NZ623089B2 - Processes for making compounds useful as inhibitors of atr kinase - Google Patents

Abstract

The present disclosure relates to processes and intermediates for preparing compounds useful as inhibitors of ATR kinase, such as aminopyrazine-isoxazole derivatives and related molecules. The present disclosure also relates to compounds useful as inhibitors of ATR protein kinase. The disclosure relates to pharmaceutically acceptable compositions comprising the compounds; methods of treating of various diseases, disorders, and conditions using the compounds; processes for preparing the compounds; intermediates for the preparation of the compounds; and solid forms of the compounds. The compounds have formula (I) or (II) wherein the variables are as defined herein. ates to pharmaceutically acceptable compositions comprising the compounds; methods of treating of various diseases, disorders, and conditions using the compounds; processes for preparing the compounds; intermediates for the preparation of the compounds; and solid forms of the compounds. The compounds have formula (I) or (II) wherein the variables are as defined herein.

Description

PROCESSES FOR NG COMPOUNDS USEFUL AS INHIBITORS OF ATR KINASE BACKGROUND OF THE ION ATR (“ATM and Rad3 related”) kinase is a protein kinase involved in cellular ses to DNA damage. ATR kinase acts with ATM (“ataxia telangiectasia mutated”) kinase and many other proteins to regulate a cell’s response to DNA damage, commonly referred to as the DNA Damage Response (“DDR”). The DDR stimulates DNA repair, promotes survival and stalls cell cycle progression by activating cell cycle oints, which provide time for repair. Without the DDR, cells are much more sensitive to DNA damage and readily die from DNA lesions induced by endogenous cellular processes such as DNA replication or ous DNA damaging agents commonly used in cancer therapy.

Healthy cells can rely on a host of different proteins for DNA repair including the DDR kinase ATR. In some cases these ns can compensate for one another by activating functionally redundant DNA repair processes. On the contrary, many cancer cells harbour defects in some of their DNA repair processes, such as ATM signaling, and therefore display a r reliance on their remaining intact DNA repair proteins which e ATR.

In addition, many cancer cells express activated oncogenes or lack key tumour suppressors, and this can make these cancer cells prone to dysregulated phases of DNA replication which in turn cause DNA damage. ATR has been implicated as a critical component of the DDR in response to disrupted DNA replication. As a result, these cancer cells are more dependent on ATR activity for survival than healthy cells. Accordingly, ATR inhibitors may be useful for cancer treatment, either used alone or in combination with DNA damaging agents, because they shut down a DNA repair mechanism that is more important for cellular al in many cancer cells than in healthy normal cells.

In fact, disruption ofATR function (e. g. by gene deletion) has been shown to promote cancer cell death both in the absence and presence ofDNA damaging agents. This ts that ATR inhibitors may be effective both as single agents and as potent sensitizers to radiotherapy or genotoxic chemotherapy.

SUBSTITUTE SHEET (RULE 26) For all of these reasons, there is a need for the development of potent and selective ATR inhibitors for the treatment of cancer, either as single agents or as combination therapies with radiotherapy or genotoxic chemotherapy. Furthermore, it would be desirable to have a synthetic route to ATR inhibitors that is le to large-scale synthesis and improves upon tly known methods.

ATR peptide can be sed and isolated using a variety of methods known in the literature (& e. g., Unsal-Kacmaz et al, PNAS 99: 10, pp6673—6678, May 14, 2002; & alfl Kumagai et al. Cill 124, 955, March 10, 2006; Unsal-Kacmaz et al. Molecular and Cellular Biology, Feb 2004, p1292-1300; and Hall—Jackson et al. Oncogene 1999, 18, 6707—6713).

DESCRIPTION OF THE FIGURES FIGURE 1a: XRPD Compound I—2 free base FIGURE 2a: TGA nd I—2 free base FIGURE 3a: DSC Compound I—2 free base FIGURE 4a: ORTEP plot of the asymmetric unit of the Compound I—2 free form single crystal structure FIGURE 1b: XRPD Compound I-2 ° HCl FIGURE 2b: TGA Compound I—2 ° HCl FIGURE 3b: DSC Compound L2 0 HCl FIGURE 4b: ORTEP plot of the asymmetric unit of the Compound I-2 ° HCl anhydrous ure.

FIGURE 1c: XRPD Compound I-2 ° 2HCl FIGURE 2c: TGA Compound I-2 ° 2HCl FIGURE 3c: DSC Compound L2 0 2HCl FIGURE 1d: XRPD Compound I-2 ° HCl monohydrate FIGURE 2d: TGA Compound I—2 ° HCl monohydrate FIGURE 3d: DSC Compound L2 0 HCl monohydrate FIGURE 1e: XRPD Compound I-2 ° HCl ° 2HZO FIGURE 2e: TGA Compound I-2 ° HCl ° 2H20 FIGURE 3e: DSC Compound L2 0 HCl 0 2HZO FIGURE 4a: Solid State Compound I—1 free base FIGURE 4b: Solid State 13CNMR of Compound 1-1 - HCl WO 49726 SUMMARY OF THE INVENTION The present ion relates to processes and intermediates for ing compounds useful as inhibitors of ATR kinase, such as aminopyrazine-isoxazole derivatives and related molecules. Aminopyrazine-isoxazole derivatives are useful as ATR inhibitors and are also useful for preparing ATR inhibitors. The present invention also relates to solid forms of ATR inhibitors as well as ated ATR inhibitors.

One aspect of the invention provides a process for preparing a compound of formula I: NH O’N 3» \\ / ”N’R KKN| J1 comprising preparing a compound of formula 4: HO—N R3 \”H/ ”N, from a compound of formula 3: R1_O 3 R2_O -’:/ PG under suitable oxime formation conditions.

Another aspect comprises preparing a compound of formula 4: from a compound of formula 3: R1—o ,R3 / Will R2_O -’:/ PG under suitable oxime formation conditions.

WO 49726 Another aspect of the present invention comprises a compound of formula II: R4 R38 O§S R3b o// R30 R1a R1 C or a pharmaceutically acceptable salt thereof,wherein each R”, Rlb, R”, R2, R3a, R3b, R36, R4, R5, R6, R7, R8, R93, R9b, R10, R“, R”, and R13 is ndently hydrogen or deuterium, andat least one of R13, Rlb, R”, R2, R33, R3b, R32 R4, R5, R6, R7, R8, R93, R9b, R10, R11, R12, and R13 is deuterium.

Yet another aspect of the invention provides solid forms of a compound of formula 1-2: NH2 O’N\ HN\ Oj:0 Other aspects of the invention are set forth herein.

The present invention has several advantages over usly known methods.

First, the present process has fewer number of total synthetic steps compared with previously disclosed processes. Second, the present process has improved yields over previously sed processes. Third, the present process is effective for compounds wherein R3 is a wide range of groups, such as alkyl groups or a large, hindered moiety, such as a ring. Fourth, the present process comprises intermediates which are more stable and have a longer shelf life. In certain embodiments, the non-acidic formation of the oxime group in the present s allows the preservation of acid—sensitive protecting groups such as Boc or CBz during the course of the synthesis. In other embodiments, the process is more easily scaled up to larger quantities due to the elimination of chromatography as a purification step.

DETAILED DESCRIPTION OF THE INVENTION One aspect of the invention provides a process for making a compound of preparing a compound of a 4: HO—N ,R3 \ / WN from a compound of formula 3: under suitable oxime formation conditions; wherein R1 is kyl; R2 is C1_6alkyl; or R1 and R2, together with the oxygen atoms to which they are attached, form an optionally tuted 5 or 6 membered saturated heterocyclic ring having two oxygen atoms; R3 is hydrogen, C‘1_6alkyl, or a 3-6 membered saturated or lly unsaturated heterocyclyl having 1-2 heteroatoms selected from the group consisting of , nitrogen, and sulfur; wherein the heterocyclyl is optionally substituted with l occurrence of halo or kyl; J1 is halo, C1_4alkyl, or C1_4alkoxy; PG is a carbamate protecting group.

Another aspect provides a process for preparing a compound of formula I: sing the steps of: preparing a compound of formula 4: HO—N R3 /fl\ r —|— PG from a compound of formula 3: R1—o R3 /fl\ I R2—O 'l— PG under suitable oxime formation conditions; wherein R1 is C1_6alkyl; R2 is C1_6alkyl; or R1 and R2, together with the oxygen atoms to which they are attached, form an optionally substituted 5 or 6 membered saturated heterocyclic ring haVing two oxygen atoms; R3 is hydrogen, C‘1_6alkyl, or a 3-6 membered ted or partially unsaturated heterocyclyl haVing 1-2 heteroatoms selected from the group consisting of , nitrogen, and sulfur; wherein the heterocyclyl is optionally substituted with l occurrence of halo or C1_3alkyl; (J2)q R4 is Q is phenyl, pyridyl, or an lated pyridine; J1 is H, halo, C1_4alkyl, or C1_4alkoxy; J2 is halo; CN; phenyl; oxazolyl; or a C1_6aliphatic group wherein up to 2 methylene units are optionally replaced with 0, NR”, C(O), S, 8(0), or S(O)2; said C1_6aliphatic group is ally substituted with 1—3 fluoro or CN; q is 0, l, or 2; PG is a carbamate protecting group.

Another embodiment further comprises the step of protecting a compound of formula 2: R1—o R3 R2_O>—Q/ ”N;H under suitable protection conditions to form the compound of formula 3.

Another embodiment r comprises the step of ng a compound of formula 1: R1—o R2-O ‘l— with a le amine under suitable reductive amination conditions to form a compound of formula 2.

In some embodiments, the suitableamine is NHCH3. In other embodiments, the suitable amine is OQNHZ.

Another embodiment further comprises the step of reacting a compound of formula 4: under suitable isoxazole formation ions to form a compound of formula 5: IU‘I r embodiment further comprises the step of reacting a compound of formula 5 under suitable coupling conditions followed by suitable deprotection conditions to form a compound of formula I.

In some embodiments, PG is Boc or Cbz. In some embodiments, PG is Boc.

In other embodiments, R1 is ethyl and R2 ethyl.

In yet other embodiments, R3 is CH3 or C .

.Ivvv (J2)q In some ments, R4 is ; wherein Q is phenyl. In some embodiments, Q is substituted in the para position with J2, wherein q is l.

In some embodiments, J1 is H or halo. In some embodiments, J1 is H. In other embodiments, J1 is halo.

In other embodiments, J2 is a C1_6aliphatic group wherein up to l methylene unit is optionally ed with S(O)2. In some embodiments, J2 is —S(O)2—(C1_5alkyl). In some embodiments, q is l.

According to another ment, R1 is ethyl; R2 is ethyl; R3 is CH3 or ECO; PG is Boc or Cbz; J1 is H; (J2)q R4 is wherein Q is phenyl; J2 is -S(O)2—CH(CH3)2; q is l; In some embodiments, R3 is CH3. In some ments, R3 is CH3. In yet another embodiments, R3 is CH3 or ECO.

According to another embodiment, R1 is ethyl; R2 is ethyl; E 0 PG is Boc; J1 is H; (J2)q Hfi<CH3$24 \\\\ R4 is wherein Q is pyridyl; J2 is N q is l; \ CH3 N CH3 . 4 In some embod1ments, R . is N Reactions Conditions In some embodiments, the le oxime formation conditions consist of either a single step sequence or a two step sequence.

In some embodiments, the two step sequence consists of first deprotecting the ketal group in the compound of formula 3 into an aldehyde under suitable deprotection conditions, and then forming the oxime of formula 4 under suitable oxime formation conditions. In some ments, suitable deprotection ions comprise adding catalytic amounts of para-toluenesulfonic acid , acetone, and water; and suitable oxime formation conditions comprise mixing together hydroxylamine, a catalytic amount of acid, a dehydrating agent, and an lic solvent. In other embodiments, the acid is pTSA or HCl, the dehydrating agent is molecular sieves or dimethoxyacetone, and the alcoholic t is methanol or ethanol.

In other embodiments, the single step sequence comprises adding NHZOHHCl and a mixture of THF and water. In other embodiments, the sequence comprises adding NHZOHHCl with a mixture of 2-methyl tetrahydrofuran and water optionally buffered with NaZSO4. In some embodiments, 1 equivalent of the compound of formula 3 is combined with a 1.1 equivalents ofNHZOHHCl in a 10:1 v/v mixture of THF and water. In some embodiments, 1 lent of the compound of formula 3 is combined with a 1.1 equivalents ofNHZOHHCl in a 10:1 v/v mixture of yl tetrahydrofuran and water optionally buffered with NaZSO4.

In other embodiments, the protection ions are selected from the group consisting of 0 R-OCOCl, a suitable tertiary amine base, and a suitable solvent; wherein R is kyl optionally substituted with phenyl; 0 R(C02)OR’, a suitable solvent, and optionally a catalytic amount of base, n R is and R’ are each independently kyl optionally substituted with ; 0 [RO(C=O)]ZO, a suitable base, and a suitable solvent.

In some embodiments, the suitable base is Et3N, ropylamine, and pyridine; and the suitable solvent is selected from a chlorinated solvent, an ether, or an aromatic hydrocarbon. In other embodiments, the suitable base is Et3N, the suitable t is a chlorinated solvent selected from DCM. In yet other embodiments, the protection conditions comprise adding 1.20 equivalents of (Boc)20 and 1.02 equivalents of Et3N in DCM.

According to another embodiment suitable ng conditions se adding a suitable metal and a suitable base in a le solvent. In other embodiments, the suitable metal is Pd[P(tBu)3]2; the suitable solvent is a mixture of itrile and water; and the suitable base is sodium carbonate. In yet other embodiments, the suitable ng conditions comprise adding 0.1 equivalents of Pd[P(tBu)3]2; 1 equivalent of boronic acid or ester; and 2 equivalents of sodium carbonate in a 2:1 ratio v/v of acetonitrile/water at 60—70°C.

According to another embodiment, suitable ection conditions comprise combining the compound of formula Q with a le acid in a suitable solvent. In some embodiments, the suitable acid is selected from para—toluenesulfonic acid (pTSA), HCl, TBAF, H3PO4, or TFA and the suitable solvent is selected from acetone, methanol, ethanol, CHzClz, EtOAc, THF, 2-MeTHF, dioxane, toluene, or diethylether.

According to another embodiment, suitable isoxazole—formation conditions consists of two steps, the first step comprising reacting the compound of formula 4 under suitable chlorooxime formation conditions to form a chlorooxime intermediate; the second step comprising reacting the chlorooxime intermediate with acetylene under suitable cycloaddition conditions to form a compound of formula 5.

According to another embodiment, suitable chlorooxime formation conditions are selected from . N—chlorosuccinimide and suitable solvent or ' potassium peroxymonosulfate, HCl, and dioxane.

In some embodiments, the suitable solvent is selected from a nonprotic solvent, an aromatic hydrocarbon, or an alkyl acetate. According to another embodiment, the suitable oxime formation conditions are 1.05 lents of N—chlorosuccinimide in isopropylacetate at 40—50°C.

According to another ment, suitable cycloaddition conditions consist of a suitable base and a suitable solvent. In some embodiments, the suitable base is selected from pyridine, DIEA, TEA, t—BuONa, and K2C03 and the suitable solvent is selected from acetonitrile, tetrahydrofuran, 2-methyltetrahydrofuran, MTBE, EtOAc, i-PrOAc, DCM, toluene, DMF, and methanol. In other embodiments, the le base is selected from Et3N and the le t is selected from DCM.

According to another ment, the second step comprises reacting 1 equivalent of acetylene with 1.2 equivalents of the oxime intermediate and 1.3 equivalents of Et3N in DCM at room temperature.

According to another embodiment, suitable isoxazole-formation conditions comprise combining the compound of a 4 with an oxidant in a suitable solvent. In som embodiments, said oxidant is [bis(trifluoroacetoxy)iodo] benzene and said solvent is a 1:1:1 mixture of ol, water, and dioxane.

Synthesis of Compounds I—2 and L3 One embodiment provides a process for preparing a compound of formula I-2: NH2 0"] HN\ SOZiPr comprising one or more of the following steps: a) Reacting a compound of formula lb: EtOé with amine under suitable reductive amination conditions to form a nd of formula 2b: b) reacting a compound of formula 2b under suitable Boc protection conditions to form the compound of formula 3b: EtO z—w8 c) reacting a compound of formula 3b under suitable oxime formation conditions to form the compound of formula 4-i: d) reacting a compound of formula 4-i under suitable chlorooxime formation conditions to form the compound of formula 4-ii: HON\ CIéBoc 4-ii e) reacting the nd of formula 4-ii with a compound of formula 4-iii Z A03O82 4-iii under suitable cycloaddition ions to form a compound of formula 4—iv: Boc / Boc N o—N \ ‘N\ KfN| 4-iv f) .reacting a compound of formula 4-iV with a compound of formula Ai: i-PrOZS A-5—i under suitable coupling conditions to form the compound of formula 5-i: Boc BOC N O’N \ \ N\ SOZiPr g) deprotecting a compound of formula 5—i under suitable Boc deprotection conditions optionally followed by ent under basic aqueous conditions to form a compound of formula 1-2.

Another embodiment provides a process for preparing a nd of formula 1-3: SOziPr comprising one or more of the following steps: a) Reacting a compound of formula A-l: EtO C 0 EC H with tetrahydro-2H—pyranamine under suitable reductive amination conditions to form a compound of formula A-2: EtO : HNCO b) reacting a compound of a A-2 under suitable Boc protection ions to form the compound of formula A—3: c) reacting a compound of formula A-3 under suitable oxime formation conditions to form the nd of formula A-4: d) reacting a compound of formula A-4: N‘Boc under suitable chlorooxime formation ions to form the compound of formula A-4—i: HO\ O cquN‘BocN A-4—i e) reacting the compound of formula Ai with a nd of formula : N(Boc)2 Aii under suitable cycloaddition conditions to form the compound of formula A—S: BOCN\/ O’N\ Br Q f) reacting a compound of formula A-5 with a compound of formula Ai: i-PrOZS A-5—i under suitable coupling conditions to form the compound of formula A-6: SOZiPr g) deprotecting a compound of formula A—6 under suitable Boc deprotection conditions optionally followed by treatment under basic aqueous conditions to form a compound of formula 1-3. le coupling conditions comprise combining a suitable palladium catalyst with a suitable base in a suitable solvent. Suitable palladium catalyst include, but are not limited to, Pd[P(tBu)3]2, pf)Clz, Pd(PPh3)2Clz, 3)2Clz and , Pd(dppf)Clz, Pd(dppe)Clz. Suitable solvents include, but are not limited to. toluene, MeCN, water, EtOH, IPA, 2—Me—THF, or IPAc. Suitable bases include, but are not limited to, K2CO3, NazCO3, or K3PO4.

Suitable oxime formation conditions consist of either a single step sequence or a two step sequence. The two step sequence consists of first deprotecting the ketal group in the compound of formula A-3 into an aldehyde under suitable deprotection conditions, and then forming the oxime of a A-4 under suitable oxime formation conditions.

The single step sequence comprises, for example, comprise mixing together hydroxylamine, an acid, an c solvent, and water. In some embodiments, NHzOHHCl is added to a mixture of THF and water. In some embodiments, 1 lent of the compound of a 3-A is combined with a 1.1 equivalents ofNHZOHHCl in a 10:1 v/v mixture of THF/water. le ection conditions comprise adding an acid, acetone, and water.

Suitable acids include pTSA or HCl, tsuitable organic solvents include chlorinated solvents (e. g., dichloromethane (DCM), roethane (DCE), CHzClz, and chloroform); an ether (e.g., THF, 2-MeTHF and dioxane); an aromatic hydrocarbons (e. g., toluene and xylenes, or other aprotic solvents.

Suitable cycloaddition conditions comprise a suitable base (e. g., pyridine, DIEA, TEA, t-BuONa, or K2C03) and a suitable solvent (e.g., acetonitrile, tetrahydrofuran, 2—methyltetrahydrofuran, MTBE, EtOAc, c, DCM, toluene, DMF, and methanol_.

Suitable chlorooxime formation conditions comprise adding HCl in e to a solution of the oxime in the presence ofNCS in a suitable solvent selected from a nonprotic solvents (DCM, DCE, THF, and dioxane), ic hydrocarbons (e. g. toluene, xylenes), and alkyl acetates (e. g., isopropyl acetate, ethyl acetate).

Suitable Boc deprotection conditions comprises adding a le Boc deprotecting agent (e. g, TMS-Cl, HCl, TBAF, H3PO4, or TFA) and a le solvent (e. g., acetone, toluene, ol, ethanol, l-propanol, panol, CHzClz, EtOAc, isopropyl acetate, tetrahydrofuran, yltetraydrofuran, dioxane, and diethylether). In some embodiments, the suitable Boc deprotection conditions comprises adding a suitable Boc deprotecting agent selected from HCl, TFA and a suitable solvent selected from acetone, toluene, isopropyl acetate, tetrahydrofuran, or 2—methyltetraydrofuran.

Suitable Boc protection conditions include (Boc)20, a suitable base, and a suitable solvent. Suitable bases include, but are not limited to, Et3N, diisopropylamine, and pyridine. Suitable solvents include, but are not limited to, chlorinated solvents (e. g., dichloromethane (DCM), dichloroethane (DCE), CHzClz, and form); an ether (e.g., THF, 2—MeTHF and dioxane); an aromatic hydrocarbons (e.g., toluene and xylenes, or other aprotic solvents. In some ments, the suitable base is Et3N, the le solvent is DCM, tetrahydrofuran or 2—methyltetrahydrofuran. In certain embodiments, the protection conditions comprise adding 1.05 equivalents of (Boc)20 in yltetrahydrofuran or DCM.

Suitable reductive amination conditions comprise adding a reducing agent selected from NaBH4 NaBH4, NaBH3CN, or NaBH(OAc)3 in the presence of a solvent selected from dichloromethane (DCM), dichloroethane (DCE), an alcoholic solvent selected from methanol, ethanol, l-propanol, panol, or a nonprotic solvent selected from dioxane, tetrahydrofuran, or 2-methyltetrahydrofuran and optionally a base selected from Et3N or diisopropylethylamine. In some embodiments, the suitable reductive amination conditions comprise adding 1.2 equivalents ofNaBH4 s in the presence Et3N in MeOH. r aspect of the present invention provides a compound of Formula II: R4 R3a o\ R3b O// R30 R1 a R1 0 R1 b or a pharmaceutically acceptable salt thereof, wherein each Rla, Rlb, Rlc, R2, R3a, R3b, R3: R4, R5, R6, R7, R8, R9a, R9b, RIOa, R10b, and R10c is independently hydrogen or deuterium, and at least one of Rla, Rlb, Rlc, R2, R3a, R3b, R36, R4, R5, R6, R7, R8, R9a, R9b, RIOa, Rlob, and R10c is deuterium.

In some embodiments, R9a and R9b are the same. In other embodiments, R9a and R9b are deuterium, and R13, Rlb,R1c, R2, R33, R3b, R32 R4, R5, R6, R7, R8, R10, R113, Rllb, R123, Rlzb, Rm, R13b, R14a, and R14b are deuterium or hydrogen. In yet another embodiment, R9a and R9b are deuterium, and R13, Rlb, R16, R2, R33, R3b, R36, R4, R5, R6, R7, R8, R10, R113, Rllb, R123, Rlzb, R133, R1”, R143, and R14b are hydrogen.

In one embodiment, Rga, Rgb, Rloa, Rlob, and R10c are the same. In another ment, Rga, Rgb, R103, Rlob, and R10c are deuterium, and R”, Rlb, R16, R2, R33, R3b, R3c, R4, R5, R6, R7, and R8 are ium or hydrogen. In some embodiments, R93, Rgb, R103, Rlob, and R10c are ium, and R”, Rlb, R16, R2, R3a, R3b, R36, R4, R5, R6, R7, and R8 are hydrogen.

In other embodiments, Rloa, Rlob, and R10c are the same. In one embodiment, R103, Rlob, and Rmc are deuterium, and R13, Rlb, R16, R2, R33, R3b, R36, R4, R5, R6, R7, R8, R93, and R9b are deuterium or hydrogen. In yet another embodiment, Rloa, Rlob, and R10c are deuterium, and R”, Rlb, R16, R2, R3a, R3b, R36, R4, R5, R6, R7, R8, Rga, and R9b are hydrogen.

In some ments, R13, Rlb, R16, R2, R33, R3b, and R3c are the same. In another embodiment Rla, Rlb, R”, R2, R3a, R3b, and R30 are deuterium, and R4, R5, R6, R7, R8, Rga, Rgb, Rloa, Rlob, and R10c are deuterium or hydrogen. In yet another embodiment, Rla, Rlb, R”, R2, R33, R3b, and R3c are deuterium, and R4, R5, R6, R7, R8, R93, R9b, R103, Rlob, and Rmc are deuterium.

In another embodiment, R6 is deuterium, and R”, Rlb, R16, R2, R33, R3b, R36, R4, R5, R7, R8, Rga, Rgb, R103, Rlob, and R10c are deuterium or hydrogen. In yet another embodiment, R6 is deuterium, and R1111”: R”, R2, R33, R3b, R32 R4, R5, R7, R8, R93, R9b, Rloa, Rlob, and R10c are hydrogen.

In other embodiments, R2 is deuterium, and R”, Rlb, Rlc, R3a, R3b, and R3c, R4, R5, R6, R7, R8, Rga, Rgb, Rloa, Rlob, and R10c are deuterium or hydrogen. In another ment, R2 is ium, and R13, Rlb, R”, R33, R3b, and R36, R4, R5, R6, R7, R8, R93, R9b, Rloa, Rlob, and R10c are hydrogen.

In another embodiment, R7 is deuterium, and R13, Rlb, R16, R2, R3a, R3b, R36, R4, R5, R6, R8, Rga, Rgb, Rloa, Rlob, and R10c are deuterium or hydrogen. In other embodiments, R7 is deuterium, and Rla, Rlb, Rlc, R2, R3a, R3b, R32 R4, R5, R6, R8, R9a, R9b, RlOa, R10b, R10c are hydrogen.

In yet another embodiment, R8 is deuterium, and R”, Rlb, R”, R2, R3a, R3b, R3c, R4, R5, R6, R7, Rga, Rgb, Rloa, Rlob, and R10c are deuterium or en. In another embodiment, R8 is deuterium, and R1111”: R”, R2, R33, R3b, R32 R4, R5, R6, R7, R93, R9b, Rloa, Rlob, R10c are hydrogen.

In some embodiments, at least one of Rloa, Rlob, or R10c are the same. In r embodiment, at least one of R103, Rlob, or R10c are deuterium, and R”, Rlb, R16, R2, R3a, R3b, R36, R4, R5, R6, R7, R8, Rga, and R9b are deuterium or hydrogen. In yet another embodiment, at least one of Rloa, Rlob, or R10c are ium, and R”, Rlb, R”, R2, R3a, R3b, R36, R4, R5, R6, R7, R8, R93, and R9b are hydrogen.

In some embodiments, at least two of Rloa, Rlob, or R10c are the same. In another ment, at least two of R103, Rlob, or R10c are deuterium, and R”, Rlb, R”, R2, R3a, R3b, R36, R4, R5, R6, R7, R8, Rga, and R9b are deuterium or hydrogen. In yet another embodiment, at least two of R103, Rlob, or R10c are deuterium, and R”, Rlb, R16, R2, R3a, R3b, R36, R4, R5, R6, R7, R8, R93, and R9b are hydrogen.

