitrogen. The mixture was stirred for 4 h at 0°C. If necessary, more lithium tert-butoxide solution (0.6 wt) was added and the mixture stirred for 4 h at 0°C. Once complete, aqueous acetic acid (6%, ~5.1 vol) was added to adjust the reaction pH to 5-7. The mixture was stirred for 1 h at 0°C, raised to 20°C, 2-MeTHF (5 vol) was charged, and the biphasic mixture was stirred for 1 h. The layers were separated. The organic layer was reserved, and the aqueous layer was extracted with 2-MeTHF (5.0 vol). The second organic layer was combined with the first and was washed in succession with 5% Na2SO4(5 vol), water (5 vol), 3% NaHCO3(5 vol, 2x), and water (5 vol). The organic layer was concentrated under vacuum to 5 vol, and two cycles of dilution with toluene (10 vol) and reconcentration (to 10 vol) was conducted. The solution was adjusted to 45°C. Seed (0.005 wt) was added, the light suspension was stirred for 2 h, heptane (2.0 vol) was added dropwise over 2 h, and then stirred for 2 h at 45°C. The suspension was cooled to 25°C over 4 h, held at 25°C for 15 h, filtered, washed with 5:1 toluene:heptane (4 vol) and then with heptane (4 vol), and dried under vacuum at 55°C for 24 h to produce (10B) as a light yellow solid.

[0137] Step 10A: Compound (10B) was recrystallized from a toluene / heptane solution. Route C:Scheme 5

[0138] Synthesis of ethyl 4-methyl-2-(methylthio)-4H-pyrrolo[2,3-d]thiazole-5- carboxylate (2C):

[0139] To a mixture of ethyl 2-bromo-4-methyl-4H-pyrrolo[2,3-d]thiazole-5-carboxylate ((1C) 500.0 mg, 1.73 mmol) in EtOH (10.0 mL) was added NaSMe (240.0 mg, 3.5 mmol). The reaction mixture was stirred at 25°C for 3 hours then quenched with ice water and extracted with DCM. The combined organic layers were washed with brine, dried over anhydrous Na2SO4and concentrated under reduced pressure to afford (2C) which was directly used in the next step without any purification. LC-MS: m / z 257 (M+H)+.

[0140] Synthesis of ethyl 6-formyl-4-methyl-2-(methylthio)-4H-pyrrolo[2,3- d]thiazole-5-carboxylate (3C):

[0141] To a solution of ethyl 4-methyl-2-(methylthio)-4H-pyrrolo[2,3-d]thiazole-5- carboxylate ((2C) 460.0 mg, 1.8 mmol) and N-methyl-N-phenylformamide (490 mg ,3.6 mmol) in DCE (10 mL) was added POCl3(550.0 mg,3.6 mmol). The resulting mixture was stirred at 130°C for 3 hours, then quenched with ice water and extracted with DCM. The combined organic layers were washed with brine, dried over anhydrous Na2SO4and concentrated under reduced pressure. The residue was purified by column chromatography on silica gel (eluent: PE / EtOAc = 8 / 1) to give the desired intermediate. LC-MS: m / z 285 (M+H)+. To a solution of the desired intermediate (300.0 mg, 1.06 mmol) in EtOH (5.0 mL) was added N2H4·H2O (2 mL, 98%wt). The reaction mixture was stirred at r.t. for 1 hour. then heated to 60°C overnight then cooled down. The solid was collected by filtration and dried under high vacuum to afford (3C). LCMS: m / z 253 (M+H)+.1H NMR (400 MHz, DMSO-d6) δ 12.61 (s, 1H), 8.48 (s, 1H), 4.22 (s, 3H), 2.81 (s, 3H).

