| WO/2019/138018 | PESTICIDALLY ACTIVE HETEROCYCLIC DERIVATIVES WITH SULFUR CONTAINING SUBSTITUENTS |
| JP5567587 | Imidazopyrazine Syk inhibitor |
| WO/1999/005107 | VITRONECTIN RECEPTOR ANTAGONIST |
VA PORINO JINJO (US)
PINCHMAN JOSEPH ROBERT (US)
TRZOSS LYNNIE (US)
DONG QING (US)
KALDOR STEPHEN W
| WO2009053373A1 | 2009-04-30 | |||
| WO2022222995A1 | 2022-10-27 | |||
| WO2022228387A1 | 2022-11-03 |
| CLAIMS WHAT IS CLAIMED IS: 1. A compound of Formula (I), or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof: wherein: R1 is hydrogen, deuterium, halogen, -CN, -ORa, -NRcRd, C1-C6alkyl, C1-C6haloalkyl, C1-C6deuteroalkyl, C1-C6hydroxyalkyl, C1-C6aminoalkyl, C1-C6heteroalkyl, C2-C6alkenyl, C2-C6alkynyl, cycloalkyl, or heterocycloalkyl; wherein the alkyl, alkenyl, alkynyl, cycloalkyl, and heterocycloalkyl is optionally substituted with one or more R; R2 is hydrogen, deuterium, halogen, -CN, -ORa, -NRcRd, C1-C6alkyl, C1-C6haloalkyl, or C1-C6deuteroalkyl; X is N or CR3; Y is N or CR4; Z is N or CR5; R3 is hydrogen, deuterium, halogen, -CN, -ORa, -NRcRd, C1-C6alkyl, C1-C6haloalkyl, C1-C6deuteroalkyl, C1-C6hydroxyalkyl, C1-C6aminoalkyl, C1-C6heteroalkyl, C2-C6alkenyl, C2-C6alkynyl, cycloalkyl, or heterocycloalkyl; wherein the alkyl, alkenyl, alkynyl, cycloalkyl, and heterocycloalkyl is optionally substituted with one or more R; R4 is hydrogen, deuterium, halogen, -CN, -ORa, -NRcRd, C1-C6alkyl, C1-C6haloalkyl, C1-C6deuteroalkyl, C1-C6hydroxyalkyl, C1-C6aminoalkyl, C1-C6heteroalkyl, C2-C6alkenyl, C2-C6alkynyl, cycloalkyl, or heterocycloalkyl; wherein the alkyl, alkenyl, alkynyl, cycloalkyl, and heterocycloalkyl is optionally substituted with one or more R; R5 is hydrogen, deuterium, halogen, -CN, -ORa, -NRcRd, C1-C6alkyl, C1-C6haloalkyl, C1-C6deuteroalkyl, C1-C6hydroxyalkyl, C1-C6aminoalkyl, C1-C6heteroalkyl, C2-C6alkenyl, C2-C6alkynyl, cycloalkyl, or heterocycloalkyl; wherein the alkyl, alkenyl, alkynyl, cycloalkyl, and heterocycloalkyl is optionally substituted with one or more R; each R6 is independently hydrogen, deuterium, halogen, C1-C6alkyl, C1-C6haloalkyl, C1-C6deuteroalkyl, C1-C6hydroxyalkyl, C1-C6aminoalkyl, C1-C6heteroalkyl, cycloalkyl, or heterocycloalkyl; wherein the alkyl, cycloalkyl, and heterocycloalkyl is optionally substituted with one or more R; or two R6 are taken together to form a cycloalkyl or a heterocycloalkyl; each optionally substituted with deuterium, halogen, -CN, -OH, -ORa, -NRcRd, C1-C6alkyl, C1-C6haloalkyl, C1-C6deuteroalkyl, C1-C6hydroxyalkyl, C1-C6aminoalkyl, or C1-C6heteroalkyl; each R7 is independently deuterium, halogen, -CN, -OH, -ORa, -NRcRd, C1-C6alkyl, C1-C6haloalkyl, C1-C6deuteroalkyl, C1-C6hydroxyalkyl, C1-C6aminoalkyl, C1-C6heteroalkyl, C2-C6alkenyl, or C2- C6alkynyl; or two R7 on the same carbon are taken together to form an oxo; or two R7 on the same carbon or different carbons are taken together to form a cycloalkyl or heterocycloalkyl; each optionally substituted with one or more R; n is 0-4; T is N or CR8; U is N or CR9; R8 is hydrogen, deuterium, halogen, -CN, -OH, -ORa, -NRcRd, C1-C6alkyl, C1-C6haloalkyl, C1-C6deuteroalkyl, C1-C6hydroxyalkyl, C1-C6aminoalkyl, C1-C6heteroalkyl, C2-C6alkenyl, C2-C6alkynyl, cycloalkyl, or heterocycloalkyl; wherein the alkyl, alkenyl, alkynyl, cycloalkyl, and heterocycloalkyl is optionally substituted with one or more R; R9 is hydrogen, deuterium, halogen, -CN, -OH, -ORa, -NRcRd, C1-C6alkyl, C1-C6haloalkyl, C1-C6deuteroalkyl, C1-C6hydroxyalkyl, C1-C6aminoalkyl, C1-C6heteroalkyl, C2-C6alkenyl, C2-C6alkynyl, cycloalkyl, or heterocycloalkyl; wherein the alkyl, alkenyl, alkynyl, cycloalkyl, and heterocycloalkyl is optionally substituted with one or more R; R10 is hydrogen, deuterium, halogen, -CN, -OH, -ORa, -NRcRd, C1-C6alkyl, C1-C6haloalkyl, C1-C6deuteroalkyl, C1-C6hydroxyalkyl, C1-C6aminoalkyl, C1-C6heteroalkyl, C2-C6alkenyl, C2-C6alkynyl, cycloalkyl, or heterocycloalkyl; wherein the alkyl, alkenyl, alkynyl, cycloalkyl, and heterocycloalkyl is optionally substituted with one or more R; R11 is hydrogen, deuterium, halogen, -CN, -OH, -ORa, -NRcRd, C1-C6alkyl, C1-C6haloalkyl, C1-C6deuteroalkyl, C1-C6hydroxyalkyl, C1-C6aminoalkyl, C1-C6heteroalkyl, C2-C6alkenyl, C2-C6alkynyl, cycloalkyl, or heterocycloalkyl; wherein the alkyl, alkenyl, alkynyl, cycloalkyl, and heterocycloalkyl is optionally substituted with one or more R; R12 is cyano or halogen; each Ra is independently C1-C6alkyl, C1-C6haloalkyl, C1-C6deuteroalkyl, C1-C6hydroxyalkyl, C1-C6aminoalkyl, C1-C6heteroalkyl, C2-C6alkenyl, C2-C6alkynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, C1-C6alkylene(cycloalkyl), C1-C6alkylene(heterocycloalkyl), C1-C6alkylene(aryl), or C1-C6alkylene(heteroaryl); wherein each alkyl, alkylene, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, and heteroaryl is independently optionally substituted with one or more R; each Rb is independently hydrogen, C1-C6alkyl, C1-C6haloalkyl, C1-C6deuteroalkyl, C1-C6hydroxyalkyl, C1-C6aminoalkyl, C1-C6heteroalkyl, C2-C6alkenyl, C2-C6alkynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, C1-C6alkylene(cycloalkyl), C1-C6alkylene(heterocycloalkyl), C1-C6alkylene(aryl), or C1-C6alkylene(heteroaryl); wherein each alkyl, alkylene, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, and heteroaryl is independently optionally substituted with one or more R; and each Rc and Rd are independently hydrogen, C1-C6alkyl, C1-C6haloalkyl, C1-C6deuteroalkyl, C1-C6hydroxyalkyl, C1-C6aminoalkyl, C1-C6heteroalkyl, C2-C6alkenyl, C2-C6alkynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, C1-C6alkylene(cycloalkyl), C1-C6alkylene(heterocycloalkyl), C1-C6alkylene(aryl), or C1-C6alkylene(heteroaryl); wherein each alkyl, alkylene, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, and heteroaryl is independently optionally substituted with one or more R; or Rc and Rd are taken together with the atom to which they are attached to form a heterocycloalkyl optionally substituted with one or more R; each R is independently deuterium, halogen, -CN, -OH, -OC1-C6alkyl, -NH2, -NHC1-C6alkyl, - N(C1-C6alkyl)2, -NHC(=O)OC1-C6alkyl, -C(=O)C1-C6alkyl, -C(=O)OH, -C(=O)OC1-C6alkyl, - C(=O)NH2, -C(=O)N(C1-C6alkyl)2, -C(=O)NHC1-C6alkyl, C1-C6alkyl, C1-C6haloalkyl, C1-C6deuteroalkyl, C1-C6hydroxyalkyl, C1-C6aminoalkyl, or C1-C6heteroalkyl; or two R on the same atom are taken together to form an oxo; provided that when X is CR3, Y is CR4, and Z is CR5; then one of R3, R4, and R5 is not hydrogen and R4 is not -OMe; and provided that when X is CH, Y is CH, and Z is CH; then R2 is not hydrogen. 2. The compound of claim 1, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein X is N. 3. The compound of claim 1, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein X is CR3. 4. The compound of any one of claims 1-3, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein Y is N. 5. The compound of any one of claims 1-3, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein Y is CR4. 6. The compound of any one of claims 1-5, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein Z is N. 7. The compound of any one of claims 1-5, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein Z is CR5. 8. The compound of claim 1, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein the compound is of Formula (Ia): ( ) 9. The compound of claim 1, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein the compound is of Formula (Ib): Formula (Ib). 10. The compound of claim 1, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein the compound is of Formula (Ic): Formula (Ic). 11. The compound of claim 1, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein the compound is of Formula (Id): Formula (Id); wherein one of R3, R4, and R5 is not hydrogen and R4 is not -OMe. 12. The compound of claim 1, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein the compound is of Formula (Ie): Formula (Ie); wherein R2 is deuterium, halogen, -CN, -ORa, -NRcRd, C1-C6alkyl, C1-C6haloalkyl, or C1-C6deuteroalkyl. 13. The compound of any one of claims 1-12, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein R1 is halogen, C1-C6alkyl, C1-C6haloalkyl, C2-C6alkynyl, or cycloalkyl. 14. The compound of any one of claims 1-12, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein R1 is C1-C6alkyl or cycloalkyl. 15. The compound of any one of claims 1-12, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein R1 is C1-C6alkyl. 16. The compound of any one of claims 1-12, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein R1 is cycloalkyl. 17. The compound of any one of claims 1-12, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein R1 is halogen. 18. The compound of any one of claims 1-17, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein R2 is hydrogen or halogen. 19. The compound of any one of claims 1-18, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein R2 is halogen. 20. The compound of any one of claims 1-19, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein R3 is hydrogen, or halogen, or C1-C6alkyl. 21. The compound of any one of claims 1-20, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein R3 is hydrogen or halogen. 22. The compound of any one of claims 1-21, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein R3 is hydrogen. 23. The compound of any one of claims 1-22, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein R4 is hydrogen, or halogen, or C1-C6alkyl. 24. The compound of any one of claims 1-23, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein R4 is hydrogen or halogen. 25. The compound of any one of claims 1-24, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein R4 is hydrogen. 26. The compound of any one of claims 1-25, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein R5 is hydrogen, or halogen, or C1-C6alkyl. 27. The compound of any one of claims 1-26, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein R5 is hydrogen or halogen. 28. The compound of any one of claims 1-27, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein R5 is hydrogen. 29. The compound of any one of claims 1-28, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein each R6 is independently hydrogen, deuterium, or C1-C6alkyl. 30. The compound of any one of claims 1-29, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein one R6 is hydrogen and the other R6 is C1-C6alkyl. 31. The compound of any one of claims 1-29, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein each R6 is deuterium. 32. The compound of any one of claims 1-29, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein each R6 is independently C1-C6alkyl. 33. The compound of any one of claims 1-29, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein each R6 is hydrogen. 34. The compound of any one of claims 1-33, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein each R7 is independently C1-C6alkyl. 35. The compound of any one of claims 1-33, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein two R7 on the same carbon or different carbons are taken together to form a cycloalkyl. 36. The compound of any one of claims 1-35, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein n is 0 or 1. 37. The compound of any one of claims 1-35, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein n is 1 or 2. 38. The compound of any one of claims 1-37, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein . 39. The compound of any one of claims 1-37, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein . 40. The compound of any one of claims 1-39, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein R8 is hydrogen, halogen, -CN, -ORa, C1-C6alkyl, or C1-C6haloalkyl. 41. The compound of any one of claims 1-40, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein R9 is hydrogen, halogen, - C1-C6alkyl, C1-C6haloalkyl, or cycloalkyl. 42. The compound of any one of claims 1-41, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein R10 is hydrogen, halogen, C1-C6alkyl, or C1-C6haloalkyl. 43. The compound of any one of claims 1-42, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein R11 is hydrogen, halogen, C1-C6alkyl, or C1-C6haloalkyl. 44. The compound of any one of claims 1-43, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein R12 is cyano. 45. The compound of any one of claims 1-43, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein R12 is halogen. 46. The compound of any one of claims 1-43, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, wherein R12 is fluoro or chloro. 47. The compound of claim 1, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, selected from a compound in table 1. 48. A pharmaceutical composition comprising a compound of any one of claims 1-47, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, and a pharmaceutically acceptable excipient. 49. A method of treating cancer in a subject in need thereof, the method comprising administering a compound of any one of claims 1-47, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof. 50. The method of claim 49, wherein the cancer is breast cancer, ovarian cancer, pancreatic cancer, prostate cancer, a hematological cancer, gastrointestinal cancer, or lung cancer. 51. A method of treating a cancer comprising a BRCA1 and/or a BRCA2 mutation in a subject in need thereof, the method comprising administering a compound of any one of claims 1-47, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof. 52. A method of treating a cancer comprising a mutation in a gene conferring homologous repair deficiency in a subject in need thereof, the method comprising administering a compound of any one of claims 1-47, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof. 53. The method of claim 52, wherein the mutation in a gene conferring homologous repair deficiency comprises ATM, BRCA1, BRCA2, BARD1, BRIP1, CDK12, CHEK1, CHEK2, FANCL, PALB2, RAD51B, RAD51C, RAD51D, or RAD54L, or any combinations thereof. 54. The method of any one of claims 49-53, wherein the cancer is bladder cancer, brain & CNS cancer, breast cancer, cervical cancer, colorectal cancer, esophagus cancer, Hodgkin lymphoma, non- Hodgkin lymphoma, kidney cancer, leukemia, lung cancer, melanoma, myeloma, oral cavity cancer, ovarian cancer, pancreatic cancer, prostate cancer, skin cancer, stomach cancer, thyroid cancer, or uterus cancer. 55. The method of any one of claims 49-54, wherein the cancer is metastatic cancer. 56. The method of any one of claims 49-55, wherein the cancer has metastasized in the brain. 57. A method of treating a cancer that is present in the brain in a subject in need thereof, the method comprising administering a compound of any one of claims 1-47, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof. 58. A method of treating brain cancer in a subject in need thereof, the method comprising administering a compound of any one of claims 1-47, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof. |
[0085] The absolute label (abs) is added to a chiral center to denote that it is unambiguously a pure sample of the drawn stereoisomer. [0086] The OR label (or) denotes a pure substance, but the absolute configuration of the stereochemical center is unknown. After chiral separation with pure structures isolated, multiple OR labels (OR indicates purity) with the same numerical value will indicates that a sample is one of a pair of pure enantiomers (but the absolute configuration of the stereochemical center is unknown). [0087] The AND label (and) denotes both isomers are present at the depicted stereochemical center. Assigning different numerical values to the AND labels denotes that they are independent of each other. The use of AND labels with the same values indicate that the two stereocenters are relative to each other and can only change in concert. [0088] In some embodiments, the compound disclosed herein, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, is selected from . Further Forms of Compounds Disclosed Herein Isomers/Stereoisomers [0089] In some embodiments, the compounds described herein exist as geometric isomers. In some embodiments, the compounds described herein possess one or more double bonds. The compounds presented herein include all cis, trans, syn, anti, entgegen (E), and zusammen (Z) isomers as well as the corresponding mixtures thereof. In some situations, the compounds described herein possess one or more chiral centers and each center exists in the R configuration, or S configuration. The compounds described herein include all diastereomeric, enantiomeric, and epimeric forms as well as the corresponding mixtures thereof. In additional embodiments of the compounds and methods provided herein, mixtures of enantiomers and/or diastereoisomers, resulting from a single preparative step, combination, or interconversion are useful for the applications described herein. In some embodiments, the compounds described herein are prepared as their individual stereoisomers by reacting a racemic mixture of the compound with an optically active resolving agent to form a pair of diastereoisomeric compounds, separating the diastereomers and recovering the optically pure enantiomers. In some embodiments, dissociable complexes are preferred. In some embodiments, the diastereomers have distinct physical properties (e.g., melting points, boiling points, solubilities, reactivity, etc.) and are separated by taking advantage of these dissimilarities. In some embodiments, the diastereomers are separated by chiral chromatography, or preferably, by separation/resolution techniques based upon differences in solubility. In some embodiments, the optically pure enantiomer is then recovered, along with the resolving agent, by any practical means that would not result in racemization. Labeled compounds [0090] In some embodiments, the compounds described herein exist in their isotopically-labeled forms. In some embodiments, the methods disclosed herein include methods of treating diseases by administering such isotopically-labeled compounds. In some embodiments, the methods disclosed herein include methods of treating diseases by administering such isotopically-labeled compounds as pharmaceutical compositions. Thus, in some embodiments, the compounds disclosed herein include isotopically-labeled compounds, which are identical to those recited herein, but for the fact that one or more atoms are replaced by an atom having an atomic mass or mass number different from the atomic mass or mass number usually found in nature. Examples of isotopes that can be incorporated into compounds disclosed herein include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorous, sulfur, fluorine, and chloride, such as 2 H, 3 H, 13 C, 14 C, l5 N, 18 O, 17 O, 31 P, 32 P, 35 S, 18 F, and 36 Cl, respectively. Compounds described herein, and the pharmaceutically acceptable salts, solvates, or stereoisomers thereof which contain the aforementioned isotopes and/or other isotopes of other atoms are within the scope of this invention. Certain isotopically-labeled compounds, for example those into which radioactive isotopes such as 3 H and 14 C are incorporated, are useful in drug and/or substrate tissue distribution assays. Tritiated, i.e., 3 H and carbon-14, i.e., 14 C, isotopes are particularly preferred for their ease of preparation and detectability. Further, substitution with heavy isotopes such as deuterium, i.e., 2 H, produces certain therapeutic advantages resulting from greater metabolic stability, for example increased in vivo half-life or reduced dosage requirements. [0091] In some embodiments, the compounds described herein are labeled by other means, including, but not limited to, the use of chromophores or fluorescent moieties, bioluminescent labels, or chemiluminescent labels. Pharmaceutically acceptable salts [0092] In some embodiments, the compounds described herein exist as their pharmaceutically acceptable salts. In some embodiments, the methods disclosed herein include methods of treating diseases by administering such pharmaceutically acceptable salts. In some embodiments, the methods disclosed herein include methods of treating diseases by administering such pharmaceutically acceptable salts as pharmaceutical compositions. [0093] In some embodiments, the compounds described herein possess acidic or basic groups and therefore react with any of a number of inorganic or organic bases, and inorganic and organic acids, to form a pharmaceutically acceptable salt. In some embodiments, these salts are prepared in situ during the final isolation and purification of the compounds disclosed herein, or a solvate, or stereoisomer thereof, or by separately reacting