NOMURA DANIEL (US)
BOIKE LYDIA (US)
MARQUESS DANIEL (US)
KEITZ PAUL (US)
HENNING NATHANIEL JAMES (US)
NOMURA DANIEL K (US)
BOIKE LYDIA (US)
MARQUESS DANIEL (US)
KEITZ PAUL (US)
CLAIMS 1. A bifunctional compound of Formula (I-c): or a pharmaceutically acceptable salt, hydrate, solvate, stereoisomer, or tautomer thereof, wherein: (i) the CFTR Ligand comprises a moiety capable of binding to the cystic fibrosis transmembrane conductance regulator (CFTR), or a mutant, fragment, or isoform thereof; (ii) L1 comprises a linker; and (iii) the OTUB1 Recruiter comprises a moiety capable of binding to the deubiquitinase OTUB1. 2. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 1, wherein the CFTR is mutated or misfolded. 3. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 1, wherein the CFTR is ubiquitinated (e.g., polyubiquitinated). 4. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 1, wherein the CFTR is ΔF508-CFTR. 5. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 1, wherein the OTUB1 Recruiter covalently binds to OTUB1. 6. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 1, wherein the OTUB1 Recruiter binds to a site other than a catalytic site within OTUB1. 7. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 1, wherein the OTUB1 Recruiter binds to an allosteric site within OTUB1. 8. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 1, wherein the OTUB1 Recruiter binds to a cysteine amino acid residue within OTUB1. 9. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 8, wherein the cysteine amino acid residue is an allosteric cysteine amino acid residue. 10. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 1, wherein the OTUB1 Recruiter preferentially binds to an allosteric amino acid residue (e.g., an allosteric amino acid residue) over a catalytic amino acid residue (e.g., a catalytic cysteine amino acid residue). 11. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 1, wherein the OTUB1 Recruiter does not substantially bind to a cysteine amino acid residue in the catalytic site of OTUB1 (e.g., a catalytic cysteine). 12. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 10, wherein OTUB1 Recruiter binds to cysteine 23 (C23) within the OTUB1 sequence. 13. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 1, wherein OTUB1 Recruiter binds preferentially to cysteine 23 (C23) over cysteine 91 (C91) within the OTUB1 sequence. 14. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 1, wherein the OTUB1 Recruiter does not substantially bind to cysteine 91 (C91) within the OTUB1 sequence. 15. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 1, wherein the CFTR Ligand binds to (e.g., covalently binds to) CFTR, or a mutant, fragment, or isoform thereof. 16. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 1, wherein the CFTR Ligand is capable of modulating CFTR, or a mutant, fragment, or isoform thereof. 17. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 16, wherein the modulating comprises one or more of: (i) modulating the folding of the target protein; (ii) modulating the half-life of the target protein; (iii) modulating trafficking of the target protein to the proteasome; (iv) modulating the level of ubiquitination of the target protein; (v) modulating degradation (e.g., proteasomal degradation) of the target protein; (vi) modulating target protein signaling; (vii) modulating target protein localization; (viii) modulating trafficking of the target protein to the lysosome; (ix) modulating trafficking of the target protein to the ER, Golgi, vesicle, plasma membrane; (x) modulating target protein interactions with another protein (e.g., other proteins in the UPS); and (xi) modulating posttranslational modifications of the target protein (e.g., SUMOlyation, phosphorylation, glycosylation). 18. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 1, wherein the CFTR Ligand has the structure of Formula (II-a): or a pharmaceutically acceptable salt, hydrate, solvate, prodrug, stereoisomer, or tautomer thereof, wherein: X and Z are each independently O, S, or C(R7a)(R7b); Y is C(R7a)(R7b) or NR7c; R1 is H or C1–6 alkyl; R3a, R3b, R4a, R4b are each independently H, C1–6 alkyl, C1–6 haloalkyl, C1–6 heteroalkyl, halo, cyano, or -ORA; each R5, R5’, and R6 is independently C1–6 alkyl, C1–6 haloalkyl, C1–6 heteroalkyl, halo, cyano, -ORA, -C(O)N(RB)(RC), or -N(RB)CO(RD); R7a and R7b are each independently H, C1–6 alkyl, C1–6 haloalkyl, C1–6 heteroalkyl, or halo; R7c is H or C1–6 alkyl; RA, RB, RC, and RD are each independently H, C1–6 alkyl, C2–6 alkenyl, C2–6 alkynyl, C1–6 haloalkyl, C1–6 heteroalkyl, cycloalkyl, heterocyclyl, aryl, or heteroaryl; p is 0, 1, 2, 3, or 4; p’ is 0, 1, 2, 3, or 4; q is 0, 1, 2, or 3; and denotes the point of attachment to L1 in Formula (I). 19. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 18, wherein each of X and Z is independently O. 20. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 18, wherein Y is C(R7a)(R7b). 21. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 18, wherein each of R7a and R7b is independently halo (e.g., fluoro). 22. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 18, wherein each of R3a, R3b, R4a, R4b is independently H. 23. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 18, wherein R1 is H. 24. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 18, wherein each of p and q is 0. 25. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 1, wherein the CFTR Ligand has the structure of Formula (II-f): or a pharmaceutically acceptable salt, hydrate, solvate, prodrug, stereoisomer, or tautomer thereof, wherein denotes the point of attachment to L1 in Formula (I). 26. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 1, wherein the bifunctional compound of Formula (I-c) has the structure (II-l): or a pharmaceutically acceptable salt, hydrate, solvate, prodrug, stereoisomer, or tautomer thereof, wherein: X and Z are each independently O, S, or C(R7a)(R7b); Y is C(R7a)(R7b) or NR7c; R1 is H or C1–6 alkyl; R3a, R3b, R4a, R4b are each independently H, C1–6 alkyl, C1–6 haloalkyl, C1–6 heteroalkyl, halo, cyano, or -ORA; each R5, R5’, and R6 is independently C1–6 alkyl, C1–6 haloalkyl, C1–6 heteroalkyl, halo, cyano, -ORA, -C(O)N(RB)(RC), or -N(RB)CO(RD); R7a and R7b are each independently H, C1–6 alkyl, C1–6 haloalkyl, C1–6 heteroalkyl, or halo; R7c is H or C1–6 alkyl; RA, RB, RC, and RD are each independently H, C1–6 alkyl, C2–6 alkenyl, C2–6 alkynyl, C1–6 haloalkyl, C1–6 heteroalkyl, cycloalkyl, heterocyclyl, aryl, or heteroaryl; p is 0, 1, 2, 3, or 4; p’ is 0, 1, 2, 3, or 4; q is 0, 1, 2, or 3; and L1 and OTUB Recruiter are as defined in claim 1. 27. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 1, wherein the OTUB1 Recruiter binds to (e.g., covalently binds to) OTUB1. 28. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 1, wherein the binding of the OTUB1 Recruiter to OTUB1 does not substantially inhibit the activity of OTUB1. 29. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 1, wherein the OTUB1 Recruiter binds to a site other than a catalytic site within OTUB1. 30. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 1, wherein the OTUB1 Recruiter binds to an allosteric site within OTUB1. 31. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 1, wherein the OTUB1 Recruiter binds to a cysteine amino acid residue within OTUB1. 32. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 1, wherein the OTUB1 Recruiter comprises an acrylamide moiety. 33. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 1, wherein the OTUB1 Recruiter comprises a furan moiety. 34. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 1, wherein the OTUB1 Recruiter has the structure of Formula (V-a): or a pharmaceutically acceptable salt, hydrate, solvate, prodrug, stereoisomer, or tautomer thereof, wherein Ring Z is cycloalkyl, heterocyclyl, aryl, or heteroaryl, each of which is substituted with 0- 12 R4; L1 is absent, -O-, C1–12 alkylene, C2–12 alkenylene, C2–12 alkynylene, C1–12 heteroalkyl, wherein each alkylene, alkenylene, and heteroalkyl is optionally substituted with one or more R5; each R1 is independently C1–6 alkyl, C1–6 haloalkyl, C1–6 heteroalkyl, halo, -ORA, - C(O)RA, -C(O)ORA, -NRBRC, -NRBC(O)RA, -C(O)NRBRC, cycloalkyl, heterocyclyl, aryl, or heteroaryl, wherein each of alkyl, haloalkyl, heteroalkyl, cycloalkyl, heterocyclyl, aryl and heteroaryl is optionally substituted with 0-12 R5; R2 is H, C1–6 alkyl, or an electrophilic moiety (e.g., C2–6 alkenyl); each R3 is independently C1–6 alkyl, C1–6 haloalkyl, C1–6 heteroalkyl, halo, or -ORA, wherein each alkyl, haloalkyl and heteroalkyl is optionally substituted with 1-6 R6; or two R3 are taken together with the atoms to which they are attached to form an oxo, cycloalkyl, or heterocyclyl, wherein each cycloalkyl and heterocyclyl is optionally substituted with 1-6 R6; each R4, R5 and R6 is independently C1–6 alkyl, C1–6 haloalkyl, C1–6 heteroalkyl, halo, cyano, oxo, -ORA, -C(O)RA, -C(O)ORA, -NRBRC, -NRBC(O)RA, -C(O)NRBRC, or wherein two of R4, R5, or R6 are taken together with the atoms to which they are attached to form a cycloalkyl, heterocyclyl, aryl, or heteroaryl; RA is H, C1–6 alkyl, C2–6 alkenyl, C2–6 alkynyl, C1–6 haloalkyl, C1–6 heteroalkyl, halo, cycloalkyl, heterocyclyl, aryl, or heteroaryl; each RB and RC is independently H, C1–6 alkyl, C2–6 alkenyl, C2–6 alkynyl, C1–6 haloalkyl, C1–6 heteroalkyl, cycloalkyl, heterocyclyl, aryl, or heteroaryl; and n is 0, 1, 2, 3, 4, or 6. 35. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 34, wherein Ring Z is heteroaryl (e.g., a monocyclic heteroaryl). 36. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 34, wherein Ring Z is a 5-membered heteroaryl (e.g., furanyl). 37. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 34, wherein Ring Z is selected from the following group: 38. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 34, wherein Ring Z is selected from . 39. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 34, wherein L1 is absent, C1-6 alkylene, C2-6 alkenylene, C2-6 alkynylene, 40. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 34, wherein L1 is absent or C1-6 alkylene. 41. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 34, wherein R1 is absent, aryl, heteroaryl, - C(O)ORA, -C(O)NH-RC, -ORA, or -NRBRC. 42. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 34, wherein R2 is an electrophilic moiety. 43. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 34, wherein R2 is C2–6 alkenyl (e.g., CH=CH2). 44. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 1, wherein the OTUB1 Recruiter has the structure of Formula (V-p): or a pharmaceutically acceptable salt, hydrate, solvate, prodrug, stereoisomer, or tautomer thereof, wherein Ring Z, R2, R3, R5, n, and subvariables thereof are as described for Formula (V- a), Ring W is cycloalkyl, heterocyclyl, aryl, or heteroaryl, wherein each of cycloalkyl, heterocyclyl, aryl and heteroaryl is optionally substituted with 0-12 R5; and p is 0, 1, 2, 3, 4, 5, or 6. 45. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 1, wherein the OTUB1 Recruiter binds to cysteine 23 (C23) within the OTUB1 sequence. 46. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 1, wherein the OTUB1 Recruiter binds preferentially to cysteine 23 (C23) over cysteine 91 (C91) within the OTUB1 sequence. 47. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 1, wherein the OTUB1 Recruiter does not substantially bind to cysteine 91 (C91) within the OTUB1 sequence. 48. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 1, wherein L1 is a non-cleavable linker. 49. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 1, wherein L1 comprises an alkylene or heteroalkylene. 50. The bifunctional compound or pharmaceutically acceptable salt hydrate, solvate, stereoisomer, or tautomer thereof according to claim 1, wherein the bifunctional compound of Formula (I-c) is selected from a bifunctional compound provided in Table 1, or a pharmaceutically acceptable salt, hydrate, solvate, prodrug, stereoisomer, or tautomer thereof. 51. A pharmaceutical composition comprising a bifunctional compound, or pharmaceutically acceptable salt, solvate, stereoisomer, or tautomer thereof according to any one of the preceding claims, and one or more pharmaceutically acceptable carriers. 52. A composition for use in providing a compound to a subject, wherein the composition comprises a bifunctional compound of Formula (I): or a pharmaceutically acceptable salt, hydrate, solvate, stereoisomer, or tautomer thereof, wherein: (i) the CFTR Ligand comprises a moiety capable of binding to the cystic fibrosis transmembrane conductance regulator (CFTR), or a mutant, fragment, or isoform thereof; (ii) L1 comprises a linker; and (iii) the OTUB1 Recruiter comprises a moiety capable of binding to the deubiquitinase OTUB1. 53. A composition for use in treating a disease, disorder, or condition in a subject, comprising a bifunctional compound of Formula (I): or a pharmaceutically acceptable salt, hydrate, solvate, stereoisomer, or tautomer thereof, wherein: (i) the CFTR Ligand comprises a moiety capable of binding to the cystic fibrosis transmembrane conductance regulator (CFTR), or a mutant, fragment, or isoform thereof; (ii) L1 comprises a linker; and (iii) the OTUB1 Recruiter comprises a moiety capable of binding to the deubiquitinase OTUB1. 54. The composition for use of claim 53, wherein administering the composition ameliorates a symptom or element of the disease, disorder, or condition. 55. The composition for use of claim 53, wherein the disease, disorder, or condition is cystic fibrosis. 56. A composition for use in treating cystic fibrosis in a subject, comprising a bifunctional compound of Formula (I): (I), or a pharmaceutically acceptable salt, hydrate, solvate, stereoisomer, or tautomer thereof, wherein: (i) the CFTR Ligand comprises a moiety capable of binding to the cystic fibrosis transmembrane conductance regulator (CFTR), or a mutant, fragment, or isoform thereof; (ii) L1 comprises a linker; and (iii) the OTUB1 Recruiter comprises a moiety capable of binding to the deubiquitinase OTUB1. 57. A composition for use in modulating a protein in a cell or subject comprising a bifunctional compound of Formula (I): (I), or a pharmaceutically acceptable salt, hydrate, solvate, stereoisomer, or tautomer thereof, wherein: (i) the CFTR Ligand comprises a moiety capable of binding to the cystic fibrosis transmembrane conductance regulator (CFTR), or a mutant, fragment, or isoform thereof; (ii) L1 comprises a linker; and (iii) the OTUB1 Recruiter comprises a moiety capable of binding to the deubiquitinase OTUB1. 58. A composition for use in recruiting a deubiquitinase to a target protein in a cell or subject, wherein the composition comprises a bifunctional compound of Formula (I): or a pharmaceutically acceptable salt, hydrate, solvate, stereoisomer, or tautomer thereof, wherein: (i) the CFTR Ligand comprises a moiety capable of binding to the cystic fibrosis transmembrane conductance regulator (CFTR), or a mutant, fragment, or isoform thereof; (ii) L1 comprises a linker; and (iii) the OTUB1 Recruiter comprises a moiety capable of binding to the deubiquitinase OTUB1. 59. The composition for use of claim 58, wherein the deubiquitinase is OTUB1. 60. A composition for use in deubiquitinating a protein comprising a bifunctional compound of Formula (I): (I), or a pharmaceutically acceptable salt, hydrate, solvate, stereoisomer, or tautomer thereof, wherein: (i) the CFTR Ligand comprises a moiety capable of binding to the cystic fibrosis transmembrane conductance regulator (CFTR), or a mutant, fragment, or isoform thereof; (ii) L1 comprises a linker; and (iii) the OTUB1 Recruiter comprises a moiety capable of binding to the deubiquitinase OTUB1. 61. A method of treating a disease, disorder, or condition in a subject, wherein the method comprises administering to the subject a bifunctional compound of Formula (I): or a pharmaceutically acceptable salt, hydrate, solvate, stereoisomer, or tautomer thereof, wherein: (i) the CFTR Ligand comprises a moiety capable of binding to the cystic fibrosis transmembrane conductance regulator (CFTR), or a mutant, fragment, or isoform thereof; (ii) L1 comprises a linker; and (iii) the OTUB1 Recruiter comprises a moiety capable of binding to the deubiquitinase OTUB1, thereby treating disease, disorder, or condition in the subject 62. The method of claim 61, wherein the disease, disorder, or condition is cystic fibrosis. 63. A method of treating cystic fibrosis in a subject, the method comprising administering to the subject a bifunctional compound of Formula (I): or a pharmaceutically acceptable salt, hydrate, solvate, stereoisomer, or tautomer thereof, wherein: (i) the CFTR Ligand comprises a moiety capable of binding to the cystic fibrosis transmembrane conductance regulator (CFTR), or a mutant, fragment, or isoform thereof; (ii) L1 comprises a linker; and (iii) the OTUB1 Recruiter comprises a moiety capable of binding to the deubiquitinase OTUB1, thereby treating cystic fibrosis. |
To a solution of 3-benzyl-1-imidazo[1,2-a]pyridin-2-yl-piperazin-2-one (A31, 60 mg, 196 µmol, 1 eq) in DCM (1 mL) was added TEA (39.6 mg, 391.7 µmol, 54.5 uL, 2 eq), followed by addition of prop-2-enoyl chloride (17.7 mg, 196 µmol, 16 uL, 1 eq) in DCM (0.5 mL) at 0 °C. The mixture was stirred at 0 °C for 1 h. Upon completion, the reaction mixture was quenched by addition H2O (0.5 mL) at 0 °C, and then concentrated under reduced pressure to give a residue, which was purified by prep-HPLC (Condition 1, Gradient d) to give 3-benzyl-1-imidazo[1,2- a]pyridin-2-yl-4-prop-2-enoyl-piperazin-2-one (Compound 109, 14.7 mg, 38.1 umol) as a solid. MS (ESI) m/z 361.1 [M+H] + . 1 H NMR (400 MHz, DMSO-d6) δ ppm 8.63 - 8.59 (m, 1 H), 8.43 - 8.40 (m, 1 H), 7.50 - 7.47 (m, 1 H), 7.27 - 7.18 (m, 6 H), 7.14 - 6.80 (m, 1 H), 6.77 - 6.31 (m, 1 H), 6.25 - 5.89 (m, 1 H), 5.75 - 5.38 (m, 1 H), 5.35 - 5.00 (m, 1 H), 4.58 - 4.30 (m, 1 H), 4.11 - 3.91 (m, 1 H), 3.91 - 3.76 (m, 1 H), 3.27 - 3.13 (m, 3 H). Example 10: Synthesis of Compound 110 Step 1: tert-butyl 3-oxo-4-([1,2,4]triazolo[4,3-a]pyridin-3-yl)piperazine-1-car boxylate A mixture of 3-bromo-[1,2,4]triazolo[4,3-a]pyridine (A31, 500 mg, 2.5 mmol, 1 eq), tert- butyl 3-oxopiperazine-1-carboxylate (A2, 607 mg, 3.0 mmol, 1.2 eq), Cs2CO3 (1.65 g, 5.0 mmol, 2 eq), iodocopper;tetrabutylammonium;diiodide (56.5 mg, 50.5 µmol, 0.02 eq) and 3,4,7,8- tetramethyl-1,10-phenanthroline (11.9 mg, 50.5 µmol, 0.02 eq) in dioxane (5 mL) was degassed and purged with N 2 (x3), and then the mixture was stirred at 90 °C for 16 h under N 2 atmosphere. The reaction mixture was quenched by addition H2O (0.5 mL) at 0 °C, and then concentrated under reduced pressure to give a residue, which was purified by prep-HPLC (Condition 6, Gradient a) to give a tert-butyl 3-oxo-4-([1,2,4]triazolo[4,3-a]pyridin-3-yl)piperazine-1-car boxylate (A32, 380 mg, 1.20 mmol) as a solid. MS (ESI) m/z 318.2 [M+H] + . To a solution of tert-butyl 3-oxo-4-([1,2,4]triazolo[4,3-a]pyridin-3-yl)piperazine-1- carboxylate (A32, 100 mg, 315 µmol, 1 eq) in DCM (2 mL) was added TFA (770 mg, 6.75 mmol, 0.5 mL, 21.4 eq), and the mixture was stirred at 20 °C for 1 h. The reaction mixture was concentrated under reduced pressure to give a residue, which contained 1-([1,2,4]triazolo[4,3- a]pyridin-3-yl)piperazin-2-one (A33, 70 mg, crude) as an oil. MS (ESI) m/z 218.2 [M+H] + . S To a solution of 1-([1,2,4]triazolo[4,3-a]pyridin-3-yl)piperazin-2-one (A33, 70 mg, 322.2 µmol, 1 eq) in DCM (2 mL) was added TEA (130.4 mg, 1.3 mmol, 179.4 uL, 4 eq) and prop-2- enoyl prop-2-enoate (40.6 mg, 322.2 µmol, 1 eq), and the mixture was stirred at -40 °C for 1 h. The reaction mixture was then concentrated under reduced pressure to give a residue, which was purified by prep-HPLC (Condition 5, Gradient b) to give 4-prop-2-enoyl-1-([1,2,4]triazolo[4,3- a]pyridin-3-yl)piperazin-2-one (Compound 110, 10 mg, 37 umol) as a solid. MS (ESI) m/z 272.2 NMR (400 MHz, CDCl 3 -d): δ = 7.87 - 7.61 (m, 2H), 7.33 (br t, J = 7.6 Hz, 1H), 6.96 - 6.86 (m, 1H), 6.70 - 6.54 (m, 1H), 6.52 - 6.38 (m, 1H), 5.88 (br d, J = 9.8 Hz, 1H), 4.60 (br s, 2H), 4.33 - 3.97 ppm (m, 4H). Example 11: Synthesis of Compound 113 Step 1: tert-butyl 4-imidazo[1,2-a]pyridin-7-yl-3-oxo-piperazine-1-carboxylate A mixture of 7-bromoimidazo[1,2-a]pyridine (A40, 300 mg, 1.52 mmol, 1 eq), tert-butyl 3-oxopiperazine-1-carboxylate (305 mg, 1.52 mmol, 1 eq), CuI (43.5 mg, 228 µmol, 0.15 eq), DMEDA (33.6 mg, 381 µmol, 41 uL, 0.25 eq) and K 2 CO 3 (421 mg, 3.05 mmol, 2 eq) in dioxane (3 mL) was degassed and purged with N2 (x3), and then the mixture was stirred at 100 °C for 12 h under N2 atmosphere. Upon completion, the reaction mixture was quenched by addition H2O (30 mL) at 20 °C, and extracted with EtOAc (30 mL x3). The combined organic layers were washed with brine (30 mL x2), dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue that was purified by prep-TLC (50% EtOAc in PE) to afford tert-butyl 4- imidazo[1,2-a]pyridin-7-yl-3-oxo-piperazine-1-carboxylate (A41, 280 mg, 885 umol) was obtained as a solid. MS (ESI) m/z 316.3 [M+H] + . Step 2: 1-imidazo[1,2-a]pyridin-7-ylpiperazin-2-one A solution of tert-butyl 4-imidazo[1,2-a]pyridin-7-yl-3-oxo-piperazine-1-carboxylate (A41, 150 mg, 474 µmol, 1 eq) in DCM (5 mL) was added TFA (1.54 g, 13.5 mmol, 1 mL, 28.5 eq) at 20 °C and stirred at 20 °C for 0.5 h. Upon completion, the reaction mixture was concentrated under reduced pressure to give A42 (1-imidazo[1,2-a]pyridin-7-ylpiperazin-2-one, 156 mg, crude, TFA) as an oil. MS (ESI) m/z 217.2 [M+H] + . Step 3: 1-imidazo[1,2-a]pyridin-7-yl-4-prop-2-enoyl-piperazin-2-one To a solution of 1-imidazo[1,2-a]pyridin-7-ylpiperazin-2-one (A42, 156 mg, 472.4 µmol, 1 eq, TFA) in DCM (3 mL) was added TEA (143.4 mg, 1.42 mmol, 197.2 uL, 3 eq) and prop-2- enoyl chloride (47 mg, 519.6 µmol, 42.4 uL, 1.1 eq) at 0 °C. The mixture was stirred at 0 °C for 1 h. Upon completion, the reaction mixture was quenched by addition MeOH (0.5 mL) at 20 °C. The reaction mixture was concentrated under reduced pressure to remove solvent, and the residue was purified by prep-HPLC (Condition 2, Gradient b) to afford 1-imidazo[1,2-a]pyridin-7-yl-4-prop- 2-enoyl-piperazin-2-one (Compound 113, 15.2 mg, 55.9 umol) as a solid. MS (ESI) m/z 271.2 [M+H] + . 1 H NMR (400 MHz, DMSO-d 6 ) δ = 8.52 (d, J = 7.3 Hz, 1H), 7.93 (s, 1H), 7.57 (s, 2H), 7.00 (br d, J = 7.1 Hz, 1H), 6.93 - 6.76 (m, 1H), 6.21 (br d, J = 16.6 Hz, 1H), 5.77 (br d, J = 10.3 Hz, 1H), 4.44 (s, 1H), 4.29 (s, 1H), 3.99 (br d, J = 4.8 Hz, 1H), 3.94 - 3.78 (m, 3H). Example 12: Synthesis of Compound 114 Step 1: tert-butyl 3-oxo-4-([1,2,4]triazolo[1,5-a]pyridin-6-yl)piperazine-1-car boxylate A mixture of 6-bromo-[1,2,4]triazolo[1,5-a]pyridine (A43, 500 mg, 2.52 mmol, 1 eq), tert- butyl 3-oxopiperazine-1-carboxylate (A2, 505.6 mg, 2.52 mmol, 1 eq), CuI (72 mg, 379 µmol, 0.15 eq), DMEDA (55.7 mg, 631.3 µmol, 67.9 uL, 0.25 eq) and K2CO3 (697.9 mg, 5.05 mmol, 2 eq) in dioxane (7 mL) was degassed and purged with N 2 (x3), and then the mixture was stirred at 100 °C for 16 h under N 2 atmosphere. The reaction mixture was quenched by water (12 mL), and then extracted with EtOAc (15 mL x3). The combined organic layers were washed with brine (15 mL), dried over Na2SO4, filtered and concentrated under reduced pressure to give a that residue was purified by column chromatography (17-100% EtOAc in PE) to give tert-butyl 3-oxo-4- ([1,2,4]triazolo[1,5-a]pyridin-6-yl)piperazine-1-carboxylate (A44, 350 mg, 1.10 mmol) as a solid. MS (ESI) m/z 318.3 [M+H] + . Step 2: 1-([1,2,4]triazolo[1,5-a]pyridin-6-yl)piperazin-2-one To a solution of tert-butyl 3-oxo-4-([1,2,4]triazolo[1,5-a]pyridin-6-yl)piperazine-1- carboxylate (A44, 350 mg, 1.10 mmol, 1 eq) in DCM (4 mL) was added TFA (1.54 g, 13.51 mmol, 1 mL, 12.25 eq). The mixture was stirred at 20 °C for 1 h. The reaction mixture was concentrated under reduced pressure to give a residue that yielded 1-([1,2,4]triazolo[1,5-a]pyridin-6- yl)piperazin-2-one (A45, 240 mg, crude) as an oil. MS (ESI) m/z 218.2 [M+H] + . Step 3: 4-prop-2-enoyl-1-([1,2,4]triazolo[1,5-a]pyridin-7-yl)piperaz in-2-one To a solution of 1-([1,2,4]triazolo[1,5-a]pyridin-6-yl)piperazin-2-one (A45, 120 mg, 552.4 µmol, 1 eq) in DCM (2.5 mL) was added TEA (223.6 mg, 2.21 mmol, 307.6 uL, 4 eq) and prop- 2-enoyl chloride (60 mg, 662.9 µmol, 54 uL, 1.2 eq). The mixture was stirred at 0 °C for 1 h. The reaction mixture was concentrated under reduced pressure to give a residue, which was purified by prep-HPLC (Condition 2, Gradient b) to give 4-prop-2-enoyl-1-([1,2,4]triazolo[1,5-a]pyridin- 7-yl)piperazin-2-one (Compound 114, 30 mg, 110.6 umol) as a solid. MS (ESI) m/z 272.2 [M+H] + . 