[0142] Synthesis of 4-methyl-2-(methylsulfonyl)-4,6-dihydro-5H- thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-5-one--4-methyl-2-(methylthio)-4,6-dihydro- 5H-thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-5-one (4C):

[0143] Three necked flask charged with 4-methyl-4,6-dihydro-5H- thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-5-one ((3C), 30 g, 0.119 mol, 1.0 eq) in DCM (600 mL) m-CPBA (61.5 g, 3 eq) was added at 20°C in three portions. The mixture was stirred at 30°C overnight, LC-MS indicated 100% consumption of starting material, 20% of sulfoxide and 80% of sulfone were formed. The mixture was cooled to r.t., another portion of m-CPBA (1.0 eq) was added. The reaction mixture was stirred at 30°C for 2 hours, LC-MS indicated sulfoxide (LCMS: m / z 269 (M+H)+.) < 8%. The mixture was cooled to r.t. and filtered. The filtered cake was suspension in MeOH (500 mL) and stirred at r.t. for 1 hour. Solid was collected by filtration, washed with ethylacetate, dried in vacuum to afford a mixture of 5% of sulfoxide and 95% of sulfone. The mixture was suspended in DMSO (600 mL), heated to 120°C ~ 130°C to form a clear solution. Then cooled to r.t., solid precipitated. The mixture was filtered and dried to provide pure 4-methyl-2-(methylsulfonyl)-4,6-dihydro-5H- thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-5-one (4C), LCMS: m / z 285 (M+H)+.1H NMR (400 MHz, DMSO) δ12.87 (s, 1H), 8.69 (s, 1H), 4.32 (s, 3H), 3.56 (s, 3H).

[0144] Synthesis of methyl 1H-pyrazole-3-carboxylate--(1-((2- (trimethylsilyl)ethoxy)methyl)-1H-pyrazol-3-yl)methanol (6C):

[0145] At 0°C under N2atmosphere, to a stirred solution of methyl 1H-pyrazole-3- carboxylate ((5C), 90 g, 0.72 mol) in THF (1 L) was added NaH (20.7 g, 0.864 mol, 60%). The resulting mixture was slowly warmed up to r.t and stirred for 1 hour. The reaction mixture was then cooled back to 0°C and SEMCl (151.5 mL, 0.842 mol) was added drop wise. The stirring was continued for another 2 hours before quenched with sat. NH4Cl and extracted with ethyl acetate (3 x). The combined organic layers were washed with brine and dried over Na2SO4. Solvents were removed under vacuum to provide crude product which was used in the next step without purification.

[0146] At 0°C under N2atmosphere, to the suspension of LAH (16.9 g, 0.44 mol) in THF (760 mL) was added the crude methyl 1-((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazole-3- carboxylate. The resulting mixture was slowly warmed up to r.t. and stirred for 1 hour. The reaction mixture was cooled back to 0°C and H2O (15.6 mL), 10% NaOH (15.6 mL), H2O (15.6 mL) was added successively. The resulting mixture was filtered through a pad of celite and washed with MTBE (4 x). The combined organic fractions were dried over Na2SO4. Solvents were removed under reduced pressure to provide crude product (6C) which was used in the next step without purification. LC-MS: m / z 229 (M+H)+.

[0147] Synthesis of (1-((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazol-3-yl)methanol-- 3-(iodomethyl)-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazole (7C): Step 5

[0148] At 0°C under N2atmosphere, to a stirred solution of (1-((2- (trimethylsilyl)ethoxy)methyl)-1H-pyrazol-3-yl)methanol ((6C), 61.5 g, theoretically 0.262 mol) in THF (310 mL) was added TEA (55.42 mL, 0.393 mol) followed by MsCl (24 mL, 0.314 mol). The reaction was warmed up to r.t and stirred for 1 hour before the introduction of NaI (196.5 g, 1.31 mol, in 310 mL DMF). The resulting mixture was stirred for another 1 hour and quenched with ice-water, extracted with MTBE (3 x). The combined organic layers were washed with sat. Na2S2O3and brine, dried over Na2SO4and concentrated to provide (7C) used in the next step without purification. LC-MS: m / z 339 (M+H)+.