a purified compound in its free form with a suitable acid or base, and isolating the salt thus formed. [0094] Examples of pharmaceutically acceptable salts include those salts prepared by reaction of the compounds described herein with a mineral, organic acid or inorganic base, such salts including, acetate, acrylate, adipate, alginate, aspartate, benzoate, benzenesulfonate, bisulfate, bisulfite, bromide, butyrate, butyn-1,4-dioate, camphorate, camphorsulfonate, caproate, caprylate, chlorobenzoate, chloride, citrate, cyclopentanepropionate, decanoate, digluconate, dihydrogenphosphate, dinitrobenzoate, dodecylsulfate, ethanesulfonate, formate, fumarate, glucoheptanoate, glycerophosphate, glycolate, hemisulfate, heptanoate, hexanoate, hexyne-1,6-dioate, hydroxybenzoate, γ-hydroxybutyrate, hydrochloride, hydrobromide, hydroiodide, 2-hydroxyethanesulfonate, iodide, isobutyrate, lactate, maleate, malonate, methanesulfonate, mandelate, metaphosphate, methanesulfonate, methoxybenzoate, methylbenzoate, monohydrogenphosphate, 1-napthalenesulfonate, 2-napthalenesulfonate, nicotinate, nitrate, palmoate, pectinate, persulfate, 3- phenylpropionate, phosphate, picrate, pivalate, propionate, pyrosulfate, pyrophosphate, propiolate, phthalate, phenylacetate, phenylbutyrate, propanesulfonate, salicylate, succinate, sulfate, sulfite, succinate, suberate, sebacate, sulfonate, tartrate, thiocyanate, tosylate, undecanoate, and xylenesulfonate. [0095] Further, the compounds described herein can be prepared as pharmaceutically acceptable salts formed by reacting the free base form of the compound with a pharmaceutically acceptable inorganic or organic acid, including, but not limited to, inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid metaphosphoric acid, and the like; and organic acids such as acetic acid, propionic acid, hexanoic acid, cyclopentanepropionic acid, glycolic acid, pyruvic acid, lactic acid, malonic acid, succinic acid, malic acid, maleic acid, fumaric acid, p-toluenesulfonic acid, tartaric acid, trifluoroacetic acid, citric acid, benzoic acid, 3-(4-hydroxybenzoyl)benzoic acid, cinnamic acid, mandelic acid, arylsulfonic acid, methanesulfonic acid, ethanesulfonic acid, 1,2-ethanedisulfonic acid, 2- hydroxyethanesulfonic acid, benzenesulfonic acid, 2-naphthalenesulfonic acid, 4-methylbicyclo-[2.2.2]oct- 2-ene-1-carboxylic acid, glucoheptonic acid, 4,4’-methylenebis-(3-hydroxy-2-ene-1 -carboxylic acid), 3- phenylpropionic acid, trimethylacetic acid, tertiary butylacetic acid, lauryl sulfuric acid, gluconic acid, glutamic acid, hydroxynaphthoic acid, salicylic acid, stearic acid and muconic acid. In some embodiments, other acids, such as oxalic, while not in themselves pharmaceutically acceptable, are employed in the preparation of salts useful as intermediates in obtaining the compounds disclosed herein, solvate, or stereoisomer thereof and their pharmaceutically acceptable acid addition salts. [0096] In some embodiments, those compounds described herein which comprise a free acid group react with a suitable base, such as the hydroxide, carbonate, bicarbonate, sulfate, of a pharmaceutically acceptable metal cation, with ammonia, or with a pharmaceutically acceptable organic primary, secondary, tertiary, or quaternary amine. Representative salts include the alkali or alkaline earth salts, like lithium, sodium, potassium, calcium, and magnesium, and aluminum salts and the like. Illustrative examples of bases include sodium hydroxide, potassium hydroxide, choline hydroxide, sodium carbonate, N + (C1-4 alkyl)4, and the like. [0097] Representative organic amines useful for the formation of base addition salts include ethylamine, diethylamine, ethylenediamine, ethanolamine, diethanolamine, piperazine and the like. It should be understood that the compounds described herein also include the quaternization of any basic nitrogen- containing groups they contain. In some embodiments, water or oil-soluble or dispersible products are obtained by such quaternization. Solvates [0098] In some embodiments, the compounds described herein exist as solvates. The invention provides for methods of treating diseases by administering such solvates. The invention further provides for methods of treating diseases by administering such solvates as pharmaceutical compositions. [0099] Solvates contain either stoichiometric or non-stoichiometric amounts of a solvent, and, in some embodiments, are formed during the process of crystallization with pharmaceutically acceptable solvents such as water, ethanol, and the like. Hydrates are formed when the solvent is water, or alcoholates are formed when the solvent is alcohol. Solvates of the compounds described herein can be conveniently prepared or formed during the processes described herein. By way of example only, hydrates of the compounds described herein can be conveniently prepared by recrystallization from an aqueous/organic solvent mixture, using organic solvents including, but not limited to, dioxane, tetrahydrofuran or methanol. In addition, the compounds provided herein can exist in unsolvated as well as solvated forms. In general, the solvated forms are considered equivalent to the unsolvated forms for the purposes of the compounds and methods provided herein. Tautomers [00100] In some situations, compounds exist as tautomers. The compounds described herein include all possible tautomers within the formulas described herein. Tautomers are compounds that are interconvertible by migration of a hydrogen atom, accompanied by a switch of a single bond and adjacent double bond. In bonding arrangements where tautomerization is possible, a chemical equilibrium of the tautomers will exist. All tautomeric forms of the compounds disclosed herein are contemplated. The exact ratio of the tautomers depends on several factors, including temperature, solvent, and pH. Method of Treatment [00101] Disclosed herein are methods of treatment of a disease in which inhibition of PARP is beneficial, the method comprising administering a compound disclosed herein. Also disclosed herein are methods of treatment of a disease in which inhibition of PARP1 is beneficial, the method comprising administering a compound disclosed herein. In some embodiments, the disease is cancer. In some embodiments, the cancer is breast cancer, ovarian cancer, pancreatic cancer, prostate cancer, a hematological cancer, a gastrointestinal cancer such as gastric cancer and colorectal cancer, or lung cancer. In some embodiments, the cancer is breast cancer, ovarian cancer, pancreatic cancer, or prostate cancer. In some embodiment, the cancer is leukemia, colon cancer, glioblastoma, lymphoma, melanoma, or cervical cancer. In some embodiments, the cancer is bladder cancer, brain & CNS cancer, breast cancer, cervical cancer, colorectal cancer, esophagus cancer, Hodgkin lymphoma, non-Hodgkin lymphoma, kidney cancer, leukemia, lung cancer, melanoma, myeloma, oral cavity cancer, ovarian cancer, pancreatic cancer, prostate cancer, skin cancer, stomach cancer, thyroid cancer, or uterus cancer. [00102] In some embodiments, the cancer is metastatic cancer. In some embodiments, the cancer has metastasized in the brain. [00103] In some embodiments, the cancer comprises a BRCA1 and/or a BRCA2 mutation. [00104] In some embodiments, the cancer comprising a BRCA1 and/or a BRCA2 mutation is bladder cancer, brain & CNS cancers, breast cancer, cervical cancer, colorectal cancer, esophagus cancer, Hodgkin lymphoma, non-Hodgkin lymphoma, kidney cancer, leukemia, lung cancer, melanoma, myeloma, oral cavity cancer, ovarian cancer, pancreatic cancer, prostate cancer, skin cancer, stomach cancer, thyroid cancer, or uterus cancer. [00105] In some embodiments, the cancer is a cancer deficient in Flomologous Recombination (FIR) dependent DNA DSB repair activity. The FIR dependent DNA DSB repair pathway repairs double-strand breaks (DSBs) in DNA via homologous mechanisms to reform a continuous DNA helix. The components of the FIR dependent DNA DSB repair pathway include, but are not limited to, ATM (NM_000051 ), RAD51 (NM_002875), RAD51 L1 (NM_002877), RAD51 C (NM_002876), RAD51 L3 (NM_002878), DMC1 (NM_007068), XRCC2 (NM_005431 ), XRCC3 (NM_005432), RAD52 (NM_002879), RAD54L (NM_003579), RAD54B (NM_012415), BRCA1 (NM_007295), BRCA2 (NM_000059), RAD50 (NM_005732), MRE11 A (NM_005590) and NBS1 (NM_002485). Other proteins involved in the FIR dependent DNA DSB repair pathway include regulatory factors such as EMSY. In some embodiments, the cancer which is deficient in FIR dependent DNA DSB repair comprises one or more cancer cells which have a reduced or abrogated ability to repair DNA DSBs through that pathway, relative to normal cells i.e. the activity of the FIR dependent DNA DSB repair pathway may be reduced or abolished in the one or more cancer cells. [00106] In some embodiments, the activity of one or more components of the FIR dependent DNA DSB repair pathway is abolished in the one or more cancer cells of an individual having a cancer which is deficient in FIR dependent DNA DSB repair. [00107] In some embodiments, the cancer cells have a BRCA1 and/or a BRCA2 deficient phenotype i.e. BRCA1 and/or BRCA2 activity is reduced or abolished in the cancer cells. Cancer cells with this phenotype may be deficient in BRCA1 and/or BRCA2, i.e. expression and/or activity of BRCA1 and/or BRCA2 may be reduced or abolished in the cancer cells, for example by means of mutation or polymorphism in the encoding nucleic acid, or by means of amplification, mutation or polymorphism in a gene encoding a regulatory factor, for example the EMSY gene which encodes a BRCA2 regulatory factor. BRCA1 and BRCA2 are known tumor suppressors whose wild-type alleles are frequently lost in tumors of heterozygous carriers. Amplification of the EMSY gene, which encodes a BRCA2 binding factor, is also known to be associated with breast and ovarian cancer. Carriers of mutations in BRCA1 and/or BRCA2 are also at elevated risk of certain cancers, including breast cancer, ovarian cancer, pancreatic cancer, prostate cancer, a hematological cancer, gastrointestinal cancer, and lung cancer. [00108] Also disclosed herein is a method of treating a cancer comprising a mutation in a gene conferring homologous repair deficiency in a subject in need thereof, the method comprising administering a compound disclosed herein, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof. In some embodiments, the mutation in a gene conferring homologous repair deficiency comprises ATM, BRCA1, BRCA2, BARD1, BRIP1, CDK12, CHEK1, CHEK2, FANCL, PALB2, RAD51B, RAD51C, RAD51D, or RAD54L, or any combinations thereof. [00109] Also disclosed herein is a method for treating a cancer that is present in the brain, the method comprising administering a compound disclosed herein, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof. [00110] In some embodiments, the cancer that is present in the brain arises from primary peripheral tumors that have metastasized to the brain. In some embodiments, the cancer that is present in the brain arises from primary brain tissues. [00111] Also disclosed herein is a method for treating brain cancer, the method comprising administering a compound disclosed herein, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof. [00112] In some embodiments, the brain cancer is a primary brain tumor that starts in the brain and tends to stay there. [00113] In some embodiments, the brain cancer is a secondary brain tumor. These cancers start somewhere else in the body and travel to the brain. Lung, breast, kidney, colon, and skin cancers are among the most common cancers that spread to the brain. [00114] In some embodiments, the compound disclosed herein, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, is capable of penetrating the blood brain barrier (BBB). In some embodiments, the ratio of compound that penetrates the BBB is > 0.1 , wherein 1 is complete BBB penetration, and 0 is no penetration. In some embodiments, the ratio of compound that penetrates the BBB is > 0.2. In some embodiments, the ratio of compound that penetrates the BBB is > 0.3. In some embodiments, the ratio of compound that penetrates the BBB is measured using the rat kp,uu assay. In some embodiments, the compound has a ratio of >0.3 (i.e. from 0.3 to 1) as determined in the rat kp,uu assay. Dosing [00115] In certain embodiments, the compositions containing the compound(s) described herein are administered for prophylactic and/or therapeutic treatments. In certain therapeutic applications, the compositions are administered to a patient already suffering from a disease or condition, in an amount sufficient to cure or at least partially arrest at least one of the symptoms of the disease or condition. Amounts effective for this use depend on the severity and course of the disease or condition, previous therapy, the patient’s health status, weight, and response to the drugs, and the judgment of the treating physician. Therapeutically effective amounts are optionally determined by methods including, but not limited to, a dose escalation and/or dose ranging clinical trial. [00116] In prophylactic applications, compositions containing the compounds described herein are administered to a patient susceptible to or otherwise at risk of a particular disease, disorder, or condition. Such an amount is defined to be a “prophylactically effective amount or dose.” In this use, the precise amounts also depend on the patient’s state of health, weight, and the like. When used in patients, effective amounts for this use will depend on the severity and course of the disease, disorder or condition, previous therapy, the patient’s health status and response to the drugs, and the judgment of the treating physician. In one aspect, prophylactic treatments include administering to a mammal, who previously experienced at least one symptom of or risk factor for the disease being treated and is currently in remission, a pharmaceutical composition comprising a compound described herein, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, in order to prevent a return of the symptoms of the disease or condition. [00117] In certain embodiments wherein the patient’s condition does not improve, upon the doctor’s discretion the administration of the compounds are administered chronically, that is, for an extended period of time, including throughout the duration of the patient’s life in order to ameliorate or otherwise control or limit the symptoms of the patient’s disease or condition. [00118] In certain embodiments wherein a patient’s status does improve, the dose of drug being administered is temporarily reduced or temporarily suspended for a certain length of time (i.e., a “drug holiday”). In specific embodiments, the length of the drug holiday is between 2 days and 1 year, including by way of example only, 2 days, 3 days, 4 days, 5 days, 6 days, 7 days, 10 days, 12 days, 15 days, 20 days, 28 days, or more than 28 days. The dose reduction during a drug holiday is, by way of example only, by 10%-100%, including by way of example only 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, and 100%. [00119] Once improvement of the patient’s conditions has occurred, a maintenance dose is administered if necessary. Subsequently, in specific embodiments, the dosage, or the frequency of administration, or both, is reduced, as a function of the symptoms, to a level at which the improved disease, disorder or condition is retained. In certain embodiments, however, the patient requires intermittent or daily treatment on a long- term basis upon any recurrence of symptoms. [00120] The amount of a given agent that corresponds to such an amount varies depending upon factors such as the particular compound, disease condition and its severity, the identity (e.g., weight, sex) of the subject or host in need of treatment, but nevertheless is determined according to the particular circumstances surrounding the case, including, e.g., the specific agent being administered, the route of administration, the condition being treated, and the subject or host being treated. [00121] In general, however, doses employed for adult human treatment are typically in the range of 0.01 mg-5000 mg per day. In one aspect, doses employed for adult human treatment are from about 1 mg to about 1000 mg per day. In one embodiment, the desired dose is conveniently presented in a single dose or in divided doses administered simultaneously or at appropriate intervals, for example as two, three, four or more sub-doses per day. [00122] In one embodiment, the daily dosages appropriate for the compound described herein, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, are from about 0.01 to about 50 mg/kg per body weight. In some embodiments, the daily dosage, or the amount of active in the dosage form are lower or higher than the ranges indicated herein, based on a number of variables in regard to an individual treatment regime. In various embodiments, the daily and unit dosages are altered depending on a number of variables including, but not limited to, the activity of the compound used, the disease or condition to be treated, the mode of administration, the requirements of the individual subject, the severity of the disease or condition being treated, and the judgment of the practitioner. [00123] Toxicity and therapeutic efficacy of such therapeutic regimens are determined by standard pharmaceutical procedures in cell cultures or experimental animals, including, but not limited to, the determination of the LD 10 and the ED 90 . The dose ratio between the toxic and therapeutic effects is the therapeutic index and it is expressed as the ratio between LD 50 and ED 50 . In certain embodiments, the data obtained from cell culture assays and animal studies are used in formulating the therapeutically effective daily dosage range and/or the therapeutically effective unit dosage amount for use in mammals, including humans. In some embodiments, the daily dosage amount of the compounds described herein lies within a range of circulating concentrations that include the ED 50 with minimal toxicity. In certain embodiments, the daily dosage range and/or the unit dosage amount varies within this range depending upon the dosage form employed and the route of administration utilized. [00124] In any of the aforementioned aspects are further embodiments in which the effective amount of the compound described herein, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, is: (a) systemically administered to the mammal; and/or (b) administered orally to the mammal; and/or (c) intravenously administered to the mammal; and/or (d) administered by injection to the mammal; and/or (e) administered topically to the mammal; and/or (f) administered non-systemically or locally to the mammal. [00125] In any of the aforementioned aspects are further embodiments comprising single administrations of the effective amount of the compound, including further embodiments in which (i) the compound is administered once a day; or (ii) the compound is administered to the mammal multiple times over the span of one day. [00126] In any of the aforementioned aspects are further embodiments comprising multiple administrations of the effective amount of the compound, including further embodiments in which (i) the compound is administered continuously or intermittently: as in a single dose; (ii) the time between multiple administrations is every 6 hours; (iii) the compound is administered to the mammal every 8 hours; (iv) the compound is administered to the subject every 12 hours; (v) the compound is administered to the subject every 24 hours. In further or alternative embodiments, the method comprises a drug holiday, wherein the administration of the compound is temporarily suspended or the dose of the compound being administered is temporarily reduced; at the end of the drug holiday, dosing of the compound is resumed. In one embodiment, the length of the drug holiday varies from 2 days to 1 year. Routes of Administration [00127] Suitable routes of administration include, but are not limited to, oral, intravenous, rectal, aerosol, parenteral, ophthalmic, pulmonary, transmucosal, transdermal, vaginal, otic, nasal, and topical administration. In addition, by way of example only, parenteral delivery includes intramuscular, subcutaneous, intravenous, intramedullary injections, as well as intrathecal, direct intraventricular, intraperitoneal, intralymphatic, and intranasal injections. [00128] In certain embodiments, a compound as described herein is