1 H NMR (400 MHz, DMSO-d 6 ): δ = 9.15 (d, J = 1.4 Hz, 1H), 8.54 (s, 1H), 7.88 (d, J = 9.5 Hz, 1H), 7.76 - 7.70 (m, 1H), 6.95 - 6.78 (m, 1H), 6.21 (br d, J = 16.6 Hz, 1H), 5.78 (br d, J = 10.4 Hz, 1H), 4.51 - 4.22 (m, 2H), 4.07 - 3.76 ppm (m, 4H). Example 13: Synthesis of Compound 115 Step 1: tert-butyl 4-(1,3-benzothiazol-2-yl)-3-oxo-piperazine-1-carboxylate A mixture of 2-bromo-1,3-benzothiazole (A45, 500 mg, 2.3 mmol, 1 eq), tert-butyl 3- oxopiperazine-1-carboxylate (468 mg, 2.3 mmol, 1 eq), CuI (66.7 mg, 350.3 µmol, 0.15 eq), DMEDA (51.5 mg, 584 µmol, 63 uL, 0.25 eq) and K 2 CO 3 (645.6 mg, 4.7 mmol, 2 eq) in dioxane (8 mL) was degassed and purged with N2 (x3), and then the mixture was stirred at 100 °C for 12 hr under N2 atmosphere. Upon completion, the reaction mixture was quenched by water (12 mL), and then extracted with EtOAc (20 mL x3). The combined organic layers were washed with brine (20 mL), dried over Na 2 SO 4 , filtered, and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (25-33% EtOAc in PE) to give tert-butyl 4- (1,3-benzothiazol-2-yl)-3-oxo-piperazine-1-carboxylate (A47, 350 mg, 892 umol) as a solid. Step 2: 1-(1,3-benzothiazol-2-yl)piperazin-2-one To a solution of tert-butyl 4-(1,3-benzothiazol-2-yl)-3-oxo-piperazine-1-carboxylate (A47, 350 mg, 1.05 mmol, 1 eq) in DCM (5 mL) was added TFA (1.54 g, 13.5 mmol, 1 mL, 12.9 eq), and the mixture was stirred at 25 °C for 2 hr . Upon completion, the reaction mixture was concentrated under reduced pressure to give a residue that yielded 1-(1,3-benzothiazol-2- yl)piperazin-2-one (A48, 240 mg, crude) as a solid. Step 3: 1-(1,3-benzothiazol-2-yl)-4-prop-2-enoyl-piperazin-2-one To a solution of 1-(1,3-benzothiazol-2-yl)piperazin-2-one (A48, 240 mg, 1.03 mmol, 1 eq) in DCM (5 mL) was added prop-2-enoyl chloride (112 mg, 1.23 mmol, 101 uL, 1.2 eq) and TEA (416.4 mg, 4.12 mmol, 573 uL, 4 eq), and the mixture was stirred at 0 °C for 1 hr. Upon completion, the reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (Condition 4, Gradient a) to give 1-(1,3-benzothiazol-2-yl)-4-prop-2- enoyl-piperazin-2-one (Compound 115, 100 mg, 340.6 umol) as a solid. MS (ESI) m/z= 288.0 [M+H] + . 1 H NMR (400 MHz, DMSO-d6) δ = 8.02 (d, J = 7.9 Hz, 1H), 7.84 (d, J = 8.1 Hz, 1H), 7.47 (dt, J = 1.2, 7.7 Hz, 1H), 7.40 - 7.31 (m, 1H), 6.86 (br d, J = 10.7 Hz, 1H), 6.21 (br d, J = 16.7 Hz, 1H), 5.87 - 5.69 (m, 1H), 4.76 - 4.41 (m, 2H), 4.39 - 4.21 (m, 2H), 4.14 - 3.93 (m, 2H). Example 14: Synthesis of Compound 117 Step 1: tert-butyl 4-imidazo[1, 2-a]pyrimidin-6-yl-3-oxo-piperazine-1-carboxylate A mixture of 6-bromoimidazo[1, 2-a]pyrimidine (A52, 1 g, 5.05 mmol, 1 eq), tert-butyl 3- oxopiperazine-1-carboxylate (1.01 g, 5.05 mmol, 1 eq), CuI (384.71 mg, 2.02 mmol, 0.4 eq), N1, N2-dimethylcyclohexane-1, 2-diamine (574.7 mg, 4.04 mmol, 0.8 eq) and K 2 CO 3 (1.4 g, 10.1 mmol, 2 eq) in toluene (10 mL) was degassed and purged with N2 (x3), and then the mixture was stirred at 120 °C for 16 h under N2 atmosphere. Upon completion, the reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-TLC (9% MeOH in EtOAc) to give tert-butyl 4-imidazo[1,2-a]pyrimidin-6-yl-3-oxo-piperazine-1- carboxylate (A53, 100 mg, 315 umol) as a solid. MS (ESI) m/z 318.3 [M+H] + . Step 2: 1-imidazo[1,2-a]pyrimidin-6-ylpiperazin-2-one A solution of tert-butyl 4-imidazo[1,2-a]pyrimidin-6-yl-3-oxo-piperazine-1-carboxylat e (A53, 100 mg, 315 µmol, 1 eq) in HCl/dioxane (4 M, 3 mL, 38.1 eq) was stirred at 25 °C for 0.5 h. Upon completion, the reaction mixture was concentrated under reduced pressure to give 1- imidazo[1,2-a]pyrimidin-6-ylpiperazin-2-one (A54, 65 mg, crude) as a solid. MS (ESI) m/z 218.2 [M+H] + . Step 3: 1-imidazo[1, 2-a]pyrimidin-6-yl-4-prop-2-enoyl-piperazin-2-one To a solution of 1-imidazo[1, 2-a]pyrimidin-6-ylpiperazin-2-one (A54, 65 mg, 299.2 µmol, 1 eq) in THF (1 mL) and H 2 O (1 mL) was added KOH (35.3 mg, 628.4 µmol, 2.1 eq) at 0 °C, then prop-2-enoyl chloride (24.4 mg, 269.3 µmol, 22 uL, 0.9 eq) in THF (1 mL) was added drop-wise at 0 °C, the mixture was stirred at 0 °C for 10 min. Upon completion, the reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (Condition 6, Gradient b) to give 1-imidazo [1, 2-a] pyrimidin-6-yl-4-prop-2-enoyl-piperazin-2- one (Compound 117, 25 mg, 91.5 umol) as a solid. MS (ESI) m/z 272.2 [M+H] + . 1 H NMR (400 MHz, DMSO-d 6 ) δ = 9.12 (d, J = 2.5 Hz, 1H), 8.57 (d, J = 2.6 Hz, 1H), 7.94 (d, J = 1.2 Hz, 1H), 7.76 (d, J = 1.2 Hz, 1H), 6.95 - 6.80 (m, 1H), 6.22 (br d, J = 16.7 Hz, 1H), 5.78 (br d, J = 10.6 Hz, 1H), 4.49 - 4.30 (m, 2H), 4.07 - 3.77 (m, 4H). Example 15: Synthesis of Compound 118 Step 1: benzyl 4-[1-(1-tert-butoxycarbonyl-4-piperidyl)pyrazol-4-yl]-3-oxo- piperazine-1- carboxylate A mixture of tert-butyl 4-(4-bromopyrazol-1-yl)piperidine-1-carboxylate (A56, 500 mg, 1.51 mmol, 1 eq), benzyl 3-oxopiperazine-1-carboxylate (354.7 mg, 1.5 mmol, 1 eq), CuI (43.3 mg, 227.1 µmol, 0.15 eq), DMEDA (33.4 mg, 378.5 µmol, 40.7 uL, 0.25 eq) and K2CO3 (418.5 mg, 3.03 mmol, 2 eq) in dioxane (7 mL) was degassed and purged with N 2 (x3), and then the mixture was stirred at 100 °C for 16 h under N 2 atmosphere. The reaction mixture was quenched by water (12 mL), and then extracted with EtOAc (15 mL x3). The combined organic layers were washed with brine (15 mL), dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by prep-TLC (100% EtOAc) to give benzyl 4-[1-(1- tert-butoxycarbonyl-4-piperidyl)pyrazol-4-yl]-3-oxo-piperazi ne-1-carboxylate (A56, 400 mg, 827 umol) as a solid. MS (ESI) m/z 484.2 [M+H] + . Step 2: tert-butyl 4-[4-(2-oxopiperazin-1-yl)pyrazol-1-yl]piperidine-1-carboxyl ate To a solution of benzyl 4-[1-(1-tert-butoxycarbonyl-4-piperidyl)pyrazol-4-yl]-3-oxo- piperazine-1-carboxylate (A56, 200 mg, 413.6 µmol, 1 eq) in 75-65-0 (10 mL) was added Pd/C (250 mg), and the mixture was stirred at 20 °C for 1 h. The reaction mixture was concentrated under reduced pressure to give a residue to give tert-butyl 4-[4-(2-oxopiperazin-1-yl)pyrazol-1- yl]piperidine-1-carboxylate (A57, 130 mg, crude) as a solid. MS (ESI) m/z 350.2 [M+H] + . Step 3: tert-butyl 4-[4-(2-oxo-4-prop-2-enoyl-piperazin-1-yl)pyrazol-1-yl]piper idine-1- carboxylate To a solution of tert-butyl 4-[4-(2-oxopiperazin-1-yl)pyrazol-1-yl]piperidine-1- carboxylate (A58, 130 mg, 372 µmol, 1 eq) was added TEA (131.8 mg, 1.3 mmol, 181 uL, 3.5 eq) and prop-2-enoyl chloride (40.4 mg, 446.5 µmol, 36.4 uL, 1.2 eq) in DCM (2.5 mL). The mixture was stirred at 0 °C for 1 h. The reaction mixture was concentrated under reduced pressure to give a residue, which was purified by prep-HPLC (Condition 2, Gradient c) to give (tert-butyl 4-[4-(2- oxo-4-prop-2-enoyl-piperazin-1-yl)pyrazol-1-yl]piperidine-1- carboxylate (Compound 118, 10 mg, 24.8 umol)) as a solid. MS (ESI) m/z 404.2 [M+H] + . 1 H NMR (400 MHz, DMSO-d 6 ): δ = 8.10 (s, 1H), 7.69 (s, 1H), 6.93 - 6.75 (m, 1H), 6.18 (br d, J = 16.5 Hz, 1H), 5.76 (br d, J = 8.8 Hz, 1H), 4.43 - 4.27 (m, 2H), 4.23 (s, 1H), 4.08 - 3.82 (m, 4H), 3.78 - 3.66 (m, 2H), 2.89 (br s, 2H), 2.01 - 1.90 (m, 2H), 1.82 - 1.66 (m, 2H), 1.41 ppm (s, 9H). Example 16: Synthesis of Compound 119 Step 1: 2-[(5-bromopyrrolo[2,3-d]pyrimidin-7-yl)methoxy]ethyl-trimet hyl-silane To a solution of 5-bromo-7H-pyrrolo[2,3-d]pyrimidine (A58, 5 g, 25.3 mmol, 1 eq) in THF (25 mL) was added dropwise NaH (1.11 g, 27.8 mmol, 60% purity, 1.1 eq) at 0 °C. After addition, the mixture was stirred at this temperature for 15 min, and then SEM-Cl (4.6 g, 27.8 mmol, 4.9 mL, 1.1 eq) was added dropwise at 0 °C. The resulting mixture was stirred at 20 °C for 1 h. The reaction mixture was quenched by water (50 mL), and then extracted with EtOAc (50 mL x3). The combined organic layers were washed with brine (50 mL), dried over Na 2 SO 4 , filtered and concentrated under reduced pressure to give a residue. The residue was purified by silica gel column chromatography (25% EtOAc in PE) to give 2-[(5-bromopyrrolo[2,3-d]pyrimidin-7- yl)methoxy]ethyl-trimethyl-silane (A59, 2.6 g, 7.9 mmol) as an oil. MS (ESI) m/z 328.1 [M+H] + . Step 2: tert-butyl 3-oxo-4-[7-(2-trimethylsilylethoxymethyl)pyrrolo[2,3-d]pyrim idin-5- yl]piperazine-1-carboxylate A mixture of 2-[(5-bromopyrrolo[2,3-d]pyrimidin-7-yl)methoxy]ethyl-trimet hyl-silane (A59, 3.3 g, 10 mmol, 1 eq), tert-butyl 3-oxopiperazine-1-carboxylate (2.01 g, 10 mmol, 1 eq), CuI (766 mg, 4 mmol, 0.4 eq), DMEDA (709 mg, 8 mmol, 866 uL, 0.8 eq) and K 2 CO 3 (2.78 g, 20 mmol, 2 eq) in dioxane (35 mL) was degassed and purged with N2 (x3), and then the mixture was stirred at 100 °C for 16 h under N 2 atmosphere. The reaction mixture was diluted with H 2 O 40 mL and extracted with EtOAc (35 mL x3). The combined organic layers were dried over sat. Na 2 SO 4 , filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (5-100% EtOAc in Petroleum ether) to give A60 (tert-butyl 3-oxo-4-[7- (2-trimethylsilylethoxymethyl)pyrrolo[2,3-d]pyrimidin-5-yl]p iperazine-1-carboxylate (3.6 g, 6.76 mmol) as a solid. MS (ESI) m/z 448.3 [M+H] + . Step 3: tert-butyl 3-oxo-4-(7H-pyrrolo[2,3-d]pyrimidin-5-yl)piperazine-1-carbox ylate To a solution of tert-butyl 3-oxo-4-[7-(2-trimethylsilylethoxymethyl)pyrrolo[2,3- d]pyrimidin-5-yl]piperazine-1-carboxylate (A60, 400 mg, 894 µmol, 1 eq) in THF (5 mL) was added TBAF (1 M, 5 mL, 5.60 eq), and the mixture was stirred at 60 °C for 16 h. The reaction mixture was diluted with H2O 15 mL and extracted with EtOAc (15mL x3). The combined organic layers were dried over sat. Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by prep-TLC (5% MeOH in EtOAc) to give tert-butyl 3-oxo-4- (7H-pyrrolo [2,3-d]pyrimidin-5-yl)piperazine-1-carboxylate (A61, 60 mg, 189 umol) as a solid. MS (ESI) m/z 318.3 [M+H] + . Step 4: 1-(7H-pyrrolo[2,3-d]pyrimidin-5-yl)piperazin-2-one To a solution of tert-butyl 3-oxo-4-(7H-pyrrolo[2,3-d]pyrimidin-5-yl)piperazine-1- carboxylate (A61, 160 mg, 504 µmol, 1 eq) was added HCl/dioxane (4 M, 4.7 mL, 37.3 eq). The mixture was stirred at 25 °C for 1 h. The reaction mixture was concentrated under reduced pressure to give a residue containing A62 (1-(7H-pyrrolo[2,3-d]pyrimidin-5-yl)piperazin-2-one (116 mg, crude)) as an oil. MS (ESI) m/z 218.3 [M+H] + . To a solution of 1-(7H-pyrrolo[2,3-d]pyrimidin-5-yl)piperazin-2-one (A62, 116 mg, 534 µmol, 1 eq) in DCM (2.5 mL) was added TEA (270 mg, 2.67 mmol, 371.6 uL, 5 eq) and prop-2- enoyl prop-2-enoate (60.6 mg, 480.6 µmol, 0.9 eq) in DCM (0.25 mL), and the mixture was stirred at 0 °C for 1 h. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (Condition 5, Gradient c) to give Compound 119 (4- prop-2-enoyl-1-(7H-pyrrolo[2,3-d]pyrimidin-5-yl)piperazin-2- one (17 mg, 59.53 umol)) as a solid. MS (ESI) m/z 272.2 [M+H] + . 1 H NMR (400 MHz, DMSO-d6): δ = 12.19 (br s, 1H), 8.94 (s, 1H), 8.82 - 8.68 (m, 1H), 7.67 (s, 1H), 6.99 - 6.72 (m, 1H), 6.37 - 6.07 (m, 1H), 5.77 (br d, J = 10.5 Hz, 1H), 4.57 - 4.22 (m, 2H), 4.14 - 3.68 ppm (m, 4H). Example 17: Synthesis of Compound 120 Step 1: tert-butyl 4-(5-methylpyrazolo[1,5-a]pyrimidin-3-yl)-3-oxo-piperazine-1 -carboxylate A mixture of 3-bromo-5-methyl-pyrazolo[1,5-a]pyrimidine (A63, 400 mg, 1.89 mmol, 1 eq), tert-butyl 3-oxopiperazine-1-carboxylate (377.7 mg, 1.89 mmol, 1 eq), iodocopper (53.9 mg, 283 µmol, 0.15 eq), N,N'-dimethylethane-1,2-diamine (41.6 mg, 471.6 µmol, 50.8 uL, 0.25 eq) and dipotassium;carbonate (521.4 mg, 3.8 mmol, 2 eq) in dioxane (6 mL) was degassed and purged with N2 (x3), and then the mixture was stirred at 100 °C for 16 hr under N2 atmosphere. Upon completion, the reaction mixture was diluted with H 2 O 10 mL and extracted with EtOAc (5 mL * 3). The combined organic layers were dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (5-100% EtOAc in PE) to give A64 (tert-butyl 4-(5-methylpyrazolo[1,5-a]pyrimidin-3-yl)-3-oxo-piperazine-1 - carboxylate (400 mg, 1.21 mmol, 64% yield) as a solid. Step 2: 1-(5-methylpyrazolo[1,5-a]pyrimidin-3-yl)piperazin-2-one To a solution of tert-butyl 4-(5-methylpyrazolo[1,5-a]pyrimidin-3-yl)-3-oxo-piperazine-1 - carboxylate (A64, 200 mg, 603.56 µmol, 1 eq) in DCM (2 mL) was added TFA (770 mg, 6.75 mmol, 0.5 mL, 11.19 eq). The mixture was stirred at 25 °C for 1 h. Upon completion, the reaction was concentrated in vacuum (30 °C), then was added DCM (2 mL), and concentrated to give A65 (1-(5-methylpyrazolo[1,5-a]pyrimidin-3-yl)piperazin-2-one (140 mg, crude)) as an oil. Step 3: 1-(5-methylpyrazolo[1,5-a]pyrimidin-3-yl)-4-prop-2-enoyl-pip erazin-2-one To a solution of 1-(5-methylpyrazolo[1,5-a]pyrimidin-3-yl)piperazin-2-one (139 mg, 601.07 µmol, 1 eq) in DCM (2 mL) was cooled to 0 °C then was added TEA (304.11 mg, 3.01 mmol, 418.31 uL, 5 eq) and prop-2-enoyl chloride (48.96 mg, 540.96 µmol, 44.11 uL, 0.9 eq). The mixture was stirred at 0 °C for 1 h. Upon completion, the reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (Condition 3, Gradient a) to give 1-(5-methylpyrazolo[1,5-a]pyrimidin-3-yl)-4-prop-2-enoyl-pip erazin-2-one (14 mg, 47.60 µmol, 8% yield, 97% purity) as a solid. MS (ESI) m/z 286.2 [M+H] + . NMR (400 MHz, CDCl3-d) δ = 8.47 (d, J = 7.3 Hz, 1H), 8.28 (br s, 1H), 6.70 (d, J = 7.1 Hz, 1H), 6.65 - 6.53 (m, 1H), 6.48 - 6.40 (m, 1H), 5.84 (br d, J = 9.6 Hz, 1H), 4.62 - 4.37 (m, 2H), 4.21 - 3.90 (m, 4H), 2.60 (s, 3H). 1 H NMR (400 MHz, MeOD-d 4 ) δ = 8.74 (d, J = 7.3 Hz, 1H), 8.21 (s, 1H), 6.96 (d, J = 7.1 Hz, 1H), 6.90 - 6.75 (m, 1H), 6.33 (dd, J = 1.9, 16.8 Hz, 1H), 5.94 - 5.72 (m, 1H), 4.57 - 4.41 (m, 2H), 4.11 (br d, J = 2.3 Hz, 2H), 4.01 - 3.88 (m, 2H), 2.61 (s, 3H). The synthesis was taken further to provide the final OTUB1 Recruiter, Compound 120. Example 18: Synthesis of Compound 121 Step 1: tert-butyl 4-(5-methylthiazol-2-yl)-3-oxo-piperazine-1-carboxylate A mixture of 2-bromo-5-methyl-thiazole (A66, 500 mg, 2.8 mmol, 1 eq), tert-butyl 3- oxopiperazine-1-carboxylate (562.3 mg, 2.8 mmol, 1 eq), CuI (80.2 mg, 421.2 µmol, 0.15 eq), DMEDA (61.9 mg, 702 µmol, 75.6 uL, 0.25 eq) and K2CO3 (776.2 mg, 5.6 mmol, 2 eq) in dioxane (7 mL) was degassed and purged with N 2 (x3), and then the mixture was stirred at 100 °C for 16 h under N 2 atmosphere. The reaction mixture was quenched by addition H 2 O 15 mL at 25 °C, and then extracted with EtOAc (15 mL * 3). The combined organic layers were dried over sat. Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (3-18% EtOAc in PE) to give A67 (tert-butyl 4-(5-methylthiazol-2-yl)- 3-oxo-piperazine-1-carboxylate (290 mg, 975 umol)) as a solid. MS (ESI) m/z 298.2 [M+H] + . Step 2: 1-(5-methylthiazol-2-yl)piperazin-2-one To a solution of tert-butyl 4-(5-methylthiazol-2-yl)-3-oxo-piperazine-1-carboxylate (A67, 100 mg, 336.3 µmol, 1 eq) in DCM (2 mL) was added TFA (550 mg, 4.82 mmol, 357.14 uL, 14.34 eq). The mixture was stirred at 25 °C for 1 h. The reaction mixture was concentrated under reduced pressure to give a residue that contained A68 (1-(5-methylthiazol-2-yl) piperazin-2-one (66 mg, crude)) as an oil. MS (ESI) m/z 198.2 [M+H] + . Step 3: 1-(5-methylthiazol-2-yl)-4-prop-2-enoyl-piperazin-2-one To a solution of 1-(5-methylthiazol-2-yl) piperazin-2-one (A68, 60 mg, 304 µmol, 1 eq) in DCM (2 mL) was added TEA (153.9 mg, 1.52 mmol, 211.7 uL, 5 eq) and prop-2-enoyl chloride (27.5 mg, 304.2 µmol, 24.8 uL, 1 eq). The mixture was stirred at -40 °C for 1 h. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (Condition 1, Gradient e) to give Compound 121 (1-(5-methylthiazol-2-yl)-4-prop- 2-enoyl-piperazin-2-one (18.6 mg, 71.1 umol)) as a solid. MS (ESI) m/z 252.2 [M+H] + . 1 H NMR (400 MHz, DMSO-d 6 ): δ = 7.25 (d, J = 0.9 Hz, 1H), 6.95 - 6.73 (m, 1H), 6.19 (br d, J = 16.8 Hz, 1H), 5.77 (br d, J = 9.0 Hz, 1H), 4.65 - 4.32 (m, 2H), 4.21 - 3.86 (m, 4H), 2.36 ppm (s, 3H) Example 19: Synthesis of Compound 123 Step 1: tert-butyl 4-(8-methylimidazo[1,2-a]pyridin-2-yl)-3-oxo-piperazine-1-ca rboxylate A mixture of 2-bromo-8-methyl-imidazo[1,2-a]pyridine (A72, 100 mg, 473.8 µmol, 1 eq), tert-butyl 3-oxopiperazine-1-carboxylate (95 mg, 473.8 µmol, 1 eq), iodocopper (36.1 mg, 189.5 µmol, 0.4 eq), N,N'-dimethylethane-1,2-diamine (33.4 mg, 379 µmol, 40.8 uL, 0.8 eq) and dipotassium;carbonate (131 mg, 947.6 µmol, 2 eq) in dioxane (2 mL) was degassed and purged with N 2 (x3), and then the mixture was stirred at 100 °C for 16 hr under N 2 atmosphere. Upon completion, the reaction mixture was diluted with aq. EDTA 10 mL and extracted with EtOAc (10 mL x2). The combined organic layers were dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (10% EtOAc in PE) to give A73 (tert-butyl 4-(8-methylimidazo[1,2-a]pyridin-2-yl)-3-oxo-piperazine-1- carboxylate (70 mg, 212 umol)) as a solid. Step 2: 1-(8-methylimidazo[1,2-a]pyridin-2-yl)piperazin-2-one To a solution of tert-butyl 4-(8-methylimidazo[1,2-a]pyridin-2-yl)-3-oxo-piperazine-1- carboxylate (A73, 70 mg, 212 µmol, 1 eq) in DCM (2 mL) was added TFA (616 mg, 5.4 mmol, 0.4 mL, 25.5 eq). The mixture was stirred at 25 °C for 1 h. Upon completion, the reaction was concentrated in vacuum (30 °C), then was added DCM(2 mL), and concentrated, and repeat to give A74 (1-(8-methylimidazo[1,2-a]pyridin-2-yl)piperazin-2-one (48 mg, crude)) as an oil. Step 3: 1-(8-methylimidazo[1,2-a]pyridin-2-yl)-4-prop-2-enoyl-pipera zin-2-one To a solution of 1-(8-methylimidazo[1,2-a]pyridin-2-yl)piperazin-2-one (A74, 48 mg, 208.5 µmol, 1 eq) in DCM (2 mL) was cooled to 0 °C then was added TEA (105.5 mg, 1.04 mmol, 145 uL, 5 eq) and prop-2-enoyl chloride (17 mg, 187.6 µmol, 15.3 uL, 0.9 eq). The mixture was stirred at 0 °C for1 h. Upon completion, the reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (Condition 3, Gradient a) to give Compound 123 (1-(8-methylimidazo[1,2-a]pyridin-2-yl)-4-prop-2-enoyl-piper azin-2-one (25.8 mg, 88.8 umol)) as a solid. MS (ESI) m/z 285.1 [M+H] + . 1 H NMR (400 MHz, DMSO-d6) δ = 8.48 - 8.41 (m, 1H), 8.33 (s, 1H), 7.10 - 7.03 (m, 1H), 6.92 - 6.77 (m, 2H), 6.25 - 6.12 (m, 1H), 5.84 - 5.68 (m, 1H), 4.56 - 4.30 (m, 2H), 4.27 - 4.16 (m, 2H), 4.03 - 3.84 (m, 2H), 2.47 (s, 3H). Example 20: Synthesis of Compound 125 Step 1: tert-butyl 4-imidazo[1,2-b]pyridazin-3-yl-3-oxo-piperazine-1-carboxylat e To a solution of 3-bromoimidazo[1,2-b]pyridazine (A78, 1 g, 5 mmol, 1 eq) and tert-butyl 3-oxopiperazine-1-carboxylate (1.01 g, 5 mmol, 1 eq) in dioxane (10 mL) was added iodocopper (385 mg, 2 mmol, 0.4 eq), N,N'-dimethylethane-1,2-diamine (356.1 mg, 4 mmol, 435 uL, 0.8 eq) and dipotassium;carbonate (1.4 g, 10 mmol, 2 eq). The mixture was degassed and purged with N2 (x3), and then the mixture was stirred at 100 °C for 16 h under N2 atmosphere. Upon completion, the reaction mixture was diluted with H 2 O 10 mL and extracted with EtOAc ( 5 mL * 3). The combined organic layers were dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (5-100% EtOAc in PE) to give A79 (tert-butyl 4-imidazo[1,2-b]pyridazin-3-yl-3-oxo-piperazine-1- carboxylate (600 mg, 1.89 mmol)) as a solid. MS (ESI) m/z 318.1 [M+H] + . Step 2: 1-imidazo[1,2-b]pyridazin-3-ylpiperazin-2-one To a solution of tert-butyl 4-imidazo[1,2-b]pyridazin-3-yl-3-oxo-piperazine-1-carboxylat e (300 mg, 945.4 µmol, 1 eq) in DCM (5 mL) was added TFA (1.54 g, 13.5 mmol, 1 mL, 14.3 eq). The mixture was stirred at 25 °C for 2 h. Upon completion, the reaction mixture was concentrated under reduced pressure to give a residue. The residue was used in the next step directly, and contained A80 (1-imidazo[1,2-b]pyridazin-3-ylpiperazin-2-one (200 mg, 921 umol). MS (ESI) m/z 218.1 [M+H] + . Step 3: 1-imidazo[1,2-b]pyridazin-3-yl-4-prop-2-enoyl-piperazin-2-on e To a solution of 1-imidazo[1,2-b]pyridazin-3-ylpiperazin-2-one (A80, 180 mg, 829 µmol, 1 eq) and prop-2-enoyl chloride (113 mg, 1.24 mmol, 101.4 uL, 1.5 eq) in DCM (5 mL) was added TEA (419 mg, 4.14 mmol, 576.7 uL, 5 eq). The mixture was stirred at 0 °C for 1 h. Upon completion, the reaction mixture was concentrated under reduced pressure to to give a residue. The residue was purified by prep-HPLC (Condition 2, Gradient d) to give Compound 125 (1- imidazo[1,2-b]pyridazin-3-yl-4-prop-2-enoyl-piperazin-2-one (118 mg, 436 umol)) as a solid. MS (ESI) m/z 272.1 [M+H] + . 1 H NMR (400 MHz, DMSO-d 6 ) δ = 8.57 (d, J = 4.4 Hz, 1H), 8.23 - 8.13 (m, 1H), 7.81 (s, 1H), 7.30 (dd, J = 4.5, 9.2 Hz, 1H), 6.97 - 6.82 (m, 1H), 6.23 (br d, J = 16.7 Hz, 1H), 5.79 (br d, J = 10.4 Hz, 1H), 4.65 - 4.30 (m, 2H), 4.15 - 3.92 (m, 2H), 3.87 - 3.69 (m, 2H). Example 21: Synthesis of Compound 126 Step 1: tert-butyl 4-imidazo[1,2-a]pyrimidin-3-yl-3-oxo-piperazine-1-carboxylat e To a solution of 3-bromoimidazo[1,2-a]pyrimidine (A81, 500 mg, 2.52 mmol, 1 eq), tert- butyl 3-oxopiperazine-1-carboxylate (505.59 mg, 2.52 mmol, 1 eq), DMEDA (178.07 mg, 2.02 mmol, 217.42 uL, 0.8 eq) and K 2 CO 3 (697.94 mg, 5.05 mmol, 2 eq) in dioxane (5 mL) was added CuI (192.35 mg, 1.01 mmol, 0.4 eq) under N2. The mixture was stirred at 100 °C for 16 h under N2 atmosphere. Upon completion, the reaction mixture was poured into H2O 30 mL, and celite filtered then extracted with EtOAc (30 mL * 3). The combined organic layers were dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (25-50% EtOAc in PE) to give A82 (tert-butyl 4-imidazo[1,2- a]pyrimidin-3-yl-3-oxo-piperazine-1-carboxylate (370 mg, 933 umol)) as a solid. MS (ESI) m/z 318.2 [M+H] + . Step 2: 1-imidazo[1,2-a]pyrimidin-3-ylpiperazin-2-one A mixture of tert-butyl 4-imidazo[1,2-a]pyrimidin-3-yl-3-oxo-piperazine-1-carboxylat e (A82, 200 mg, 630.2 µmol, 1 eq) in HCl/dioxane (4 M, 30 mL, 190.4 eq) was stirred at 25 °C for 1 h. Upon completion, the reaction mixture was concentrated under reduced pressure to give a residue. The crude was used to next step without further purification. A83 (1-imidazo[1,2- a]pyrimidin-3-ylpiperazin-2-one (136 mg, crude)) was obtained as a solid. MS (ESI) m/z 218.2 [M+H] + . Step 3: 4-acryloyl-1-(5-methylfuran-2-yl)piperazin-2-one To a solution of 1-imidazo[1,2-a]pyrimidin-3-ylpiperazin-2-one (A83, 136 mg, 626 µmol, 1 eq) in DCM (3 mL) was added TEA (190 mg, 1.88 mmol, 261.4 uL, 3 eq) and prop-2-enoyl chloride (51 mg, 563.5 µmol, 45.9 uL, 0.9 eq) at 0 °C. The mixture was stirred at 0 °C for 1 h. Upon completion, the reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (Condition 4, Gradient b) to give Compound 126 (1- imidazo[1,2-a]pyrimidin-3-yl-4-prop-2-enoyl-piperazin-2-one (10.48 mg, 37.4 umol)) as a solid. MS (ESI) m/z 272.4 [M+H] + . 1 H NMR (400 MHz, DMSO-d 6 ) δ = 9.01 (dd, J = 1.9, 6.7 Hz, 1H), 8.50 (dd, J = 1.9, 4.2 Hz, 1H), 8.33 (s, 1H), 7.09 (dd, J = 4.2, 6.7 Hz, 1H), 6.87 (br dd, J = 10.4, 16.7 Hz, 1H), 6.20 (br d, J = 16.7 Hz, 1H), 5.85 - 5.69 (m, 1H), 4.59 - 4.29 (m, 2H), 4.28 - 4.10 (m, 2H), 4.05 - 3.87 (m, 2H) An analogous method was performed to obtain the following compounds. Example 22: Synthesis of Compound 127 Step 1: tert-butyl 2-(4-benzyloxycarbonyl-2-oxo-piperazin-1-yl)-6,7-dihydro-4H- thiazolo[4,5- c]pyridine-5-carboxylate A mixture of tert-butyl 2-bromo-6,7-dihydro-4H-thiazolo[4,5-c]pyridine-5-carboxylate (400 mg, 1.25 mmol, 1 eq), benzyl 3-oxopiperazine-1-carboxylate (293.53 mg, 1.25 mmol, 1 eq), CuI (35.8 mg, 188 µmol, 0.15 eq), DMEDA (27.6 mg, 313.3 µmol, 33.7 uL, 0.25 eq) and K2CO3 (346.4 mg, 2.5 mmol, 2 eq) in dioxane (6 mL) was degassed and purged with N 2 (x3), and then the mixture was stirred at 100 °C for 16 h under N2 atmosphere. The reaction mixture was quenched by water (15 mL), and then extracted with EtOAc (20 mL *3). The combined organic layers were washed with brine (20 mL), dried over Na 2 SO 4 , filtered and concentrated under reduced pressure to give a residue. The residue was purified by prep-TLC (25% EtOAc in PE) to give A85 (tert- butyl 2-(4-benzyloxycarbonyl-2-oxo-piperazin-1-yl)-6,7-dihydro-4H- thiazolo[4,5-c]pyridine-5- carboxylate (220 mg, 465.6 umol)) as a solid. MS (ESI) m/z 473.2 [M+H] + . Step 2: tert-butyl 2-(2-oxopiperazin-1-yl)-6,7-dihydro-4H-thiazolo[4,5-c]pyridi ne-5-carboxylate To a solution of tert-butyl 2-(4-benzyloxycarbonyl-2-oxo-piperazin-1-yl)-6,7-dihydro-4H- thiazolo[4,5-c]pyridine-5-carboxylate (150 mg, 317.42 µmol, 1 eq) in i-PrOH (5 mL) was added Pd/C (200 mg, 10% purity).The mixture was stirred at 20 °C for 1 h. The reaction mixture was concentrated under reduced pressure to give a residue that contains A86 (tert-butyl 2-(2- oxopiperazin-1-yl)-6,7-dihydro-4H-thiazolo[4,5-c]pyridine-5- carboxylate (84 mg, crude). MS (ESI) m/z 339.2 [M+H] + . Step 3: tert-butyl 2-(2-oxo-4-prop-2-enoyl-piperazin-1-yl)-6,7-dihydro-4H-thiaz olo[4,5- c]pyridine-5-carboxylate To a solution of tert-butyl 2-(2-oxopiperazin-1-yl)-6,7-dihydro-4H-thiazolo[4,5- c]pyridine-5-carboxylate (84 mg, 248.21 µmol, 1 eq) in DCM (2 mL) was added TEA (125.58 mg, 1.24 mmol, 172.74 uL, 5 eq) and prop-2-enoyl chloride (22.46 mg, 248.21 µmol, 20.24 uL, 1 eq) in DCM (0.25 mL).The mixture was stirred at 0 °C for 1 h. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (Condition 4, Gradient g) to give Compound 127 (tert-butyl 2-(2-oxo-4-prop-2-enoyl-piperazin-1-yl)-6,7- dihydro-4H-thiazolo[4,5-c]pyridine-5-carboxylate (17.13 mg, 43.65 umol)) as a solid. MS (ESI) m/z 393.1 [M+H] + . 