In another embodiment, Rla, Rlb, R16, R3a, R3b, and R30 are the same. In some embodiments, Rla, Rlb, R16, R3a, R3b, and R30 are deuterium, and R2, R4, R5, R6, R7, R8, Rga, Rgb, Rloa, Rlob, and R10c are deuterium or hydrogen. In yet another embodiment, Rla, Rlb, Rlc, R3a, R3b, and R30 is ium, and R2, R4, R5, R6, R7, R8, Rga, Rgb, R103, Rlob, and R10c are hydrogen.

In yet another embodiment, R4 is deuterium, and R”, Rlb, R”, R2, R3a, R3b, R3c, R5, R6, R7, R8, Rga, Rgb, Rloa, Rlob, and R10c are deuterium or hydrogen. In other embodiments, R4 is deuterium, and R13, Rlb, R”, R2, R33, R3b, R32 R5, R6, R7, R8, R93, R9b, Rloa, Rlob, and R10c are hydrogen.

In another embodiment, R5 is deuterium, and R”, Rlb, R16, R2, R33, R3b, R36, R4, R6, R7, R8, Rga, Rgb, Rloa, Rlob, and R10c are deuterium or hydrogen. In yet another embodiment, R5 is deuterium, and R13, Rlb, R”, R2, R33, R3b, R36, R4, R6, R7, R8, R93, R9b, Rloa, Rlob, and R10c are hydrogen.

In another embodiment, at least one of R9a or R9b are the same. In other embodiments, at least one of R9a or R9b are deuterium, and R13, Rlb, R”, R2, R3a, R3b, R36, R4, R5, R6, R7, R8, R103, Rlob, and R10c are ium or en. In some embodiments, at least one of R9a and R9b are deuterium, and R13, Rlb, R”, R2, R33, R3b, R36, R4, R5, R6, R7, R8, R103, Rlob, R10c are en.

In one embodiment, R6, R9a and R9b are the same. In some embodiments, R6, R9a and R9b are deuterium, and R13, Rlb, R”, R2, R33, R3b, R36, R4, R5, R7, R8, R103, Rlob, Rmc are deuterium or hydrogen. In other embodiments, R6, R9a and R9b are deuterium, and R”, Rlb, R”, R2, R33, R3b, R32 R4, R5, R7, R8, R103, Rlob, and Rmc are hydrogen.

In some embodiments, R2, Rloa, Rlob, and R10c are the same. In another embodiment, R2, R103, Rlob, and R10c are ium, and R13, Rlb, Rlc, R3a, R3b, R3c, R4, R5, R6, R7, R8, Rga, and R9b are deuterium or en. In yet another embodiment, R2, R103, Rlob, and Rmc are deuterium, and R13, Rlb, R”, R33, R3b, R36, R4, R5, R6, R7, R8, R93, and R9b are hydrogen.

In some embodiments, R7 and at least two of Rloa, Rlob, or R10c are the same. In another embodiment, R7 and at least two of Rloa, Rlob, or R10c are deuterium, and R”, Rlb, R16, R2, R3a, R3b, R36, R4, R5, R6, R8, Rga, and R9b are deuterium or hydrogen. In yet another embodiment, R7 and at least two of R103, Rlob, or R10c are ium, and R”, Rlb, R”, R2, R33, R3b, R32 R4, R5, R6, R8, R93, and R9b are hydrogen.

In some embodiments, Rla, Rlb, R”, R2, R3a, R3b, R3c, and at least one of Rloa, Rlob, or R10c are the same. In another embodiment, Rla, Rlb, R16, R2, R3a, R3b, R36, and at least one of Rloa, Rlob, or R10c are deuterium, and R4, R5, R6, R7, R8, Rga, and R9b are deuterium or hydrogen. In yet another embodiment, Rla, Rlb, R”, R2, R3a, R3b, R3c, and at least one of Rloa, Rlob, or R10c are deuterium, and R4, R5, R6, R7, R8, Rga, and R9b are en.

In some embodiments, Rla, Rlb, R16, R3a, R3b, R36, and R5 are the same. In another embodiment, Rla, Rlb, Rlc, R3a, R3b, R3c, and R5 are deuterium, and R2, R4, R6, R7, R8, Rga, Rgb, Rloa, Rlob, and R10c are deuterium or en. In yet r embodiment, Rla, Rlb, R”, R33, R3b, R36, and R5 are deuterium, and R2, R4, R6, R7, R8, R93, R9b, R103, Rlob, and Rmc are hydrogen.

In other embodiments, R4 and R6 are the same. In another embodiment, R4 and R6 are deuterium, and R13, Rlb, R”, R2, R33, R3b, R32 R5, R7, R8, R93, R9b, R103, Rlob, and Rmc are deuterium or hydrogen. In yet another embodiment, R4 and R6 are deuterium, and R”, Rlb, Rlc, R2, R3a, R3b, R36, R5, R7, R8, R9a, R9b, RlOa, R10b, and RlOc are hydrogen.

In one embodiment, R2, R5, Rga, and R9b are the same. In some embodiments, R2, R5, R93, and R9b are deuterium, and R13, Rlb, R”, R33, R3b, R36, R4, R6, R7, R8, R103, Rlob, and R10c are deuterium or hydrogen. In another embodiment, R2, R5, Rga, and R9b are deuterium, and R”, Rlb, R16, R3a, R3b, R36, R4, R6, R7, R8, R103, Rlob, and R10c are hydrogen.

In yet another embodiment, Rla, Rlb, R16, R2, R3a, R3b, R36, R5, R6, Rga, Rgb, Rloa, Rlob, and R10c are the same. In some embodiments, Rla, Rlb, R16, R2, R3a, R3b, R3 , R5, R6, Rga, Rgb, Rloa, Rlob, and R10c are deuterium, and R4, R7, and R8 are deuterium or hydrogen. In other embodiments, R13, Rlb, R16, R2, R33, R3b, R3 and Rmc is , R5, R6, R93, R9b, R103, Rlob, deuterium, and R4, R7, and R8 are en.

In some embodiments, the variables are as depicted in the nds of the disclosure including compounds in the tables below.

TableI NH2 0,N , \ HNCO NH2 0 N\ HN\ \ \ N \ \ | | /N /N ll O=S=O N 4\ 1—1 1—2 WO 49726 SOziPr Table II NWDNH20’N\ HN‘ —N D NHZ 0:1 NH2 o\\ HN+D ' D N \ D \ HN‘gD /N D | D | /N /N 11—1 11—2 11—3 NH o—N NI \ \ /N NW I NI \ /N D /N 02820 WD 8:3 D D D D Y 83fD 11—4 11—5 11—6 NH2 0, "i HN‘ NH2 0 Q, HN‘ NH2 04‘: \ HN\< \ \ N \ N \ N \ D I I I /N D /N D /N O\\ O¢ 0931/ 0437/ 048Q 11—7 11—8 11—9 WO 49726 11—12 NH2 04‘: HN‘ I D /N D OGSY 11—15 NH2 04‘: HNJD Q‘s D 0’ $0 D D 11—18 NH2 04‘: HN‘ | D 11-20 11—21 11-22 Compounds of this invention include those described generally , and are further illustrated by the classes, subclasses, and species sed herein. As used herein, the following definitions shall apply unless ise indicated. For purposes of this invention, the al elements are identified in accordance with the Periodic Table of the Elements, CAS version, Handbook of Chemistry and Physics, 75th Ed. Additionally, general principles of organic try are described in “Organic Chemistry”, Thomas Sorrell, University Science Books, ito: 1999, and “March’s Advanced Organic Chemistry”, 5th Ed., Ed.: Smith, MB. and March, J., John Wiley & Sons, New York: 2001, the entire contents of which are hereby incorporated by reference.

As described herein, a specified number range of atoms includes any integer therein. For example, a group having from l-4 atoms could have 1, 2, 3, or 4 atoms.

As described , compounds of the invention may optionally be substituted with one or more substituents, such as are illustrated lly herein, or as exemplified by particular classes, subclasses, and species of the invention. It will be appreciated that the phrase “optionally substituted” is used interchangeably with the phrase “substituted or unsubstituted.” In general, the term “substituted”, whether preceded by the term “optionally” or not, refers to the replacement of en radicals in a given structure with the radical of a specified substituent. Unless otherwise indicated, an optionally substituted group may have a substituent at each substitutable position of the group, and when more than one position in any given structure may be substituted with more than one substituent ed from a ied group, the substituent may be either the same or different at every position.

Combinations of substituents envisioned by this invention are preferably those that result in the formation of stable or chemically feasible compounds.

Unless ise indicated, a substituent connected by a bond drawn from the center of a ring means that the substituent can be bonded to any position in the ring. In example i below, for instance, I1 can be bonded to any position on the pyridyl ring. For bicyclic rings, a bond drawn through both rings indicates that the tuent can be bonded from any position of the bicyclic ring. In example ii below, for ce, I1 can be bonded to the 5-membered ring (on the nitrogen atom, for instance), and to the 6-membered ring. (9—065/ E PM / '_NA “”05 N H 2012/058127 The term “stable”, as used herein, refers to compounds that are not substantially altered when subjected to conditions to allow for their production, detection, recovery, purification, and use for one or more of the purposes sed herein. In some embodiments, a stable compound or chemically feasible compound is one that is not substantially altered when kept at a ature of 40°C or less, in the absence of re or other chemically reactive conditions, for at least a week.

The term “aliphatic” or “aliphatic group”, as used herein, means a straight-chain (i.e., unbranched), branched, or cyclic, substituted or unsubstituted hydrocarbon chain that is completely saturated or that contains one or more units of unsaturation that has a single point of attachment to the rest of the le.

Unless otherwise specified, aliphatic groups contain 1—20 aliphatic carbon atoms.

In some embodiments, aliphatic groups contain l-lO aliphatic carbon atoms. In other embodiments, aliphatic groups contain 1-8 aliphatic carbon atoms. In still other embodiments, aliphatic groups n 1-6 aliphatic carbon atoms, and in yet other embodiments aliphatic groups contain 1-4 aliphatic carbon atoms. Aliphatic groups may be linear or ed, substituted or unsubstituted alkyl, alkenyl, or alkynyl groups. Specific examples include, but are not limited to, methyl, ethyl, isopropyl, n-propyl, sec-butyl, vinyl, n-butenyl, ethynyl, and tert-butyl. Aliphatic groups may also be cyclic, or have a combination of linear or branched and cyclic groups. Examples of such types of aliphatic groups include, but are not limited to cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cyclohexenyl, —CH2— cyclopropyl, CHzCHzCH(CH3)—cyclohexyl.

The term “cycloaliphatic” (or cycle” or “carbocyclyl”) refers to a monocyclic C3—C8 hydrocarbon or bicyclic C8-C12 hydrocarbon that is completely saturated or that contains one or more units of unsaturation, but which is not aromatic, that has a single point of attachment to the rest of the molecule n any individual ring in said bicyclic ring system has 3-7 members. Examples of cycloaliphatic groups include, but are not limited to, lkyl and cycloalkenyl groups. Specific es include, but are not limited to, cyclohexyl, ropenyl, and cyclobutyl.

The term “heterocycle”, “heterocyclyl”, or “heterocyclic” as used herein means non-aromatic, monocyclic, bicyclic, or tricyclic ring systems in which one or more ring members are an independently selected heteroatom. In some embodiments, the “heterocycle”, “heterocyclyl”, or “heterocyclic” group has three to en ring members in which one or more ring members is a heteroatom independently selected from , sulfur, nitrogen, or orus, and each ring in the system ns 3 to 7 ring members.

WO 49726 Examples of heterocycles include, but are not limited to, 3—lH-benzimidazol—2— one, 3-(l-alkyl)-benzimidazolone, 2-tetrahydrofuranyl, 3-tetrahydrofuranyl, 2- tetrahydrothiophenyl, 3-tetrahydrothiophenyl, holino, 3-morpholino, holino, 2- thiomorpholino, morpholino, morpholino, l-pyrrolidinyl, 2-pyrrolidinyl, 3- pyrrolidinyl, l-tetrahydropiperazinyl, 2-tetrahydropiperazinyl, 3-tetrahydropiperazinyl, l- piperidinyl, 2-piperidinyl, 3-piperidinyl, l-pyrazolinyl, 3-pyrazolinyl, 4-pyrazolinyl, 5- pyrazolinyl, l-piperidinyl, 2-piperidinyl, 3-piperidinyl, ridinyl, zolidinyl, 3- thiazolidinyl, 4-thiazolidinyl, l-imidazolidinyl, 2-imidazolidinyl, 4-imidazolidinyl, 5- imidazolidinyl, indolinyl, tetrahydroquinolinyl, ydroisoquinolinyl, benzothiolane, benzodithiane, and l,3-dihydro-imidazol-2—one.

Cyclic groups, (e.g. cycloaliphatic and heterocycles), can be linearly fused, bridged, or spirocyclic.

The term “heteroatom” means one or more of oxygen, sulfur, nitrogen, phosphorus, or silicon (including, any oxidized form of nitrogen, sulfur, phosphorus, or silicon; the quaternized form of any basic nitrogen or; a substitutable nitrogen of a heterocyclic ring, for example N (as in 3,4-dihydro-2H—pyrrolyl), NH (as in pyrrolidinyl) or NR+ (as in N—substituted pyrrolidinyl)).

The term "unsaturated", as used herein, means that a moiety has one or more units of unsaturation. As would be known by one of skill in the art, unsaturated groups can be partially unsaturated or fully unsaturated. Examples of partially unsaturated groups include, but are not limited to, butene, cyclohexene, and tetrahydropyridine. Fully unsaturated groups can be aromatic, anti-aromatic, or non-aromatic. Examples of fully rated groups include, but are not limited to, phenyl, cyclooctatetraene, pyridyl, thienyl, and l- methylpyridin-2(lH)-one.

The term “alkoxy”, or “thioalkyl”, as used herein, refers to an alkyl group, as preViously defined, attached through an oxygen (“alkoxy”) or sulfur (“thioalkyl”) atom.

The terms “haloalkyl”, “haloalkenyl”, liphatic”, and “haloalkoxy” mean alkyl, alkenyl or alkoxy, as the case may be, substituted with one or more halogen atoms.

This term includes rinated alkyl groups, such as —CF3 and —CF2CF3.

The terms “halogen”, , and “hal” mean F, Cl, Br, or I.

The term “aryl” used alone or as part of a larger moiety as in “aralkyl”, “aralkoxy”, or “aryloxyalkyl”, refers to monocyclic, ic, and tricyclic ring systems haVing a total of five to fourteen ring members, wherein at least one ring in the system is WO 49726 aromatic and wherein each ring in the system contains 3 to 7 ring members. The term “aryl” may be used interchangeably with the term “aryl ring”.

The term “heteroaryl”, used alone or as part of a larger moiety as in “heteroaralkyl” or “heteroarylalkoxy”, refers to monocyclic, bicyclic, and tricyclic ring s having a total of five to fourteen ring s, wherein at least one ring in the system is ic, at least one ring in the system contains one or more heteroatoms, and wherein each ring in the system contains 3 to 7 ring members. The term “heteroaryl” may be used interchangeably with the term “heteroaryl ring” or the term “heteroaromatic”. Examples of heteroaryl rings include, but are not limited to, 2-furanyl, 3-furanyl, N—imidazolyl, 2- imidazolyl, 4-imidazolyl, 5-imidazolyl, benzimidazolyl, 3-isoxazolyl, 4-isoxazolyl, 5- isoxazolyl, 2-oxazolyl, 4-oxazolyl, olyl, N—pyrrolyl, 2-pyrrolyl, 3-pyrrolyl, 2-pyridyl, 3-pyridyl, 4-pyridyl, 2-pyrimidinyl, midinyl, midinyl, pyridazinyl (e. g., 3- pyridazinyl), 2-thiazolyl, 4-thiazolyl, 5-thiazolyl, tetrazolyl (e. g., 5-tetrazolyl), triazolyl (e. g., 2-triazolyl and 5-triazolyl), 2-thienyl, 3-thienyl, benzofuryl, benzothiophenyl, indolyl (e. g., 2— indolyl), pyrazolyl (e.g., 2-pyrazolyl), isothiazolyl, 1,2,3-oxadiazolyl, 1,2,5-oxadiazolyl, 1,2,4-oxadiazolyl, 1,2,3-triazolyl, 1,2,3-thiadiazolyl, 1,3,4-thiadiazolyl, 1,2,5-thiadiazolyl, l, pyrazinyl, 1,3,5-triazinyl, quinolinyl (e.g., 2-quinolinyl, 3-quinolinyl, 4-quinolinyl), and isoquinolinyl (e.g., uinolinyl, 3-isoquinolinyl, or 4-isoquinolinyl).

It shall be understood that the term “heteroaryl” includes certain types of heteroaryl rings that exist in equilibrium between two different forms. More specifically, for example, s such hydropyridine and pyridinone (and likewise hydroxypyrimidine and pyrimidinone) are meant to be encompassed within the definition of “heteroaryl.” |\ |\ /N‘_ NH OH 0 The term “protecting group” and “protective group” as used herein, are interchangeable and refer to an agent used to temporarily block one or more desired functional groups in a nd with le reactive sites. In certain embodiments, a protecting group has one or more, or preferably all, of the following characteristics: a) is added selectively to a functional group in good yield to give a protected ate that is b) stable to reactions occurring at one or more of the other reactive sites; and c) is selectively removable in good yield by reagents that do not attack the regenerated, deprotected functional group. As would be tood by one skilled in the art, in some cases, the reagents do not WO 49726 attack other reactive groups in the nd. In other cases, the reagents may also react with other reactive groups in the compound. Examples of ting groups are detailed in Greene, T.W., Wuts, P. G in “Protective Groups in Organic Synthesis”, Third Edition, John Wiley & Sons, New York: 1999 (and other editions of the book), the entire contents of which are hereby incorporated by reference. The term “nitrogen protecting group”, as used herein, refers to an agent used to temporarily block one or more desired nitrogen reactive sites in a multifunctional compound. red nitrogen protecting groups also possess the characteristics exemplified for a protecting group above, and certain exemplary nitrogen protecting groups are also detailed in Chapter 7 in Greene, T.W., Wuts, P. G in “Protective Groups in Organic Synthesis”, Third Edition, John Wiley & Sons, New York: 1999, the entire contents of which are hereby incorporated by reference.

In some embodiments, a methylene unit of an alkyl or aliphatic chain is optionally replaced with r atom or group. Examples of such atoms or groups include, but are not limited to, nitrogen, oxygen, , —C(O)—, —C(=N—CN)—, )—, —C(=NOR)—, —SO—, and —SOz—. These atoms or groups can be combined to form larger groups. Examples of such larger groups include, but are not limited to, —OC(O)—, —C(O)CO—, —COz—, —C(O)NR—, —C(=N— CN), —NRCO—, —NRC(O)O—, —SOzNR—, —NRSOz—, —NRC(O)NR—, —OC(O)NR—, and -NRSOZNR-, wherein R is, for example, H or C1_6aliphatic. It should be understood that these groups can be bonded to the methylene units of the aliphatic chain via single, double, or triple bonds. An example of an optional replacement (nitrogen atom in this case) that is bonded to the aliphatic chain via a double bond would be =N—CH3. In some cases, especially on the terminal end, an optional replacement can be bonded to the aliphatic group via a triple bond. One example of this would be CHZCHZCHZCEN. It should be understood that in this situation, the terminal nitrogen is not bonded to another atom.

It should also be tood that, the term “methylene unit” can also refer to branched or substituted ene units. For example, in an isopropyl moiety [-CH(CH3)2], a nitrogen atom (e.g. NR) replacing the first recited “methylene unit” would result in dimethylamine [-N(CH3)2]. In instances such as these, one of skill in the art would understand that the nitrogen atom will not have any additional atoms bonded to it, and the “R” from “NR” would be absent in this case.

Unless otherwise indicated, the optional replacements form a chemically stable compound. al replacements can occur both within the chain and/or at either end of the chain; i.e. both at the point of attachment and/or also at the terminal end. Two al ements can also be nt to each other within a chain so long as it results in a ally stable compound. For example, a C3 aliphatic can be optionally replaced by 2 nitrogen atoms to form —C—NEN. The optional replacements can also tely replace all of the carbon atoms in a chain. For example, a C3 aliphatic can be optionally replaced by —NR—, —C(O)—, and —NR— to form —NRC(O)NR— (a urea).

] Unless otherwise indicated, if the replacement occurs at the al end, the replacement atom is bound to a hydrogen atom on the terminal end. For example, if a methylene unit of -CH2CH2CH3 were optionally replaced with -O-, the resulting compound could be —OCH2CH3, —CHZOCH3, or ZOH. It should be understood that if the terminal atom does not contain any free valence electrons, then a hydrogen atom is not required at the terminal end (e. g., —CH2CH2CH=O or 2CEN).

Unless ise indicated, structures depicted herein are also meant to include all isomeric (e. g., enantiomeric, diastereomeric, geometric, conformational, and rotational) forms of the structure. For example, the R and S configurations for each asymmetric , (Z) and (E) double bond isomers, and (Z) and (E) conformational isomers are included in this invention. As would be understood to one skilled in the art, a substituent can freely rotate around any rotatable bonds. \ For example, a substituent drawn as also represents Therefore, single stereochemical isomers as well as enantiomeric, diastereomeric, geometric, conformational, and rotational mixtures of the present compounds are within the scope of the invention.

Unless otherwise indicated, all tautomeric forms of the nds of the invention are within the scope of the invention.

In the compounds of this invention any atom not specifically designated as a particular e is meant to represent any stable isotope of that atom. Unless otherwise stated, when a position is designated specifically as "H" or "hydrogen", the position is understood to have hydrogen at its natural nce isotopic composition. Also unless otherwise stated, when a position is designated specifically as "D" or "deuterium", the position is understood to have deuterium at an abundance that is at least 3340 times r than the natural abundance of deuterium, which is 0.015% (i.e., at least 50.1% incorporation of deuterium).

"D" and "d" both refer to deuterium.

] Additionally, unless ise indicated, structures depicted herein are also meant to include nds that differ only in the presence of one or more isotopically enriched atoms. For example, compounds having the present structures except for the replacement of hydrogen by deuterium or tritium, or the replacement of a carbon by a 13C— or 14C—enriched carbon are within the scope of this invention. Such nds are useful, for example, as analytical tools or probes in ical assays.

Processes Processes and compounds described herein are useful for producing ATR inhibitors that contain an aminopyrazine—isoxazole core. The general synthetic procedures shown in schemes herein are useful for ting a wide array of chemical species which can be used in the manufacture of pharmaceutical compounds.

SCHEME A R1-O>_<__\>/%O R1_ Step1 Step2 Rl-O N,R3 —> —> RZ'O __l O>_</:\>/\NIR3H R2'0 __ Reductive | Protection R2-O PG J1 amination J1 J1 l 2 § , 33 N O’N .

Step 3 / \/\'il HO—N 3 Step 5 W,R Step 4 \ N PG —> \ /| ‘ — 9 —> N \ _> Oxime ' PG I Isoxazole J1 / N Suzuki formation J1 formation (when R4 is Br) 4 (1 or 2 steps) R4 Deprotection NH ON 3 NH O’N 3 2 2 NH / X\N'R \\ / X\N'R 2 ON \\ Step6 Step7 \\ N’R3 N| \ H —> H —> N \ H Y NI \ I N 1:) free base N 1:) salt formation N J1 formation Rf J1 S? ' acid J1 R4 R4 R4 1 LA 1-3 $124 The compound of formula I can be made according to the steps outlined in Scheme A. Step 1 depicts the use of a readily available aldehyde/ketal as a starting point for the preparation of compounds of formula I, I-A, and LB. Reductive amination between compound 1 and a suitable primary amine, under conditions known to those d in the art leads to compound 2 where a benzylamine motif has been installed. For example, imines can be formed by ing an amine and an aldehyde in a suitable solvent, such as dichloromethane (DCM), dichloroethane (DCE), an alcoholic solvent (e. g., ol, ethanol), or a nonprotic solvent (e. g., dioxane or tetrahydrofuran (THF)). These imines can then be reduced by known reducing agents including, but not limited to, NaBH4, N, and NaBH(OAc)3 (@ JOC 1996, 3849). In some embodiments, 1.05 lents of amine is combined with 1 equivalent of aldehyde in methanol. In other embodiments, 1.2 equivalents of amine is combined with 1 equivalent of aldehyde in methanol. This step is then ed by reduction with 0.6 to 1.4 (such as 1.2) lents 4. In some cases, if an amine salt is used, base (e. g., Et3N or diisopropylethylamine) can also be added.

Step 2 depicts the protection of the benzylamine 1 prepared above, using a carbamate—based protecting group, under suitable protection conditions known to those skilled in the art. Various ting groups, such as Cbz and Boc, can be used. tion conditions include, but are not d to the following: a) R-OCOCl, a suitable tertiary amine base, and a suitable solvent; wherein R is C1_6alkyl optionally substituted with phenyl; b) R(COz)OR’, a suitable solvent, and optionally a catalytic amount of base, wherein R is and R’ are each independently C1_6alkyl optionally substituted with phenyl; c) [RO(C=O)]ZO, a suitable base, and a suitable solvent.

Examples of suitable bases include, but are not limited to, Et3N, diisopropylamine, and pyridine. Examples of suitable solvents include chlorinated ts (e. g., dichloromethane (DCM), dichloroethane (DCE), CHzClz, and chloroform), ethers (e. g., THF and dioxane), aromatic hydrocarbons (e.g., e, xylenes) and other , 2-MeTHF, aprotic solvents.

In some ments, tion can be done by reacting the benzylamine with (Boc)20 and Et3N in DCM. In some embodiments, 1.02 equivalents of (Boc)20 and 1.02 equivalents of Et3N 1.02 are used. . In another embodiment, protection can be done by reacting the benzylamine with (Boc)20 in 2-MeTHF. In some embodiments, 1.05 equivalents of (Boc)20 are used.

Step 3 Step 3 shows how the ketal functional group in 3 is then converted into the oxime 4 in a single step. This direct conversion from ketal to oxime is not extensively described in the literature and it will be appreciated that this step could also be conducted in a two—step sequence, transiting through the aldehyde after ection of the ketal using methodologies known to those skilled in the art.

Oxime formation conditions comprise mixing together hydroxylamine, acid, optionally a dehydrating agent, and an alcoholic solvent. In some embodiments, the acid is a catalytic amount. In some embodiments, the acid is pTSA or HCl, the dehydrating agent is molecular sieves or dimethoxyacetone, and the alcoholic solvent is methanol or ethanol. In some embodiments, the hydroxylamine hydrochloride is used in which case no additional acid is required. In other embodiments, the desired t is isolated via a biphasic work up and optionally precipitation or crystallization. If a ic work up is used, a ating agent is not needed.

In another embodiment, the oxime formation conditions comprise of mixing together hydroxylamine, an acid, an organic solvent and water. es of suitable organic solvents e chlorinated solvents (e. g., dichloromethane (DCM), dichloroethane (DCE), CHzClz, and chloroform), ethers (e. g., THF, 2-MeTHF and dioxane), aromatic hydrocarbons (e.g., toluene, xylenes) and other aprotic solvents. In some embodiments, 1.5 equivalents of hydroxylamine hydrochloride are used, the c solvent is 2-MeTHF and the water is ed with NaZSO4. In another embodiment, 1.2 lents of hydroxylamine hloride are used, the organic solvent is THF.

In some embodiments, suitable ection conditions comprise adding tic amounts of para-toluenesulfonic acid (pTSA), acetone, and water; and then forming the oxime using conditions known to one skilled in the art. In other embodiments, a single step ce is used. In some embodiments, the single step sequence comprises adding NHZOHHCl and a mixture of THF and water. In some embodiments, 1 equivalent of the compound of formula 3 is combined with a 1.1 equivalents ofNHZOHHCl in a 10:1 v/v mixture of ter.