administered in a local rather than systemic manner, for example, via injection of the compound directly into an organ, often in a depot preparation or sustained release formulation. In specific embodiments, long acting formulations are administered by implantation (for example subcutaneously or intramuscularly) or by intramuscular injection. Furthermore, in other embodiments, the drug is delivered in a targeted drug delivery system, for example, in a liposome coated with organ specific antibody. In such embodiments, the liposomes are targeted to and taken up selectively by the organ. In yet other embodiments, the compound as described herein is provided in the form of a rapid release formulation, in the form of an extended release formulation, or in the form of an intermediate release formulation. In yet other embodiments, the compound described herein is administered topically. Pharmaceutical Compositions/Formulations [00129] The compounds described herein are administered to a subject in need thereof, either alone or in combination with pharmaceutically acceptable carriers, excipients, or diluents, in a pharmaceutical composition, according to standard pharmaceutical practice. In one embodiment, the compounds of this invention may be administered to animals. The compounds can be administered orally or parenterally, including the intravenous, intramuscular, intraperitoneal, subcutaneous, rectal, and topical routes of administration. [00130] In another aspect, provided herein are pharmaceutical compositions comprising a compound described herein, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, and at least one pharmaceutically acceptable excipient. Pharmaceutical compositions are formulated in a conventional manner using one or more pharmaceutically acceptable excipients that facilitate processing of the active compounds into preparations that can be used pharmaceutically. Proper formulation is dependent upon the route of administration chosen. A summary of pharmaceutical compositions described herein can be found, for example, in Remington: The Science and Practice of Pharmacy, Nineteenth Ed (Easton, Pa.: Mack Publishing Company, 1995); Hoover, John E., Remington’s Pharmaceutical Sciences, Mack Publishing Co., Easton, Pennsylvania 1975; Liberman, H.A. and Lachman, L., Eds., Pharmaceutical Dosage Forms, Marcel Decker, New York, N.Y., 1980; and Pharmaceutical Dosage Forms and Drug Delivery Systems, Seventh Ed. (Lippincott Williams & Wilkins1999), herein incorporated by reference for such disclosure. [00131] In some embodiments, the pharmaceutically acceptable excipient is selected from carriers, binders, filling agents, suspending agents, flavoring agents, sweetening agents, disintegrating agents, dispersing agents, surfactants, lubricants, colorants, diluents, solubilizers, moistening agents, plasticizers, stabilizers, penetration enhancers, wetting agents, anti-foaming agents, antioxidants, preservatives, and any combinations thereof. [00132] The pharmaceutical compositions described herein are administered to a subject by appropriate administration routes, including, but not limited to, oral, parenteral (e.g., intravenous, subcutaneous, intramuscular), intranasal, buccal, topical, rectal, or transdermal administration routes. The pharmaceutical formulations described herein include, but are not limited to, aqueous liquid dispersions, liquids, gels, syrups, elixirs, slurries, suspensions, self-emulsifying dispersions, solid solutions, liposomal dispersions, aerosols, solid oral dosage forms, powders, immediate release formulations, controlled release formulations, fast melt formulations, tablets, capsules, pills, powders, dragees, effervescent formulations, lyophilized formulations, delayed release formulations, extended release formulations, pulsatile release formulations, multiparticulate formulations, and mixed immediate and controlled release formulations. [00133] Pharmaceutical compositions including compounds described herein, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof are manufactured in a conventional manner, such as, by way of example only, by means of conventional mixing, dissolving, granulating, dragee-making, levigating, emulsifying, encapsulating, entrapping, or compression processes. [00134] Pharmaceutical compositions for oral use are obtained by mixing one or more solid excipient with one or more of the compounds described herein, optionally grinding the resulting mixture, and processing the mixture of granules, after adding suitable auxiliaries, if desired, to obtain tablets or dragee cores. Suitable excipients include, for example, fillers such as sugars, including lactose, sucrose, mannitol, or sorbitol; cellulose preparations such as, for example, maize starch, wheat starch, rice starch, potato starch, gelatin, gum tragacanth, methylcellulose, microcrystalline cellulose, hydroxypropylmethylcellulose, sodium carboxymethylcellulose; or others such as polyvinylpyrrolidone (PVP or povidone) or calcium phosphate. If desired, disintegrating agents are added, such as the cross-linked croscarmellose sodium, polyvinylpyrrolidone, agar, or alginic acid or a salt thereof such as sodium alginate. In some embodiments, dyestuffs or pigments are added to the tablets or dragee coatings for identification or to characterize different combinations of active compound doses. [00135] Pharmaceutical compositions that are administered orally include push-fit capsules made of gelatin, as well as soft, sealed capsules made of gelatin and a plasticizer, such as glycerol or sorbitol. The push-fit capsules contain the active ingredients in admixture with filler such as lactose, binders such as starches, and/or lubricants such as talc or magnesium stearate and, optionally, stabilizers. In soft capsules, the active compounds are dissolved or suspended in suitable liquids, such as fatty oils, liquid paraffin, or liquid polyethylene glycols. In some embodiments, stabilizers are added. [00136] Pharmaceutical compositions for parental use are formulated as infusions or injections. In some embodiments, the pharmaceutical composition suitable for injection or infusion includes sterile aqueous solutions, or dispersions, or sterile powders comprising a compound described herein, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof. In some embodiments, the pharmaceutical composition comprises a liquid carrier. In some embodiments, the liquid carrier is a solvent or liquid dispersion medium comprising, for example, water, saline, ethanol, a polyol (for example, glycerol, propylene glycol, liquid polyethylene glycols, and the like), vegetable oils, nontoxic glyceryl esters, and any combinations thereof. In some embodiments, the pharmaceutical compositions further comprise a preservative to prevent growth of microorganisms. Combination [00137] Disclosed herein are methods of treating cancer using a compound disclosed herein, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, in combination with an additional therapeutic agent. [00138] In some embodiments, the additional therapeutic agent is an anticancer agent. [00139] In some embodiments, the additional therapeutic agent is administered at the same time as the compound disclosed herein. In some embodiments, the additional therapeutic agent and the compound disclosed herein are administered sequentially. In some embodiments, the additional therapeutic agent is administered less frequently than the compound disclosed herein. In some embodiments, the additional therapeutic agent is administered more frequently than the compound disclosed herein. In some embodiments, the additional therapeutic agent is administered prior than the administration of the compound disclosed herein. In some embodiments, the additional therapeutic agent is administered after the administration of the compound disclosed herein. EXAMPLES [00140] The compounds described herein are synthesized as generally described in general scheme 1 and general scheme 2. General Scheme 1 General Scheme 2 Example 1
Step 1: Preparation of 3-bromo-2-methoxy-6-methyl-5-nitropyridine: [00141] To a stirred mixture of 3-bromo-2-chloro-6-methyl-5-nitropyridine (20.00 g, 79.54 mmol, 1.00 equiv) in MeOH (50 mL) was added NaOMe (15.76 g, 87.49 mmol, 1.10 equiv, 30%wt) dropwise at 0°C under nitrogen atmosphere. The resulting mixture was stirred overnight at rt under nitrogen atmosphere. The reaction was monitored by TLC (Pet. Ether : EtOAc=1:1, R f = 0.4). Upon completion, the reaction was concentrated under reduced pressure and water (100 mL) was added. The resulting mixture was extracted with EtOAc (3x100 mL). The combined organic layers were washed with brine (1x200 mL), dried over anhydrous Na 2 SO 4 , filtered and concentrated to afford 3-bromo-2-methoxy-6-methyl-5-nitropyridine (20 g, 99%). 1 H NMR (400 MHz, DMSO-d 6 ) δ 8.66 (s, 1H), 4.04 (s, 3H), 2.70(s, 3H). Step 2: Preparation of (E)-2-(5-bromo-6-methoxy-3-nitropyridin-2-yl)-N,N-dimethylet hen-1-amine: [00142] A mixture of 3-bromo-2-methoxy-6-methyl-5-nitropyridine (15.00 g, 60.72 mmol, 1.00 equiv) in DMF-DMA (100 mL) and DMF (100 mL) was stirred overnight at 100°C under nitrogen atmosphere. The reaction was monitored by TLC (Pet. Ether : EtOAc =1:1, R f = 0.5). The mixture was allowed to cool down to rt and then concentrated under reduced pressure to afford crude (E)-2-(5-bromo-6-methoxy-3- nitropyridin-2-yl)-N,N-dimethylethen-1-amine. The crude product was used in the next step directly without further purification. Step 3: Preparation of 5-bromo-6-methoxy-3-nitropicolinaldehyde: [00143] To a stirred mixture of (E)-2-(5-bromo-6-methoxy-3-nitropyridin-2-yl)ethenyl]dimethy lamine (18.01 g, crude) in THF (100 mL) and H2O (100 mL) was added NaIO4 (28.00 g, 131.07 mmol, 2.20 equiv) in portions at 0°C under nitrogen atmosphere. The resulting mixture was stirred for 2h at rt under nitrogen atmosphere. The reaction was monitored by TLC (Pet. Ether : EtOAc = 5:1, Rf = 0.2). The reaction was quenched by the addition of sat. sodium hyposulfite (aq.) (100 mL) at rt. The resulting mixture was extracted with EtOAc (3x100 mL). The combined organic layers were washed with brine (2x100 mL), dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The crude product was used in the next step directly without further purification. 1 H NMR (400 MHz, DMSO-d 6 ) δ 10.16 (s, 1H), 8.87 (s, 1H), 4.10 (s, 3H). Step 4: Preparation of ethyl 7-bromo-6-methoxy-1,5-naphthyridine-3-carboxylate: [00144] To a stirred mixture of 5-bromo-6-methoxy-3-nitropyridine-2-carbaldehyde (7.00 g, crude) and ethyl 3,3-diethoxypropanoate (20.40 g, 107.27 mmol, 4.00 equiv) in EtOH (100 mL) were added SnCl2 (26.25 g, 134.09 mmol, 5.00 equiv) in portions at rt under nitrogen atmosphere. The resulting mixture was stirred overnight at 90°C under nitrogen atmosphere. The reaction was monitored by LCMS. The mixture was allowed to cool down to rt. The resulting mixture was concentrated under reduced pressure. The crude mixture was poured into saturated sodium bicarbonate (100 mL). The resulting mixture was extracted with EtOAc (3x100 mL). The combined organic layers were concentrated under reduced pressure and the residue was purified by silica gel column chromatography to afford the crude product. The crude product was further purified by trituration with hexane (50 mL) to afford ethyl 7-bromo-6-methoxy-1,5-naphthyridine-3- carboxylate (3.50 g, 18.5%, over three steps). LC-MS: (ES+H, m/z): [M+H] + =311.0 Step 5: Preparation of ethyl 7-bromo-6-oxo-5H-1,5-naphthyridine-3-carboxylate: [00145] A solution of ethyl 7-bromo-6-methoxy-1,5-naphthyridine-3-carboxylate (5.00 g, 16.07 mmol, 1.00 equiv) in ACN (400 mL) were added TMSI (13.8 mL, 96.42 mmol, 6.00 equiv) dropwise at rt. The final reaction mixture was stirred for 2h at 80°C under nitrogen atmosphere. The mixture was allowed to cool down to rt. The reaction was monitored by LCMS. The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford ethyl 7-bromo-6- oxo-5H-1,5-naphthyridine-3-carboxylate (3.8 g, 80%). LC-MS: (ES+H, m/z): [M+H] + =296.95. Step 6: Preparation of ethyl 7-[2-(tert-butyldimethylsilyl)ethynyl]-6-oxo-5H-1,5-naphthyr idine-3- carboxylate: [00146] A mixture of ethyl 7-bromo-6-oxo-5H-1,5-naphthyridine-3-carboxylate (1.50 g, 5.05 mmol, 1.00 equiv), tert-butyl(ethynyl)dimethylsilane (850 mg, 6.05 mmol, 1.20 equiv), CuI (0.19 g, 1.01 mmol, 0.20 equiv), Et 3 N (1.53 g, 15.15 mmol, 3 equiv) and Pd(PPh 3 ) 2 Cl 2 (0.35 g, 0.51 mmol, 0.10 equiv) in DMF (45 mL) was stirred for 2h at 50°C under nitrogen atmosphere. The mixture was allowed to cool down to rt. The reaction was monitored by LCMS. The resulting mixture was diluted with water (450 mL). The resulting mixture was extracted with EtOAc (3 x 450 mL). The combined organic layers were washed with brine (3x 450 mL), dried over anhydrous Na 2 SO 4 . After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford ethyl 7-[2-(tert- butyldimethylsilyl)ethynyl]-6-oxo-5H-1,5-naphthyridine-3-car boxylate (1.5 g, 83%). LC-MS: (ES+H, m/z): [M+H] + =357.2. Step 7: Preparation of 3-[2-(tert-butyldimethylsilyl)ethynyl]-7-(hydroxymethyl)-1H- 1,5-naphthyridin- 2-one: [00147] To a stirred mixture of ethyl 7-[2-(tert-butyldimethylsilyl)ethynyl]-6-oxo-5H-1,5-naphthyr idine-3- carboxylate (1.20 g, 3.37 mmol, 1.00 equiv) in THF (30 mL) was added LiEt3BH (13.46 mL, 13.46 mmol, 4.00 equiv, 1M in THF) dropwise at 0°C under nitrogen atmosphere. The resulting mixture was stirred for 1h at 0°C under nitrogen atmosphere. The reaction was monitored by LCMS. The reaction was quenched by the addition of citric acid (5.05 mL, 5.05 mmol, 1.50 equiv, 1 M) at 0°C. The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford 3-[2-(tert-butyldimethylsilyl)ethynyl]-7-(hydroxymethyl)-1H- 1,5-naphthyridin-2-one (840 mg, 79%). LC-MS: (ES+H, m/z): [M+H] + =315.0. Step 8: Preparation of 3-[2-(tert-butyldimethylsilyl)ethynyl]-7-(chloromethyl)-1H-1 ,5-naphthyridin-2- one: [00148] To a stirred mixture of 3-[2-(tert-butyldimethylsilyl)ethynyl]-7-(hydroxymethyl)-1H- 1,5- naphthyridin-2-one (92 mg, 0.29 mmol, 1.00 equiv) and DMF (1 mg, 0.01 mmol, 0.05 equiv) in DCM (5 mL) was added SOCl2 (0.06 mL, 0.88 mmol, 3.00 equiv, 1.64g/mL) dropwise at 0°C under nitrogen atmosphere. The resulting mixture was stirred overnight at rt under nitrogen atmosphere. The reaction was monitored by LCMS. The resulting mixture was concentrated under reduced pressure to afford 3-[2-(tert- butyldimethylsilyl)ethynyl]-7-(chloromethyl)-1H-1,5-naphthyr idin-2-one (97 mg, crude). The crude product was used in the next step directly without further purification. LC-MS: (ES+H, m/z): [M+H+MeCN] + = 374.2. Step 9: Preparation of 5-[4-({7-[2-(tert-butyldimethylsilyl)ethynyl]-6-oxo-5H-1,5-n aphthyridin-3- yl}methyl)piperazin-1-yl]pyridine-2-carbonitrile: [00149] To a stirred mixture of 5-(piperazin-1-yl)pyridine-2-carbonitrile hydrochloride (119 mg, 0.53 mmol, 1.48 equiv), DIEA (232 mg, 1.80 mmol, 5.00 equiv) and 3-[2-(tert-butyldimethylsilyl)ethynyl]-7- (chloromethyl)-1H-1,5-naphthyridin-2-one (120 mg, 0.36 mmol, 1.00 equiv) in MeCN (5 mL) was added KI (11 mg, 0.07 mmol, 0.20 equiv) portion wise at rt under nitrogen atmosphere. The resulting mixture was stirred for 2h at 80°C under nitrogen atmosphere. The reaction was monitored by LCMS. The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford 5-[4-({7-[2-(tert-butyldimethylsilyl)ethynyl]-6-oxo-5H-1,5-n aphthyridin-3- yl}methyl)piperazin-1-yl]pyridine-2-carbonitrile (120 mg, 69%). Step 10: Preparation of 5-{4-[(7-ethynyl-6-oxo-5H-1,5-naphthyridin-3-yl)methyl]piper azin-1- yl}pyridine-2-carbonitrile: [00150] To a stirred mixture of 5-[4-({7-[2-(tert-butyldimethylsilyl)ethynyl]-6-oxo-5H-1,5-n aphthyridin-3- yl}methyl)piperazin-1-yl]pyridine-2-carbonitrile (120 mg, 0.24 mmol, 1.00 equiv) in THF (2.5 mL) was added TBAF (0.27 mL, 0.27 mmol, 1.10 equiv) dropwise at rt under nitrogen atmosphere. The resulting mixture was stirred for 1h at rt under nitrogen atmosphere. The reaction was monitored by LCMS. The resulting mixture was concentrated under reduced pressure. The resulting mixture was diluted with water (50 mL). The resulting mixture was extracted with EtOAc (3 x 25mL). The combined organic layers were washed with brine (1x50 mL), dried over anhydrous Na 2 SO 4 . After filtration, the filtrate was concentrated under reduced pressure. The crude product (120 mg) was purified by prep HPLC to afford 5-{4-[(7-ethynyl- 6-oxo-5H-1,5-naphthyridin-3-yl)methyl]piperazin-1-yl}pyridin e-2-carbonitrile (31.3 mg, 34%). LC-MS: (ES-H, m/z): [M-H]- =369.1; 1 H NMR (300 MHz, DMSO-d6) δ 12.17 (s, 1H), 8.48 (d, 1H), 8.42 (d, 1H), 8.15 (s, 1H), 7.75 (d, 1H), 7.64 (d, 1H), 7.36 (dd, 1H), 4.51 (s, 1H), 3.67 (s, 2H), 3.47 - 3.38 (m, 4H), 2.60 - 2.52 (m, 4H). Example 2 Step 1: Preparation of methyl 5-nitro-6-(prop-1-en-2-yl)pyridine-3-carboxylate: [00151] A mixture of methyl 6-chloro-5-nitropyridine-3-carboxylate (10.00 g, 46.17 mmol, 1.00 equiv.), 4,4,5,5-tetramethyl-2-(prop-1-en-2-yl)-1,3,2-dioxaborolane (15.52 g, 92.34 mmol, 2.00 equiv.), K2CO3 (12.76 g, 92.34 mmol, 2.00 equiv.) and Pd(dppf)Cl2 (3.38 g, 4.62 mmol, 0.10 equiv.) in dioxane (150 mL) and water (15 mL) was stirred for 3h at 100°C under nitrogen atmosphere. The reaction was monitored by LCMS. The mixture was allowed to room temperature. The resulting mixture was diluted with water (300 mL) and extracted with EtOAc (3x300 mL). The combined organic layers were washed with sat. NaCl (aq.) (3x100 mL), dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure. The filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with PE / EA (1:1) to afford methyl 5-nitro-6-(prop-1-en-2-yl)pyridine-3-carboxylate (5.00 g, 48.74%) as a light yellow oil. LC-MS: (ES+H, m/z): [M+H] + =222.95; 1 H NMR (300 MHz, DMSO-d6) δ 9.25 (d, 1H), 8.74 (d, 1H), 5.41 – 5.47 (m, 1H), 5.13 – 5.21 (m, 1H), 3.94 (s, 3H), 2.16 (dd, 3H). Step 2: Preparation of methyl 5-nitro-6-(prop-1-en-2-yl)pyridine-3-carboxylate: [00152] To a stirred solution of methyl 5-nitro-6-(prop-1-en-2-yl)pyridine-3-carboxylate (5.00 g, 22.50 mmol, 1.00 equiv.) in MeOH (100 mL) were added NH 4 Cl (25 mL, sat. aq.) and Fe (5.03 g, 90.01 mmol, 4.00 equiv.). The reaction was stirred at 80°C for 4h under nitrogen atmosphere. The reaction was monitored by LCMS. The mixture was allowed to room temperature then concentrated under reduced pressure. The residue was diluted with CH 2 Cl 2 /2-Propanol (5:1, 200 mL) and washed with water (250 mL) and brine (250 mL). The organic layers were dried over anhydrous Na 2 SO 4 , filtered and concentrated under reduced pressure. This resulted in methyl 5-amino-6-(prop-1-en-2-yl)pyridine-3-carboxylate (3.90 g, 90.17%) which was used directly without further purification. LC-MS: (ES+H, m/z): [M+H] + =193.15; 1 H NMR (300 MHz, DMSO-d 6 ) δ 8.29 (d, 1H), 7.60 (d, 1H), 5.51-5.47 (m, 1H), 5.41 (s, 2H), 5.39 – 5.36 (m, 1H), 3.84 (s, 3H), 2.08 (t, 3H). Step 3: Preparation of methyl 8-methyl-6-oxo-5H-1,5-naphthyridine-3-carboxylate: [00153] A solution of triphosgene (1.54 g, 5.20 mmol, 0.50 equiv.) in toluene (20 mL) was added to the solution of methyl 5-amino-6-(prop-1-en-2-yl)pyridine-3-carboxylate (3.90 g, 20.29 mmol, 1.00 equiv.) and Et3N (6.16 g, 60.87 mmol, 3.00 equiv.) in toluene (40 mL) at 0°C under nitrogen atmosphere. The resulting mixture was stirred overnight at 60°C under nitrogen. The reaction was monitored by LCMS. The reaction was quenched with MeOH (30 mL) at 0°C. The resulting mixture was diluted with water (200 mL) and extracted with CH2Cl2/2-Propanol (5:1, 3x200 mL). The combined organic layers were washed with water (3x100 mL), dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford methyl 8-methyl-6-oxo-5H-1,5-naphthyridine-3- carboxylate (1.80 g, 40.66%). LC-MS: (ES+H, m/z): [M+H] + =219.1; 1 H NMR (400 MHz, DMSO-d6) δ 11.92 (s, 1H), 8.92 (d, 1H), 8.15 (d, 1H), 6.79 (s, 1H), 3.93 (s, 3H), 2.48 (s, 3H). Step 4: Preparation of methyl 7-chloro-8-methyl-6-oxo-5H-1,5-naphthyridine-3-carboxylate: [00154] To a solution of methyl 8-methyl-6-oxo-5H-1,5-naphthyridine-3-carboxylate (600 mg, 2.75 mmol, 1.00 equiv.) and NCS (587 mg, 4.40 mmol, 1.60 equiv.) in CH3COOH (7 mL) was added 2,2-dichloroacetic acid (71 mg, 0.55 mmol, 0.20 equiv.) dropwise at room temperature under nitrogen atmosphere. The resulting mixture was stirred overnight at 100°C under nitrogen atmosphere. The reaction was monitored by LCMS. The mixture was allowed to cool down to room temperature. The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford methyl 7- chloro-8-methyl-6-oxo-5H-1,5-naphthyridine-3-carboxylate (650 mg, 93.5%). LC-MS: (ES+H, m/z): [M+H] + =253.0; 1 H NMR (300 MHz, DMSO-d 6 ) δ 8.90 (d, 1H), 8.13 (d, 1H), 3.93 (s, 3H), 2.60 (s, 3H). Step 5: Preparation of 3-chloro-7-(hydroxymethyl)-4-methyl-1H-1,5-naphthyridin-2-on e: [00155] To a stirred solution of methyl 7-chloro-8-methyl-6-oxo-5H-1,5-naphthyridine-3-carboxylate (600 mg, 2.38 mmol, 1.00 equiv.) in THF (5 mL) was added LiAlH 4 (2 mL, 2.5 M in THF, 4.75 mmol, 2.00 equiv.) dropwise at 0°C under nitrogen atmosphere. The resulting mixture was stirred for 1 h at room temperature under nitrogen atmosphere. The reaction was monitored by LCMS. The reaction was quenched by the addition of HCl (1 mL, 12M) at 0°C. The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford 3-chloro-7-(hydroxymethyl)-4- methyl-1H-1,5-naphthyridin-2-one (230 mg, 43.1%). LC-MS: (ES+H, m/z): [M+H] + =225.1; 1 H NMR (300 MHz, DMSO-d 6 ) δ 12.24 (br, 1H), 8.48 (d, 1H), 7.77 (d, 1H), 5.69 (s, 1H), 4.63 (s, 2H), 2.63 (s, 3H). Step 6: Preparation of 3-chloro-7-(chloromethyl)-4-methyl-1H-1,5-naphthyridin-2-one : [00156] To a stirred solution of 3-chloro-7-(hydroxymethyl)-4-methyl-1H-1,5-naphthyridin-2-on e (200 mg, 0.89 mmol, 1.00 equiv.) and DMF (7 mg, 0.09 mmol, 0.10 equiv.) in DCM (10 mL) were added SOCl2 (318 mg, 2.67 mmol, 3.00 equiv.) dropwise at 0°C under nitrogen atmosphere. The resulting mixture was stirred for 10 h at room temperature. The reaction was monitored by LCMS. The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford 3-chloro-7-(chloromethyl)-4-methyl-1H-1,5-naphthyridin-2-one (98 mg, 45.2%). LC-MS: (ES+H, m/z): [M+H] + =243.0. Step 7: Preparation of 4-{4-[(7-chloro-8-methyl-6-oxo-5H-1,5-naphthyridin-3-yl)meth yl]piperazin-1- yl}benzonitrile: [00157] A solution of 3-chloro-7-(chloromethyl)-4-methyl-1H-1,5-naphthyridin-2-one (25 mg, 0.10 mmol, 1.00 equiv.), DIEA (66 mg, 0.51 mmol, 5.00 equiv.), KI (4 mg, 0.02 mmol, 0.20 equiv.) and 4-(piperazin-1- yl)benzonitrile (19 mg, 0.10 mmol, 1.00 equiv.) in MeCN (2 mL) was stirred for 2h at 80°C under nitrogen atmosphere. The mixture was allowed to cool down to room temperature. The reaction was monitored by LCMS. The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography. The resulting mixture was concentrated under reduced pressure. The pure fraction was concentrated then lyophilized to afford 4-{4-[(7-chloro-8-methyl-6-oxo-5H-1,5-naphthyridin-3- yl)methyl]piperazin-1-yl}benzonitrile (12 mg, 29.6%). LC-MS: (ES-H, m/z): [M-H]-=392.15; 1 H NMR (400 MHz, DMSO-d6) δ 12.30 (s, 1H), 8.51 (d, J = 1.8 Hz, 1H), 7.69 (d, J = 1.9 Hz, 1H), 7.58 (d, J = 8.7 Hz, 2H), 7.02 (d, J = 8.7 Hz, 2H), 3.67 (s, 2H), 3.38 – 3.34(m, 4H), 2.64 (s, 3H), 2.57-2.54 (m, 4H). Example 3 Step 1: Preparation of methyl 6-methyl-5-nitropyridine-3-carboxylate: [00158] To a stirred mixture of methyl 6-chloro-5-nitropyridine-3-carboxylate (5 g, 273.08 mmol, 1.00 equiv) and methylboronic acid (2.75 g, 46.17 mmol, 2.00 equiv) in Toluene (60 mL) and H 2 O (3 mL) were added Pd(dppf)Cl 2 (0.83 g, 1.15 mmol, 0.05 equiv) and CsF (7 g, 46.17 mmol, 2.00 equiv) at rt under nitrogen atmosphere. The resulting mixture was stirred overnight at 100 o C under nitrogen atmosphere. The resulting mixture was added to 500 mL water and extracted with EtOAc (3 x 200 mL). The combined organic layers were washed with brine (200 mL), dried over anhydrous Na2SO4, filtered and concentrated. The crude product was purified by silica gel column chromatography to afford methyl 6-methyl-5- nitropyridine-3-carboxylate (2.9 g, 64%). LC-MS: (ES+H, m/z): [M+H] + =197.0; 1H NMR (400 MHz, Chloroform-d) δ 9.29 (d, 1H), 8.84 (d, 1H), 4.01 (s, 3H), 2.94 (s, 3H). Step 2: Preparation of methyl 6-formyl-5-nitropyridine-3-carboxylate: [00159] A mixture of methyl 6-methyl-5-nitropyridine-3-carboxylate (35 g, 178.42 mmol, 1.00 equiv) and SeO2 (30 g, 267.64 mmol, 1.50 equiv) in dioxane (200 mL) was stirred overnight at 110°C under nitrogen atmosphere. The resulting mixture was then filtered and the filter cake was washed with EtOAc (5x200 mL). The filtrate was concentrated under reduced pressure and then the crude product was dissolved in THF (200 mL). The resulting mixture was filtered and the filter cake was washed with THF (3x100 mL). The filtrate was concentrated under reduced pressure to afford nitropyridine-3-carboxylate (40 g, crude). LC-MS: (ES+H, m/z): [M+H] + =211.1. Step 3: Preparation of methyl (Z)-6-(2-(ethoxycarbonyl)but-1-en-1-yl)-5-nitronicotinate: [00160] To a stirred solution of NaH (11.42g, 285.52 mmol, 1.50 equiv, 60% wt) in THF (500 mL) was added ethyl 2-(diethoxyphosphoryl)butanoate (72 g, 285.52 mmol, 1.50 equiv) in THF (50 mL) dropwise at 0°C under nitrogen atmosphere. The resulting mixture was stirred for 10 min at 0°C and then warmed to 40°C and stirred for 10 min under nitrogen atmosphere. The resulting mixture was cooled to -78°C and then methyl 6-formyl-5-nitropyridine-3-carboxylate (40 g, 190.35 mmol, 1.00 equiv) in THF (50 mL) was added dropwise. The resulting mixture was stirred for 30 min at -78°C under nitrogen atmosphere. The reaction was monitored by LCMS. The reaction was quenched by the addition of sat. NH 4 Cl (aq.) (100 mL) at 0°C. The resulting mixture was added 400 mL water and extracted with EtOAc (3 x 500 mL). The combined organic layers were washed with brine (1x500 mL), dried over anhydrous Na 2 SO 4 , filtered and concentrated. The residue was purified by silica gel column chromatography to afford methyl (Z)-6-(2- (ethoxycarbonyl)but-1-en-1-yl)-5-nitronicotinate (20 g, 36%, two steps) and methyl (E)-6-(2- (ethoxycarbonyl)but-1-en-1-yl)-5-nitronicotinate (8.8 g). Data for methyl (Z)-6-(2-(ethoxycarbonyl)but-1- en-1-yl)-5-nitronicotinate: LC-MS: (ES+H, m/z): [M+H] + =309.1; 1H NMR (300 MHz, Chloroform-d) δ 9.27 (d, 1H), 8.88 (d, 1H), 7.10 (t, 1H), 4.22 – 4.16 (m, 2H), 4.03 (s, 3H), 2.59 (qd, 2H), 1.25 (t, 3H), 1.19 (t, 3H). Data for methyl (E)-6-(2-(ethoxycarbonyl)but-1-en-1-yl)-5-nitronicotinate: LC-MS: (ES+H, m/z): [M+H] + =309.1; 1 H NMR (300 MHz, Chloroform-d) δ 9.45 (d, 1H), 8.88 (d, 1H), 7.87 (s, 1H), 4.34 (q, 2H), 4.05 (s, 3H), 2.67 (q, 2H), 1.39 (t, 3H), 1.15 (t, 3H). Step 4: Preparation of ethyl 7-ethyl-6-oxo-5H-1,5-naphthyridine-3-carboxylate: [00161] To a stirred mixture of methyl 6-[(1Z)-3-ethoxy-2-ethyl-3-oxoprop-1-en-1-yl]-5-nitropyridin e-3- carboxylate (9.00 g, 29.19 mmol, 1.00 equiv) and Fe (16.30 g, 291.93 mmol, 10.00 equiv) in EtOH (200 mL) was added CaCl 2 (19.44 g, 175.16 mmol, 6.00 equiv) at rt under nitrogen atmosphere. The resulting mixture was stirred overnight at 90°C under nitrogen atmosphere. The reaction was monitored by LCMS. The resulting mixture was filtered and the filter cake was washed with EtOAc (5x100 mL) and the combined filtrate was concentrated. The resulting mixture was added to 250 mL water and extracted with EtOAc (3x250 mL). The combined organic layers were washed with brine (1x250 mL), dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure to afford ethyl 7-ethyl-6-oxo- 5H-1,5-naphthyridine-3-carboxylate (5.2 g, 72%). LC-MS: (ES+H, m/z): [M+H] + =247.1; 1H NMR (400 MHz, DMSO-d6) δ 12.06 (s, 1H), 8.90 (s, 1H), 8.16 (s, 1H), 7.83 (s, 1H), 4.38 (q, 2H), 2.58 (q, 2H), 1.36 (t, 3H), 1.20 (t, 3H). Step 5: Preparation of 3-ethyl-7-(hydroxymethyl)-1H-1,5-naphthyridin-2-one: [00162] To a stirred solution of ethyl 7-ethyl-6-oxo-5H-1,5-naphthyridine-3-carboxylate (4.00 g, 16.24 mmol, 1.00 equiv) in THF (50 mL) was added LiAlH4 (13 mL, 32.49 mmol, 2.00 equiv, 2.5M in THF) dropwise at 0°C under nitrogen atmosphere. The resulting mixture was stirred for 1 h at 0°C. The reaction was monitored by LCMS. The reaction was quenched by the addition of 1M aq HCl (16 mL) at 0°C.The resulting mixture was concentrated under reduced pressure and the crude product was purified by silica gel column chromatography to afford 3-ethyl-7-(hydroxymethyl)-1H-1,5-naphthyridin-2-one (2.00 g, 60%). LC- MS: (ES+H, m/z): [M+H] + =204.8; 1H NMR (400 MHz, DMSO-d6) δ 11.91 (s, 1H), 8.38 (d, 1H), 7.74 (s, 1H), 7.62 (d, 1H), 5.49 (t, 1H), 4.62 (d, 2H), 2.55 (dd, 2H), 1.18 (t, 3H). Step 6: Preparation of 7-(chloromethyl)-3-ethyl-1H-1,5-naphthyridin-2-one: [00163] To a stirred mixture of 3-ethyl-7-(hydroxymethyl)-1H-1,5-naphthyridin-2-one (300 mg, 1.47 mmol, 1.00 equiv) and DMF (11 mg, 0.15 mmol, 0.10 equiv) in DCM (10 mL) was added SOCl 2 (1.05 g, 8.81 mmol, 6.00 equiv) dropwise at 0°C under nitrogen atmosphere. The resulting mixture was stirred overnight at rt and then concentrated under vacuum. The residue was purified by silica gel column chromatography to afford 7-(chloromethyl)-3-ethyl-1H-1,5-naphthyridin-2-one (300 mg, 92%). LC-MS: (ES+H, m/z): [M+H] + =222.8; 1H NMR (400 MHz, DMSO-d 6 ) δ 11.99 (s, 1H), 8.50 (d, 1H), 7.76 (s, 1H), 7.70 (d, 1H), 4.93 (s, 2H), 2.56 (td, 2H), 1.19 (t, 3H). Step 7: Preparation of 5-{4-[(7-ethyl-6-oxo-5H-1,5-naphthyridin-3-yl)methyl]piperaz in-1-yl}pyridine- 2-carbonitrile: [00164] To a stirred mixture of 5-(piperazin-1-yl)pyridine-2-carbonitrile, HCl salt (600 mg) and 7- (chloromethyl)-3-ethyl-1H-1,5-naphthyridin-2-one (500 mg, 2.25 mmol, 1.00 equiv) in ACN (4 mL) were added DIEA (1.45 g, 11.25 mmol, 5.00 equiv) and KI (75 mg, 0.45 mmol, 0.20 equiv) in portions at rt. The resulting mixture was stirred for 1h at 80°C under nitrogen atmosphere. The reaction was monitored by LCMS. The mixture was allowed to cool down to rt. The reaction was poured into water (20 mL). The aqueous layer was extracted with EtOAc (3x20 mL). The combined organic layers were concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford 5-{4-[(7-ethyl-6- oxo-5H-1,5-naphthyridin-3-yl)methyl]piperazin-1-yl}pyridine- 2-carbonitrile (650 mg, 77%). LC-MS: (ES+H, m/z): [M+H] + = 375.2; 1 H NMR (300 MHz, DMSO-d 6 ) δ 11.86 (s, 1H), 8.41-8.35 (m, 2H), 7.75 (t, 2H), 7.61 (s, 1H), 7.36 (dd, 1H), 3.64 (s, 2H), 3.52-3.42 (m, 4H), 2.61-2.54 (m, 6H), 1.18 (t, 3H). Example 5 Step 1: Preparation of methyl 2-[(4-bromo-2-fluoro-6-nitrophenyl)amino]butanoate: [00165] To a stirred mixture of 5-bromo-1,2-difluoro-3-nitrobenzene (10.00 g, 42.02 mmol, 1.00 equiv) and Methyl 2-aminobutanoate hydrochloride (6.43 g, 42.02 mmol, 1.00 equiv) in NMP (200 mL) was added DIEA (27.15 g, 210.10 mmol, 5.00 equiv) dropwise at rt. The resulting mixture was diluted with water (500 mL). The resulting mixture was extracted with EtOAc (3 x 500 mL). The combined organic layers were washed with brine (8x300 mL), dried over anhydrous Na 2 SO 4 . After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, the pure fraction was concentrated under vacuum to afford methyl 2-[(4-bromo-2-fluoro-6-nitrophenyl)amino]butanoate (13.00 g, 92%). LC-MS: (ES+H, m/z): [M+H] + = 334.9; 1 H NMR (400 MHz, DMSO-d6) δ 8.09 (t, 1H), 7.88 (dd, 1H), 7.79 (dd, 1H), 4.56 (dtd, 1H), 3.69 (s, 3H), 1.97 – 1.79 (m, 2H), 0.91 (t, 3H). Step 2: Preparation of 7-bromo-3-ethyl-5-fluoro-3,4-dihydro-1H-quinoxalin-2-one: [00166] To a stirred mixture of Fe (5.00 g, 89.52 mmol, 5.00 equiv) in AcOH (100 mL) was added methyl 2-[(4-bromo-2-fluoro-6-nitrophenyl)amino]butanoate (6.00 g, 17.90 mmol, 1.00 equiv) in AcOH (20 mL) dropwise at 80°C under nitrogen atmosphere. The resulting mixture was stirred for 2 h at 80°C under nitrogen atmosphere. The reaction was monitored by TLC (CH2Cl2/MeOH=10/1). The mixture was allowed to cool down to rt. The resulting mixture was filtered, the filter cake was washed with EtOAc (3x50 mL). The filtrate was concentrated under reduced pressure. The residue was purified by trituration with water (3 x 100 mL). The precipitated solids were collected by filtration and washed with water (3x20 mL). This resulted in 7-bromo-3-ethyl-5-fluoro-3,4-dihydro-1H-quinoxalin-2-one (4.5 g, 92%). 1 H NMR (400 MHz, DMSO-d6) δ 10.52 (s, 1H), 6.99 (d, 1H), 6.72 (s, 1H), 6.17 (s, 1H), 3.78 (d, 1H), 1.72 – 1.62 (m, 2H), 0.88 (t, 3H). Step 3: Preparation of 7-bromo-3-ethyl-5-fluoro-1H-quinoxalin-2-one: [00167] To a stirred solution of 7-bromo-3-ethyl-5-fluoro-3,4-dihydro-1H-quinoxalin-2-one (4.40 g, 16.11 mmol, 1.00 equiv) in DCM (100 mL) was added DDQ (4.39 g, 19.33 mmol, 1.20 equiv) in portions at rt under nitrogen atmosphere. The resulting mixture was stirred for 2 h at rt under nitrogen atmosphere. The reaction was monitored by LCMS. The reaction was quenched by the addition of sat. NaHCO3 (aq.) (100 mL) at 0 °C. The residue was purified by trituration with sat. NaHCO3 (aq.) (5 x 100 mL). The precipitated solids were collected by filtration and washed with water (3x100 mL) to afford 7-bromo-3-ethyl-5-fluoro- 1H-quinoxalin-2-one (3.50 g, 80%). LC-MS: (ES-H, m/z): [M-H]- = 269.0; 1 H NMR (400 MHz, DMSO-d6) δ 12.51 (s, 1H), 7.44 (d, 1H), 7.24 (s, 1H), 2.79 (q, 2H), 1.21 (t, 3H). Step 4: Preparation of 2-ethyl-8-fluoro-3-oxo-4H-quinoxaline-6-carbaldehyde: [00168] To a solution of 7-bromo-3-ethyl-5-fluoro-1H-quinoxalin-2-one (2.90 g, 10.70 mmol, 1.00 equiv) in Toluene (100 mL) was added TMEDA (1.49 g, 12.84 mmol, 1.20 equiv), bis(adamantan-1- yl)(butyl)phosphane (0.77 g, 2.14 mmol, 0.20 equiv), Pd(OAc)2 (0.24 g, 1.07 mmol, 0.10 equiv) in a pressure tank. The mixture was purged with nitrogen for 5 min and then was pressurized to 30 atm with (CO:H2=1:1). The mixture was stirred overnight at 100°C. The reaction was monitored by LCMS. The reaction mixture was cooled to rt and filtered to remove insoluble solids. The resulting mixture was concentrated under vacuum. The residue was purified by silica gel column chromatography, the pure fraction was concentrated under vacuum to afford 2-ethyl-8-fluoro-3-oxo-4H-quinoxaline-6-carbaldehyde (1.40 g, 59%). LC-MS: (ES-H, m/z): [M-H]- = 219.0; 1 H NMR (300 MHz, DMSO-d6) δ 12.74 (s, 1H), 10.01 (d, 1H), 7.65 – 7.47 (m, 2H), 2.86 (q, 2H), 1.24 (t, 3H). Step 5: Preparation of 6-(4-((2-ethyl-8-fluoro-3-oxo-3,4-dihydroquinoxalin-6-yl)met hyl)piperazin-1- yl)nicotinonitrile: [00169] To a stirred solution of 6-(piperazin-1-yl)pyridine-3-carbonitrile (200 mg, 1.06 mmol, 1.00 equiv) and 2-ethyl-8-fluoro-3-oxo-3,4-dihydroquinoxaline-6-carbaldehyde (304 mg, 1.38 mmol, 1.30 equiv) in THF (10 mL) was added Ti(OiPr) 4 (453 mg, 1.60 mmol, 1.50 equiv) at rt under nitrogen atmosphere. After 2 h, NaBH(AcO) 3 (901 mg, 4.25 mmol, 4.00 equiv) was added at 0°C and the reaction was warmed to rt. After 2 h, the reaction was quenched by the addition of water (10 mL) at 0°C. The aqueous layer was extracted with EtOAc (5x30 mL). The combined organic phase was concentrated under reduced pressure and the crude product was purified by prep HPLC to afford 6-(4-((2-ethyl-8-fluoro-3-oxo-3,4-dihydroquinoxalin-6- yl)methyl)piperazin-1-yl)nicotinonitrile (56.1 mg, 13 %). LC-MS: (ES+H, m/z): [M+H] + =393.2; 1H NMR (300 MHz, DMSO-d 6 ) δ 12.43 (s, 1H), 8.48 (d, 1H), 7.85 (dd, 1H), 7.16-7.04 (m, 2H), 6.93 (d, 1H), 3.77 – 3.52 (m, 6H), 2.87-2.75 (m, 2H), 2.50-2.43 (m, 4H), 1.22 (t, 3H); 19 F NMR (282 MHz, DMSO-d6) δ - 125.38. Example 12 Step1: Preparation of methyl 2-fluoro-4-[(1-methoxy-1-oxobutan-2-yl)amino]-5-nitrobenzoat e: [00170] To a stirred mixture of methyl 2,4-difluoro-5-nitrobenzoate (10.00 g, 46.06 mmol, 1.00 equiv) and methyl 2-aminobutanoate, HCl salt (5.40 g, 46.06 mmol, 1.00 equiv) in NMP (100 mL) was added DIEA (48.1 mL, 276.34 mmol, 6.00 equiv) dropwise at rt under nitrogen atmosphere. The resulting mixture was stirred for 4h at rt under nitrogen atmosphere. The reaction was monitored by LCMS. The reaction was quenched by the addition of water (200 mL) at rt. The resulting mixture was extracted with EtOAc (3 x100 mL). The combined organic layers were washed with brine (8x200 mL), dried over anhydrous Na 2 SO 4 , filtered and concentrated. The crude product was purified by silica gel column chromatography to afford methyl 2-fluoro-4-[(1-methoxy-1-oxobutan-2-yl)amino]-5-nitrobenzoat e (12 g, 83%). LC-MS: (ES+H, m/z): [M+H] + = 315.1; 1 H NMR (400 MHz, DMSO-d6) δ 8.80-8.56 (m, 2H), 7.21-6.99 (m, 1H), 4.84-4.74 (m, 1H), 3.85-3.81 (s, 3H), 3.77-3.74 (s, 3H), 2.05 – 1.82 (m, 2H), 0.91-0.85 (m, 3H). Step 2: Preparation of methyl 2-ethyl-7-fluoro-3-oxo-2,4-dihydro-1H-quinoxaline-6-carboxyl ate: [00171] To a stirred mixture of methyl 2-fluoro-4-[(1-methoxy-1-oxobutan-2-yl)amino]-5-nitrobenzoat e (12.00 g, 38.18 mmol, 1.00 equiv) in MeOH (180 mL) and EtOAc (30 mL) was added Pd(OH)2 /C (2.40 g) portion-wise at rt under a nitrogen atmosphere. The resulting mixture was stirred overnight at rt under hydrogen atmosphere. The reaction was monitored by LCMS. Upon completion, the resulting mixture was filtered and the filter cake was washed with EtOAc (3x300 mL). The filtrate was concentrated and purified by silica gel column chromatography, (Pet. Ether / EtOAc) to afford methyl 2-ethyl-7-fluoro-3-oxo-2,4- dihydro-1H-quinoxaline-6-carboxylate (8 g, 83%). LC-MS: (ES+H, m/z): [M+H] + = 253.0; 1 H NMR (400 MHz, DMSO-d6) δ 10.44 (s, 1H), 7.25-7.20 (m, 1H), 7.19-7.14 (s, 1H), 4.00-3.91 (m, 1H), 3.75 (s, 3H), 1.76 – 1.60 (m, 2H), 0.94-0.86 (m, 3H). Step 3: Preparation of methyl 2-ethyl-7-fluoro-3-oxo-4H-quinoxaline-6-carboxylate: [00172] To a stirred mixture of methyl 2-ethyl-7-fluoro-3-oxo-2,4-dihydro-1H-quinoxaline-6-carboxyl ate (7.00 g, 27.75 mmol, 1.00 equiv) in DCM (70 mL) at rt was added DDQ (7.56 g, 33.30 mmol, 1.20 equiv) in DCM (70 mL). The resulting mixture was stirred for 2 h at rt under a nitrogen atmosphere. The reaction was monitored by LCMS. The residue was then dissolved in sat. aq. NaHCO 3 (200 mL). The precipitated solids were collected by filtration and washed with sat. aq. NaHCO 3 (3x200 mL) and water (3x200 mL). The solids were purified by trituration with EtOAc (30 mL)/hexane (100 mL). The resulting mixture was filtered, and the filter cake was washed with hexane (3x100 mL). The filter cake was dried under reduced pressure to afford methyl 2-ethyl-7-fluoro-3-oxo-4H-quinoxaline-6-carboxylate (6 g, 86 %). LC-MS: (ES+H, m/z): [M+H] + =251.1; 1 H NMR (400 MHz, DMSO-d 6 ) δ 12.44 (s, 1H), 7.78 (d, 1H), 7.63 (d, 1H), 3.89 (s, 3H), 2.82 (q, 2H), 1.23 (t, 3H). Step 4: Preparation of 3-ethyl-6-fluoro-7-(hydroxymethyl)-1H-quinoxalin-2-one: [00173] To a stirred mixture of methyl 2-ethyl-7-fluoro-3-oxo-4H-quinoxaline-6-carboxylate (5.00 g, 19.98 mmol, 1.00 equiv) in THF (100 mL) was added LiAlH4 (40 mL, 39.96 mmol, 2.00 equiv, 1 moL/L in THF) dropwise at 0 o C under nitrogen atmosphere. The reaction was monitored by LCMS. The reaction was quenched by the addition of water (1.5 mL), 15% NaOH (1.5 mL) and water (4.5 ml) at 0 o C. The precipitated solids were collected by filtration and washed with water (3x30 mL). The solids were dissolved in water (500 mL) and EtOAc (200 mL). The resulting mixture was extracted with EtOAc (8 x 200 mL). The filtrate was concentrated under reduced pressure. The residue was purified by trituration with MeOH (5 mL) and EtOAc (20 ml). The resulting mixture was filtered and the filter cake was washed with EtOAc (3x10 mL). The solid was dried under reduced pressure to afford 3-ethyl-6-fluoro-7-(hydroxymethyl)-1H- quinoxalin-2-one (1.2 g, 27 %). LC-MS: (ES+H, m/z): [M+H] + =223.2; 1 H NMR (400 MHz, DMSO-d6) δ 12.38 (s, 1H), 7.48 (d, 1H), 7.41 (d, 1H), 5.52 (brs, 1H), 4.64 (s, 2H), 2.80 (q, 2H), 1.18 (t, 3H). Step 5: Preparation of 7-(bromomethyl)-3-ethyl-6-fluoro-1H-quinoxalin-2-one: [00174] To a stirred solution of 3-ethyl-6-fluoro-7-(hydroxymethyl)-1H-quinoxalin-2-one (150 mg, 0.68 mmol, 1.00 equiv) in HBr (4 mL, 33%wt in AcOH) at rt under nitrogen atmosphere. The resulting mixture was stirred for 1h at 80 o C under nitrogen atmosphere. The reaction was monitored by LCMS. The mixture was allowed to cool down to rt. The resulting mixture was concentrated under reduced pressure. The crude product was used in the next step directly without further purification. LC-MS: (ES+H, m/z): [M+H] + =285.0. Step 6: Preparation of 6-{4-[(2-ethyl-7-fluoro-3-oxo-4H-quinoxalin-6-yl)methyl]pipe razin-1- yl}pyridine-3-carbonitrile: [00175] A mixture of 7-(bromomethyl)-3-ethyl-6-fluoro-1H-quinoxalin-2-one (190 mg, 0.67 mmol, 1.00 equiv), 6-(piperazin-1-yl)pyridine-3-carbonitrile (150 mg, 0.80 mmol, 1.20 equiv) and DIEA (431 mg, 3.33 mmol, 5.00 equiv) in NMP (5 mL) was stirred for 1h at 80 o C under nitrogen atmosphere. The mixture was allowed to cool down to rt. The reaction was poured into Water (30 mL) and the aqueous layer was extracted with EtOAc (3x20 mL). The combined organic layers were concentrated under reduced pressure. The crude product (200 mg) was purified by prep HPLC to afford 6-{4-[(2-ethyl-7-fluoro-3-oxo-4H-quinoxalin-6- yl)methyl]piperazin-1-yl}pyridine-3-carbonitrile (100.2 mg, 38%). LC-MS: (ES+H, m/z): [M+H] + =393.1; 1 H NMR (300 MHz, DMSO-d 6 ) δ 12.31 (s, 1H), 8.48 (d, 1H), 7.85 (dd, 1H), 7.54 (d, 1H), 7.38 (d, 1H), 6.94 (d, 1H), 3.75-3.67 (m, 6H), 2.81 (q, 2H), 2.56-2.51 (m, 4H), 1.21 (t, 3H); 19 F NMR (282 MHz, DMSO-d6) δ -124.28. Example 16 Step 1: Preparation of ethyl 2-bromo-2-cyclopropylacetate: [00176] To a stirred solution of ethyl 2-cyclopropylacetate (10.00 g, 78.02 mmol, 1.00 equiv) in THF (100 mL) was added LDA (42.9 mL, 85.82 mmol, 1.10 equiv, 2.0 M in THF) dropwise at -78°C under nitrogen atmosphere. The reaction was stirred for 1 hour then TMSCl (8.48 g, 78.02 mmol, 1.00 equiv) added dropwise and the reaction was stirred for 3 hours as it was warmed to rt. The reaction was cooled to -78 °C and NBS (15.28 g, 85.82 mmol, 1.10 equiv) in 50 mL THF added dropwise. The reaction was then stirred for 2 hours and allowed to warm to rt. The reaction was monitored by LCMS. The reaction was quenched by the addition of sat. NH 4 Cl (aq.) (50 mL) at 0°C. The resulting mixture was extracted with Et 2 O (3 x 200 mL). The combined organic layers were washed with brine (3 x 200 mL), dried over anhydrous Na 2 SO 4 . After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by reversed phase combi-flash chromatography to afford ethyl 2-bromo-2-cyclopropylacetate (5.00 g, 31%). 