[0149] Synthesis of 3-((phenylsulfonyl)methyl)-1-((2-(trimethylsilyl)ethoxy)methyl)- 1H-pyrazole (8C):

[0150] At 0°C under N2atmosphere, to a stirred solution of (1-((2- (trimethylsilyl)ethoxy)methyl)-1H-pyrazol-3-yl)methanol ((7C) 77.5 g, theoretically 0.229 mol) in DMF (600 mL) was added sodium benzenesulfinate (53.5 g, 0.32 mol) and stirred for 1 hour at 0°C. After warmed up to r.t., the reaction mixture was quenched with ice-water and sat. Na2S2O3, extracted with ethyl acetate (3 x). The combined organic layers were washed with sat. NaHCO3and brined successively, dried over Na2SO4. Solvents were removed undervacuum and the residue was purified by flash chromatography (silica gel, 20% ~ 70% ethyl acetate in petroleum ether) to provide (8C) as a light-yellow oil. LCMS: [M + H]+353.1H NMR (400 MHz, DMSO) δ 7.85–7.77 (m, 4H), 7.62 (dd, 2H), 6.19 (d, 1H), 5.35 (d, 2H), 4.70 (d, 2H), 3.44-3.38 (m, 2H), 0.88–0.77 (m, 2H), -0.01 (s, 9H).

[0151] Synthesis of tert-butyl (tert-butoxycarbonyl)(3- (iodomethyl)phenyl)carbamate--tert-butyl (tert-butoxycarbonyl)(6-((4-methyl-2- (methylsulfonyl)-5-oxo-4,5-dihydro-6H-thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-6- yl)methyl)pyridin-2-yl)carbamate--4-methyl-2-(methylsulfonyl)-4,6-dihydro-5H- thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-5-one (9C):

[0152] A mixture of 4-methyl-2-(methylsulfonyl)-4H-thiazolo[5',4':4,5]pyrrolo[2,3- d]pyridazin-5(6H)-one ((4C), 7.5 g, 26.4^mmol) and K3PO4(8.3 g, 39.3^mmol) in anhydrous MeCN (300 mL) was stirred at 70°C for 1 hour under N2. Followed a solution of tert-butylN- [(tert-butoxy)carbonyl]-N-[6-(bromomethyl)pyridin-2-yl]carbamate (11.2 g, 29.0^mmol) in MeCN (30 mL) was added. After stirred at 70°C for 2.5 hours under N2, the reaction mixture was quenched with sat. NH4Cl and extracted with EA (300 mL X 3). The combined organic layers were washed with water and brine, dried over Na2SO4, filtered and the organic phase was concentrated. The crude product was purified by flash chromatography (silica gel, 0 ~ 50% ethyl acetate in petroleum ether) to give tert-butylN-[(tert-butoxy)carbonyl]-N-[6-({4- methanesulfonyl-7-methyl-9-oxo-3-thia-5,7,10,11-tetraazatricyclo[6.4.0.0{2,6}]dodeca- 1(8),2(6),4,11-tetraen-10-yl}methyl)pyridin-2-yl]carbamate (9C). LC-MS (ESI) found: 591.1 (M+H)+.

[0153] Synthesis of tert-butyl (6-((4-methyl-5-oxo-2-((phenylsulfonyl)(1-((2- (trimethylsilyl)ethoxy)methyl)-1H-pyrazol-3-yl)methyl)-4,5-dihydro-6H- thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-6-yl)methyl)pyridin-2-yl)carbamate--tert- butyl (tert-butoxycarbonyl)(6-((4-methyl-2-(methylsulfonyl)-5-oxo-4,5-dihydro-6H- thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-6-yl)methyl)pyridin-2-yl)carbamate--3- ((phenylsulfonyl)methyl)-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazole (10C):