Step 4 Step 4 illustrates how the oxime fi is then transformed and engaged in a [3+2] cycloaddition to for the isoxazole g. This transformation can be conducted in one pot but requires two distinct steps. The first step is an oxidation of the oxime functional group into a nitrone, or a similar intermediate with the same degree of oxidation, for example a chlorooxime. This reactive species then reacts with an alkyne in a [3+2] cycloaddition to form the isoxazole adduct.

In some embodiments, the suitable isoxazole-formation conditions consists of two steps, the first step comprising reacting the compound of formula 4 under suitable chlorooxime formation ions to form a chlorooxime intermediate; the second step comprising reacting the chlorooxime intermediate with acetylene under suitable cycloaddition conditions to form a nd of formula 5.

In some embodiments, the oxime ion conditions are selected from a) N—chlorosuccinimide and suitable solvent; b) potassium peroxymonosulfate, HCl, and dioxane; and c) Sodium hypochlorite and a suitable solvent Examples of suitable solvents e, but are not limited to, nonprotic solvents (e.g., DCM, DCE, THF, 2-MeTHF, MTBE and dioxane), aromatic hydrocarbons (e.g. toluene, xylenes), and alkyl acetates (e. g., isopropyl acetate, ethyl acetate).

Isolation of the product can be achieved by adding an antisolvent to a solution of a compound of formula 5. Examples of suitable solvents for ing the chlorooxime intermediate include mixtures of suitable solvents (EtOAc, IPAC) with hydrocarbons (e.g., hexanes, heptane, cyclohexane), or ic hydrocarbons (e. g., toluene, xylenes). In some embodiments, heptane is added to a solution of chlorooxime in IPAC.

Suitable cycloaddition conditions consist of combining the chlorooxime with acetylene with a suitable base and a suitable t. Suitable solvents include protic solvents, aproptic solvents, polar solvents, and nonpolar ts. Examples of suitable solvent include, but are not limited to, acetonitrile, ydrofuran, 2-methyltetrahydrofuran, MTBE, EtOAc, i—PrOAc, DCM, toluene, DMF, and ol. Suitable bases include, but are not d to, ne, DIEA, TEA, t—BuONa, and K2C03. In some embodiments, suitable cycloaddition conditions se adding 1.0 equivalents of chlorooxime, 1.0 equivalents of acetylene, l.l equivalents of Et3N in DCM.

Isolation of the product can be achieved by adding an antisolvent to a solution of a compound of formula 5. Examples of suitable solvents for isolating the chlorooxime include mixtures of suitable solvents (EtOAc, IPAC) with hydrocarbons (e. g., s, heptane, cyclohexane), or aromatic hydrocarbons (e.g., toluene, xylenes). In some embodiments, heptane is added to a solution of chlorooxime in IPAC.

Step_ 5 Step 5 depicts the final step(s) of the preparation of compounds of formula I.

When the R4 group is bromo, intermediate Q can be subjected to a Suzuki cross—coupling with boronic acid or esters, under conditions known to those skilled in the art, to form compounds where R4 an aryl, heteroaryl or alternative moieties ing from the metal- assisted coupling reaction. When intermediate Q is suitably functionalised, a deprotection step can be carried out to remove the protecting groups and generate the compounds of formula I.

Metal ed coupling reactions are known in the art (& e. g., Org.Proc. Res.

Dev. 2010, 30—47). In some ments, suitable coupling conditions comprise adding 0.1 equivalents of Pd[P(tBu)3]2; 1 equivalent of boronic acid or ester; and 2 equivalents of sodium carbonate in a 2:1 ratio v/v of acetonitrile/water at 60-70°C. In other ments, suitable coupling conditions comprise adding 0.010—0.005 equivalents Pd(dtbpf)Clz, 1 equivalent of c acid or ester, and 2 equivalents of potassium carbonate in a 7:2 v/v of toluene and water at 70 °C The final product can treated with a metal scavenger (silica gel, functionalized resins, charcoal) (flegu Org. Proc. Res. Dev. 2005, 198—205). In some embodiments, the solution of the product is treated with Biotage MP-TMT resin.

The product can also be isolated by crystallization from an alcoholic solvent (e.g. ol, ethanol, isopropanol). In some embodiments the solvent is ethanol. In other embodiments the solvent is isopropanol.

Deprotection of Boc groups is known in the art (see e. g. Protecting Groups in Organic sis, Greene and Wuts). In some embodiments, suitable deprotection conditions are hydrochloric acid in e at 35-45 °C. In other embodiments, suitable deprotection conditions are TFA in DCM.

Step 6 Step 6 illustrates how compounds of formula I are ted to compounds of a I-A using a base under le conditions known to those skilled in the art. In some ments, isolation of the free—base form of compounds of formula I may be achieved by adding suitable base, such as NaOH to an alcoholic acidic solution of compounds of formula Ito precipitate the t.

Step_ 7 Step 7 illustrates how compounds of formula I—A are converted to compounds of formula I-B using an acid under syuitable conditions known to those skilled in the art.

In some ments suitable conditions involve adding aqueous HCl, to a suspension of compounds of formula I-A in acetone at 35 °C then heating at 50 °C.

SCHEME B: Formation of dl-boronate x W x O\,0 1) Base Boronate 2) D20 Formation —> —> O=S=O Scheme B shows a general synthetic method for the ation of onate intermediates. A suitable l—halo—(is0pr0pylsulf0nyl)benzene is treated with a base such as, but not d to NaH, LiHMDS 0r KHMDS followed by quenching of the anion with deuterium source such as D20. The halogen is then transformed into a suitable boronate derivative via, for example, metal mediated cross-coupling catalyzed by, for instance, Pd(tBu3)2 0r Pd(dppf)Clz-DCM.

SCHEME C: ion of d6-boronate x W x O\,0 1) Base Boronate 2) DSCX Formation —> —> 02520 o=s=o | DWD 025:0 Do DD 3W3 D D Scheme C shows a general synthetic method for the ation of d6—b0r0nate intermediates. A suitable l-halo-(methylsulfonyl)benzene is treated with a base such as, but not limited to NaH, LiHMDS 0r KHMDS followed by quenching of the anion with deuterium source such as D3CI. This reaction is repeated until the desired amount of deuterium has been incorporated into the molecule. The halogen is then transformed into a suitable boronate tive via, for example, metal mediated cross-coupling catalyzed by, for instance, Pd(tBu3)2 0r Pd(dppf)Clz-DCM.

WO 49726 SCHEME D: Formation of d7-boronate Br Br Br W 0\ ’0 Alkylatlon. Oxudatlon_ . Boronate _ _ Formation D>Hs\'< D D:i':D <j O=S=O D D D D D D D D D D DWI)D D D D D Scheme D shows a general synthetic method for the ation of d7-boronate ediates. 4-Bromobenzenethiol is treated with a base such as, but not limited to NaH, LiHMDS or KHMDS followed by quenching of the anion with deuterium source such as 1,1,l,2,3,3,3-heptadeuterio—2—iodo—propane. The sulfide is then oxidized to the corresponding sulfone using, for example, mCPBA or Oxone. The halogen is then transformed into a suitable boronate derivative via, for example, metal mediated cross-coupling catalyzed by, for ce, Pd(tBu3)2 or Pd(dppf)Clz-DCM.

SCHEME E: Formation of aryl ring deuterated boronate Br Br )< )< Br 0 O O 0 B B / / I I :—:X m \TX Oxidation \TX Egrrorgitgn /—'X Deuterogenation —> /4D | | S O=S=0 I A A O;S:O o;s:o 1)Base 2) D20 \ f , , : Br QB’O O‘B,O Boronate - —:X / Formation Deuterogenatlon / \ —> —'X ’ 4D I I \ \ O=S=O a; fi?0%,,O ‘1?UM0 Scheme E shows a general synthetic method for the preparation of boronate intermediates where the aryl ring is substituted with a deuterium. A suitable l—iodo—4—bromo— aryl derivative is treated with a substituted thiol such as propanethiol under metal catalyzed coupling conditions using a catalyst such as CuI. The sulfide is then oxidized to the corresponding sulfone using, for example, mCPBA or Oxone. The bromide is then ormed into a suitable boronate derivative via, for example, metal mediated cross— coupling zed by, for instance, Pd(tBu3)2 or Pd(dppf)Clz-DCM. The remaining substituent is then converted into deuterium by, for instance, metal zed halogen- deuterium exchange using a suitbale metal st, such as Pd on C under an atmposhere of deuterium gas. In addition, the l-bromo-(isopropylsulfonyl)benzene can be treated with a base such as, but not limited to NaH, LiHMDS or KHMDS followed by quenching of the anion with deuterium source such as D20. The bromide is then transformed into a le te derivative via, for example, metal mediated cross-coupling catalyzed by, for instance, Pd(tBu3)2 or Pd(dppf)Clz-DCM. The remaining substituent is then converted into deuterium by, for instance, metal catalyzed halogen-deuterium exchange using a suitbale metal catalyst, such as Pd on C under an here of deuterium gas.

SCHEME F: Formation of aryl ring deuterated boronate Br Br Br W W /—:X / 0~ ,O I —:X O‘B,O —|X \ \ \ / . . . BFglfiriilignt ojs::_' Deuterogenatlon- <3D/ Alkylatlon Ion DhiflD DO:S:O D DDDDD DDDDD DWI) DozszoD D D D D D DWD D D D Scheme F shows another general synthetic method for the preparation of te intermediates where the aryl ring is substituted with a deuterium. A substituted 4— bromobenzenethiol is treated with a base such as, but not limited to NaH, LiHMDS or KHMDS followed by quenching of the anion with deuterium source such as l,l,l,2,3,3,3— heptadeuterio—2—iodo—propane. The sulfide is then oxidized to the ponding sulfone using, for example, mCPBA or Oxone. The halogen is then transformed into a suitable te derivative via, for e, metal mediated cross-coupling catalyzed by, for instance, Pd(tBu3)2 or Pd(dppf)Clz-DCM. The remaining substituent is then converted into deuterium by, for instance, metal zed halogen-deuterium exchange using a suitbale metal catalyst, such as Pd on C under an atmposhere of deuterium gas.

SCHEME G: Formation of aryl ring deuterated boronate Br Br Br / / / .—X / . . \ , Alkylatlon —'X OXIdatIon. —'x 1)Ba.se Boronate —|x l | — ’ \ ’ \ 2)D30X Formation \ —> O=S=O S O=S=O SH D D I I D D D D O‘ ,0 0‘ ,0 B B fix/ / Deuterogenatlon_ —:D \ —> \ O=S=O 0=S=O ”W”D D D”W”D D D D D Scheme G shows another general synthetic method for the preparation of boronate intermediates where the aryl ring is substituted with a deuterium. A substituted 4— bromobenzenethiol is treated with a base such as, but not limited to NaH, LiHMDS or KHMDS ed by quenching of the anion with for instance Mel. The sulf1de is then oxidized to the corresponding sulfone using, for example, mCPBA or Oxone. The sulfone is treated with a base such as, but not limited to NaH, LiHMDS or KHMDS ed by quenching of the anion with deuterium source such as D3CI. This reaction is repeated until the desired amount of deuterium has been orated into the molecule. The n is then transformed into a suitable boronate derivative via, for example, metal mediated cross— coupling catalyzed by, for instance, Pd(tBu3)2 or Pd(dppf)Clz-DCM. The ing substituent is then converted into deuterium by, for instance, metal catalyzed halogen- deuterium exchange using a suitbale metal catalyst, such as Pd on C under an atmposhere of deuterium gas.

SCHEME H: Formation of aryl ring deuterated oxime intermediates 3, L / 0/ / O I 3" / I X\ o\ ation I Hydrolysis 7‘ 0\ Protection /\0 / 7‘ o —> \ X | o X o *4 °\ /\00 o .

Deuterogenation Reductlve Reduction /\0 Amination D7 I D// 0 Lo Lo \I/ HoeN \I/ 300 _ . o 0 /\° \ T \ Oxlme Formation Protection /\0 \ 0T0 3/ ”KW D/I/ IV ”KR” ”K”11 DRW‘1 R9bR1§11RasnsbR1?“ 39a Rab R1 Scheme H shows a general synthetic method for the preparation of oxime intermediates where the aryl ring is tuted with a deuterium. The methyl group of an appropriately substituted methyl 4-methylbenzoate derivative can be converted into the corresponding ide under conditions such as AIBN catalyzed bromination with NBS.

This di-bromide is then hydrolysed to the corresponding aldehyde, for ce using AgNO3 in acetone/water. Protection of the aldehyde as a le acetal, for instance the diethyl acetal and subsequent conversion of the remaining substituent into deuterium by, for instance, metal catalyzed halogen-deuterium exchange using a suitable metal catalyst, such as Pd on C under an atmosphere of deuterium gas gives the deuterated ester intermediate. The ester functionality can be reduced using reagents such as LiAlH4, NaBH4, NaBD4 or LiAlD4 to give corresponding aldehyde. This can be reacted under reductive amination conditions using a le amine, such as methylamine or d3-methylamine using a reducing agent such as NaBH4 or NaBD4 to give the corresponding amine derivative. This can be protected with, for instance a Boc group and the acetal converted into the oxime using, for ce, hydroxylamine hydrochloride in THF/water.

SCHEME 1: Formation of aryl ring deuterated oxime intermediates \ 0/ / \O I . . Br / . | ’ o\ Bromination I ysus )ZA 0\ Protection. l \0 X 7 O \ /| O x 0 /\ O \ \ o \ o o Deuterogenaiion\0 / \ I Amide Formation \0 Reduction \0 \ BOC ) 0\ * I I 7‘ // NH2 ’ D// NH2 m) D D o 0 R93 R9!) \0 4/ \o 4/ ”°‘~ 4/ o o o o \ 0Y0 Alkylaiion \o \ Oxime Formation \ | —> | —> | // NH D D// N\I<R1° D// N\I<R“° Rea R9b R95 RED RIB" R9“ Rsb R1511 ] Scheme 1 shows another general synthetic method for the preparation of oxime intermediates where the aryl ring is substituted with a deuterium. The methyl group of an appropriately substituted methyl 4-methylbenzoate derivative can be converted into the corresponding dibromide under conditions such as AIBN zed bromination with NBS.

This di-bromide is then hydrolysed to the corresponding aldehyde, for instance using AgNO3 in acetone/water. Protection of the aldehyde as a suitable acetal, for instance the dimethyl acetal and subsequent conversion of the remaining subtituent into deuterium by, for instance, metal catalyzed halogen-deuterium exchange using a suitable metal catalyst, such as Pd on C under an atmosphere of ium gas gives the deuterated ester intermediate. The ester onality can be converted into the corresponding primary amide under standard conditions, such as heating with a solution of ammonia in methanol. The amide can be reduced to the ponding amine using reagents not limited to LiAlH4 or LiAlD4. This can be ted with, for instance a Boc group. The carbamate NH can be alkylated under basic conditions using for instance NaH, LiHMDS or KHMDS followed by quenching of the anion with deuterium source such Mel or D3CI. The acetal can be ted into the oxime using, for instance, hydroxylamine hydrochloride in THF/water.

WO 49726 SCHEME J: Formation of aryl ring deuterated oxime intermediates / 0/ / O X\‘ o\ Bromination 3" : I I Hydrolysis )Efi 0\ Protection \0 / \ I o x 0 >24 0\ \o \o \o \ \ - Deuterogenation 0 / \ I Amide Formation 0 I Reduction \0 \ —> —> —> I 2;?ggi'3: 7~ O\ // NHZ D// NHZ —> D D o 0 ma R9" o \O \I/ HotIN \I/ \o \ Boc \ o o \ OXIme Formation. . o o l/ / H - O \ Protection Nj<R1° |/ / —* N R10 I, / T R10 R95 R9b R1?” 11 D R95 R9b\R'<1 R93 R9:Rl<1511 Scheme J shows another general synthetic method for the preparation of oxime intermediates where the aryl ring is substituted with a deuterium. The methyl group of an appropriately substituted methyl 4-methylbenzoate derivative can be converted into the corresponding dibromide under conditions such as AIBN catalyzed bromination with NBS.

This mide is then hydrolysed to the ponding aldehyde, for instance using AgNO3 in acetone/water. tion of the aldehyde as a suitable acetal, for instance the dimethyl acetal and subsequent conversion of the remaining tuent into deuterium by, for instance, metal catalyzed halogen-deuterium exchange using a suitable metal catalyst, such as Pd on C under an atmposhere of deuterium gas gives the deuterated ester intermediate. The ester onality can be converted into the corresponding primary amide under standard conditions, such as heating with a on of ammonia in methanol. The amide can be reduced to the corresponding amine using reagents not limited to LiAlH4 or LiAlD4. This can be reacted under reductive amination conditions using a suitable amine, such as amine, d3 -methylamine, formaldehyde or d2-formaldehyde using a ng agent such as NaBH4 or NaBD4 to give the corresponding amine derivative. This can be protected with, for instance a Boc group. The acetal can be converted into the oxime using, for instance, hydroxylamine hydrochloride in THF/water.

SCHEME K: Formation of aryl ring deuterated oxime ediates Rsa R93 \ 0 / o / I Br / I I / 0\ N ysis Deuterogenation X I O\ , O\ 7\ O \ )Zfi I? X O O 0 0+ 0+ Reduciive \o \ \ O 0 - \ O O —>Ammat'°n. . I// Protection O \ Alkylation o \ ””2 —> I// TIT-l —> I // N R10 R95 R9” D D 11 Rea Ran R95 R9:RI<1 \I/ 9 \I/ \I/ 0 O O O O O Reduction HO \ Oxidation \ Oxime Formation \ —> I —> —> N\'<R10 I 10 I NKR D// NKR10 D// // D 11 Rga Rab R15" Rea Rab R1511 Rsa Rab R1 Scheme K shows another l synthetic method for the preparation of oxime intermediates where the aryl ring is substituted with a deuterium. The methyl group of an appropriately substituted methyl ylbenzoate derivative can be converted into the corresponding dibromide under conditions such as AIBN catalyzed bromination with NBS.

This di-bromide is then hydrolysed to the corresponding aldehyde, for instance using AgNO3 in e/water. Protection of the aldehyde as a le acetal, for instance the dimethyl acetal and subsequent sion of the remaining substituent into deuterium by, for instance, metal zed halogen-deuterium exchange using a suitable metal catalyst, such as Pd on C under an atmosphere of deuterium gas gives the deuterated ester intermediate. This can be reacted under reductive amination conditions using a suitable amine, such as ammonium hydroxide using a reducing agent such as NaBH4 or NaBD4 to give the corresponding amine derivative. This can be protected with, for instance a Boc group and the carbamate NH alkylated under basic conditions using for instance NaH, LiHMDS or KHMDS followed by quenching of the anion with ium source such Mel or D3CI. The ester can be reduced to the corresponding alcohol using a suitable reducing agent such as LiBH4 or NaBH4. The alcohol can be oxidized to the aldehyde using regeants such as MnOz or Dess-Martin periodane. The acetal can be converted into the oxime using, for instance, aqueous hydroxylamine.

SCHEME L: Formation of deuterated oXime intermediates k L0 to O \l/ Reductive BOC O O Amination /\O Protection /\0 /\0 —> H —> N R10 N R10 0 11 \'< 11 Rsa R9b R15 Rea R9b R15 H0\|N \l/ O O Oxime Formation N R10 Baa 393:5“ Scheme L shows a general synthetic method for the preparation of ated oxime intermediates. 4—(diethoxymethyl)benzaldehyde can be reacted under reductive amination conditions using a suitable amine, such as methylamine or d3 -methylamine using a reducing agent such as NaBH4 or NaBD4 to give the corresponding amine derivative. This can be protected with, for ce a Boc group and the acetal converted into the oxime using, for ce, hydroxylamine hydrochloride in THF/water.

SCHEME M: Formation of deuterated oXime intermediates \ \ o O \o \0 \4/ \ \ 0 Amide Formation 0 Reduction \0 Eggction \O —> —> 0Y0 o\ NH2 NH2 —> NH 0 0 R93 Rab R93 R9” \0 \P HO\|N \~/ Alkylaton' \ O O O O o 7: 0xim9 F0rmatoi n R10 N H10 Rsa F193;?“ R95 391:7: ] Scheme M shows another general synthetic method for the preparation of deuterated oxime intermediates. The ester functionality of methyl 4- (dimethoxymethyl)benzoate can be converted into the corresponding primary amide under standard conditions, such as heating with a solution of ammonia in methanol. The amide can be reduced to the corresponding amine using reagents not limited to LiAlH4 or LiAlD4. This can be protected with, for instance a Boc group. The carbamate NH can be alkylated under basic ions using for instance NaH, LiHMDS or KHMDS followed by quenching of the anion with ium source such Mel or D3CI. The acetal can be converted into the oxime using, for instance, hydroxylamine hloride in THF/water.

SCHEME N: Formation of deuterated oXime intermediates \0 \o \ \ \ \0 Rafictlve' 0 Amide Formation 0 Reduction 0\ —> atlon NHz —> NH2 —> o o R95 R9” 0 \O \‘/ HoxlN \l/ \O Eggcfion \O O O 0 O H Oxime Formation NKR10 N R10 N R10 Fi9‘I R9h 31511 11 Rea Raul? Rea 39:55“ Scheme N shows another general synthetic method for the ation of deuterated oxime intermediates. The ester functionality of methyl 4- (dimethoxymethyl)benzoate can be converted into the corresponding primary amide under standard conditions, such as heating with a solution of ammonia in methanol. The amide can be reduced to the corresponding amine using reagents not limited to LiAlH4 or LiAlD4. This can be reacted under reductive amination conditions using a suitable amine, such as methylamine, d3 -methylamine, formaldehyde or d2-formaldehyde using a reducing agent such as NaBH4 or NaBD4 to give the corresponding amine derivative. This can be protected with, for instance a Boc group. The acetal can be converted into the oxime using, for instance, hydroxylamine hydrochloride in ter.

SCHEME 0: ion of deuterated oXime intermediates o \l/ 0 \l/ 0 Protection \0 Alkylation \0 ””2 —> 0Y0 —> 0Y0 NH N R10 Rea R9b \|< Rsa Rsb R93 Rab R1?“ i , + i . o o o o Reduction Y Oxudation. . Y Oxume Formation. . o 0 N R10 N R10 N R10 11 11 mfi/ 11 R93 R9:R'<1 R9: Ref]: R93 Rabi-E1 Scheme 0 shows another general synthetic method for the preparation of deuterated oxime intermediates. A 4-substituted benzylmine can be ted with, for instance a Boc group. The carbamate NH can be alkylated under basic conditions using for ce NaH, LiHMDS or KHMDS followed by quenching of the anion with deuterium source such Mel or D3CI. The ester can be reduced to the corresponding alcohol using a suitable reducing agent such as LiBH4 or NaBH4. The alcohol can be ed to the aldehyde using ts such as MnOz or Dess-Martin periodane. The acetal can be ted into the oxime using, for instance, aqueous hydroxylamine.

SCHEME P: Formation of isoxazole atives (BOC)2N NH2 NH2 TMS (BOC)2N TMS Br é é , R5 N \ Sono ashira N \ BOG N \ 1)Suzukl | —>g| Protection | 2) TMS Removal /N / N —> /N _ ) Br 3' 3' o=s=o R18 R81; R1!) R2 Rah R1c Rae [3+2] cycloadditon | \‘/ 0Y0 Chlorination C| 0Y0 N R10 N R10 R7 \l< R7 Ra RSa R9!) R1 HE R95 Rsb R1K511 a Ha /N R :‘9 Rab R R5 Deproteciion R5 —> 34 o=s=o o=s=o R13 R30 R14: R30 Fl‘lb R2 Rab R1“ R3“ Rm R2 Ran R1c R3: Scheme P shows a general synthetic method for the preparation of deuterated pyrazine-isoxazole derrivatives. 3,5-Dibromopyrazinamine is converted into the corresponding silyl-protected alkyne under standard Sonagashira conditions utilizing, for example, Pd(PPh3)4 and CuI as catalysts. The pyrazine NHZ can then be protected as, for example the di-Boc derivative. Coupling of the ne bromide with a boronate, for instance those outlined in Schemes l to 6 above, under standard Suzuki cross—coupling conditions ed by removal of the silyl protecting group give the desired alkyne intermediate. Oximes, such as those outline in Schems 7 to 14 above, can be converted into the corresponding chlorooximes using, for instance, NCS. The alkyne and chlorooxime intermediates can undergo a [3+2] cycloaaditon to give corresponding ole under standard conditions, for instance by the addition of Et3N. The Boc protecting groups can be removed under acicid conditions such as TFA in DCM or HCl in MeOH/DCM to give the ated pyrazine isoxale derrivatives.

SCHEME Q: Formation of deuterated isoxazole derrivatives O 0 NH2 o—N F30 \\ HN‘éR" FSCJkNH O"! \ R12 ‘(NTQRRH10 R93 Hm: N \ R12 /N R7 R93 RB I 9b /N R7 R Protection R5 Halogenation —> —> O=S=O 0—8—0_ _ R13 Rae Ria Rae R1b H2 Rab H10 R“ WRa Rab R16 Raa kyN R93 X R7 R9” R8 Deprotection Deuterogenation —> —> RHZ)4 O=S=O O—S—O_ _ R1a R3: R18 R30 Rn: R2 Rab R1“ R2 Rab R10 Raa R11: Raa N R10 R1 a Rae R1 b R2 Rab R1 0 Raa Scheme Q shows a general synthetic method for the preparation of deuterated isoxazole derrivatives. The pyrazine NHZ and benzylamineamine NH can be protected under standard conditions using roacetic anhydride. Halogenation 0f the isoxazole ring with, for example NIS followed by removal of the trifluoroacetate protecting group under basic conditions provides the desired halogenated interemdiates. The halogen can then converted into deuterium by, for instance, metal catalyzed n-deuterium exchange using a suitbale metal catalyst, such as Pd on C under an atmposhere of ium gas.

Abbreviations The following abbreviations are used: ATP adenosine triphosphate Boc tert-butyl ate Cbz Carboxybenzyl DCM dichloromethane DMSO dimethyl ide Et3N triethylamine 2-MeTHF 2-methyltetrahydrofuran NMM N—Methyl morpholine DMAP 4-Dimethylaminopyridine TMS Trimethylsilyl MTBE methyl tyl ether EtOAc ethyl acetate i-PrOAc isopropyl acetate IPAC isopropyl acetate DMF dimethylformamide DIEA diisopropylethylamine TEA triethylamine t—BuONa sodium tertbutoxide K2CO3 potassium carbonate PG ting group pTSA para-toluenesulfonic acid TBAF Tetra-n-butylammonium fluoride 1HNMR proton r magnetic resonance HPLC high performance liquid chromatography LCMS liquid chromatography-mass spectrometry TLC thin layer chromatography Rt retention time SCHEMES AND EXAMPLES The compounds of the disclosure may be prepared in light of the specification using steps generally known to those of ordinary skill in the art. Those compounds may be analyzed by known s, ing but not limited to LCMS (liquid chromatography mass spectrometry) and NMR (nuclear magnetic resonance). The following c schemes and examples illustrate how to prepare the compounds of the present disclosure. The examples are for the purpose of illustration only and are not to be construed as limiting the scope of the invention in any way. H-NMR spectra were recorded at 400 MHz using a Bruker DPX 400 instrument. Mass spec. s were analyzed on a MicroMass Quattro Micro mass spectrometer operated in single MS mode with electrospray ionization.

Compound 1—1 Method 1: To a solution of tetrahydropyranamine (100 g, 988.7 mmol) in MeOH (3.922 L) was added 4—(diethoxymethyl)benzaldehyde (196.1 g, 941.6 mmol) over 2 min at RT. The reaction mixture was stirred at RT for 80 min, until the aldimine formation was complete (as seen by NMR). NaBH4 (44.49 g, 1.176 mol) was carrefully added over 45 min, maintaining the temperature between 24 OC and 27 0C by mean of an ice bath. After 75 min at RT, the reaction has gone to completion. The reaction mixture was quenched with 1M NaOH (1 L).