1 H NMR (300 MHz, Chloroform-d) δ 4.25 (q, 2H), 3.58 (d, 1H), 1.65 – 1.55 (m, 1H), 1.31 (t, 3H), 0.92 – 0.76 (m, 2H), 0.61 – 0.53 (m, 1H), 0.48 – 0.40 (m, 1H). Step 2: Preparation of ethyl 2-cyclopropyl-2-(diethoxyphosphoryl)acetate: [00177] A solution of ethyl 2-bromo-2-cyclopropylacetate (5.00 g, 24.14 mmol, 1.00 equiv) and triethyl phosphite (5.22 g, 31.39 mmol, 1.30 equiv) was stirred for 24 h at 130 °C under a nitrogen atmosphere. The residue was purified by reversed phase combi-flash chromatography to afford ethyl 2-cyclopropyl-2- (diethoxyphosphoryl)acetate (2.40 g, 38%). 1 H NMR (300 MHz, Chloroform-d) δ 4.26 – 4.07 (m, 6H), 2.19 (dd, 1H), 1.30 (dt, 10H), 0.71 (dddd, 1H), 0.60 (ddddd, 1H), 0.47 – 0.37 (m, 1H), 0.24 (ddtd, 1H). Step 3: Preparation of methyl 6-[(1Z)-2-cyclopropyl-3-ethoxy-3-oxoprop-1-en-1-yl]-5-nitrop yridine-3- carboxylate: [00178] To a stirred mixture of NaH (0.29 g, 7.14 mmol, 1.50 equiv, 60%wt) in THF (20 mL) was added ethyl 2-cyclopropyl-2-(diethoxyphosphoryl)acetate (1.89 g, 7.14 mmol, 1.50 equiv) dropwise at 0°C under nitrogen atmosphere. The resulting mixture was stirred for 10 min at 0°C and then warmed to 40°C stirred for 10 min under nitrogen atmosphere. The resulting mixture was cooled to -78°C followed by the addition of methyl 6-formyl-5-nitropyridine-3-carboxylate (1.00 g, 4.76 mmol, 1.00 equiv) in THF (20 mL) dropwise. The resulting mixture was stirred for 30 min at -78°C under nitrogen atmosphere. The reaction was monitored by LCMS. The reaction was quenched by the addition of sat. NH 4 Cl (aq.) (5 mL) at 0°C. The resulting mixture was added 20 mL water and extracted with EtOAc (3 x 20 mL). The combined organic layers were washed with brine (1 x 50 mL), dried over anhydrous Na 2 SO 4 . After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford methyl 6-[(1Z)-2-cyclopropyl-3-ethoxy-3-oxoprop-1-en-1-yl]-5-nitrop yridine-3-carboxylate (700 mg, 46%). LC-MS: (ES+H, m/z): [M+H] + =320.8. Step 4: Preparation of ethyl 7-cyclopropyl-6-oxo-5H-1,5-naphthyridine-3-carboxylate: [00179] To a stirred mixture of methyl 6-[(1Z)-2-cyclopropyl-3-ethoxy-3-oxoprop-1-en-1-yl]-5- nitropyridine-3-carboxylate (600 mg, 1.87 mmol, 1.00 equiv) and Fe (1.04 g, 18.73 mmol, 10.00 equiv) in EtOH (10 mL) was added CaCl2 (1.24 g, 11.24 mmol, 6.00 equiv) at rt under nitrogen atmosphere. The resulting mixture was stirred overnight at 90°C under nitrogen atmosphere. The reaction was monitored by LCMS. The mixture was allowed to cool down to rt. The resulting mixture was filtered, the filter cake was washed with EtOAc (2 x 50 mL). The filtrate was concentrated under reduced pressure. The resulting mixture was added 50 mL water and extracted with EtOAc (2 x 50 mL). The combined organic layers were washed with brine (2 x 50 mL) and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford ethyl 7-cyclopropyl-6-oxo-5H-1,5-naphthyridine-3-carboxylate (200 mg, 41%). LC-MS: (ES+H, m/z): [M+H] + =259.0. Step 5: Preparation of 3-cyclopropyl-7-(hydroxymethyl)-1H-1,5-naphthyridin-2-one: [00180] To a stirred solution of ethyl 7-cyclopropyl-6-oxo-5H-1,5-naphthyridine-3-carboxylate (160 mg, 0.62 mmol, 1.00 equiv) was added LiAlH 4 (0.50 mL, 1.23 mmol, 2.00 equiv, 2.5M in THF) dropwise at 0°C under nitrogen atmosphere. The resulting mixture was stirred for 1 h at 0°C under nitrogen atmosphere. The reaction was monitored by LCMS. The reaction was quenched by the addition of 1M aq HCl (1 mL) at 0°C. The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford 3-cyclopropyl-7-(hydroxymethyl)-1H-1,5-naphthyridin-2-one (100 mg, 75%). LC-MS: (ES+H, m/z): [M+H] + =217.2. Step 6: Preparation of 7-(chloromethyl)-3-cyclopropyl-1H-1,5-naphthyridin-2-one: [00181] To a stirred mixture of 3-cyclopropyl-7-(hydroxymethyl)-1H-1,5-naphthyridin-2-one (80 mg, 0.37 mmol, 1.00 equiv) and DMF (3 mg, 0.04 mmol, 0.10 equiv) in DCM (10 mL) was added SOCl 2 (264 mg, 2.22 mmol, 6.00 equiv) dropwise at 0°C under nitrogen atmosphere. The resulting mixture was stirred overnight at rt under nitrogen atmosphere. The reaction was monitored by LCMS. The resulting mixture was concentrated under reduced pressure to afford 7-(chloromethyl)-3-cyclopropyl-1H-1,5-naphthyridin-2-one. The crude product was used in the next step directly without further purification. LC-MS: (ES+H, m/z): [M+H] + =235.0. Step 7: Preparation of 3-cyclopropyl-7-{[4-(5-fluoropyridin-2-yl)piperazin-1-yl]met hyl}-1H-1,5- naphthyridin-2-one: [00182] To a stirred solution of 1-(5-fluoropyridin-2-yl)piperazine (100 mg, 0.55 mmol, 1.00 equiv) and 7- (chloromethyl)-3-cyclopropyl-1H-1,5-naphthyridin-2-one (130 mg, 0.55 mmol, 1.00 equiv) in ACN (3 mL) was added DIEA (357 mg, 2.76 mmol, 5.00 equiv) and KI (18 mg, 0.11 mmol, 0.20 equiv) at rt. The resulting mixture was stirred for 1h at 80°C under nitrogen atmosphere. The reaction was monitored by LCMS. The mixture was allowed to cool down to rt and then poured into H2O (10mL), The resulting mixture was extracted with EtOAc (3 x 10mL). The combined organic layers were concentrated and the crude product was purified by prep HPLC to afford 3-cyclopropyl-7-{[4-(5-fluoropyridin-2-yl)piperazin-1- yl]methyl}-1H-1,5-naphthyridin-2-one (122.8 mg, 58%).LC-MS: (ES+H, m/z): [M+H] + =380.3; 1 H NMR (300 MHz, DMSO-d6) δ 11.88 (s, 1H), 8.38 (d, 1H), 8.09 (d, 1H), 7.61 (d, 1H), 7.55-7.45 (m, 1H), 7.42 (s, 1H), 6.86 (dd, 1H), 3.62 (s, 2H), 3.49-2.38 (m, 4H), 2.50-2.45 (m, 4H), 2.20-2.08 (m, 1H), 1.02 – 0.92 (m, 2H), 0.86 – 0.78 (m, 2H); 19 F NMR (282 MHz, DMSO-d6) δ -143.41. Examples 28 and 29 Step 1: Preparation of 7-(1-ethoxyvinyl)-3-methyl-1,5-naphthyridin-2(1H)-one: [00183] To a stirred mixture of 7-bromo-3-methyl-1H-1,5-naphthyridin-2-one (3.00 g, 12.54 mmol, 1.00 equiv) and tributyl(1-ethoxyethenyl)stannane (13.60 g, 37.64 mmol, 3.00 equiv) in 1,4-dioxane (20 mL) was added Pd(PPh3)2Cl2 (0.44 g, 0.62 mmol, 0.05 equiv) at rt. The resulting mixture was stirred for 2h at 100°C under nitrogen atmosphere. Upon completion, the reaction was cooled to rt and the resulting solution was used in the next step directly without further purification. LC-MS: (ES+H, m/z): [M+H] + =231.1 Step 2: Preparation of 7-acetyl-3-methyl-1,5-naphthyridin-2(1H)-one: [00184] The solution of 7-(1-ethoxyvinyl)-3-methyl-1,5-naphthyridin-2(1H)-one from Step 1 was cooled to 0 °C and treated withed conc. HCl (4 mL) dropwise. The resulting reaction mixture was stirred for 1h at rt and then basified to pH 8 with sat. NaHCO 3 (aq.). The resulting mixture was then diluted with water (100 mL) and extracted with CH 2 Cl 2 (3 x 200 mL). The combined organic layers were washed with brine (1x300 mL), dried over anhydrous Na2SO4, filtered, and concentrated. The crude product was purified by silica gel column chromatography to afford 7-acetyl-3-methyl-1,5-naphthyridin-2(1H)-one (1.39 g, 55% over two steps). LC-MS: (ES+H, m/z): [M+H] + =203.2 Step 3: Preparation of 6-[(2R)-2-methyl-4-[1-(7-methyl-6-oxo-5H-1,5-naphthyridin-3- yl)ethyl]piperazin-1-yl]pyridine-3-carbonitrile: [00185] 7-acetyl-3-methyl-1H-1,5-naphthyridin-2-one (350 mg, 1.73 mmol, 1.00 equiv) was added to a solution 6-[(2R)-2-methylpiperazin-1-yl]pyridine-3-carbonitrile (455 mg, 2.25 mmol, 1.30 equiv) in CH2Cl2 (8 mL). The resulting mixture was then concentrated under reduced pressure. To the above mixture was added Ti(OiPr)4 (4.6 mL, 15.58 mmol, 9.00 equiv) dropwise at rt. The resulting mixture was stirred for additional 4 h at 80°C and then cooled to rt. To the above mixture was added EtOH (5 mL) and NaBH3CN (217 mg, 3.46 mmol, 2.00 equiv) portion-wise at rt. The resulting mixture was stirred for an additional 2h at 80°C and then cooled down to rt. The reaction mixture was poured into water (100 mL) and stirred for 1h, filtered through a plug of Celite and washed with DCM/MeOH (3/1; 300 mL). The aqueous layer was extracted with DCM/i-PrOH (5/1; 2x100 mL) and the combined organic layers were washed with brine (2x50 mL), dried over Na2SO4., filtered and concentrated. The residue was purified by silica gel column chromatography to afford 6-[(2R)-2-methyl-4-[1-(7-methyl-6-oxo-5H-1,5-naphthyridin-3- yl)ethyl]piperazin-1-yl]pyridine-3-carbonitrile (280 mg, 41.6%). LC-MS: (ES-H, m/z): [M+H] + = 389.1. The diasteromers were separated by Chiral-HPLC to afford rel-6-[(2R)-2-methyl-4-[(1R*)-1-(7-methyl-6-oxo- 5H-1,5-naphthyridin-3-yl)ethyl]piperazin-1-yl]pyridine-3-car bonitrile (Example 2890.6 mg, de = 100%) and rel-6-[(2R)-2-methyl-4-[(1R*)-1-(7-methyl-6-oxo-5H-1,5-napht hyridin-3-yl)ethyl]piperazin-1- yl]pyridine-3-carbonitrile (Example 29, 69.5 mg, de = 100%). Data for Example 28: LC-MS: (ES+H, m/z): [M+H] + = 389.2; 1 H NMR (300 MHz, DMSO-d 6 ) δ 11.90 (s, 1H), 8.48 – 8.43 (dd, 2H), 7.85 – 7.82 (m, 2H), 7.65 – 7.64 (d, 1H), 6.86 – 6.83 (d, 1H), 4.51 (s, 1H), 4.26 – 4.22 (d, 1H), 3.59 – 3.57 (q, 1H), 3.14 – 3.11 (m, 2H), 2.58 – 2.50 (d, 1H), 2.18 – 2.06 (m, 5H), 1.36 – 1.34 (d, 3H), 1.18 – 1.15 (d, 3H). Data for Example 29: LC-MS: (ES+H, m/z): [M+H] + = 389.1; 1 H NMR (300 MHz, DMSO-d 6 ) δ 11.85 (s, 1H), 8.48 – 8.46(dd, 2H), 7.85 – 7.81 (m, 2H), 7.66 (d, 1H), 6.86 – 6.83 (d, 1H), 4.62 (s, 1H), 4.16 – 4.12 (d, 1H), 3.70 – 3.64 (q, 1H), 3.11 – 3.01 (td, 1H), 2.88 – 2.76 (dd, 2H), 2.26 – 2.03 (m, 5H), 1.34 (d, 3H), 1.22 (d, 3H). The relative stereochemistry for Examples 28 and 29 was arbitrarily assigned. Example 36 Step 1: Preparation of methyl 5-nitro-6-(prop-1-en-2-yl)pyridine-3-carboxylate: [00186] A mixture of methyl 6-chloro-5-nitropyridine-3-carboxylate (10.00 g, 46.17 mmol, 1.00 equiv), 4,4,5,5-tetramethyl-2-(prop-1-en-2-yl)-1,3,2-dioxaborolane (15.52 g, 92.34 mmol, 2.00 equiv), K2CO3 (12.76 g, 92.34 mmol, 2.00 equiv) and Pd(dppf)Cl2 (3.38 g, 4.62 mmol, 0.10 equiv) in dioxane (150 mL) and water (15 mL) was stirred for 3h at 100°C under nitrogen atmosphere. The reaction was monitored by LCMS. The mixture was allowed to rt. The resulting mixture was diluted with water (300 mL) and extracted with EtoAc (3x300 mL). The combined organic layers were washed with sat. NaCl (aq.) (3x100 mL), dried over anhydrous Na 2 SO 4 , filtered and concentrated under reduced pressure. The filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford methyl 5- nitro-6-(prop-1-en-2-yl)pyridine-3-carboxylate (5.00 g, 49%) as a light yellow oil. LC-MS: (ES+H, m/z): [M+H] + =222.95. Step 2: Preparation of methyl 5-nitro-6-(prop-1-en-2-yl)pyridine-3-carboxylate: [00187] To a stirred solution of methyl 5-nitro-6-(prop-1-en-2-yl)pyridine-3-carboxylate (5.00 g, 22.50 mmol, 1.00 equiv) in MeOH (100 mL) were added NH 4 Cl (25 mL, sat. aq.) and Fe (5.03 g, 90.01 mmol, 4.00 equiv). The reaction was stirred at 80°C for 4h under nitrogen atmosphere. The reaction was monitored by LCMS. The mixture was allowed to rt then concentrated under reduced pressure. The residue was diluted with CH 2 Cl 2 /2-Propanol (5:1, 200 mL) and washed with water (250 mL) and brine (250 mL). The organic layers were dried over anhydrous Na 2 SO 4 , filtered and concentrated under reduced pressure. This resulted in methyl 5-amino-6-(prop-1-en-2-yl)pyridine-3-carboxylate (3.90 g, 90%) which was used directly without further purification. LC-MS: (ES+H, m/z): [M+H] + =193.15. 1 H NMR (300 MHz, DMSO-d 6 ) δ 8.29 (d, 1H), 7.60 (d, 1H), 5.51-5.47 (m, 1H), 5.41 (s, 2H), 5.39 – 5.36 (m, 1H), 3.84 (s, 3H), 2.08 (t, 3H). Step 3: Preparation of methyl 8-methyl-6-oxo-5H-1,5-naphthyridine-3-carboxylate: [00188] A solution of triphosgene (1.54 g, 5.20 mmol, 0.50 equiv) in toluene (20 mL) was added to the solution of methyl 5-amino-6-(prop-1-en-2-yl)pyridine-3-carboxylate (3.90 g, 20.29 mmol, 1.00 equiv) and Et 3 N (6.16 g, 60.87 mmol, 3.00 equiv) in toluene (40 mL) at 0°C under nitrogen atmosphere. The resulting mixture was stirred overnight at 60°C under nitrogen. The reaction was monitored by LCMS. The reaction was quenched with MeOH (30 mL) at 0°C. The resulting mixture was diluted with water (200 mL) and extracted with CH2Cl2/2-Propanol (5:1, 3x200 mL). The combined organic layers were washed with water (3x100 mL), dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford methyl 8-methyl-6-oxo-5H-1,5-naphthyridine-3- carboxylate (1.80 g, 41%). LC-MS: (ES+H, m/z): [M+H] + =219.1; 1 H NMR (400 MHz, DMSO-d6) δ 11.92 (s, 1H), 8.92 (d, 1H), 8.15 (d, 1H), 6.79 (s, 1H), 3.93 (s, 3H), 2.48 (s, 3H). Step 4: Preparation of methyl 7-chloro-8-methyl-6-oxo-5H-1,5-naphthyridine-3-carboxylate: [00189] To a solution of methyl 8-methyl-6-oxo-5H-1,5-naphthyridine-3-carboxylate (600 mg, 2.75 mmol, 1.00 equiv) and NCS (587 mg, 4.40 mmol, 1.60 equiv) in CH3COOH (7 mL) was added 2,2-dichloroacetic acid (71 mg, 0.55 mmol, 0.20 equiv) dropwise at rt under nitrogen atmosphere. The resulting mixture was stirred overnight at 100°C and then cooled to rt. The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford methyl 7-chloro-8-methyl- 6-oxo-5H-1,5-naphthyridine-3-carboxylate (650 mg, 94%). LC-MS: (ES+H, m/z): [M+H] + =253.0; 1 H NMR (300 MHz, DMSO-d6) δ 8.90 (d, 1H), 8.13 (d, 1H), 3.93 (s, 3H), 2.60 (s, 3H). Step 5: Preparation of 3-chloro-7-(hydroxymethyl)-4-methyl-1H-1,5-naphthyridin-2-on e: [00190] To a stirred solution of methyl 7-chloro-8-methyl-6-oxo-5H-1,5-naphthyridine-3-carboxylate (600 mg, 2.38 mmol, 1.00 equiv) in THF (5 mL) was added LiAlH 4 (2 mL, 2.5 M in THF, 4.75 mmol, 2.00 equiv) dropwise at 0°C under nitrogen atmosphere. The resulting mixture was stirred for 1 h at rt under nitrogen atmosphere. The reaction was monitored by LCMS. The reaction was quenched by the addition of HCl (1 mL, 12M) at 0°C. The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford 3-chloro-7-(hydroxymethyl)-4-methyl-1H-1,5- naphthyridin-2-one (230 mg, 43%). LC-MS: (ES+H, m/z): [M+H] + =225.1; 1 H NMR (300 MHz, DMSO-d6) δ 12.24 (br, 1H), 8.48 (d, 1H), 7.77 (d, 1H), 5.69 (s, 1H), 4.63 (s, 2H), 2.63 (s, 3H). Step 6: Preparation of 3-chloro-7-(chloromethyl)-4-methyl-1H-1,5-naphthyridin-2-one : [00191] To a stirred solution of 3-chloro-7-(hydroxymethyl)-4-methyl-1H-1,5-naphthyridin-2-on e (200 mg, 0.89 mmol, 1.00 equiv) and DMF (7 mg, 0.09 mmol, 0.10 equiv) in DCM (10 mL) were added SOCl2 (318 mg, 2.67 mmol, 3.00 equiv) dropwise at 0°C under nitrogen atmosphere. The resulting mixture was stirred for 10 h at rt. The reaction was monitored by LCMS. The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford 3-chloro-7- (chloromethyl)-4-methyl-1H-1,5-naphthyridin-2-one (98 mg, 45%). LC-MS: (ES+H, m/z): [M+H] + =243.0. Step 7: Preparation of 6-{4-[(7-chloro-8-methyl-6-oxo-5H-1,5-naphthyridin-3-yl)meth yl]piperazin-1- yl}pyridine-3-carbonitrile: [00192] To a stirred mixture of 3-chloro-7-(chloromethyl)-4-methyl-1H-1,5-naphthyridin-2-one (85 mg, 0.35 mmol, 1.00 equiv) and 6-(piperazin-1-yl)pyridine-3-carbonitrile (66 mg, 0.35 mmol, 1.00 equiv) in ACN (1 mL) were added DIEA (136 mg, 1.05 mmol, 3.00 equiv) and KI (1 mg, 0.01 mmol, 0.10 equiv) at rt under nitrogen atmosphere. The resulting mixture was stirred for 1 h at 80°C. The reaction was monitored by LCMS. The mixture was allowed to cool down to rt. The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford the crude product (90 mg), which was further purified by prep HPLC to afford 6-{4-[(7-chloro-8-methyl-6-oxo-5H-1,5-naphthyridin-3- yl)methyl]piperazin-1-yl}pyridine-3-carbonitrile (52.5 mg, 28%). LC-MS: (ES+H, m/z): [M+H] + =395.10; 1 H NMR (300 MHz, DMSO-d6) δ 12.30 (br, 1H), 8.50 (m, 2H), 7.85 (dd, 1H), 7.69 (d, 1H), 6.94 (d, 1H), 3.67-3.62 (m, 6H), 2.65 (s, 3H), 2.50-2.49 (m, 4H). Example 42 Step 1: Preparation of methyl 7-(difluoromethyl)-8-methyl-6-oxo-5H-1,5-naphthyridine-3- carboxylate: [00193] A mixture of methyl 8-methyl-6-oxo-5H-1,5-naphthyridine-3-carboxylate (800 mg, 3.67 mmol, 1.00 equiv), difluoromethanesulfinic acid sodium (1012 mg, 7.33 mmol, 2.00 equiv), 2-methylpropane-2- peroxol (991 mg, 11.00 mmol, 3.00 equiv) and TFA (418 mg, 3.67 mmol, 1.00 equiv) in CH2Cl2/H2O (2.5:1, 120 mL) was stirred overnight at rt under nitrogen atmosphere. The reaction was monitored by LCMS. The resulting mixture was then diluted with water (100 mL) and extracted with CH 2 Cl 2 /2-Propanol (5:1, 3x100mL). The combined organic layers were washed with water (3x50 mL) and dried over anhydrous Na 2 SO 4 , filtered and concentrated under reduced pressure to afford methyl 7-(difluoromethyl)-8-methyl-6- oxo-5H-1,5-naphthyridine-3-carboxylate (650 mg, 66%). LC-MS: (ES+H, m/z): [M+H] + =269.1; 1 H NMR (400 MHz, DMSO-d 6 ) δ 12.39 (s, 1H), 8.99 (d, 1H), 8.19 (m, 1H), 7.26 (t, 1H), 3.94 (s, 3H), 2.72 (s, 3H). Step 2: Preparation of 3-(difluoromethyl)-7-(hydroxymethyl)-4-methyl-1H-1,5-naphthy ridin-2-one: [00194] To a stirred solution of methyl 7-(difluoromethyl)-8-methyl-6-oxo-5H-1,5-naphthyridine-3- carboxylate (650 mg, 2.42 mmol,1.00 equiv) in THF (65 mL) was added LiAlH 4 solution (2.5M in THF, 3.9 mL, 9.75 mmol 4.00 equiv) dropwise at 0°C under nitrogen atmosphere. The resulting mixture was stirred for 1h at rt under nitrogen atmosphere. The reaction was monitored by LCMS. The reaction was quenched by the addition of saturated aqueous NH 4 Cl (10 mL) at 0°C and dried over anhydrous Na 2 SO 4 . The mixture was filtered and washed with CH 2 Cl 2 / MeOH (10:1, 100 mL) and the combined organic layers were concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford 3-(difluoromethyl) -7-(hydroxymethyl)-4-methyl-1H-1,5-naphthyridin-2-one (520 mg, 89%). LC- MS: (ES+H, m/z): [M+H] + =241.1. 1 H NMR (400 MHz, Methanol-d4) δ 8.58 (d, 1H), 7.73 (d, 1H), 7.20 (t, 1H), 4.79 (s, 2H), 2.81 (t, 3H). Step 3: Preparation of 7-(chloromethyl)-3-(difluoromethyl)-4-methyl-1H-1,5-naphthyr idin-2-one: [00195] To a stirred mixture of 3-(difluoromethyl)-7-(hydroxymethyl)-4-methyl-1H-1,5-naphthy ridin-2- one (320 mg, 1.33 mmol, 1.00 equiv) and DMF (5 mg, 0.07 mmol, 0.05 equiv) in CH2Cl2 (32 mL) was added SOCl2 (475 mg, 4.00 mmol, 3.00 equiv) dropwise at 0°C under nitrogen atmosphere. The resulting mixture was stirred for 2 h at 50°C under nitrogen atmosphere. The reaction was monitored by LCMS. The mixture was allowed to rt then concentrated under reduced pressure to afford 7-(chloromethyl)-3- (difluoromethyl)-4-methyl-1H-1,5-naphthyridin-2-one (crude, 400 mg). LC-MS: (ES+H, m/z): [M+H] + =259.1 Step 4: Preparation of 6-(4-{[7-(difluoromethyl)-8-methyl-6-oxo-5H-1,5-naphthyridin -3-yl]methyl} piperazin-1-yl)pyridine-3-carbonitrile: [00196] A mixture of 7-(chloromethyl)-3-(difluoromethyl)-4-methyl-1H-1,5-naphthyr idin-2-one (380 mg, 1.47 mmol, 1.00 equiv), 6-(piperazin-1-yl)pyridine-3-carbonitrile (415 mg, 2.20 mmol, 1.50 equiv), KI (365 mg, 2.20 mmol, 1.50 equiv) and DIEA (569 mg, 4.40 mmol, 3.00 equiv) in MeCN (4mL) was stirred for 1h at 80°C under nitrogen atmosphere. The reaction was monitored by LCMS. The mixture was allowed to rt. The resulting mixture was diluted with water (50 mL) and extracted with EtOAc (3x50 mL). The combined organic layers were washed with water (3x10 mL), dried over anhydrous Na2SO4. The mixture was filtered and washed with EtOAc (50 mL) and the filtrate was concentrated under reduced pressure. The residue was purified by prep HPLC to afford 6-(4-{[7-(difluoromethyl)-8-methyl-6-oxo-5H-1,5-naphthyridin -3- yl]methyl} piperazin-1-yl)pyridine-3-carbonitrile (31 mg, 5%). LC-MS: (ES+H, m/z): [M+H] + =411.2; 1 H NMR (300 MHz, DMSO-d 6 ) δ 12.17 (br, 1H), 8.57 (d, 1H), 8.41 (d, 1H), 7.85 (dd, 1H), 7.68 (d, 1H), 7.24 (t, 1H), 6.94 (d, 1H),3.80-3.60 (m, 6H), 2.72 (s, 3H), 2.60-2.40 (m, 4H); 19 F NMR (282 MHz, DMSO-d6) δ - 115.51.