[0154] To a stirred mixture of 3-((phenylsulfonyl)methyl)-1-((2- (trimethylsilyl)ethoxy)methyl)-1H-pyrazole ((9C), 11.9 g, 33.8^mmol) in anhydrous THF (200 mL) was added LiHMDS (50 mL, 1 M in THF) at -40°C under argon. After 10^min, the mixture was warmed up to 10°C and stirred for 1 hour, then (9) (9.1 g, 15.4^mmol in 35 mL THF) was added. The reaction was stirred at 10°C for another 30 min. The reaction mixture was poured into aq. NH4Cl, extracted with EtOAc (200 mL X 3). The combined organic layers were washed with water and brine, dried over anhydrous Na2SO4and concentrated. The crude product was purified by^flash chromatography (silica gel, 0 ~ 50% ethyl acetate in petroleum ether) to give tert-butyl (6-((4-methyl-5-oxo-2-((phenylsulfonyl)(1-((2- (trimethylsilyl)ethoxy)methyl)-1H-pyrazol-3-yl)methyl)-4H-thiazolo[5',4':4,5]pyrrolo[2,3- d]pyridazin-6(5H)-yl)methyl)pyridin-2-yl)carbamate (10C). LC-MS (ESI) found: 763.2 (M+H)+.

[0155] Synthesis of tert-butyl (6-((4-methyl-5-oxo-2-((1-((2- (trimethylsilyl)ethoxy)methyl)-1H-pyrazol-3-yl)methyl)-4,5-dihydro-6H- thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-6-yl)methyl)pyridin-2-yl)carbamate--tert- butyl (6-((4-methyl-5-oxo-2-((phenylsulfonyl)(1-((2-(trimethylsilyl)ethoxy)methyl)-1H- pyrazol-3-yl)methyl)-4,5-dihydro-6H-thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-6- yl)methyl)pyridin-2-yl)carbamate (11C):

[0156] A solution of (10C) (6.0 g, 7.86^mmol) in EtOH / AcOH (35 mL / 50 mL) was heated to 50°C with vigorously stirred in the presence of Zn (2.55 g, 117.9^mmol) for 40 min. Additional zinc were added every 40 min (2.55 g, twice, monitor the reaction by TLC / LC- MS to avoid the by-product and over reduced product). The solution was filtered and the filter cake was washed with DCM. The filtrate was partly evaporated, neutralized with saturated NaHCO3solution, dried over MgSO4and the solvent was removed under vacuum.The crude product was purified by flash chromatography (silica gel, DCM: MeOH = 40:1) to give (11C). LC-MS (ESI) found: 623.3 (M+H)+.

[0157] Synthesis of tert-butyl (6-((4-methyl-5-oxo-2-((1-((2- (trimethylsilyl)ethoxy)methyl)-1H-pyrazol-3-yl)methyl)-4,5-dihydro-6H- thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-6-yl)methyl)pyridin-2-yl)carbamate--2-((1H- pyrazol-3-yl)methyl)-6-((6-aminopyridin-2-yl)methyl)-4-methyl-4,6-dihydro-5H- thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-5-one (13):

[0158] To a mixture of (11C) (3.0 g, 4.8^mmol) in ethanol (30 mL) was added HCl (30 mL, 4 M in dioxane). The reaction mixture was stirred at 80°C for 40 min. The reaction mixture was cooled down to r.t., filtered and the solid was collected, suspended in water and neutralized with aqueous NaHCO3at 10°C. Filtered to give the desired compound (13). LC- MS (ESI) found: 393.2 (M+H)+.1HNMR (400 MHz, DMSO-d6) δ 12.78 (s, 1H), 8.53 (s, 1H), 7.72(s, 1H), 7.25 (dd, 1H), 6.33–6.24 (m, 2H), 6.08 (d, 1H), 5.90 (s, 2H), 5.19 (s, 2H), 4.49 (s, 2H), 4.26 (s, 3H).