1 H NMR (400 MHz, DMSO-d 6 ): δ = 6.96 - 6.69 (m, 1H), 6.26 - 6.09 (m, 1H), 5.78 (br d, J = 9.6 Hz, 1H), 4.67 - 4.28 (m, 4H), 4.23 - 3.88 (m, 4H), 3.63 (t, J = 5.6 Hz, 2H), 2.74 (br t, J = 5.4 Hz, 2H), 1.42 ppm (s, 9H). Example 23: Synthesis of Compound 129 Step 1: 2-[(2-bromopyrrolo[2,3-b]pyridin-1-yl)methoxy]ethyl-trimethy l-silane A solution of 2-bromo-1H-pyrrolo[2,3-b]pyridine (2 g, 10.15 mmol, 1 eq) in THF (30 mL) was added NaH (609.04 mg, 15.23 mmol, 60% purity, 1.5 eq) stirred at 0 °C for 0.5 h, then added SEM-Cl (2.54 g, 15.23 mmol, 2.69 mL, 1.5 eq) was drop-wise, the mixture was stirred at 25 °C for 1 h. Upon completion, the reaction mixture was quenched by addition H 2 O 60 mL, and then extracted with EtOAc (40 mL * 3). The combined organic layers were washed with brine 90 mL, dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (0-20% EtOAc in PE) to give A88 (2-[(2- bromopyrrolo[2,3-b]pyridin-1-yl)methoxy]ethyl-trimethyl-sila ne (2.1 g, 5.8 mmol)) as a solid. MS (ESI) m/z 327.0 [M+H] + . Step 2: tert-butyl 3-oxo-4-[1-(2-trimethylsilylethoxymethyl)pyrrolo[2,3-b]pyrid in-2- yl]piperazine-1-carboxylate A solution of 2-[(2-bromopyrrolo[2,3-b]pyridin-1-yl)methoxy]ethyl-trimethy l-silane (2.1 g, 6.42 mmol, 1 eq), tert-butyl 3-oxopiperazine-1-carboxylate (1.54 g, 7.70 mmol, 1.2 eq) in dioxane (30 mL) was added K 2 CO 3 (1.77 g, 12.83 mmol, 2 eq), CuI (488.79 mg, 2.57 mmol, 0.4 eq) and N,N'-dimethylethane-1,2-diamine (452.48 mg, 5.13 mmol, 552.48 uL, 0.8 eq) under N 2 , the mixture was stirred at 100 °C for 16 h. Upon completion, the reaction mixture was quenched by addition H 2 O 50 mL, and then extracted with EtOAc (40 mL * 3). The combined organic layers were washed with brine 70 mL, dried over Na 2 SO 4 , filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (0-20% EtOAc in Petroleum ether), TLC (50% EtOAc in PE, Rf = 0.46) to give A89 (tert-butyl 3-oxo-4-[1-(2- trimethylsilylethoxymethyl)pyrrolo[2,3-b]pyridin-2-yl]pipera zine-1-carboxylate (890 mg, 1.79 mmol)) as a solid.. MS (ESI) m/z 447.2 [M+H] + . Step 3: 1-(1H-pyrrolo[2,3-b]pyridin-2-yl)piperazin-2-one A solution of tert-butyl 3-oxo-4-[1-(2-trimethylsilylethoxymethyl)pyrrolo[2,3-b]pyrid in- 2-yl]piperazine-1-carboxylate (200 mg, 223.9 µmol, 1 eq) in DCM (0.5 mL) and TFA (3.08 g, 27 mmol, 2 mL, 120.6 eq) was stirred at 25 °C for 1 h. Upon completion, the reaction mixture was concentrated under reduced pressure to remove DCM. A90 (1-(1H-pyrrolo[2,3-b]pyridin-2- yl)piperazin-2-one (100 mg, crude) was obtained as a solid. MS (ESI) m/z 217.1 [M+H] + . Step 4: 4-prop-2-enoyl-1-(1H-pyrrolo[2,3-b]pyridin-2-yl)piperazin-2- one A solution of prop-2-enoyl chloride (37.7 mg, 416.2 µmol, 33.9 uL, 0.9 eq) in DCM (2 mL) was added TEA (93.6 mg, 924.9 µmol, 128.7 uL, 2 eq), then 1-(1H-pyrrolo[2,3-b]pyridin-2- yl)piperazin-2-one (100 mg, 462.5 µmol, 1 eq) in DCM (0.2 mL) was added drop-wise, the mixture was stirred at 0 °C for 1 h. Upon completion, the reaction mixture was quenched by addition H 2 O 0.2 mL and remove DCM. The residue was purified by prep-HPLC (Condition 7, Gradient a), then the residue was purified by prep-HPLC (Condition 2, Gradient e) to give Compound 129 (1-prop- 2-enoyl-1-(1H-pyrrolo[2,3-b]pyridin-2-yl)piperazin-2-one (12 mg, 43.5 umol)) as a solid. MS (ESI) m/z 271.1 [M+H] + . 1 H NMR (400 MHz, DMSO-d6) δ = 11.65 (br d, J = 1.9 Hz, 1H), 8.16 (dd, J = 1.5, 4.8 Hz, 1H), 7.88 (dd, J = 1.5, 7.8 Hz, 1H), 7.06 (dd, J = 4.8, 7.8 Hz, 1H), 6.95 - 6.78 (m, 1H), 6.35 (s, 1H), 6.21 (br d, J = 16.5 Hz, 1H), 5.78 (br d, J = 9.6 Hz, 1H), 4.50 (br s, 1H), 4.35 (s, 1H), 4.03 (br d, J = 4.3 Hz, 1H), 3.96 - 3.84 (m, 3H). An analogous method was performed to obtain the following compounds. Example 24: Synthesis of Compound 128 Step 1: tert-butyl 3-oxo-4-([1,2,4]triazolo[1,5-a]pyridin-2-yl)piperazine-1-car boxylate To a solution of 2-bromo-[1,2,4]triazolo[1,5-a]pyridine (90 mg, 454.50 µmol, 1 eq) and tert-butyl 3-oxopiperazine-1-carboxylate (91.01 mg, 454.50 µmol, 1 eq) in dioxane (3 mL) was added Cs 2 CO 3 (444.25 mg, 1.36 mmol, 3 eq), then Pd 2 (dba) 3 (41.62 mg, 45.45 µmol, 0.1 eq) and Xantphos (52.60 mg, 90.90 µmol, 0.2 eq) was added under N2. The mixture was stirred at 130 °C for 2 h under N 2 . Upon completion, the reaction mixture was filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by prep-TLC (100% EtOAc) to give tert-butyl 3-oxo-4-([1,2,4]triazolo[1,5-a]pyridin-2-yl)piperazine-1- carboxylate (130 mg, 287 µmol, 63% yield, 70% purity) as a solid. MS (ESI) m/z 318.2 [M+H] + . Step 2: 1-([1,2,4]triazolo[1,5-a]pyridin-2-yl)piperazin-2-one A mixture of tert-butyl 3-oxo-4-([1,2,4]triazolo[1,5-a]pyridin-2-yl)piperazine-1- carboxylate (100 mg, 315 µmol, 1 eq) in DCM (1.5 mL) was added drop-wise TFA (462 mg, 4.05 mmol, 0.3 mL, 12.86 eq), the mixture was stirred at 25 °C for 1 h. Upon completion, the reaction mixture was concentrated under reduced pressure to give 1-([1,2,4]triazolo[1,5-a]pyridin-2- yl)piperazin-2-one (65 mg, crude) as an oil. MS (ESI) m/z 218.2 [M+H] + . Step 3: 4-prop-2-enoyl-1-([1,2,4]triazolo[1,5-a]pyridin-2-yl)piperaz in-2-one To a solution of 1-([1,2,4]triazolo[1,5-a]pyridin-2-yl)piperazin-2-one (65 mg, 299.2 µmol, 1 eq) in THF (1 mL) and H 2 O (1 mL) was added K 2 CO 3 (91 mg, 658 µmol, 2.2 eq) at 0 °C, then prop-2-enoyl chloride (27.1 mg, 299.2 µmol, 24.4 uL, 1 eq) in THF (0.5 mL) was added drop- wise, the mixture was stirred at 0 °C for 0.5 h. Upon completion, the reaction mixture was filtered to give a residue. The residue was purified by prep-HPLC (Condition 8, Gradient a) to give the 4- prop-2-enoyl-1-([1,2,4]triazolo[1,5-a]pyridin-2-yl)piperazin -2-one (Compound 128, 11 mg, 39.9 µmol, 13% yield, 98% purity) as a solid. MS (ESI) m/z 272.1 [M+H] + . 1 H NMR (400 MHz, DMSO-d 6 ) δ = 8.93 (d, J = 6.8 Hz, 1H), 7.84 - 7.75 (m, 1H), 7.74 - 7.66 (m, 1H), 7.26 - 7.17 (m, 1H), 6.91 - 6.77 (m, 1H), 6.20 (br d, J = 16.6 Hz, 1H), 5.77 (br d, J = 10.1 Hz, 1H), 4.55 - 4.32 (m, 2H), 4.09 - 3.87 (m, 4H). Example 25: Synthesis of Compound 133 A solution of (2R)-2-amino-4-phenyl-butanoic acid (5 g, 27.9 mmol, 1 eq) in HCl/MeOH (4 M, 50 mL, 7.17 eq) was stirred at 25 °C for 4 h. Upon completion, the mixture was diluted with 200 mL EtOAc, washed with NaHCO 3 , water, brine, then dried over Na 2 SO 4 . To give methyl (2R)- 2-amino-4-phenyl-butanoate (5.28 g, crude) as an oil. Step 2: methyl (2S)-2-[2-(tert-butoxycarbonylamino)ethylamino]-4-phenyl-but anoate
To a solution of tert-butyl N-(2-oxoethyl)carbamate (1 g, 6.28 mmol, 1 eq) and methyl (2R)-2-amino-4-phenyl-butanoate (3.46 g, 7.54 mmol, 50% purity, 1.2 eq, HCl) in MeOH (10 mL), was added AcOH (528.14 mg, 8.79 mmol, 502.99 uL, 1.4 eq) and NaBH3CN (592.15 mg, 9.42 mmol, 1.5 eq) at 0 °C under N 2 , the mixture was stirred at 25 °C for 1 h. Upon completion, the reaction mixture was quenched by addition Sat. aq. NaHCO 3 (15 mL) at 0 °C, and then diluted with H2O (10 mL) and extracted with DCM (30 mL * 3). The combined organic layers were washed with brine (30 mL), dried over Na 2 SO 4 , filtered and concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (Condition 3, Gradient c) to afford methyl (2S)-2-[2-(tert-butoxycarbonylamino)ethylamino]-4-phenyl-but anoate (1.45 g, 3.88 mmol, 62% yield, 90% purity) as an oil. Step 3: methyl (2R)-2-[benzyloxycarbonyl-[2-(tert-butoxycarbonylamino)ethyl ]amino]-4-phenyl- butanoate To a soution of methyl (2S)-2-[2-(tert-butoxycarbonylamino)ethylamino]-4-phenyl- butanoate (1.45 g, 4.31 mmol, 1 eq) in DCM (15 mL) was added CbzCl (1.10 g, 6.47 mmol, 919 uL, 1.5 eq) and DIEA (2.23 g, 17.2 mmol, 3 mL, 4 eq) at 0 °C, the mxture was stirred at 25 °C for 2 h. Upon completion, the reaction mixture was diluted with H 2 O (10 mL) and extracted with DCM (30 mL * 3). The combined organic layers were washed with brine (30 * 3 mL), dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (33-50% EtOAc in PE) to give methyl (2R)-2- [benzyloxycarbonyl-[2-(tert-butoxycarbonylamino)ethyl]amino] -4-phenyl-butanoate (2 g, 3.95 mmol, 92% yield, 93% purity) as an oil. To a solution of methyl (2R)-2-[benzyloxycarbonyl-[2-(tert-butoxycarbonylamino) ethyl]amino]-4-phenyl-butanoate (2 g, 4.3 mmol, 1 eq) in DCM (15 mL) was added TFA (7.7 g, 67.5 mmol, 5 mL, 15.9 eq), the mixture was stirred at 25 °C for 1 h. Upon completion, the reaction concentrated in vacuo. To give methyl (2R)-2-[2-aminoethyl(benzyloxycarbonyl)amino]-4- phenyl-butanoate (1.5 g, crude) as an oil. To solution of methyl (2R)-2-[2-aminoethyl(benzyloxycarbonyl)amino]-4-phenyl- butanoate (1.5 g, 4.05 mmol, 1 eq) in DMF (20 mL) was added Cs2CO3 (3.30 g, 10.12 mmol, 2.5 eq), the mixture was stirred at 80 °C for 2 h. Upon completion, the reaction mixture was diluted with H 2 O (10 mL) and extracted with DCM (30 mL * 3). The combined organic layers were washed with brine (30 mL), dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (50-100% EtOAc in PE) to give benzyl (2R)-3-oxo-2-(2-phenylethyl)piperazine-1-carboxylate (1.25 g, 2.96 mmol, 73% yield, 80% purity) as a solid. Step 6: benzyl (2R)-4-imidazo[1,2-a]pyridin-2-yl-3-oxo-2-(2-phenylethyl)pip erazine-1- carboxylate A mixture of 2-bromoimidazo[1,2-a]pyridine (698.7 mg, 3.55 mmol, 1 eq), benzyl (2R)-3- oxo-2-(2-phenylethyl)piperazine-1-carboxylate (1.2 g, 3.55 mmol, 1 eq), CuI (101.3 mg, 532 µmol, 0.15 eq), DMEDA (78.2 mg, 886.5 µmol, 95.4 uL, 0.25 eq) and K2CO3 (980.2 mg, 7.1 mmol, 2 eq) in dioxane (12 mL) was degassed and purged with N 2 (x3), and then the mixture was stirred at 100 °C for 12 h under N2 atmosphere. Upon completion, the reaction mixture was diluted with H2O (10 mL) and extracted with DCM (30 mL * 3). The combined organic layers were washed with brine (30 mL), dried over Na 2 SO 4 , filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (33-50% EtOAc in Petroleum ether) to afford benzyl (2R)-4-imidazo[1,2-a]pyridin-2-yl-3-oxo-2-(2- phenylethyl)piperazine-1-carboxylate (600 mg, 1.23 mmol, 35% yield, 93% purity) as a solid. A solution of benzyl (2R)-4-imidazo[1,2-a]pyridin-2-yl-3-oxo-2-(2- phenylethyl)piperazine-1-carboxylate (150 mg, 330 µmol, 1 eq) in TFA (2 mL) was stirred at 70 °C for 4 h. Upon completion, the solution was concentrated to dryness to give crude. The crude was used directly for the next step. To give (3R)-1-imidazo[1,2-a]pyridin-2-yl-3-(2- phenylethyl)piperazin-2-one (100 mg, crude) as an oil. Step 8: (3R)-1-imidazo[1,2-a]pyridin-2-yl-3-(2-phenylethyl)-4-prop-2 -enoyl-piperazin-2-one To a solution of (3R)-1-imidazo[1,2-a]pyridin-2-yl-3-(2-phenylethyl)piperazin -2-one (100 mg, 312.12 µmol, 1 eq) in DCM (13 mL) was added TEA (94.75 mg, 936.37 µmol, 130.33 uL, 3 eq) and then prop-2-enoyl chloride (33.90 mg, 374.55 µmol, 30.54 uL, 1.2 eq) was added at 0 °C. The solution was stirred for 1 h at 0 °C. Upon completion, the reaction concentrated in vacuo. The residue was purified by prep-HPLC (Condition 2, Gradient f) to afford (3R)-1-imidazo[1,2- a]pyridin-2-yl-3-(2-phenylethyl)-4-prop-2-enoyl-piperazin-2- one (Compound 133 (R), 30 mg, 79.4 µmol, 25% yield, 99% purity) as a solid. MS (ESI) m/z= 375.2 [M+H] + . 1 H NMR (400 MHz, DMSO-d 6 ) δ = 8.71 - 8.33 (m, 2H), 7.51 (br d, J = 8.9 Hz, 1H), 7.36 - 7.14 (m, 6H), 6.92 (dt, J = 0.9, 6.7 Hz, 2H), 6.22 (dd, J = 2.3, 16.6 Hz, 1H), 5.79 (br d, J = 10.1 Hz, 1H), 5.17 - 4.78 (m, 1H), 4.60 - 4.22 (m, 2H), 4.07 - 3.41 (m, 2H), 2.81 - 2.56 (m, 2H), 2.38 - 2.11 (m, 2H). An analogous method was performed to obtain the following compounds. Step 9: 1-imidazo[1,2-a]pyridin-2-yl-3-(2-phenylethyl)-4-prop-2-enoy l-piperazin-2-one
(3S)-1-imidazo[1,2-a]pyridin-2-yl-3-(2-phenylethyl)-4-prop-2 -enoyl-piperazin-2-one (Compound 133-(S), 10 mg, 26.7 µmol, 1 eq) and (3R)-1-imidazo[1,2-a]pyridin-2-yl-3-(2- phenylethyl)-4-prop-2-enoyl-piperazin-2-one (Compound 133-(R), 10 mg, 26.7 µmol, 1 eq) was dissolved in MeCN 0.5 mL and H 2 O 2 mL, then lyophilized to give 1-imidazo[1,2-a]pyridin-2-yl- 3-(2-phenylethyl)-4-prop-2-enoyl-piperazin-2-one (Compound 133, 15 mg, 40 umol). MS (ESI) m/z 375.2 [M+H] + . 1 H NMR (400 MHz, DMSO-d 6 ) δ = 8.58 (br d, J = 6.6 Hz, 1H), 8.37 (br s, 1H), 7.50 (br d, J = 8.9 Hz, 1H), 7.31 - 7.15 (m, 6H), 6.91 (br t, J = 6.8 Hz, 2H), 6.21 (dd, J = 2.1, 16.6 Hz, 1H), 5.78 (br d, J = 9.8 Hz, 1H), 5.15 - 4.78 (m, 1H), 4.28 (br d, J = 10.3 Hz, 2H), 4.08 - 3.75 (m, 2H), 2.71 - 2.57 (m, 2H), 2.33 - 2.14 (m, 2H) Example 26: Synthesis of Compound 134 To a solution of 2-amino-5-phenyl-pentanoic acid (5 g, 25.9 mmol, 1 eq) in MeOH (50 mL) was added SOCl2 (9.23 g, 77.6 mmol, 5.6 mL, 3 eq) at 0 °C, the mixture was stirred at 80 °C for 3 h. Upon completion, the mixture was concentrated in vacuum and was diluted with DCM and then was concentrated in vacuum to give methyl 2-amino-5-phenyl-pentanoate (6 g, crude, HCl) as a solid. MS (ESI) m/z 208.2 [M+H] + . Step 2: methyl 2-[2-(tert-butoxycarbonylamino)ethylamino]-5-phenyl-pentanoa te
To a solution of methyl 2-amino-5-phenyl-pentanoate (2 g, 8.21 mmol, 1 eq, HCl) and tert- butyl N-(2-oxoethyl)carbamate (1.31 g, 8.21 mmol, 1 eq) in MeOH (20 mL) was added NaBH 3 CN (773.51 mg, 12.31 mmol, 1.5 eq) and AcOH (689.89 mg, 11.49 mmol, 657.04 uL, 1.4 eq) at 0 °C under N2, the mixture was stirred at 25 °C for 1 h. Upon completion, the reaction mixture was quenched by addition sat. NaHCO3 (100 mL) at 0 °C, and then diluted with H2O (100 mL) and extracted with DCM (100 mL * 3). The combined organic layers were washed with brine (50 mL), dried over Na2SO4, filtered and concentrated under reduced pressure. The crude product was purified by silica gel column chromatography (0-50% PE in EtOAc) to give methyl 2-[2-(tert- butoxycarbonylamino)ethylamino]-5-phenyl-pentanoate (1.3 g, 2.97 mmol, 36% yield, 80% purity) as an oil. MS (ESI) m/z 351.3 [M+H] + . Step 3: methyl 2-[benzyloxycarbonyl-[2-(tert-butoxycarbonylamino)ethyl]amin o]-5-phenyl- pentanoate To a solution of methyl 2-[2-(tert-butoxycarbonylamino)ethylamino]-5-phenyl- pentanoate (1.3 g, 3.71 mmol, 1 eq) in DCM (30 mL) was added DIEA (1.92 g, 14.84 mmol, 2.58 mL, 4 eq) and CbzCl (949.22 mg, 5.56 mmol, 791.02 uL, 1.5 eq) at 0 °C under N2. The mixture was stirred at 25 °C for 3 h. Upon completion, the reaction mixture was diluted with H 2 O (200 mL) and extracted with DCM (100 mL * 3). The combined organic layers were dried over Na 2 SO 4 , filtered and concentrated under reduced pressure to give a residue. The crude product was purified by column (0-25% EtOAc in PE) to give methyl 2-[benzyloxycarbonyl-[2-(tert- butoxycarbonylamino)ethyl]amino]-5-phenyl-pentanoate (1.5 g, 2.2 mmol, 58% yield, 70% purity) as a gum. MS (ESI) m/z 485.2 [M+H] + . A solution of methyl 2-[benzyloxycarbonyl-[2-(tert-butoxycarbonylamino)ethyl]amin o]- 5-phenyl-pentanoate (1.5 g, 3.1 mmol, 1 eq) in HCl/dioxane (20 mL, 4M) was stirred at 25 °C for 1 h. Upon completion, the mixture was concentrated in vacuum to give methyl 2-[2- aminoethyl(benzyloxycarbonyl)amino]-5-phenyl-pentanoate (1.3 g, crude, HCl) as an oil. MS (ESI) m/z 385.2 [M+H] + . Step 5: benzyl 3-oxo-2-(3-phenylpropyl)piperazine-1-carboxylate To a solution of methyl 2-[2-aminoethyl(benzyloxycarbonyl)amino]-5-phenyl-pentanoate (1.3 g, 3.38 mmol, 1 eq) in DMF (15 mL) was added Cs 2 CO 3 (2.75 g, 8.45 mmol, 2.5 eq), the mixture was stirred at 25 °C for 2 h. Upon completion, the reaction mixture was diluted with H 2 O 200 mL and extracted with DCM (100 mL * 3). The combined organic layers were washed with brine 50 mL, dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The crude product was purified by column (0-33% EtOAc in PE) to give benzyl 3-oxo-2-(3- phenylpropyl)piperazine-1-carboxylate (1.2 g, 2.4 mmol, 71% yield, 70% purity) as an oil. MS (ESI) m/z 353.2 [M+H] + . Step 6: benzyl 4-imidazo[1,2-a]pyridin-2-yl-3-oxo-2-(3-phenylpropyl)piperaz ine-1-carboxylate
To a solution of 2-bromoimidazo[1,2-a]pyridine (375.70 mg, 1.91 mmol, 1.2 eq), benzyl 3-oxo-2-(3-phenylpropyl)piperazine-1-carboxylate (800 mg, 1.59 mmol, 70% purity, 1 eq) in dioxane (16 mL) was added K2CO3 (439.21 mg, 3.18 mmol, 2 eq), CuI (121.05 mg, 635.59 µmol, 0.4 eq) and N,N'-dimethylethane-1,2-diamine (112.06 mg, 1.27 mmol, 136.82 uL, 0.8 eq) under N 2 , the mixture was stirred at 100 °C for 20 h. Upon completion, the reaction mixture was concentrated under reduced pressure to remove solvent. The residue was diluted with H2O 100 mL and extracted with EtOAc (40 mL * 3). The combined organic layers were washed with NaCl 20 mL, dried over Na 2 SO 4 , filtered and concentrated under reduced pressure to give a residue. The crude product was purified by column (0-25-50% EtOAc in PE) to give benzyl 4-imidazo[1,2- a]pyridin-2-yl-3-oxo-2-(3-phenylpropyl)piperazine-1-carboxyl ate (280 mg, 568 µmol, 36% yield, 95% purity) as a gum. MS (ESI) m/z 469.2 [M+H] + . A solution of benzyl 4-imidazo[1,2-a]pyridin-2-yl-3-oxo-2-(3-phenylpropyl)piperaz ine-1- carboxylate (250 mg, 534 µmol, 1 eq) in TFA (4 mL) was stirred at 70 °C for 2 h. Upon completion, the mixure was dried by blowing N 2 to obtained 1-imidazo[1,2-a]pyridin-2-yl-3-(3- phenylpropyl)piperazin-2-one (180 mg, crude) as a gum. MS (ESI) m/z 335.2 [M+H] + . Step 8: 1-imidazo[1,2-a]pyridin-2-yl-3-(3-phenylpropyl)-4-prop-2-eno yl-piperazin-2-one
To a solution of 1-imidazo[1,2-a]pyridin-2-yl-3-(3-phenylpropyl)piperazin-2-o ne (240 mg, 473.66 µmol, 66% purity, 1 eq) in DCM (25 mL) was added TEA (239.65 mg, 2.37 mmol, 329.64 uL, 5 eq) and then was added prop-2-enoyl chloride (42.87 mg, 473.66 µmol, 38.62 uL, 1 eq) dropwised at 0 °C, the mixture was stirred at 0 °C for 0.5 h. Upon completion, the mixture was quenched with water (0.5 mL) and was dried by blowing N 2 . The crude product was purified by prep-HPLC (Condition 4, Gradient f) to give 1-imidazo[1,2-a]pyridin-2-yl-3-(3-phenylpropyl)-4- prop-2-enoyl-piperazin-2-one (Compound 134, 40 mg, 103 µmol, 22% yield, 100% purity) as a solid. MS (ESI) m/z 389.2 [M+H] + . 1 H NMR (400 MHz, DMSO-d 6 ) δ ppm 8.57 (d, J = 6.6 Hz, 1H), 8.35 (s, 1H), 7.50 (br d, J = 8.8Hz, 1H), 7.31 - 7.21 (m, 3H), 7.21 - 7.12 (m, 3H), 6.96 - 6.81 (m, 2H), 6.20 (br d, J = 16.6 Hz, 1H), 5.84 - 5.68 (m, 1H), 5.10 - 4.77 (m, 1H), 4.59 - 4.17 (m, 2H), 4.08 - 3.64 (m, 2H), 2.72 - 2.53 (m, 2H), 2.05 - 1.84 (m, 2H), 1.77 - 1.50 (m, 2H). Example 27: Synthesis of Compound 135 Step1 : tert-butyl 3-oxo-2-(4-phenylbutyl)piperazine-1-carboxylate To a solution of tert-butyl 3-oxopiperazine-1-carboxylate (1 g, 4.99 mmol, 1 eq) in THF (10 mL) was added LDA (2.5 M, 4 mL, 2 eq). The mixture was stirred at -70 °C for 30 min, then a solution of (4-bromobutyl) benzene (745 mg, 3.5 mmol, 0.7 eq) in THF (2 mL) was added to the mixture. The mixture was stirred for 1 h. Upon completion, the mixture was quenched by NaHCO3 aq. (10 mL), then extract with EtOAc (20 mL * 3). The combined organic layers were washed with sat. NaCl (20 mL * 3), dried over Na 2 SO 4 to give a residue. The residue was purified by silica gel column (9-50% EtOAc in PE) to give tert-butyl 3-oxo-2-(4-phenylbutyl)piperazine-1-carboxylate as a solid. MS (ESI) m/z 333.2 [M+H] + . Step2 : tert-butyl 4-imidazo[1 To a solution of tert-butyl 3-oxo-2-(4-phenylbutyl)piperazine-1-carboxylate (350 mg, 1.05 mmol, 1 eq) and 2-bromoimidazo[1,2-a]pyridine (207.4 mg, 1.05 mmol, 1 eq) in dioxane (5 mL) was added K2CO3 (291 mg, 2.11 mmol, 2 eq), DMEDA (23.2 mg, 263.2 µmol, 28.3 uL, 0.25 eq), then exchanged N 2 (x3), then added CuI (30.1 mg, 157.9 µmol, 0.15 eq) and exchange N 2 (x3), The mixture was stirred at 100 °C for 12 h. Upon completion, the reaction mixture was diluted with H2O (10 mL) and extracted with DCM (10 mL * 3). The combined organic layers were washed with sat. NaCl (15 mL), dried over Na2SO4, filtered, and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (0-50% EtOAc in PE) to give tert-butyl 4-imidazo[1,2-a]pyridin-2-yl-3-oxo-2-(4-phenylbutyl)piperazi ne-1- carboxylate (210 mg, 468 µmol, 45% yield) as a solid. MS (ESI) m/z 449.3[M+H] + To a solution of tert-butyl 4-imidazo[1,2-a]pyridin-2-yl-3-oxo-2-(4- phenylbutyl)piperazine-1-carboxylate (200 mg, 446 µmol, 1 eq) in DCM (3 mL) was added TFA (1.54 g, 13.51 mmol, 30.29 eq). The mixture was stirred at 25 °C for 1 h. Upon completion, the resulting solution was concentrated in vacuum to give 1-imidazo [1, 2-a] pyridin-2-yl-3-(4- phenylbutyl) piperazin-2-one (150 mg, crude) as a solid. MS (ESI) m/z 349.3[M+H] + Step4 : 1-imidazo[1,2-a]pyridin-2-yl-3-(4-phenylbutyl)-4-prop-2-enoy l-piperazin-2-one
To a solution of 1-imidazo[1,2-a]pyridin-2-yl-3-(4-phenylbutyl)piperazin-2-on e (150 mg, 430.5 µmol, 1 eq) in DCM (2 mL) was added TEA (217.8 mg, 2.15 mmol, 299.6 uL, 5 eq) and prop-2-enoyl chloride (39 mg, 430 µmol, 35 uL, 1 eq) at 0 °C under N2 protection. The mixture was stirred at 0 °C for 1 h. Upon completion, the reaction mixture was concentrated in vacuum to give a residue. The residue was purified by prep-HPLC (Condition 1, Gradient g) to give 1- imidazo[1,2-a]pyridin-2-yl-3-(4-phenylbutyl)-4-prop-2-enoyl- piperazin-2-one (Compound 135, 62.3 mg, 154.7 µmol, 36% yield, 100% purity) as a solid. MS (ESI) m/z 403.2 [M+H] + . 1 H NMR (400 MHz, DMSO-d 6 ) δ = 8.62 - 8.55 (m, 1H), 8.41 - 8.33 (m, 1H), 7.55 - 7.45 (m, 1H), 7.29 - 7.11 (m, 6H), 6.95 - 6.80 (m, 2H), 6.27 - 6.09 (m, 1H), 5.82 - 5.66 (m, 1H), 4.80 (br s, 1H), 4.25 (br d, J = 9.7 Hz, 2H), 4.12 - 3.68 (m, 2H), 2.60 - 2.53 (m, 2H), 2.03 - 1.86 (m, 2H), 1.68 - 1.52 (m, 2H), 1.46 - 1.23 (m, 2H). An analogous method was followed to obtain the following compounds. Example 28: Synthesis of Compound 139 Step 1: methyl (R)-2-amino-3-(4-fluorophenyl)propanoate To a solution of (2R)-2-amino-3-(4-fluorophenyl)propanoic acid (5 g, 27.3 mmol, 1 eq) in MeOH (60 mL) was added SOCl 2 (13 g, 109 mmol, 7.9 mL, 4 eq) at 0 °C. And then, the reaction mixture was stirred at 60 °C for 2 h. Upon completion, the reaction mixture was concentrated under reduced pressure to give methyl (2R)-2-amino-3-(4-fluorophenyl)propanoate (6.2 g, crude, HCl) as a solid. Step 2: methyl (2R)-2-[2-(tert-butoxycarbonylamino)ethylamino]-3-(4-fluorop henyl)propanoate
To a solution of tert-butyl N-(2-oxoethyl)carbamate (3.5 g, 22 mmol, 1 eq) and methyl (2R)-2-amino-3-(4-fluorophenyl)propanoate (6.17 g, 26 mmol, 1.2 eq, HCl) in MeOH (50 mL) was added NaBH 3 CN (2.07 g, 33 mmol, 1.5 eq) and AcOH (1.85 g, 30.8 mmol, 1.76 mL, 1.4 eq) at 0 °C. The mixture was stirred at 25 °C for 1 h. Upon completion, the mixture was adjusted pH to 7 by NaHCO3 aq., then extracted with EtOAc (50 mL * 3), then washed with brine (50 mL), dried over Na 2 SO 4 , filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (5-100% EtOAc in PE) to give methyl (2R)-2-[2-(tert- butoxycarbonylamino)ethylamino]-3-(4-fluorophenyl)propanoate (5 g, 13.2 mmol, 60% yield, 90% purity) as an oil. MS (ESI) m/z 341.3 [M+H] + . Step 3: methyl (2R)-2-[benzyloxycarbonyl-[2-(tert-butoxycarbonylamino)ethyl ]amino]-3-(4- fluorophenyl)propanoate To a solution of methyl (2R)-2-[2-(tert-butoxycarbonylamino)ethylamino]-3-(4- fluorophenyl)propanoate (5 g, 14.7 mmol, 1 eq) and in DCM (50 mL) was added DIEA (7.59 g, 58.8 mmol, 10.2 mL, 4 eq) and CbzCl (3.8 g, 22 mmol, 3.13 mL, 1.5 eq) at 0 °C. The mixture was stirred at 25 °C for 16 h. Upon completion, the reaction mixture was poured into H 2 O (150 mL) at 20 °C, and then extracted with DCM (50 mL * 3). The combined organic layers were washed with brine (50 mL), dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column (5-100% EtOAc in PE) to give methyl (2R)-2- [benzyloxycarbonyl-[2-(tert-butoxycarbonylamino)ethyl]amino] -3-(4-fluorophenyl)propanoate (5 g, 9.5 mmol, 65% yield, 90% purity) as an oil. MS (ESI) m/z 375.2 [M+H-100] + . Step 4: methyl (2R)-2-[2-aminoethyl(benzyloxycarbonyl)amino]-3-(4-fluorophe nyl)propanoate A solution of methyl (2R)-2-[benzyloxycarbonyl-[2-(tert-butoxycarbonylamino)- ethyl]amino]-3-(4-fluorophenyl)propanoate (5 g, 10.5 mmol, 1 eq) in HCl/dioxane (4 M, 50 mL, 19 eq) was stirred at 25 °C for 1 h. Upon completion, the reaction mixture was concentrated under reduced pressure to give methyl (2R)-2-[2-aminoethyl(benzyloxycarbonyl)amino]-3-(4- fluorophenyl)propanoate (4.8 g, crude, HCl) as an oil. MS (ESI) m/z 375.2 [M+H] + . Step 5: benzyl (2R)-2-[(4-fluorophenyl)methyl]-3-oxo-piperazine-1-carboxyla te A solution of methyl (2R)-2-[2-aminoethyl(benzyloxycarbonyl)amino]-3-(4- fluorophenyl)propanoate (4.8 g, 11.7 mmol, 1 eq, HCl) in MeOH (70 mL) was added DBU (1.78 g, 11.7 mmol, 1.76 mL, 1 eq), the reaction mixture was stirred at 25 °C for 1 h. Upon completion, the reaction mixture was poured into H 2 O (50 mL) at 20 °C, and then extracted with EtOAc (70mL * 3). The combined organic layers were washed with brine (100 mL), dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column (5- 75% EtOAc in PE) to give benzyl (2R)-2-[(4-fluorophenyl)methyl]-3-oxo-piperazine-1- carboxylate (3.2 g, 9.4 mmol, 80% yield) as a solid. MS (ESI) m/z 343.3 [M+H] + . Step 6: benzyl (2R)-2-[(4-fluorophenyl)methyl]-4-imidazo[1,2-a]pyridin-2-yl -3-oxo-piperazine- 1-carboxylate A solution of benzyl (2R)-2-[(4-fluorophenyl)methyl]-3-oxo-piperazine-1-carboxyla te (1.2 g, 3.5 mmol, 1 eq) and 2-bromoimidazo[1,2-a]pyridine (828.7 mg, 4.2 mmol, 1.2 eq) in dioxane (15 mL) and dipotassium;carbonate (968.84 mg, 7.01 mmol, 2 eq) was gassed and purged with N 2 (x3). Then CuI (267 mg, 1.4 mmol, 0.4 eq) and N,N'-dimethylethane-1,2-diamine (247.2 mg, 2.8 mmol, 302 uL, 0.8 eq) was added under N2, the mixture was stirred at 100 °C for 16 h. Upon completion, the reaction mixture was poured into H2O (10 mL) at 20 °C, and then extracted with EtOAc (20 mL * 3). The combined organic layers were washed with brine (40 mL), dried over Na 2 SO 4 , filtered and concentrated under reduced pressure to give a residue. The residue was purified by column (5-35% EtOAc in PE) to give benzyl (2R)-2-[(4-fluorophenyl)methyl]-4- imidazo[1,2-a]pyridin-2-yl-3-oxo-piperazine-1-carboxylate (750 mg, 1.6 mmol, 47% yield) as a solid. MS (ESI) m/z 459.2 [M+H] + . Step 7: (3R)-3-[(4-fluorophenyl)methyl]-1-imidazo[1,2-a]pyridin-2-yl -piperazin-2-one