Example 47 Step 1: Preparation of 3-bromo-2-methoxy-6-methyl-5-nitropyridine: [00197] To a stirred mixture of 3-bromo-2-chloro-6-methyl-5-nitropyridine (20.00 g, 79.54 mmol, 1.00 equiv) in MeOH (50 mL) was added NaOMe (15.76 g, 87.49 mmol, 1.10 equiv, 30%wt) dropwise at 0°C under nitrogen atmosphere. The resulting mixture was stirred overnight at rt under nitrogen atmosphere. The reaction was monitored by TLC (Pet. Ether : EtOAc=1:1, R f = 0.4). Upon completion, the reaction was concentrated under reduced pressure and water (100 mL) was added. The resulting mixture was extracted with EtOAc (3x100 mL). The combined organic layers were washed with brine (1x200 mL), dried over anhydrous Na 2 SO 4 , filtered and concentrated to afford 3-bromo-2-methoxy-6-methyl-5-nitropyridine (20 g, 99%). 1 H NMR (400 MHz, DMSO-d 6 ) δ 8.66 (s, 1H), 4.04 (s, 3H), 2.70(s, 3H). Step 2: Preparation of (E)-2-(5-bromo-6-methoxy-3-nitropyridin-2-yl)-N,N-dimethylet hen-1-amine: [00198] A mixture of 3-bromo-2-methoxy-6-methyl-5-nitropyridine (15.00 g, 60.72 mmol, 1.00 equiv) in DMF-DMA (100 mL) and DMF (100 mL) was stirred overnight at 100°C under nitrogen atmosphere. The reaction was monitored by TLC (Pet. Ether : EtOAc =1:1, Rf = 0.5). The mixture was allowed to cool down to rt and then concentrated under reduced pressure to afford crude (E)-2-(5-bromo-6-methoxy-3- nitropyridin-2-yl)-N,N-dimethylethen-1-amine. The crude product was used in the next step directly without further purification. Step 3: Preparation of 5-bromo-6-methoxy-3-nitropicolinaldehyde: [00199] To a stirred mixture of (E)-2-(5-bromo-6-methoxy-3-nitropyridin-2-yl)ethenyl]dimethy lamine (18.01 g, crude) in THF (100 mL) and H2O (100 mL) was added NaIO4 (28.00 g, 131.07 mmol, 2.20 equiv) in portions at 0°C under nitrogen atmosphere. The resulting mixture was stirred for 2h at rt under nitrogen atmosphere. The reaction was monitored by TLC (Pet. Ether : EtOAc = 5:1, R f = 0.2). The reaction was quenched by the addition of sat. sodium hyposulfite (aq.) (100 mL) at rt. The resulting mixture was extracted with EtOAc (3x100 mL). The combined organic layers were washed with brine (2x100 mL), dried over anhydrous Na 2 SO 4 . After filtration, the filtrate was concentrated under reduced pressure. The crude product was used in the next step directly without further purification. 1 H NMR (400 MHz, DMSO-d 6 ) δ 10.16 (s, 1H), 8.87 (s, 1H), 4.10 (s, 3H). Step 4: Preparation of ethyl 7-bromo-6-methoxy-1,5-naphthyridine-3-carboxylate: [00200] To a stirred mixture of 5-bromo-6-methoxy-3-nitropyridine-2-carbaldehyde (7.00 g, crude) and ethyl 3,3-diethoxypropanoate (20.40 g, 107.27 mmol, 4.00 equiv) in EtOH (100 mL) were added SnCl2 (26.25 g, 134.09 mmol, 5.00 equiv) in portions at rt under nitrogen atmosphere. The resulting mixture was stirred overnight at 90°C under nitrogen atmosphere. The reaction was monitored by LCMS. The mixture was allowed to cool down to rt. The resulting mixture was concentrated under reduced pressure. The crude mixture was poured into saturated sodium bicarbonate (100 mL). The resulting mixture was extracted with EtOAc (3x100 mL). The combined organic layers were concentrated under reduced pressure and the residue was purified by silica gel column chromatography to afford the crude product. The crude product was further purified by trituration with hexane (50 mL) to afford ethyl 7-bromo-6-methoxy-1,5-naphthyridine-3- carboxylate (3.50 g, 18.5%, over three steps). LC-MS: (ES+H, m/z): [M+H] + =311.0 Step 5: Preparation of ethyl 7-chloro-6-methoxy-1,5-naphthyridine-3-carboxylate: [00201] To a stirred mixture of ethyl 7-bromo-6-methoxy-1,5-naphthyridine-3-carboxylate (1.20 g, 3.85 mmol, 1.00 equiv) in DMF (10 mL) was added CuCl (0.57 g, 5.78 mmol, 1.50 equiv) at rt under nitrogen atmosphere. The resulting mixture was stirred overnight at 120°C. The reaction was monitored by LCMS. The mixture was allowed to cool down to rt. The resulting mixture was diluted with EtOAc (20 mL). The resulting mixture was washed with 3x30 mL of Water (10% NH 3 ^H 2 O). The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford ethyl 7-chloro-6-methoxy-1,5-naphthyridine-3-carboxylate (800 mg, 77.78%). LC-MS: (ES+H, m/z): [M+H] + =267.0; 1 H NMR (300 MHz, DMSO-d6) δ 9.27 (d, 1H), 8.63 (d, 1H), 8.57 (s, 1H), 4.41 (q, 2H), 4.12 (s, 3H), 1.37 (t, 3H). Step 6: Preparation of ethyl 7-chloro-6-oxo-5H-1,5-naphthyridine-3-carboxylate: [00202] To a stirred mixture of ethyl 7-chloro-6-methoxy-1,5-naphthyridine-3-carboxylate (800 mg, 3.00 mmol, 1.00 equiv) in CH 3 CN (8 mL) was added TMSI (1.80 g, 9.00 mmol, 3.00 equiv) at rt under nitrogen atmosphere. The resulting mixture was stirred for 2 h at 50°C.The reaction was monitored by LCMS. The mixture was allowed to cool down to rt. The resulting mixture was diluted with EtOAc (50 mL). The aqueous layer was washed with 3x50 mL of water (10% Et 3 N). The combined organic layers were washed with brine (50 mL), dried over anhydrous Na 2 SO 4 , filtered and concentrated. The residue was purified by silica gel column chromatography to afford ethyl 7-chloro-6-oxo-5H-1,5-naphthyridine-3-carboxylate (740 mg, 97.64%). LC-MS: (ES+H, m/z): [M+H] + =252.9; 1 H NMR (300 MHz, DMSO-d 6 ) δ12.61 (s, 1H), 8.94 (d, 1H), 8.37 (d, 1H), 8.20 (s, 1H), 4.39 (q, 2H), 1.36 (t, 3H). Step 7: Preparation of 3-chloro-7-(hydroxymethyl)-1H-1,5-naphthyridin-2-one: [00203] To a stirred mixture of ethyl 7-chloro-6-oxo-5H-1,5-naphthyridine-3-carboxylate (740 mg, 2.92 mmol, 1.00 equiv) in THF (6 mL) was added LiAlH4 (2.5 mL, 5.85 mmol, 2.00 equiv) dropwise at 0°C under nitrogen atmosphere. The resulting mixture was stirred for additional 2 h at 0°C. The reaction was monitored by LCMS. The mixture was acidified to pH 5 with 1 M HCl(aq.). The resulting mixture was concentrated under reduced pressure and the residue was purified by silica gel column chromatography to afford 3-chloro-7-(hydroxymethyl)-1H-1,5-naphthyridin-2-one (250 mg, 40.53%). LC-MS: (ES+H, m/z): [M+H] + =211.0; 1 H NMR (400 MHz, DMSO-d6) δ12.49 (s, 1H), 8.45 (d, 1H), 8.28 (s, 1H), 7.69 (d, 1H), 5.53 (t, 1H), 4.64 (d, 2H). Step 4: Preparation of 3-chloro-7-(chloromethyl)-1H-1,5-naphthyridin-2-one: [00204] To a stirred mixture of 3-chloro-7-(hydroxymethyl)-1H-1,5-naphthyridin-2-one (250 mg, 1.18 mmol, 1.00 equiv) in CH2Cl2 (5 mL) was added SOCl2 (423 mg, 3.56 mmol, 3.00 equiv) and DMF (8 mg, 0.11 mmol, 0.10 equiv) dropwise at rt under nitrogen atmosphere. The resulting mixture was stirred for 3 h at rt. The reaction was monitored by LCMS. The resulting mixture was concentrated under reduced pressure. This resulted in 3-chloro-7-(chloromethyl)-1H-1,5-naphthyridin-2-one (280 mg, crude). The crude product was used in the next step directly without further purification. LC-MS: (ES+H, m/z): [M+H] + =228.95. Step 9: Preparation of 3-chloro-7-{[4-(5-fluoropyridin-2-yl)piperazin-1-yl]methyl}- 1H-1,5- naphthyridin-2-one: [00205] To a stirred mixture of 3-chloro-7-(chloromethyl)-1H-1,5-naphthyridin-2-one (200 mg, 0.87 mmol, 1.00 equiv) and 1-(5-fluoropyridin-2-yl)piperazine (126 mg, 0.69 mmol, 0.80 equiv) in CH 3 CN (5 mL) were added DIEA (564 mg, 4.36 mmol, 5 equiv) and KI (14 mg, 0.08 mmol, 0.10 equiv) at rt. The resulting mixture was stirred for 2 h at 50°C. The reaction was monitored by LCMS. The resulting mixture was cooled down to rt and poured into 20 mL of water. The resulting mixture was extracted with EtOAc (3 x 20 mL). The combined organic layers were washed with brine (50 mL), dried over anhydrous Na 2 SO 4 . After filtration, the filtrate was concentrated under reduced pressure. The crude product was purified by prep HPLC to afford 3-chloro-7-{[4-(5-fluoropyridin-2-yl)piperazin-1-yl]methyl}- 1H-1,5-naphthyridin-2-one (42.7 mg, 13.08%). LC-MS: (ES+H, m/z): [M+H] + =374.0; 1 1H NMR (300 MHz, DMSO-d 6 ) δ 12.48 (s, 1H), 8.48 (d, 1H), 8.29 (s, 1H), 8.09 (d, 1H), 7.70 (d, 1H), 7.55 – 7.47 (m, 1H), 6.87 (dd, 1H), 3.66 (s, 2H), 3.46 – 3.42 (m, 4H), 2.53–2.50(m, 4H); 19 F NMR (282 MHz, DMSO-d 6 ) δ -143.39. Example 48 Step 1: Preparation of ethyl 7-(difluoromethyl)-6-methoxy-1,5-naphthyridine-3-carboxylate : [00206] A solution of ethyl 7-bromo-6-methoxy-1,5-naphthyridine-3-carboxylate (300 mg, 0.96 mmol, 1.00equiv), [1,3-Bis[2,6-bis(i-propyl)phenyl]-2-imidazolidinylidene]difl uoromethylsilver(I) (97 mg, 0.17 mmol, 1.10 equiv), [2-[2-(diphenylphosphanyl)phenoxy]phenyl]diphenylphosphane (181 mg, 0.33 mmol, 0.35 equiv) and Pd(dba)2 (177 mg, 0.30 mmol, 0.32 equiv) in toluene (5 ml) was stirred for 1.5 h at 80 °C under nitrogen atmosphere. The reaction was monitored by LCMS. The mixture was allowed to cool down to rt. The resulting mixture was diluted with EtOAc (50 mL). The resulting mixture was filtered and the filter cake was washed with EtOAc (3x50 mL). The filtrate was concentrated under reduced pressure. The residue was purified by Prep-TLC (Pet. Ether / EtOAc 5:1) to afford ethyl 7-(difluoromethyl)-6-methoxy- 1,5-naphthyridine-3-carboxylate (235 mg, 86%). LC-MS: (ES+H, m/z): [M+H] + =283.1; 1 H NMR (300 MHz, DMSO-d6) δ 9.25 (d, 1H), 8.59 (d, 1H), 8.53 (s, 1H), 7.24 (t, 1H), 4.42 (q, 2H), 4.12 (s, 3H), 1.40 (t, 3H); 19 F NMR (282 MHz, DMSO-d6) δ -117.83. Step 2: Preparation of ethyl 7-(difluoromethyl)-6-oxo-5H-1,5-naphthyridine-3-carboxylate: [00207] A solution of ethyl 7-(difluoromethyl)-6-methoxy-1,5-naphthyridine-3-carboxylate (240 mg, 0.85 mmol, 1.00 equiv) in ACN (7 mL) was added TMSI (680 mg, 3.40 mmol, 4.00 equiv) dropwise at rt. The reaction mixture was then stirred for 5 h at 50 o C under nitrogen atmosphere, cooled to rt and concentrated under reduced pressure. The residue was purified by silica gel column chromatography, (Pet. Ether / EtOAc) to afford ethyl 7-(difluoromethyl)-6-oxo-5H-1,5-naphthyridine-3-carboxylate (225 mg, 99%). LC- MS: (ES+H, m/z): [M+H] + = 269.1. Step 3: Preparation of 3-(difluoromethyl)-7-(hydroxymethyl)-1H-1,5-naphthyridin-2-o ne: [00208] To a stirred solution of ethyl 7-(difluoromethyl)-6-oxo-5H-1,5-naphthyridine-3-carboxylate (230 mg, 0.85 mmol, 1.00 equiv) in THF (5 ml) was added LiAlH 4 (0.69 mL, 2.5 mol/L in THF, 2.00 equiv) dropwise at 0 °C under nitrogen atmosphere. The resulting mixture was stirred for 1 h at rt. The reaction was monitored by LCMS. The resulting mixture was quenched with MeOH (10 mL) and then DCM (50 mL) was added. The solution was filtered, the filter cake was washed with DCM / MeOH (5:1) (3x50 mL). The filtrate was concentrated under reduced pressure. The residue was purified by silica gel column (CH 2 Cl 2 / MeOH) to afford 3-(difluoromethyl)-7-(hydroxymethyl)-1H-1,5-naphthyridin-2-o ne (50 mg, 26%). LC-MS: (ES+H, m/z): [M+H] + = 227.0; 1 H NMR (400 MHz, DMSO-d 6 ) δ 12.36 (s, 1H), 8.50 (d, 1H), 8.16 (d, 1H), 7.70 (s, 1H), 6.98 (t, 1H), 5.57 (t, 1H), 4.67 (d, 2H). Step 4: Preparation of 7-(chloromethyl)-3-(difluoromethyl)-1H-1,5-naphthyridin-2-on e: [00209] To a stirred solution of 3-(difluoromethyl)-7-(hydroxymethyl)-1H-1,5-naphthyridin-2-o ne (80 mg, 0.35 mmol, 1.00 equiv) and DMF (1 mg, 0.02 mmol, 0.05 equiv) in DCM (4 ml) was added SOCl2 (126 mg, 1.06 mmol, 3.00 equiv) dropwise at rt under nitrogen atmosphere. The resulting mixture was stirred for 5 h at rt. The reaction was monitored by LCMS. The resulting mixture was concentrated under reduced pressure. The crude product was used in the next step directly without further purification. LC-MS: (ES+H, m/z): [M+H] + = 245.0. Step 5: Preparation of 6-(4-{[7-(difluoromethyl)-6-oxo-5H-1,5-naphthyridin-3-yl]met hyl}piperazin-1- yl)pyridine-3-carbonitrile: [00210] To a stirred solution of 7-(chloromethyl)-3-(difluoromethyl)-1H-1,5-naphthyridin-2-on e (86 mg, assumed 100% yield, 0.35 mmol, 1.00 equiv), 6-(piperazin-1-yl)pyridine-3-carbonitrile (66 mg, 0.35 mmol, 1.00 equiv), KI (11 mg, 0.07 mmol, 0.20 equiv) in ACN(5 ml) was added DIEA (227 mg, 1.76 mmol, 5.00 equiv) dropwise at rt. The resulting mixture was stirred for 1 h at 80 o C. The reaction was monitored by LCMS. The mixture was allowed to cool down to rt. The resulting mixture was diluted with water (50 mL). The solution was extracted with EtOAc (3 x 100 mL). The combined organic layers were concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford the crude product (80 mg). The crude product was purified by prep HPLC to afford 6-(4-{[7-(difluoromethyl)-6-oxo- 5H-1,5-naphthyridin-3-yl]methyl}piperazin-1-yl)pyridine-3-ca rbonitrile (42.9 mg, 33% over two steps). LC- MS: (ES+H, m/z): [M+H] + = 397.1 ; 1 H NMR (300 MHz, DMSO-d 6 ) δ 12.33 (s, 1H), 8.54 (d, 1H), 8.48 (d, 1H), 8.16 (s, 1H), 7.85 (dd, 1H), 7.71 (d, 1H), 7.20 – 6.73 (m, 2H), 3.72 – 3.68 (m, 4H), 3.67 (s, 2H), 2.51 – 2.49 (m, 4H); 19 F NMR (282 MHz, DMSO-d 6 ) δ -119.30. Example 49 Step 1: Preparation of methyl 2-[(4-bromo-2-fluoro-6-nitrophenyl)amino]propanoate: [00211] To a stirred mixture of 5-bromo-1,2-difluoro-3-nitrobenzene (10.00 g, 42.01 mmol, 1.00 equiv) and methyl 2-aminopropanoate hydrochloride (2.93 g, 21.00 mmol, 1.00 equiv) in NMP (200 mL) was added DIEA (27.15 g, 210.09 mmol, 5.00 equiv) dropwise at rt under nitrogen atmosphere. The resulting mixture was stirred for 2h at 100°C under nitrogen atmosphere. The reaction was monitored by TLC (Pet. Ether/EtOAc=10/1). The mixture was allowed to cool down to rt. The resulting mixture was diluted with EtOAc (1L). The resulting mixture was washed with water (3x300 mL), dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford methyl 2-[(4-bromo-2-fluoro-6-nitrophenyl)amino]propanoate (7.5 g, 56%) as a red oil. 1 H NMR (400 MHz, DMSO-d6) δ 8.08 (t, 1H), 7.87 (dd, 1H), 7.77 (dd, 1H), 4.67 - 4.55 (m, 1H), 3.68 (s, 3H), 1.48 (dd, 3H); 19 F NMR (377 MHz, DMSO-d6) δ -122.19. Step 2: Preparation of 7-bromo-5-fluoro-3-methyl-3,4-dihydro-1H-quinoxalin-2-one: [00212] To a stirred solution of methyl 2-[(4-bromo-2-fluoro-6-nitrophenyl)amino]propanoate (6.00 g, 18.68 mmol, 1.00 equiv) in HOAc (200 mL) was added Fe (5.22 g, 93.43 mmol, 5.00 equiv) at rt under nitrogen atmosphere. The resulting mixture was stirred for 2h at 80°C under nitrogen atmosphere. The reaction was monitored by LCMS. The mixture was allowed to cool down to rt. The resulting mixture was diluted with EtOAc (50 mL). The resulting mixture was filtered, the filter cake was washed with DCM:MeOH=4:1 (3x100 mL). The filtrate was concentrated under reduced pressure. The pH of the residue was adjusted to pH 7 with sat. NaHCO3 (aq.). The resulting mixture was then extracted with CH2Cl2 (3 x 300mL). The combined organic layers were washed with brine (1x100 mL) and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure to afford 7-bromo-5-fluoro-3- methyl-3,4-dihydro-1H-quinoxalin-2-one (4.2 g, 87%). LC-MS: (ES-H, m/z): [M-H]- =257.1; 1 H NMR (400 MHz, DMSO-d 6 ) δ 10.54 (s, 1H), 7.01 (dd, 1H), 6.76 (t, 1H), 6.20 (s, 1H), 3.90 - 3.80 (m, 1H), 1.28 (d, 3H). 19 F NMR (377 MHz, DMSO-d 6 ) δ -132.99. Step 3: Preparation of 7-bromo-5-fluoro-3-methyl-1H-quinoxalin-2-one: [00213] To a stirred solution of 7-bromo-5-fluoro-3-methyl-3,4-dihydro-1H-quinoxalin-2-one (4.50 g, 17.36 mmol, 1.00 equiv) in DCM (500 mL) was added DDQ (4.34 g, 19.10 mmol, 1.10 equiv) at rt. The resulting mixture was stirred for 2h at rt under nitrogen atmosphere. The reaction was monitored by LCMS. The resulting mixture was concentrated under reduced pressure. The resulting mixture was diluted with sat. NaHCO 3 (aq) (500 mL). The resulting mixture was stirred for 30min at rt. The resulting mixture was filtered, the filter cake was washed with sat. NaHCO 3 (aq) (3x100 mL). The filter cake was dried under reduced pressure. The residue was purified by silica gel column chromatography to afford 7-bromo-5-fluoro-3- methyl-1H-quinoxalin-2-one (3 g, 67%). LC-MS: (ES-H, m/z): [M-H]- =255.0; 1 H NMR (400 MHz, DMSO- d 6 ) δ 12.52 (s, 1H), 7.49 - 7.40 (m, 1H), 7.24 (q, 1H), 2.40 (q, 3H). Step 4: Preparation of 8-fluoro-2-methyl-3-oxo-4H-quinoxaline-6-carbaldehyde: [00214] To a solution of 7-bromo-5-fluoro-3-methyl-1H-quinoxalin-2-one (500 mg, 1.94 mmol, 1.00 equiv) in Toluene (100 mL) was added bis(adamantan-1-yl)(butyl)phosphane (349 mg, 0.97 mmol, 0.50 equiv), TMEDA (1.13 g, 9.72 mmol, 5.00 equiv) and Pd(OAc)2 (218 mg, 0.97 mmol, 0.50 equiv) in a pressure tank. The mixture was purged with nitrogen for 5 min and then was pressurized to 30Mpa with CO2:H2=1:1 at rt. The resulting mixture was stirred overnight at 100°C. The reaction mixture was cooled to rt and filtered to remove insoluble solids. The reaction was monitored by LCMS. The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford 8-fluoro-2-methyl-3-oxo-4H-quinoxaline-6-carbaldehyde (300 mg, 74.8%). LC-MS: (ES-H, m/z): [M-H]- =205.2; 1 H NMR (400 MHz, DMSO-d 6 ) δ 12.75 (s, 1H), 10.02 (q, 1H), 7.67 – 7.52 (m, 2H), 2.49 – 2.44 (m, 3H). Step 5: Preparation of 6-{4-[(8-fluoro-2-methyl-3-oxo-4H-quinoxalin-6-yl)methyl]pip erazin-1- yl}pyridine-3-carbonitrile: [00215] To a stirred mixture of 8-fluoro-2-methyl-3-oxo-4H-quinoxaline-6-carbaldehyde (200 mg, 0.97 mmol, 1.00 equiv) and 6-(piperazin-1-yl)pyridine-3-carbonitrile (274 mg, 1.45 mmol, 1.50 equiv) in THF (20 mL) was added tetrakis(propan-2-yloxy)titanium (551 mg, 1.94 mmol, 2.00 equiv) at rt under nitrogen atmosphere. The resulting mixture was stirred for 4h at rt under nitrogen atmosphere. To the above mixture was added NaBH(OAc)3 (822 mg, 3.88 mmol, 4.00 equiv) at rt. The resulting mixture was stirred for additional 4h at rt. The reaction was monitored by LCMS. The reaction was quenched by the addition of Water (80 mL) at 0°C. The resulting mixture was extracted with EtOAc (3 x 100 mL). The combined organic layers were washed with brine (1x50 mL), dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography followed by trituration with MeCN (5mL) to afford 6-{4-[(8-fluoro-2-methyl-3-oxo-4H-quinoxalin-6- yl)methyl]piperazin-1-yl}pyridine-3-carbonitrile (51.1 mg, 14%). LC-MS: (ES+H, m/z): [M+H] + =379.2; 1 H NMR (400 MHz, DMSO-d6) δ 12.44 (s, 1H), 8.48 (d, 1H), 7.85 (dd, 1H), 7.09 (d, 2H), 6.93 (d, 1H), 3.68 (t, 4H), 3.60 - 3.55 (m,2H), 2.47 (d, 4H), 2.41 (s, 3H); 19 F NMR (377 MHz, DMSO-d6) δ -125.51. Example 60 Step 1: Preparation of 3-bromo-2-methoxy-6-methyl-5-nitropyridine: [00216] To a stirred mixture of 3-bromo-2-chloro-6-methyl-5-nitropyridine (20.00 g, 79.54 mmol, 1.00 equiv.) in MeOH (50 mL) was added NaOMe (15.76 g, 87.49 mmol, 1.10 equiv., 30%wt.) dropwise at 0°C under nitrogen atmosphere. The resulting mixture was stirred overnight at room temperature under nitrogen atmosphere. The reaction was monitored by TLC (PE:EA=1:1, R f = 0.4). The resulting mixture was concentrated under reduced pressure and water (100 mL) was added. The resulting mixture was extracted with EtOAc (3x100 mL). The combined organic layers were washed with brine (1x200 mL), dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure to afford 3-bromo- 2-methoxy-6-methyl-5-nitropyridine (20 g, 99%). 1 H NMR (400 MHz, DMSO-d6) δ 8.66 (s, 1H), 4.04 (s, 3H), 2.70(s, 3H). Step 2: Preparation of (E)-2-(5-bromo-6-methoxy-3-nitropyridin-2-yl)-N,N-dimethylet hen-1-amine: [00217] A mixture of 3-bromo-2-methoxy-6-methyl-5-nitropyridine (15.00 g, 60.72 mmol, 1.00 equiv.) in DMF-DMA (100 mL) and DMF (100 mL) was stirred overnight at 100°C under nitrogen atmosphere. The reaction was monitored by TLC (PE:EA=1:1, Rf = 0.5). The mixture was allowed to cool down to room temperature. The resulting mixture was concentrated under reduced pressure. The crude product was used in the next step directly without further purification. Step 3: Preparation of 5-bromo-6-methoxy-3-nitropicolinaldehyde: [00218] To a stirred mixture of (E)-2-(5-bromo-6-methoxy-3-nitropyridin-2-yl)ethenyl]dimethy lamine (18.01 g, crude) in THF (100 mL) and H2O (100 mL) was added NaIO4 (28.00 g, 131.07 mmol, 2.20 equiv.) in portions at 0°C under nitrogen atmosphere. The resulting mixture was stirred for 2h at room temperature under nitrogen atmosphere. The reaction was monitored by TLC (PE:EA= 5:1, R f = 0.2). The reaction was quenched by the addition of sat. sodium hyposulfite (aq.) (100 mL) at room temperature. The resulting mixture was extracted with EtOAc (3x100 mL). The combined organic layers were washed with brine (2x100 mL), dried over anhydrous Na 2 SO 4 . After filtration, the filtrate was concentrated under reduced pressure. The crude product was used in the next step directly without further purification. 