The reaction mixture was partitioned n brine (2.5 L) and TBDME (4 L then 2 x 1 L).

The c phase was washed with brine (500 mL) and trated in vacuo. The crude mixture was redisolved in DCM (2 L). The aqueous phase was separated, the organic phase was dried over MgSO4, filtered and concentrated in vacuo to give the title compound as a yellow oil (25299 g, 91%).

Method 2: A solution of N-[[4-(diethoxymethyl)phenyl]methyl] tetrahydropyranamine (252.99 g, 862.3 mmol) and Boc anhydride (191.9 g, 202.0 mL, 879.5 mmol) in DCM (2.530 L) was cooled down to 3.3 °C. Et3N (89.00 g, 122.6 mL, 879.5 mmol) was added over 4 min, keeping the al temperature below 5 °C. The bath was removed 45 min after the end of the addition. And the reaction mixture was stirred at RT overnight. The reaction mixture was sequentially washed with 0.5 M citric acid (1 L), saturated NaHCO3 solution (1 L) and brine (1 L). The organic phase was dried (MgSO4), filtered and concentrated in vacuo to give a colourless oil (372.38 g, 110%). 1H NMR (400.0 MHz, DMSO); MS (ES+) Method 3: tert-butylN—[[4-(diethoxymethyl)phenyl]methyl]-N-tetrahydropyranyl-carbamate (372.38 g, 946.3 mmol) was dissolved in THF (5 L) and water (500 mL). Hydroxylamine hloride (72.34 g, 1.041 mol) was added in one portion and the reaction mixture was stirred overnight at RT. The reaction mixture was partitioned between DCM (5 L) and water.

The combined organic extract was washed with water (1L x 2). The c phase was concentrated in vacuo to a volume of about 2L. The organic layer was dried over MgSO4, filtered and concentrated in vacuo to give a sticky colourless oil that crystallized on standing under vacuo. (334.42g, 106%). 1H NMR (400.0 MHz, ; MS (ES+) Method 4: utylN—[[4-[(E)-hydroxyiminomethyl]phenyl]methyl]-N-tetrahydropyranyl- carbamate (334.13 g, 999.2 mmol) was dissolved in isopropyl acetate (3.0 L) (the e was warmed to 40 °C to allow all the solids to go into on). N—chlorosuccinimide (140.1 g, 1.049 mol) was added nwise over 5 min and the reaction mixture was heated to 55 OC (external block temperature). After 45 min at 55°C The reaction had gone to completion.

The reaction mixture was cooled down to RT. The solids were d off and rinsed with Isopropyl acetate (1 L). Combined organic extract was sequentially washed with water (1.5 L, 5 times) and brine, dried over MgSO4, filtered and concentrated in vacuo to give a viscous yellow oil (355.9 g; 96%). 1H NMR (400.0 MHz, CDC13); MS (ES+) Method 5: Et3N (76.97 g, 106.0 mL, 760.6 mmol) was added over 20 minutes to a solution of tert—butyl N—(5 -bromo-3 -ethynyl-pyrazinyl)-N-tert-butoxycarbonyl-carbamate (23 3 .0 g, 585.1 mmol) and tert—butyl N—[[4-[(Z)—C—chloro-N—hydroxy-carbonimidoyl]phenyl]methyl]- N—tetrahydropyranyl-carbamate (269.8 g, 731.4 mmol) in DCM (2.330 L) at RT. During addition of triethylamine, the rm was stabilised by cooling the mixture in an ice bath, then the reaction mixture was gradually warmed up to RT and the mixture was stirred at RT ght. The reaction e was sequentially washed with water (1.5 L, 3 times ) and brine. The organic extract was dried over MgSO4, filtered and partially concentrated in vacuo. Heptane (1.5L) was added and the concentration was ued yielding 547.63 g of a yellow—orange solid. 542.12 g was taken up into ~2 vol (1 L) of ethyl acetate. The mixture was heated to 74-75 0C internally and stirred until all the solid went into solution. Heptane (3.2 L) was added slowly via addition funnel to the hot solution keeping the internal temperature between 71 °C and 72 °C . At the end of the addition, the dark brown solution was seeded with some tallised product, and the reaction mixture was allowed to cool down to RT without any stirring to crystallise O/N. The solid was filtered off and rinsed with heptane (2 x 250 mL), then dried in vacuo to yield 307.38 g of the title product (72 %). %). 1H NMR (400.0 MHz, CDCl3); MS (ES+) Method 6: tert—butyl N—[[4—[5—[3—[bis(tert-butoxycarbonyl)amino]bromo-pyrazinyl] isoxazol-3 -yl]phenyl]methyl]-N-tetrahydropyranyl-carbamate (303 g, 414.7 mmol) and 2- methyl[4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolanyl)pyridyl] propanenitrile (112.9 g, 414.7 mmol) were suspended in MeCN (2 L) and H20 (1 L). NazCO3 (414.7 mL of 2 M, 829.4 mmol) followed by Bu)3]2 (21.19 g, 41.47 mmol) were added and the reaction mixture was degassed with N2 for 1 h. The reaction mixture was placed under a nitrogen atmosphere and heated at 70 OC (block temperature) for 4 h (internal ature fluctuated between 60 °C and 61 OC). The reaction was cooled down to room temperature and stirred at RT overnight. The reaction mixture was partitioned between EtOAc (2 L) and water (500 mL). The combined organic extract was washed with brine (500 mL), filtered h a short pad of celite and concentrated under reduced pressure to a volume of about 3 L. The on was dried over MgSO4, filtered and partially concentrated in vacuo. iPrOH (1.5 L) was added and the solvent was removed in vacuo to yield the desired product as a light brown foam (405 400 g was taken up into ~5 vol (2 L) of iPrOH and the mixture was heated to 80 0C until all the solid went into solution. The dark brown solution was seeded, and the reaction mixture was d to slowly cool down to RT overnight. The solid was d off and rinsed with iPrOH (2 x 250 mL) and Petroleum ether (2x200 mL). The resulting solid was 2012/058127 slurried in petroleum ether (2.5 L), filtered off and dried in vacuo. The resulting solid was dissolved in DCM (2.5 L) and stirred slowly for 1 h with 30 g of SPM32 ( 3-mercaptopropyl ethyl sulfide silica). The silica was filtered through a pad of florisil and rinsed with DCM.

The procedure was ed twice, then the DCM solution was trated in vacuo to give 238.02 g of a light yellow solid.

Method 7: tert—butyl N—[[4—[5—[3—[bis(tert-butoxycarbonyl)amino][2-(1-cyanomethylethyl )pyridyl]pyrazinyl]isoxazolyl]phenyl]methyl]-N-tetrahydropyranyl- carbamate (238 g, 299.0 mmol) was dissolved in DCM (2.380 L). TFA (500 mL, 6.490 mol) was added at RT over 3 min. The reaction mixture was stirred at RT for 3.5 h. The reaction mixture was concentrated under reduced pressure then azeotroped with e (2x3 00ml).

The oil was then slurried in abs. EtOH (2.5 L) and filtered . The solid was dissolved in a mixture of ethanol (1.190 L) and water (1.190 L). potassium carbonate (124.0 g, 897.0 mmol) in water (357.0 mL) was added to the solution and the mixture was stirred at RT overnight.

The solid was filtered off, was washed with water (2.5 L), and dried at 50 °C in vacuo to give 108.82 g of the title compound (Compound 1—1) as a yellow powder. (73 %) Methods 6a and 7a 0\ /O B Method 6a Boc\ ,B NWNNC(53'N\ / \ \N CN | Boc /N (3O —> 1. cat. pf)C|2, PhCH3, aq. K2C03 2. crystallization Boc\ Boc O’N Method 7a W \ 1. TFA \ N \ NI N\Boc DCM I NH /” 6 / °C 2. NaOH / GO / o I 90A)0 I \N CN \N CN Compound 1-] A mixture of tert—butyl N—[[4-[5-[3-[bis(tert-butoxycarbonyl)amino]bromo- pyrazin-2—yl] isoxazolyl]phenyl]methyl]-N-tetrahydropyranyl-carbamate (110.0 g, 151 mmol), K2C03 (41.6 g, 301 mmol), and 2-methyl-2—[4-(4,4,5,5-tetramethyl-1,3,2- dioxaborolan-2—yl)—2—pyridyl] propanenitrile (41.0 g, 151 mmol) in toluene (770 mL) and water (220 mL) is d and degassed with N2 for 30 min. at 20 OC. The catalyst Pd(dtbpf)C12 (1.96 g, 3.01 mmol) is added and the mixture is degassed for an additional 10 min. The mixture is heated at 70 0C until the reaction is complete. The mixture is cooled to ambient temperature, diluted with water (220 mL), and filtered through a bed of Celite. The organic phase is concentrated to remove most of the solvent. The trate is diluted with i-PrOH (550 mL). The resultant suspension is stirred for at least 1 h and then the solid is collected by ion to afford a tan . The solid is dissolved in toluene (990 mL) and stirred with Biotage MP-TMT resin (18.6 g) for 2 h at ambient temperature. The resin is d by filtration. The filtrate is concentrated then diluted with i-PrOH (550 mL) and then re-concentratd. Add i-PrOH (550 mL) and stir for 1 h at ambient temperature. Cool the suspension to 5 °C and collect the solid by filtration then dry to afford tert—butyl N—[[4—[5—[3- [bis(tert—butoxycarbonyl)amino][2-(1-cyanomethyl-ethyl)pyridyl]pyrazin xazolyl]phenyl]methyl]-N-tetrahydropyranyl-carbamate und 1-1) (81.9 g; 68%, yield, 98.7 area % purity by HPLC) as a cream—colored powder.

Form Chan e to Com ound I-1°HC1°1.5 H20 A suspension of tert—butyl N—[[4—[5—[3—[bis(tert—butoxycarbonyl)amino]—6—[2—(1— cyanomethyl-ethyl)pyridyl]pyrazinyl]isoxazol-3 -yl]phenyl]methyl] -N- ydropyranyl-carbamate (Compound 1-1) (36.0 g, 72.6 mmol) in CH3CN (720 mL) is stirred at ambient temperature (20 0C) in a flask equipped with mechanical stirring. A 1 M aqueous solution of HCl (72.6 mL; 72.6 mmol) is added. The suspension is stirred at ambient temperature for 20 h. The solid is collected by filtration. The filter-cake is washed with CH3CN (3 x 50 mL) then dried under vacuum with high humidity for 2 h to afford 2012/058127 Compound I—1°HCl°1.5 H20 (30.6 g; 74%) yield, 98.8 area % purity by HPLC) as a yellow powder. .1H NMR (400 MHz, DMSO) 5 9.63 (d, J= 4.7 Hz, 2H), 9.05 (s, 1H), 8.69 (d, J= .2 Hz, 1H), 8.21 (s, 1H), 8.16 — 8.03 (m, 3H), 7.84 (t, J= 4.1 Hz, 3H), 7.34 (br s, 2H), 4.40 — 4.18 (m, 2H), 3.94 (dd, J: 11.2, 3.9 Hz, 2H), 3.32 (t, J: 11.2 Hz, 3H), 2.17 — 2.00 (m, 2H), 1.81 (s, 6H), 1.75 (dd, J= 12.1, 4.3 Hz, 2H). - 4-is0 r0 lsulfon l hen l razin-Z-amine Com ound II-1 (BOC)2N NHZ TMS N TMS I NJYBr // /N N \ 300 N \ 1) Suzuki | Sonogashira /N —> | Protection | 2) TMS Removal / N —> / N —> Br Br Br i ii iii 'V \O \o \ \ \ O Amide Formation 0 Reduction \0 —> —> ngection\ o\ NH2 NH D D 0 0 v vi VII VIII HO\IN Alkylation \OJWO7< Oxime Formation Hormo\'< Egiii'irgtcigiime Cl)\©::/O\i<N\ D D §< NW”\ H (BOC)2N O’N \\ N~ D | D | D [3+2] cycloadditon Deprotection —> —> O=S=O Oj:0 A xii 11-1 Step 1: S-Brom0((trimethylsilyl)ethynyl)pyrazinamine (Trimethylsilyl)acetylene (1.845 g, 2.655 mL, 18.78 mmol) was added dropwise to a on of 3,5-dibromopyrazinamine (compound i) (5 g, 19.77 mmol) in DMF (25 mL). Triethylamine (10.00 g, 13.77 mL, 98.85 mmol), copper(1) iodide (451.7 mg, 2.372 mmol) and Pd(PPh3)4 (1.142 g, 0.9885 mmol) were then added and the resulting solution stirred at RT for 30 minutes.The reaction mixture was diluted with EtOAc and water and the layers separated. The aqueous layer was extracted further with EtOAc and the combined organic layers washed with water, dried (MgSO4) and trated in vacuo. The residue was purified by column chromatography eluting with 15% EtOAc/Petroleum ether to give the product as a yellow solid (3.99 g, 75% Yield). 1H NMR (400.0 MHz, DMSO) 5 0.30 (9H, s), 8.06 (1H, s); MS (ES+) 271.82.

Step 2: tert—Butyl N-tert-butoxycarbonyI-N-[S-br0mo((trimethylsilyl)ethyynyl) pyrazin-Z-yl]carbamate (BOChN TMS 5-Bromo(2-trimethylsilylethynyl)pyrazinamine (2.85 g, 10.55 mmol) was dissolved in DCM (89.06 mL) and d with Boc anhydride (6.908 g, 7.272 mL, 31.65 mmol) ed by DMAP (128.9 mg, 1.055 mmol). The reaction was allowed to stir at ambient temperature for 2 hours. The mixture was then diluted with DCM and NaHCO3 and the layers separated. The aqueous layer was extracted further with DCM, dried (MgSO4), filtered and trated in vacuo. The resultant residue was d by column chromatography eluting with dichloromethane to give the d product as a colourless oil (4.95g, 99% Yield). 1H NMR (400.0 MHz, DMSO) 5 0.27 (9H, s), 1.42 (18H, s), 8.50 (1H, s); MS (ES+) 472.09.

Step 3: tert-Butyl N-(3-ethynyl—5—(4-(is0propylsulfonyl)phenyl)pyrazinyl)N- tertbutoxycarbonyl-carbamate tert-butyl (BOChN O=S=O WO 49726 N— [5 -Bromo(2-trimethylsilylethynyl)pyrazinyl]-N- tertbutoxycarbonylcarbamate (3 g, 6.377 mmol) and (4-isopropylsulfonylphenyl)boronic acid (1.491 g, 6.536 mmol) were dissolved in MeCN/water (60/12 mL). K3PO4 (2.706 g, 12.75 mmol) was added and the reaction mixture was degassed with a flow of nitrogen (5 cycles).

Pd[P(tBu)3]2 (162.9 mg, 0.3188 mmol) was added and the resulting mixture was stirred at room temperature for 1h. The reaction e was poured quickly into a mixture of ethyl e (500 mL), water (90 mL) and 1% aqueous sodium metabisulphite at 4 °C, shaken well and the layer separated. The organic fraction was dried over MgSO4, filtered and the filtrate was treated with 3-mercaptopropyl ethyl sulphide on silica (0.8mmol/g, l g), sorbed onto silica gel then purified by column chromatography on silica gel eluting with 30-40% EtOAc/petroleum ether. The solvents were concentrated in vacuo to leave the product as a yellow viscous oil that was triturated with petroleum ether to yield the product as beige crystals (1.95 g, 61% Yield); 1H NMR (400 MHz, DMSO) 5 1.20 (m, 6H), 1.39 (s, 18H), 3.50 (m, 1H), 5.01 (s, 1H), 8.03 (m, 2H), 8.46 (m, 2H) and 9.37 (s, 1H).

Step 4: 4-(Dimethoxymethyl)benzamide A mixture of methyl 4-(dimethoxymethyl)benzoate (3.8 g, 18.08 mmol) and 7M NH3 in MeOH (30 mL of 7 M, 210.0 mmol) in a sealed tube was heated at 110 0C for 22 hours. A further portion of 7M NH3 in MeOH (20 mL of 7 M, 140.0 mmol) was added and the reaction heated at 135 0C for 23 hours. The reaction was cooled to ambient temperature and the solvent removed in vacuo. The residue was mitted to the reaction conditions (7M NH3 in MeOH (30 mL of7 M, 210.0 mmol) at 115 0C) for a further 16 hours. The solvent was removed in vacuo and the e tritruated from EtZO. The resultant itate was ed by filtration to give the sub—title compound as a white solid (590 mg, 17% yield).

The filtrate was purified by column chromatography (ISCO Companion, 40 g column, eluting with 0 to 100% EtOAc/Petroleum Ether to 10% MeOH/EtOAc, loaded in EtOAc/MeOH) to give a further protion of the sub-title t as a white solid (225 mg, 6% Yield). Total isolated (815 mg, 23% Yield); 1H NMR (400 MHz, DMSO) 5 3.26 (s, 6H), 5.44 (s, 1H), 7.37 (s, 1H), 7.46 (d, J = 8.0 Hz, 2H), 7.84 — 7.91 (m, 2H) and 7.98 (s, 1H) ppm; MS (ES+) 196.0.

Steo 5: Dideuterio—[4—(dimethoxymethyl)phenyl]methanamine LiDH4 (12.52 mL of 1 M, 12.52 mmol) was added dropwise to a stirred solution of 4— (dimethoxymethyl)benzamide (815 mg, 4.175 mmol) in THF (20 mL) at 0 0C under an atmosphere of nitrogen. The reaction was heated at reflux for 16 hours then cooled to ambient temperature. The reaction was quenched by the sequential addition of D20 (1 mL), 15% NaOH in D20 (1 mL) and D20 (4 mL). The resultant solid was removed by filtration and washed with EtOAc. The filtrate was concentrated in vacuo and the residue dried by azeotropic lation with toluene (x 3) to give the sub—title compound as a yellow oil (819 mg) that was used without further purification; 1H NMR (400 MHz, DMSO) 5 3.23 (s, 6H), .36 (s, 1H) and 7.30 — 7.35 (m, 4H) ppm; MS (ES+) 167.0.

Step 6: tert—Butyl N-[dideuterio—[4-(dimethoxymethyl)phenyl]methyl]carbamate D D Viii Et3N (633.7 mg, 872.9 11L, 6.262 mmol) was added to a stirred suspension of dideuterio-[4-(dimethoxymethyl)phenyl]methanamine (765 mg, 4.175 mmol) in THF (15 mL) at 0 OC. The on was d to stir at this temperature for 30 minutes then Boc20 (956.8 mg, 1.007 mL, 4.384 mmol) was added in portions. The reaction was allowed to warm to ambient temperature and stirred for 18 hours. The solvent was removed in vacuo and the residue was d by column chromatography (ISCO Companion, 120 g , eluting with 0 to 50% EtOAc/Petroleum Ether, loaded in DCM) to give the sub-title product as a colourless oil (1.04 g, 88% Yield); 1H NMR (400 MHz, DMSO) 5 1.40 (s, 9H), 3.23 (s, 6H), .36 (s, 1H), 7.24 (d, J = 8.2 Hz, 2H), 7.33 (d, J = 8.0 Hz, 2H) and 7.38 (s, 1H) ppm. 2012/058127 Step 7: tert—Butyl N-[dideuterio-[4-(dimeth0xymethyl)phenyl]methyl]-N-methyl— carbamate D D LiHMDS (1M in THF) (1.377 mL of 1 M, 1.377 mmol) was added dropwise to a sittred solution of tert-butyl N-[dideuterio-[4-(dimethoxymethyl)phenyl]methyl]carbamate (300 mg, 1.059 mmol) in THF (5 mL) at -78 OC. The solution was stirred at this temperature for 10 minutes then iodomethane (225.4 mg, 98.86 uL, 1.588 mmol) was added dropwise and the mixture allowed to warm to ambient temperature over 1 hour. The reaction was again cooled to —78 °C and LiHMDS (1M in THF) (635.4 uL ofl M, 0.6354 mmol) was added.

After 10 minutes iodomethane (105.2 mg, 46.14 uL, 0.7413 mmol) was added and the reaction allowed to warm to t temperature over 6 hours. The mixture was diluted with EtOAc and the organic layer washed with saturated aqueous NaHCO3 (x 2), brine (x 1), dried (MgSO4) filtered and concentrated in vacuo. The residue was purified by column tography (ISCO Companion, 24 g column, eluting with 0 to 30% EtOAc/Petroleum Ether, loaded in DCM) to give the sub—title product as a colourless oil (200 mg, 63% Yield); 1H NMR (400 MHz, DMSO) 5 1.41 (d, J = 27.7 Hz, 9H), 2.76 (s, 3H), 3.24 (s, 6H), 5.37 (s, 1H), 7.23 (d, J = 7.9 Hz, 2H) and 7.37 (d, J = 8.0 Hz, 2H) ppm.

Step 8: tert—Butyl N-[dideuterio-[4-[hydroxyiminomethyl]phenyl]methyl]-N— (methyl)carbamate D D Hydroxylamine hydrochloride (51.15 mg, 0.7361 mmol) was added to a stirred solution of tert—butyl N—[dideuterio-[4-(dimethoxymethyl)phenyl]methyl]—N—methyl- ate (199 mg, 0.6692 mmol) in THF (10 mL)/water (1.000 mL) and the reaction d to stir at ambient ature for 4 hours. The on was partitioned between DCM and brine and the layers separated. The aqueous layer was extracted with DCM (x 2) and the combined organic extracts washed with brine (x 1), dried (MgSO4), filtered and concentrated in vacuo to give the sub—title compound as a white solid (180 mg, 100 % Yield); 1H NMR (400 MHz, DMSO) 5 1.41 (d, J = 24.6 Hz, 9H) 2.76 (s, 3H), 7.25 (d, J = 8.1 Hz, 2H), 7.58 (d, J = 8.0 Hz, 2H), 8.13 (s, 1H) and 11.20 (s, 1H) ppm; MS (ES+) 211.0 ).

Step 9: tert—ButylN-[[4-[ch10ro-N—hydroxy-carbonimidoyl]phenyl]-dideuterio-methyl]- N—methyl—carbamate HO\IN D D tert—Butyl N—[dideuterio-[4-[hydroxyiminomethyl]phenyl]methyl]-N- (methyl)carbamate (178 mg, 0.6683 mmol) in DMF (2 mL) was treated with NCS (89.24 mg, 0.6683 mmol) and the reaction warmed to 65 °C for 1 hour. The reaction was cooled to ambient temperature and diluted with water. The mixture was extracted with EtOAc (x 2) and the combined organic ts washed with brine (x 4), dried (MgSO4), filtered and concentrated in vacuo to give the sub—title compound as a white solid (188 mg, 94% Yield); 1H NMR (400 MHz, DMSO) 5 1.42 (d, J = 24.7 Hz, 9H), 2.78 (s, 3H), 7.32 (d, J = 8.4 Hz, 2H), 7.78 (d, J = 8.2 Hz, 2H) and 12.36 (s, 1H) ppm.

Step 10: tert—Butyl N-[[4-[5-[3-[bis(tert—butoxycarbonyl)amin0](4- pylsulfonylphenyl)pyrazinyl]isoxazol—3-yl]phenyl]-dideuterio-methyl]-N— methyl-carbamate O=S=O Et3N (36.31 mg, 50.01 uL, 0.3588 mmol) was added dropwise to a stirred solution of utyl N—tert-butoxycarbonyl—N—[3—ethynyl—5-(4- isopropylsulfonylphenyl)pyrazinyl]carbamate (150 mg, 0.2990 mmol) and tert—Butyl N- [[4-[chloro-N—hydroxy-carbonimidoyl]phenyl]-dideuterio-methyl]-N—methyl-carbamate (89.93 mg, 0.2990 mmol) in anhydrous THF (3 mL) and the reaction mixture heated at 65 °C for 3 hours. The reaction mixture was cooled to ambient temperature and diluted with brine. Water was added until the aqueous layer became clear and the layers were separated. The aqueous layer was extracted with EtOAc (x 1) and the combined organic extracts were washed with brine (x 1), dried (MgSO4), filtered and concentrated in vacuo.

The residue was purified by column chromatography (ISCO Companion, 40 g column, elueting with 0 to 30% EtOAc/Petroleum Ether, loaded in DCM) to give the sub—title product as a white solid (134 mg, 59% Yield); 1H NMR (400 MHz, DMSO) 5 1.22 (d, J = 6.8 Hz, 6H) 1.32 (s, 18H), 1.43 (d, J = 23.1 Hz, 9H), 2.82 (s, 3H), 3.56 (pent, 1H), 7.43 (d, J = 8.3 Hz, 3H), 8.02 - 8.03 (m 3H), 8.06 - 8.11 (m, 2H), 8.62 - 8.67 (m, 2H) and 9.51 (s, 1H) ppm; MS (ES+) 666.2 ).

Step 11: 4-[Dideuteri0(methylamin0)methyl]phenyl]isoxazol—S-yl](4- isopropylsulfonylphenyl)pyrazin-Z-amine (compound II-l) NH2 0’N \ HN\ N| \ D Oj:0 II-l 3M HCl in MeOH (1.167 mL of3 M, 3.500 mmol) was added to a stirred solution of tert—butyl N—[[4—[5—[3—[bis(tert-butoxycarbonyl)amino](4- isopropylsulfonylphenyl)pyrazinyl]isoxazolyl]phenyl]-dideuterio-methyl]-N—methyl- carbamate (134 mg, 0.1750 mmol) in DCM (5 mL) and the reaction heated at reflux for 16 hours. The reaction was cooled to t temperature and the resultant precipitate was isolated by filtration and dried under vacuum at 40 0C to give the di-HCl salt of the title compound as a yellow solid (58.8 mg, 62% Yield); 1H NMR (400 MHz, DMSO) 5 1.20 (d, J = 6.8 Hz, 6H), 2.60 (t, J = 5.4 Hz, 3H), 3.48 (hept, J = 6.8 Hz, 1H), 7.22 (br s, 2H), 7.69 — 7.75 (m, 2H), 7.85 (s, 1H), 7.92 — 7.99 (m, 2H), 8.08 — 8.15 (m, 2H) 8.37 — 8.42 (m, 2H), 8.97 (s, 1H) and 9.10 (d, J = 5.8 Hz, 2H) ppm; MS (ES+) 466.2.

D D \ O O \ I D .

O Y \,< Alkylation O Oxime Formation Chi orooxume —> \{/D Formation NH N N viii xiii XIV x D o ””2 OK HN+D O=<N D \ D N \ D 0WI D D [3+2] cycloadditon / N Deprotection —> —> o=3=o xv xvi "-2 Step 1: tert—Butyl N-[dideuterio-[4-(dimethoxymethyl)phenyl]methyl]-I- (trideuteriomethyl)carbamate LiHMDS (1M in THF) (1.181 mL of 1 M, 1.181 mmol) was added dropwise to a sittred solution of tert—butyl N—[dideuterio—[4-(dimethoxymethyl)phenyl]methyl]carbamate (300 mg, 1.059 mmol) in THF (5 mL) at -78 OC. The on was stirred at this temperature for 30 minutes then trideuterio(iodo)methane (198.0 mg, 84.98 uL, 1.366 mmol) was added dropwise and the mixture allowed to warm to t temperature over 21 hours. The reaction was again cooled to -78 OC and a further portion of LiHMDS (1M in THF) (635.4 uL of 1 M, 0.6354 mmol) was added. After 15 minutes more terio(iodo)methane (76.75 mg, 32.94 ”L, 0.5295 mmol) was added and the reaction allowed to warm to ambient ature over 5 hours. The mixture was diluted with EtOAc and the organic layer washed with ted aqueous NaHCO3 (x 2), brine (x 1), dried (MgSO4) filtered and concentrated in vacuo. The residue was purified by column chromatography (ISCO Companion, 24 g column, eluting with 0 to 30% EtOAc/Petroleum Ether, loaded in DCM) to give the sub-title product as a colourless oil (213 mg, 67% Yield); 1H NMR (400 MHz, DMSO) 5 1.36 - 1.42 (m, 9H) 3.22 (s, 6H), 5.35 (s, 1H), 7.21 (d, J = 7.8 Hz, 2H) and 7.35 (d, J = 7.7 Hz, 2H) ppm.

