Heterocyclic PLA2g15 inhibitors and their use in therapy, in the treatment of diseases characterized by lysosomal dysregulation

Abstract

The current invention relates to PLA2G15 inhibitors represented by formula (I), and corresponding compositions and uses. Preferably, the inhibitors and compositions are for use in the treatment of lysosomal storage diseases, Alzheimer's disease and Parkinson's disease; in particular for use in the treatment of Niemann Pick type C or a neuronal ceroid lipofuscinosis such as CLN3 disease or Batten disease, CLN5 disease, or GRN frontotemporal dementia.

Description

[0001] HETEROCYCLIC PLA2G15 INHIBITORS AND THEIR USE IN THERAPY, IN THE TREATMENT OF DISEASES CHARACTERIZED BY LYSOSOMAL

[0002] Field DYSREGULATION

[0003] The current invention relates to PLA2G15 inhibitors represented by formula (I), and corresponding compositions and uses. Preferably, the inhibitors and compositions are for use in the treatment of lysosomal storage diseases, HIV, Alzheimer’s disease and Parkinson’s disease; in particular for use in the treatment of Niemann Pick type C or a neuronal ceroid lipofuscinosis such as CLN3 disease or Batten disease, CLN5 disease, or GRN frontotemporal dementia.

[0004] Background of the invention

[0005] PLA2G15 proteins are phospholipase A2 group XV enzymes that occur naturally in humans. These proteins have been identified as clinically relevant, for example for their role in drug-induced phospholipidosis (Hinkovska-Galcheva, Vania, et al. "Inhibition of lysosomal phospholipase A2 predicts drug-induced phospholipidosis." Journal of lipid research 62 (2021)). Nonetheless, few potent and / or selective PLA2G15 inhibitors have been identified to the best of our knowledge.

[0006] Simona Di Martino et al. ‘Lead Optimization of Benzoxazoione Carboxamides as Oraiiy Bioavaiiabie and CNS Penetrant Acid Ceramidase Inhibitors’ Journal of Medicinal Chemistry (2020) discloses acide ceramidase (AC) inhibitors for use in the treatment of lysosomal storage diseases such as Gaucher disease.

[0007] Edward A. Dennis et al. ‘Phospholipase A 2 Enzymes: Physical Structure, Biological Function, Disease Implication, Chemical Inhibition, and Therapeutic Intervention’ (2011) discloses the phospholipase A2 superfamily, in particular group XV and chemical inhibitors such as darapladib.

[0008] US2009 / 324575A1 discloses therapeutic implication of lysosomal phospholipase A2 group XV (LPLA2) and inhibitors thereof.

[0009] WO2014 / 141035A2 discloses heterocycle derivatives which are NAMPT inhibitors which are useful in the treatment or prevention of diseases or disorder caused by an elevated level of nicotinamide phosphoribosyltransferase.

[0010] W02014 / 065270A1 discloses tetrahydrooxazolo-pyridine derivatives which are useful in the treatment of Parkinson disease or Alzheimer's disease.

[0011] Hence, there is a continuing need in the art for novel potent and / or selective PLA2G15 inhibitors.

[0012] Description of the invention

[0013] Inhibitors In an aspect, the invention provides a PLA2G15 inhibitor represented by formula (I), or a salt or solvate thereof:

[0014] (I), wherein X is S or O; wherein Y is CH or N; wherein L is a single bond or a linker moiety; wherein RXYis H, a C1-4 alkyl, a halogen, or a pseudohalogen selected from the group consisting of -CN, -CP, -NC, -OH, -SH, -SeH,-TeH, -OCN, -SCN, -NCS, -SeCN, -TeCN, -N3, -NO, or -NO; wherein RNis a C1-4 alkyl, a C3-6 cycloalkyl, a 5-membered aromatic ring, or a 6-membered aromatic ring; wherein RLcomprises a 3- to 6-membered carbocyclic ring or a 4-membered heterocyclic ring; wherein each 5-membered aromatic ring and 6-membered aromatic ring may be independently substituted with one or more halogens, one or more pseudohalogens selected from the group consisting of -CN, -CP, -NC, -OH, -SH, -SeH,-TeH, -OCN, -SCN, -NCS, -SeCN, -TeCN, -N3, -NO, or -NO, C1-4 alkyls, O-C1-4 alkyls, C3-6 cycloalkyls, or O-C3-6 cycloalkyls; and wherein each C1-4 alkyl and C3-6 cycloalkyl may be independently substituted with one or more halogens or one or more pseudohalogens selected from the group consisting of -CN, -CP, -NC, -OH, - SH, -SeH,-TeH, -OCN, -SCN, -NCS, -SeCN, -TeCN, -N3, -NO, or -NO.