Claims

Listing of Claims:

1. A method of preparing a compound of Formula (A-1): or a salt thereof, whereinR1is hydrogen, C1-C6alkyl, C1-C6haloalkyl, C1-C6hydroxyalkyl, C1-C6aminoalkyl, C1-C6cyanoalkyl C2-C6alkenyl, C2-C6alkynyl, or C3-C6cycloalkyl, wherein said C3-C6cycloalkyl is optionally substituted with 1 to 3 groups selected from halo, hydroxyl, NH2, and CN; X is halo; L is a bond or C1-C6alkylene; and Q is C3-C6cycloalkyl, 5-14 membered heterocyclyl, 6-12 membered aryl, or 5-14 membered heteroaryl, each of which is optionally substituted with 1-3 groups selected from halo, C1-C6alkyl, C1-C6haloalkyl, C1-C6hydroxyalkyl, hydroxyl, C1-C6aminoalkyl, -NH2, - NH(C1-C6alkyl), -N(C1-C6alkyl)2, CN, and NO2; the method comprising reacting a compound of Formula (A-2):(A-2) or a salt thereof, with a compound of Formula (A-3):to form the compound of Formula (A-1), or a salt thereof.

2. The method of claim 1, wherein R1is C1-C6alkyl or C1-C6haloalkyl.

3. The method of claim 1 or claim 2, wherein R1is methyl.

4. The method of any one of claims 1-3, wherein X is Cl.

5. The method of any one of claims 1-4, wherein L is C1-C6alkylene.

6. The method of any one of claims 1-5, wherein L is –CH2–.

7. The method of any one of claims 1-6, wherein Q is a 6-12 membered aryl, or 5-14 membered heteroaryl, each of which is optionally substituted with 1 to 3 groups selected from halo, C1-C6alkyl, C1-C6haloalkyl, C1-C6hydroxyalkyl, hydroxyl, C1-C6aminoalkyl, NH2, -NH(C1-C6alkyl), N(C1-C6alkyl)2, CN, and NO2.

8. The method of any one of claims 1-7, wherein Q is a 5-7 membered heteroaryl substituted with -NH2.

9. The method of any one of claims 1-8, wherein the compound of Formula (A-1) is:

10. The method of any one of claims 1-9, wherein the compound of Formula (A-2) is:or a bis-acid salt thereof.

11. The method of any one of claims 1-9, wherein the compound of Formula (A-2) is: a tautomer or hydrate thereof.

12. The method of any one of claims 1-11, wherein the compound of Formula (A-3) is:.

13. The method of any one of claims 1-12, wherein the compound of Formula (A-2) or salt thereof is reacted with the compound of Formula (A-3) in the presence of an acid and one or more solvents.

14. The method of any of claims 1-13, wherein the compound of Formula (A-3) is reacted with a bis acid salt of the compound of Formula (A-2) in one or more solvents.

15. The method of claim 11 or 12, wherein the acid is selected from hydrochloric acid, acetic acid, phosphoric acid, sulfuric acid, citric acid, formic acid, methanesulfonic acid and p-toluenesulfonic acid.

16. The method of any one of claims 1-15, wherein at least one solvent is a polar protic solvent.

17. The method of claim 16, wherein at least one polar protic solvent is selected from water, methanol, ethanol, n-propanol, i-propanol, and t-butanol, or any combination thereof.

18. The method of claim 17, wherein at least one polar protic solvent is ethanol or n- propanol.

19. The method of claim 17, wherein the polar protic solvent is a mixture of ethanol and i-propanol.

20. A method of preparing a compound of Formula (A-3),wherein R1is hydrogen, C1-C6alkyl, C1-C6haloalkyl, C1-C6hydroxyalkyl, C1-C6aminoalkyl, C1-C6cyanoalkyl C2-C6alkenyl, C2-C6alkynyl, or C3-C6cycloalkyl, wherein said C3-C6cycloalkyl is optionally substituted with 1-3 groups selected from halo, hydroxyl, NH2, and CN; and X is halo;the method comprising: hydrolyzing a compound of Formula (A-6):wherein R2is C1-C6alkyl, to form compound of Formula (A-3).