WO 49726 Step 2: utyl N-[dideuterio-[4-[hydroxyiminomethyl]phenyl]methyl]-N- (trideuteriomethyl)carbamate N O E? D ‘5 K Hydroxylamine hydrochloride (53.95 mg, 0.7763 mmol) was added to a stirred solution of tert—butyl N—[dideuterio-[4-(dimethoxymethyl)phenyl]methyl]-N— (trideuteriomethyl)carbamate (212 mg, 0.7057 mmol) in THF (10 mL)/water (1.000 mL) and the reaction allowed to stir at ambient temperature for 22 hours. The reaction was partitioned between DCM and brine and the layers separated. The aqueous layer was extracted with DCM (x 2) and the combined organic extracts washed with brine (x 1), dried ), filtered and concentrated in vacuo to give the sub—title compound as a white solid (190 mg, 100% Yield). ); 1H NMR (400 MHz, DMSO) 5 1.41 (d, J = 24.2 Hz, 9H) ), 7.25 (d, J = 8.1 Hz, 2H), 7.58 (d, J = 8.0 Hz, 2H), 8.13 (s, 1H) and 11.20 (s, 1H) ppm.

Step 3: tert-ButylN—[[4-[ch10ro-N—hydroxy-carbonimidoyl]phenyl]-dideuterio-methyl]- N—(trideuteriomethyl)carbamate l D D c. + N O . . r r tert—Butyl N-[dideuterio-[4-[hydroxyiminomethyl]phenyl]methyl]-N- (trideuteriomethyl)carbamate (190.0 mg, 0.7054 mmol) in DMF (2 mL) was treated with NCS (94.19 mg, 0.7054 mmol) and the reaction warmed to 65 °C for 1 hour. The reaction was cooled to ambient ature and diluted with water. The mixture was extracted with EtOAc (x 2) and the combined organic extracts washed with brine (x 4), dried (MgSO4), filtered and concentrated in vacuo to give the sub—title nd as a white solid (198 mg, 93% Yield); 1H NMR (400 MHz, DMSO) 5 1.41 (d, J = 26.0 Hz, 9H), 7.32 (d, J = 8.3 Hz, 2H), 7.78 (d, J = 8.2 Hz, 2H) and 12.36 (s, 1H) ppm.

Step 4: tert-Butyl N—[[4-[5-[3-[bis(tert-butoxycarb0nyl)amin0](4- isopropylsulfonylphenyl)pyrazinyl]isoxazol—3-yl]phenyl]-dideuterio-methyl]-N— (trideuteriomethyl)carbamate O=< D (Boc)2N O’N N+D \ D N \ D | D o=s=o Et3N (36.31 mg, 50.01 uL, 0.3588 mmol) was added dropwise to a stirred solution of utyl N—tert—butoxycarbonyl—N—[3—ethynyl—5-(4- isopropylsulfonylphenyl)pyrazinyl]carbamate (150 mg, 0.2990 mmol) and utyl N— [[4-[chloro-N—hydroxy-carbonimidoyl]phenyl]-dideuterio-methyl]-N— (trideuteriomethyl)carbamate (90.84 mg, 0.2990 mmol) in anhydrous THF (3 mL) and the reaction e heated at 65 0C for 3.5 hours. The reaction mixture was cooled to ambient temperature and diluted with EtOAc/brine. Water was added until the aqueous layer became clear and the layers were separated. The aqueous layer was extracted with EtOAc (x 1) and the combined c extracts were washed with brine (x 1), dried (MgSO4), filtered and concentrated in vacuo. The residue was purified by column chromatography (ISCO Companion, 40 g column, ng with 0 to 35% EtOAc/Petroleum Ether, loaded in DCM) to give the sub—title product as a white solid (158 mg, 69% Yield); 1H NMR (400 MHz, DMSO) 5 1.22 (d, J = 6.8 Hz, 6H) ), 1.44 (d, J = 22.0 Hz, 9H), 3.56 (dt, J = 13.5, 6.7 Hz, 2H), 7.43 (d, J = 8.2 Hz, 3H), 8.02 (d, J = 6.9 Hz, 2H), 8.08 (d, J = 8.7 Hz, 2H), 8.65 (d, J = 8.8 Hz, 2H) and 9.51 (s, 1H) ppm; MS (ES+) 669.3 (M—Boc).

Step 5: 3-[3-[4-[dideuterio-(trideuteriomethylamino)methyl]phenyl]isoxazol—5—yl](4- isopr0pylsulfonylphenyl)pyrazin-Z-amine (compound 11-2) NH2 O’N HN\< D \ D N \ D I D O=S=O II-2 3M HCl in MeOH (1.361 mL of3 M, 4.084 mmol) was added to a stirred solution of tert—butyl N—[[4—[5—[3—[bis(tert-butoxycarbonyl)amino](4- pylsulfonylphenyl)pyrazinyl]isoxazolyl]phenyl]-dideuterio-methyl]—N— (trideuteriomethyl)carbamate (157 mg, 0.2042 mmol) in DCM (5 mL) and the reaction heated at reflux for 16 hours. The reaction was cooled to ambient temperature and the resultant precipitate was isolated by filtration and dried under vacuum at 40 0C to give the di-HCl salt of the title compound as a yellow solid (72.5 mg, 66% Yield); 1H NMR (400 MHz, DMSO) 1.20 (d, J = 6.8 Hz, 6H), 3.48 (dq, J = 13.6, 6.7 Hz, 1H), 7.21 (s, 2H), 7.68 — 7.78 (m, 2H), 7.85 (s, 1H), 7.91 — 7.99 (m, 2H), 8.08 — 8.13 (m, 2H), 8.36 — 8.42 (m, 2H), 8.96 (s, 1H) and 9.14 (s, 2H) ppm; MS (ES+) 469.1.

Exam 1e 4: S nthesis of 5- 4-is0 r0 n l hen 1 3- 4- trideuteriometh lamino meth l hen lisoxazol—S- l razin-Z-amine 1C0mpound 11-31 0 O O D D D BOC \o \o . \0 Protection JK(>\/H A'kY'at'O” 7i: Reduction 0 ’ NH2 NTOK 7r 7< xvii xviii xix O HON D D N HO/\©\Dj’D I D\|,D | D\|D/D NTO\|< Oxidation NTO Oxime Formation N O Egggggme o O )< r 7< xx xxi xxii O HN+D HO‘N 0% \\ D \ D D (BOC)2NWmflON\ I D D ”I CI \1/ D NTO\|< [3+2]cycloadditon /N Deprotection xxiii xxiv A 11—3 Step 1: Methyl 4-[(tert—butoxycarbonylamin0)methyl]benzoate xv111 Et3N (1.882 g, 2.592 mL, 18.60 mmol) was added to a stirred suspension of methyl 4-(aminomethyl)benzoate (Hydrochloric Acid (1)) (1.5 g, 7.439 mmol) in THF (20 mL) at 0 °C. The reaction was d to stir at this ature for 30 minutes then BoczO (1.705 g, 1.795 mL, 7.811 mmol) was added in portions. The reaction was allowed to warm to ambient temperature and stirred for 18 hours. The mixture was diluted with EtOAc. The organic layer was washed with 1M aqueous HCl (x 2), saturated aqueous NaHC03 (x 2) and brine (x 1). The organic layer was dried (MgSO4), filtered and concentrated in vacuo to give the sub—title compound as a white solid that was used without further purification (1.93 g, 98% Yield); 1H NMR (400 MHz, DMSO) 5 1.40 (s, 9H), 3.85 (s, 3H), 4.20 (d, J = 6.1 Hz, 2H), 7.38 (d, J = 8.2 Hz, 2H), 7.49 (t, J = 6.1 Hz, 1H) and 7.92 (d, J = 8.2 Hz, 2H) ppm; MS (ES+) 251.1 (M—Me).

Step 2: Methyl 4-[[tert-butoxycarbonyl(trideuteri0methyl)amino]methyl]benzoate LiHMDS (1M in THF) (8.112 mL of 1 M, 8.112 mmol) was added se to a stirred solution of methyl 4-[(tert—butoxycarbonylamino)methyl]benzoate (1.93 g, 7.275 mmol) in THF (10 mL) at -78 °C. The solution was stirred at this temperature for 30 s then trideuterio(iodo)methane (1.360 g, 9.385 mmol) was added dropwise and the mixture allowed to warm to ambient temperature over 3 hours. The reaction was cooled to —78 °C and a further portion of LiHMDS (1M in THF) (2.182 mL of 1 M, 2.182 mmol) was added. After minutes a further portion of trideuterio(iodo)methane (527.4 mg, 3.638 mmol) was added and the reaction d to warm to ambient temperature over 17 hours. The mixture was diluted with EtOAc and the organic layer washed with saturated aqueous NaHC03 (x 2), brine (x 1), dried (MgSO4) d and concentrated in vacuo. The residue was purified by column chromatography (ISCO Companion, 120 g , eluting with 0 to 30% EtOAc/Petroleum Ether, loaded in DCM) to give the sub-title product as a pale yellow oil (1.37 g, 67% Yield); 1H NMR (400 MHz, DMSO) 5 1.38 (d, J = 44.2 Hz, 9H), 3.83 (s, 3H), 4.43 (s, 2H), 7.33 (d, J = 8.2 Hz, 2H) and 7.94 (d, J = 8.1 Hz, 2H) ppm; MS (ES+) 268.1 (M— Step 3: tert—Butyl N—[[4-(hydr0xymethyl)phenyl]methyl]-N- (trideuteriomethyl)carbamate LiBH4 (158.5 mg, 7.278 mmol) was added to a stirred solution of methyl 4— —butoxycarbonyl(trideuteriomethyl)amino]methyl]benzoate (1.37 g, 4.852 mmol) in THF (10 mL) and the reaction warmed to 85 0C for 15 hours. A further portion of LiBH4 (158.5 mg, 7.278 mmol) was added and the reaction stirred at 65 °C for a further 7 hours. The reaction mixture was cooled to ambient ature then poured onto crushed ice and whilst stirring, 1M HCl was added dropwise until no effervescence was ed. The mixture was stirred for 10 minutes then saturated aqueous NaHCO3 was added until the mixture was at pH 8. The aqueous layer was extracted with EtOAc (x 3) and the combined organic extracts dried (MgSO4), filtered and concentrated in vacuo. The e was purified by column chromatography (ISCO Companion, 120 g column, elueting with 0 to 100% EtOAc/Petroleum Ether, loaded in DCM) to give the sub—title product as a colourless oil (1.03 g, 84% Yield); 1H NMR (400 MHz, DMSO) 5 1.42 (d, J = 14.6 Hz, 9H), 4.35 (s, 2H), 4.48 (d, J = 5.7 Hz, 2H), 5.15 (t, J = 5.7 Hz, 1H), 7.18 (d, J = 7.9 Hz, 2H) and 7.30 (d, J = 7.7 Hz, 2H) ppm; MS (ES+) 181.1 (M-OtBu).

Step 4: tert—Butyl N-[(4-f0rmylphenyl)methyl]-N—(trideuteriomethyl)carbamate Em.)DD D O \i< MnOz (5.281 g, 1.051 mL, 60.75 mmol) was added to a stirred solution of tert— butyl N—[[4-(hydroxymethyl)phenyl]methyl]-N—(trideuteriomethyl)carbamate (1.03 g, 4.050 mmol) in DCM (10 mL) and the reaction stirred at ambient temperature for 20 hours. The reaction was filtered through a pad of Celite and washed with DCM. The filtrate was concentrated in vacuo to give the sub—title compound as a colourless oil (891 mg, 88% Yield); 1H NMR (400 MHz, DMSO) 5 1.40 (d, J = 43.4 Hz, 9H), 4.48 (s, 2H), 7.43 (d, J = 8.0 Hz, 2H), 7.91 (d, J = 7.9 Hz, 2H) and 10.00 (s, 1H), ppm.

Step 5: tert-butyl N-[[4-[hydroxyiminomethyl]phenyl]methyl]-N- (trideuteriomethyl)carbamate XXii Hydroxylamine (466.0 uL of 50 %w/V, 7.054 mmol) was added to a stirred solution of tert-butyl formylphenyl)methyl]-N—(trideuteriomethyl)carbamate (890 mg, 3.527 mmol) in ethanol (5 mL) and the reaction e stirred at ambient temperature for 45 minutes. The reaction mixture was concentrated in vacuo and the residue taken up in water and extracted with EtOAc (x 3). The combined organic extracts were washed with brine (x 1), dried (MgSO4), filtered and concentrated in vacuo. The residue was ated from petroleum ether and the precipitate isolated by ion to give the sub-title product as a white solid (837 mg, 89% Yield); 1H NMR (400 MHz, DMSO) 5 1.41 (d, J = 25.8 Hz, 9H), 4.38 (s, 2H), 7.24 (d, J = 8.0 Hz, 2H), 7.58 (d, J = 8.0 Hz, 2H), 8.13 (s, 1H) and 11.20 (s, 1H) ppm; MS (ES+) 212.0 (M—tBu).

Step 6: tert—ButylN—[[4-[ch10ro-N—hydroxy-carbonimidoyl]phenyl]methyl]-N— (trideuteriomethyl)carbamate xxiii tert—butyl N-[[4-[hydroxyiminomethyl]phenyl]methyl]-N— (trideuteriomethyl)carbamate (250 mg, 0.9351 mmol) in DMF (2.5 mL) was treated with NCS (124.9 mg, 0.9351 mmol) and the reaction warmed to 65 0C for 1 hour. The on was cooled to ambient temperature and diluted with water. The mixture was extracted with EtOAc (x 2) and the ed organic extracts washed with brine (x 4), dried (MgSO4), filtered and concentrated in vacuo to give the sub—title nd as a white solid (259 mg, 92% Yield); 1H NMR (400 MHz, DMSO) 5 1.41 (d, J = 29.6 Hz, 9H), 4.42 (s, 2H), 7.31 (d, J = 8.3 Hz, 2H), 7.78 (d, J = 8.0 Hz, 2H), and12.38 (s, 1H), ppm.

Step 7: tert-Butyl N—[[4-[5-[3-[bis(tert-butoxycarb0nyl)amin0](4- isopropylsulfonylphenyl)pyrazinyl]isoxazol—3-yl]phenyl]methyl]-N- (trideuteriomethyl)carbamate O=S=O xxiV Et3N (48.41 mg, 66.68 uL, 0.4784 mmol) was added dropwise to a stirred solution of tert—butyl N—tert—butoxycarbonyl—N—[3—ethynyl—5 —(4— isopropylsulfonylphenyl)pyrazinyl]carbamate (200 mg, 0.3987 mmol) and tert—butyl N— [[4-[chloro-N—hydroxy-carbonimidoyl]phenyl]methyl]-N—(trideuteriomethyl)carbamate (120.3 mg, 0.3987 mmol) in anhydrous THF (5 mL) and the reaction mixture heated at 65 0C for 2.5 hours. The reaction mixture was cooled to ambient temperature and diluted with EtOAc/brine. Water was added until the aqueous layer became clear and the layers were separated. The aqueous layer was extracted with EtOAc (x 1) and the combined organic ts were washed with brine (x 1), dried (MgSO4), filtered and concentrated in vacuo.

The residue was d by column chromatography (ISCO Companion, 40 g column, elueting with 0 to 20% EtOAc/Petroleum Ether, loaded in DCM) to give the sub—title product as a white solid (213.5 mg, 70% Yield); 1H NMR (400 MHz, DMSO) 5 1.22 (d, J = 6.8 Hz, 6H), 1.31 (s, 18H), 1.43 (d, J = 26.2 Hz, 9H), 3.51 = - 3.60 (m, 1H), 4.47 (s, 2H), 7.42 (d, J 8.1 Hz, 2H), 8.03 (d, J = 5.2 Hz, 3H), 8.08 (d, J = 8.6 Hz, 2H), 8.65 (d, J = 8.6 Hz, 2H) and 9.52 (s, 1H) ppm; MS (ES+) 667.4 (M-Boc).

Step 8: 5—(4—Is0pr0pylsulf0nylphenyl)[3-[4-[(trideuteriomethylamino)methyl] phenyl]isoxazol—S-yl]pyrazin-Z-amine (Compound 11-3) WO 49726 NHZO’N\ HN {D N\ D o=s=o II-3 3M HCl in MeOH (1.5 mL of 3 M, 4.500 mmol) was added to a stirred solution of tert—butyl N—[[4—[5—[3—[bis(tert-butoxycarbonyl)amino](4- isopropylsulfonylphenyl)pyrazinyl]isoxazolyl]phenyl]methyl]—N— (trideuteriomethyl)carbamate (213 mg, 0.2777 mmol) in DCM (6 mL) and the reaction heated at reflux for 15 hours. A further portion of 3M HCl in MeOH (0.5 mL of 3 M, 1.500 mmol) was added and the reaction heated at reflux for a further 7 hours. The reaction was cooled to ambient temperature and the resultant precipitate was isolated by filtration and dried under vacuum at 40 °C to give the di—HCl salt of the title nd as a yellow solid (97.6 mg, 65% Yield); 1H NMR (400 MHz, DMSO) 5 1.20 (d, J = 6.8 Hz, 6H), 3.47 (tt, J = 14.0, 6.9 Hz, 1H), 4.19 — 4.25 (m, 2H), 7.23 (s, 2H), 7.72 (d, J = 8.4 Hz, 2H), 7.85 (s, 1H), 7.95 (d, J = 8.7 Hz, 2H), 8.11 (d, J = 8.4 Hz, 2H), 8.39 (d, J = 8.7 Hz, 2H), 8.97 (s, 1H) and 9.11 (s, 2H) ppm; MS (ES+) 467.2. tetradeuterio-l- trideuteriometh leth lsulfon l hen l razin-Z-amine Com ound L \o Reductive /\0 B00 Amination )\©\/ OXime Formation 21 Protection AOW? ,0 —> \ I \i/ o o —N o=< OYO Y (BOC Ch'°r°°x'me C' )2 N o N\ [3+2] cycloadditon \\ Formation N —> \ N \ xxviii xxix Br xxx Br Br Br W Alkylation s ion o=s=o 30mm? Suzuki grim, —> Formation SH —> D D DWD Q 02520 D D D xxxi xxxii xxxiii xxxiv 0=<o NH2 o—Q HN‘ ()Boc2\0—N\N\ N|\ Deprotection xxxv 11—4 :W: D D D D D XXVI 2M methylamine in MeOH (288.1 mL, 576.2 mmol) was diluted with methanol (1.000 L) and stirred at ~20 °C. 4-(Diethoxymethyl)benzaldehyde (100 g, 480.2 mmol) was added dropwise over 1 minute and the reaction stirred at ambient temperature for 1.25 hours.

Sodium borohydride (29.07 g, 30.76 mL, 768.3 mmol) was added portionwise over 20 minutes while maintaining the ature between 20 and 30 0C with an ice-water bath. The reaction on was stirred at t temperature overnight then quenched by the dropwise addition ofNaOH (960.4 mL of 1.0 M, 960.4 mmol) over 20 minutes. The reaction was stirred for 30 minutes and concentrated in vacuo to remove MeOH. The reaction was partitioned with MTBE (1.200 L) and the phases separated. The organic phase was washed with water (3 00.0 mL), dried (Na2S04), and trated in vacuo to give the title compound as a yellow oil (102.9 g, 96% Yield); 1H NMR (400 MHz, CDCl3) 5 1.25 (t, 6H), 2.46 (s, 3H), 3.45 — 3.65 (m, 4H), 3.75 (s, 2H), 5.51 (s, 1H), 7.32 (d, 2H) and 7.44 (d, 2H) ppm.

Step 2: tert—Butyl N—[[4-(dieth0xymethyl)phenyl]methyl]-N—methyl-carbamate LO \p ”coco”N\ xxvn A l-L glass-j acketed reactor was fitted with an overhead stirrer, thermocouple, and chiller. A solution of 1-[4-(diethoxymethyl)phenyl]-N—methyl-methanamine (80.0 g, 358.2 mmol) in DCM (480.0 mL) was d at 18 °C. A solution of Boc ide (79.75 g, 83.95 mL, 365.4 mmol) in DCM (160.0 mL) was added over 10 minutes and the solution was stirred at 20 - 25 OC overnight. The reaction mixture was filtered, rinsed with DCM (3 x 50 mL) and the filtrate concentrated in vacuo to afford give the title compound as a pale yellow liquid (116.6 g, quantitative yield); 1H NMR (400 MHz, CDCl3) 5 1.25 (t, 6H), 1.49 — 1.54 (2 x s, 9H), 2.78 — 2.83 (2 x s, 3H), 3.50 — 3.66 (m, 4H), 4.42 (s, 2H), 5.49 (s, 1H), 7.22 (d, 2H) and 7.45 (d, 2H) ppm.

Step 3: tert—Butyl [hydroxyiminomethyl]phenyl]methyl]-N—methyl-carbamate XXViii A biphasic solution of tert—butyl N—[[4—(diethoxymethyl)phenyl]methyl]—N— -carbamate (50.0 g, 154.6 mmol) in 2-MeTHF (400.0 mL) and NaZSO4 (100.0 mL of %w/v, 70.40 mmol) was stirred at 8 — 10 °C in a 1—L, glass—jacketed reactor.

Hydroxylamine hydrochloride (46.38 mL of 5.0 M, 231.9 mmol) was added and the biphasic solution was stirred at 30 °C for 16 hours. The reaction was diluted with MTBE (200.0 mL) and the layers separated. The organic phase was washed with water (200.0 mL), dried (NaZSO4), filtered and concentrated in vacuo. The e was diluted with heptane (200.0 mL) and the resultant suspension was stirred at ambient temperature for 30 minutes. The solid was collected by filtration to give the title compound as a white solid (36.5 g, 89% Yield); 1H NMR (400 MHz, CDC13) 5 1.50 (s, 9H), 2.88 (br s, 3H), 4.60 (s, 2H), 7.26 (d, 2H), 7.52 (d, 2H) and 8.15 (s, 1H) ppm.

Step 4: tert—ButylN—[[4-[ch10ro-N—hydroxy-carbonimidoyl]phenyl]methyl]-N—methyl— XXiX A sion of tert—butyl [hydroxyiminomethyl]phenyl]methyl]—N— methyl-carbamate (100 g, 378.3 mmol) in isopropyl acetate (1.000 L) was stirred at ambient temperature. rosuccinimide (53.04 g, 397.2 mmol) was added and stirred at ambient temperature for 16 hours. The reaction was partitioned with water (500.0 mL) and the phases separated. The organic phase was washed with water (500.0 mL) (2 X), dried (NaZSO4), filtered and concentrated in vacuo to remove most of the solvent. Heptane (1.000 L) was added and the mixture concentrated in vacuo to remove most of the solvent. Heptane (1.000 L) was added and the ant precipitate isolated by filtration. The filter-cake was washed with heptane (500 mL) and air-dried to give the title compound as an off—white powder (105.45 g, 93% Yield); 1H NMR (400 MHz, CDC13) 5 1.48 (2 x s, 9H), 2.90 (2 x s, 3H), 4.47 (s, 2H), 7.26 (d, 2H), 7.77 (d, 2H) and 8.82 (s, 1H) ppm.

Step 5: tert—Butyl N—[[4-[5-[3-[bis(tert-but0xycarbonyl)amino]-6—br0mo-pyrazin xazol—3-yl]phenyl]methyl]-N-methyl-carbamate A suspension of tert—butyl N—[[4—[chloro—N—hydroxy— carbonimidoyl]phenyl]methyl]-N—methyl-carbamate (100.0 g, 334.7 mmol) and tert—butyl N— tert—butoxycarbonyl-N—[3-ethynyl(4-isopropylsulfonylphenyl)pyrazinyl]carbamate (121.2 g, 304.3 mmol) in DCM (1.212 L) was stirred at ambient temperature. Triethylamine (33.87 g, 46.65 mL, 334.7 mmol) was added in one portion and the reaction stirred at ambient temperature for 16 hours. The reaction was partitioned with water (606.0 mL) and the phases separated. The organic phase was washed with water (606.0 mL), dried (NaZSO4), filtered and concentrated in vacuo to near dryness. e (3 63.6 mL) was added and the mixture concentrated to about 300 mL. Further heptane (1.212 L) was added and the mixture heated to 90 0C with stirring. The mixture was slowly cooled to ambient temperature and stirred at this temperature for 1 hour. The resultant precipitate was isolated by filtration and the filter- cake washed with heptane (2 x 363.6 mL) and air—dried to give the title compound as a beige solid (181.8 g, 90% Yield); 1H NMR (400 MHz, CDC13) 5 1.41 (s, 18H), 1.51 (s, 9H), 2.88 (2 x s, 3H), 4.50 (s, 2H), 7.36 — 7.38 (m, 3H), 7.86 (d, 2H) and 8.65 (s, 1H) ppm.

Step 6: 1-Bromo-4—[1,2,2,2-tetradeuteri0(trideuteriomethyl)ethyl]sulfanyl—benzene XXXii ] Sodium e (246.5 mg, 6.163 mmol) was added portionwise to a stirred on of 4—bromobenzenethiol (compound xxxi) (970.9 mg, 5.135 mmol) in DMF (10 mL) at 0 °C. After stirring at this temperature for 15 minutes 1,1,1,2,3,3,3-heptadeuterio—2—iodo— propane (1 g, 5.649 mmol) was added and the reaction allowed to warm to ambient ature over 18 hours. The reaction was quenched by the addition of water and the mixture stirred for 10 minutes. The mixture was extracted with diethyl ether (x 3) and the combined organic extracts washed with water (x 2), brine (x 2), dried (MgSO4), d and concentrated in vacuo to give the sub—title compound that was used directly without further purification assuming 100% Yield and purity; 1H NMR (500 MHz, DMSO) 5 7.25 — 7.37 (m, 2H) and 7.48 - 7.55 (m, 2H) ppm. 2012/058127 Step 7: 1-Bromo-4—[1,2,2,2-tetradeuteri0(trideuteriomethyl)ethyl]sulfonyl—benzene XXXiii mCPBA (2.875 g, 12.83 mmol) was added in portions to a stirred solution of 1- 4-[1,2,2,2-tetradeuterio(trideuteriomethyl)ethyl]sulfanyl-benzene (1.223 g, 5.134 mmol) in DCM (20 mL) at 0 OC and the reaction allowed to warm to ambient temperature over 17 hours. The mixture was washed 1M aqueous NaOH (X 2), saturated aqueous Na2S203 (x 3), brine (x 1), dried (MgSO4), filtered and concentrated in vacuo. The residue was purified by column chromatography (ISCO Companion, 80 g , eluting with 0 to 40% EtOAc/Petroleum Ether, loaded in DCM) to give the sub—title nd as a colourless oil (1.19 g, 86% Yield); 1H NMR (500 MHz, DMSO) 5 7.77 — 7.81 (m, 2H) and 7.88 — 7.92 (m, 2H) ppm.