A solution of benzyl (2R)-2-[(4-fluorophenyl)methyl]-4-imidazo[1,2-a]pyridin-2-yl -3- oxo-piperazine-1-carboxylate (400 mg, 872.4 µmol, 1 eq) in TFA (5 mL) was stirred at 70 °C for 4 h. Upon completion, the reaction mixture was concentrated under reduced pressure to give (3R)- 3-[(4-fluorophenyl)methyl]-1-imidazo[1,2-a]pyridin-2-yl-pipe razin-2-one (280 mg, crude) was obtained as an oil. MS (ESI) m/z 325.3 [M+H] + . Step 8: (3R)-3-[(4-fluorophenyl)methyl]-1-imidazo[1,2-a]pyridin-2-yl -4-prop-2-enoyl-piperazin- 2-one To a solution of (3R)-3-[(4-fluorophenyl)methyl]-1-imidazo[1,2-a]pyridin-2-yl -piperazin- 2-one (280 mg, 863.3 µmol, 1 eq) in DCM (4 mL) was added TEA (262 mg, 2.6 mmol, 360.5 uL, 3 eq) and then prop-2-enoyl chloride (78.1 mg, 863.3 µmol, 70.4 uL, 1 eq) in DCM (1 mL) was added at 0 °C. The solution was stirred for 0.5 h at 0 °C. Upon completion, the reaction mixture was quenched by addition H2O (0.5 mL) at 0 °C, and then concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (Condition 4, Gradient i) to give (3R)-3- [(4-fluorophenyl)methyl]-1-imidazo[1,2-a]pyridin-2-yl-4-prop -2-enoyl-piperazin-2-one (Compound 139-(R), 128 mg, 338 µmol, 39% yield) as a solid. MS (ESI) m/z 379.1 [M+H] + . An analogous method was followed to obtain the following compound. Step 9: 3-[(4-fluorophenyl)methyl]-1-imidazo[1,2-a]pyridin-2-yl-4-pr op-2-enoyl-piperazin-2- one (3R)-3-[(4-fluorophenyl)methyl]-1-imidazo[1,2-a]pyridin-2-yl -4-prop-2-enoyl-piperazin- 2-one (Compound 139-(R), 10 mg, 26.4 μmol, 1 eq) and (3S)-3-[(4-fluorophenyl)methyl]-1- imidazo[1,2-a]pyridin-2-yl-4-prop-2-enoyl-piperazin-2-one (Compound 139-(S), 10 mg, 26.4 μmol, 1 eq) was dissolved in MeCN (0.5 mL) and H2O (2 mL), The mixture was stirred at 25 °C for 5 min. Upon completion, the mixture was obtained by freeze-drying to give 3-[(4- fluorophenyl)methyl]-1-imidazo[1,2-a]pyridin-2-yl-4-prop-2-e noyl-piperazin-2-one (Compound 139, 18 mg, 47.6 μmol, 90% yield, 100% purity) as a solid. MS (ESI) m/z 379.1 [M+H] + . 1 H NMR (400 MHz, DMSO-d6) δ = 8.62 (t, J = 7.3 Hz, 1H), 8.42 (d, J = 12.6 Hz, 1H), 7.52 (t, J = 10.0 Hz, 1H), 7.31 - 7.14 (m, 3H), 7.13 - 7.04 (m, 2H), 6.94 (t, J = 6.7 Hz, 1H), 6.88 - 6.19 (m, 1H), 6.17 - 5.82 (m, 1H), 5.81 - 5.32 (m, 1H), 5.22 (s, 1H), 4.68 - 4.27 (m, 1H), 4.24 - 4.03 (m, 1H), 4.01 - 3.70 (m, 1H), 3.47 - 3.37 (m, 1H), 3.29 - 3.17 (m, 2H). An analogous method was followed to obtain the following compounds. Example 29: Synthesis of Compound 144 Step 1: methyl (2R)-2-amino-3-(4-nitrophenyl)propanoate A solution of (2R)-2-amino-3-(4-nitrophenyl)propanoic acid (10 g, 47.6 mmol, 1 eq) in MeOH (100 mL) was added SOCl2 (22.6 g, 190.3 mmol, 13.8 mL, 4 eq). The mixture was stirred at 80 °C for 2 hr. Upon completion, the reaction mixture was poured into H2O 300 mL at 20 °C, and then extracted with EtOAc (200 mL * 3). The combined organic layers were washed with brine 300 mL, dried over Na 2 SO 4 , filtered and concentrated under reduced pressure to give methyl (2R)-2-amino-3-(4-nitrophenyl)propanoate (12 g, crude) as a solid. MS (ESI) m/z 225.2 [M+H] + . Step 2: methyl (2R)-2-[2-(tert-butoxycarbonylamino)ethylamino]-3-(4-nitroph enyl)propanoate
A solution of methyl (2R)-2-amino-3-(4-nitrophenyl)propanoate (6 g, 26.8 mmol, 1.2 eq) and tert-butyl N-(2-oxoethyl)carbamate (3.55 g, 22.3 mmol, 1 eq) in MeOH (60 mL) was added NaBH 3 CN (2.1 g, 33.5 mmol, 1.5 eq) and AcOH (1.87 g, 31.2 mmol, 1.79 mL, 1.4 eq) at 0 °C. The mixture was stirred at 25 °C for 1 h. Upon completion, the reaction mixture was poured into H2O 200 mL at 20 °C, and then extracted with EtOAc (100 mL * 3). The combined organic layers were washed with brine 100 mL, dried over Na 2 SO 4 , filtered and concentrated under reduced pressure. The residue was purified by column chromatography (0-100% EtOAc in PE) (TLC, 100% EtOAc, Rf = 0.5) to give methyl (2R)-2-[2-(tert-butoxycarbonylamino)ethylamino]-3-(4- nitrophenyl)propanoate (6 g, 15.2 mmol, 70% yield, 93% purity) as a solid. MS (ESI) m/z 368.2 [M+H] + . Step 3: methyl (2R)-2-[benzyloxycarbonyl-[2-(tert-butoxycarbonylamino)ethyl ]amino]-3-(4- nitrophenyl)propanoate A solution of methyl (2R)-2-[2-(tert-butoxycarbonylamino)ethylamino]-3-(4- nitrophenyl)propanoate (5.5 g, 15 mmol, 1 eq) in DCM (50 mL) was added DIEA (7.74 g, 59.88 mmol, 10.43 mL, 4 eq) and CbzCl (3.8 g, 22.5 mmol, 3.2 mL, 1.5 eq) at 0 °C. The mixture was stirred at 25 °C for 16 hr. Upon completion, the reaction mixture was poured into H2O 100 mL at 20 °C, and then extracted with EtOAc (50 mL * 3). The combined organic layers were washed with brine 100 mL, dried over Na 2 SO 4 , filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (0-100% EtOAc in PE) to give methyl (2R)-2-[benzyloxycarbonyl-[2-(tert-butoxycarbonylamino)-ethy l]amino]-3-(4- nitrophenyl)propanoate (6.3 g, 12 mmol, 81% yield, 96% purity) as an oil. MS (ESI) m/z 524.2 [M+Na] + . Step 4: methyl (2R)-2-[2-aminoethyl(benzyloxycarbonyl)amino]-3-(4-nitrophen yl)propanoate A solution of methyl (2R)-2-[benzyloxycarbonyl-[2-(tert-butoxycarbonylamino)ethyl ]- amino]-3-(4-nitrophenyl)propanoate (6.3 g, 12.6 mmol, 1 eq) in HCl/dioxane (4 M, 60 mL) was stirred at 25 °C for 1 hr. Upon completion, the mixture was concentrated under reduced pressure to give methyl (2R)-2-[2-aminoethyl(benzyloxycarbonyl)amino]-3-(4-nitrophen yl)propanoate (4.6 g, crude) as an oil. MS (ESI) m/z 402.2 [M+H] + . Step 5: benzyl (2R)-2-[(4-nitrophenyl)methyl]-3-oxo-piperazine-1-carboxylat e A solution of methyl (2R)-2-[2-aminoethyl(benzyloxycarbonyl)amino]-3-(4- nitrophenyl)propanoate (4.6 g, 11.5 mmol, 1 eq) in DMF (60 mL) was added Cs 2 CO 3 (7.47 g, 22.9 mmol, 2 eq). The mixture was stirred at 80 °C for 2 hr. Upon completion, the reaction mixture was quenched by addition H2O 100 mL at 25 °C, and extracted with EtOAc (50 mL * 3). The combined organic layers were washed with brine (100 mL * 1), dried over Na 2 SO 4 , filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (0- 50% EtOAc in PE) to give benzyl (2R)-2-[(4-nitrophenyl)methyl]-3-oxo-piperazine-1-carboxylat e (1.96 g, 3.9 mmol, 25% yield, 74% purity) as an oil. MS (ESI) m/z 370.2 [M+H] + . Step 6: benzyl (2R)-4-imidazo[1,2-a]pyridin-2-yl-2-[(4-nitrophenyl)methyl]- 3-oxo-piperazine-1- carboxylate A mixture of benzyl (2R)-2-[(4-nitrophenyl)methyl]-3-oxo-piperazine-1-carboxylat e (1.96 g, 5.3 mmol, 1 eq), 2-bromoimidazo[1,2-a]pyridine (1.25 g, 6.4 mmol, 1.2 eq), K 2 CO 3 (1.47 g, 10.6 mmol, 2 eq), CuI (404.2 mg, 2.12 mmol, 0.4 eq) and DMEDA (374.2 mg, 4.25 mmol, 456.9 uL, 0.8 eq) in dioxane (20 mL) was degassed and purged with N2 (x3), and then the mixture was stirred at 100 °C for 16 hr under N 2 atmosphere. Upon completion, the reaction mixture was quenched by addition H 2 O 60 mL, and then extracted with EtOAc (40 mL * 3). The combined organic layers were washed with brine 70 mL, dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (0-50% EtOAc in PE) to give benzyl (2R)-4-imidazo[1,2-a]pyridin-2-yl-2-[(4-nitrophenyl)methyl]- 3-oxo- piperazine-1-carboxylate (1 g, 1.85 mmol, 35% yield, 90% purity) as a solid. MS (ESI) m/z 486.2 [M+H] + . Step 7: benzyl (2R)-2-[(4-aminophenyl)methyl]-4-imidazo[1,2-a]pyridin-2-yl- 3-oxo-piperazine- 1-carboxylate
A solution of benzyl (2R)-4-imidazo[1,2-a]pyridin-2-yl-2-[(4-nitrophenyl)methyl]- 3-oxo- piperazine-1-carboxylate (500 mg, 1.03 mmol, 1 eq) and NH 4 Cl (551 mg, 10.3 mmol, 10 eq) in EtOH (15 mL) and H2O (3 mL) then Fe (287.6 mg, 5.15 mmol, 5 eq) was added at 60 °C and was stirred at 80 °C for 2 h. Upon completion, the solids were filtered out and the resulting solution was concentrated under vacuum. The residue was purified by column chromatography (0-100% EtOAc in PE) to give benzyl (2R)-2-[(4-aminophenyl)methyl]-4-imidazo[1,2-a]pyridin-2-yl- 3- oxo-piperazine-1-carboxylate (500 mg, crude) as an oil. MS (ESI) m/z 456.2 [M+H] + . Step 8: benzyl (2R)-2-[(4-acetamidophenyl)methyl]-4-imidazo[1,2-a]pyridin-2 -yl-3-oxo- piperazine-1-carboxylate A solution of benzyl (2R)-2-[(4-aminophenyl)methyl]-4-imidazo[1,2-a]pyridin-2-yl- 3- oxo-piperazine-1-carboxylate (500 mg, 1.10 mmol, 1 eq) in DCM (5 mL) was added Py (260.48 mg, 3.29 mmol, 265.79 uL, 3 eq) then Ac2O (123.27 mg, 1.21 mmol, 113.09 uL, 1.1 eq) was added, and was stirred at 20°C for 1 h. Upon completion, the reaction mixture was poured into H 2 O 20 mL at 20 °C, and then extracted with EtOAc (10 mL * 3). The combined organic layers were washed with brine 20 mL, dried over Na2SO4, filtered and concentrated under reduced pressure to give benzyl (2R)-2-[(4-acetamidophenyl)methyl]-4-imidazo[1,2-a]pyridin-2 -yl-3-oxo-piperazine- 1-carboxylate (660 mg, crude) as an oil. MS (ESI) m/z 498.2 [M+H] + . Step 9: N-[4-[[(2R)-4-imidazo[1,2-a]pyridin-2-yl-3-oxo-piperazin-2- yl]methyl]phenyl]acetamide A mixture of benzyl (2R)-2-[(4-acetamidophenyl)methyl]-4-imidazo[1,2-a]pyridin-2 -yl-3- oxo-piperazine-1-carboxylate (660 mg, 1.33 mmol, 1 eq), Pd/C (660 mg, 1.33 mmol, 10% purity, 1 eq) in EtOH (5 mL) was degassed and purged with H 2 (x3), and then the mixture was stirred at 20 °C for 10 min under H2 (15 Psi) atmosphere. Upon completion, the reaction mixture was filtered and concentrated under reduced pressure to give N-[4-[[(2R)-4-imidazo[1,2-a]pyridin-2- yl-3-oxo-piperazin-2-yl]methyl]phenyl]acetamide (290 mg, crude) as an oil. MS (ESI) m/z 364.2 [M+H] + . Step 10: N-[4-[[(2R)-4-imidazo[1,2-a]pyridin-2-yl-3-oxo-1-prop-2-enoy l-piperazin-2- yl]methyl]phenyl]acetamide A solution of N-[4-[[(2R)-4-imidazo[1,2-a]pyridin-2-yl-3-oxo-piperazin-2- yl]methyl]phenyl]acetamide (540 mg, 1.60 mmol, 1 eq) in DCM (6 mL) was added TEA (807.48 mg, 7.98 mmol, 1.11 mL, 5 eq) and prop-2-enoyl chloride (216.67 mg, 2.39 mmol, 194.50 μL, 1.5 eq) at 0 °C. The mixture was stirred at 0 °C for 0.5 hr. Upon completion, the reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (Condition 1, Gradient h) to give N-[4-[[(2R)-4-imidazo[1,2-a]pyridin-2-yl-3-oxo-1- prop-2-enoyl-piperazin-2-yl]methyl]phenyl]acetamide (Compound 152, 38.6 mg, 92.5 μmol, 6% yield, 100% purity) as a solid. MS (ESI) m/z 418.2 [M+H] + . 1 H NMR (400 MHz, DMSO-d6) δ = 9.86 (d, J = 7.4 Hz, 1H), 8.61 (t, J = 6.7 Hz, 1H), 8.41 (d, J = 10.4 Hz, 1H), 7.55 - 7.37 (m, 3H), 7.25 (t, J = 7.9 Hz, 1H), 7.05 (br dd, J = 8.3, 18.8 Hz, 2H), 6.93 (t, J = 6.6 Hz, 1H), 6.86 - 6.20 (m, 1H), 6.17 - 5.85 (m, 1H), 5.77 - 5.35 (m, 1H), 5.20 - 4.89 (m, 1H), 4.61 - 4.22 (m, 1H), 4.13 - 3.70 (m, 2H), 3.30 - 3.06 (m, 3H), 2.00 (d, J = 3.3 Hz, 3H). An analogous method was followed to obtain the following compound. Step 11: N-[4-[(4-imidazo[1,2-a]pyridin-2-yl-3-oxo-1-prop-2-enoyl-pip erazin-2- yl)methyl]phenyl]acetamide N-[4-[[(2R)-4-imidazo[1,2-a]pyridin-2-yl-3-oxo-1-prop-2-enoy l-piperazin-2-yl]methyl]- phenyl]acetamide (Compound 152, 6 mg, 14.4 µmol, 1 eq) and N-[4-[[(2S)-4-imidazo[1,2- a]pyridin-2-yl-3-oxo-1-prop-2-enoyl-piperazin-2-yl]methyl]ph enyl]acetamide (Compound 151, 6 mg, 14.37 µmol, 1 eq) was dissolved in MeCN 0.5 mL and H2O 2 mL, then lyophilized to give N-[4-[(4-imidazo[1,2-a]pyridin-2-yl-3-oxo-1-prop-2-enoyl-pip erazin-2-yl)methyl]phenyl] acetamide (Compound 144, 10.4 mg, 25 µmol, 100% yield). MS (ESI) m/z 418.2 [M+H] + . 1 H NMR (400 MHz, DMSO-d6) δ = 9.85 (br d, J = 7.2 Hz, 1H), 8.61 (br t, J = 6.4 Hz, 1H), 8.41 (d, J = 10.2 Hz, 1H), 7.55 - 7.41 (m, 3H), 7.25 (br t, J = 7.8 Hz, 1H), 7.05 (br dd, J = 8.2, 18.8 Hz, 2H), 6.93 (t, J = 6.6 Hz, 1H), 6.86 - 6.21 (m, 1H), 6.16 - 5.85 (m, 1H), 5.77 - 5.36 (m, 1H), 5.18 - 4.91 (m, 1H), 4.58 - 4.24 (m, 1H), 4.12 - 3.71 (m, 2H), 3.29 - 3.06 (m, 3H), 2.00 (br d, J = 3.0 Hz, 3H). Example 30: Synthesis of Compound 150 Step 1: methyl (2R)-2-[2-(tert-butoxycarbonylamino)ethylamino]-3-(1H-indol- 3-yl)propanoate To a solution of tert-butyl N-(2-oxoethyl)carbamate (1 g, 6.3 mmol, 1 eq) and methyl (2R)- 2-amino-3-(1H-indol-3-yl)propanoate (1.9 g, 7.5 mmol, 1.2 eq, HCl) in MeOH (40 mL), was added NaBH 3 CN (592 mg, 9.42 mmol, 1.5 eq), AcOH (528.2 mg, 8.79 mmol, 503 uL, 1.4 eq) at 0 °C under N 2 atmosphere, and then the mixture was stirred at 25 °C for 1 h under N 2 atmosphere. Upon completion, the reaction mixture was quenched by addition of sat. aq. NaHCO3 (10 mL) at 0 °C, and then diluted with H2O (20 mL) and extracted with DCM (20 mL x3). The combined organic layers were washed with brine (10 mL), dried over Na 2 SO 4 , filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (0-50% EtOAc in PE) to give methyl (2R)-2-[2-(tert-butoxycarbonylamino)ethylamino]-3-(1H-indol- 3- yl)propanoate (1.78 g, 4.9 mmol, 78% yield, 97% purity) as an oil. MS (ESI) m/z 362.2 [M+H] + . Step 2: methyl (2R)-2-[benzyloxycarbonyl-[2-(tert-butoxycarbonylamino)ethyl ]amino]-3-(1H- indol-3-yl)propanoate To a solution of methyl (2R)-2-[2-(tert-butoxycarbonylamino)ethylamino]-3-(1H-indol- 3- yl)propanoate (1.7 g, 4.70 mmol, 1 eq) in DCM (20 mL), was added CbzCl (1.20 g, 7 mmol, 1 mL, 1.5 eq) and DIEA (2.43 g, 18.8 mmol, 3.28 mL, 4 eq) at 0 °C. The mixture was stirred at 25 °C for 2.5h. Upon completion, the reaction mixture was diluted with H 2 O (10 mL) and extracted with DCM (10 mL * 3). The combined organic layers were dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (0-20% EtOAc in PE) to give methyl (2R)-2-[benzyloxycarbonyl-[2-(tert- butoxycarbonylamino)ethyl]amino]-3-(1H-indol-3-yl)propanoate (1.54 g, 3.1 mmol, 66% yield, 100% purity) as an oil. MS (ESI) m/z 396.3 [M+H-Boc] + . Step 3: methyl (2R)-2-[2-aminoethyl(benzyloxycarbonyl)amino]-3-(1H-indol-3- yl)propanoate A solution of methyl (2R)-2-[benzyloxycarbonyl-[2-(tert-butoxycarbonylamino)- ethyl]amino]-3-(1H-indol-3-yl)propanoate (1.54 g, 3.11 mmol, 1 eq) and TFA (11.86 g, 104 mmol, 7.70 mL, 33.5 eq) in DCM (24 mL), the mixture was stirred at 25 °C for 1 h. Upon completion, the reaction mixture was concentrated under reduced pressure to give methyl (2R)-2-[2- aminoethyl(benzyloxycarbonyl)amino]-3-(1H-indol-3-yl)propano ate (1.2 g, crude) as an oil. MS (ESI) m/z 396.2 [M+H] + . To solution of methyl (2R)-2-[2-aminoethyl(benzyloxycarbonyl)amino]-3-(1H-indol-3- yl)propanoate (1.2 g, 3.03 mmol, 1 eq) in DMF (25 mL), was added Cs 2 CO 3 (2.47 g, 7.6 mmol, 2.5 eq), the mixture was stirred at 80 °C for 2 h. Upon completion, the reaction mixture was diluted with H 2 O (30 mL) and extracted with DCM (30 mL * 3). The combined organic layers were washed with brine (30 mL), dried over Na 2 SO 4 , filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (50-100% EtOAc in PE) to give benzyl (2R)-2-(1H-indol-3-ylmethyl)-3-oxo-piperazine-1-carboxylate (1.1 g, 3 mmol, 99.8% yield, 97.7% purity) as a solid. MS (ESI) m/z 364.2 [M+H] + . Step 5: tert-butyl 3-[[(2R)-1-benzyloxycarbonyl-3-oxo-piperazin-2-yl]methyl]ind ole-1- carboxylate To a solution of benzyl (2R)-2-(1H-indol-3-ylmethyl)-3-oxo-piperazine-1-carboxylate (1 g, 2.75 mmol, 1 eq) and DMAP (33.6 mg, 275.2 µmol, 0.1 eq) in DCM (10 mL), was added tert- butoxycarbonyl tert-butyl carbonate (540.5 mg, 2.48 mmol, 568.9 uL, 0.9 eq) at 0 °C, the mixture was stirred at 25 °C for 1 h. Upon completion, the reaction mixture was added aq.citric acid (5 mL), and extracted with DCM (10 mL * 2). The combined organic layers were washed with NaHCO3 (10 mL) and brine (20 mL), dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (50-100% EtOAc in PE) to give tert-butyl 3-[[(2R)-1-benzyloxycarbonyl-3-oxo-piperazin-2-yl]methyl]ind ole-1- carboxylate (400 mg, 863 µmol, 31% yield) as an oil. MS (ESI) m/z 464.2 [M+H] + . Step 6: tert-butyl 3-[[(2R)-1-benzyloxycarbonyl-4-imidazo[1,2-a]pyridin-2-yl-3- oxo-piperazin-2- yl]methyl]indole-1-carboxylate To a solution of tert-butyl 3-[[(2R)-1-benzyloxycarbonyl-3-oxo-piperazin-2- yl]methyl]indole-1-carboxylate (410 mg, 884.5 µmol, 1 eq), 2-bromoimidazo[1,2-a]pyridine (174.3 mg, 884.5 µmol, 1 eq), CuI (25.3 mg, 132.7 µmol, 0.15 eq), DMEDA (19.5 mg, 221 µmol, 23.8 uL, 0.25 eq) and K2CO3 (244.5 mg, 1.77 mmol, 2 eq) in dioxane (10 mL) was degassed and purged with N 2 (x3), and then the mixture was stirred at 100 °C for 16 h under N 2 atmosphere. Upon completion, the reaction mixture was quenched by addition H 2 O (10 mL), and extracted with DCM (10 mL * 3). The combined organic layers were, dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (0- 50% EtOAc in PE) to give tert-butyl 3-[[(2R)-1-benzyloxycarbonyl-4-imidazo[1,2-a]pyridin-2-yl- 3-oxo-piperazin-2-yl]methyl]indole-1-carboxylate (250 mg, 431 µmol, 49% yield) as an oil. MS (ESI) m/z 580.2 [M+H] + . Step 7: tert-butyl 3-[[(2R)-4-imidazo[1,2-a]pyridin-2-yl-3-oxo-piperazin-2-yl]m ethyl]indole-1- carboxylate
To a solution of tert-butyl 3-[[(2R)-1-benzyloxycarbonyl-4-imidazo[1,2-a]pyridin-2-yl-3- oxo-piperazin-2-yl]methyl]indole-1-carboxylate (250 mg, 431.3 µmol, 1 eq) in i-PrOH (5 mL) was added and Pd/C (100 mg, 10% purity), the mixture was degassed and purged with H 2 (x3), and stirred at 25 °C for 30 min at 15 Psi under H2. Upon completion, the reaction mixture was filtered and concentrated under reduced pressure to give tert-butyl 3-[[(2R)-4-imidazo[1,2-a]pyridin-2-yl- 3-oxo-piperazin-2-yl]methyl]indole-1-carboxylate (185 mg, crude) as an oil. MS (ESI) m/z 446.3 [M+H] + . To a solution of tert-butyl 3-[[(2R)-4-imidazo[1,2-a]pyridin-2-yl-3-oxo-piperazin-2- yl]methyl]indole-1-carboxylate (180 mg, 404 µmol, 1 eq) in DCM (3 mL) was added TFA (1.54 g, 13.5 mmol, 1 mL, 33.4 eq), the mixture was stirred at 25 °C for 30 min. Upon completion, the reaction mixture was concentrated under reduced pressure to give (3R)-1-imidazo[1,2-a]pyridin- 2-yl-3-(1H-indol-3-ylmethyl)piperazin-2-one (135 mg, crude) as an oil. MS (ESI) m/z 346.3 [M+H] + . Step 9: (3R)-1-imidazo[1,2-a]pyridin-2-yl-3-(1H-indol-3-ylmethyl)-4- prop-2-enoyl-piperazin-2- one
To a solution of (3R)-1-imidazo[1,2-a]pyridin-2-yl-3-(1H-indol-3-ylmethyl)pip erazin-2- one (135 mg, 391 µmol, 1 eq) and TEA (237 mg, 2.35 mmol, 326.4 uL, 6 eq) in DCM (10 mL) and then prop-2-enoyl chloride (31.8 mg, 352 µmol, 29 uL, 0.9 eq) was added at 0 °C, the solution was stirred for 30 min at 0 °C. Upon completion, the reaction mixture was concentrated to give a residue. The residue was purified by prep-HPLC (Condition 4, Gradient j) to give (3R)-1- imidazo[1,2-a]pyridin-2-yl-3-(1H-indol-3-ylmethyl)-4-prop-2- enoyl-piperazin-2-one (Compound 150, 45 mg, 113 μmol, 29% yield, 100% purity) as a solid. MS (ESI) m/z 400.2 [M+H] + . 1 H NMR (400 MHz, DMSO-d 6 ) δ = 10.99 - 10.70 (m, 1H), 8.62 (t, J = 7.4 Hz, 1H), 8.45 (d, J = 17.8 Hz, 1H), 7.61 - 7.38 (m, 2H), 7.37 - 7.15 (m, 2H), 7.14 - 6.68 (m, 4H), 6.33 - 4.86 (m, 3H), 5.20 - 4.86 (m, 1H), 4.57 - 4.18 (m, 1H), 4.06 - 3.63 (m, 2H), 3.60 - 3.35 (m, 2H), 3.30 - 2.91 (m, 1H). An analogous method was followed to obtain the following compound. Step 10: 1-imidazo[1,2-a]pyridin-2-yl-3-(1H-indol-3-ylmethyl)-4-prop- 2-enoyl-piperazin-2-one
(3R)-1-imidazo[1,2-a]pyridin-2-yl-3-(1H-indol-3-ylmethyl)-4- prop-2-enoyl-piperazin-2-one (Compound 150, 7 mg, 17.5 μmol, 1 eq) and (3S)-1-imidazo[1,2-a]pyridin-2-yl-3-(1H-indol-3- ylmethyl)-4-prop-2-enoyl-piperazin-2-one (Compound 149, 7 mg, 17.5 μmol, 1 eq) was dissolved in ACN (2 mL) and H 2 O (10 mL) and lyophilized to give 1-imidazo[1,2-a]pyridin-2-yl-3-(1H- indol-3-ylmethyl)-4-prop-2-enoyl-piperazin-2-one (Compound 143, 12 mg, crude) as a solid. MS (ESI) m/z 400.1 [M+H] + . 1 H NMR (400 MHz, DMSO-d6) δ = 10.99 - 10.72 (m, 1H), 8.62 (t, J = 7.4 Hz, 1H), 8.45 (d, J = 17.8 Hz, 1H), 7.60 - 7.39 (m, 2H), 7.35 - 7.16 (m, 2H), 7.08 - 6.88 (m, 4H), 6.83 - 5.22 (m, 3H), 5.21 - 4.88 (m, 1H), 4.55 - 4.16 (m, 1H), 4.06 - 3.63 (m, 2H), 3.55 - 3.34 (m, 2H), 3.18 - 2.96 (m, 1H). An analogous method to the last step was followed to obtain the following compound. Example 31: Synthesis of Compound 154 Step 1: methyl 2-(3-oxopiperazin-2-yl)acetate To a solution of dimethyl maleate (10 g, 69.38 mmol, 8.70 mL, 1 eq) in i-PrOH (100 mL) was added ethane-1, 2-diamine (4.57 g, 76.04 mmol, 5.09 mL, 1.10 eq) at 25 °C. The mixture was stirred at 50 °C for 16 h. Upon completion, the reaction mixture filtered and concentrated under reduced pressure to give a residue. The crude product was triturated with PE: EA = 1:1 at 25 °C for 30 min to give methyl 2-(3-oxopiperazin-2-yl)acetate (9.3 g, 54 mmol, 78% yield, 100% purity) as a solid. MS (ESI) m/z 173.2 [M+H] + . Step.2: tert-butyl 2-(2-methoxy-2-oxoethyl)-3-oxopiperazine-1-carboxylate To a solution of methyl 2-(3-oxopiperazin-2-yl)acetate (8.8 g, 51.1 mmol, 1 eq) in DCM (150 mL) was added TEA (7.24 g, 71.5 mmol, 9.96 mL, 1.4 eq) and Boc2O (11.15 g, 51.1 mmol, 11.7 mL, 1 eq). The mixture was stirred at 25 °C for 1 h. Upon completion, the reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (33-100% EtOAc in Petroleum ether) to give tert-butyl 2-(2-methoxy-2- oxoethyl)-3-oxopiperazine-1-carboxylate (13.2 g, 48.5 mmol, 95% yield, 100% purity) as a solid. MS (ESI) m/z 273.1 [M+H] + . Step.3: tert-butyl 4-(imidazo[1,2-a]pyridin-2-yl)-2-(2-methoxy-2-oxoethyl)-3-ox opiperazine-1- carboxylate A solution of tert-butyl 2-(2-methoxy-2-oxoethyl)-3-oxopiperazine-1-carboxylate (5 g, 18.4 mmol, 1 eq) in dioxane (20 mL) was added K 2 CO 3 (5.1 g, 36.7 mmol, 2 eq) and N,N'- dimethylethane-1,2-diamine (1.29 g, 14.7 mmol, 1.58 mL, 0.8 eq). Then CuI (1.40 g, 7.34 mmol, 0.4 eq) and 2-bromoimidazo[1,2-a]pyridine (4.34 g, 22.03 mmol, 1.2 eq) was added. The mixture was stirred at 100 °C for 16 h. Upon completion, the reaction mixture was quenched by addition H2O (20 mL) at 25 °C, and then diluted with EtOAc (15 mL) and extracted with EtOAc (15 mL * 3). The combined organic layers were washed with brine (15 mL * 3), dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (18-50% EtOAc in PE) to give tert-butyl 4-(imidazo[1,2-a]pyridin-2-yl)-2-(2- methoxy-2-oxoethyl)-3-oxopiperazine-1-carboxylate (2.5 g, 6.2 mmol, 34% yield, 97% purity) as a solid. MS (ESI) m/z 389.1 [M+H] + . Step.4: methyl 2-(4-(imidazo[1,2-a]pyridin-2-yl)-3-oxopiperazin-2-yl)acetat e A mixture of tert-butyl 4-(imidazo[1,2-a]pyridin-2-yl)-2-(2-methoxy-2-oxoethyl)-3- oxopiperazine-1-carboxylate (500 mg, 1.29 mmol, 1 eq) in HCl/dioxane (4 M, 25 mL, 77.7 eq) was stirred at 25 °C for 1 h. Upon completion, the reaction mixture was concentrated under N2 to give methyl 2-(4-(imidazo[1,2-a]pyridin-2-yl)-3-oxopiperazin-2-yl)acetat e (400 mg, crude) as an oil. MS (ESI) m/z 289.1 [M+H] + . Step 5: methyl 2-(1-acryloyl-4-(imidazo[1,2-a]pyridin-2-yl)-3-oxopiperazin- 2-yl)acetate To a solution of methyl 2-(4-(imidazo[1,2-a]pyridin-2-yl)-3-oxopiperazin-2-yl)acetat e (371 mg, 1.29 mmol, 1 eq) in DCM (2 mL) was added TEA (390.64 mg, 3.86 mmol, 537.34 uL, 3 eq) at 0 °C, then added a solution of prop-2-enoyl chloride (139.76 mg, 1.54 mmol, 125.91 uL, 1.2 eq) in DCM (0.5 mL) to the mixture and was stirred at 25 °C for 1 h. Upon completion, the reaction mixture was quenched by addition H2O (1 mL) at 25 °C and concentrated under N2 to give a residue. The residue was purified by prep-HPLC (Condition 6, Gradient c) to give methyl 2-(1-acryloyl-4-(imidazo[1,2-a]pyridin-2-yl)-3-oxopiperazin- 2-yl)acetate (Compound 154, 500 mg, crude) as an oil. MS (ESI) m/z 343.1 [M+H] + . 1 H NMR (400 MHz, DMSO-d6): δ = 8.54 (d, J = 6.8 Hz, 1H), 8.32 (s, 1H), 7.49 (d, J = 9.0 Hz, 1H), 7.18-7.27 (m, 1H), 6.74-6.95 (m, 2H), 6.17 (dd, J = 16.8, 2.1 Hz, 1H), 5.76 (dd, J = 10.6, 2.2 Hz, 1H), 5.20 (t, J = 6.0 Hz, 1H), 4.28-4.49 (m, 2H), 3.81-3.98 (m, 1H), 3.63-3.76 (m, 1H), 3.62 (s, 3H), 2.98 ppm (t, J = 5.8 Hz, 2H). Example 32: Synthesis of Compound 155 Step 1: 2-(1-tert-butoxycarbonyl-4-imidazo[1,2-a]pyridin-2-yl-3-oxo- piperazin-2-yl)acetic acid To a solution of tert-butyl 4-imidazo[1,2-a]pyridin-2-yl-2-(2-methoxy-2-oxo-ethyl)-3- oxo- piperazine-1-carboxylate (1.5 g, 3.86 mmol, 1 eq) in THF (15 mL) and H2O (5 mL) was added LiOH.H2O (486.17 mg, 11.59 mmol, 3 eq). The mixture was stirred at 25 °C for 2 h. Upon completion, the reaction mixture was quenched by addition HCl (1M, 15 mL) at 0 °C, and then extracted with DCM (6 mL * 8). The combined organic layers were washed with brine (15 mL), dried over Na2SO4, filtered and concentrated under reduced pressure to give the residue 2-(1-tert- butoxycarbonyl-4-imidazo[1,2-a] pyridin-2-yl-3-oxo-piperazin-2-yl)acetic acid (1.14 g, crude) as a solid. MS (ESI) m/z 375.2 [M+H] + . Step 2: 2-(4-imidazo[1,2-a]pyridin-2-yl-3-oxo-piperazin-2-yl)acetic acid A mixture of 2-(1-tert-butoxycarbonyl-4-imidazo[1,2-a]pyridin-2-yl-3-oxo- piperazin-2- yl)acetic acid (140 mg, 373.94 μmol, 1 eq) in TFA (0.5 mL) and DCM (2.5 mL). The mixture was stirred at 25 °C for 0.5 hr. Upon completion, the reaction mixture was concentrated under the reduced pressure to give 2-(4-imidazo[1,2-a]pyridin-2-yl-3-oxo-piperazin-2-yl)acetic acid (100 mg, crude) as an oil. MS (ESI) m/z 275.1 [M+H] + . Step 3: 2-(4-imidazo[1,2-a]pyridin-2-yl-3-oxo-1-prop-2-enoyl-piperaz in-2-yl)acetic acid To a solution of 2-(4-imidazo[1,2-a]pyridin-2-yl-3-oxo-piperazin-2-yl)acetic acid (100 mg, 257.53 μmol, 1 eq, TFA) in DCM (5 mL) was added TEA (78.18 mg, 772.60 μmol, 107.54 μL, 3 eq) at 0 °C, then added to a solution of prop-2-enoyl chloride (13.99 mg, 154.52 μmol, 12.60 μL, 0.6 eq) in DCM (1 mL).The mixture was stirred at 0 °C for 0.5 hr. Upon completion, the reaction mixture was concentrated under the reduced pressure to give the residue. The residue was purified by prep-HPLC (Condition 4, Gradient b) to give 2-(4-imidazo[1,2-a]pyridin-2-yl-3-oxo- 1-prop-2-enoyl-piperazin-2-yl)acetic acid (Compound 155, 24 mg, 73 μmol, 28% yield, 100% purity) as a solid. MS (ESI) m/z 329.1 [M+H] + . 