1 H NMR (400 MHz, DMSO-d 6 ) δ 10.16 (s, 1H), 8.87 (s, 1H), 4.10 (s, 3H). Step 4: Preparation of ethyl 7-bromo-6-methoxy-1,5-naphthyridine-3-carboxylate: [00219] To a stirred mixture of 5-bromo-6-methoxy-3-nitropyridine-2-carbaldehyde (7.00 g, crude) and ethyl 3,3-diethoxypropanoate (20.40 g, 107.27 mmol, 4.00 equiv.) in EtOH (100 mL) were added SnCl 2 (26.25 g, 134.09 mmol, 5.00 equiv.) in portions at room temperature under nitrogen atmosphere. The resulting mixture was stirred overnight at 90°C under nitrogen atmosphere. The reaction was monitored by LCMS. The mixture was allowed to cool down to room temperature. The resulting mixture was concentrated under reduced pressure. The crude mixture was poured into saturated sodium bicarbonate (100 mL). The resulting mixture was extracted with EtOAc (3x100 mL). The combined organic layers were concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford crude product. The crude product was purified by trituration with hexane (50 mL) to afford ethyl 7-bromo-6- methoxy-1,5-naphthyridine-3-carboxylate (3.50 g, 18.5% yield over three steps). LC-MS: (ES+H, m/z): [M+H] + =311.0/313.0; 1 H NMR (400 MHz, DMSO-d 6 ) δ 9.22 (s, 1H), 8.78 (s, 1H), 8.58 (s, 1H), 4.42 (q, 2H), 4.12 (s, 3H), 1.39 (t 3H). Step 5: Preparation of ethyl 7-bromo-6-oxo-5H-1,5-naphthyridine-3-carboxylate: [00220] A solution of ethyl 7-bromo-6-methoxy-1,5-naphthyridine-3-carboxylate (5.00 g, 16.07 mmol, 1.00 equiv.) in ACN (400 mL) were added TMSI (13.8 mL, 96.42 mmol, 6.00 equiv.) dropwise at room temperature. The final reaction mixture was stirred for 2h at 80°C under nitrogen atmosphere. The mixture was allowed to cool down to room temperature. The reaction was monitored by LCMS. The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography with the following conditions: column, silica gel; mobile phase, EA in DCM, 45% to 60% gradient in 20 min; detector, UV 254 nm. The resulting mixture was concentrated under reduced pressure to afford ethyl 7-bromo-6-oxo-5H-1,5-naphthyridine-3-carboxylate (3.8 g, 79.5%). LC-MS: (ES+H, m/z): [M+H] + =296.95/298.95 Step 6: Preparation of 3-bromo-7-(hydroxymethyl)-1H-1,5-naphthyridin-2-one: [00221] To a stirred mixture of ethyl 7-bromo-6-oxo-5H-1,5-naphthyridine-3-carboxylate (450 mg, 1.51 mmol, 1.00 equiv.) in THF (8 mL) was added LiAlH4 (1.21 mL, 3.03 mmol, 2.00 equiv, 2.5M in THF) dropwise at 0°C under nitrogen atmosphere. The resulting mixture was stirred for 1h at 0°C under nitrogen atmosphere. The reaction was monitored by LCMS. The reaction was quenched by the addition of HCl (3.03 mL, 3.03 mmol, 2.00 equiv.) at 0°C. The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford 3-bromo-7-(hydroxymethyl)-1H-1,5- naphthyridin-2-one (220 mg, 56.9%). LC-MS: (ES+H, m/z): [M+H] + =255.0/257.0. Step 7: Preparation of 3-bromo-7-(chloromethyl)-1H-1,5-naphthyridin-2-one: [00222] To a stirred mixture of 3-bromo-7-(hydroxymethyl)-1H-1,5-naphthyridin-2-one (230 mg, 0.90 mmol, 1.00 equiv.) and DMF (6 mg, 0.09 mmol, 0.10 equiv.) in DCM (7 mL) was added SOCl 2 (321 mg, 2.70 mmol, 3.00 equiv.) dropwise at 0°C under nitrogen atmosphere. The resulting mixture was stirred overnight at room temperature under nitrogen atmosphere. The reaction was monitored by LCMS. The resulting mixture was concentrated under reduced pressure to afford 3-bromo-7-(chloromethyl)-1H-1,5- naphthyridin-2-one (220 mg, 89.2%). LC-MS: (ES+H, m/z): [M+H] + =272.9/274.9. Step 8: Preparation of 6-{4-[(7-bromo-6-oxo-5H-1,5-naphthyridin-3-yl)methyl]piperaz in-1-yl}pyridine- 3-carbonitrile: [00223] To a stirred mixture of 3-bromo-7-(chloromethyl)-1H-1,5-naphthyridin-2-one (150 mg, 0.54 mmol, 1.00 equiv.), DIEA (354 mg, 2.74 mmol, 5.00 equiv.) and 6-(piperazin-1-yl)pyridine-3-carbonitrile (113 mg, 0.60 mmol, 1.10 equiv.) in MeCN (7 mL) was added KI (18 mg, 0.11 mmol, 0.20 equiv.) in portions at room temperature under nitrogen atmosphere. The resulting mixture was stirred for 2h at 80°C under nitrogen atmosphere. The reaction was monitored by LCMS. The mixture was allowed to cool down to room temperature. The resulting mixture was concentrated under reduced pressure. The crude product (120mg) was purified by Prep-HPLC, the pure fraction was concentrated then lyophilized to afford 6-{4-[(7- bromo-6-oxo-5H-1,5-naphthyridin-3-yl)methyl]piperazin-1-yl}p yridine-3-carbonitrile (25.6 mg , 10.9%). LC-MS: (ES+H, m/z): [M+H] + =425.00/427.00; 1 H NMR (400 MHz, DMSO-d 6 ) δ 12.41 (s, 1H), 8.47 (s, 3H), 7.85 (dd, J = 9.1, 2.4 Hz, 1H), 7.68 (s, 1H), 6.93 (d, J = 9.1 Hz, 1H), 3.78 – 3.59 (m, 6H), 2.48 (d, J = 4.7 Hz, 4H). Example 68 and 69 Step 1: Preparation of ethyl 6-methoxy-7-(oxolan-2-yl)-1,5-naphthyridine-3-carboxylate: [00224] To a stirred solution of ethyl 7-bromo-6-methoxy-1,5-naphthyridine-3-carboxylate (5.00 g, 16.07 mmol, 1.00 equiv.) and (4-methoxyphenyl)[4-(trifluoromethyl)phenyl]methanone (1.12 g, 4.01 mmol, 0.25 equiv.) in THF were added 5,5'-dimethyl-2,2'-bipyridine (0.74 g, 4.01 mmol, 0.25 equiv.), nickel acetylacetonate (1.03 g, 4.01 mmol, 0.25 equiv.) and Na2CO3 (1.70 g, 16.07 mmol, 1.00 equiv.) at room temperature under nitrogen atmosphere. The resulting mixture was placed approximately ~3 cm away from two 40W Blue LED and stirred for 7 days at room temperature under nitrogen atmosphere. The reaction was monitored by LCMS. The resulting mixture was extracted with EtOAc (3 x 100 mL). The combined organic layers were washed with brine (50 mL), dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford ethyl 6-methoxy-7-(oxolan-2-yl)-1,5-naphthyridine-3-carboxylate (1.6 g, 32.93%). LC-MS: (ES+H, m/z): [M + H] + = 303.15; 1 H NMR (400 MHz, DMSO-d 6 ) δ 9.20 (d, J = 2.0 Hz, 1H), 8.57 (dd, J = 2.1, 0.8 Hz, 1H), 8.27 – 8.10 (m, 1H), 5.13 – 5.04 (m, 1H), 4.41 (q, J = 7.1 Hz, 2H), 4.18 – 4.09 (m, 1H), 4.09 (s, 3H), 3.95 – 3.85 (m, 1H), 2.49 – 2.39 (m, 1H), 2.05 – 1.82 (m, 2H), 1.79 – 1.67 (m, 1H), 1.39 (t, J = 7.1 Hz, 3H). Step 2 : Preparation of ethyl 6-oxo-7-(oxolan-2-yl)-5H-1,5-naphthyridine-3-carboxylate: [00225] To a stirred solution of ethyl 6-methoxy-7-(oxolan-2-yl)-1,5-naphthyridine-3-carboxylate (1.30 g, 4.30 mmol, 1.00 equiv.) in MeCN were added HBr in AcOH (0.25 ml, 33 wt.%, 2.00 equiv.) at 0°C under nitrogen atmosphere. The resulting mixture was stirred for 0.5 h at 0°C. The reaction was monitored by LCMS. The resulting mixture was diluted with water (20 mL) and basified to pH 8 with Et 3 N. The resulting mixture was extracted with EtOAc (3 x 50mL). The combined organic layers were washed with brine (50 mL), dried over anhydrous Na 2 SO 4 . After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford ethyl 6-oxo-7-(oxolan-2-yl)-5H-1,5- naphthyridine-3-carboxylate (730 mg, 58.89%). LC-MS: (ES+H, m/z): [M+H] + = 289.10; 1 H NMR (400 MHz, DMSO-d 6 ) δ 12.14 (s, 1H), 8.91 (d, J = 1.9 Hz, 1H), 8.18 (d, J = 1.9 Hz, 1H), 7.84 (d, J = 1.1 Hz, 1H), 4.96 – 4.87 (m, 1H), 4.39 (q, J = 7.1 Hz, 2H), 4.12 – 4.04 (m, 1H), 3.91 – 3.79 (m, 1H), 2.48 – 2.32 (m, 1H), 1.99 – 1.81 (m, 2H), 1.74 – 1.63 (m, 1H), 1.36 (t, J = 7.1 Hz, 3H). Step 3: Preparation of 7-(hydroxymethyl)-3-(oxolan-2-yl)-1H-1,5-naphthyridin-2-one: [00226] To a stirred mixture of ethyl 6-oxo-7-(oxolan-2-yl)-5H-1,5-naphthyridine-3-carboxylate (730 mg, 2.53 mmol, 1.00 equiv.) in THF (8 mL) was added LiAlH4 (192 mg, 5.06 mmol, 2.00 equiv.) dropwise at 0°C under nitrogen atmosphere. The resulting mixture was stirred for 1h at 0°C under nitrogen atmosphere. The reaction was monitored by LCMS. The reaction was quenched by the addition of 1M HCl (1 mL) at 0°C. The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford 7-(hydroxymethyl)-3-(oxolan-2-yl)-1H-1,5-naphthyridin-2-one (280 mg, 44.90%). LC-MS: (ES+H, m/z): [M+H] + = 247.00; 1 H NMR (300 MHz, DMSO-d6) δ 11.97 (s, 1H), 8.40 (d, J = 1.9 Hz, 1H), 7.80 (s, 1H), 7.64 (d, J = 1.9 Hz, 1H), 5.48 (t, J = 5.6 Hz, 1H), 4.98 – 4.84 (m, 1H), 4.63 (d, J = 5.3 Hz, 2H), 4.18 – 4.01 (m, 1H), 3.91 – 3.75 (m, 1H), 2.45 – 2.31 (m, 1H), 2.02 – 1.78 (m, 2H), 1.74 – 1.58 (m, 1H). Step 4: Preparation of 7-(chloromethyl)-3-(oxolan-2-yl)-1H-1,5-naphthyridin-2-one: [00227] To a stirred solution of 7-(hydroxymethyl)-3-(oxolan-2-yl)-1H-1,5-naphthyridin-2-one (280 mg, 1.13 mmol, 1.00 equiv.) and DMF (8 mg, 0.11 mmol, 0.10 equiv.) in DCM were added SOCl2 (0.25 mL, 3.41 mmol, 3.00 equiv.) dropwise at 0°C under nitrogen atmosphere. The resulting mixture was stirred for 5h at room temperature under nitrogen atmosphere. The reaction was monitored by LCMS. The resulting mixture was concentrated under reduced pressure, to afford 7-(chloromethyl)-3-(oxolan-2-yl)-1H-1,5- naphthyridin-2-one (300 mg, crude). LC-MS: (ES+H, m/z): [M+H] + =265.05. Step 5: Preparation of 6-(4-{[6-oxo-7-(oxolan-2-yl)-5H-1,5-naphthyridin-3-yl]methyl }piperazin-1- yl)pyridine-3-carbonitrile: [00228] To a stirred mixture of 7-(chloromethyl)-3-(oxolan-2-yl)-1H-1,5-naphthyridin-2-one (150 mg, 0.56 mmol, 1.00 equiv.) and 6-(piperazin-1-yl)pyridine-3-carbonitrile (106 mg, 0.56 mmol, 1.00 equiv.) in MeCN (10 mL) were added DIEA (292 mg, 2.26 mmol, 4.00 equiv.) and KI (18 mg, 0.11 mmol, 0.20 equiv.) at room temperature under nitrogen atmosphere. The resulting mixture was stirred for 2 h at 80°C under nitrogen atmosphere. The mixture was allowed to cool down to room temperature. The reaction was monitored by LCMS. The resulting mixture was extracted with EtOAc (3 x 20 mL). The combined organic layers were washed with brine (20 mL) and dried over anhydrous Na 2 SO 4 . After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by reverse flash chromatography to afford 6- (4-{[6-oxo-7-(oxolan-2-yl)-5H-1,5-naphthyridin-3-yl]methyl}p iperazin-1-yl)pyridine-3-carbonitrile (88 mg, 37.29%). LC-MS: (ES+H, m/z): [M + H] + = 417.10. The racemate (88 mg) was separated by prep-chiral- HPLC with the following conditions: Column: CHIRALPAK IH, 2*25 cm, 5 μm; Mobile Phase A: Hex(10mM NH 3 -MeOH), Mobile Phase B: EtOH: ACN=5: 1; Flow rate: 20 mL/min; Gradient: 50% B to 50% B in 12 min; Wave Length: 218/282 nm; RT1(min): 4.62; RT2(min): 6.76; Sample Solvent: MeOH: DCM=1: 1--HPLC; Injection Volume: 0.75 mL; Number Of Runs: 6, the pure fraction was concentrated under vacuum then lyophilized to afford Example 68 (34.2 mg, 99.7% purity, ee=100%) and Example 69 (34.1 mg, 99.3% purity, ee=99.7%). Example 68: LC-MS: (ES+H, m/z): [M+H] + = 417.10; 1 H NMR (300 MHz, DMSO-d 6 ) δ 11.95 (s, 1H), 8.47 (d, J = 2.4 Hz, 1H), 8.43 (d, J = 1.8 Hz, 1H), 7.84 (dd, J = 9.1, 2.4 Hz, 1H), 7.80 (d, J = 1.3 Hz, 1H), 7.64 (d, J = 1.9 Hz, 1H), 6.93 (d, J = 9.2 Hz, 1H), 4.89 (t, J = 6.8 Hz, 1H), 4.10 – 4.03 (m, 1H), 3.87-3.79 (m, 1H), 3.73 – 3.62 (m, 6H), 2.58 – 2.49 (m, 4H), 2.43 – 2.30 (m, 1H), 2.02 – 1.77 (m, 2H), 1.73 – 1.57 (m, 1H). Example 69: LC-MS: (ES+H, m/z): [M+H] + =417.10; 1 H NMR (300 MHz, DMSO-d6) δ 11.95 (s, 1H), 8.48 (d, J = 2.4 Hz, 1H), 8.43 (d, J = 1.8 Hz, 1H), 7.85 (dd, J = 9.1, 2.4 Hz, 1H), 7.82 – 7.77 (m, 1H), 7.64 (d, J = 1.9 Hz, 1H), 6.93 (d, J = 9.1 Hz, 1H), 4.90 (t, J = 6.8 Hz, 1H), 4.15 – 4.00 (m, 1H), 3.92 – 3.76 (m, 1H), 3.74 – 3.59 (m, 6H), 2.50 – 2.45 (m, 4H), 2.44 – 2.31 (m, 1H), 2.02 – 1.78 (m, 2H), 1.74 – 1.57 (m, 1H). Example 71 Step 1: Preparation of (2E)-N-(3-bromo-2-fluorophenyl)-2-methyl-3-phenylprop-2-enam ide: [00229] A solution of α-methylcinnamic acid (2.56 g, 15.78 mmol, 1.00 equiv.) in DCM (30 mL) was treated with DIEA (8.16 g, 63.15 mmol, 4.00 equiv.), T 3 P (15.07 g, 23.68 mmol, 1.50 equiv., 50wt% in DCM) for 5 min at room temperature under nitrogen atmosphere followed by the addition of 3-bromo-2- fluoroaniline (3.00 g, 15.78 mmol, 1.00 equiv.) at room temperature. The resulting mixture was stirred overnight at room temperature under nitrogen atmosphere. The reaction was monitored by LCMS. The resulting mixture was diluted with water (100 mL). The resulting mixture was extracted with EtOAc (3 x 100 mL). The combined organic layers were washed with brine (2 x 100 mL), dried over anhydrous Na 2 SO 4 . After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford (2E)-N-(3-bromo-2-fluorophenyl)-2-methyl-3-phenylprop-2-enam ide (2.00 g, 37.9%). LC-MS: (ES+H, m/z): [M+H] + =333.95/335.95; 1 H NMR (400 MHz, DMSO-d 6 ) δ 9.89 (s, 1H), 7.62 –7.41 (m, 7H), 7.21 -7.13 (m, 1H), 6.85 –6.69 (m, 1H), 2.12 (d, J=1.4 Hz, 3H). Step 2: Preparation of 7-bromo-8-fluoro-3-methyl-1H-quinolin-2-one: [00230] To a stirred solution (2E)-N-(3-bromo-2-fluorophenyl)-2-methyl-3-phenylprop-2-enam ide (2.00 g, 5.98 mmol, 1.00 equiv.) in chlorobenzene (20 mL) was added AlCl 3 (2.39 g, 17.95 mmol, 3.00 equiv.) at 0°C under nitrogen atmosphere. The resulting mixture was stirred for 3h at 120°C under nitrogen atmosphere. The reaction was monitored by LCMS. The mixture was allowed to cool down to room temperature. The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford 7-bromo-8-fluoro-3-methyl-1H-quinolin-2-one (1.00 g, 65.2%). LC-MS: (ES+H, m/z): [M+H] + = 255.80/257.80; 1 H NMR (400 MHz, DMSO-d6) δ 11.94 (s, 1H), 7.82 (s, 1H), 7.44 -7.34 (m, 2H), 2.09 (s, 3H). Step 3: Preparation of 8-fluoro-7-(hydroxymethyl)-3-methyl-1H-quinolin-2-one: [00231] A solution of 7-bromo-8-fluoro-3-methyl-1H-quinolin-2-one (800 mg, 3.12 mmol, 1.00 equiv.) and (tributylstannyl)methanol (1.10 g, 3.43 mmol, 1.10 equiv.), 2nd Generation XPhos Precatalyst (123 mg, 0.15 mmol, 0.05 equiv.) in 1,4-dioxane (10 mL) was stirred overnight at 80°C under nitrogen atmosphere. The reaction was monitored by LCMS. The mixture was allowed to cool down to room temperature. The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford 8-fluoro-7-(hydroxymethyl)-3-methyl-1H-quinolin-2-one (400 mg, 61.7%). LC- MS: (ES+H, m/z): [M+H] + =208.15 Step 4: Preparation of 7-(chloromethyl)-8-fluoro-3-methyl-1H-quinolin-2-one: [00232] To a stirred solution 8-fluoro-7-(hydroxymethyl)-3-methyl-1H-quinolin-2-one (400 mg, 1.93 mmol, 1.00 equiv.) and DMF (14 mg, 0.19 mmol, 0.10 equiv.) in DCM (5 mL) was added SOCl 2 (2.30 g, 19.30 mmol, 10.00 equiv.) dropwise at 0°C under nitrogen atmosphere. The resulting mixture was stirred for 2h at room temperature under nitrogen atmosphere. The reaction was monitored by LCMS. The resulting mixture was concentrated under reduced pressure to afford 7-(chloromethyl)-8-fluoro-3-methyl-1H- quinolin-2-one (400 mg, 91.8%). LC-MS: (ES+H, m/z): [M+H] + =226.3 Step 5: Preparation of 6-{4-[(8-fluoro-3-methyl-2-oxo-1H-quinolin-7-yl) methyl] piperazin-1-yl} pyridine-3-carbonitrile: [00233] A solution of 6-(piperazin-1-yl) pyridine-3-carbonitrile hydrochloride (229 mg, assumed 100% yield, 0.88 mmol, 1.00 equiv.) in MeCN (10 mL) was treated with DIEA (573 mg, 4.43 mmol, 5.00 equiv.) for 5 min at room temperature under nitrogen atmosphere followed by the addition of KI (17 mg, 0.10 mmol, 0.10 equiv.) and 7-(chloromethyl)-8-fluoro-3-methyl-1H-quinolin-2-one (200 mg, 0.88 mmol, 1.00 equiv.). The resulting mixture was stirred for 2h at 100°C under nitrogen atmosphere. The reaction was monitored by LCMS. The mixture was allowed to cool down to room temperature. The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford 6-{4-[(8-fluoro-3-methyl-2-oxo-1H-quinolin-7-yl) methyl] piperazin-1-yl} pyridine-3-carbonitrile (92.1 mg, 23.8%). LC-MS: (ES+H, m/z): [M+H] + =378.10; 1 H NMR (400 MHz, DMSO-d 6 ) δ 11.77 (s, 1H), 8.47 (d, J = 4.0 Hz, 1H), 7.87 – 7.76 (m, 2H), 7.38 (d, J = 8.1 Hz, 1H), 7.18 (dd, J = 8.1, 6.4 Hz, 1H), 6.91 (d, J = 8.0 Hz, 1H), 3.66 – 3.65 (m, 6H), 2.51-2.48 (m, 4H), 2.10 (d, J = 1.3 Hz, 3H). 19 F NMR (377 MHz, DMSO-d6) δ -135.84. Example 82 Step 1: Preparation of 1-(tert-butyl) 3-ethyl 2-(5-bromo-3-nitropyridin-2-yl)malonate: [00234] To a stirred solution of potassium tert-butoxide (21.27 g, 189.52 mmol, 1.50 equiv.) in THF (500 ml) was added 1-tert-butyl 3-ethyl propanedioate (35.67 g, 189.52 mmol, 1.50 equiv.) dropwise at room temperature under nitrogen atmosphere. To the above mixture was added 5-bromo-2-chloro-3-nitropyridine (30.00 g, 126.34 mmol, 1.00 equiv.) in THF (50 ml) dropwise over 15 min at 60 o C. The resulting mixture was stirred for additional 3h at 60 o C. The reaction was monitored by LCMS. The mixture was allowed to cool down to room temperature. The resulting mixture was concentrated under vacuum. The resulting mixture was diluted with HCl (1M in water) (200 mL). The resulting mixture was extracted with EtOAc (3 x 300 mL). The combined organic layers were washed with brine (2x200 mL), dried over anhydrous Na 2 SO 4 . After filtration, the filtrate was concentrated under reduced pressure, to afford 1-(tert-butyl) 3-ethyl 2-(5- bromo-3-nitropyridin-2-yl)malonate (45 g, 91.5%). LC-MS: (ES-H, m/z): [M-H]- =387.1 Step 2: Preparation of ethyl 2-(5-bromo-3-nitropyridin-2-yl)acetate: [00235] To a stirred solution of 1-(tert-butyl) 3-ethyl 2-(5-bromo-3-nitropyridin-2-yl)malonate (45.00 g, 115.62 mmol, 1.00 equiv.) in CH2Cl2 (300 ml) was added TFA (200 mL) at room temperature under nitrogen atmosphere. The resulting mixture was stirred overnight at room temperature under nitrogen atmosphere. The reaction was monitored by LCMS. The resulting mixture was concentrated under reduced pressure. The residue was diluted with saturated NaHCO3 aqueous solution (100 mL). The resulting mixture was extracted with EtOAc (3 x 500 mL). The combined organic layers were washed with brine (1x200 mL), dried over anhydrous Na 2 SO 4 . After filtration, the filtrate was concentrated under reduced pressure to afford ethyl 2-(5-bromo-3-nitropyridin-2-yl)acetate (35 g, crude). LC-MS: (ES+H, m/z): [M+H] + =290.9 Step 3: Preparation of ethyl 2-(3-amino-5-bromopyridin-2-yl)acetate: [00236] To a stirred solution of ethyl 2-(5-bromo-3-nitropyridin-2-yl)acetate (35.00 g, 121.07 mmol, 1.00 equiv.) and Fe (79.72 g, 1219.36 mmol, 7.50 equiv.) in EtOH (250 ml) were added saturated NH4Cl aqueous solution (250 mL) at room temperature under nitrogen atmosphere. The resulting mixture was stirred for 2h at 80 o C under nitrogen atmosphere. The reaction was monitored by LCMS. The mixture was allowed to cool down to room temperature. The resulting mixture was diluted with EtOAc (300 mL). The resulting mixture was stirred for 10 min at room temperature. The resulting mixture was filtered, the filter cake was washed with EtOAc (3x70 mL). The resulting mixture was extracted with EtOAc (3x300 mL). The combined organic layers were washed with brine (2x200 mL), then dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford ethyl 2-(3-amino-5-bromopyridin-2-yl)acetate (9.3 g, 29.7%) as a light yellow oil. LC-MS: (ES+H, m/z): [M+H] + =259.1 Step 4: Preparation of ethyl 2-(5-bromo-3-(2,2-dimethoxypropanamido)pyridin-2-yl)acetate: [00237] To a stirred solution of methyl 2,2-dimethoxypropanoate (12.00 g, 80.99 mmol, 1.00 equiv.) in MeOH (120 mL) were added NaOH (9.72 g, 242.98 mmol, 3.00 equiv.) in H2O (120 mL) dropwise at room temperature under nitrogen atmosphere. The resulting mixture was stirred for 4h at room temperature under nitrogen atmosphere. The reaction was monitored by TLC. The mixture was acidified to pH 8 with HCl (2M in water). The resulting mixture was concentrated under reduced pressure. The resulting mixture was diluted with EtOH (500mL). The resulting mixture was stirred for 2h at room temperature. The resulting mixture was filtered, the filter cake was washed with EtOH (3x100 mL). The filtrate was concentrated under reduced pressure to afford sodium 2,2-dimethoxypropanoate (8.5 g, 78.2%). 