Step 8: 4,4,5,5-Tetramethyl—2-[4-[1,2,2,2-tetradeuteri0(trideuteriomethyl)ethyl] sulfonylphenyl]-1,3,2-dioxab0rolane XXXiV Pd(dppf)Clz.DCM (179.8 mg, 0.2202 mmol) was added to a stirred suspension of 1-bromo[1,2,2,2-tetradeuterio(trideuteriomethyl)ethyl]sulfonyl-benzene (1. 19 g, 4.404 mmol), pinacolato)diboron (1.342 g, 5.285 mmol) and KOAc (1.296 g, 13.21 mmol) in dioxane (10 mL). The reaction placed under an atmosphere of nitrogen via 5 x nitrogen/vacuum cycles and the mixture was heated at 80 °C for 4.5 hours. The reaction was cooled to ambient temperature and the solvent removed in vacuo. The residue was partitioned between EtZO and water and the layers separated. The organic layer was dried (MgSO4), filtered and concentrated in vacuo. The residue was dissolved in 30% EtOAc/Petroleum ether (35 mL) and 1.2 g of Florosil was added. The mixture was stirred for 30 minutes then filtered, washing the solids with further alliquots of 30% EtOAc/Petrol (x 3). The filtrate was concentrated in vacuo and tritruated from 10% EtOAc/petroleum ether. The resultant solid was isolated by filtration, washed with petroleum ether and dried in vacuo to give the sub— title nd as an off-white solid (1052.1 mg, 75% Yield); 1H NMR (400 MHz, DMSO) 5 1.33 (s, 12H), 7.87 (d, J = 8.4 Hz, 2H) and 7.94 (d, J = 8.4 Hz, 2H) ppm.

Step 9: utyl N—[[4-[5-[3-[bis(tert-butoxycarbonyl)amino][4-[1,2,2,2- tetradeuterio-l-(trideuteriomethyl)ethyl]sulfonylphenyl]pyrazin-Z-yl]isoxazol yl]phenyl]methyl]-N—methyl-carbamate N 0% (BOC)2N 0' \ N\ XXXV [1,1’-Bis(di—tert—butylphosphino)ferrocene]dichloropalladium(H) (106.8 mg, 0.1639 mmol) was added to a e of 4,4,5,5-tetramethyl[4-[1,2,2,2-tetradeuterio (trideuteriomethyl)ethyl]sulfonylphenyl]—1,3,2-dioxaborolane (1.3 g, 4.098 mmol), tert-butyl N—[[4—[5—[3—[bis(tert—butoxycarbonyl)amino]bromo-pyrazinyl]isoxazol yl]phenyl]methyl]-N-methyl-carbamate (2.707 g, 4.098 mmol) and K2CO3 (1.133 g, 8.200 mmol) in toluene (9.100 mL), EtOH (2.600 mL) and water (2.600 mL) and the reaction mixture was ed with a flow of nitrogen (5 cycles).

The mixture was heated at 75 °C for 1.5 hours. The reaction was ccoled to ambient temperature and water (5.2 mL) was added. After stirring the layers were separated and the organic layer dried (NaZSO4), filtered, and trated in vacuo. The e was triturated with IPA and the resultant precipitate isolated by filtration, washed with IPA (3 x 4 mL) and dried in vacuo at 50 °C to give the title compound as a white solid (2.4 g, 76% Yield); 1H NMR (400 MHz, CDCl3) 5 1.41 (s, 18H), 1.50 (s, 9H), 2.85 — 2.89 (m, 3H), 4.50 (s, 2H), 7.36 — 7.38 (m, 3H), 7.87 (d, 2H), 8.09 (d, 2H), 8.35 (d, 2H) and 9.06 (s, 1H) ppm.

Step 10: 3-[3-[4-(Methylaminomethyl)phenyl]isoxazol—S-yl][4—[1,2,2,2-tetradeuterio— 1-(trideuteriomethyl)ethyl]sulfonylphenyl]pyrazin-Z-amine (compound II-4) NH2 O’N\ HN‘ II-4 Concentrated HCl (3.375 g, 2.812 mL of 37 %w/w, 34.25 mmol) was added to a solution of tert—butyl N—[[4—[5—[3—[bis(tert-butoxycarbonyl)amino][4-[1,2,2,2-tetradeuterio- 1-(trideuteriomethyl)ethyl] sulfonylphenyl]pyrazinyl] ol-3 -yl]phenyl]methyl] -N— methyl-carbamate (2.2 g, 2.854 mmol) in acetone (28.60 mL) and the reaction heated at reflux for 7 hours. The reaction was cooled to t temperature and the resultant precipitate isolated by filtration, washed with acetone (2 X 4.5 mL) and dried in vacuo at 50 0C to give the di-HCl salt of the title compound as a yellow solid (1.42 g, 92% Yield); 1H NMR (400 MHz, DMSO) 5 2.58 (t, 3H), 4.21 (t, 2H), 5.67 (br s, 2H), 7.74 (d, 2H), 7.85 (s, 1H), 7.94 (d, 2H), 8.10 (d, 2H), 8.38 (d, 2H), 8.96 (s, 1H) and 9.33 (br s, 2H) ppm; MS (ES+) 471.8. meth lamino meth l hen lisoxazol—S- l razin-Z-amine Com ound I-2 HO\|N Step 1 HOxN Step 2 Boo Ncs, IPAc CI Boo TEA, DCM I —> I —> N\ N\ N(Boc)2 4-I 4-ll 4-III Step 3 B I800 800 om” O’N [Boo B OH( )2 \ \N\ Boo Boo \ \N O,N\ \N\ \ |'P O Sr NI \ 2 /N KKN| —> 4-iv 1. cat. pf)C|2, PhCH3, 5-i Br aQ- K2C03 2. IPA crystallization SOZiPr NH NH 2 O’N O’N \ 2 \ HN \ HN\ Step 4 \ Step 5 \ \ \ conc. HCI NI NI acetone / N 4:1 IPA/water -2HC| / N cHCI —> —> SOZiPr SOZIPF Compound I-2°2HC1 nd I-2~HC1 Step 1: Preparation of Compound 4-ii HoxlN Step 1 HO\|N EEOC NCS, IPAc N\ 02mg“;N\ 4-i 4-ii A suspension of tert—butyl 4-((hydroxyimino)methyl)benzyl (methy1)carbamate (Compound 4-i) (650 g, 2.46 mol) in isopropyl acetate (6.5 L) is stirred at ambient ature. N—Chlorosuccinimide (361 g, 2.71 mol) is added and the reaction temperature maintained overnight at 20—28 0C to ensure complete reaction. The reaction mixture is diluted with water (3.25 L) and EtOAc (1.3 L) and the phases are separated. The organic phase is washed with water (2 x 3.25 L), dried (NaZSO4), and concentrated to a ke. The trate is d with heptane (9.1 L), ~2 L of solvent removed, and then stirred at ambient temperature for 2—20 h. The solid is collected by filtration. The filter-cake is washed with heptane (2 x 975 mL) and dried to afford Compound 4—ii (692 g; 94% yield, 99.2 area % purity by HPLC) as a colorless powder.

Ste 2: Pre aration of tert—but l 5—bromo—3— 3— 4— tert—butox carbon 1 meth lamino meth l hen lisoxazol l razin l tert—butox carbon lcarbamate Com ound 4-iv .N —» M N N(Boc) Cl B00 5 NW '{K N \ I l /N 4-ii .

Br 4-lll 4-IV A suspension of Iert—butyl N—(5 —bromo—3—ethj,lnj,rlpyrazin—2—yl)—N—tert— butoxycarbonylcarbamate (Compound (1.59 kg, 3.99 mol) and tert—butyl 4— (chloro(hydroxyimino)methyl)benzyl(tetrahydro-2H—pyranyl)carbamate (1.31 kg, 4.39 mol; 1.10 equiv.) in CHzClz (12.7 L) is stirred at ambient temperature. ylamine (444 g, 611 mL, 4.39 mol) is added to the suspension and the reaction temperature is maintained between 20—30 °C for 20—48 h to ensure te reaction. The reaction mixture is diluted with water (8 L) and thoroughly mixed, then the phases are separated. The organic phase is washed with water (8 L), dried (NaZSO4), and then concentrated until about 1 L of CHzClz remains. The concentrate is diluted with heptane (3.2 L) and re-concentrated at 40 oC/200 torr until no distillate is observed. The concentrate is stirred and further d with heptane (12.7 L) to precipitate a solid. The suspension is stirred overnight. The solid is collected by filtration, washed with heptane (2 x 3 L) then dried to afford Compound 4-iv (2.42 kg; 92% yield, 100 area % purity by HPLC) as a light tan powder. 1H NMR (400 MHz, CDCl3) 5 8.61 (s, 1H), 7.82 (d, J: 8.2 Hz, 2H), 7.31 (m, 3H), 4.46 (br s, 2H), 2.84 (br d, 3H), 1.57 (s, 2H), 1.44 (br s, 9H), 1.36 (s, 18H).

WO 49726 Ste 3: Pre aration of Com ound 5—i B(OH>2 o"‘{ °°‘N\ Boc Boc \ / Boc \ N O’N \ \ N\ iPrOZS NI \ \ “N I —> N 5". / 1. cat. Pd(dtbpf)C|2, PhCH3, Br aq. K2003 2. i-PrOH crystallization SOZiPr ] A mixture of tert—butyl mo—3—(3—(4—(((tert— butoxycarbonyl)(methyl)amino)methyl)phenyl)isoxazolyl)pyrazinyl)(tertbutoxycarbonyl )carbamate (Compound 4-iv )(1.00 kg, 1.51 mol), K2CO3 (419 g, 3.02 mol), and (4-(isopropylsulfonyl)phenyl)boronic acid (345 g, 1.51 mol) in toluene (7.0 L) and water (2.0 L) was stirred and degassed with N2 for 30 min. 1,1’—bis(di—t—butylphosphino)ferrocen— dichloro—palladium(H) [Pd(dtbpf)Clz; 19.7 g, 30.3 mmol] was then added and degassed an additional 20 min. The reaction mixture was warmed at 70 0C for at least 1 h to ensure complete reaction. The reaction e was cooled to ambient ature then filtered h Celite. The reaction vessel and filter pad are rinsed with toluene (2 x 700 mL). The filtrates are combined and the phases are separated. The organic phase is stirred with Biotage MP-TMT resin (170 g) for 4—20 h. The resin is removed by filtration through Celite and the filter pad is washed with toluene (2 x 700 mL). The filtrate and gs are combined and concentrated to near dryness then diluted with i-PrOH (5.75 L) and re-concentrated. The concentrate is again dissolved in warm (45 oC) i-PrOH (5.75 L) and then cooled to ambient temperature with stirring to induce crystallization then stirred for around 16 — 20 h. The solid is collected by filtration, washed with i-PrOH (2 x 1 L), and dried to afford VRT-1018729 (967 g; 84%) as a beige powder. 1H NMR (400 MHz, CDCl3) 5 9.04 (s, 1H), 8.33 (d, J= 8.6 Hz, 2H), 8.06 (d, J: 8.5 Hz, 2H), 7.85 (d, J: 8.1 Hz, 2H), 7.34 (m, 3H), 4.47 (br s, 2H), 3.25 (hept, J: 7.0 Hz, 1H), 2.85 (br d, 3H), 1.47 (s, 9H), 1.38 (s, 18H), 1.33 (d, J: 6.9 Hz, 6H).

Step 4: Preparation of Compound I-2 0 2HCl A solution of Compound 5-i (950 g, 1.24 mol) in acetone (12.35 L) is warmed to 40 0C then concentrated HCl (1.23 kg, 1.02 L of37 %w/w, 12.4 mol) is added at a rate to maintain the reaction temperature between 40 — 45 °C for at least 5 h to ensure complete reaction. The suspension is cooled to below 30 °C and the solid collected by filtration. The filter-cake is washed with acetone (2 x 950.0 mL) then dried to afford Compound 1-2 ° 2HCl (578 g; 87% yield, 99.5 area % purity by HPLC) as a yellow powder. 1H NMR (400 MHz, DMSO) 5 9.53 (br d, J= 4.8 Hz, 2H), 8.93 (s, 1H), 8.37 (d, J= 8.5 Hz, 2H), 8.07 (d, J= 8.3 Hz, 2H), 7.92 (d, J= 8.6 Hz, 2H), 7.84 (s, 1H), 7.75 (d, J: 8.3 Hz, 2H), 4.23 — 4.15 (m, 2H), 3.43 (hept, J= 6.8 Hz, 1H), 2.55 (t, J= 5.3 Hz, 3H), 1.17 (d, J= 6.8 Hz, 6H).

Ste 5: Pre aration of Com ound I-2 0 HCl from Com ound I-2 0 2HCl NH2 0’N\ HN\ NH2 0’N\ HN\ \ \ | . NI \ 4.1- / N I PrOH/water- / N . HCI 02HC| SOZiPr SOZiPr Compound l-2 - 2HC| nd l-2 - HCI Two-potprocess A stirred sion of Compound 1—2 ° 2HCl (874 g, 1.63 mol) in i-PrOH (3.50 L) and water (0.87 L) is warmed at 50 0C for 1—2 h, cooled to ambient temperature, and stirred for 1—20 h. XRPD is performed on a small sample to ensure that Compound 1-2 ° 2HCl has been converted to another form. The suspension is cooled to 5 °C and stirred for 1 h. The solid is collected by filtration then the filter-cake is washed with 80/20 i-PrOH/water (2 x 874 mL), and briefly dried.

IfXRPD shows the Compound 1-2 ° HCl/anhydrate form, the solid is dried to afford Compound 1-2 °HCl/anhydrate (836 g, 99% yield, 99.2 area % purity by HPLC) as a yellow solid. 1H NMR (400 MHz, DMSO) 5 9.38 (s, 2H), 8.96 (s, 1H), 8.46 — 8.34 (m, 2H), 8.10 (d, J= 8.3 Hz, 2H), 7.94 (d, J= 8.6 Hz, 2H), 7.85 (s, 1H), 7.75 (d, J= 8.3 Hz, 2H), 7.23 (br s, 2H), 4.21 (s, 2H), 3.47 (hept, J= 6.7 Hz, 1H), 2.58 (s, 3H), 1.19 (d, J= 6.8 Hz, 6H).

IfXRPD shows the Compound 1-2 °HCl/hydrate form the solid is stirred in fresh i-PrOH (3.50 L) and water (0.87 L) at 50 0C for at least 2 h until XRPD shows complete conversion to Compound 1—2 nhydrate. The suspension is then cooled to 5 °C and d for 1 h. The solid is collected by filtration then the filter-cake is washed with 80/20 1'- PrOH/water (2 x 874 mL) then dried to afford Compound I—2°HCl/anhydrate.

WO 49726 ative procedure (Single pot) used Compound 1—2 ° 2HCl (3 92 g) is charged to the reactor. 4:1 IPA/water (8 L) is charged to a reactor and stirred at ambient temperature overnight. XRPD is used to confirm the conversion to the mono-HCl salt mono-hydrate form. The mixture is heated to 50 OC.

Seeds of Compound 1—2 ° HCl/anhydrate (16 g) are added and the mixture heated at 50 0C until XRPD confirms complete conversion to the d anhydrate form. The mixture is to cooled to ambient, filtered and the solid washed with 4:1 IPA/water (2 x 800 mL) then dried to afford Compound 1-2 ° HCl/anhydrate (343 g, 94% yield).

Ste 4: ate Method 1: Pre aration of Com ound I-2 free base 1. TFA 0CN/Me /N NH2 DCM | HN/Me - 25 °C \N / . 2. NaOH . 0~N IPI'OZS IPFOZS EtOH/water i Compound l-2 (free base) A solution of Compound 5—i (100 g, 131 mmol) in DCM (200 mL) was stirred at t temperature then TFA (299 g, 202 mL, 2.62 mol) was added. After 2 h reaction solution was cooled to 5 °C. The reaction mixture was diluted with EtOH (1.00 L) over about min resulting in a bright yellow suspension. The suspension was cooled to 10 °C then NaOH (1.64 L of 2.0 M, 3.28 mol) was added over 30 min then stirred at ambient temperature overnight. The solid was collected by filtration then washed with water (2 x 400 mL), EtOH (2 x 200 mL) then dried to afford Compound 1—2 free—base (57.0 g, 94% yield, 99.7 area % purity by HPLC) as a fine, yellow powder. 1H NMR (400 MHz, DMSO) 5 8.95 (s, 1H), 8.39 (d, J= 8.5 Hz, 2H), 7.95 (dd, J= 11.6, 8.4 Hz, 4H), 7.78 (s, 1H), 7.51 (d, J= 8.2 Hz, 2H), 7.21 (br s, 2H), 3.72 (s, 2H), 3.47 (hept, J= 6.8 Hz, 1H), 2.29 (s, 3H), 1.19 (d, J= 6.8 Hz, 6H).

Ste 4: Alternate Method 2: Pre aration of Com ound I-2 0 HCl /N “HZ -HC| HN’Me | HN/Me aq HCI \ N / Acetone O iPrOZS N iPrOZS Compound |-2 Compound l-2 HCI A suspension of Compound 1—2 free base (10.0 g, 21.6 mmol) in e (80 mL) was stirred and heated to 35 °C. An aqueous solution of HCl (11.9 mL of 2.0 M, 23.8 mmol) diluted with water (8.0 mL) was added and the mixture heated at 50 0C for 4 h. The suspension was allowed to cool to ambient temperature then stirred overnight. The solid was ted by filtration. The filter-cake was washed with acetone (2 x 20 mL) then dried to afford 10.2 g Compound 1—2 hydrochloride (95% yield) as a yellow powder.

Exam le 7: S nthesis of 5- 4- Iso r0 lsulfon l hen l 3- 4- tetrah dro ran-4— lamino meth l hen l isoxazol—S- l razinamine Com ound Scheme: Exam le S nthesis of Com ound 1—3 B(OH)2 QNBoo QN[Boo NVI N(BOC)}/ / N eozi-Pr A'5" Aii NCS, c, Br _ , 1. cat. Pd(dtbpf)C|2, ,N_ 20 - 30 °c ,N— TEA, DCM PhCH3, aq. ch03 HO HO Cl 20 - 30 °C 2. EtOH crystallization 3. MP-TMT resin A-4 Ai A-5 1. TFA - 25 °C 2. NaOH EtOH/water SOzi—Pr 302“” A 6 I-3 Ste 1: Pre n of N— 4- diethox meth l benz l tetrah dro-ZH— ranamine A-Z EtN(i-Pr)2, NaBH4 MeOH NH2°HC| 20- 25 °C tAZQ A on of tetrahydro-2H—pyranamine hydrochloride (1.13 kg, 8.21 mol) in MeOH (14.3 L) is stirred at about 20 0C. then Et3N (1.06 kg, 1.43 L, 8.21 mol) is added.

The mixture is d for at least 5 min then terephthalaldehyde diethyl acetal (1.43 kg, 6.84 mol) is added while maintaining the reaction ature between 20—25 °C. The mixture is stirred for at least 45 min to form the imine. NaBH4 caplets (414 g, 11.0 mol) are added while maintaining the reaction temperature below about 25 OC. The mixture is stirred for 1 h after the addition is completed. The reaction mixture is quenched by adding 1 M NaOH (13.7 L) then extracted with MTBE. The organic solution was washed with brine (7.13 L) then dried (NaZSO4) and concentrated to afford Compound A—2 (2197 g; 109% yield, 94.4 area % purity by HPLC) as a hazy oil. 1H NMR (400 MHz, CDCl3) 5 7.43 (d, J: 8.1 Hz, 2H), 7.31 (d, J: 8.1 Hz, 2H), 5.49 (s, 1H), 4.66 (br s, 1H), 4.03 — 3.91 (m, 2H), 3.82 (s, 2H), 3.69 — 3.47 (m, 4H), 3.38 (td, J: 11.6, 2.1 Hz, 2H), 2.78 — 2.65 (m, 1H), 1.90 — 1.81 (m, 2H), 1.53 — 1.37 (m, 2H), 1.23 (t, J: 7.1 Hz, 6H).

Ste 2: Pre aration of tert-but l4— diethox meth lben ltetrah dro-ZH— ran ylgcarbamate gA-31 DCM ‘ - 25 °C A mixture of N—(4-(diethoxymethyl)benzyl)tetrahydro-2H—pyranamine (A-2) (2195 g, 7.48 mol) in CHzClz (22.0 L) is stirred at 25 0C then di-t-butyl dicarbonate (1.71 kg, 7.86 mol) is added. Et3N (795 g, 1.10 L) is then added while maintaining the reaction temperature n 20 — 25 OC. The reaction mixture is d at about 25 0C for 12 — 20 h.

After the reaction is completed, the mixture is cooled to about 20 °C and quenched with 0.5 M aqueous citric acid (7.48 L, 3.74 mol) while maintaining the reaction temperature between — 25 °C. The c phase is collected, washed with sat. NaHCO3 (6.51 L, 7.48 mol), washed with brine (6.59 L), and dried (NaZSO4) then concentrated to afford tert—butyl 4— (diethoxymethyl)benzyl(tetrahydro-2H—pyranyl)carbamate (A—3) (2801 g; 95% yield, 98.8 area % purity by HPLC) as a thick, amber oil. 1H NMR (400 MHz, CDCl3) 5 7.40 (d, J = 8.1 Hz, 2H), 7.21 (d, J: 7.9 Hz, 2H), 5.49 (s, 1H), 4.39 (br s, 3H), 3.93 (br dd, J= 10.8, 3.8 Hz, 2H), 3.67 — 3.47 (m, 4H), 3.40 (br m, 2H), 1.68 — 1.59 (m, 4H), 1.39 (br s, 9H), 1.23 (t, J= 7.1 Hz, 6H).

Ste 3: Pre n of tert-but 14- h drox imino meth lben h dro-ZH— ran- 4-y11carbamate gA-41 Eto/KfljV NH20HHCI N‘Boc THF/H20 HOIK©VZBOC - 25 °C A-3 A-4 A solution of tert—butyl 4-(diethoxymethyl)benzyl(tetrahydro-2H—pyran yl)carbamate (A-3) (2.80 kg, 7.12 mol) in THF (28.0 L) and water (2.80 L) is stirred at about °C. Hydroxylamine hydrochloride (593 g, 8.54 mol) is added while maintaining the reaction temperature between 20—25 0C. The reaction mixture is stirred at about 20 °C for 16 - 20 h then diluted with CHzClz (8.4 L) and 50% brine (11.2 L) and stirred for at least 5 min.

The phases are separated then the organic phase is washed with 50% brine (2 X 2.8 L), dried (Na2SO4) and trated. The concentrate is diluted with MeOH (1.4 L) and re— trated. The concentrate is d with MeOH (14.0 L) and transferred to a reaction vessel. The solution is warmed to about 25 °C then water (14.0 L) is added over about 1 — 1.5 h; after about 10 L of water is added, the mixture is seeded and a hazy suspension is observed. Additional water (8.4 L) is added over 1.5 h to further precipitate the product. After aging, the solid is collected by filtration. The filter-cake is washed with heptane (5.6 L) and dried to afford tert—butyl 4-((hydroxyimino)methyl)benzyl(tetrahydro-2H—pyran yl)carbamate (A—4) (1678 g; 71%, 91.5 area % purity by HPLC) as an off—white powder. 1H NMR (400 MHz, CDCl3) 5 8.12 (s, 1H), 7.51 (d, J: 8.2 Hz, 2H), 7.24 (d, J: 7.9 Hz, 2H), 4.40 (br s, 3H), 3.96 (dd, J= 10.4, 3.6 Hz, 2H), 3.41 (br m, 2H), 1.69 — 1.61 (m, 4H), 1.39 (br s, 9H).

Ste 4: Pre aration of tert—but l4- chloro h drox imino meth lbenz l tetrah dro- 2H- ran lcarbamate Ai HO‘N O NCS i-PrOAc B00 20- 30 °C HOT)\©\/:BOC A-4 A-l ] A suspension of (E)—tert—butyl 4—((hydroxyimino)methyl)benzyl(tetrahydro—2H— pyranyl)carbamate (A-4) (1662 g, 4.97 mol) in i-PrOAc (16.6 L) is stirred at 20 0C in a reactor. N—chlorosuccinimide (730 g, 5.47 mol) is added maintaining about 20 °C. The suspension is d at about 20 °C to complete the reaction. The suspension is diluted with water (8.3 L) and stirred to dissolve the solid. The phases are separated and the organic phase is washed with water (8.3 L). The c phase is concentrated then diluted with i-PrOAc (831 mL). e (13.3 L; 8 V) is slowly added to induce crystallization. The thick sion is then stirred for 1 h. The solid is collected by filtration; the filter-cake is washed with heptane (2 x 1.6 L; 2 x 1 V) and dried to afford (Z)—tert—butyl 4— (chloro(hydroxyimino)methyl)benzyl (tetrahydro-2H—pyranyl)carbamate (Ai) (1628 g; 89%, 98.0 area % purity by HPLC) as a white powder.

Ste 5: Pre aration of tert—but l 5-br0m0 3- 4- tert-butox carbon 1 tetrah dro- 2H- ran lamino meth l hen zol—S- l razin-Z- l tert- butox carbon lcarbamate A-5 TEA DCM Boc\ / 00/ N \ NBoc / Br 0O Al 19;:Aii A-5 A solution of tert—butyl 4-(chloro(hydroxyimino)methyl)benzyl(tetrahydro-2H- pyranyl)carbamate (Ai) (1.60 kg, 4.34 mol) and tert—butyl N—(S—bromo—3— cthynylpyrazin—2—yl)—N~rent—butoxycarbonylcarbamate und A—4—ii) (1.73 kg, 4.34 mol) in CHzClz (12.8 L) is stirred at 20 °C. Et3N (483 g, 665 mL; 4.77 mol) is added and the reaction temperature maintained below 30 °C. The suspension stirred at 20 °C to complete the reaction then diluted with water (8.0 L) and agitated. The phases are separated and the organic phase is washed with water (8.0 L) and then concentrated. i-PrOAc (1.6 L) is added and the mixture and heated at 50 °C. Heptane (4.0 L) was slowly added then the suspension is allowed to cool to ambient temperature and stirred overnight. Additional heptane (7.2 L) is added to the suspension and it is stirred for 1 h. The solid is collected by filtration. The filter— cake is washed with heptane (2 x 1.6 L) and dried to afford tert—butyl (5—bromo—3—(3 —(4— (((tert—butoxycarbonyl)(tetrahydro-2H—pyranyl)amino)methyl)phenyl)isoxazol yl)pyrazin—2—yl)(tert—butoxycarbonyl)carbamate (A—5) (2.478 kg; 78%, 97.8 area % purity by HPLC) as a fine, tan powder. 1H NMR (400 MHZ, CDCl3) 5 8.60 (s, 1H), 7.78 (d, J = 8.3 Hz, 2H), 7.31 (m, 3H), 4.42 (br m, 3H), 4.03 — 3.82 (m, 2H), 3.38 (br s, 2H), 1.60 (m, 4H), 1.36 (s, 27H).