1 H NMR (400 MHz, DMSO+D2O-d6): δ = 8.51 (d, J = 6.8 Hz, 1H), 8.29 (s, 1H), 7.47 (d, J = 8.9 Hz, 1H), 7.16 - 7.30 (m, 1H), 6.80 - 7.04 (m, 2H), 6.14 (dd, J = 16.7, 2.2 Hz, 1H), 5.71 (dd, J = 10.5, 2.1 Hz, 1H), 5.09 (t, J = 5.5 Hz, 1H), 4.17 - 4.50 (m, 2H), 3.53 - 3.94 (m, 2H), 2.64 - 2.91 (m, 2H). Example 33: Synthesis of Compound 156 Step 1: tert-butyl 2-[2-(cyclopropylamino)-2-oxo-ethyl]-4-imidazo[1,2-a]pyridin -2-yl-3-oxo- piperazine-1-carboxylate To a solution of 2-(1-tert-butoxycarbonyl-4-imidazo[1,2-a]pyridin-2-yl-3-oxo- piperazin- 2-yl)acetic acid (200 mg, 534.2 μmol, 1 eq) in DMF (10 mL) was added HATU (203.1 mg, 534.2 μmol, 1 eq) and DIEA (69 mg, 534 μmol, 93 μL, 1 eq). The mixture was stirred at 25 °C for 0.5 hr, then cyclopropanamine (61 mg, 1.07 mmol, 74 μL, 2 eq) was added, the mixture was stirred at 25 °C for 0.5 hr. Upon completion, the reaction mixture was poured into H2O (100 mL) at 20 °C, and then extracted with EtOAc (30 mL x3). The combined organic layers were washed with brine (30 mL), dried over Na 2 SO 4 , filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (9-100% EtOAc in PE) to give tert-butyl 2- [2-(cyclopropylamino)-2-oxo-ethyl]-4-imidazo[1,2-a]pyridin-2 -yl-3-oxo-piperazine-1- carboxylate (160 mg, 348.3 μmol, 65% yield, 90% purity) as a solid. MS (ESI) m/z 414.2 [M+H] + . Step 2: N-cyclopropyl-2-(4-imidazo[1,2-a]pyridin-2-yl-3-oxo-piperazi n-2-yl)acetamide To a solution of tert-butyl 2-[2-(cyclopropylamino)-2-oxo-ethyl]-4-imidazo[1,2-a]pyridin - 2-yl-3-oxo-piperazine-1-carboxylate (160 mg, 387 μmol, 1 eq) in DCM (3 mL) and TFA (4.6 g, 40.4 mmol, 3 mL, 104.4 eq). The mixture was stirred at 20 °C for 1 hr. Upon completion, the reaction mixture was concentrated under the reduced pressure to give N-cyclopropyl-2-(4- imidazo[1,2-a]pyridin-2-yl-3-oxo-piperazin-2-yl)acetamide (100 mg, crude) as a solid. MS (ESI) m/z 314.2 [M+H] + . Step 3: N-cyclopropyl-2-(4-imidazo[1,2-a]pyridin-2-yl-3-oxo-1-prop-2 -enoyl-piperazin-2- yl)acetamide To a solution of N-cyclopropyl-2-(4-imidazo[1,2-a]pyridin-2-yl-3-oxo-piperazi n-2- yl)acetamide (100 mg, 319 μmol, 1 eq) in DCM (5 mL) was added TEA (161.5 mg, 1.6 mmol, 222 μL, 5 eq) at 0 °C, then prop-2-enoyl chloride (43.3 mg, 478.7 μmol, 39 μL, 1.5 eq) in DCM (1 mL) was added. The mixture was stirred at 0 °C for 1 hr. Upon completion, the reaction mixture was concentrated under the reduced pressure to give the residue. The residue was purified by prep- HPLC (Condition 7, Gradient b) to give N-cyclopropyl-2-(4-imidazo[1,2-a]pyridin-2-yl-3-oxo-1- prop-2-enoyl-piperazin-2-yl)acetamide (Compound 156, 15 mg, 39 μmol, 12% yield, 95% purity) as a solid. MS (ESI) m/z 368.2 [M+H] + . 1 H NMR (400 MHz, DMSO-d6) δ = 8.59 (d, J = 6.7 Hz, 1H), 8.36 (s, 1H), 8.24 - 7.91 (m, 1H), 7.51 (d, J = 8.9 Hz, 1H), 7.34 - 7.17 (m, 1H), 6.97 - 6.78 (m, 2H), 6.29 - 6.06 (m, 1H), 5.75 (d, J = 10.1 Hz, 1H), 5.07 (s, 1H), 4.69 - 4.13 (m, 2H), 3.99 - 3.57 (m, 2H), 3.04 - 2.58 (m, 3H), 0.57 (d, J = 6.4 Hz, 2H), 0.33 (d, J = 2.1 Hz, 2H). Example 34: Synthesis of Compound 160 Step 1: methyl 2-[[3-(nitromethyl)oxetan-3-yl]amino]acetate oxetan-3-one (15 g, 208.2 mmol, 1 eq) nitromethane (19.5 g, 320 mmol, 17.3 mL, 1.54 eq) and TEA (4.21 g, 41.6 mmol, 5.8 mL, 0.2 eq) were stirred at 25 °C for 60 min. DCM (200 mL) was added and the reaction mixture cooled to -70 °C. TEA (42 g, 416.3 mmol, 57.9 mL, 2 eq) was added followed by the dropwise addition of a solution of MsCl (23.16 g, 202.2 mmol, 15.7 mL, 0.97 eq) in DCM (200 mL). The reaction mixture was left to stir at -70 °C for 90 min. Meanwhile, to a solution of methyl 2-aminoacetate; hydrochloride (52.3 g, 416.3 mmol, 2 eq) in DCM (200 mL) was added TEA (42.13 g, 416.3 mmol, 57.9 mL, 2 eq) and stirred at room temperature for 10 min. This solution was added to the oxetane mixture via syringe at -78 °C. The reaction mixture was allowed to warm to 25 °C and stirred for 16 h. Upon completion, a saturated solution of NH 4 Cl (200 mL) was added to the reaction mixture and stirred for 10 min. The layers were separated and the aqueous extracted with DCM (2 * 50 mL). The combined organic layers were washed with saturated NaHCO 3 (50 mL), dried over Na 2 SO 4 , filtered and concentrated under reduced pressure. The residue was purified by silica gel column (30% EtOAc in PE) to give methyl 2-[[3- (nitromethyl)oxetan-3-yl]amino]acetate (11 g, 54 mmol, 26% yield) as an oil. Step 2: 2-oxa-5,8-diazaspiro[3.5]nonan-7-one To a solution of methyl 2-[[3-(nitromethyl)oxetan-3-yl]amino]acetate (8 g, 39.2 mmol, 1 eq) in MeOH (400 mL) was added Raney-Ni (8 g, 93.4 mmol, 2.4 eq) under N 2 atmosphere. The suspension was degassed and purged with H 2 (x3). The mixture was stirred under H 2 (15Psi) at 25 °C for 6 h. Upon completion, the reaction was filtered and washed with MeOH (50 mL * 3), then was concentrated under reduced pressure to give 2-oxa-5,8-diazaspiro[3.5]nonan-7-one (5.5 g, crude) as an oil. Step 3: tert-butyl 7-oxo-2-oxa-5,8-diazaspiro[3.5]nonane-5-carboxylate Boc 2 O (12.7 g, 58 mmol, 13.3 mL, 1.5 eq) was added to a suspension of 2-oxa-5,8- diazaspiro[3.5]nonan-7-one (5.5 g, 39 mmol, 1 eq) in DCM (50 mL). The mixture was stirred at 25 °C for 12 h. Upon completion, the solvent was concentrated in vacuum. The crude product was triturated with PE (100 mL) at 25 o C for 2 h to give tert-butyl 7-oxo-2-oxa-5,8- diazaspiro[3.5]nonane-5-carboxylate (7.5 g, 31 mmol, 80% yield) as a solid. Step 4: tert-butyl 4-(5-methyl-2-furyl)-3-oxo-piperazine-1-carboxylate To a solution of tert-butyl 7-oxo-2-oxa-5,8-diazaspiro[3.5]nonane-5-carboxylate (800 mg, 3.30 mmol, 1 eq) and 2-bromo-5-methyl-furan (797.44 mg, 4.95 mmol, 1.5 eq) in dioxane (10 mL) was added K2CO3 (1.37 g, 9.91 mmol, 3 eq) and CuI (251.55 mg, 1.32 mmol, 0.4 eq) was degassed and purged with N 2 (x3), then N,N'-dimethylethane-1,2-diamine (232.9 mg, 2.6 mmol, 284.3 uL, 0.8 eq) was added under N2, the mixture was stirred at 100 °C for 16 h. Upon completion, the reaction mixture was concentrated under reduced pressure to remove dioxane. The residue was purified by prep-TLC (50% EtOAC in PE) to give tert-butyl 8-(5-methyl-2-furyl)-7-oxo-2-oxa- 5,8-diazaspiro[3.5]nonane-5-carboxylate (0.8 g, 2.23 mmol, 68% yield, 90% purity) as a solid. MS (ESI) m/z 323.2 [M+H] + . Step 5: 8-(5-methyl-2-furyl)-2-oxa-5,8-diazaspiro[3.5]nonan-7-one To a solution of tert-butyl 8-(5-methyl-2-furyl)-7-oxo-2-oxa-5,8-diazaspiro[3.5]nonane-5 - carboxylate (0.5 g, 1.55 mmol, 1 eq) in DCM (3 mL) was added TFA (4.62 g, 40.52 mmol, 3 mL, 26.12 eq), the solution was stirred at 25 °C for 1 h. The reaction mixture was concentrated under reduced pressure to give a 8-(5-methyl-2-furyl)-2-oxa-5,8-diazaspiro[3.5]nonan-7-one (0.4 g, crude, HCl) as an oil. MS (ESI) m/z 223.2 [M+H] + . Step 6: 8-(5-methyl-2-furyl)-5-prop-2-enoyl-2-oxa-5,8-diazaspiro[3.5 ]nonan-7-one To a solution of 8-(5-methyl-2-furyl)-2-oxa-5,8-diazaspiro[3.5]nonan-7-one (0.3 g, 1.35 mmol, 1 eq) in DCM (5 mL) was added TEA (682.98 mg, 6.75 mmol, 939.45 μL, 5 eq) at 0 °C, and prop-2-enoyl chloride (183.26 mg, 2.02 mmol, 165.10 μL, 1.5 eq) in DCM (1 mL) was added, the mixture was stirred at 0 °C for 1 h. Upon completion, the reaction mixture filtered and concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (Condition 1, Gradient h) to give 8-(5-methyl-2-furyl)-5-prop-2-enoyl-2-oxa-5,8-diazaspiro[3.5 ]- nonan-7-one (Compound 160, 20 mg, 69 μmol, 5% yield, 95% purity) as a solid. MS (ESI) m/z 277.1 [M+H] + . 1 H NMR (400 MHz, DMSO-d 6 ) δ = 6.78 - 6.58 (m, 1H), 6.26 - 6.13 (m, 2H), 6.10 (dd, J = 1.0, 3.0 Hz, 1H), 5.85 - 5.71 (m, 1H), 4.88 (d, J = 7.0 Hz, 2H), 4.45 (d, J = 6.9 Hz, 2H), 4.32 (s, 2H), 4.14 (s, 2H), 2.24 (s, 3H). An analogous method was followed to obtain the following compounds. Example 35: Synthesis of Compound 163 Step 1: 5-methylpiperazin-2-one and 6-methylpiperazin-2-one A solution of ethyl 2-chloroacetate (16.5 g, 134.9 mmol, 14.4 mL, 0.2 eq) in EtOH (600 mL) was added the solution of propane-1,2-diamine (50 g, 674.5 mmol, 57.6 mL, 1 eq) in EtOH (100 mL) drop-wise at 20 °C over 1.5 h, after 2 h K2CO3 (18.7 g, 134.9 mmol, 0.2 eq) was added. And the mixture was stirred at 20 °C for another 2 h. Upon completion, insoluble material was removed by filteration, and the filtrare was concentrated under reduced pressure to give the mixture of 5-methylpiperazin-2-one (B56A, 12.5 g, crude) and 6-methylpiperazin-2-one (B56B, 37.5 g, crude) as an oil. Step 2: tert-butyl 2-methyl-5-oxo-piperazine-1-carboxylate and tert-butyl 3-methyl-5-oxo- piperazine-1-carboxylate A mixture of 6-methylpiperazin-2-one (37.5 g, 328.53 mmol, 1 eq) and 5-methylpiperazin- 2-one (12.5 g, 109.51 mmol, 3.33e-1 eq) in DCM (1000 mL) was added (Boc)2O (93.21 g, 427.09 mmol, 98.12 mL, 1.3 eq) and the mixture was sitrred at 20 °C for 16 h. Upon completion, the reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (9-50% EtOAc in PE) to give the mixture tert-butyl 2-methyl- 5-oxo-piperazine-1-carboxylate (4 g, 18.7 mmol, 6% yield) and tert-butyl 3-methyl-5-oxo- piperazine-1-carboxylate (6 g, 28 mmol, 9% yield) as a solid. To a mixture of tert-butyl 3-methyl-5-oxo-piperazine-1-carboxylate (500 mg, 2.33 mmol, 1 eq) and tert-butyl 2-methyl-5-oxo-piperazine-1-carboxylate (500 mg, 2.33 mmol, 1 eq) and 2- bromo-5-methyl-furan (902 mg, 5.60 mmol, 2.4 eq) in dioxane (20 mL) was added CuI (178 mg, 933 μmol, 0.4 eq) and K 2 CO 3 (645 mg, 4.67 mmol, 2 eq) under N 2 . Then DMEDA (164.6 mg, 1.87 mmol, 201 μL, 0.8 eq) was added to the mixture and was stirred at 100 °C for 16 h. Upon completion, the solids were filtered out and the resulting solution was concentrated under vacuum to give a residue. The residue was purified by column chromatography (0-20% EtOAc in PE) to give tert-butyl 3-methyl-4-(5-methyl-2-furyl)-5-oxo-piperazine-1-carboxylate (240 mg, 815 μmol, 48% yield) and tert-butyl 2-methyl-4-(5-methyl-2-furyl)-5-oxo-piperazine-1-carboxylate (250 mg, 849.3 μmol, 50% yield) as an oil. MS (ESI) m/z 295.2 [M+H] + . Step 4: 5-methyl-1-(5-methyl-2-furyl)piperazin-2-one A solution of tert-butyl 2-methyl-4-(5-methyl-2-furyl)-5-oxo-piperazine-1-carboxylate (100 mg, 339.7 μmol, 1 eq) in DCM (2 mL) was added TFA (200 mg, 1.75 mmol, 130 μL, 5.15 eq) and stirred at 25 °C for 1 h. Upon completion, the reaction mixture was concentrated under reduced pressure to give 5-methyl-1-(5-methyl-2-furyl)piperazin-2-one (60 mg, crude) as an oil. MS (ESI) m/z 195.3 [M+H] + . Step 5: 5-methyl-1-(5-methyl-2-furyl)-4-prop-2-enoyl-piperazin-2-one A solution of 5-methyl-1-(5-methyl-2-furyl)piperazin-2-one (60 mg, 308.9 μmol, 1 eq) in DCM (3 mL) was added TEA (31.3 mg, 308.9 μmol, 43 μL, 1 eq) at 0 °C, then prop-2-enoyl chloride (28 mg, 308.9 μmol, 25.1 μL, 1 eq) was added. The mixture was stirred at 0 °C for 1 h. Upon completion, the reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (Condition 4, Gradient e) to give 5-methyl- 1-(5-methyl-2-furyl)-4-prop-2-enoyl-piperazin-2-one (Compound 163, 11 mg, 44 μmol, 14% yield, 100% purity) as an oil. MS (ESI) m/z 249.1 [M+H] + . 1 H NMR (400 MHz, DMSO-d 6 ) δ = 6.77 - 6.70 (m, 1H), 6.19 - 6.14 (m, 2H), 6.10 (s 1H), 5.75 (d, J = 10.6 Hz, 1H), 4.63 (s, 1H), 4.50 - 4.46 (m, 1H), 4.08 - 4.03 (m, 1H), 3.98 - 3.94 (m, 1H), 3.67 - 3.63 (m, 1H), 2.24 (s, 3H), 1.27 (d, J = 6.4 Hz, 3H). An analogous method was followed to obtain the following compounds. Example 36: Synthesis of Compound 167 Step 1: tert-butyl 4-(2,1,3-benzoxadiazol-5-yl)-3-oxo-piperazine-1-carboxylate To a solution of 5-bromo-2,1,3-benzoxadiazole (350 mg, 1.76 mmol, 1 eq) and tert-butyl 3-oxopiperazine-1-carboxylate (528.2 mg, 2.6 mmol, 1.5 eq) in dioxane (14 mL) was added Pd 2 (dba) 3 (161 mg, 176 µmol, 0.1 eq) and Xantphos (203.5 mg, 352 µmol, 0.2 eq) and then K 2 CO 3 (729 mg, 5.3 mmol, 3 eq), and the mixture was stirred for 2 h at 130 °C. Upon completion, the reaction was filtered to give crude. The crude was purified by pre-TLC (50% EtOAc in PE) to give tert-butyl 4-(2,1,3-benzoxadiazol-5-yl)-3-oxo-piperazine-1-carboxylate (400 mg, 1.26 mmol, 72% yield) as a solid. MS (ESI) m/z 319.1 [M+H] + . Step 2: 1-(2,1,3-benzoxadiazol-5-yl)piperazin-2-one A solution of tert-butyl 4-(2,1,3-benzoxadiazol-5-yl)-3-oxo-piperazine-1-carboxylate (159 mg, 499 µmol, 1 eq) in HCl/dioxane (3 mL) was stirred for 1 h at 20 °C. Upon completion, the reaction was concentrated to give 1-(2,1,3-benzoxadiazol-5-yl)piperazin-2-one (108 mg, 495 µmol, 99% yield) as a solid. MS (ESI) m/z 219.2 [M+H] + . Step 3: 1-(2,1,3-benzoxadiazol-5-yl)-4-prop-2-enoyl-piperazin-2-one To a solution of 1-(2,1,3-benzoxadiazol-5-yl)piperazin-2-one (108 mg, 495 µmol, 1 eq) in DCM (10 mL) was added TEA (150 mg, 1.5 mmol, 207 uL, 3 eq). Prop-2-enoyl chloride (58 mg, 643 µmol, 52.5 uL, 1.3 eq) was added. The mixture was stirred for 1 h at 0 °C. Upon completion, the reaction was concentrated to give crude. The crude was purified by pre-HPLC (Condition 4, Gradient b) to give 1-(2,1,3-benzoxadiazol-5-yl)-4-prop-2-enoyl-piperazin-2-one (Compound 167, 44 mg, 161.6 µmol, 33% yield) as a solid. MS (ESI) m/z 273.0 [M+H] + . 1 H NMR (400MHz, DMSO-d6) δ = 8.12 - 7.91 (m, 2H), 7.69 (d, J=9.4 Hz, 1H), 6.96 - 6.75 (m, 1H), 6.21 (d, J=16.6 Hz, 1H), 5.78 (d, J=10.0 Hz, 1H), 4.56 - 4.28 (m, 2H), 4.02 (br s, 1H), 3.99 - 3.86 (m, 3H). Example 37: Synthesis of Compound 169 Step 1: 6-bromo-2-[(4-methoxyphenyl)methyl]-[1,2,4]triazolo[4,3-a]py ridin-3-one To a solution of 6-bromo-2H-[1,2,4]triazolo[4,3-a]pyridin-3-one (800 mg, 3.74 mmol, 1 eq) and 1-(chloromethyl)-4-methoxy-benzene (643.9 mg, 4.11 mmol, 560 μL, 1.1 eq) in DMF (12 mL) was added K2CO3 (1.55 g, 11.2 mmol, 3 eq). The mixture was stirred for 5 h at 50 °C. Upon completion, the reaction was diluted with H 2 O (60mL) and extracted with EtOAc (60mL*2) and washed with brine (60mL) and concentrated to give crude. The crude was triturated with petroleum ether (20mL) to give product 6-bromo-2-[(4-methoxyphenyl)methyl]-[1,2,4]triazolo-[4,3- a]pyridin-3-one (850 mg, 2.5 mmol, 68% yield) as a solid. MS (ESI) m/z 334.1 [M+H] + . Step 2: tert-butyl 4-[2-[(4-methoxyphenyl)methyl]-3-oxo-[1,2,4]triazolo[4,3-a]p yridin-6-yl]-3- oxo-piperazine-1-carboxylate To a solution of 6-bromo-2-[(4-methoxyphenyl)methyl]-[1,2,4]triazolo[4,3-a]py ridin-3- one (230 mg, 688.28 μmol, 1 eq) and tert-butyl 3-oxopiperazine-1-carboxylate (551.27 mg, 2.75 mmol, 4 eq) in dioxane (30 mL) was added Xantphos (79.65 mg, 137.66 μmol, 0.2 eq) and Pd 2 (dba) 3 (63.03 mg, 68.83 μmol, 0.1 eq) and then K 2 CO 3 (475.62 mg, 3.44 mmol, 5 eq). The mixture was stirred for 12 h at 130 °C. Upon completion, the reaction was filtered and concentrated to give crude. The crude was purified by prep-TLC (100% EtOAc) to give tert-butyl 4-[2-[(4- methoxyphenyl)methyl]-3-oxo-[1,2,4]triazolo[4,3-a]pyridin-6- yl]-3-oxo-piperazine-1- carboxylate (170 mg, 375 μmol, 55% yield) as a solid. MS (ESI) m/z 454.2 [M+H] + . Step 3: 2-[(4-methoxyphenyl)methyl]-6-(2-oxopiperazin-1-yl)-[1,2,4]t riazolo[4,3-a]pyridin-3- one A solution of tert-butyl 4-[2-[(4-methoxyphenyl)methyl]-3-oxo-[1,2,4]triazolo[4,3- a]pyridin-6-yl]-3-oxo-piperazine-1-carboxylate (400 mg, 882 μmol, 1 eq) in DCM (15 mL) and TFA (5 mL) was stirred for 1 h at 20 °C. Upon completion, the reaction was concentrated to give 2-[(4-methoxyphenyl)methyl]-6-(2-oxopiperazin-1-yl)-[1,2,4]t riazolo[4,3-a]pyridin-3-one (311 mg, 880 μmol, 99.8% yield) as a solid and used directly in the next step. MS (ESI) m/z 354.1 [M+H] + . Step 4: 2-[(4-methoxyphenyl)methyl]-6-(2-oxo-4-prop-2-enoyl-piperazi n-1-yl)- [1,2,4]triazolo[4,3-a]pyridin-3-one To a solution of 2-[(4-methoxyphenyl)methyl]-6-(2-oxopiperazin-1-yl)- [1,2,4]triazolo[4,3-a]pyridin-3-one (311 mg, 880 μmol, 1 eq) and prop-2-enoyl chloride (95.6 mg, 1.06 mmol, 86.1 μL, 1.2 eq) in DCM (30 mL) was added TEA (267.2 mg, 2.6 mmol, 367.5 μL, 3 eq). Then prop-2-enoyl chloride (95.6 mg, 1.06 mmol, 86.1 μL, 1.2 eq) was added. The mixture was stirred for 1 h at 0 °C. Upon completion, the reaction was concentrated to give crude. The crude was purified by prep-TLC (100% EtOAc) to give 2-[(4-methoxyphenyl)methyl]-6-(2-oxo- 4-prop-2-enoyl-piperazin-1-yl)-[1,2,4]triazolo[4,3-a]pyridin -3-one (240 mg, 589 μmol, 67% yield) as a solid. MS (ESI) m/z 408.2 [M+H] + . Step 5: 6-(2-oxo-4-prop-2-enoyl-piperazin-1-yl)-2H-[1,2,4]triazolo[4 ,3-a]pyridin-3-one A solution of 2-[(4-methoxyphenyl)methyl]-6-(2-oxo-4-prop-2-enoyl-piperazi n-1-yl)- [1,2,4]triazolo[4,3-a]pyridin-3-one (110 mg, 269.99 μmol, 1 eq) in methanesulfonic acid (3 mL) was stirred for 1 h at 70°C. Upon completion, the solution was filtered to give crude. The crude was purified by prep-HPLC (Condition 9, Gradient a) to give 6-(2-oxo-4-prop-2-enoyl-piperazin- 1-yl)-2H-[1,2,4]triazolo[4,3-a]pyridin-3-one (Compound 169, 25 mg, 84.4 μmol, 31% yield, 97% purity) as a solid. MS (ESI) m/z 288.1 [M+H] + . 1 H NMR (400MHz, DMSO-d 6 ) δ = 12.55 (br s, 1H), 7.98 (s, 1H), 7.14-7.35 (m, 2H), 6.95-6.72 (m, 1H), 6.19 (br d, J = 16.6 Hz, 1H), 5.76 (br d, J = 10.4 Hz, 1H), 4.45-4.22 (m, 2H), 4.04-3.85 (m, 2H), 3.79 - 3.67 (m, 2H). Example 38: Synthesis of Compound 170 Step 1: tert-butyl 4-[5-ethoxycarbonyl-4-methyl-1-(2-trimethylsilylethoxymethyl )imidazol-2-yl]- 3-oxo-piperazine-1-carboxylate
A solution of ethyl 2-bromo-5-methyl-3-(2-trimethylsilylethoxymethyl)imidazole- 4-carboxylate (1 g, 2.75 mmol, 1 eq) and tert-butyl 3-oxopiperazine-1-carboxylate (1.65 g, 8.3 mmol, 3 eq) in DMF (8 mL) was added K 3 PO 4 (1.17 g, 5.5 mmol, 2 eq), then CuI (52.4 mg, 275.2 μmol, 0.1 eq) and 1,10-phenanthroline (148.8 mg, 826 μmol, 0.3 eq) was added under N 2 , the mixture was stirred at 100 °C for 16 h. Upon completion, the reaction mixture was poured into H 2 O 10 mL at 20 °C, and then extracted with EtOAc (20 mL * 3). The combined organic layers were washed with brine 60 mL, dried over Na 2 SO 4 , filtered and concentrated under reduced pressure to give tert-butyl 4-[5-ethoxycarbonyl-4-methyl-1-(2-trimethylsilylethoxymethyl ) imidazol-2-yl]-3-oxo-piperazine-1-carboxylate (1.1 g, crude) as a solid. MS (ESI) m/z 483.2 [M+H] + . Step 2: ethyl 4-methyl-2-(2-oxopiperazin-1-yl)-1H-imidazole-5-carboxylate A solution of tert-butyl 4-[5-ethoxycarbonyl-4-methyl-1-(2-trimethylsilylethoxymethyl )- imidazol-2-yl]-3-oxo-piperazine-1-carboxylate (100 mg, 207 μmol, 1 eq) in DCM (5 mL) was added diethyloxonio(trifluoro)boranuide (117.6 mg, 829 μmol, 102 μL, 4 eq) and the mixture was stirred at 25 °C for 1 h. Upon completion, the reaction mixture was concentrated under reduced pressure to give ethyl 4-methyl-2-(2-oxopiperazin-1-yl)-1H-imidazole-5-carboxylate (50 mg, crude) as an oil. MS (ESI) m/z 253.2 [M+H] + . Step 3: ethyl 4-methyl-2-(2-oxo-4-prop-2-enoyl-piperazin-1-yl)-1H-imidazol e-5-carboxylate
A solution of ethyl 4-methyl-2-(2-oxopiperazin-1-yl)-1H-imidazole-5-carboxylate (50 mg, 198.2 μmol, 1 eq) and TEA (60 mg, 595 μmol, 82.8 μL, 3 eq) in DCM (3 mL) was added prop-2- enoyl chloride (21.5 mg, 237.8 μmol, 19.3 μL, 1.2 eq) at 0 °C and the mixture was stirred at 0 °C for 1 h. Upon completion, the reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (Condition 2, Gradient b) to give ethyl 4-methyl- 2-(2-oxo-4-prop-2-enoyl-piperazin-1-yl)-1H-imidazole-5-carbo xylate (Compound 170, 10.8 mg, 35 μmol, 18% yield, 100% purity) as a solid. MS (ESI) m/z 307.1 [M+H] + . 1 H NMR (400 MHz, MeOD) δ = 6.90 - 6.68 (m, 1H), 6.30 (br d, J = 16.6 Hz, 1H), 5.84 (dd, J = 1.8, 10.6 Hz, 1H), 4.59 - 4.42 (m, 2H), 4.32 (q, J = 7.1 Hz, 2H), 4.17 - 3.95 (m, 4H), 2.49 (s, 3H), 1.37 (t, J = 7.1 Hz, 3H). Example 39: Synthesis of Compound 171 Step 1: methyl 2-(4-tert-butoxycarbonyl-2-oxo-piperazin-1-yl)imidazo[1,2-a] pyridine-6- carboxylate A solution of methyl 2-bromoimidazo[1,2-a]pyridine-6-carboxylate (5 g, 19.6 mmol, 1 eq) , tert-butyl 3-oxopiperazine-1-carboxylate (4.7 g, 23.5 mmol, 1.2 eq) in dioxane (100 mL) was added K2CO3 (5.4 g, 39.2 mmol, 2 eq) , CuI (1.49 g, 7.8 mmol, 0.4 eq) and N,N'-dimethylethane- 1,2-diamine (1.38 g, 15.7 mmol, 1.7 mL, 0.8 eq) under N2, the mixture was stirred at 110 °C for 16 h. Upon completion, the reaction mixture was quenched by addition H 2 O (130 mL), and then extracted with EtOAc (80 mL*3). The combined organic layers were washed with brine 200 mL, dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (15-20% EtOAc in PE) to give methyl 2-(4-tert- butoxycarbonyl-2-oxo-piperazin-1-yl)imidazo[1,2-a]pyridine-6 -carboxylate (4.8 g, 10.1 mmol, 52% yield, 79% purity) as a solid. MS (ESI) m/z 375.2 [M+H] + . Step 2: 2-(4-tert-butoxycarbonyl-2-oxo-piperazin-1-yl)imidazo[1,2-a] pyridine-6-carboxylic acid A solution of methyl 2-(4-tert-butoxycarbonyl-2-oxo-piperazin-1-yl)imidazo[1,2- a]pyridine-6-carboxylate (2.15 g, 4.6 mmol, 80% purity, 1 eq) in MeOH (40 mL) and H2O (13 mL) was added K 2 CO 3 (1.9 g, 13.8 mmol, 3 eq), the mixture was stirred at 50 °C for 2 h. Upon completion, the two batches of reaction mixtures were quenched by addition H2O 100 mL. The Two reaction mixtures were combined and added 1M HCl adjust to pH~3 and extracted with EtOAc (50mL x5). The combined organic layers were washed with brine (100 mL), dried over Na2SO4, filtered and concentrated under reduced pressure to give 2-(4-tert-butoxycarbonyl-2-oxo- piperazin-1-yl)imidazo[1,2-a]pyridine-6-carboxylic acid (2.6 g, crude) as a solid. MS (ESI) m/z 361.4 [M+H] + . Step 3: tert-butyl 4-[6-(methylcarbamoyl)imidazo[1,2-a]pyridin-2-yl]-3-oxo-pipe razine-1- carboxylate A solution of 2-(4-tert-butoxycarbonyl-2-oxo-piperazin-1-yl)imidazo[1,2-a] pyridine-6- carboxylic acid (200 mg, 554.99 µmol, 1 eq), HATU (316.5 mg, 832.5 µmol, 1.5 eq) and DIEA (143.5 mg, 1.1 mmol, 193.3 uL, 2 eq) in DCM (3 mL) was stirred at 25 °C for 0.5 h, then added methanamine;hydrochloride (45 mg, 666 µmol, 1.2 eq), the mixture was stirred at 25 °C for 1 h. Upon completion, the reaction mixture was quenched by addition H2O 30 mL, and then extracted with DCM (25 mL x3). The combined organic layers were washed with brine 50 mL, dried over Na 2 SO 4 , filtered, and concentrated under reduced pressure to give a residue. The residue was purified by prep-TLC (100% EtOAc, Rf = 0.64) to give tert-butyl 4-[6-(methylcarbamoyl) imidazo[1,2-a]pyridin-2-yl]-3-oxo-piperazine-1-carboxylate (109 mg, 277 µmol, 50% yield, 95% purity) as a solid. MS (ESI) m/z 374.2 [M+H] + . Step 4: N-methyl-2-(2-oxopiperazin-1-yl)imidazo[1,2-a]pyridine-6-car boxamide A solution of tert-butyl 4-[6-(methylcarbamoyl)imidazo[1,2-a]pyridin-2-yl]-3-oxo- piperazine-1-carboxylate (109 mg, 292 µmol, 1 eq) in DCM (0.1 mL) and TFA (3.36 g, 29.4 mmol, 2.18 mL, 101 eq) was stirred at 25 °C for 1 h. Upon completion, the reaction mixture was concentrated under reduced pressure to remove DCM. N-methyl-2-(2-oxopiperazin-1- yl)imidazo[1,2-a]pyridine-6-carboxamide (80 mg, crude) was obtained as a solid. MS (ESI) m/z 274.1 [M+H] + . Step 5: N-methyl-2-(2-oxo-4-prop-2-enoyl-piperazin-1-yl)imidazo[1,2- a]pyridine-6- carboxamide A solution of N-methyl-2-(2-oxopiperazin-1-yl)imidazo[1,2-a]pyridine-6-car boxamide (80 mg, 293 µmol, 1 eq) in DCM (7 mL) was added TEA (89 mg, 878 µmol, 122.2 uL, 3 eq), then prop-2-enoyl chloride (26.5 mg, 293 µmol, 24 uL, 1 eq) in DCM (1 mL) was added drop-wise at 0 °C, the mixture was stirred at 0 °C for 1 h. Upon completion, the reaction mixture was concentrated under reduced pressure to remove DCM. The residue was purified by prep-HPLC (Condition 7, Gradient c) to give a N-methyl-2-(2-oxo-4-prop-2-enoyl-piperazin-1-yl)imidazo- [1,2-a]pyridine-6-carboxamide (Compound 171, 30 mg, 89.3 μmol, 31% yield, 97% purity) as a solid. MS (ESI) m/z 328.1 [M+H]+. 1H NMR (400 MHz, DMSO-d 6 ) δ = 9.11 (s, 1H), 8.54 - 8.42 (m, 2H), 7.67 - 7.62 (m, 1H), 7.58 - 7.53 (m, 1H), 6.91 - 6.77 (m, 1H), 6.20 (br d, J = 16.8 Hz, 1H), 5.80 - 5.72 (m, 1H), 4.51 (s, 1H), 4.34 (s, 1H), 4.24 - 4.12 (m, 2H), 4.03 - 3.85 (m, 2H), 2.80 (d, J = 4.5 Hz, 3H). Example 40: Synthesis of Compound 172 Step 1: tert-butyl 3-oxo-2-(2-phenoxyethyl)piperazine-1-carboxylate
To solution of tert-butyl 3-oxopiperazine-1-carboxylate (8 g, 39.95 mmol, 1 eq) in THF (80 mL), was added LDA (2 M, 39.95 mL, 2 eq). The mixture was stirred at -78 °C for 30 min, then to a solution of 2-bromoethoxybenzene (5.62 g, 27.97 mmol, 0.7 eq) in THF (6 mL) was added to the mixture and stirred for 5 h. Upon completion, the mixture quenched with NH4Cl (100 mL), extrated with EtOAc (50 mL*2), the combined organic phase was washed with brine (50 mL), dried over Na 2 SO 4 , filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (33-50% EtOAc in PE) to give tert-butyl 3-oxo- 2-(2-phenoxyethyl)piperazine-1-carboxylate (1.41 g, 4.3 mmol, 11% yield, 97% purity) as a solid. MS (ESI) m/z 321.2 [M+H] + . Step 2: methyl 2-[4-tert-butoxycarbonyl-2-oxo-3-(2-phenoxyethyl)piperazin-1 -yl]imidazo[1,2- a]pyridine-6-carboxylate To a solution of methyl A mixture of tert-butyl 3-oxo-2-(2-phenoxyethyl)piperazine-1- carboxylate (300 mg, 936 μmol, 1 eq), methyl 2-bromoimidazo[1,2-a]pyridine-6-carboxylate (239 mg, 936 μmol, 1 eq), CuI (27 mg, 140.5 μmol, 0.15 eq), N,N'-dimethylethane-1,2-diamine (20.6 mg, 234 μmol, 25 μL, 0.25 eq) and K 2 CO 3 (259 mg, 1.87 mmol, 2 eq) in dioxane (10 mL) was degassed and purged with N 2 (x3), and then the mixture was stirred at 100 °C for 16 h under N 2 atmosphere. Upon completion, the reaction mixture was quenched by EDTA (10 mL), extracted with EtOAc (30 mL x3). The combined organic layers were washed with brine (30 mL), dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (30-50% EtOAc in PE) to give methyl 2-[4-tert- butoxycarbonyl-2-oxo-3-(2-phenoxyethyl)piperazin-1-yl]imidaz o[1,2-a]pyridine-6-carboxylate (100 mg, 137.5 μmol, 15% yield, 68% purity) as a solid. MS (ESI) m/z 495.2 [M+H] + . Step 3: tert-butyl 4-[6-(methylcarbamoyl)imidazo[1,2-a]pyridin-2-yl]-3-oxo-2-(2 - To a solution of methyl 2-[4-tert-butoxycarbonyl-2-oxo-3-(2-phenoxyethyl)piperazin-1 - yl]imidazo[1,2-a]pyridine-6-carboxylate (100 mg, 202.2 μmol, 1 eq) in EtOH (1 mL), was added MeNH 2 (1 g, 9.66 mmol, 30% purity, 47.8 eq). The mixture was stirred at 50 °C for 16 h. Upon completion, the reaction mixture was concentrated under reduced pressure to give tert-butyl 4-[6- (methylcarbamoyl)imidazo[1,2-a]pyridin-2-yl]-3-oxo-2-(2-phen oxyethyl)piperazine-1- carboxylate (135 mg, crude) as an oil. MS (ESI) m/z 494.2 [M+H] + . Step 4: N-methyl-2-[2-oxo-3-(2-phenoxyethyl)piperazin-1-yl]imidazo[1 ,2-a]pyridine-6- carboxamide