21. The method of claim 20, wherein the compound of Formula (A-6) is hydrolyzed in the presence of a base and one or more solvents.

22. The method of claim 21, wherein the base is selected from lithium hydroxide (LiOH), calcium hydroxide (Ca(OH)2), potassium hydroxide (KOH), sodium hydroxide (NaOH), and barium hydroxide (Ba(OH)2).

23. The method of claim 22, wherein the base is sodium hydroxide (NaOH).

24. The method of any one of claims 20-23, wherein the compound of Formula (A-6) is hydrolyzed in the presence of water and one or more solvents.

25. The method of any one of claims 20-24, wherein the compound of Formula (A-6) is hydrolyzed in the presence of water and at least one solvent that is a polar aprotic solvent.

26. The method of claim 25, wherein at least one polar aprotic solvent is selected from dimethylacetamide (DMAC), 2-methyl tetrahydrofuran (2-Me THF), tetrahydrofuran (THF), acetone, dimethylformamide (DMF), acetonitrile (ACN), and dimethylsulfoxide (DMSO), or any combination thereof.

27. The method of claim 26, wherein at least one polar aprotic solvent is THF.

28. The method of any one of claims 20-27, wherein the compound of Formula (A-6) is prepared by reacting a compound of Formula (A-5):with phosphorus oxychloride (POCl3), dimethylformamide (DMF), and one or more additional solvents to form the compound of Formula (A-6).

29. The method of claim 28, wherein one or more additional solvents is a non-polar solvent.

30. The method of claim 29, wherein the non-polar solvent is selected from dioxane, dichloroethane, benzene, toluene, chlorobenzene, dichlorobenzene, and xylene.

31. The method of claim 30, wherein the non-polar solvent is chlorobenzene.

32. The method of any one of claims 28-31, wherein the compound of Formula (A-5) is prepared by reacting a compound of Formula (A-4):with a halogenating agent, a base and at least one solvent to form the compound of Formula (A-5).

33. The method of claim 32, wherein the base is selected from, lithium diisopropylamide (LDA), sodium bis(trimethylsilyl)amide (NaHMDS), lithium bis(trimethylsilyl)amide (LiHMDS), potassium bis(trimethylsilyl)amide (KHMDS), and sodium hydride (NaH).

34. The method of claim 33, wherein the base is lithium bis(trimethylsilyl)amide (LiHMDS).

35. The method of any one of claims 31-34, wherein the halogenating reagent is selected from elemental halogen, carbon tetrachloride, hexachloroethane, carbon tetrabromide, n- bromosuccinmide (NBS), and n-chlorosuccinimide (NCS).

36. The method of claim 35, wherein the halogenating reagent is hexachloroethane.

37. The method of any one of claims 31-36, wherein at least one solvent is a polar aprotic solvent.

38. The method of claim 37, wherein the polar aprotic solvent is selected from dimethylacetamide (DMAC), 2-methyl tetrahydrofuran (2-Me THF), tetrahydrofuran (THF), acetonitrile (ACN), and dimethylsulfoxide (DMSO), or any combination thereof.