Ste 6: Pre aration of tert—bu ltert—butox carbon 1 3- 3- 4- tert- butox carbon 1 tetrah dro-ZH- ran lamino meth l hen lisoxazol—S- 1 4- iso r0 lsulfon l hen l razin-Z- lcarbamate KfNI (j 1. cat. Pd(dtbpf)C|2, PhCH3, aq. K2CO3 2. EtOH llization 3. MP-TMT resin SOzi-Pr A mixture of tert—butyl (5 (3-(4-(((tert—butoxycarbonyl)(tetrahydro— 2H—pyranyl)amino)methyl)phenyl)isoxazolyl)pyrazinyl)(tert— butoxycarbonyl)carbamate (A-5) (425 g, 582 mmol), K2CO3 (161 g, 1.16 mol; 2.0 ), and (4-(isopropylsulfonyl)phenyl)boronic acid (133 g, 582 mmol) in toluene (2.98 L) and water (850 mL) is stirred and degassed with N2 at ambient ature. The catalyst [11- bis(di—tert—butylphosphino)ferrocene]dichloropalladium(II), (Pd(dtbpf)Clz; 1.90 g, 2.91 mmol) is added and the mixture is ed for an additional 10 min. The e is heated at 70 °C until the reaction is complete. The mixture is cooled to 50 0C, diluted with water (850 mL) and filtered through a bed of Celite. The phases are separated. The organic phase is concentrated then the residue is diluted with EtOH (1.70 L) and re—concentrated. With mixing at 40 0C, the concentrate is diluted with EtOH (1.70 L) to induce crystallization. The suspension is cooled to 20 °C and stirred for 4 h. The solid is collected by filtration. The filter-cake is washed with EtOH (2 x 425 mL) and air-dried to afford tert—butyl tert— butoxycarbonyl(3 —(3 —(4—(((tert—butoxycarbonyl)(tetrahydro-2H—pyran yl)amino)methyl)phenyl)isoxazolyl)-5 sopropylsulfonyl)phenyl)pyrazin yl)carbamate (A-6) as a beige powder. The solid is dissolved in THF (2.13 L) and slurried with Biotage MP-TMT resin (48 g) at ambient temperature. The resin is removed by filtration and the filtrate concentrated to remove most of the THF. The concentrate is d with EtOH (970 mL) and re-concentrated to about half the original volume. The concentrate is diluted again with EtOH (970 mL) and mixed for 1 h at 40 °C. The suspension is cooled to ambient temperature and the solid is collected by tion then dried to afford tert—butyl tert— butoxycarbonyl(3 —(3 —(4—(((tert—butoxycarbonyl)(tetrahydro-2H—pyran yl)amino)methyl)phenyl)isoxazolyl)-5 -(4-(isopropylsulfonyl)phenyl)pyrazin yl)carbamate (A—6) (416 g; 86% yield, 99.3 area % purity by HPLC) as a white powder. 1H NMR (400 MHz, CDCl3) 5 9.04 (s, 1H), 8.38 — 8.28 (m, 2H), 8.10 — 8.01 (m, 2H), 7.82 (d, J = 8.2 Hz, 2H), 7.34 (m, 3H), 4.44 (br s, 2H), 3.94 (dd, J: 10.5, 3.5 Hz, 2H), 3.40 (br s, 2H), 3.25 (hept, J: 6.8 Hz, 1H), 1.65 (m, 4H), 1.38 (br s, 27H), 1.33 (d, J: 6.9 Hz, 6H).

Ste 7: Pre aration of 5- 4- iso r0 lsulfon l hen 1 3- 4- tetrah dro-ZH— ran- 4- lamino meth l hen lisoxazol—S- l razin-Z-amine I-3 se form Boc\ ,Boc NWNO’N NH2 0’\ ‘\ 1 TFA \\ - NW | N\Boc DCM I NH /'N </::§ N -25°C / o 2.NaOH </::So ater A-6 I-3 SOflPr SOjPr A suspension of tert—butyl tert—butoxycarbonyl(3 —(3—(4—(((tert— butoxycarbonyl)(tetrahydro-2H—pyranyl)amino)methyl)phenyl)isoxazolyl)(4- (isopropylsulfonyl)phenyl)pyrazinyl)carbamate (A-6) (410 g; 492 mmol) in CHzClz (410 mL) is stirred at ambient temperature in a flask. TFA (841 g, 568 mL; 7.4 mol) is added while maintaining the on temperature between 20—25 °C. The solution is stirred at ambient temperature for about 3 h when analysis shows on completion. The solution is cooled to about 5—10 °C and diluted with EtOH (3.3 L) while maintaining the temperature below 20 0C. A 5.0 M s solution ofNaOH (1.77 L; 8.85 mol) is added while allowing the reaction temperature to rise from about 14°C to about 42 °C. The suspension is heated at 70 — 75 °C for 6 h while removing distillate. The sion is d to cool to ambient temperature. The solid is collected by filtration and the filter-cake is washed with water (4 x 1.64 L). The filter-cake is washed with EtOH (2 x 820 mL) and dried to afford 5-(4- (isopropylsulfonyl)phenyl)(3-(4-(((tetrahydro-2H—pyranyl)amino)methyl)phenyl) isoxazol—5—yl)pyrazin—2—amine (Compound 1—1) (257 g; 98% yield, 99.5 area % purity by HPLC) as a yellow powder. 1H NMR (400 MHz, DMSO) 5 8.94 (s, 1H), 8.44 — 8.33 (m, 2H), 7.94 (t, J= 8.2 Hz, 4H), 7.76 (s, 1H), 7.53 (d, J= 8.2 Hz, 2H), 7.20 (s, 2H), 3.83 (m, 1H), 3.80 (s, 3H), 3.46 (hept, J= 6.8 Hz, 1H), 3.25 (td, J= 11.4, 2.1 Hz, 2H), 2.66 — 2.54 (m, 1H), 1.79 (br dd, 2H), 1.36 — 1.22 (m, 2H), 1.19 (d, .1: 6.8 Hz, 6H).13C NMR (101 MHz, DMSO) 5 167.57, 151.76, 141.07, 137.58, 135.75, 129.16, 128.53, 126.57, 126.41, 125.69, 124.52, 102.13, 65.83, 54.22, 52.60, 49.19, 33.18, 15.20.

Compound Analytical Data Cmpd LCMS LCMS HNMR No. BS + Rt min H NMR (400 MHz, DMSO) 5 9.63 (d, J = 4.7 Hz, 2H), 9.05 (s, 1H), 8.69 (d, J= 5.2 Hz, 1H), 8.21 (s, 1H), 8.16 — 8.03 (m, 3H), 7.84 (t, J= 4.1 Hz, 3H), 7.34 (br s, 2H), 4.40 — 4.18 (m, 2H), 3.94 (dd, J= 11.2, 3.9 Hz, 2H), 3.32 (t, J= 11.2 Hz, 3H), 2.17 — 2.00 (m, 2H), 1.81 (s, 6H), 1.75 (dd, J = 12.1, 4.3 Hz, 2H. 1H NMR (400 MHz, DMSO) 5 8.94 (s, 1H), 8.44 — 8.33 (m, 2H), 7.94 (t, J: 8.2 Hz, 4H), 7.76 (s, 1H), 7.53 (d, J= 8.2 Hz, 2H), 7.20 (s, 2H), 3.83 (m, 1H), 3.80 (s, 3H), 3.46 (hept, J= 6.8 Hz, 1H), 3.25 (td, J: 11.4, 2.1 Hz, 2H), 2.66 — 2.54 (m, 1H), 1.79 (br dd, 2H), 1.36 — 1.22 (m, 2H), 1.19 d, J= 6.8 Hz, 6H. 1H NMR (500 MHz, DMSO) 9.10 (d, J = 5.8 Hz, 2H), 8.97 (s, 1H), 8.42 — 8.37 (m, 2H), 8.15 — 8.08 (m, 2H), 7.99 — 7.92 II—l 466.2 0.83 (m, 2H), 7.85 (s, 1H), 7.75 — 7.69 (m, 2H), 7.22 (br s, 2H), 3.48 (hept, J = 6.8 Hz, 1H), 2.60 (t, J = 5.4 Hz, 3H), 1.20 (d, J = 6.8 Hz, 6H). 1H NMR (500 MHz, DMSO) 9.14 (s, 2H), 8.96 (s, 1H), 8.42 — 8.36 (m, 2H), 8.13 — 8.08 (m, 2H), 7.99 — 7.91 (m, H—2 469.1 0.82 2H), 7.85 (s, 1H), 7.78 — 7.68 (m, 2H), 7.21 (s, 2H), 3.48 (dq, J = 13.6, 6.7 Hz, 1H), 1.20 (d, J = 6.8 Hz, 6H). 1H NMR (500 MHz, DMSO) 9.11 (s, 2H), 8.97 (s, 1H), 8.39 (d, J = 8.7 Hz, 2H), 8.11 (d, J = 8.4 Hz, 2H), 7.95 (d, J 11—3 467.2 0.78 = 8.7 Hz, 2H), 7.85 (s, 1H), 7.72 (d, J = 8.4 Hz, 2H), 7.23 (s, 2H), 4.25 — 4.19 (m, 2H), 3.47 (tt, J = 14.0, 6.9 Hz, 1H), 1.20 (d, J = 6.8 Hz, 6H). 1H NMR (400 MHz, DMSO) 8 2.58 (t, 3H), 4.21 (t, 2H), 114 471.8 0.83 5.67 (br s, 2H), 7.74 (d, 2H), 7.85 (s, 1H), 7.94 (d, 2H), 8.10 (d, 2H), 8.38 (d, 2H), 8.96 (s, 1H) and 9.33 (br s, 2H) ppm INTERMEDIATES Example 8: Preparation of Oxime Sa SCHEME BB: Step 1b OEt Step 2b NaBH4 Nx DCM MeOH OEt HO\ EtO)\©\/II3OC Step 3b IN NHZOH-HCI Boc N —> ' THF/water N\ 3b 5a Step 1b Add MeOH (28.00 L) and 4—(diethoxymethyl)benzaldehyde (Compound 1b) (3500 g, 16.81 mol) into a reactor at 20 °C. Add methylamine, 33% in EtOH (1.898 kg, 2.511 L of 33 %w/w, 20.17 mol) maintaining 20-30 0C then stir for 1.5 h to form the imine. Add NaBH4 (381.7 g, 10.09 mol) caplets maintaining the temperature between 20 - 30 °C. Stir at room temperature for at least 30 min to ensure complete reaction. Add s NaOH (16.81 L of 2.0 M, 33.62 mol) maintaining approximately 20 °C. Add MTBE (17.50 L) and brine (7.0 L), stir for at least 5 min then allow the phases to separate. Extract the aqueous layer with MTBE (7.0 L) then combine the c phases and wash with brine (3.5 L) then dry (NaZSO4) then concentrate to 6 L. The biphasic mixture was transferred to a separatory funnel and the aqueous phase d. The organic phase was concentrated to afford 1—(4- (diethoxymethyl)phenyl)-N-methylmethanamine (Compound 2b) (3755 g, 16.82 mol, 100% yield) as an oil. 1H NMR (400 MHz, CDCl3) 5 7.43 (d, J: 8.1 Hz, 2H), 7.31 (d, J: 8.1 Hz, 2H), 5.49 (s, 1H), 3.75 (s, 2H), 3.68 — 3.46 (m, 4H), 2.45 (s, 3H), 1.23 (t, J: 7.1 Hz, 6H).

Steps 2b and 3b Add 2-MeTHF (15.00 L) and 1-(4-(diethoxymethyl)phenyl)-N- methylmethanamine (Compound 2b) (3750 g, 16.79 mol) to a reactor at 20 °C. Add a solution of Boc anhydride (3.848 kg, 4.051 L, 17.63 mol) in 2—MeTHF (7.500 L) maintaining approximately 25 OC. Stir for at least 30 min to ensure complete sion to utyl 4— (diethoxymethyl)benzyl(methyl)carbamate (Compound 3b), then add a solution ofNaZSO4 (1.192 kg, 8.395 mol) in water (11.25 L). Heat to 35 0C then add a on of hydroxylamine hydrochloride (1.750 kg, 25.18 mol) in water (3.75 L) then stir for at least 6 h to ensure complete reaction. Cool to 20 OC, stop the stirring and remove the aqueous phase. Wash the organic layer with brine (3.75 L), dry (NaZSO4), filter and concentrate to about 9 L. Add heptane (15.00 L) and crystalline tert—butyl 4-((hydroxyimino)methyl)benzyl(methyl) carbamate (Compound 5a) (1.0g portions every 10 min) until nucleation was evident, then concentrate to afford a solid slurry. Add heptane (3.75 L) then cool to room temp and filter.

Wash with heptane (5.625 L) then dry to afford tert—butyl 4-((hydroxyimino)methyl) (methyl)carbamate (Compound 5a) (4023 g, 15.22 mol, 91 % yield, 97.2 area % purity by HPLC) as a colorless solid. 1H NMR (400 MHz, CDC13) 5 8.13 (s, 1H), 7.54 (d, J: 8.1 Hz, 2H), 7.25 (br d, 2H), 4.44 (br s, 2H), 2.83 (br d, 3H), 1.47 (br s, 9H).

Scheme CC: Synthesis of Intermediate Aii ' l /N ,N -PTSA Sonogashira BOC protection Br —> Br —> N(Boc)2 TMS N(Boc)2 / / N \ NV KfN| TMS removal KKN| Br Br C-3 Aii The compound of formula Aii may be made ing to the steps outlined in Scheme C. Sonogashira coupling reactions are known in the art (see e. g., Chem. Rev.

WO 49726 2007, 874—922). In some embodiments, suitable Sonogashira coupling conditions comprise adding 1 equivalent of the compound of formula C-1, 1 equivalent of TMS—acetylene, 0.010 equivalents of 3)2C12, 0.015 equivalents of CuI and 1.20 equivalents ofNMM in isopropanol. The product can be ed by adding water to the alcoholic reaction mixture.

Amine salts of a product maybe formed by dissolving the amine in a common organic solvent and adding an acid. Examples of suitable solvents include chlorinated solvents (e. g., dichloromethane (DCM), dichloroethane (DCE), , and chloroform), ethers (e. g., THF, 2-MeTHF and dioxane), esters (e. g., EtOAc, IPAC) and other c solvents. Examples of suitable acids include but are not limited to HCl, H3PO4, H2804, MSA, and PTSA. In some embodiments, the solvent is IPAC and the acid is PTSA. In some embodiments, the acid addition salt is converted back to the free amine base in the ce of a suitable solvent and a suitable base. Suitable solvents e EtOAc, IPAC, romethane (DCM), dichloroethane (DCE), CHzClz, chloroform, 2-MeTHF, and suitable bases include NaOH, NaHCO3, NazCO3, KOH, KHCO3, K2CO3, and CszCO3. In some embodiments, the suitable solvent is EtOAc and the suitable base is KHCO3_ The amine of Compound C-2 may be protected with various amine protecting groups, such as Boc (tert—butoxycarbonyl). Introduction of Boc protecting groups is known in the art (see e. g. Protecting Groups in Organic Synthesis, Greene and Wuts). In some embodiments, suitable conditions e adding 1.00 equivalents of the amine, 2.10 equivalents of di—tert—butyl dicarbonate, and 0.03 equivalents of DMAP in EtOAc.

Reduction in Pd is achieved by treating with a metal scavenger (silica gel, functionalized resins, charcoal). In some embodiments, suitable conditions involve adding charcoal.

The TMS (trimethylsilyl) protecting group on Compound C—3 may be removed via conditions known to one of skill in the art. In some embodiments, TMS removal conditions comprise reacting the TMS-protected compound with a suitable base in a le solvent. Examples of suitable solvents include chlorinated solvents (e. g., dichloromethane (DCM), dichloroethane (DCE), CHzClz, and form), ethers (e. g., THF, 2-MeTHF and dioxane), esters (e. g., EtOAc, IPAC), other c solvents and alcohol solvents (e. g., MeOH, EtOH, iPrOH). Examples of suitable bases include but are not limited to (e. g., NaOH, KOH, K2C03, NazCO3). In certain embodiments, suitable ions comprise adding 1.00 equivalents of the TMS—protected ene, 1.10 equivalents of K2C03, EtOAc and EtOH. In some emboments, the alcoholic solvent, such as EtOH, is added last in the reaction. In some ments the product acetylene is isolated by adding water.

Scheme DD: Example Synthesis of Compound Aii NH2 1. TMS-acetylene NH2 TMS N)\( cat. 3)2C|2 ¢ 1. EtOAc, aq. KHC03 cat. Cul, NMM, IPA N \ 2. 80020, cat. DMAP, RN 2. water RN °PTSA EtOAc 3. PTSA, EtOAc 3. charcoal Br —> Br —> c-1 (65-75%) 0-2 0%) M80095 TMS 1-K2003, EtOAc, N(Boc)}/ EtOH N \ NV M 2W_t, 9N Br (75-80%) Br c-3 Aii Exam le 9: S nthesis of Com ound Aii NH2 1. TMS-acetylene NH2 TMS cat. Pd(PPh3)2CI2 Br- é NI \ cat. Cul, NMM, IPA N \ RN 2. water RN ‘PTSA 3. PTSA, EtOAC 31‘ —> Br c-1 (65-75%) c-2 Charge panol (8.0 L) to a reactor the stir and sparge with a stream of N2.

Add 3,5—dibromopyrazin—2—amine (Compound C—l) (2000 g, 7.91 moles), Pd(PPh3)2Clz (56 g, 0.079 , CuI (23 g, 0.119 moles), and NMM (1043 mL, 9.49 moles) to the reactor under a N2 atmosphere. Adjust the reaction temperature to 25 °C. Purge the reactor with N2 by doing at least three vacuum/N2 purge cycles. Charge TMS—acetylene (1.12 L, 7.91 moles) to the reaction mixture and maintain the reaction temperature below 30 °C. When the reaction is complete lower the temperature of the reaction mixture to 15 0C then add water (10 L) and stir for at least 2 h. The solid is ted by filtration washing the solid with 1:1 IPA/water (2 x 6 L). The filter cake is dried under vacuum then charged to a reactor and dissolved in EtOAc (12.5 L). PTSA hydrate (1.28 kg, 6.72 mol) is charged as a solid to the reactor. The mixture is stirred at ambient temperature for at least 5 h then the solid is collected by filtration, washed with 1:1 heptane/EtOAc (3.5 L) followed by e (3.5 L). The filter cake is dried to afford o((trimethylsilyl)ethynyl)pyrazinamine(Compound C-2) as a PTSA salt (2356 g, 67% yield, 98.9 area % purity by HPLC).1H NMR (400 MHz, DMSO) 5 8.12 (s, 1H), 7.48 (d, J: 8.1 Hz, 2H), 7.12 (d, J: 8.0 Hz, 2H), 2.29 (s, 3H), 0.26 (s, 9H).

Steps 2 and 3 1. EtOAc, aq. KHC03 2. B0020, cat. DMAP, 1_ K2C03 EtOAc ””2 T'V'S é EtOAc N(Boc)} TMS EtOH N(Boc)} N \ 3. charcoal NV 2_ water NV ka -PTSA —> | —> I (95—1000/0) YN (75-80%) Br V” Br Br c_3 Aii A solution of5-bromo((trimethylsilyl)ethynyl)pyrazinamine PTSA salt (Compound C—2) (2350 g, 5.31 mol) in EtOAc (11.5 L) is stirred with a 20% w/w aq. solution of KHC03 (4.5 kg, 1.5 eq.) for at least 30 min. The layers are separated and the organic layer is concentrated then dissolved in EtOAc (7 L) and added to a reactor. DMAP (19.5 g, 0.16 mol) is added followed a solution of BoczO (2436 g, 11.16 mol) in EtOAc (3 L) is added lowly. The reaction is stirred for at least 30 min to ensure complete reaction then activated charcoal (Darco G—60, 720 g) and Celite (720 g) are added and stirred for at least 2 h. The mixture is ed washing the solid pad with EtOAc (2 x 1.8 L). The filtrate is concentrated to afford tert—butyl N—tert—butoxycarbonyl-N—[5-bromo((trimethylsilyl)ethynyl) pyrazin bamate (Compound C—3) that is used directly in the next step.

Ste 3: Pre aration of tart-bu '1 Nu Subromo—3—ethvnvl vrazin-Z-vl)—N-tert—= butox a carbon learbamate -'Com n ound Aii K2CO3 (811 g, 5.87 mol) is charged to a reactor ed by a solution of Compound C—3 (2300 g, 4.89 mol) dissolved in EtOAc (4.6 L) agitation started. EtOH (9.2 L) is added slowly and the e stirred for at least 1 h to ensure that the reaction is complete then water (4.6 L) is added and stirred for at least 2 h. The solid is collected by filtration and washed with 1:1 EtOH/water (4.6 L followed by 2.3 L) followed by EtOH (2.3 L). The filter cake is dried to afford zerr-butyl N—(S—bromo—3—ethynylpyrazin—2—yl)—N—tert— butoxycarbonylcarbamate (Compound A—4—ii) (1568 g, 78% yield, 97.5 area % by HPLC).1H NMR (400 MHz, CDCl3) 5 8.54 (s, 1H), 3.52 (s, 1H), 1.42 (s, 18H).

Solid Forms of Compound I-2 Compound 1-2 has been prepared in various solid forms, including salts and co- solvates. The solid forms of the present invention are useful in the manufacture of medicaments for the treatment of cancer. One embodiment provides use of a solid form described herein for treating cancer. In some embodiments, the cancer is pancreatic cancer or non-small cell lung . Another embodiment provides a pharmaceutical composition comprising a solid form described herein and a ceutically able carrier. ] ants describe herein five novel solid forms of Compound 1—2. The names and stoichiometry for each of these solid forms are provided in Table S—l below: Table S—l Examole 13 Comoound 1—2 free base Exam-16 14 : Example 15 Compound 1-2 ° 2HCl 1:2 Example 16 Compound 1-2 ° HCl ° H20 121 Example 17 Compound 1-2 ° HCl ° 2H20 1:1:2 Solid state NMR spectra were acquired on the Bruker-Biospin 400 MHz e III wide—bore spectrometer equipped with Bruker—Biospin 4mm HFX probe.

Samples were packed into 4mm ZrOz rotors (approximately 70mg or less, depending on sample availability). Magic angle spinning (MAS) speed of typically 12.5 kHz was applied.

The temperature of the probe head was set to 275K to ze the effect of frictional heating during spinning. The proton relaxation time was ed using 1H MAS T1 saturation recovery relaxation experiment in order to set up proper recycle delay of the 13 C cross-polarization (CP) MAS ment. The recycle delay of 13C CPMAS experiment was WO 49726 adjusted to be at least 1.2 times longer than the ed 1H T1 relaxation time in order to maximize the carbon spectrum signal-to-noise ratio. The CP contact time of 13C CPMAS experiment was set to 2 ms. A CP proton pulse with linear ramp (from 50% to 100%) was employed. The Hartmann-Hahn match was optimized on external reference sample (glycine). Carbon spectra were ed with SPINAL 64 ling with the field strength of approximately 100 kHz. The chemical shift was referenced against external standard of tane with its upf1eld resonance set to 29.5 ppm.

XRPD data for Examples 13—14 were measured on Bruker D8 Advance System (Asset V014333) equipped with a sealed tube Cu source and a Vantec—l detector (Bruker AXS, Madison, WI) at room temperature. The X-ray generator was operating at a voltage of 40 kV and a current of 40 mA. The powder sample was placed in a shallow silicon .

The data were recorded in a reflection scanning mode (locked coupled) over the range of 3°— 400 2 theta with a step size of 0.01440 and a dwell time of 0.25s (105 s per step). Variable divergence slits were used.

Example 10: Compound I-2 [free base] Compound 1—2 free base can be formed according to the methods described in e 6, Step 4: Alternate Method 1.

XRPD of Compound 1—2 [free base] Figure 1a shows the X-ray powder diffractogram of the sample which is characteristic of crystalline drug substance.

Representative XRPD peaks from Compound 1—2 free base: XRPD Angle Intensity % Peaks (2-Theta :: 0.2) 1 23.8 100.0 *2 14.2 43.9 3 22.5 39.3 *4 25.6 31.1 19.3 28.6 6 27.2 27.6 7 17.0 25.4 *8 18.1 25.2 9 17.6 19.6 20.2 17.2 11 28.3 15.6 12 20.8 14.5 13 29.9 14.5 14 33.2 14.3 30.1 13.5 XRPD Angle Intensity % Peaks (2-Theta :: 0.2) 16 26.8 13.4 *17 22.0 12.3 18 36.5 12.3 19 31.8 12.2 34.6 11.5 21 31.1 11.2 22 34.0 11.0 23 30.6 10.9 *24 11.1 10.6 13.3 10.6 Thermo Analysis of Compound 1—2 free base A thermal graVimetric is of Compound 1—2 free base was performed to determine the percent weight loss as a on of time. The sample was heated from ambient temperature to 350°C at the rate of 10°C/min on TA Instrument TGA Q5000 (Asset V01425 8). Figure 2a shows the TGA result with a one-step weight loss before evaporation or thermal decomposition. From ambient temperature to 215°C, the weight was ~1.9 %.

Differential Scanning Calorimetm of Compound 1—2 free base The thermal properties of Compound 1—2 free base were measured using the TA Instrument DSC Q2000 (Asset V014259). A Compound 1—2 free base sample (1.6900 mg) was weighed in a pre-punched pinhole aluminum hermetic pan and heated from ambient temperature to 350°C at 10°C/min. One endothermic peak is observed at 210°C with its onset temperature at 201°C e 3a). The enthalpy associated with the ermic peak is 78 J/g.

Solid State NMR of nd 1—2 free base 13C CPMAS on Compound 1—2 free base 275K; 1H T1=1.30s 12.5 kHz spinning; ref adamantane 29.5 ppm For the full spectrum, see Figure 4a. entative Peaks Chem Shift Intensity Peak # [o om] [rel] 1* 171.0 2 163.7 3* 152.1 4 143.1 57.3 * 141.2 38.8 6 138.8 30.0 7 132.4 62.1 Cmstal ure of Compound 1—2 free base The free form of Compound 1-2 was prepared from the Compound 1-2 HCl salt. 200mg Compound 1—2 HCl salt was added to 1mL of 6N NaOH solution. 20mL of dichloromethane was used to extract the free form. The dichloromethane layer was dried over K2CO3. The solution was filtered off and 5mL of n—heptane was added to it. Crystals were obtained by slow evaporation of the solution at room temperature over night.

Most crystals obtained were thin plates. Some prismatic shape crystals were found among them.

A yellow prismatic crystal with dimensions of 0.2>< 0.1><0.1 mm3 was selected, mounted on a MicroMount and centered on a Bruker APEX II diffractometer. Three batches of 40 frames separated in reciprocal space were obtained to provide an orientation matrix and initial cell parameters. Final cell parameters were obtained and refined after data collection was completed based on the full data set.

A ction data set of ocal space was obtained to a resolution of 116.960 20 angle using 0.50 steps with 10 s exposure for each frame. Data were collected at 100 (2) K temperature with a nitrogen flow stem. Integration of intensities and refinement of cell parameters were accomplished using APEXII software.

CRYSTAL DATA C24H25N503S Mr = 463.55 inic, P21/n a = 8.9677 (1)A b= 10.1871 (1M c=24.59l4 (3)A B= 100.213 (1)0 V= 2210.95 (4) A3 Example 11: Compound I-2 0 HCl Compound 1—2 ° HCl can be formed according to the methods described in Example 6, Step 4: Alternate Method 2 and Example 6, Step 5.

XRPD of nd 1-2 ° HCl Figure lb shows the X-ray powder diffractogram of the sample which is characteristic of crystalline drug nce.

Representative XRPD peaks from Compound 1—2 0 HCl XRPD Angle ity % Peaks (2-Theta : 100.0 89.1 45.8 41.9 33.8 29.6 26.0 23.3 21.3 *10 21.1 *11 20.8 13.3 *13 13.0 12.7 Thermo Analysis of Compound 1-2 ° HCl A thermal graVimetric analysis of Compound 1-2 ° HCl was performed to determine the percent weight loss as a function of time. The sample was heated from ambient temperature to 350°C at the rate of 10°C/min on TA Instrument TGA Q5000 (Asset V01425 8). Figure 2b shows the TGA result with a two—step weight loss before evaporation or thermal decomposition. From ambient temperature to 100°C, the weight was ~1.1 %, and from 110°C to 240°C the weight loss is ~0.8%.