To a solution of tert-butyl 4-[6-(methylcarbamoyl)imidazo[1,2-a]pyridin-2-yl]-3-oxo-2- (2-phenoxyethyl)piperazine-1-carboxylate (135 mg, 273.5 μmol, 1 eq) in DCM (3 mL), was added TFA (1.54 g, 13.5 mmol, 1 mL, 49.2 eq). The mixture was stirred at 25 °C for 1 h. Upon completion, the reaction mixture was concentrated under reduced pressure to give N-methyl-2-[2-oxo-3-(2- phenoxyethyl)piperazin-1-yl]imidazo[1,2-a]pyridine-6-carboxa mide (87 mg, 221 μmol, 81% yield) as an oil. MS (ESI) m/z 394.2 [M+H] + . Step 5: N-methyl-2-[2-oxo-3-(2-phenoxyethyl)-4-prop-2-enoyl-piperazi n-1-yl]imidazo[1,2- To a solution of N-methyl-2-[2-oxo-3-(2-phenoxyethyl)piperazin-1-yl]imidazo[1 ,2- a]pyridine-6-carboxamide (77 mg, 195.7 μmol, 1 eq) in DCM (8 mL) was added TEA (99 mg, 978.6 μmol, 391.4 μL, 5 eq). Then was dropwise added prop-2-enoyl chloride (15.9 mg, 176.1 μmol, 14.4 μL, 0.9 eq) in DCM (5 ml) , the mixture was stirred at 0 °C for 1 h. Upon completion, the reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (Condition 4, Gradient c) to give N-methyl-2-[2-oxo-3-(2-phenoxyethyl)- 4-prop-2-enoyl-piperazin-1-yl]imidazo[1,2-a]pyridine-6-carbo xamide (Compound 172, 16.8 mg, 38 μmol, 19% yield, 100% purity) as a solid. MS (ESI) m/z 448.2 [M+H] + . 1 H NMR (400 MHz, DMSO-d 6 ) δ = 9.11 (s, 1H), 8.47 (br s, 2H), 7.67 - 7.62 (m, 1H), 7.60 - 7.53 (m, 1H), 7.32 - 7.21 (m, 2H), 6.91 (br s, 4H), 6.14 (br s, 1H), 5.78 - 5.51 (m, 1H), 5.24 - 4.95 (m, 1H), 4.63 - 4.27 (m, 2H), 4.22 - 3.57 (m, 4H), 2.81 (s, 3H), 2.45 - 2.36 (m, 2H). Example 41: Synthesis of Compound 173 Step 1: tert-butyl 2-(2-hydroxyethyl)-3-oxo-piperazine-1-carboxylate
A solution of tert-butyl 2-(2-methoxy-2-oxoethyl)-3-oxo-piperazine-1-carboxylate (1 g, 3.67 mmol, 1 eq) in THF (15 mL), was cooled to 0 °C and was added LAH (2.5 M, 1.76 mL, 1.2 eq) and was stirred at 0 °C for 1 h. Upon completion, the reaction mixture was quenched by addition H 2 O (0.2 mL) at 0 °C, then was added 15% NaOH(aq.) (0.2 ml) and then added H 2 O (0.6 mL) then the mixture was filtered out and the resulting solution was concentrated under vacuum to give tert-butyl 2-(2-hydroxyethyl)-3-oxo-piperazine-1-carboxylate (900 mg, crude) as an oil. MS (ESI) m/z 245.2 [M+H] + . Step 2: tert-butyl 2-(2-hydroxyethyl)-4-[6-(methylcarbamoyl)imidazo[1,2-a]pyrid in-2-yl]-3-oxo- piperazine-1-carboxylate A solution of tert-butyl 2-(2-hydroxyethyl)-3-oxo-piperazine-1-carboxylate (900 mg, 3.7 mmol, 1.1 eq) and 2-bromo-N-methyl-imidazo[1,2-a]pyridine-6-carboxamide (851 mg, 3.35 mmol, 1 eq) in dioxane (30 mL) was added K2CO3 (926 mg, 6.7 mmol, 2 eq) and DMEDA (236.2 mg, 2.7 mmol, 288 μL, 0.8 eq) then CuI (255.2 mg, 1.34 mmol, 0.4 eq) was added and stirred under N 2 at 100 °C for 16 h. Upon completion, the solids were filtered out and the resulting solution was concentrated under vacuum. The residue was purified by column chromatography (9% EtOAc in PE to 9% MEOH in EtOAc) to give tert-butyl 2-(2-hydroxyethyl)-4-[6-(methylcarbamoyl) imidazo[1,2-a]pyridin-2-yl]-3-oxo-piperazine-1-carboxylate (1 g, 2.16 mmol, 64% yield, 90% purity) as an oil. MS (ESI) m/z 418.3 [M+H] + . Step 3: tert-butyl 4-[6-(methylcarbamoyl)imidazo[1,2-a]pyridin-2-yl]-3-oxo-2-[2 -(p- tolylsulfonyloxy)ethyl]piperazine-1-carboxylate
A solution of tert-butyl 2-(2-hydroxyethyl)-4-[6-(methylcarbamoyl)imidazo[1,2- a]pyridin-2-yl]-3-oxo-piperazine-1-carboxylate (1 g, 2.40 mmol, 1 eq) in DCM (20 mL) was added TEA (363.59 mg, 3.59 mmol, 500.13 μL, 1.5 eq) and DMAP (29.27 mg, 239.55 μmol, 0.1 eq) was stirred at 0 °C. Then 4-methylbenzenesulfonyl chloride (685.03 mg, 3.59 mmol, 1.5 eq) was added and the mixture was sitrred at 0 °C for 1 h, then the mixture was stirred at 20 °C for 15 h. Upon completion, the reaction mixture was quenched by addition H2O (50 mL) at 0°C, and extracted with DCM (30 mL x3). The combined organic layers were washed with brine (50 mL), dried over Na 2 SO 4 , filtered and concentrated under reduced pressure to give tert-butyl 4-[6- (methylcarbamoyl)imidazo[1,2-a]pyridin-2-yl]-3-oxo-2-[2-(p-t olylsulfonyloxy)ethyl]piperazine- 1-carboxylate (900 mg, crude) as an oil. MS (ESI) m/z 572.3 [M+H] + . Step 4: tert-butyl 2-[2-(3-methoxycarbonylphenoxy)ethyl]-4-[6-(methylcarbamoyl) imidazo[1,2- a]pyridin-2-yl]-3-oxo-piperazine-1-carboxylate A solution of methyl 3-hydroxybenzoate (95.8 mg, 630 μmol, 1.2 eq) and t-BuOK (88.3 mg, 787 μmol, 1.5 eq) in DMF (3 mL), was added tert-butyl 4-[6-(methylcarbamoyl)imidazo[1,2- a]pyridin-2-yl]-3-oxo-2-[2-(p-tolylsulfonyloxy)ethyl]piperaz ine-1-carboxylate (300 mg x3, 524.8 μmol, 1 eq) and stirred at 20 °C for 16 h. Upon completion, the reaction mixture was poured into H2O (6 mL) at 20 °C, and then extracted with EtOAc (3 mL x3). The combined organic layers were washed with brine 5 mL, dried over Na 2 SO 4 , filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (3-100% EtOAc in PE) to give tert-butyl 2-[2-(3-methoxycarbonylphenoxy)ethyl]-4-[6-(methylcarbamoyl) imidazo[1,2- a]pyridin-2-yl]-3-oxo-piperazine-1-carboxylate (350 mg, 381 μmol, 24% yield, 60% purity) as an oil. MS (ESI) m/z 552.3 [M+H] + . Step 5: 3-[2-[1-tert-butoxycarbonyl-4-[6-(methylcarbamoyl)imidazo[1, 2-a]pyridin-2-yl]-3-oxo- piperazin-2-yl]ethoxy]benzoic acid A solution of tert-butyl 2-[2-(3-methoxycarbonylphenoxy)ethyl]-4-[6-(methylcarbamoyl) imidazo[1,2-a]pyridin-2-yl]-3-oxo-piperazine-1-carboxylate (350 mg, 634.5 μmol, 1 eq) in THF (2 mL), then LiOH.H 2 O (79.9 mg, 1.9 mmol, 3 eq) in H 2 O (0.6 mL) was added and stirred at 20 °C for 16 h. Upon completion, the reaction mixture was quenched by addition H2O (2 mL). The reaction mixture was added 1M HCl adjust to pH=3 and extracted with EtOAc (1 mL x3). The combined organic layers were washed with brine (1 mL), dried over Na 2 SO 4 , filtered and concentrated under reduced pressure to give 3-[2-[1-tert-butoxycarbonyl-4-[6- (methylcarbamoyl)imidazo[1,2-a]pyridin-2-yl]-3-oxo-piperazin -2-yl]ethoxy]benzoic acid (150 mg, crude) as an oil. MS (ESI) m/z 538.2 [M+H] + . Step 6: 3-[2-[4-[6-(methylcarbamoyl)imidazo[1,2-a]pyridin-2-yl]-3-ox o-piperazin-2- yl]ethoxy]benzoic acid
A solution of 3-[2-[1-tert-butoxycarbonyl-4-[6-(methylcarbamoyl)imidazo[1, 2-a]pyridin- 2-yl]-3-oxo-piperazin-2-yl]ethoxy]benzoic acid (150 mg, 279 μmol, 1 eq) in DCM (1 mL), was added TFA (0.2 mL) was stirred at 20 °C for 1 h. The reaction mixture was concentrated under reduced pressure to give 3-[2-[4-[6-(methylcarbamoyl)imidazo[1,2-a]pyridin-2-yl]-3-ox o- piperazin-2-yl]ethoxy]benzoic acid (120 mg, crude) as an oil. MS (ESI) m/z 438.3 [M+H] + . Step 7: 3-[2-[4-[6-(methylcarbamoyl)imidazo[1,2-a]pyridin-2-yl]-3-ox o-1-prop-2-enoyl- piperazin-2-yl]ethoxy]benzoic acid A solution of 3-[2-[4-[6-(methylcarbamoyl)imidazo[1,2-a]pyridin-2-yl]-3-ox o-piperazin- 2-yl]ethoxy]benzoic acid (120 mg, 274.3 μmol, 1 eq) in DCM (1 mL) was added TEA (138.8 mg, 1.4 mmol, 191 μL, 5 eq) and was cooled to 0 °C, then prop-2-enoyl chloride (24.8 mg, 274.3 μmol, 22.3 μL, 1 eq) in DCM (0.5 mL) was added dropwise and was stirred at 0 °C for 0.5 h. Upon completion, the reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (Condition 9, Gradient b) to give 3-[2-[4-[6- (methylcarbamoyl)imidazo[1,2-a]pyridin-2-yl]-3-oxo-1-prop-2- enoyl-piperazin-2-yl]ethoxy]- benzoic acid (Compound 173, 10.1 mg, 20.6 μmol, 7.5% yield, 100% purity) as a solid. MS (ESI) m/z 492.2 [M+H] + . 1 H NMR (400 MHz, DMSO-d6) δ = 13.14 - 12.83 (s, 1H), 9.11 (s, 1H), 8.59 - 8.44 (m, 2H), 7.69 - 7.62 (m, 1H), 7.61 - 7.47 (m, 2H), 7.46 - 7.34 (m, 2H), 7.24 - 7.06 (m, 1H), 6.97 - 6.74 (m, 1H), 6.22 - 6.02 (m, 1H), 5.80 - 5.47 (m, 1H), 5.27 - 4.98 (m, 1H), 4.67 - 4.18 (m, 3H), 4.13 - 3.80 (m, 3H), 2.80 (d, J = 4.5 Hz, 3H), 2.48 - 2.36 (m, 2H). Example 42: Synthesis of Compound 174 Step 1 : dimethyl (2-((tert-butoxycarbonyl)amino)ethyl)glutamate To a solution of tert-butyl N-(2-oxoethyl)carbamate (7 g, 43.97 mmol, 1 eq) in MeOH (100 mL) was added dimethyl 2-aminopentanedioate;hydrochloride (10.9 g, 44 mmol, 1 eq, HCl), NaBH 3 CN (4.15 g, 66 mmol, 1.5 eq) and AcOH (3.7 g, 61.6 mmol, 3.5 mL, 1.4 eq) at 0 °C, the mixture was stirred at 25 °C for 1 h. Upon completion, the reaction was poured into H2O (200 mL), then extracted with DCM (100 mL*3), the combined organic layers were washed with sat. NaCl (100 mL*3), dried over Na 2 SO 4 , filtered and concentrated under reduced pressure to give dimethyl 2-[2-(tert-butoxycarbonylamino)ethylamino]pentanedioate (14 g, crude) as an oil. Step 2 : dimethyl N-((benzyloxy)carbonyl)-N-(2-((tert-butoxycarbonyl)amino)eth yl)glutamate To a solution of dimethyl 2-[2-(tert-butoxycarbonylamino)ethylamino]pentanedioate (14 g, 44 mmol, 1 eq) in DCM (100 mL) was added CbzCl (11.3 g, 66 mmol, 9.4 mL, 1.5 eq) and DIEA (17 g, 131.9 mmol, 23 mL, 3 eq) at 0 °C, the mixture was stirred at 25 °C for 1 h. Upon completion, the reaction mixture was diluted with H 2 O (150 mL), extracted with DCM (100 mL*2) and washed with sat. NaCl (100 mL*2). The combined organic layers were dried over Na 2 SO 4 , filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (0-50% EtOAc in PE) to give dimethyl 2-[benzyloxycarbonyl-[2-(tert- butoxycarbonylamino)ethyl]amino]pentanedioate (11.6 g, 26 mmol, 58% yield) as an oil. Step3 : dimethyl N-(2-aminoethyl)-N-((benzyloxy)carbonyl)glutamate A mixture of dimethyl 2-[benzyloxycarbonyl-[2-(tert-butoxycarbonylamino) ethyl]amino]pentanedioate (11.6 g, 25.64 mmol, 1 eq) and HCl/dioxane (4 M, 116 mL, 18.1 eq) was stirred at 25 °C for 1 h. Upon completion, the reaction was concentrated in vacuum to give dimethyl 2-[2-aminoethyl (benzyloxycarbonyl) amino] pentanedioate (9 g, crude) as an oil. Step4 : benzyl 2-(3-methoxy-3-oxopropyl)-3-oxopiperazine-1-carboxylate To a solution of dimethyl 2-[2-aminoethyl(benzyloxycarbonyl)amino]pentanedioate (9 g, 25.5 mmol, 1 eq) in DMF (90 mL) was added Cs2CO3 (20.8 g, 64 mmol, 2.5 eq). The mixture was stirred at 25 °C for 2 h. Upon completion, the reaction mixture was diluted with H 2 O (100 mL) and extracted with DCM (100 mL*3). The combined organic layers were washed with sat.NaCl (100 mL*2), dried over Na2SO4, filtered and concentrated under reduced pressure to give benzyl 2-(3-methoxy-3-oxo-propyl)-3-oxo-piperazine-1-carboxylate (7 g, crude) as a solid. Step5 : 3-(1-((benzyloxy)carbonyl)-3-oxopiperazin-2-yl)propanoic acid To a solution of benzyl 2-(3-methoxy-3-oxo-propyl)-3-oxo-piperazine-1-carboxylate (1 g, 3.1 mmol, 1 eq) in H 2 O (2.3 mL) and MeOH (7 mL) was added K 2 CO 3 (1.3 g, 9.4 mmol, 3 eq). The mixture was stirred at 50 °C for 1 h. Upon completion, the reaction mixture was added 1M HCl adjust to pH~2, then extracted with EtOAc (15 mL*3). The combined organic layers were washed with sat. NaCl (20 mL*2), dried over Na 2 SO 4 , filtered and concentrated under reduced pressure to give 3-(1-benzyloxycarbonyl-3-oxo-piperazin-2-yl) propanoic acid (800 mg, crude) as an oil. Step6 : 3-(1-((benzyloxy)carbonyl)-4-(6-(methoxycarbonyl)imidazo[1,2 -a]pyridin-2-yl)-3- oxopiperazin-2-yl)propanoic acid To a solution of 3-(1-benzyloxycarbonyl-3-oxo-piperazin-2-yl)propanoic acid (100 mg, 327 μmol, 1 eq) and methyl 2-bromoimidazo[1,2-a]pyridine-6-carboxylate (83.3 mg, 326.5 μmol, 1 eq) in dioxane (20 mL) was added K 2 CO 3 (90.2 mg, 653 μmol, 2 eq) and DMEDA (7.2 mg, 81.6 μmol, 8.8 μL, 0.25 eq), then exchange N2 (x3), then added CuI (9.33 mg, 49 μmol, 0.15 eq) and exchange N2 (x3), the mixture was stirred at 100 °C for 16 h. Upon completion, the reaction mixture was diluted with H 2 O (30 mL) and extracted with DCM (30 mL*3). The combined organic layers were washed with sat. NaCl (30 mL*2), dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (0-100% EtOAc in PE) to give 3-[1-benzyloxycarbonyl-4-(6-methoxycarbonylimidazo[1,2-a]pyr idin-2-yl)- 3-oxo-piperazin-2-yl]propanoic acid (110 mg, 229 μmol, 9% yield) as a solid. Step7 : 3-(1-((benzyloxy)carbonyl)-4-(6-(methylcarbamoyl)imidazo[1,2 -a]pyridin-2-yl)-3- oxopiperazin-2-yl)propanoic acid
3-[1-benzyloxycarbonyl-4-(6-methoxycarbonylimidazo[1,2-a]pyr idin-2-yl)-3-oxo- piperazin-2-yl]propanoic acid (110 mg, 229 μmol, 1 eq) was added to MeNH2/EtOH (2 g, 19.3 mmol, 30% purity, 84.4 eq), the mixture was stirred for 12 h at 50 °C. Upon completion, the reaction mixture was concentrated in vacuum to give 3-[1-benzyloxycarbonyl-4-[6- (methylcarbamoyl)imidazo[1,2-a]pyridin-2-yl]-3-oxo-piperazin -2-yl]propanoic acid (105 mg, crude) as an oil. Step8 : 3-[4-[6-(methylcarbamoyl)imidazo[1,2-a]pyridin-2-yl]-3-oxo-p iperazin-2-yl]propanoic acid To a solution of 3-[1-benzyloxycarbonyl-4-[6-(methylcarbamoyl)imidazo[1,2-a]p yridin-2- yl]-3-oxo-piperazin-2-yl]propanoic acid (105 mg, 219 μmol, 1 eq) in EtOH (2 mL) was added H2(15 psi) and Pd/C (105 mg, 219 μmol, 10% purity). The mixture was stirred at 25 °C for 3 h. Upon completion, the residue was filtered and washed by H 2 O and EtOH, then the filtrate was concentrated in vacuum to give 3-[4-[6-(methylcarbamoyl)imidazo[1,2-a]pyridin-2-yl]-3-oxo- piperazin-2-yl]propanoic acid (70 mg, crude) as a solid. Step 9 : 3-(1-acryloyl-4-(6-(methylcarbamoyl)imidazo[1,2-a]pyridin-2- yl)-3-oxopiperazin-2- yl)propanoic acid To a solution of 3-[4-[6-(methylcarbamoyl)imidazo[1,2-a]pyridin-2-yl]-3-oxo-p iperazin- 2-yl]propanoic acid (70 mg, 202.7 μmol, 1 eq) in THF (1 mL) was added KOH (17 mg, 304 μmol, 1.5 eq) in H2O (1 mL) and prop-2-enoyl chloride (18.4 mg, 202.7 μmol, 16.5 μL, 1 eq) at 0 °C. The mixture was stirred at 0 °C for 0.5 h. Upon completion, the mixture was concentrated in vacuum to give a residue. The residue was purified by prep-HPLC (Condition 9, Gradient c) to give 3-[4-[6-(methylcarbamoyl)imidazo[1,2-a]pyridin-2-yl]-3-oxo-1 -prop-2-enoyl-piperazin-2- yl]propanoic acid (Compound 174, 12.8 mg, 31.5 μmol, 16% yield, 98.7% purity) as a solid. MS (ESI): m/z 400.2 [M+H] + . 1 H NMR (400 MHz, DMSO-d 6 ) δ = 12.59 - 11.65 (m, 1H), 9.13 - 9.09 (m, 1H), 8.57 - 8.42 (m, 2H), 7.68 - 7.51 (m, 2H), 6.97 - 6.81 (m, 1H), 6.30 - 6.12 (m, 1H), 5.85 - 5.69 (m, 1H), 5.12 - 4.76 (m, 1H), 4.54 - 4.19 (m, 2H), 4.06 - 3.69 (m, 2H), 2.83 - 2.78 (m, 3H), 2.37 - 2.06 (m, 4H). Example 43: Synthesis of Compound 175 Step 1: benzyl 2-(3-hydroxypropyl)-3-oxo-piperazine-1-carboxylate To a solution of benzyl 2-(3-methoxy-3-oxo-propyl)-3-oxo-piperazine-1-carboxylate (1.2 g, 3.75 mmol, 1 eq) in THF (15 mL) was added LiAlH 4 (284.32 mg, 7.49 mmol, 2 eq) at 0 °C, the mixture was stirred at 0 °C for 1 h. Upon completion, the reaction mixture was quenched by addition H2O (0.3 mL) and 15% NaOH (0.3 mL) and H2O (0.9 mL) at 20 °C, and then diluted with EtOAc (20 mL) and extracted with EtOAc (20 mL * 3). The combined organic layers were washed with brine (30 mL), dried over Na 2 SO 4 , filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (0-9% MeOH in EtOAc) to obtain benzyl 2-(3-hydroxypropyl)-3-oxo-piperazine-1-carboxylate (600 mg, 1.3 mmol, 36% yield, 65% purity) as an oil. MS (ESI) m/z 293.2 [M+H] + . Step 2: methyl 2-[4-benzyloxycarbonyl-3-(3-hydroxypropyl)-2-oxo-piperazin-1 -yl]imidazo[1,2- a]pyridine-6-carboxylate
To a solution of benzyl 2-(3-hydroxypropyl)-3-oxo-piperazine-1-carboxylate (600 mg, 2.05 mmol, 1 eq) in dioxane (15 mL) was added K 2 CO 3 (567.35 mg, 4.10 mmol, 2 eq), methyl 2- bromoimidazo[1,2-a]pyridine-6-carboxylate (575.87 mg, 2.26 mmol, 1.1 eq), N,N'- dimethylethane-1,2-diamine (144.74 mg, 1.64 mmol, 176.73 μL, 0.8 eq) and CuI (156.36 mg, 820.99 μmol, 0.4 eq) under N 2 atmosphere, the mixture was stirred at 100 °C for 16 h under N 2 atmosphere. Upon completion, the mixture was diluted with H2O (30 mL), extracted with ethyl acetate (20 mL * 3), the organic layer was washed with brine, dried over Na2SO4, filtered and concentrated under reduced pressure to give the residue. The residue was purified by column chromatography (0-50% EtOAc in PE) to obtained methyl 2-[4-benzyloxycarbonyl-3-(3- hydroxypropyl)-2-oxo-piperazin-1-yl]imidazo[1,2-a]pyridine-6 -carboxylate (400 mg, 772 μmol, 38% yield, 90% purity) as a solid. MS (ESI) m/z 467.2 [M+H] + . Step 3: benzyl 2-(3-hydroxypropyl)-4-[6-(methylcarbamoyl)imidazo[1,2-a]pyri din-2-yl]-3-oxo- piperazine-1-carboxylate A solution of methyl 2-[4-benzyloxycarbonyl-3-(3-hydroxypropyl)-2-oxo-piperazin-1 - yl]imidazo[1,2-a]pyridine-6-carboxylate (350 mg, 750.3 μmol, 1 eq) in MeNH 2 (8 mL, 30% purity in EtOH) was stirred at 80 °C for 16 h. Upon completion, the reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-TLC (9% MeOH in EtOAc) to obtained benzyl 2-(3-hydroxypropyl)-4-[6-(methylcarbamoyl)imidazo[1,2-a]pyri din-2- yl]-3-oxo-piperazine-1-carboxylate (210 mg, 406 μmol, 54% yield, 90% purity) as a solid. MS (ESI) m/z 466.2 [M+H] + . Step 4: benzyl 4-[6-(methylcarbamoyl)imidazo[1,2-a]pyridin-2-yl]-3-oxo-2-[3 -(p- tolylsulfonyloxy)propyl]piperazine-1-carboxylate To a solution of benzyl 2-(3-hydroxypropyl)-4-[6-(methylcarbamoyl)imidazo[1,2- a]pyridin-2-yl]-3-oxo-piperazine-1-carboxylate (210 mg, 451.1 μmol, 1 eq) and DMAP (5.51 mg, 45.1 μmol, 0.1 eq) in DCM (5 mL) was stirred at 0 °C, was added Et 3 N (68.5 mg, 676.7 μmol, 94.2 μL, 1.5 eq) and TosCl (129 mg, 676.7 μmol, 1.5 eq) slowly, the mixture was stirred at 20 °C for 8 h. Upon completion, the mixture was diluted with H2O (20 mL) and extracted with DCM (10 mL x3). The organic layer was washed with brine, dried over Na 2 SO 4 , filtered, and concentrated under reduced pressure to obtained benzyl 4-[6-(methylcarbamoyl)imidazo[1,2-a]pyridin-2-yl]-3- oxo-2-[3-(p-tolylsulfonyloxy)propyl]piperazine-1-carboxylate (310 mg, crude) as a solid. MS (ESI) m/z 620.2 [M+H] + . Step 5: benzyl 2-[3-(dimethylamino)propyl]-4-[6-(methylcarbamoyl)imidazo[1, 2-a]pyridin-2- yl]-3-oxo-piperazine-1-carboxylate A mixture of benzyl 4-[6-(methylcarbamoyl)imidazo[1,2-a]pyridin-2-yl]-3-oxo-2-[3 -(p- tolylsulfonyloxy)propyl]piperazine-1-carboxylate (310 mg, 500.3 μmol, 1 eq) in N- methylmethanamine (2 M in THF, 2.5 mL, 50% purity) was stirred at 50 °C for 16 h. Upon completion, the reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (Condition 7, Gradient d) to obtained benzyl 2-[3- (dimethylamino)propyl]-4-[6-(methylcarbamoyl)imidazo[1,2-a]p yridin-2-yl]-3-oxo-piperazine-1 carboxylate (230 mg, 444 μmol, 89% yield, 95% purity) as a solid. MS (ESI) m/z 493.2 [M+H] + . Step 6: 2-[3-[3-(dimethylamino)propyl]-2-oxo-piperazin-1-yl]-N-methy l-imidazo[1,2-a]pyridine- 6-carboxamide To a solution of benzyl 2-[3-(dimethylamino)propyl]-4-[6-(methylcarbamoyl)imidazo[1, 2- a]pyridin-2-yl]-3-oxo-piperazine-1-carboxylate (100 mg, 203 μmol, 1 eq) in EtOH (6 mL) was added Pd/C (108 mg, 101.5 μmol, 10% purity, 0.5 eq), the mixture was stirred at 20 °C for 0.5 h under H 2 (15 psi) atmosphere. Upon completion, the mixture was filtered and concentrated under reduced pressure to obtained 2-[3-[3-(dimethylamino)propyl]-2-oxo-piperazin-1-yl]-N-methy l- imidazo[1,2-a]pyridine-6-carboxamide (50 mg, crude) as a gum. MS (ESI) m/z 359.3 [M+H] + . Step 7: 2-[3-[3-(dimethylamino)propyl]-2-oxo-4-prop-2-enoyl-piperazi n-1-yl]-N-methyl- i To a solution of 2-[3-[3-(dimethylamino)propyl]-2-oxo-piperazin-1-yl]-N-methy l- imidazo[1,2-a]pyridine-6-carboxamide (60 mg, 167.4 μmol, 1 eq, 2 batches in parallel) in DCM (10 mL) was added TEA (50.8 mg, 502.2 μmol, 69.9 μL, 3 eq), then the mixture was cooled to 0 °C and was added prop-2-enoyl chloride (18.18 mg, 200.87 μmol, 16.32 μL, 1.2 eq) drop-wised at 0 °C, the mixture was stirred at 0 °C for 1 h. Upon completion, the mixture was quenched by water (0.1 mL) and was dried by blowing N 2 . The crude product was purified by prep-HPLC (Condition 4, Gradient b) to obtained 2-[3-[3-(dimethylamino)propyl]-2-oxo-4-prop-2-enoyl- piperazin-1-yl]-N-methyl-imidazo[1,2-a]pyridine-6-carboxamid e (Compound 175, 18 mg, 43.6 μmol, 13% yield) as a solid. MS (ESI) m/z 413.2 [M+H] + . 1 H NMR (400 MHz, MeOD) δ ppm 8.94 (s, 1H), 8.41 (s, 1H), 7.66 (dd, J = 1.6, 9.4 Hz, 1H), 7.52 (d, J = 9.4 Hz, 1H), 7.01 - 6.78 (m, 1H), 6.41 - 6.24 (m, 1H), 5.85 (dd, J = 1.8, 10.6 Hz, 1H), 5.24 - 5.12 (m, 1H), 4.44 - 4.22 (m, 2H), 4.19 - 3.75 (m, 2H), 2.94 (s, 3H), 2.54 - 2.38 (m, 2H), 2.33 - 2.25 (m, 6H), 2.17 - 1.97 (m, 2H), 1.83 - 1.57 (m, 2H). Example 44: Synthesis of Compound 176 Step 1 : 4-bromo-1-methyl-imidazole-2-carbaldehyde LDA (2 M, 10.25 mL, 1.1 eq) was added to a solution of 4-bromo-1-methyl-imidazole (3 g, 18.6 mmol, 1 eq) in THF (90 mL) at -10 °C, after 1 h, DMF (2.04 g, 28 mmol, 2.15 mL, 1.5 eq) was added, then the mixture was stirred for another 1 h at 0 °C. Upon completion, the reaction mixture was quenched by saturated aqueous citric acid solution (30 Ml), and then extracted with ethyl acetate (30 mL x2). The combined organic layers were washed with aqueous NaCl (30 mL * 3), dried over Na2SO4, filtered and concentrated under reduced pressure to 4-bromo-1-methyl- imidazole-2-carbaldehyde (2.7 g, crude) as a solid. Step 2 : methyl (E)-3-(4-bromo-1-methyl-imidazol-2-yl)prop-2-enoate To a solution of methyl 2-diethoxyphosphorylacetate (3.56 g, 16.93 mmol, 1 eq) in THF (60 mL) was added NaH (1.22 g, 30.60 mmol, 60 % purity, 1.81 eq) at 0 °C for 0.5 h, then a solution of 4-bromo-1-methyl-imidazole-2-carbaldehyde (3.2 g, 16.93 mmol, 1 eq) in THF (60 mL) was added and the mixture was stirred at 0 °C for 2 h. Upon completion, the reaction mixture was poured into H2O 90 mL, and then extracted with ethyl acetate (90 mL * 2). The combined organic layers were washed with aqueous NaCl (60 mL*3), dried over Na 2 SO 4 , filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (25-100% EtOAc in PE) to give methyl (E)-3-(4-bromo-1-methyl-imidazol-2- yl)prop-2-enoate (3.11 g, 12.7 mmol, 75% yield) as a solid. Step 3 : tert-butyl 4-[2-[(E)-3-methoxy-3-oxo-prop-1-enyl]-1-methyl-imidazol-4-y l]-3-oxo- piperazine-1-carboxylate A mixture of methyl (E)-3-(4-bromo-1-methyl-imidazol-2-yl)prop-2-enoate (3.11 g, 12.7 mmol, 1 eq), tert-butyl 3-oxopiperazine-1-carboxylate (2.54 g, 12.7 mmol, 1 eq), CuI (362.5 mg, 1.9 mmol, 0.15 eq), DMEDA (279.7 mg, 3.17 mmol, 341.5 μL, 0.25 eq) and K 2 CO 3 (3.51 g, 25.4 mmol, 2 eq) in dioxane (50 mL) was degassed and purged with N 2 (x3), and then the mixture was stirred at 100 °C for 12 h under N2 atmosphere. Upon completion, the reaction mixture was quenched by H2O 100 mL, and then extracted with EtOAc (80 mL*2). The combined organic layers were washed with aqueous NaCl (50 mL*3), dried over Na 2 SO 4 , filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (18-100% EtOAc in PE) to give tert-butyl 4-[2-[(E)-3-methoxy-3-oxo-prop-1-enyl]-1-methyl- imidazol-4-yl]-3-oxo-piperazine-1-carboxylate (1.7 g, 4.7 mmol, 37% yield) as a solid. Step 4 : tert-butyl 4-[2-(3-methoxy-3-oxo-propyl)-1-methyl-imidazol-4-yl]-3-oxo- piperazine-1- carboxylate To a solution of tert-butyl 4-[2-[(E)-3-methoxy-3-oxo-prop-1-enyl]-1-methyl-imidazol-4- yl]-3-oxo-piperazine-1-carboxylate (500 mg, 1.37 mmol, 1 eq) in EtOH (10 mL) was added Pd/C (500 mg, 10% purity) under N 2 atmosphere. The suspension was degassed and purged with H 2 (x3). The mixture was stirred under H2 (15 psi) at 25 °C for 1 h. Upon completion, the mixture was filtered and concentrated in vacuum to give tert-butyl 4-[2-(3-methoxy-3-oxo-propyl)-1- methyl-imidazol-4-yl]-3-oxo-piperazine-1-carboxylate (500 mg, crude) as an oil. Step5 : tert-butyl 4-[1-methyl-2-[3-(methylamino)-3-oxo-propyl]imidazol-4-yl]-3 -oxo- piperazine-1-carboxylate tert-butyl 4-[2-(3-methoxy-3-oxo-propyl)-1-methyl-imidazol-4-yl]-3-oxo- piperazine-1- carboxylate (500 mg, 1.4 mmol, 1 eq) was added to methanamine (12.4 g, 120 mmol, 30% purity, 88 eq) (methylamine ethanol solution), and then the temperature was raised to 50 °C for 16 h. Upon completion, the residue was evaporated to dryness to give tert-butyl 4-[1-methyl-2-[3- (methylamino)-3-oxo-propyl]imidazol-4-yl]-3-oxo-piperazine-1 -carboxylate (500 mg, crude) as an oil. Step6 : N-methyl-3-[1-methyl-4-(2-oxopiperazin-1-yl)imidazol-2-yl]pr opanamide To a solution of tert-butyl 4-[1-methyl-2-[3-(methylamino)-3-oxo-propyl]imidazol-4-yl]- 3-oxo-piperazine-1-carboxylate (500 mg, 1.37 mmol, 1 eq) in TFA (2 mL) and DCM (6 mL), the mixture was stirred for 1 h at 25 °C. Upon completion, the resulting solution was concentrated in vacuum to give N-methyl-3-[1-methyl-4-(2-oxopiperazin-1-yl) imidazol-2-yl] propanamide (400 mg, crude) as an oil. Step 7 : N-methyl-3-[1-methyl-4-(2-oxo-4-prop-2-enoyl-piperazin-1-yl) imidazol-2- yl]propanamide To a solution of N-methyl-3-[1-methyl-4-(2-oxopiperazin-1-yl)imidazol-2- yl]propanamide (400 mg, 1.5 mmol, 1 eq) in DCM (5 mL) was added TEA (610 mg, 6 mmol, 839 μL, 4 eq) and prop-2-enoyl chloride (164 mg, 1.8 mmol, 147.5 μL, 1.2 eq), then the mixture was stirred for 1 h at 0 °C. Upon completion, the resulting solution was quenched with H 2 O (0.1 mL), then was concentrated in vacuum (25 °C). The residue was purified by prep-HPLC (Condition 4, Gradient b) to give N-methyl-3-[1-methyl-4-(2-oxo-4-prop-2-enoyl-piperazin-1-yl) imidazol-2- yl]propanamide (Compound 176, 206.5 mg, 622 μmol, 41% yield, 96% purity) as a solid. MS (ESI) m/z 320.2 NMR (400 MHz, MeOD) δ = 7.24 (s, 1H), 6.82 (br dd, J = 10.6, 16.9 Hz, 1H), 6.29 (dd, J = 1.6, 16.8 Hz, 1H), 5.82 (dd, J = 1.8, 10.6 Hz, 1H), 4.48 - 4.35 (m, 2H), 4.04 - 3.92 (m, 4H), 3.63 (s, 3H), 2.95 (t, J = 7.4 Hz, 2H), 2.60 (s, 3H), 2.63 - 2.51 (m, 2H). Example 45: Synthesis of Compound 177 Step 1: 2-[(2-bromoimidazol-1-yl)methoxy]ethyl-trimethyl-silane To a solution of 2-bromo-1H-imidazole (10.5 g, 71.44 mmol, 1 eq) in THF (100 mL) was added dropwise NaH (3.43 g, 85.73 mmol, 60% purity, 1.2 eq) at 0 °C. After addition, the mixture was stirred at this temperature for 0.5 h and then was added dropwise SEMCl (15.48 g, 92.87 mmol, 16.44 mL, 1.3 eq). The mixture was stirred at 25 °C for 1.5 h. Upon completion, the reaction mixture was quenched by aqueous NH4Cl 200 mL, and then extracted with EtOAc (200 mL*2). The combined organic ayers were washed with aqueous NaCl (150 mL * 3), dried over Na 2 SO 4 , filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (18-50% EtOAc in PE) to give 2-[(2-bromoimidazol-1-yl)methoxy]ethyl- trimethyl-silane (12 g, 43.3 mmol, 61% yield) as an oil. MS (ESI) m/z 279.1 [M+H] + . Step 2: 2-bromo-3-(2-trimethylsilylethoxymethyl)imidazole-4-carbalde hyde To a solution of 2-[(2-bromoimidazol-1-yl)methoxy]ethyl-trimethyl-silane (12 g, 43.28 mmol, 1 eq) in THF (250 mL) was added LDA (2 M, 23.81 mL, 1.1 eq). The mixture was stirred at -65 °C for 1 h. Then was added DMF (6.33 g, 86.57 mmol, 6.66 mL, 2 eq). The mixture was stirred at -65 °C for 0.5 h. Upon completion, the reaction mixture was quenched by aqueous 1 M NH 4 Cl 100 mL and extracted with EtOAc (200 mL*2). The combined organic ayers were washed with aqueous NaCl (150 mL * 3). The combined organic layers were dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (9-50% EtOAc in PE) to give 2-bromo-3-(2-trimethylsilylethoxymethyl)- imidazole-4-carbaldehyde (6.5 g, 21.3 mmol, 49% yield) as an oil. MS (ESI) m/z 307.2 [M+H] + . Step 3: methyl (E)-3-[2-bromo-3-(2-trimethylsilylethoxymethyl)imidazol-4-yl ]prop-2-enoate To a solution of methyl 2-diethoxyphosphorylacetate (4.48 g, 21.29 mmol, 1 eq) in THF (60 mL) was added DBU (6.48 g, 42.59 mmol, 6.42 mL, 2 eq). The mixture was stirred at 0 °C for 0.5 h. Then was added 2-bromo-3-(2-trimethylsilylethoxymethyl)imidazole-4-carbalde hyde (6.5 g, 21.29 mmol, 1 eq). The mixture was stirred at 0 °C for 1 h. Upon completion, the reaction mixture was quenched by aqueous NH4Cl 100 mL, and then extracted with EtOAc (150 mL*2). The combined organic ayers were washed with aqueous NaCl (200 mL*3). The combined organic layers were dried over Na 2 SO 4 , filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (9-50% EtOAc in PE) to give methyl (E)-3- [2-bromo-3-(2-trimethylsilylethoxymethyl)imidazol-4-yl]prop- 2-enoate (5 g, 13.8 mmol, 65% yield) as an oil. MS (ESI) m/z 363.1 [M+H] + . Step 4: methyl (E)-3-(2-bromo-1H-imidazol-5-yl)prop-2-enoate To a solution of methyl (E)-3-[2-bromo-3-(2-trimethylsilylethoxymethyl)imidazol-4- yl]prop-2-enoate (5 g, 13.84 mmol, 1 eq) in THF (30 mL) was added TBAF (1 M, 37.36 mL, 2.7 eq). The mixture was stirred at 60 °C for 16 h. Upon completion, the reaction mixture was quenched by aqueous H 2 O 20 mL and then extracted with EtOAc (50 mL * 2). The combined organic ayers were washed with aqueous NaCl (40 mL * 3), dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The crude product was triturated with (9% EtOAc in PE, 10 mL) to give methyl (E)-3-(2-bromo-1H-imidazol-5-yl)prop-2-enoate (2 g, 6.9 mmol, 50% yield, 80% purity) as a solid. MS (ESI) m/z 233.0 [M+H] + . Step 5: methyl (E)-3-(2-bromo-3-methyl-imidazol-4-yl)prop-2-enoate To a solution of methyl (E)-3-(2-bromo-1H-imidazol-5-yl)prop-2-enoate (2 g, 8.66 mmol, 1 eq) in DMF (25 mL) was added dropwise NaOH (519 mg, 12.98 mmol, 1.5 eq) at 0 °C. After addition, the mixture was stirred at this temperature for 0.5 h and then was added dropwise CH3I (2.58 g, 18.18 mmol, 1.13 mL, 2.1 eq). The mixture was stirred at 50 °C for 1 h. Upon completion, the reaction mixture was quenched by aqueous H 2 O 5 mL and then extracted with EtOAc (10 mL * 2). The combined organic ayers were washed with aqueous NaCl (15 mL * 3), dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (column: Welch Xtimate C18180*70mm*10um; mobile phase: [H 2 O (0.01%TFA)- ACN]; gradient: 10%-40% B over 0.1 min) to give methyl (E)-3-(2-bromo-3-methyl-imidazol-4- yl)prop-2-enoate (700 mg, 2.86 mmol, 83% yield) as a solid. MS (ESI) m/z 247.1 [M+H] + . Step 6: tert-butyl 4-[5-[(E)-3-methoxy-3-oxo-prop-1-enyl]-1-methyl-imidazol-2-y l]-3-oxo- piperazine-1-carboxylate To a solution of methyl (E)-3-(2-bromo-3-methyl-imidazol-4-yl)prop-2-enoate (700 mg, 2.9 mmol, 1 eq) and tert-butyl 3-oxopiperazine-1-carboxylate (1.7 g, 8.6 mmol, 3 eq) in dioxane (50 mL) was added CuI (272 mg, 1.4 mmol, 0.5 eq) 1,10-phenanthroline (360.3 mg, 2 mmol, 0.7 eq) and K2CO3 (395 mg, 2.9 mmol, 1 eq). The mixture was stirred at 100 °C for 16 h. Upon completion, the reaction mixture was quenched by aqueous H 2 O 60 mL, and then extracted with EtOAc (100 mL * 2). The combined organic ayers were washed with aqueous NaCl (100mL * 3), dried over Na 2 SO 4 , filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (18-90% EtOAc in PE) to give tert-butyl 4-[5-[(E)-3- methoxy-3-oxo-prop-1-enyl]-1-methyl-imidazol-2-yl]-3-oxo-pip erazine-1-carboxylate (500 mg, 1.37 mmol, 48% yield) as a solid. MS (ESI) m/z 365.2 [M+H] + . Step 7: tert-butyl 4-[5-(3-methoxy-3-oxo-propyl)-1-methyl-imidazol-2-yl]-3-oxo- piperazine-1- carboxylate To a solution of tert-butyl 4-[5-[(E)-3-methoxy-3-oxo-prop-1-enyl]-1-methyl-imidazol-2- yl]-3-oxo-piperazine-1-carboxylate (500 mg, 1.37 mmol, 1 eq) in EtOH (10 mL) was added Pd/C (706.6 mg, 664 μmol, 10% purity). The suspension was degassed and purged with H 2 (x3). The mixture was stirred under H2 (15 Psi) at 25 °C for 1 h. Upon completion, the solution was filtered and concentrated to give tert-butyl 4-[5-(3-methoxy-3-oxo-propyl)-1-methyl-imidazol-2-yl]-3- oxo-piperazine-1-carboxylate (500 mg, 1.36 mmol, 99.5% yield) as a solid and used directly for the next step. MS (ESI) m/z 367.2 [M+H] + . Step 8: tert-butyl 4-[1-methyl-5-[3-(methylamino)-3-oxo-propyl]imidazol-2-yl]-3 -oxo- piperazine-1-carboxylate To a solution of tert-butyl 4-[5-(3-methoxy-3-oxo-propyl)-1-methyl-imidazol-2-yl]-3-oxo- piperazine-1-carboxylate (500 mg, 1.36 mmol, 1 eq) in MeNH2/EtOH (15 mL) was stirred for 4 h at 50 °C. Upon completion, the solution was concentrated to dryness to give tert-butyl 4-[1-methyl- 5-[3-(methylamino)-3-oxo-propyl]imidazol-2-yl]-3-oxo-piperaz ine-1-carboxylate (495 mg, 1.35 mmol, 99.3% yield) as an oil and used directly for the next step. MS (ESI) m/z 366.2 [M+H] + . Step 9: N-methyl-3-[3-methyl-2-(2-oxopiperazin-1-yl)imidazol-4-yl]pr opanamide To a solution of tert-butyl 4-[1-methyl-5-[3-(methylamino)-3-oxo-propyl]imidazol-2-yl]- 3-oxo-piperazine-1-carboxylate (300 mg, 821 μmol, 1 eq) in DCM (7 mL) and TFA (2 mL) was stirred for 2 h at 20 °C. Upon completion, the solution was concentrated to dryness to give N- methyl-3-[3-methyl-2-(2-oxopiperazin-1-yl)imidazol-4-yl]prop anamide (217 mg, 818 μmol, 99.6% yield) as an oil and used directly for the next step. MS (ESI) m/z 266.3 [M+H] + . Step 10: N-methyl-3-[3-methyl-2-(2-oxo-4-prop-2-enoyl-piperazin-1-yl) imidazol-4- yl]propanamide To a solution of N-methyl-3-[3-methyl-2-(2-oxopiperazin-1-yl)imidazol-4- yl]propanamide (210 mg, 791.5 μmol, 1 eq) in DCM (6 mL) was added TEA (240.3 mg, 2.4 mmol, 330.5 μL, 3 eq) and then prop-2-enoyl chloride (85.97 mg, 949.8 μmol, 77.2 μL, 1.2 eq) was added at 0 °C. The solution was stirred for 1 h at 0 °C. Upon completion, the solution was quenched with H2O (1 mL) and filtered to give crude. The crude was purified by prep-HPLC (Condition 1, Gradient c) to give N-methyl-3-[3-methyl-2-(2-oxo-4-prop-2-enoyl-piperazin-1-yl) imidazol-4- yl]propanamide (Compound 177, 55 mg, 172.2 μmol, 22% yield) as a solid. MS (ESI) m/z 320.1 [M+H] + . 1 H NMR (400 MHz, MeOD) δ = 6.89 - 6.71 (m, 2H), 6.31 (dd, J = 1.8, 16.8 Hz, 1H), 5.84 (dd, J = 1.7, 12.0 Hz, 1H), 4.55 - 4.41 (m, 2H), 4.06 (br d, J = 2.4 Hz, 2H), 3.75 (br d, J = 3.6 Hz, 2H), 3.42 (s, 3H), 2.97 (t, J = 7.6 Hz, 2H), 2.71 (s, 3H), 2.60 (t, J = 7.6 Hz, 2H). Example 46: Synthesis of Compound 178 Step 1: 2-chlorooxazole-4-carbaldehyde To a solution of ethyl 2-chlorooxazole-4-carboxylate (3 g, 17.09 mmol, 1 eq) in THF (30 mL), was added DIBAL-H (1 M, 68.35 mL, 4 eq), the mixture was stirred at -78 °C for 2 h. Upon completion, the reaction mixture was quenched by addition seignette salt 150 mL at -78 °C, and then diluted with EtOAc 200 mL and extracted with EtOAc (200 mL * 3). The combined organic layers were washed with NaCl aq. (200 mL * 2), dried over Na 2 SO 4 , filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (25-50% EtOAc in PE) to give 2-chlorooxazole-4-carbaldehyde (965 mg, 7.34 mmol, 43% yield, 100% purity) as a solid. Step 2: benzyl (E)-3-(2-chlorooxazol-4-yl)prop-2-enoate To a solution of benzyl 2-diethoxyphosphorylacetate (652.98 mg, 2.28 mmol, 1 eq) in THF (5 mL) was added NaH (165.13 mg, 4.13 mmol, 60% purity, 1.81 eq) at 0 °C for 0.5 h, then a solution of 2-chlorooxazole-4-carbaldehyde (300 mg, 2.28 mmol, 1 eq) in THF (5 mL) was added and the mixture was stirred at 25 °C for 1 h. Upon completion, the reaction mixture was poured into NH4Cl 70 mL, and then extracted with EtOAc (100 mL*2). The combined organic layers were washed with aqueous NaCl (100 mL*3), dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (25-100% EtOAc in PE) to give benzyl (E)-3-(2-chlorooxazol-4-yl)prop-2-enoate (600 mg, 2.07 mmol, 91% yield, 91% purity) as a solid. Step 3: tert-butyl 4-[4-[(E)-3-benzyloxy-3-oxo-prop-1-enyl]oxazol-2-yl]-3-oxo-p iperazine-1- carboxylate To solution of benzyl (E)-3-(2-chlorooxazol-4-yl)prop-2-enoate (600 mg, 2.28 mmol, 1 eq), tert-butyl 3-oxopiperazine-1-carboxylate (546.8 mg, 2.7 mmol, 1.2 eq) in dioxane (15 mL), was added Cs2CO3 (1.48 g, 4.55 mmol, 2 eq), tBuXPhos Pd G3 (180.8 mg, 227.6 μmol, 0.1 eq). The mixture was stirred at 90 °C for 1 h. Upon completion, the mixture was concentrated in vauum. The residue was purified by column chromatography (25-50% EtOAc in PE) to give tert-butyl 4- [4-[(E)-3-benzyloxy-3-oxo-prop-1-enyl]oxazol-2-yl]-3-oxo-pip erazine-1-carboxylate (698 mg, 1.52 mmol, 67% yield, 93% purity) as a solid. Step 4: 3-[2-(4-tert-butoxycarbonyl-2-oxo-piperazin-1-yl)oxazol-4-yl ]propanoic acid To a solution of tert-butyl 4-[4-[(E)-3-benzyloxy-3-oxo-prop-1-enyl]oxazol-2-yl]-3-oxo- piperazine-1-carboxylate (698 mg, 1.63 mmol, 1 eq) in EtOH (15 mL) was added Pd/C (608.72 mg, 572 μmol, 10% purity, 0.35 eq) under N2. The suspension was degassed under vacuum and purged with H 2 several times. The mixture was stirred under H 2 (15 psi) at 25°C for 30 min. Upon completion, the reaction mixture is filtered with diatomaceous earth and spin dried. The residue was purified by column chromatography (50-100% EtOAc in PE) to give 3-[2-(4-tert- butoxycarbonyl-2-oxo-piperazin-1-yl)oxazol-4-yl]propanoic acid (320 mg, 698 μmol, 43% yield, 74% purity) as an oil. Step 5: tert-butyl 4-[4-[3-(methylamino)-3-oxo-propyl]oxazol-2-yl]-3-oxo-pipera zine-1- carboxylate To a solution of 3-[2-(4-tert-butoxycarbonyl-2-oxo-piperazin-1-yl)oxazol-4-yl ]propanoic acid (310 mg, 913.5 μmol, 1 eq) in DMF (1 mL), was added HATU (416.8 mg, 1.1 mmol, 1.2 eq), DIPEA (354.2 mg, 2.74 mmol, 477.4 μL, 3 eq), the mixture was stirred at 25 °C for 30 min, methanamine; hydrochloride (95 mg, 913.5 μmol, 1 eq, HCl) was added, the mixture was stirred at 25 °C for 30 min. Upon completion, the reaction mixture was diluted with H2O 10 mL and extracted with EtOAc (30 mL*3). The combined organic layers were dried over sat.Na 2 SO 4 , filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (0-9% MeOH in EtOAc) to afford tert-butyl 4-[4-[3-(methylamino)-3- oxo-propyl]oxazol-2-yl]-3-oxo-piperazine-1-carboxylate (217 mg, 431 μmol, 47% yield, 70% purity) as an oil. Step 6: N-methyl-3-[2-(2-oxopiperazin-1-yl)oxazol-4-yl]propanamide To solution of tert-butyl 4-[4-[3-(methylamino)-3-oxo-propyl]oxazol-2-yl]-3-oxo- piperazine-1-carboxylate (217 mg, 615.8 μmol, 1 eq) in TFA (1.73 g, 15.2 mmol, 1.13 mL, 24.7 eq), DCM (3 mL), the mixture was stirred at 25 °C for 1 h. Upon completion, the mixture was concentrated in vacuum. To give N-methyl-3-[2-(2-oxopiperazin-1-yl)oxazol-4-yl]propanamide (150 mg, crude) as an oil. Step 7: N-methyl-3-[2-(2-oxo-4-prop-2-enoyl-piperazin-1-yl)oxazol-4- yl]propanamide To solution of N-methyl-3-[2-(2-oxopiperazin-1-yl)oxazol-4-yl]propanamide (120 mg, 475.7 μmol, 1 eq) in ACN (12 mL), was added K 2 CO 3 (197.2 mg, 1.43 mmol, 3 eq), then prop-2- enoyl chloride (30 mg, 333 μmol, 27 μL, 0.7 eq) was added, the mixture was stirred at 0 °C for 1 h. Upon completion, the mixture was concentrated in vacuum. The residue was purified by prep- HPLC (Condition 9, Gradient c) to give N-methyl-3-[2-(2-oxo-4-prop-2-enoyl-piperazin-1- yl)oxazol-4-yl]propanamide (Compound 178, 20 mg, 63.4 μmol, 13% yield, 97% purity) as a solid. MS (ESI) m/z= 307.1 [M+H] + . 1 H NMR (400 MHz, DMSO-d6) δ = 7.79 (br d, J = 4.0 Hz, 1H), 7.68 (s, 1H), 6.86 - 6.75 (m, 1H), 6.18 (br d, J = 16.8 Hz, 1H), 5.77 (br d, J = 9.0 Hz, 1H), 4.49 - 4.27 (m, 2H), 3.97 - 3.83 (m, 4H), 2.65 (t, J = 7.6 Hz, 2H), 2.56 (d, J = 4.6 Hz, 3H), 2.35 (t, J = 7.7 Hz, 2H). Example 47: Synthesis of Compound 181 Step 1: benzyl 4-hydroxy-3-nitro-benzoate To a solution of 4-hydroxy-3-nitro-benzoic acid (10 g, 54.6 mmol, 1 eq) in DMF (500 mL) was added NaHCO3 (6.88 g, 81.9 mmol, 3.2 mL, 1.5 eq) and bromomethylbenzene (14 g, 81.9 mmol, 9.73 mL, 1.5 eq). The mixture was stirred at 50 °C for 16 h. Upon completion, the reaction mixture was quenched by addition H 2 O 800 mL, and then extracted with EtOAc 2.4 L (800 mL * 3). The combined organic layers were washed with brine (1L x3), dried over Na2SO4, filtered, and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (20% EtOAc in PE) to give benzyl 4-hydroxy-3-nitro-benzoate (6.4 g, 23.4 mmol, 43% yield) as a solid. MS (ESI) m/z 272.2 [M-H] + . Step 2: benzyl 3-amino-4-hydroxy-benzoate A mixture of benzyl 4-hydroxy-3-nitro-benzoate (2 g, 7.32 mmol, 1 eq) and NH4Cl (3.9 g, 73.2 mmol, 10 eq) in EtOH (20 mL) and H2O (4 mL) was degassed and purged with N2 (x3), then added Fe (2 g, 36.6 mmol, 5 eq) at 60 °C. Then the mixture was stirred at 80 °C for 1 h under N 2 atmosphere. Upon completion, the reaction mixture was filtered and concentrated under reduced pressure to give a residue, then added H2O 30 mL, and then extracted with EtOAc (50 mL * 3). The combined organic layers were washed with brine 150 mL, dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (18% EtOAc in PE) to give benzyl 3-amino-4-hydroxy-benzoate (940 mg, 3.86 mmol, 53% yield) as a solid. MS (ESI) m/z 244.2 [M+H] + . Step 3: benzyl 1,3-benzoxazole-5-carboxylate
A solution of benzyl 3-amino-4-hydroxy-benzoate (850 mg, 3.5 mmol, 1 eq) in trimethoxymethane (5.81 g, 54.7 mmol, 6 mL, 15.66 eq) was stirred at 80 °C for 16 h. Upon completion, the solution was concentrated to dryness to give the residue. The residue was purified by column chromatography (10% EtOAc in PE) to give benzyl 1,3-benzoxazole-5-carboxylate (650 mg, 2.57 mmol, 74% yield) as a solid. MS (ESI) m/z 254.1 [M+H] + . Step 4: benzyl 2-bromo-1,3-benzoxazole-5-carboxylate To a solution of benzyl 1,3-benzoxazole-5-carboxylate (600 mg, 2.37 mmol, 1 eq) in THF (6 mL) was added LiHMDS (1 M, 2.84 mL, 1.2 eq) at -25 °C for 1 h under N 2 atmosphere. Then added NBS (632.5 mg, 3.6 mmol, 1.5 eq) at -25 °C. The mixture was stirred at 20 °C for 1 h. Upon completion, the reaction mixture was quenched by addition NH4CI 15 mL, and then extracted with EtOAc (20 mL * 3). The combined organic layers were washed with brine 60 mL, dried over Na 2 SO 4 , filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (100% PE) to give benzyl 2-bromo-1,3-benzoxazole-5- carboxylate (710 mg, 2.14 mmol, 90% yield) as a solid. MS (ESI) m/z 332.0 [M+H] + . Step 5: benzyl 2-(4-tert-butoxycarbonyl-2-oxo-piperazin-1-yl)-1,3-benzoxazo le-5-carboxylate To a solution of benzyl 2-bromo-1,3-benzoxazole-5-carboxylate (200 mg, 602.14 μmol, 1 eq) in ACN (14 mL) was added K 2 CO 3 (249.67 mg, 1.81 mmol, 3 eq) and tert-butyl 3- oxopiperazine-1-carboxylate (241.14 mg, 1.20 mmol, 2 eq). The mixture was stirred at 80 °C for 16 h. Upon completion, the solution was concentrated to dryness to give crude. The residue was purified by column chromatography (70-40% EtOAc in PE) to give benzyl 2-(4-tert- butoxycarbonyl-2-oxo-piperazin-1-yl)-1,3-benzoxazole-5-carbo xylate (450 mg, 209 μmol, 12% yield, 21% purity) as a solid. MS (ESI) m/z 452.2 [M+H] + . Step 6: 2-(4-tert-butoxycarbonyl-2-oxo-piperazin-1-yl)-1,3-benzoxazo le-5-carboxylic acid A mixture of Pd/C (420 mg, 394.7 μmol, 10% purity, 0.4 eq) in i-PrOH (60 mL) was added benzyl 2-(4-tert-butoxycarbonyl-2-oxo-piperazin-1-yl)-1,3-benzoxazo le-5-carboxylate (420 mg, 930.3 μmol, 1 eq) was degassed and purged with H2 (x3), and then the mixture was stirred at 20 °C for 0.5 h under H 2 atmosphere. Upon completion, the reaction was filtered and concentrated to dryness to give 2-(4-tert-butoxycarbonyl-2-oxo-piperazin-1-yl)-1,3-benzoxazo le-5-carboxylic acid (280 mg, crude) as a solid. MS (ESI) m/z 362.1 [M+H] + . Step 7: tert-butyl 4-[5-(methylcarbamoyl)-1,3-benzoxazol-2-yl]-3-oxo-piperazine -1-carboxylate To a solution of 2-(4-tert-butoxycarbonyl-2-oxo-piperazin-1-yl)-1,3-benzoxazo le-5- carboxylic acid (280 mg, 774.9 μmol, 1 eq) in DCM (3 mL) was added HATU (353.6 mg, 929.9 μmol, 1.2 eq) and DIEA (300.4 mg, 2.3 mmol, 404.9 μL, 3 eq) at 20 °C for 0.5 h. Then added methanamine;hydrochloride (52.3 mg, 774.9 μmol, 1 eq). The mixture was stirred at 20 °C for 1 h. Upon completion, the reaction mixture was filtered and concentrated under reduced pressure to give a residue, then added H2O 10 mL, and then extracted with DCM (10 mL*3). The combined organic layers were washed with brine 30 mL, dried over Na2SO4, filtered, and concentrated under reduced pressure to give tert-butyl 4-[5-(methylcarbamoyl)-1,3-benzoxazol-2-yl]-3-oxo- piperazine-1-carboxylate (350 mg, crude) as a solid. MS (ESI) m/z 375.2 [M+H] + . Step 8: N-methyl-2-(2-oxopiperazin-1-yl)-1,3-benzoxazole-5-carboxami de To a solution of tert-butyl 4-[5-(methylcarbamoyl)-1,3-benzoxazol-2-yl]-3-oxo- piperazine-1-carboxylate (400 mg, 1.07 mmol, 1 eq) in DCM (5 mL) was added TFA (1.23 g, 10.77 mmol, 1 mL, 10.1 eq) .The mixture was stirred at 20 °C for 1 h. Upon completion, the reaction mixture was concentrated under reduced pressure to give N-methyl-2-(2-oxopiperazin-1- yl)-1,3-benzoxazole-5-carboxamide (293 mg, crude) as an oil. MS (ESI) m/z 275.2 [M+H] + . Step 9: N-methyl-2-(2-oxo-4-prop-2-enoyl-piperazin-1-yl)-1,3-benzoxa zole-5-carboxamide To a solution of N-methyl-2-(2-oxopiperazin-1-yl)-1,3-benzoxazole-5-carboxami de (293 mg, 534 μmol, 1 eq) in DCM (3 mL) was added TEA (270.2 mg, 2.7 mmol, 371.7 μL, 5 eq) and prop-2-enoyl chloride (43.5 mg, 481 μmol, 39 μL, 0.9 eq) in DCM (0.3 mL) at 0 °C. The mixture was stirred at 0 °C for 1 h. Upon completion, the reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (Condition 9, Gradient d) to give N-methyl-2-(2-oxo-4-prop-2-enoyl-piperazin-1-yl)-1,3-benzoxa zole-5-carboxamide (Compound 181, 12.5 mg, 38.1 μmol, 7% yield, 94% purity) as a solid. MS (ESI) m/z 329.1 [M+H] + . 1 H NMR (400 MHz, CDCl 3 ): δ = 8.02 (d, J = 1.0 Hz, 1H), 7.78 (br d, J = 7.8 Hz, 1H), 7.58 (d, J = 8.4 Hz, 1H), 6.54 - 6.52 (m, 1H), 6.50 - 6.41 (m, 1H), 6.15 (br d, J = 3.2 Hz, 1H), 5.97 - 5.77 (m, 1H), 4.56 (br s, 2H), 4.28 (br s, 2H), 4.05 (s, 2H), 3.06 ppm (d, J = 4.8 Hz, 3H). An analogous method was followed to obtain the following compound. Example 48: Synthesis of Compound 183 Step 1: methyl 3-(3-iodopyrazol-1-yl)propanoate To a solution of 3-iodo-1H-pyrazole (3 g, 15.47 mmol, 1 eq) in ACN (45 mL) was added DBU (1.18 g, 7.73 mmol, 1.17 mL, 0.5 eq) at 0 °C, followed by methyl prop-2-enoate (2.66 g, 30.93 mmol, 2.79 mL, 2 eq) then was stirred at 25 °C for 2 h. Upon completion, the reaction mixture was quenched with 1 M HCl (30 mL) and extracted with EtOAc (30 mL*2). The combined organic layers were washed with water (20 mL) and brine (20 mL*2), dried over Na2SO4 and filtered. The filtrate was concentrated under reduced pressure to give methyl 3-(3-iodopyrazol-1- yl)propanoate (4 g, crude) as an oil. Step 2: tert-butyl 4-[1-(3-methoxy-3-oxo-propyl)pyrazol-3-yl]-3-oxo-piperazine- 1-carboxylate A mixture of methyl 3-(3-iodopyrazol-1-yl)propanoate (2 g, 7.14 mmol, 1 eq), tert-butyl 3- oxopiperazine-1-carboxylate (1.43 g, 7.14 mmol, 1 eq), iodocopper (204 mg, 1.07 mmol, 0.15 eq), N,N'-dimethylethane-1,2-diamine (157.4 mg, 1.79 mmol, 192.2 uL, 0.25 eq) and dipotassium;carbonate (1.97 g, 14.3 mmol, 2 eq) in dioxane (20 mL) was degassed and purged with N 2 (x3), and then the mixture was stirred at 100 °C for 16 h under N 2 atmosphere. Upon completion, the reaction mixture was diluted with aq. EDTA 30 mL and extracted with EtOAc (20 mL*2). The combined organic layers were dried over sat.Na 2 SO 4 , filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (10-30% EtOAc in PE) to give tert-butyl 4-[1-(3-methoxy-3-oxo-propyl)pyrazol-3-yl]-3-oxo-piperazine- 1- carboxylate (1.9 g, 5.4 mmol, 76% yield) as an oil. Step 3: tert-butyl 4-[1-[3-(methylamino)-3-oxo-propyl]pyrazol-3-yl]-3-oxo-piper azine-1- carboxylate tert-butyl 4-[1-(3-methoxy-3-oxo-propyl)pyrazol-3-yl]-3-oxo-piperazine- 1-carboxylate (1 g, 2.84 mmol, 1 eq) was added to methanamine (25.860 g, 249.80 mmol, 30% purity, 88.03 eq) (methylamine ethanol solution), and then the temperature was raised to 50 °C for 16 h. Upon completion, the residue was evaporated to dryness to give tert-butyl 4-[1-[3-(methylamino)-3-oxo- propyl]pyrazol-3-yl]-3-oxo-piperazine-1-carboxylate (0.9 g, crude) as an oil. Step 4: N-methyl-3-[3-(2-oxopiperazin-1-yl)pyrazol-1-yl]propanamide To a solution of tert-butyl 4-[1-[3-(methylamino)-3-oxo-propyl]pyrazol-3-yl]-3-oxo- piperazine-1-carboxylate (300 mg, 854 μmol, 1 eq) in DCM (5 mL) was added TFA (2.30 g, 20.19 mmol, 1.5 mL, 23.65 eq). The mixture was stirred at 25 °C for 1 h. Upon completion, the reaction was concentrated in vacuum (30 °C), then was added DCM (2 mL), and concentrated, and repeat to give N-methyl-3-[3-(2-oxopiperazin-1-yl)pyrazol-1-yl]propanamide (220 mg, crude) as an oil. Step 5: N-methyl-3-[3-(2-oxo-4-prop-2-enoyl-piperazin-1-yl)pyrazol-1 -yl]propanamide To a solution of N-methyl-3-[3-(2-oxopiperazin-1-yl)pyrazol-1-yl]propanamide (210 mg, 835.7 μmol, 1 eq) in DCM (5 mL) was added TEA (422.8 mg, 4.2 mmol, 581.6 μL, 5 eq) and prop- 2-enoyl chloride (83.2 mg, 919.3 μmol, 74.7 μL, 1.1 eq). The mixture was stirred at 0 °C for 1 h. Upon completion, the reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (Condition 6, Gradient d) to give N-methyl-3-[3-(2-oxo- 4-prop-2-enoyl-piperazin-1-yl)pyrazol-1-yl]propanamide (Compound 183, 130 mg, 426 μmol, 51% yield, 100% purity) as a solid. MS (ESI) m/z 306.2 [M+H] + . 