1 H NMR (300 MHz, D 2 O) δ 3.12 (d, J = 1.7 Hz, 6H), 1.33 (s, 3H). [00238] To a stirred solution of sodium 2,2-dimethoxypropanoate (7.59 g, 48.63 mmol, 1.50 equiv.) in dioxane (100 ml) were added DIEA (12.57 g, 97.26 mmol, 3.00 equiv.) and T 3 P (30.95 g, 97.26 mmol, 3.00 equiv, 50 wt% in EtOAc) at room temperature under nitrogen atmosphere. The resulting mixture was stirred for 1h at room temperature under nitrogen atmosphere. To the above mixture was added ethyl 2- (3-amino-5-bromopyridin-2-yl)acetate (8.40 g, 32.42 mmol, 1.00 equiv.) at room temperature. The resulting mixture was stirred overnight at 100 o C. The reaction was monitored by LCMS. The mixture was allowed to cool down to room temperature. The resulting mixture was diluted with water (100 mL). The resulting mixture was extracted with EtOAc (3 x 200 mL). The combined organic layers were washed with brine (1x100 mL), dried over anhydrous Na 2 SO 4 . After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford ethyl 2-(5-bromo- 3-(2,2-dimethoxypropanamido)pyridin-2-yl)acetate (9.3 g, 79.4%). LC-MS: (ES+H, m/z): [M+H] + =375.1 Step 5: Preparation of ethyl 7-bromo-3-methyl-2-oxo-1,2-dihydro-1,5-naphthyridine-4-carbo xylate: [00239] To a stirred solution of ethyl 2-(5-bromo-3-(2,2-dimethoxypropanamido)pyridin-2-yl)acetate (7.00 g, 18.65 mmol, 1.00 equiv.) in TFA(100 ml) were added H 2 O (7 mL) and followed by 2 drops of I2 fresh solution (30 mg of I2 was suspended in TFA (10 mL)) at room temperature under nitrogen atmosphere. The resulting mixture was stirred overnight at 50 o C under nitrogen atmosphere. The resulting mixture was concentrated under vacuum. The residue was dissolved in toluene (100 mL) and piperidine (6 mL). The resulting mixture was refluxed at 120 o C for 2h, then concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford ethyl 7-bromo-3-methyl-2-oxo-1H-1,5- naphthyridine-4-carboxylate (3.3 g, 56.9%). LC-MS: (ES+H, m/z): [M+H] + =313.0; 1 H NMR (400 MHz, DMSO-d6) δ 12.20 (s, 1H), 8.55 (d, J = 2.1 Hz, 1H), 7.86 (d, J = 2.1 Hz, 1H), 4.42 (q, J = 7.1 Hz, 2H), 2.06 (s, 3H), 1.33 (t, J = 7.1 Hz, 3H). Step 6: Preparation of 7-bromo-4-(hydroxymethyl)-3-methyl-1,5-naphthyridin-2(1H)-on e: To a stirred solution of ethyl 7-bromo-3-methyl-2-oxo-1H-1,5-naphthyridine-4-carboxylate (3.00 g, 9.64 mmol, 1.00 equiv.) in THF (50 ml) were added LiEt3BH (30 ml, 1M in THF) dropwise at 0 o C under nitrogen atmosphere. The resulting mixture was stirred for 2h at 0 o C under nitrogen atmosphere. The reaction was monitored by LCMS. The reaction was quenched by the addition of Water/ice (6 mL) at 0 o C. The resulting mixture was concentrated under vacuum. The residue was purified by silica gel column chromatography, eluted with CH2Cl2 / MeOH (0 to 10:1 gradient in 30 min) to afford 7-bromo-4- (hydroxymethyl)-3-methyl-1,5-naphthyridin-2(1H)-one (1.8 g, 69.4%). LC-MS: (ES-H, m/z): [M-H]- =267.0/269.0; 1 H NMR (300 MHz, DMSO-d6) δ 11.95 (s, 1H), 8.57 (d, J = 2.1 Hz, 1H), 7.83 (d, J = 2.1 Hz, 1H), 5.04 (t, J = 5.5 Hz, 1H), 4.90 (d, J = 5.2 Hz, 2H), 2.21 (s, 3H). Step 7: Preparation of 7-bromo-3-methyl-2-oxo-1,2-dihydro-1,5-naphthyridine-4-carba ldehyde: [00240] To a stirred mixture of 7-bromo-4-(hydroxymethyl)-3-methyl-1,5-naphthyridin-2(1H)-on e (1.80 g, 6.68 mmol, 1.00 equiv.) in THF (20 ml) was added 1,1-bis(acetyloxy)-3-oxo-3H-1l^[5],2-benziodaoxol-1-yl acetate (3.40 g, 8.02 mmol, 1.20 equiv.) at room temperature under nitrogen atmosphere. The resulting mixture was stirred for 2h at room temperature under nitrogen atmosphere. The reaction was monitored by LCMS. The resulting mixture was diluted with water (10 mL). The precipitated solids were collected by filtration and washed with water (3x5 mL). The residue was purified by reversed combi-flash chromatography to afford 7-bromo-3-methyl-2-oxo-1,2-dihydro-1,5-naphthyridine-4-carba ldehyde (760 mg, 42.5%).LC-MS: (ES-H, m/z): [M-H]- = 264.9; 1 H NMR (300 MHz, DMSO-d 6 ) δ 12.28 (s, 1H), 10.88 (s, 1H), 8.62 (d, J = 2.1 Hz, 1H), 7.89 (d, J = 2.1 Hz, 1H), 2.25 (s, 3H). Step 8: Preparation of 7-bromo-4-(difluoromethyl)-3-methyl-1,5-naphthyridin-2(1H)-o ne: [00241] To a stirred solution of 7-bromo-3-methyl-2-oxo-1,2-dihydro-1,5-naphthyridine-4-carba ldehyde (750 mg, 2.80 mmol, 1 equiv.) in THF (5 ml) were added BAST (2.49 g, 11.23 mmol, 4 equiv.) dropwise at 0 o C under nitrogen atmosphere. The resulting mixture was stirred for 2h at 50 o C under nitrogen atmosphere. The reaction was monitored by LCMS. The mixture was allowed to cool down to room temperature. The reaction was quenched with sat. NH4Cl (aq.) (3 ml) at 0 o C. The resulting mixture was extracted with EtOAc (3 x 20 mL), dried over anhydrous Na 2 SO 4 . After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford 7-bromo-4- (difluoromethyl)-3-methyl-1H-1,5-naphthyridin-2-one (320 mg, 39.4%). LC-MS: (ES-H, m/z): [M-H]- =287.0 Step 9: Preparation of 4-(difluoromethyl)-7-(hydroxymethyl)-3-methyl-1,5-naphthyrid in-2(1H)-one: [00242] To a stirred solution of 7-bromo-4-(difluoromethyl)-3-methyl-1H-1,5-naphthyridin-2-on e (360 mg, 1.24 mmol, 1.00 equiv.) and 2nd Generation XPhos Precatalyst/ (195 mg, 0.24 mmol, 0.20 equiv.) in dioxane (5 ml) were added (Tributylstannyl)methanol (959 mg, 2.98 mmol, 2.40 equiv.) at room temperature under nitrogen atmosphere. The resulting mixture was stirred for 2h at 80 o C under nitrogen atmosphere. The reaction was monitored by LCMS. The resulting mixture was concentrated under vacuum. The residue was purified by silica gel column chromatography to afford 4- (difluoromethyl)-7-(hydroxymethyl)-3-methyl-1,5-naphthyridin -2(1H)-one (195 mg, 65.2%). LC-MS: (ES+H, m/z): [M+H] + =241.1. Step 10: Preparation of 7-(chloromethyl)-4-(difluoromethyl)-3-methyl-1,5-naphthyridi n-2(1H)-one: [00243] To a stirred solution of 4-(difluoromethyl)-7-(hydroxymethyl)-3-methyl-1,5-naphthyrid in-2(1H)- one (185 mg, 0.25 mmol, 1.00 equiv.) and DMF (2 mg, 0.02 mmol, 0.10 equiv.) in CH2Cl2 (3 ml) were added SOCl2 (183 mg, 1.54 mmol, 6.00 equiv.) dropwise at 0 o C under nitrogen atmosphere. The resulting mixture was stirred overnight at room temperature under nitrogen atmosphere. The reaction was monitored by LCMS. The resulting mixture was concentrated under vacuum to afford 7-(chloromethyl)-4- (difluoromethyl)-3-methyl-1,5-naphthyridin-2(1H)-one (195 mg, crude). The crude product was used in the next step directly without further purification. LC-MS: (ES+H, m/z): [M+H] + =259.0 Step 11: Preparation of 6-(4-((8-(difluoromethyl)-7-methyl-6-oxo-5,6-dihydro-1,5-nap hthyridin-3- yl)methyl)piperazin-1-yl)nicotinonitrile: [00244] To a stirred solution of 7-(chloromethyl)-4-(difluoromethyl)-3-methyl-1,5-naphthyridi n-2(1H)-one (104 mg, 0.40 mmol, 1.00 equiv.) and 6-(piperazin-1-yl)pyridine-3-carbonitrile (91 mg, 0.48 mmol, 1.20 equiv.) in MeCN(5 ml) were added DIEA (260 mg, 2.01 mmol, 5.00 equiv.) and KI (13 mg, 0.08 mmol, 0.20 equiv.) at room temperature under nitrogen atmosphere. The resulting mixture was stirred for 3h at 80 o C under nitrogen atmosphere. The reaction was monitored by LCMS. The mixture was allowed to cool down to room temperature. The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford 6-(4-((8-(difluoromethyl)-7-methyl-6-oxo-5,6- dihydro-1,5-naphthyridin-3-yl)methyl)piperazin-1-yl)nicotino nitrile (crude). The crude product was further purified by trituration with MeOH (6 mL) to afford 6-(4-((8-(difluoromethyl)-7-methyl-6-oxo-5,6-dihydro- 1,5-naphthyridin-3-yl)methyl)piperazin-1-yl)nicotinonitrile (28.3 mg, 16.6%). LC-MS: (ES+H, m/z): [M+H] + =411.10; 1 H NMR (300 MHz, DMSO-d 6 ) δ 12.25 (s, 1H), 8.48 (t, J = 1.8 Hz, 2H), 8.14 – 7.74 (m, 2H), 7.70 (d, J = 1.9 Hz, 1H), 6.94 (d, J = 9.1 Hz, 1H), 3.39-3.67 (m, 6H), 2.50-2.48 (m, 4H), 2.33 (t, J = 2.8 Hz, 3H). 19 F NMR (282 MHz, DMSO-d 6 ) δ-117.71. Example 83 Step 1: Preparation of (2E)-N-(3-bromo-2-fluorophenyl)-3-ethoxyprop-2-enamide: [00245] To a stirred mixture of 3-bromo-2-fluoroaniline (20.00 g, 105.25 mmol, 1.00 equiv.) in DCM (300 mL) was added pyridine (14.99 g, 189.45 mmol, 1.80 equiv.) dropwise at room temperature under nitrogen atmosphere. The resulting mixture was stirred for 5 min at room temperature under nitrogen atmosphere. To the above mixture was added (2E)-3-ethoxyprop-2-enoyl chloride (21.24 g, 157.88 mmol, 1.50 equiv.) dropwise over 5 min at room temperature. The resulting mixture was stirred for additional 2h at room temperature. The reaction was monitored by LCMS. The resulting mixture was concentrated under reduced pressure. The residue was dissolved in EtOAc (500 mL). The resulting mixture was washed with water (3x500 mL). The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford (2E)-N-(3-bromo-2-fluorophenyl)-3-ethoxyprop-2-enamide (24.6 g, 81.1%). LC-MS: (ES+H, m/z): [M+H] + =288.0/290.0. Step 2: Preparation of 7-bromo-8-fluoro-1H-quinolin-2-one: [00246] A mixture of (2E)-N-(3-bromo-2-fluorophenyl)-3-ethoxyprop-2-enamide (17.00 g, 59.00 mmol, 1.00 equiv.) in H 2 SO 4 (85 mL) was stirred for 3h at room temperature under nitrogen atmosphere. The resulting mixture was added to ice water (1L) dropwise and stirred for 1h. The precipitated solids were collected by filtration and washed with water (3x200 mL). The resulting mixture was concentrated under reduced pressure to afford 7-bromo-8-fluoro-1H-quinolin-2-one (14.30 g, crude). LC-MS: (ES+H, m/z): [M+H] + =242.0/244.0. Step 3: Preparation of 7-bromo-3-chloro-8-fluoro-1H-quinolin-2-one: [00247] To a stirred mixture of 7-bromo-8-fluoro-1H-quinolin-2-one (3.00 g, 12.39 mmol, 1.00 equiv.) and NCS (2.65 g, 19.83 mmol, 1.60 equiv.) in CH3COOH (50 mL) was added 2,2-dichloroacetic acid (0.32 g, 2.47 mmol, 0.20 equiv.) dropwise at room temperature under nitrogen atmosphere. The resulting mixture was stirred overnight at 100°C under nitrogen atmosphere. The reaction was monitored by LCMS. The mixture was allowed to cool down to room temperature. The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford 7-bromo-3- chloro-8-fluoro-1H-quinolin-2-one (2.48 g, crude). LC-MS: (ES+H, m/z): [M+H] + =275.9/277.9; 1 H NMR (400 MHz, DMSO-d6) δ 12.52 (s, 1H), 8.38 (d, J = 1.6 Hz, 1H), 7.52 – 7.42 (m, 2H). Step 4: Preparation of 3-chloro-7-ethenyl-8-fluoro-1H-quinolin-2-one: [00248] To a stirred mixture of 7-bromo-3-chloro-8-fluoro-1H-quinolin-2-one (2.48 g, 8.97 mmol, 1.00 equiv.), CsF (4.09 g, 26.91 mmol, 3.00 equiv.), Pd(dppf)Cl 2 (0.33 g, 0.44 mmol, 0.05 equiv.) and 2-ethenyl- 4,4,5,5-tetramethyl-1,3,2-dioxaborolane (1.38 g, 8.97 mmol, 1.00 equiv.) in dioxane (50 mL) was added H 2 O (5 mL) dropwise at room temperature under nitrogen atmosphere. The resulting mixture was stirred for 2h at 100°C under nitrogen atmosphere. The reaction was monitored by LCMS. The mixture was allowed to cool down to room temperature. The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford 3-chloro-7-ethenyl-8-fluoro-1H- quinolin-2-one (750 mg, 37.3%). LC-MS: (ES+H, m/z): [M+H] + =224.0; 1 H NMR (300 MHz, DMSO-d6) δ 12.38 (s, 1H), 8.34 (d, J = 1.6 Hz, 1H), 7.53-7.46 (m, 2H), 6.95 (dd, J = 17.7, 11.2 Hz, 1H), 6.07 (dd, J = 17.7, 1.0 Hz, 1H), 5.57 (dd, J = 11.2, 1.0 Hz, 1H). Step 5: Preparation of 3-chloro-8-fluoro-2-oxo-1H-quinoline-7-carbaldehyde: [00249] To a stirred mixture of 3-chloro-7-ethenyl-8-fluoro-1H-quinolin-2-one (750 mg, 3.35 mmol, 1.00 equiv.), K2OsO2(OH)4 (123 mg, 0.33 mmol, 0.10 equiv.), NaIO4 (2.87 g, 13.41 mmol, 4.00 equiv.) and 2,6- dimethylpyridine (718 mg, 6.70 mmol, 2.00 equiv.) in THF (15 mL) was added H2O (1.5 mL) dropwise at room temperature under nitrogen atmosphere. The resulting mixture was stirred overnight at room temperature under nitrogen atmosphere. The reaction was monitored by LCMS. The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford 3-chloro-8-fluoro-2-oxo-1H-quinoline-7-carbaldehyde (630 mg, 83.2%). LC-MS: (ES-H, m/z): [M- H]- =224.1. Step 6: Preparation of 6-{4-[(3-chloro-8-fluoro-2-oxo-1H-quinolin-7-yl)methyl]piper azin-1- yl}pyridine-3-carbonitrile: [00250] A mixture of 3-chloro-8-fluoro-2-oxo-1H-quinoline-7-carbaldehyde (150 mg, 0.66 mmol, 1.00 equiv.) and 6-(piperazin-1-yl)pyridine-3-carbonitrile (137 mg, 0.73 mmol, 1.10 equiv.) in DCM(2 ml) was stirred for 10 min at room temperature. The resulting mixture was concentrated under reduced pressure. The resulting mixture was added HOAc (19 mg, 0.33 mmol, 0.50 equiv.) in EtOH (3 mL) and stirred for 4h at 50°C under nitrogen atmosphere. The mixture was allowed to cool down to room temperature. To the stirred mixture was added NaBH 3 CN (83 mg, 1.33 mmol, 2.00 equiv.) in portions at 0°C under nitrogen atmosphere. The resulting mixture was stirred overnight at room temperature under nitrogen atmosphere. The precipitated solids were collected by filtration and washed with EtOH (3x2 mL). The residue was purified by silica gel column chromatography. The resulting mixture was concentrated under reduced pressure to afford 6-{4-[(3-chloro-8-fluoro-2-oxo-1H-quinolin-7-yl)methyl]piper azin-1-yl}pyridine-3- carbonitrile (69.3 mg, 26.2%). LC-MS: (ES+H, m/z): [M+H] + =398.10; 1 H NMR (400 MHz, DMSO-d 6 ) δ 12.46 (s, 1H), 8.47 (d, J = 2.0 Hz, 1H), 8.36 (d, J = 1.5 Hz, 1H), 7.85 (dd, J = 9.1, 2.4 Hz, 1H), 7.49 (d, J = 8.1 Hz, 1H), 7.28 (dd, J = 8.1, 6.3 Hz, 1H), 6.97 – 6.87 (m, 1H), 3.71 – 3.63 (m, 6H), 2.50 – 2.44 (m, 4H); 19 F NMR (282 MHz, DMSO-d6) δ -134.50. [00251] The following compound were synthesized as described above. Table 2. Characterization Data
Example A: Cell Growth Inhibition Assay [00252] Cell proliferation was measured through the cell viability assay using DLD-1 BRCA2(-/-) and parental isogenic pair and MDA-MB-436 (mutated BRCA1) cell lines. The CellTiter-Glo (CTG) based cell viability assay is designed to determine the number of viable cells in the culture because of compound effect, by quantifying ATP, which indicates the presence of metabolically active cells. [00253] DLD-1 BRCA2(-/-) and parental isogenic pair were cultured in RPMI 1640 supplemented with 10% fetal bovine serum (FBS), and MDA-MB-436 cells were cultured in DMEM supplemented with 10% FBS. Both were cultured at 37℃ with 5% CO2. Invention compounds were distributed to the 384 well plate (Corning, 3764) using Echo acoustic liquid handler to form a 1:3 serially diluted final concentration with top dose of 10 or 30 µM. The cells were seeded into the plate in the density of 50 cells/well (DLD-1 parental), 200 cells/well (DLD-1 BRCA2-/-), or 500 cells/well (MDA-MB-436). After a short spun, the cells were cultured in a well moisturized incubator at 37℃ with 5% CO2 for 7 days without disturbance. The cell viability was measured by CellTiter Glo 2.0 assay kit (Promega, G9243), and growth inhibition rate was calculated and plotted against final compound concentration, and the data were fitted in Xfit to generate IC50 values. Example B: Biochemical (FP) Assay [00254] Assays based on fluorescent polarization (FP) have been widely utilized in drug discovery due to the homogenous format, robust performance and lack of interference seen in other assays.. Compounds were characterized using an assay measuring the displacement of a commercially available fluorescently labeled PARP 1/2 inhibitor (PARPi-FL, Tocris Biosciences, #6461) as exemplified in assays performed in WO2014/064149 and WO2021/013735A1. The assay was performed utilizing the following method: [00255] Compounds were dissolved in DMSO an Echo550 liquid handler was utilized to make serial dilations in the desired concentration range in Optiplate-384F plates.100% DMSO was used for the high (with protein) and low (without protein) control samples.20nL of compound or DMSO alone was added to individual assay plate wells. [00256] PARP1 and PARP2 protein were expressed, purified, and diluted in assay buffer containing 50 mM Tris pH 8.0, 0.001% Triton X-100, 10 mM MgCl 2 , 150 mM NaCl to a final concentration of 20nM. The PARPi-FL was then added at a final concentration of 3nM. [00257] The assay plate was centrifuged at 1000rpm for 1 min and incubated for 4 h at room temperature. [00258] The fluorescent polarization was read using an Envision plate reader using the following settings: Excitation filter - FITC FP 480-Ex Slot 3 Emission filter - FITC FP P-pol 535-Em Slot 4 2nd Emission filter - FITC FP S-pol 535-Em Slot 3 Mirror module - FITC FP Dual Enh-Slot 1 [00259] The inhibition rate was calculated using the percentage of permuted Mahalanobis distances greater than the control samples (mP value) following the equation below: [00260] XLFit (equation 201) is used to calculate a reported IC 50 for each compound. [00261] The data from examples A and B are provided in Table 3. TABLE 3
Example C: In Vitro Human Transporter Efflux [00262] Madin-Darby canine kidney (MDCKII) cells expressing MDR1 and BCRP were seeded onto Corning HTS Transwell® 96-well polycarbonate permeable (0.4 µm pore) supports at a density of 545,000 cells/cm2. Cells were incubated for 4-8 days prior to assay, and monolayer integrity was assessed by measuring transepithelial electrical resistance (TEER). Test and reference compounds were diluted with the transport buffer (HBSS HEPES pH7.4) to concentrations of 10 and 1 µM, respectively. The final organic solvent concentration was 0.5% (v/v). Bidirectional (apical-to-basolateral and basolateral-to-apical) flux of the test and reference compounds was determined over a 2-hour incubation at 37°C and 5% CO2 with a relative humidity of 95%. At the end of the incubation, samples from the apical and basolateral side were taken and then precipitated with acetonitrile containing internal standard. After centrifugation at 3200 x g, supernatants were diluted 1:1 (v/v) with water and subjected to analysis via HPLC-MS/MS. The integrity [00263] The apparent permeability (Papp, units of x 10-6 cm/s) was calculated using the following equation: Papp =(dQ/dt)/(AxD0) Where dQ/dt was the rate of drug transport (pmol/s), A was the surface area of the membrane (0.143 cm 2 ), and D0 was the initial donor concentration (nM or pmol/cm3). Efflux ratio = Papp(B→A) / Papp(A→B) Where Papp(B→A) is the apparent permeability in the basolateral-to-apical direction, and Papp(A→B) is the apparent permeability in the apical-to-basolateral direction. Example D: In vivo determination of rat Kp,uu [00264] Determination of Fraction Unbound in Plasma (Pu) [00265] The equilibrium dialysis method was used to investigate the in vitro binding of test articles and reference compounds to plasma proteins. Plasma samples containing 5 μM test article or blank dialysis buffer solution (PBS, pH 7.4) were added to separate chambers of the dialysis wells of the High Throughput equilibrium Dialysis (HTD) device. The dialysis plate was sealed and placed in an incubator at 37°C with 5% CO2 with shaking at approximately 100 rpm for 6 hours. All experiments were performed in duplicate. Ketoconazole (5 μM) was used as the reference compound. After incubation, the seal was removed, and 50 μL of post-dialysis samples were pipetted from both buffer and plasma chambers into fresh 96-well plates. Samples were equimatrilyzed by either addition of blank plasma to buffer samples or the addition of blank buffer to plasma samples. Subsequently, 400 μL (4 volumes) of acetonitrile containing internal were added to all samples to precipitate proteins prior to analysis by UPLC-MS/MS to determine the relative concentrations of test articles. The unbound fractions in plasma were calculated using the concentrations of test articles in buffer and plasma samples according to the following equation: [00266] Determination of Fraction Unbound in Brain Homogenate (Bu) [00267] The equilibrium dialysis method was used to investigate the in vitro binding of test articles and reference compounds to rodent brain homogenate. Brains collected from naïve animals were weighed and homogenized in 4 volumes of PBS, pH 7.4. Brain homogenate samples containing 1 μM test article or blank dialysis buffer solution (PBS, pH 7.4) were added to separate chambers of the dialysis wells of the High Throughput equilibrium Dialysis (HTD) device. The dialysis plate was sealed and placed in an incubator at 37°C with 5% CO2 with shaking at approximately 100 rpm for 6 hours. All experiments were performed in duplicate. Telmisartan (5 μM) was used as the reference compound. After incubation, the seal was removed, and 50 μL of post-dialysis samples were pipetted from both buffer and brain homogenate chambers into fresh 96-well plates. Samples were equimatrilyzed by either addition of blank homogenate to buffer samples or the addition of blank buffer to homogenate samples. Subsequently, 400 μL (4 volumes) of acetonitrile containing internal were added to all samples to precipitate proteins prior to analysis by UPLC-MS/MS to determine the relative concentrations of test articles. The unbound fractions in diluted brain homogenate were calculated using the concentrations of test articles in buffer and homogenate samples according to the following equation: Percentage unbound homogenate (%) = x 100 [00268] Correction to percentage unbound in undiluted brain was achieved with the following equation: g ( ) [00269] Determination of the Drug Brain-to-Plasma Partition Coefficient (Kp) and Drug Unbound Kp (Kp,uu) in Rat [00270] Compounds were formulated either individually or in a cassette (as a mixture) at a concentration of 0.1 mg/mL/compound in sterile water containing 0.5% (w/v) methylcellulose 400 cP and administered to male Sprague-Dawley rats via oral gavage at a dose volume of 10 mL/kg. One animal was sacrificed at each of the following time points: 0.5, 1, 2, 4, 8, and 24 hours post-dose, and brain and blood samples were collected. Plasma was prepared from blood via refrigerated centrifugation, and plasma samples were stored frozen at -80°C until bioanalysis. Brain samples were rinsed with saline to remove residual blood and blotted dry with a paper wipe. Brain samples were then weighed and homogenized with 3 volumes (v/w) of water and stored frozen at -80°C until bioanalysis. [00271] Prior to bioanalysis, plasma and brain samples were extracted with 4 volumes of acetonitrile containing internal standard and centrifuged for 15 minutes. Supernatants were diluted with 2 volumes of water and injected for analysis via HPLC-MS/MS. Plasma and brain homogenate drug concentrations were determined against calibration curves generated by spiking blank rat plasma or brain homogenate with drug across an appropriate concentration range. The brain homogenate concentration was corrected for the homogenization buffer dilution factor yielding total brain drug concentrations. [00272] The brain-to-plasma partition coefficient (Kp) was determined for each compound, calculated as: AUCbrain:AUCplasma, provided tlast was identical in each matrix. If the drug concentration versus time profile for one matrix fell below the lower limit of quantification at a time point earlier than in the other matrix, then the brain Kp was calculated as the average of the ratios of total brain drug concentration to total plasma drug concentration measured at each time point where drug concentrations in both matrices were quantifiable. [00273] The Kp,uu was then calculated from the Kp using the following equation: Kp,uu = Kp * (fraction unbound in brain homogenate/fraction unbound in plasma). [00274] The data from examples C and D are provided in Table 4. Table 4
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