39. The method of claim 38, wherein the polar aprotic solvent is THF.

40. The method of any one of claims 22-39, wherein R1is C1-C6alkyl or C1-C6haloalkyl.

41. The method of any one of claims 20-40, wherein R1is C1-C6alkyl.

42. The method of any one of claims 20-41, wherein R1is methyl.

43. The method of any one of claims 20-42, wherein X is –Cl.

44. The method of any one of claims 20-43, wherein R2is ethyl.

45. The method of any one of claims 20-44, wherein the compound of Formula (A-3) is.

46. The method of any one of claims 32-45, wherein the compound of Formula (A-4) is.

47. The method of any one of claims 20-46, wherein the compound of Formula (A-6) is.

48. The method of any one of claims 28-47, wherein the compound of Formula (A-5) is. \49. A method of preparing a compound of Formula (B-1):(B-1), wherein R1is hydrogen, C1-C6alkyl, C1-C6haloalkyl, C1-C6hydroxyalkyl, C1-C6aminoalkyl, C1-C6cyanoalkyl C2-C6alkenyl, C2-C6alkynyl, or C3-C6cycloalkyl, wherein said C3-C6cycloalkyl is optionally substituted with 1-3 groups selected from halo, hydroxyl, NH2, and CN; R4is C1-C6alkyl, C1-C6haloalkyl, 6-12 membered aryl, or 5-14 membered heteroaryl, wherein said 6-12 membered aryl or 5-14 membered heteroaryl is optionally substituted with 1 to 3 groups selected from halo or C1-C6alkyl; L is a bond or C1-C6alkylene; and Q is C3-C6cycloalkyl, 5-14 membered heterocyclyl, 6-12 membered aryl, or 5-14 membered heteroaryl, each of which is optionally substituted with 1-3 groups selected from halo, C1-C6alkyl, C1-C6haloalkyl, C1-C6hydroxyalkyl, hydroxyl, C1-C6aminoalkyl, -NH2, - NH(C1-C6alkyl), -N(C1-C6alkyl)2, CN, and NO2; the method comprising: reacting a compound of Formula (A-2):(A-2) or a salt thereof, with a compound of Formula (B-5):to form the compound of Formula (B-1).

50. The method of claim 49, wherein the compound of Formula (A-2) or a salt thereof is reacted with the compound of Formula (B-5) in the presence of an acid and at least one solvent.

51. The method of claim 50, wherein the acid is selected from hydrochloric acid, trifluoroacetic acid, acetic acid, phosphoric acid, sulfuric acid, citric acid, formic acid, and carbonic acid.

52. The method of any one of claims 49-51, wherein at least one solvent is a polar protic solvent.

53. The method of claim 52 wherein the polar protic solvent is an alcohol.

54. The method of claim 53, wherein at least one polar protic solvent is selected from methanol, ethanol, n-propanol, i-propanol, sec- butanol, and t-butanol, or any combination thereof.

55. The method of claim 54, wherein at least one polar protic solvent is ethanol, i- propanol or n-propanol or a mixture thereof.

56. The method of any one of claims 49-55, wherein the compound of Formula (B-5) is prepared by hydrolyzing a compound of the Formula (B-4):wherein R2is C1-C6alkyl.

57. The method of claim 56, wherein the compound of Formula (B-4) is hydrolized in the presence of a base and one or more solvents.

58. The method of claim 57, wherein the base is selected from lithium hydroxide (LiOH), calcium hydroxide (Ca(OH)2), potassium hydroxide (KOH), sodium hydroxide (NaOH), and barium hydroxide (Ba(OH)2).

59. The method of claim 58, wherein the base is lithium hydroxide (LiOH) or sodium hydroxide (NaOH).

60. The method of any one of claims 56-59, wherein at least one solvent is water.

61. The method of any one of claims 56-60, wherein the compound of Formula (B-4) is hydrolyzed in water and a polar aprotic solvent.

62. The method of claim 61, wherein the polar aprotic solvent is selected from dimethylacetamide (DMAC), 2-methyl tetrahydrofuran (2-Me THF), tetrahydrofuran (THF), dimethylformamide (DMF), acetonitrile (ACN), and dimethylsulfoxide (DMSO), or any combination thereof.

63. The method of claim 62, wherein the polar aprotic solvent is THF.

64. The method of any one of claims 56-63, wherein the compound of Formula (B-4) is prepared by reacting a compound of Formula (B-3):with an oxidizing agent in at least one solvent to form a compound of Formula (B-4).