1 H NMR (400 MHz, DMSO-d6) δ = 7.85 (br d, J = 3.9 Hz, 1H), 7.60 (d, J = 2.3 Hz, 1H), 6.82 (dt, J = 10.6, 16.3 Hz, 1H), 6.59 (d, J = 2.3 Hz, 1H), 6.18 (br d, J = 16.6 Hz, 1H), 5.80 - 5.67 (m, 1H), 4.50 - 4.19 (m, 4H), 4.02 - 3.75 (m, 4H), 2.57 (t, J = 7.0 Hz, 2H), 2.54 - 2.53 (m, 3H). An analogous method was followed to obtain the following compound. Example 49: Synthesis of Compound 185 Step 1: 1-benzyl-3,5-dibromo-1H-pyrazole To a solution of 3,5-dibromo-1H-pyrazole (5 g, 22.1 mmol, 1 eq) in DMF (30 mL) was added NaH (2.2 g, 55.3 mmol, 60% purity, 2.5 eq) and BnBr (4.54 g, 26.6 mmol, 3.16 mL, 1.2 eq) at 0 °C. The mixture was then allowed to 25 °C and stirred for 12 h. Upon completion, the reaction mixture was quenched by addition aq.NH4Cl at 0 °C, and then diluted with water 10 mL and extracted with EtOAc (30 mL x3). The combined organic layers were washed with brine (10 mL), dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, Petroleum ether/Ethyl acetate=1/0 to 200/1) to give 1-benzyl-3,5-dibromo-pyrazole (1.7 g, 5 mmol, 23% yield, 94% purity) was obtained as an oil. MS (ESI) m/z 315.0 [M+H] + . Step 2: 2-benzyl-5-bromo-pyrazole-3-carbaldehyde To a solution of 1-benzyl-3,5-dibromo-pyrazole (1.5 g, 4.75 mmol, 1 eq) in THF (22.5 mL) was added chloro(isopropyl)magnesium (2 M, 2.85 mL, 1.2 eq) at -78 ℃ and stirred for 30 min. And then DMF (2.43 g, 33.23 mmol, 2.56 mL, 7 eq) was added. The mixture was stirred at 25 ℃ for 4.5 h. Upon completion, the reaction mixture was quenched by addition aq.NH4Cl at 0 ℃, and then diluted with water 30 mL and extracted with EtOAc (30 mL*3). The combined organic layers were washed with brine (30 mL), dried over Na 2 SO 4 , filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO 2 , Petroleum ether/Ethyl acetate = 1/0 to 200/1) to give 2-benzyl-5-bromo-pyrazole-3-carbaldehyde (1.03 g, 3.4 mmol, 72% yield, 88% purity) as a solid. MS (ESI) m/z 265.1 [M+H] + . Step 3: methyl (E)-3-(2-benzyl-5-bromo-pyrazol-3-yl)prop-2-enoate To a solution of methyl 2-diethoxyphosphorylacetate (575 mg, 2.73 mmol, 1 eq) in THF (32 mL) was added DBU (833 mg, 5.47 mmol, 824 μL, 2 eq) at 25℃ for 30 min. And then 2- benzyl-5-bromo-pyrazole-3-carbaldehyde (870 mg, 3.3 mmol, 1.2 eq) in THF (2 mL) was added. The mixture was stirred at 25 ℃ for 15.5 h. Upon completion, the reaction mixture was diluted with water 20 mL and extracted with ethyl acetate (30 mL*3). The combined organic layers were washed with brine (20 mL), dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO 2 , Petroleum ether/EtOAc = 200/1 to 30/1) to give methyl (E)-3-(2-benzyl-5-bromo-pyrazol-3-yl)prop-2-enoate (820 mg, 2.5 mmol, 92% yield, 99% purity) as a solid. MS (ESI) m/z 321.1 [M+H] + . Step 4: tert-butyl 4-[1-benzyl-5-[(E)-3-methoxy-3-oxo-prop-1-enyl]pyrazol-3-yl] -3-oxo- piperazine-1-carboxylate A mixture of methyl (E)-3-(2-benzyl-5-bromo-pyrazol-3-yl)prop-2-enoate (820 mg, 2.6 mmol, 1 eq), tert-butyl 3-oxopiperazine-1-carboxylate (613.5 mg, 3.06 mmol, 1.2 eq), K 2 CO 3 (705.7 mg, 5.1 mmol, 2 eq), DMEDA (180 mg, 2.04 mmol, 219.8 μL, 0.8 eq) and CuI (194.5 mg, 1 mmol, 0.4 eq) in dioxane (41 mL) was degassed and purged with N2 (x3), and then the mixture was stirred at 100 ℃ for 16 h under N 2 atmosphere. Upon completion, the reaction mixture was diluted with water 30 mL and extracted with EtOAc (30 mL x3). The combined organic layers were washed with brine (30 mL), dried over Na2SO4, filtered, and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO 2 , Petroleum ether/Ethyl acetate = 60/1 to 3/1) to give tert-butyl 4-[1-benzyl-5-[(E)-3-methoxy-3-oxo-prop-1- enyl]pyrazol-3-yl]-3-oxo-piperazine-1-carboxylate (570 mg, 1.3 mmol, 51% yield, 100% purity) as an oil. MS (ESI) m/z 441.2 [M+H] + . Step 5: tert-butyl 4-[5-(3-methoxy-3-oxo-propyl)-1H-pyrazol-3-yl]-3-oxo-piperaz ine-1- carboxylate A solution of tert-butyl 4-[1-benzyl-5-[(E)-3-methoxy-3-oxo-prop-1-enyl]pyrazol-3-yl] -3- oxo-piperazine-1-carboxylate (450 mg, 1.02 mmol, 1 eq) in AcOH (10 mL) was added Pd/C (450 mg, 10% purity), then the mixture was stirred at 40 ℃ for 16 h under H2 (2.06 mg, 1.02 mmol, 1 eq) (15 Psi). Upon completion, the mixture was filted with MeOH and concentrated under reduced pressure to give the crude. The crude was diluted with NaHCO 3 20 mL and extracted with EtOAc (30 mL * 3). The combined organic layers were washed with brine (20 mL), dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO 2 , Petroleum ether/Ethyl acetate = 15/1 to 3/1) to give tert-butyl 4- [5-(3-methoxy-3-oxo-propyl)-1H-pyrazol-3-yl]-3-oxo-piperazin e-1-carboxylate (285 mg, 785 μmol, 77% yield, 97% purity) as a solid. MS (ESI) m/z 353.2 [M+H] + . Step 6: tert-butyl 4-[5-[3-(methylamino)-3-oxo-propyl]-1H-pyrazol-3-yl]-3-oxo-p iperazine-1- carboxylate A solution tert-butyl 4-[5-(3-methoxy-3-oxo-propyl)-1H-pyrazol-3-yl]-3-oxo-piperaz ine- 1-carboxylate (285 mg, 808.8 μmol, 1 eq) in MeNH 2 (7.37 g, 71.17 mmol, 30% purity, 88 eq) (ethanol solution) was stirred at 50 ℃ for 2 h. Upon completion, the residue was evaporated to dryness. The crude was used directly. To give crude product tert-butyl 4-[5-[3-(methylamino)-3- oxo-propyl]-1H-pyrazol-3-yl]-3-oxo-piperazine-1-carboxylate (280 mg, crude) as a solid. MS (ESI) m/z 352.2 [M+H] + . Step 7: N-methyl-3-[3-(2-oxopiperazin-1-yl)-1H-pyrazol-5-yl]propanam ide To a solution of tert-butyl 4-[5-[3-(methylamino)-3-oxo-propyl]-1H-pyrazol-3-yl]-3-oxo- piperazine-1-carboxylate (280 mg, 796.8 μmol, 1 eq) in DCM (10 mL) was added TFA (4.6 g, 40.4 mmol, 3 mL, 50.7 eq). The mixture was stirred at 25 ℃ for 0.5 h. Upon completion, the residue was evaporated to dryness. The crude product N-methyl-3-[3-(2-oxopiperazin-1-yl)-1H- pyrazol-5-yl]propanamide (290 mg, crude, TFA) was obtained as an oil and used the next step without further purification. MS (ESI) m/z 252.3 [M+H] + . Step 8: N-methyl-3-[3-(2-oxo-4-prop-2-enoyl-piperazin-1-yl)-1H-pyraz ol-5-yl]propanamide To a solution of N-methyl-3-[3-(2-oxopiperazin-1-yl)-1H-pyrazol-5-yl]propanam ide (250 mg, 995 μmol, 1 eq) in DCM (12 mL) was added TEA (403 mg, 3.98 mmol, 554 μL, 4 eq) andprop- 2-enoyl chloride (45 mg, 497.4 μmol, 40.4 μL, 0.5 eq). The mixture was stirred at 0 ℃ for 1 h. Upon completion, the reaction mixture was diluted with water 20 mL and extracted with EtOAc (40 mL x3). The combined organic layers were washed with brine (20 mL), dried over Na2SO4, filtered, and concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (Condition 7, Gradient b) to give N-methyl-3-[3-(2-oxo-4-prop-2-enoyl-piperazin-1- yl)-1H-pyrazol-5-yl]propanamide (Compound 185, 16 mg, 51.4 μmol, 5% yield, 98% purity) was obtained as a solid. MS (ESI) m/z 306.2 [M+H] + . 1 H NMR (400 MHz, MeOD) δ = 6.86 - 6.68 (m, 1H), 6.50 (s, 1H), 6.29 (dd, J = 1.6, 16.8 Hz, 1H), 5.82 (dd, J = 1.8, 10.6 Hz, 1H), 4.49 - 4.36 (m, 2H), 4.04 - 3.93 (m, 4H), 2.93 (t, J = 7.6 Hz, 2H), 2.70 (s, 3H), 2.51 (t, J = 7.5 Hz, 2H). Example 50: Synthesis of Compound 186 Step 1: methyl 2-[allyl(tert-butoxycarbonyl)amino]acetate To a solution of methyl 2-(tert-butoxycarbonylamino)acetate (3.7 g, 19.6 mmol, 2.9 mL, 1 eq) in DMF (37 mL) was cooled to 0 °C then NaH (938.6 mg, 23.5 mmol, 60% purity, 1.2 eq) was added and stirred at 0 °C for 0.5 h. Then 3-bromoprop-1-ene (2.84 g, 23.5 mmol, 1.2 eq) was added and the mixture was stirred at 0 °C for 1.5 h. Upon completion, the reaction mixture was diluted with water (150 mL) and extracted with EtOAc (100 mL x3). The combined organic layers were washed with sat.aq. NaCl (60 mL x3), dried over Na 2 SO 4 , filtered, and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (0-8% EtOAc in PE) to give methyl 2-[allyl(tert-butoxycarbonyl)amino]acetate (3.2 g, 14 mmol, 71% yield) as an oil. Step 2: methyl 2-[tert-butoxycarbonyl(2-oxoethyl)amino]acetate To a solution of methyl 2-[allyl(tert-butoxycarbonyl)amino]acetate (3 g, 13.08 mmol, 1 eq) in MeOH (30 mL), cooled to -60 °C, passed through O 3 (13.08 mmol, 1 eq) under 0.1 MPa until the solution turn blue, and then pass into the N2 until the reaction solution becomes no color. The mixture was stirred at 20 °C for 16 h. Upon completion, the reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (0- 20% EtOAc in PE) to give methyl 2-[tert-butoxycarbonyl(2-oxoethyl)amino]acetate (2.1 g, 5.5 mmol, 42% yield, 60% purity) as an oil. Step 3: amino 2,4,6-trimethylbenzenesulfonate To a solution of (tert-butoxycarbonylamino) 2,4,6-trimethylbenzenesulfonate (60 g, 190.2 mmol, 1 eq) in TFA (460.5 g, 4.04 mol, 300 mL, 21.23 eq) was stirred at 0 °C for 1 h. Upon completion, the reaction mixture was quenched by ice and water (400 mL) and maintained inter- temperature below 5 °C. The mixture was stirred for 15 min to form a slurry. The slurry was filtered and the filter cake was washed with water (10 mL x3) and air dried to afford amino 2,4,6- trimethylbenzenesulfonate (46 g, crude) as a solid. Step 4: 2-(4-bromo-2-pyridyl)acetonitrile To a solution of 4-bromo-2-fluoro-pyridine (15 g, 85.2 mmol, 1 eq) and acetonitrile (7 g, 170.5 mmol, 8.97 mL, 2 eq) in THF (200 mL) was cooled to -60 °C, then added LiHMDS (1 M, 29.8 mL, 0.35 eq) dropwise for 1 h. The mixture was stirred at 20 °C for 1 h. Upon completion, the reaction mixture was saturated ammonium chloride aqueous solution (500 mL) was added thereto, then the mixture was extracted with EtOAc (200 mL x3), the combined organic phase was dried over sodium sulfate, filtered and concentrated, and the residue was purified by column chromatography (10-25% EtOAc in PE) to give 2-(4-bromo-2-pyridyl)acetonitrile (17 g, 69 mmol, 81% yield, 80% purity) as an oil. MS (ESI) m/z 199.2 [M+H] + . Step 5: 2-(1-amino-4-bromo-pyridin-1-ium-2-yl)acetonitrile To a solution of 2-(4-bromo-2-pyridyl)acetonitrile (23 g, 116.73 mmol, 1 eq) in DCM (100 mL) was cooled to 0 °C, then amino 2,4,6-trimethylbenzenesulfonate (46 g, 213.7 mmol, 1.83 eq) in DCM (150 mL) was added dropwise and stirred at 20 °C for 2 h. Upon completion, the reaction mixture was filtered and the filter cake was washed with DCM (15 ml * 2) and dried to give 2-(1- amino-4-bromo-pyridin-1-ium-2-yl)acetonitrile (10.5 g, crude) as a solid. Step 6: 5-bromopyrazolo[1,5-a]pyridin-2-amine To a solution of 2-(1-amino-4-bromo-pyridin-1-ium-2-yl)acetonitrile (10.5 g, 49.3 mmol, 1 eq) in MeOH (200 mL) was added K2CO3 (27.3 g, 197.1 mmol, 4 eq). The mixture was stirred at 20 °C for 16 h. Upon completion, the reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (2-50% EtOAc in PE) to give 5-bromopyrazolo[1,5-a]pyridin-2-amine (7 g, 31.4 mmol, 64% yield, 95% purity) as a solid. Step 7: methyl 2-aminopyrazolo[1,5-a]pyridine-5-carboxylate A mixture of 5-bromopyrazolo[1,5-a]pyridin-2-amine (3.5 g, 16.5 mmol, 1 eq), carbon monoxide;molybdenum (871.5 mg, 3.3 mmol, 444.7 μL, 0.2 eq) and TEA (5.01 g, 49.5 mmol, 6.9 mL, 3 eq) in DMF (35 mL) and MeOH (35 mL) was degassed and purged with N2 (x3), and then cyclopentyl(diphenyl)phosphane;dichloropalladium;iron (1.21 g, 1.65 mmol, 0.1 eq) was added under N 2 atmosphere. The mixture was stirred at 100 °C for 16 h under N 2 atmosphere. Upon completion, the reaction mixture was quenched by addition water 50 mL at 25 °C, and then diluted with ethyl acetate 5 mL and extracted with ethyl acetate (20 mL x3). The combined organic layers were washed with sat. aq. NaCl (6 mL x3), dried over Na 2 SO 4 , filtered, and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (2-50% EtOAc in PE) to give methyl 2-aminopyrazolo[1,5-a]pyridine-5-carboxylate (1.6 g, 8.4 mmol, 51% yield) as a solid. MS (ESI) m/z 192.4 [M+H] + . Step 8: 2-amino-N-methyl-pyrazolo[1,5-a]pyridine-5-carboxamide To a solution of methyl 2-aminopyrazolo[1,5-a]pyridine-5-carboxylate (0.3 g, 1.6 mmol, 1 eq) in EtOH (7 mL) was added MeNH 2 (162.5 mg, 1.6 mmol, 30% purity, 1 eq). The mixture was stirred at 80 °C for 16 h. Upon completion, the reaction mixture was concentrated under reduced pressure to give 2-amino-N-methyl-pyrazolo[1,5-a]pyridine-5-carboxamide (1.5 g, crude) as a solid. Step 9: methyl 2-[tert-butoxycarbonyl-[2-[[5-(methylcarbamoyl)pyrazolo[1,5- a]pyridin-2- yl]amino]ethyl]amino]acetate To a mixture of methyl 2-[tert-butoxycarbonyl(2-oxoethyl)amino]acetate (182.4 mg, 788.6 μmol, 1.5 eq) and 2-amino-N-methyl-pyrazolo[1,5-a]pyridine-5-carboxamide (0.1 g, 525.8 μmol, 1 eq) in MeOH (3 mL) under Nitrogen, a drop of AcOH (44.2 mg, 736.1 μmol, 42.14 μL, 1.4 eq) was added and then was stirred at 0 °C for 0.5 h. NaBH3CN (66.1 mg, 1.05 mmol, 2 eq) was then added and the resulting reaction mixture was stirred at 20 °C for 15.5 h under N 2 . Upon completion, the reaction mixture was quenched by addition water 0.2 mL, and then concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (column: Phenomenex luna C18 (250*70 mm, 15 um); mobile phase: [H 2 O (0.2% FA) - ACN]; gradient: 30%-60% B in 20 min) to give methyl 2-[tert-butoxycarbonyl-[2-[[5-(methylcarbamoyl)pyrazolo[1,5- a]pyridin-2- yl]amino]ethyl]amino]acetate (0.62 g, 1.45 mmol, 23% yield, 95% purity) as a solid. MS (ESI) m/z 406.4 [M+H] + . Step 10: tert-butyl 4-[5-(methylcarbamoyl)pyrazolo[1,5-a]pyridin-2-yl]-3-oxo-pip erazine-1- carboxylate To a solution of methyl 2-[tert-butoxycarbonyl-[2-[[5-(methylcarbamoyl)pyrazolo[1,5- a]pyridin-2-yl]amino]ethyl]amino]acetate (0.2 g, 493.3 μmol, 1 eq) in MeOH (3 mL) was added K2CO3 (204.5 mg, 1.5 mmol, 3 eq). The mixture was stirred at 20 °C for 16 h. Upon completion, the reaction mixture was quenched by addition of water (1 mL). The combined organic layers were concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (Condition 9, Gradient e) to give tert-butyl 4-[5-(methylcarbamoyl)pyrazolo[1,5-a]pyridin-2-yl]- 3-oxo-piperazine-1-carboxylate (40 mg, 107 μmol, 22% yield) as a solid. MS (ESI) m/z 374.3 [M+H] + . Step 11: N-methyl-2-(2-oxopiperazin-1-yl)pyrazolo[1,5-a]pyridine-5-ca rboxamide To a solution of tert-butyl 4-[5-(methylcarbamoyl)pyrazolo[1,5-a]pyridin-2-yl]-3-oxo- piperazine-1-carboxylate (30 mg, 80.34 μmol, 1 eq) in TFA (460.50 mg, 4.04 mmol, 0.3 mL, 50.27 eq) and DCM (1 mL) was stirred at 20 °C for 1 h. Upon completion, the reaction mixture was concentrated under reduced pressure to give N-methyl-2-(2-oxopiperazin-1-yl)pyrazolo[1,5- a]pyridine-5-carboxamide (20 mg, crude) as an oil. MS (ESI) m/z 274.3 [M+H] + . Step 12: N-methyl-2-(2-oxo-4-prop-2-enoyl-piperazin-1-yl)pyrazolo[1,5 -a]pyridine-5- carboxamide A solution of N-methyl-2-(2-oxopiperazin-1-yl)pyrazolo[1,5-a]pyridine-5-ca rboxamide (20 mg, 73.18 μmol, 1 eq) in ACN (0.8 mL) and H 2 O (0.2 mL) was added K 2 CO 3 (30.34 mg, 219.55 μmol, 3 eq) and prop-2-enoyl chloride (4.97 mg, 54.89 μmol, 4.46 μL, 0.75 eq). The mixture was stirred at 0 °C for 1 h. Upon completion, the reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (Condition 9, Gradient c) to give N-methyl-2-(2-oxo-4-prop-2-enoyl-piperazin-1-yl)pyrazolo[1,5 -a]pyridine-5- carboxamide (Compound 186, 18 mg, 53.3 μmol, 73% yield, 97% purity) as a solid. MS (ESI) m/z 328.1 [M+H] + . 1 H NMR (400 MHz, DMSO-d6) δ = 8.76 - 8.51 (m, 2H), 8.12 (s, 1H), 7.31 - 7.10 (m, 2H), 6.97 - 6.69 (m, 1H), 6.20 (d, J = 17.2 Hz, 1H), 5.89 - 5.66 (m, 1H), 4.52 (s, 1H), 4.35 (s, 1H), 4.17 - 4.07 (m, 2H), 4.01 (s, 1H), 3.91 (d, J = 4.0 Hz, 1H), 2.80 (d, J = 4.4 Hz, 3H). Example 51: Synthesis of Compound 187 Step 1: tert-butyl 3-oxo-4-prop-2-ynyl-piperazine-1-carboxylate A solution of tert-butyl 3-oxopiperazine-1-carboxylate (5 g, 24.97 mmol, 1 eq) in DMF (60 mL) was added NaH (1.50 g, 37.46 mmol, 60% purity, 1.5 eq) under N 2 , stirring at 25 °C for 1 h. And then, 3-bromoprop-1-yne (4.08 g, 27.47 mmol, 2.96 mL, 1.1 eq) was added. The reaction mixture was stirred at 25 °C for 0.5 h. Upon completion, the reaction mixture was quenched by addition H 2 O (60 mL) at 0 °C, and then extracted with EtOAc (80 mL * 3). The combined organic layers were washed with brine (100 mL * 3), dried over Na 2 SO 4 , filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (5-19% EtOAc in PE) to give tert-butyl 3-oxo-4-prop-2-ynyl-piperazine-1-carboxylate (3.1 g, 13 mmol, 52% yield) as a solid. MS (ESI) m/z 239.2 [M+H] + . Step 2: tert-butyl 4-[[1-(2-methoxy-2-oxo-ethyl)triazol-4-yl]methyl]-3-oxo-pipe razine-1- carboxylate A solution of tert-butyl 3-oxo-4-prop-2-ynyl-piperazine-1-carboxylate (500 mg, 2.10 mmol, 1 eq) and methyl 2-azidoacetate (241.5 mg, 2.1 mmol, 1 eq) in dioxane (8 mL) was added CuI (119.9 mg, 629.5 μmol, 0.3 eq). The reaction mixture was stirred at 80 °C for 2 h. Upon completion, the reaction mixture was poured into H2O (10 mL) at 20 °C, and then extracted with EtOAc (20 mL x3). The combined organic layers were washed with brine (40 mL), dried over Na 2 SO 4 , filtered, and concentrated under reduced pressure to give a residue. The residue was purified by column (5-20% EtOAc in PE) to give tert-butyl 4-[[1-(2-methoxy-2-oxo-ethyl)triazol- 4-yl]methyl]-3-oxo-piperazine-1-carboxylate (520 mg, 1.5 mmol, 70% yield) as a solid. MS (ESI) m/z 354.2 [M+H] + . Step 3: tert-butyl 4-[[1-[2-(methylamino)-2-oxo-ethyl]triazol-4-yl]methyl]-3-ox o-piperazine-1- carboxylate tert-butyl 4-[[1-(2-methoxy-2-oxo-ethyl)triazol-4-yl]methyl]-3-oxo-pipe razine-1- carboxylate (300 mg, 848.96 μmol, 1 eq) was added to MeNH 2 (4 g, 38.64 mmol, 4 mL, 30% purity, 45.5 eq) (methylamine ethanol solution), and then the mixture was stirred at 50 °C for 16 hours. Upon completion, the reaction mixture was concentrated under reduced pressure to give tert-butyl 4-[[1-[2-(methylamino)-2-oxo-ethyl]triazol-4-yl]methyl]-3-ox o-piperazine-1- carboxylate (300 mg, crude) as an oil. MS (ESI) m/z 353.2 [M+H] + . Step 4: N-methyl-2-[4-[(2-oxopiperazin-1-yl)methyl]triazol-1-yl]acet amide A solution of tert-butyl 4-[[1-[2-(methylamino)-2-oxo-ethyl]triazol-4-yl]methyl]-3-ox o- piperazine-1-carboxylate (300 mg, 851.3 μmol, 1 eq) in TFA (1 mL) and DCM (3 mL) was stirred at 25 °C for 0.5 h. Upon completion, the reaction mixture was concentrated under reduced pressure to give N-methyl-2-[4-[(2-oxopiperazin-1-yl)methyl]triazol-1-yl]acet amide (220 mg, crude) as an oil. MS (ESI) m/z 253.2 [M+H] + . Step 5: N-methyl-2-[4-[(2-oxo-4-prop-2-enoyl-piperazin-1-yl)methyl]t riazol-1-yl]acetamide To a solution of N-methyl-2-[4-[(2-oxopiperazin-1-yl)methyl]triazol-1-yl]acet amide (220 mg, 872.1 μmol, 1 eq) in DCM (3 mL) was added TEA (264.73 mg, 2.6 mmol, 364.1 μL, 3 eq) and then prop-2-enoyl chloride (78.9 mg, 872.1 μmol, 70.9 μL, 1 eq) in DCM (1 mL) was added at 0 °C. The solution was stirred for 0.5 h at 0 °C. Upon completion, the reaction mixture was quenched by addition H 2 O (0.2 mL) at 0 °C, and then concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (Condition 4, Gradient b) to give N-methyl-2- [4-[(2-oxo-4-prop-2-enoyl-piperazin-1-yl)methyl]triazol-1-yl ]acetamide (Compound 187, 44 mg, 133 μmol, 34% yield, 93% purity) as a solid. MS (ESI) m/z 307.2 [M+H] + . 1 H NMR (400 MHz, DMSO-d6) δ = 8.20 (d, J = 3.3 Hz, 1H), 7.97 (s, 1H), 6.87 - 6.70 (m, 1H), 6.15 (dd, J = 1.8, 16.6 Hz, 1H), 5.72 (br d, J = 10.5 Hz, 1H), 5.03 (s, 2H), 4.59 (s, 2H), 4.26 (s, 1H), 4.11 (s, 1H), 3.81 (d, J = 4.6 Hz, 1H), 3.72 (br s, 1H), 3.46 - 3.35 (m, 2H), 2.62 (d, J = 4.6 Hz, 3H). Example 52: Gel-Based Activity-Based Protein Profiling (ABPP) to assess DUB Recruiter binding to OTUB1 Recombinant OTUB1 from Sino Biological (12927-H07E) or produced and purified in- house (0.08μg/sample) was pre-treated with either DMSO vehicle or covalent ligand or bifunctional compounds at 37℃ for 30 min in 49 μL of PBS, and subsequently treated with of IA- Rhodamine (Setareh Biotech) at room temperature for 1 h. The reaction was stopped by addition of 4×reducing Laemmli SDS sample loading buffer (Alfa Aesar). After boiling at 95℃ for 5 min, the samples were separated on precast 4−20% Criterion TGX gels (Bio-Rad). Probe-labeled proteins were analyzed by in-gel fluorescence using an Azure 600 to provide a IC50 value for binding to OTUB1. These results are summarized in Table 3, wherein compounds that compete with IA-rhodamine at 50 µM to show between 100–80% of DMSO control fluorescence intensity are annotated “-“; compounds that show between 80-50% of DMSO control fluorescence intensity are annotated “ ”; compound that show between 50-30% of DMSO control fluorescence intensity are annotated “ ” and compounds that show less than 30% of DMSO control fluorescence intensity are annotated “+++”. Table 3. OTUB1 binding of Exemplary Compounds (IC50) Example 53: Intact-MS Assay via RapidFire-TOF Sample Preparation: Compound (1 μL of 5mM DMSO solution – final concentration 100 μM) was added to recombinant protein (1 μM, 49 μL) in buffer (25 mM Tris HCl; 10 mM NaCl; pH 7.5) in 96 well plates on ice. The reaction was mixed and incubated at 4C for 16 hours. Formic acid solution (1% in water, 50 μL) was added to quench the reaction and the samples were mixed, then centrifuged for 3 minutes at 3000xg and stored at 4C until analysis. Sample Analysis: A volume of 50 μL sample was aspirated from the 96-well plate by using an aspiration time of 300 ms, and 10 μL (limited by the sample loop size) was injected and concentrated on a C4 SPE cartridge (Agilent #G9203A, 20 μm, 4 μL). The sample load/wash time was 8000 ms at a flow rate of 0.6 mL/min (10% acetonitrile with 0.1% formic acid); elution time was 7000 ms at a flow rate of 0.5 mL/min (80% acetonitrile, 0.1% formic acid); re-equilibration time was 500 ms at a flow rate of 0.6 mL/min (10% acetonitrile, 0.1% formic acid). An Agilent 6230 TOF mass spectrometer was operated with a dual Agilent Jet Spray (AJS) ion source in the positive ionization mode and 3200 m/z range. The source parameters were as follows: gas temperature 325 °C, drying gas 8 L/min, nebulizer 35 psi, sheath gas 350 °C, sheath gas flow 11 L/min, capillary 3500V, nozzle 1000 V, fragmentor 175 V, and skimmer 65 V. Data Analysis: Data files were parsed into individual injections using the RapidFire UI software. In Agilent BioConfirm 12.0 TICs for individual injection files were integrated (with only one peak allowed per injection) and MS spectra extracted, using a designated background time range of 0.0 – 0.05 min. Decovolution for OTUB1 samples was performed using the maximum entropy method, filtering for peak signal-to-noise of >30, calculating average mass using the top 25% of peak height, using an explicit output mass range of 28,000 – 40,000 Da and a mass step of 0.2 Da, using a limited input range of 850 – 1800 m/z, and subtracting baseline with a baseline factor of 1.0. Biomolecule filters were set to a minimum consecutive charge states of 4, minimum fit score of 2, and minimum height threshold of 5%. Deconvolution was followed by bulk export of “Biomolecules” csv lists. Peaks present in the Biomolecules lists were compared to expected masses of protein + compound adducts for up to 5 modifications allowing for a window of +/- 2 amu in Excel. Percent modification values were calculated by comparing the peak height for a given species (i.e. a given degree of modification) versus the total peak height of unmodified and modified protein peaks in the deconvoluted spectrum. These results are summarized in Table 4, wherein compounds that demonstrated less than 20% modification for a single cysteine are annotated “ ; and compounds that demonstrated more than 20% modification for a single cysteine are annotated “++”. Table 4. Mass Spectrometry Based Deubiquitinase Screening Assay Example 53: Deubiquitinase Activity Assay Recombinant OTUB1 (500 nM) was pre-incubated with DMSO or Compound 100 (50 mM) for 1 hr. To initiate assay pre-treated OTUB1 enzyme was mixed 1:1 with di-Ub reaction mix for final concentrations of 250 nM OTUB1, 1.5 µM di-Ub, 12.5 µM UBE2D1 and 5 mM DTT. The appearance of mono-Ub was monitored by Western blotting over time by removing a portion of the reaction mix and adding Laemmli’s buffer to terminate the reaction. Blot shown is a representative gel from n=3 biologically independent experiments/group. Additionally, recombinant OTUB1 activity will be assessed using commercially available fluorescently-labeled di-Ubiquitin substrates from LifeSensors (DU0101). Western Blotting Proteins were resolved by SDS/PAGE and transferred to nitrocellulose membranes using the Trans-Blot Turbo transfer system (Bio-Rad). Membranes were blocked with 5% BSA in Tris- buffered saline containing Tween 20 (TBS-T) solution for 30 min at RT, washed in TBS-T, and probed with primary antibody diluted in recommended diluent per manufacturer overnight at 4℃. After 3 washes with TBS-T, the membranes were incubated in the dark with IR680- or IR800- conjugated secondary antibodies at 1:10,000 dilution in 5 % BSA in TBS-T at room temperature for 1 h. After 3 additional washes with TBST, blots were visualized using an Odyssey CLx Li-Cor fluorescent scanner. The membranes were stripped using ReBlot Plus Strong Antibody Stripping Solution (EMD Millipore) when additional primary antibody incubations were performed. Antibodies used in this study were CFTR (Cell Signaling Technologies, Rb mAb #78335), GAPDH (Proteintech, Ms mAb, #60004-1-Ig), OTUB1 (Abcam, Rb mAb, #ab175200, [EPR13028(B)]). Alternatively, target protein (e.g. CFTR) abundance was analyzed via Simple Western on a Jess instrument (Protein Simple) using the same primary antibodies and normalized to total protein abundance using a total protein detection kit (Protein Simple; DMTP01). Example 54: Quantitative TMT Proteomics Analysis Quantitative TMT-based proteomic analysis will be performed to assess relative protein abundance. Acquired MS data will be processed using a standard software package and searched against a protein database. Data analysis will be done to determine relative protein abundance of different target proteins. Example 55. Assessing protein abundance in ΔF508-CFTR human cystic fibrosis bronchial epithelial cells CFBE41o-4.7 with treatment of exemplary bifunctional compounds. CFBE41o-4.7 cells were seeded at 600k cells per 6cm 2 plate and were treated with either DMSO vehicle or 10uM DUBTAC (total of 0.1% DMSO) for 24 hours, then harvested, lysed, and cell lysate normalized to total protein content. Proteins were resolved by SDS–PAGE and transferred to nitrocellulose membranes. Membranes were blocked with 5% BSA in Tris-buffered saline containing Tween 20 (TBS-T) solution for 30 min at room temperature, washed in TBS-T and probed with primary antibody against CFTR (Cell Signaling Technologies, rabbit monoclonal antibody 78335, 1:1,000 dilution in 5% BSA) and GAPDH (Proteintech, mouse monoclonal antibody, 1:10,000 dilution in 5% BSA, 60004-1-Ig), and incubated overnight at 4 °C. After three washes with TBS-T, the membranes were incubated in the dark with IR680- or IR800-conjugated secondary antibodies at a 1:10,000 dilution in 5% BSA in TBS-T at room temperature for 1 h. After three additional washes with TBS-T, blots were visualized using an Odyssey CLx Li-Cor fluorescent scanner. CFTR abundance was quantified relative to GAPDH control.
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