Differential Scanning Calorimetg of Compound 1-2 ° HCl The thermal properties of Compound 1-2 ° HCl were measured using the TA ment DSC Q2000 (Asset V014259). A Compound 1—2 ° HCl sample (3.8110 mg) was weighed in a nched pinhole aluminum hermetic pan and heated from ambient ature to 350°C at 10°C/min. One endothermic peak is observed at 293°C with its onset temperature at 291°C (Figure 3b). The enthalpy associated with the endothermic peak is 160.3 J/g. The second endothermic peak is around 321°C. Both peaks were coupled with sample ation and decomposition.

Solid State NMR of Compound 1—2 0 HCl CPMAS on Compound 1-2 ° HCl 275K;_12.5 kHz spinning; ref adamantane 29.5 ppm For the full spectrum, & Figure 4b.

Representative Peaks Peak Chem Shift Intensity # [o om] [rel] 1* 171.7 47.42 2 161.9 28.72 3* 153.4 28.94 4 144.8 42.57 142.9 54.14 6 138.7 44.06 7 136.7 60.06 8* 132.9 100 9 131.2 72.62 129.8 73.58 11 127.9 63.71 12 125.4 79.5 13 124.1 34.91 14 100.7 53.52 54.5 62.56 16 53.9 61.47 17* 31.8 61.15 18 17.0 74.78 19* 15.7 77.79 Cmstal Structure of nd 1—2 0 HCl 180mg Compound 1-2 ° HCl was added to a vial with 0.8mL 2-propanol and 0.2 mL water. The sealed vial was kept in an oven at 70°C for two weeks. ction quality crystals were observed.

A yellow needle shape crystal with dimensions of 0.15>< 0.02 ><0.02 mm3 was selected, mounted on a MicroMount and centered on a Bruker APEX II diffractometer (V011510). Three batches of 40 frames separated in reciprocal space were obtained to provide an orientation matrix and initial cell parameters. Final cell parameters were collected and refined was completed based on the full data set.

A diffraction data set of reciprocal space was obtained to a resolution of 1060 20 angle using 0.50 steps with exposure times 20 s each frame for low angle frames and 60s each frame for high angle frames. Data were collected at room temperature.

To obtain the data in table 1, dry nitrogen was blown to the crystal at 6 Litre/min speed to keep the ambient re out. Data in table 2 was obtained without en. Integration of intensities and refinement of cell parameters were conducted using the APEXH software. The water occupancy can vary between 0 and 1.

Table 1 Table 2 C1N503S C24H28C1N504S Mr = 500.01 Mr = 518.02 Monoclinic, P21/n Monoclinic, P21/n a = 5.3332 (2) A a = 5.4324 (5) A b = 35.4901 (14) A b = 35.483 (4) A c = 13.5057 (5) A c = 13.3478 (12) A [3: 100.818 (2)0 [3: 100.812 (5)0 V= 7 (17) A3 V = 2527.2 (4) A3 CHN Elemental Analysis CHN tal analysis of Compound 1-2 ° HCl suggest a mono HCl salt.

Element C H N Cl % Theory 57 60' 5 20' 14 00- 7 10.

C24H25N503S.HC1 56.52 5.38 13.69 7.18 e 12: Compound I-2 0 2HCl Compound 1—2 ° 2HCl can be formed according to the methods described in Example 6, Step 4.

XRPD of Compound 1-2 ° 2HCl ] The XRPD patterns are acquired at room temperature in reflection mode using a Bruker D8 Discover system (Asset Tag V012842) equipped with a sealed tube source and a Hi-Star area detector (Bruker AXS, Madison, WI). The X-Ray generator is operated at a voltage of 40 kV and a current of 35 mA. The powder sample is placed in a nickel holder.

Two frames are registered with an exposure time of 120 s each. The data is subsequently ated over the range of 4.5°-39° 2 theta with a step size of 0.020 and merged into one continuous pattern.

Figure 1c shows the X-ray powder ctogram of the sample which is characteristic of crystalline drug substance.

Representative XRPD peaks from Compound 1-2 ° 2HCl Peaks 2-Theta::() =_100.092-2 =—91.3 _—91.3 _—91.2 _—89.0 _—89.0 _—88-8 88.1 87.5 87.4 86.6 86.0 86.0 86.0 85.9 85.9 85.7 85.7 85.4 85 2 _—85.2 84.9 84.7 84-1 Thermo Analysis of Compound 1-2 ° 2HCl A thermal gravimetric analysis of Compound 1-2 ° 2HCl was performed on the TA Instruments TGA model Q5000. nd 1-2 ° 2HCl was placed in a platinum sample pan and heated at 10°C/min to 350°C from room temperature. Figure 2c shows the TGA result, which demonstrates a weight loss of 7.0% from room temperature to 188°C, which is consistent with the loss of 1 equivalent of HCl (6.8%). The onset temperature of degradation/melting is 263°C.

Differential Scanning Calorimetg of Compound 1-2 ° 2HCl A DSC thermogram for Compound 1—2 ° 2HC1 drug substance lot 3 was obtained using TA ments DSC Q2000. Compound 1-2 ° 2HCl was heated at n to 275°C from —20°C, and modulated at i 1°C every 60 sec. The DSC thermogram e 3c) reveals an endothermic peak below 200°C, which could corresponds to the loss of 1 equivalent of HCl. Melting/recrystallization occurs n 215—245°C, followed by degradation.

Solid State NMR of Compound 1—2 0 2HC1 13C CPMAS on Compound 12 - 2HCl 275K; 1H Tl=l.7s 12.5 kHz spinning; ref adamantane 29.5 ppm For the full spectrum, & Figure 4c.

Peak # Chem Shift [ppm] Intensity [rel] 1* 166.5 32.6 WO 49726 Cmstal Structure of Compound 1—2 0 2HCl 180mg Compound 1—2 ° HCl was added to a vial with 0.8mL 2-propanol and 0.2 mL water. The sealed vial was kept in an oven at 70°C for two weeks. Diffraction quality crystals were ed.

A yellow needle shape crystal with dimensions of 0.15>< 0.02 ><0.02 mm3 was selected, mounted on a MicroMount and centered on a Bruker APEX II diffractometer (V011510). Three batches of 40 frames separated in reciprocal space were obtained to provide an ation matrix and initial cell parameters. Final cell parameters were collected and refined was completed based on the full data set.

A diffraction data set of reciprocal space was obtained to a resolution of 1060 20 angle using 0.50 steps with exposure times 20 s each frame for low angle frames and 60s each frame for high angle frames. Data were collected at room temperature. Dry en was blown to the crystal at 6 Litre/min speed to keep the ambient moisture out. Integration of intensities and refinement of cell parameters were conducted using the APEXII software.

CRYSTAL DATA ClN503S Mr = 500.01 inic, P21/n a = 5.3332 (2) A b= 35.4901 (14)A c= 13.5057 (5M B= 100.818 (2)0 V= 2510.87 (17) A3 Example 13: Compound I-2 0 HCl 0 H20 Compound 1—2 ° HCl° H20 can be formed from Compound 1—2 0 2 HCl.

(E29244-17) A suspension of Compound 1-2 ° 2 HCl (10.0 g, 18.6 mmol) in pyl alcohol (40 mL) and water (10 mL) is warmed at 50 0C for about 1 h and then cooled to below 10 OC. The solid is collected by filtration. The filter—cake is washed with 80/20 isopropyl alcohol/water (2 x 10 mL) and air-dried to afford Compound 1-2 ° HCl° 2H20 as a yellow powder.

XRPD of nd 1-2 ° HCl ° ng The XRPD patterns are acquired at room temperature in reflection mode using a Bruker D8 Discover system (Asset Tag V012842) equipped with a sealed tube source and a Hi-Star area detector (Bruker AXS, Madison, WI). The X-Ray generator is operated at a voltage of 40 kV and a current of 35 mA. The powder sample is placed in a nickel holder.

Two frames are registered with an exposure time of 120 s each. The data is subsequently integrated over the range of 4.50-39O 2 theta with a step size of 0.020 and merged into one continuous pattern.

Figure 1d shows the X-ray powder diffractogram of the sample which is characteristic of crystalline drug substance.

Representative XRPD peaks from Compound 1—2 0 HCl 0 H20 XRPD Angle ity % Peaks (2-Theta :: 0.2) *1 6-6 3 16.8 37.8 13.9 27.0 7 13.0 22.7 8 16-5 17.7 20.8 11 31-1 12 15-8 *13 8-1 12.7 17.2 16 16-0 18 20.6 16.0 *20 11.2 15.2 21 33.9 11.3 Thermo Analysis of Compound 1-2 ° HCl ° H20 Thermogravimetric analysis (TGA) for nd 1-2 ° HCl ° H20 was med on the TA Instruments TGA model Q5000. Compound 1—2 ° HCl ° H20 was placed in a platinum sample pan and heated at 10°C/min to 400°C from room temperature.

The thermogram (Figure 2d) demonstrates a weight loss of 2.9% from room temperature to 100°C, and a weight loss of 0.6% from 100°C to 222°C, which is consistent with tical monohydrate (3.5%).

Differential Scanning Calorimetg of Compound 1-2 ° HC1 ° Hgg A DSC thermogram for Compound 1-2 ° HC1 ° H20 was obtained using TA Instruments DSC Q2000. Compound 1—2 0 HCl 0 H20 was heated at 2°C/min to 275°C from - °C, and modulated at i 1°C every 60 sec. The DSC gram (Figure 3d) reveals an endothermic peak below 200°C, which could corresponds to the loss of 1 equivalent of HCl.

Melting/recrystallization occurs between 215—245°C, followed by degradation.

Example 14: Compound I-2 0 HC1 0 2H20 Compound 1—2 ° HCl° 2H20 can be formed from Compound 1-2 ° 2 HC1.

(E29244-17) A suspension of nd 1-2 ° 2 HC1 (10.0 g, 18.6 mmol) in isopropyl alcohol (40 mL) and water (10 mL) is warmed at 50 °C for about 1 h and then cooled to below 10 °C. The solid is collected by filtration. The —cake is washed with 80/20 isopropyl alcohol/water (2 x 10 mL) and air-dried to afford Compound 1-2 ° HCl° 2H20 as a yellow .

XRPD of Compound 1-2 ° HC1 ° 2H;Q The powder X—ray diffraction measurements were performed using PANalytical’s X-pert Pro diffractometer at room temperature with copper ion (1.54060 °A). The incident beam optic was comprised of a variable divergence slit to ensure a constant illuminated length on the sample and on the diffracted beam side, a fast linear solid state detector was used with an active length of 2. 12 degrees 2 theta measured in a scanning mode.

The powder sample was packed on the ed area of a zero background silicon holder and spinning was performed to achieve better statistics. A rical scan was measured from 4 — 40 degrees 2 theta with a step size of 0.017 degrees and a scan step time of 15.5s.

Figure 1d shows the X-ray powder diffractogram of the sample which is characteristic of crystalline drug substance.

] Representative XRPD peaks from Compound 1—2 0 HCl 0 2HZO XRPD Angle (2- Intensity % Peaks Theta :: 0.2) 100.0 82.0 *#WNHOQOONOU‘I-RWNHOQOONOU‘I-bw 69.2 .9 66.3 .5 63.8 17.1 55.3 55-3 7.2 51.8 .7 49.7 NNNNNfi—‘fi—‘fi—‘fi—‘h—‘h—‘h—‘h—‘h—‘t—t 15.3 46.2 .2 44.6 28.0 41.2 19.9 40.4 17.6 39.1 39.0 * 36.1 33.6 33.5 32.2 29.1 28.7 28.0 23.8 * 22.6 22.6 22.4 .8 19.7 19.0 17.4 16.2 .7 .1 14.9 14.8 14.8 13.3 13.1 11.1 2012/058127 Thermo Analysis of Compound 1-2 ° HCl ° 2H_2Q The TGA (Thermogravimetric Analysis) thermographs were obtained using a TA instrument TGA Q500 respectively at a scan rate of 10°C/min over a temperature range of 25—300°C. For TGA analysis, samples were placed in an open pan. The thermogram demonstrates a weight loss of ~6 from room temperature to 100°C, which is consistent with theoretical dihydrate (6.7%).

Differential Scanning Calorimetg of nd 1-2 ° HCl ° 2H;Q A DSC (Differential Scanning metry) thermographs were obtained using a TA instruments DSC Q2000 at a scan rate of 10°C/min over a temperature range of 25- 300°C. For DSC analysis, samples were d into aluminum hermetic T—zero pans that were sealed and punctured with a single hole. The DSC thermogram reveals dehydration between room ature and 120°C followed by melting/recrystallization between 170— 250°C.

Cmstal Structure of Compound 1—2 0 HCl with water 180mg Compound 1—2 ° HCl was added to a vial with 0.8mL 2-propanol and 0.2 mL water.

The sealed vial was kept in an oven at 70°C for two weeks. Diffraction y crystals were observed.

A yellow needle shape crystal with dimensions of 0.15>< 0.02><0.02 mm3 was selected, mounted on a MicroMount and centered on a Bruker APEX II diffractometer (V011510). Then a kapton tube with water inside covered the pin. The tube was sealed to make sure the crystal is brated with water for two days before the diffraction experiments. Three batches of 40 frames separated in reciprocal space were obtained to provide an orientation matrix and initial cell parameters. Final cell parameters were collected and refined was completed based on the full data set.

A diffraction data set of ocal space was obtained to a resolution of 1060 20 angle using 0.5° steps with exposure times 20 s each frame for low angle frames and 60s each frame for high angle frames. Data were collected at room ature. Integration of intensities and refinement of cell parameters were conducted using the APEXII software.

CRYSTAL DATA C24H28C1N504S Mr = 518.02 Monoclinic, P21/n a = 5.4324 (5) A b = 35.483 (4) A c= 13.3478 (12) A B= 100.812 (5)0 V= 2527.2 (4) A3 Example 15: Cellular ATR Inhibition Assay: Compounds can be screened for their ability to inhibit intracellular ATR using an immunofiuorescence microscopy assay to detect phosphorylation of the ATR substrate histone H2AX in hydroxyurea treated cells. HT29 cells are plated at 14,000 cells per well in 96-well black imaging plates (BD 353219) in s 5A media (Sigma M8403) supplemented with 10% foetal bovine serum (JRH Biosciences 12003), Penicillin/Streptomycin solution diluted 1:100 (Sigma P7539), and 2mM L-glumtamine (Sigma G7513), and allowed to adhere overnight at 37°C in 5% C02. Compounds are then added to the cell media from a final concentration of 25HM in 3-fold serial dilutions and the cells are incubated at 37°C in 5% C02. After 15min, hydroxyurea (Sigma H8627) is added to a final tration of 2mM.

After 45min of treatment with hydroxyurea, the cells are washed in PBS, fixed for 10min in 4% formaldehyde diluted in PBS (Polysciences Inc 18814), washed in 0.2% Tween-20 in PBS (wash buffer), and permeabilised for 10min in 0.5% Triton X-100 in PBS, all at room ature. The cells are then washed once in wash buffer and blocked for 30min at room ature in 10% goat serum (Sigma G9023) diluted in wash buffer (block buffer). To detect H2AX phosphorylation levels, the cells are then incubated for 1h at room temperature in primary antibody (mouse monoclonal anti-phosphorylated histone H2AX Serl39 antibody; Upstate 05—636) diluted 1:250 in block buffer. The cells are then washed five times in wash buffer before incubation for 1h at room ature in the dark in a mixture of ary antibody (goat anti-mouse Alexa Fluor 488 conjugated dy; lnvitrogen ) and Hoechst stain (lnvitrogen H3570); diluted 1:500 and 1:5000, respectively, in wash buffer. The cells are then washed five times in wash buffer and finally 100ul PBS is added to each well before imaging.

] Cells are imaged for Alexa Fluor 488 and Hoechst intensity using the BD Pathway 855 Bioimager and Attovision software (BD Biosciences, Version 1.6/855) to quantify orylated H2AX Serl39 and DNA staining, respectively. The percentage of phosphorylated H2AX-positive nuclei in a montage of 9 images at 20x magnification is then calculated for each well using BD Image Data Explorer software (BD Biosciences Version 2.2.15). Phosphorylated H2AX—positive nuclei are defined as Hoechst—positive s of st containing Alexa Fluor 488 intensity at 1.75-fold the average Alexa Fluor 488 intensity in cells not treated with hydroxyurea. The percentage of H2AX positive nuclei is finally plotted against concentration for each compound and IC50s for intracellular ATR inhibition are determined using Prism software (GraphPad Prism version 3.0cx for Macintosh, GraphPad Software, San Diego rnia, USA).

The compounds described herein can also be tested according to other methods known in the art (& Sarkaria et al, “Inhibition ofATM and ATR Kinase Activities by the Radiosensitizing Agent, Caffeine: Cancer Research 59: 4375—5382 ; Hickson et al, “Identification and terization of a Novel and Specific Inhibitor of the Ataxia- iectasia Mutated Kinase ATM” Cancer Research 64: 9152—9159 (2004); Kim et al, “Substrate Specificities and Identification of Putative Substrates of ATM Kinase Family Members” The Journal ogical Chemistry, 274(53): 37543 (1999); and Chiang et al, “Determination of the catalytic ties of mTOR and other members of the phosphoinositidekinase-related kinase family” Methods M01. Biol. 281 : 125—41 (2004)).

Example 16: ATR Inhibition Assay: Compounds can be screened for their ability to inhibit ATR kinase using a ctive-phosphate incorporation assay. Assays are carried out in a mixture of 50mM Tris/HCl (pH 7.5), 10mM MgClz and 1mM DTT. Final substrate concentrations are 10uM [y-33P]ATP (3mCi 33P ATP/mmol ATP, Perkin Elmer) and 800 ”M target peptide (ASELPASQPQPFSAKKK).

Assays are carried out at 25°C in the presence of 5 nM full—length ATR. An assay stock buffer solution is prepared containing all of the reagents listed above, with the exception of ATP and the test compound of interest. 13.5 [1L of the stock on is placed in a 96 well plate followed by addition of 2 [1L of DMSO stock containing serial dilutions of the test compound (typically starting from a final concentration of 15 ”M with 3-fold serial dilutions) in duplicate (final DMSO concentration 7%). The plate is pre-incubated for 10 minutes at 25°C and the reaction initiated by on of 15 [LL [y-33P]ATP (final WO 49726 concentration 10 uM).

The reaction is stopped after 24 hours by the addition of 30uL 0.1M phosphoric acid ning 2mM ATP. A multiscreen phosphocellulose filter 96-well plate (Millipore, Cat no. MAPHNOB50) is pretreated with 100uL 0.2M phosphoric acid prior to the addition of 45 [LL of the stopped assay mixture. The plate is washed with 5 x 200uL 0.2M phosphoric acid. After drying, 100 [1L Optiphase ‘SuperMix’ liquid scintillation cocktail (Perkin Elmer) is added to the well prior to scintillation counting (1450 Microbeta Liquid Scintillation Counter, Wallac).

After removing mean background values for all of the data points, Ki(app) data are calculated from non-linear regression analysis of the initial rate data using the Prism software e (GraphPad Prism version 3.0cx for Macintosh, GraphPad Software, San Diego California, USA).

In general, the compounds of the t invention are effective for inhibiting ATR.

Compounds 1-1, 1-2, 11-1, 11-2, 11-3 and 11-4 t ATR at Ki values below 0.001 uM.

Example 17: Cisplatin Sensitization Assay Compounds can be screened for their ability to sensitize HCT116 colorectal cancer cells to tin using a 96h cell viability (MTS) assay. HCT116 cells, which possess a defect in ATM signaling to Cisplatin (see, Kim et al.; Oncogene 21 :3864 (2002); see also, Takemura et al.; JBC 281:30814 (2006)) are plated at 470 cells per well in 96—well polystyrene plates (Costar 3596) in 150ul of McCoy’s 5A media (Sigma M8403) supplemented with 10% foetal bovine serum (JRH Biosciences 12003), Penicillin/Streptomycin solution diluted 1:100 (Sigma P7539), and 2mM L-glumtamine (Sigma , and allowed to adhere overnight at 37°C in 5% C02. Compounds and Cisplatin are then both added simultaneously to the cell media in 2-fold serial dilutions from a top final concentration of 10uM as a full matrix of concentrations in a final cell volume of 200ul, and the cells are then incubated at 37°C in 5% C02. After 96h, 40ul of MTS t (Promega G358a) is added to each well and the cells are ted for 1h at 37°C in 5% C02. y, absorbance is measured at 490nm using a aMax Plus 384 reader (Molecular Devices) and the concentration of compound required to reduce the IC50 of Cisplatin alone by at least 3—fold (to 1 decimal place) can be reported.

Example 18: Single Agent HCT116 ty ] Compounds can be screened for single agent activity against HCT116 colorectal cancer cells using a 96h cell viability (MTS) assay. HCT116 are plated at 470 cells per well in l yrene plates (Costar 3596) in 150ul of s 5A media (Sigma M8403) supplemented with 10% foetal bovine serum (JRH Biosciences 12003), Penicillin/ Streptomycin solution diluted 1:100 (Sigma P7539), and 2mM L-glumtamine (Sigma G7513), and allowed to adhere overnight at 37°C in 5% C02. Compounds are then added to the cell media in 2-fold serial dilutions from a top final concentration of lOuM as a full matrix of concentrations in a final cell volume of 200ul, and the cells are then incubated at 37°C in 5% C02. After 96h, 40ul of MTS reagent (Promega G358a) is added to each well and the cells are incubated for 1h at 37°C in 5% C02. Finally, absorbance is measured at 490nm using a SpectraMax Plus 384 reader (Molecular Devices) and IC50 values can be calculated.

Data for Examples 18-21 ATR inhibition ATR cellular Cisplatin Ki (nM) lC50 (nM) sensitization While we have described a number of embodiments of this invention, it is apparent that our basic examples may be altered to provide other embodiments that e the compounds, methods, and processes of this invention. Therefore, it will be appreciated that the scope of this ion is to be defined by the appended claims rather than by the specific embodiments that have been represented by way of example herein.

Claims (25)

1. A process for preparing a compound of formula I-2: comprising the steps of a) reacting a compound of formula 4-i under chlorooxime formation ions to form the compound of formula 4-ii: b) reacting the compound of formula 4-ii with a compound of formula 4-iii: 4-iii under ddition conditions to form the compound of formula 4-iv: c) reacting a compoundof formula 4-iv with under coupling conditions to form the nd of formula 5-I: and deprotecting the compound of formula 5-I under Boc deprotection conditions optionally followed by treatment under basic aqueous conditions to form a compound of a I-2.

2. The process of claim 1, further comprising the step of reacting a compound of formula 3b: under oxime formation ions to form the compound of formula 4-i.

3. The process of any one of claims 2 further comprising the step of reacting a compound of formula 2b: under Boc protection conditions to form the compound of a 3b.

4. The process of claim 3, further comprising the step of reacting a compound of formula 1b: with amine under reductive amination conditions to form the compound of formula 2b.

5. The process of claim 1, further comprising the step a) ng a compound of formula C-1: under metal-mediated couplings conditions with TMS-acetylene to form a nd of formula C-2: b) reacting a compound of formula C-2 under Boc protection conditions to form a nd of formula C-3: c) reacting a compound of formula C-3 TMS ection conditions to form a compound of formula Aii.

6. The process of claim 1, wherein the coupling conditions comprise combining a palladium catalyst with a base in a solvent.

7. The process of claim 6, wherein the palladium catalyst is selected from Pd[P(tBu)3]2, Pd(dtbpf)Cl2, Pd(PPh3)2Cl2, Pd(PCy3)2Cl2 , Pd(dppf)Cl2, and Pd(dppe)Cl2; the solvent is selected from one or more of the following: toluene, MeCN, water, EtOH, IPA, 2- Me-THF, or IPAc; and the base is selected from K2CO3, Na2CO3, or K3PO4.

8. The process of claim 1, wherein the cycloaddition conditions comprises a base selected from pyridine, DIEA, TEA, t-BuONa, or K2CO3 and a solvent ed from acetonitrile, tetrahydrofuran, 2-methyltetrahydrofuran, MTBE, EtOAc, i-PrOAc, DCM, toluene, DMF, and methanol.

9. The process of claim 1, wherein the chlorooxime formation conditions comprise adding HCl in dioxane to a solution of the oxime in the ce of NCS in a t selected from DCM, DCE, THF, dioxane, an ic arbon, and an alkyl acetate.

10. The process of claim 9, wherein the oxime formation conditions consist of either a single step sequence or a two step sequence.

11. The s of claim 10, wherein the single step sequence comprises adding NH2OH.HCl to a mixture of THF and water.

12. The process of claim 11, wherein 1 equivalent of the compound of formula 3b is combined with a 1.1 equivalents of NH2OH.HCl in a 10:1 v/v mixture of THF/water.

13. The process of claim 10, wherein the two step sequence consists of first deprotecting the ketal group in the compound of formula 3b: into an de under deprotection conditions, and then forming the oxime of formula 4i: under oxime formation ions.

14. The process of claim 13, wherein a) deprotection conditions comprise adding an acid, acetone, and water; and b) oxime formation conditions comprise mixing er hydroxylamine, an optional acid, an organic solvent, and water.

15. The s of claim 14, wherein the acid is pTSA or HCl, the organic solvent a chlorinated solvents selected from dichloromethane (DCM), roethane (DCE), CH 2Cl 2, and chloroform; an ether selected from THF, 2-MeTHF and dioxane; or an aromatic hydrocarbons selected from toluene and xylenes.

16. The s of claim 15, wherein oxime formation conditions comprise adding hydroxylamine hydrochloride to a solution of the compound of 3b in THF and water.

17. The s of claim 4, wherein the reductive amination conditions comprise adding a reducing agent selected from NaBH4 NaBH4, NaBH3CN, or NaBH(OAc)3 in the presence of a solvent selected from dichloromethane, dichloroethane, an alcoholic t selected from methanol, ethanol, 1-propanol, isopropanol, or a nonprotic solvent selected from e, tetrahydrofuran, or 2-methyltetrahydrofuran and optionally a base selected from Et3N or diisopropylethylamine.

18. The process of any one of claim 1, 2, and 3-4, wherein the Boc ection conditions comprises adding a Boc deprotecting agent selected from TMS-Cl, HCl, TBAF, H3PO 4, or TFA and the solvent is selected from acetone, toluene, ol, ethanol, 1-propanol, isopropanol, CH2Cl 2, EtOAc, pyl acetate, tetrahydrofuran, 2-methyltetraydrofuran, dioxane, or diethylether.

19. The process of claim 18, wherein the Boc deprotecting agent is HCl or TFA and the solvent is acetone or CH2Cl 2.

20. The process of claim 3, wherein the Boc protection conditions comprise adding (Boc) 2O, a base, and a solvent.

21. The process of claim 20, wherein the base is Et3N, diisopropylamine, and pyridine; and the solvent is selected from a chlorinated solvent, an ether, or an aromatic hydrocarbon.

22. The process of claim 21, wherein the base is Et3N, the solvent is DCM, tetrahydrofuran, or 2-methyltetrahydrofuran.

23. The process of claim 22, n the Boc protection conditions comprise adding 1.05 equivalents of (Boc)2O per equivalent of nd 2b in 2-methyltetrahydrofuran or DCM.

24. The process of claim 5, wherein the metal-mediated coupling conditions are Sonogashira coupling conditions; the Boc protection conditions comprise adding (Boc) 2O in 2-methyltetrahydrofuran or DCM; and the TMS deprotection conditions se adding a base, organic solvent, and water.

25. A process according to claim 1, substantially as herein described with nce to any one of the accompanying examples and or figures thereof.