65. The method of claim 64, wherein the oxidizing agent is hydrogen peroxide and sodium tungstate (Na2WO4).

66. The method of claim 63-65, wherein at least one solvent is a non-polar solvent.

67. The method of claim 66, wherein at least one non-polar solvent is selected from dichloromethane, toluene, chlorobenzene, dichlorobenzene, and xylene, or any combination thereof.

68. The method of claim 67, wherein the non-polar solvent is chlorobenzene.

69. The method of any one of claims 63-68, wherein the method further comprises at least one phase transfer catalyst.

70. The method of claim 69, wherein the phase transfer catalyst is methyl tri- octylammonium hydrogen sulfate.

71. The method of any one of claims 63-70, wherein the method further comprises at least one acid.

72. The method of claim 71, wherein the acid is phenyl phosphonic acid.

73. The method of any one of claims 63-72, wherein the compound of Formula (B-3) is prepared by reacting a compound of Formula (B-2):with phosphorus oxychloride (POCl3), dimethylformamide (DMF), and one or more solvent(s) to form the compound of Formula (B-3).

74. The method of claim 73, wherein one solvent is a non-polar solvent.

75. The method of claim 74, wherein the non-polar solvent is selected from toluene, chlorobenzene, dichlorobenzene, and xylene, or any combination thereof.

76. The method of claim 75, wherein the non-polar solvent is chlorobenzene.

77. The method of any one of claims 47-76, wherein R1is C1-C6alkyl or C1-C6haloalkyl.

78. The method of any one of claims 47-77, wherein R1is C1-C6alkyl.

79. The method of any one of claims 47-78, wherein R1is methyl.

80. The method of any one of claims 56-79, wherein R2is ethyl.

81. The method of any one of claims 56-80, wherein R4is 6-12 membered aryl optionally substituted with 1 to 3 groups selected from halo and C1-C6alkyl.

82. The method of any one of claims 56-81, wherein R4is 4-methyl benzene.

83. The method of any one of claims 49-82, wherein L is C1-C6alkylene.

84. The method of any one of claims 49-83, wherein L is –CH2–.

85. The method of any one of claims 49-84, wherein Q is a 6-12 membered aryl, or 5-14 membered heteroaryl, each of which is optionally substituted with 1-3 groups selected from halo, C1-C6alkyl, C1-C6haloalkyl, C1-C6hydroxyalkyl, hydroxyl, C1-C6aminoalkyl, -NH2, - NH(C1-C6alkyl), N(C1-C6alkyl)2, -CN, and –NO2.

86. The method of any one of claims 49-85, wherein Q is a 5-7 membered heteroaryl substituted with -NH2.

87. The method of any one of claims 49-86, wherein the compound of Formula (B-1) is.

88. The method of any one of claims 73-87, wherein the compound of Formula (B-2) is.

89. The method of any one of claims 64-88, wherein the compound of Formula (B-3) is.

90. The method of any one of claims 56-89, wherein the compound of Formula (B-4) is.

91. The method of any one of claims 49-90, wherein the compound of Formula (B-5) is.

92. The method of any one of claims 49-91, wherein the compound of Formula (A-2) is:.

93. A compound having the structure:or a salt, tautomer or hydrate thereof.

94. A compound having the structure:or a salt, tautomer or hydrate thereof.

95. A compound having the structure:or a salt, tautomer or hydrate thereof.

96. A compound having the structure:or a salt, tautomer or hydrate thereof.

97. A compound having the structure:or a salt, tautomer or hydrate thereof.

98. A compound having the structure:or a salt, tautomer or hydrate thereof.

99. A compound having the structure:or a salt, tautomer or hydrate thereof.

100. A compound having the structure:bis acid salt thereof.

101. A compound having the structure:or a tautomer or hydrate thereof.