[0015] In a second aspect, the invention provides a PLA2G15 inhibitor, or a salt or solvate thereof, as defined in the first aspect, for use as a medicament.

[0016] In a third aspect, the invention provides a PLA2G15 inhibitor, or a salt or solvate thereof, as defined in the first aspect, for use in the treatment of a disease characterized by lysosomal dysregulation.

[0017] Inhibitors according to the first, the second aspect or the third aspect are referred to in the current application as (PLA2G15) inhibitors according to or of the invention. Wherever a(n) (PLA2G15) inhibitor is mentioned in this application, reference is made to an inhibitor according to the invention, unless explicitly mentioned otherwise.

[0018] Wherever embodiments and preferences are disclosed in this application, for example in relation to X, Y, RXY, RN, L or RL, explicit reference is made to inhibitors according to the first, the second and the third aspect of the invention. In other words, such embodiments and preferences relate to the inhibitors, and to the inhibitors for use as a medicament, and to the inhibitors for use in the treatment of a disease characterized by lysosomal dysregulation. Relatedly, any embodiment aimed at inhibitors mentioned herein also discloses an embodiment for the inhibitors for use as a medicament and an embodiment for the inhibitors for use in the treatment of a disease characterized by lysosomal dysregulation. Likewise, such embodiments and preferences may be applied mutatis mutandis to related compositions, uses, therapeutic uses, etc. of the inhibitors according to the invention.

[0019] A halogen is F, Cl, Br, I, or At. Preferably, a halogen is F, Cl, Br, or I. More preferably, a halogen is F, Cl, or Br. Even more preferably, a halogen is F or Cl. Most preferably, a halogen is F.

[0020] A pseudohalogen is -CN, -CP, -NC, -OH, -SH, -SeH,-TeH, -OCN, -SCN, -NCS, -SeCN, -TeCN, -N3, -NO, or -NO2. Preferably, a pseudohalogen is -CN, -NC, -OH, -SH, -OCN, -SCN, -NCS, -N3, -NO, or - NO2.

[0021] LINKER L AND FORMULAE (II) AND (IV)

[0022] L may be a single bond. This means that carbonyl group explicitly drawn in formula (I) is directly to RL.

[0023] In embodiments, the linker moiety is an organic linker.

[0024] In embodiments, the linker moiety comprises 1 to 20, 1 to 19, 1 to 18, 1 to 17, 1 to 16, 1 to 15, 1 to 14, 1 to 13, 1 to 12, 1 to 11 , 1 to 10, 5 to 20, 5 to 19, 5 to 18, 5 to 17, 5 to 16, 5 to 15, 5 to 14, 5 to 13, 5 to 12, 5 to 11 , or 5 to 10 non-hydrogen atoms.

[0025] In embodiments, the linker moiety comprises a C=O, S(=O) or S(=O)2 group. Preferably, the C=O, S(=O) or S(=O)2group is covalently bound to RL.

[0026] In embodiments, the linker moiety comprises a carbon atom covalently bound to the carbonyl group explicitly drawn in formula (I).

[0027] In embodiments, the PLA2G15 inhibitor is represented by formula (II):

[0028] (II), wherein RAand RBare independently H, a C1-4 alkyl, a halogen, or a pseudohalogen, or wherein RAand RBare fused to form a C3-6 cycloalkyl; wherein Z is O or NR’, wherein R’ is H or a C1-4 alkyl; wherein i is 0 or 1 ; and wherein each C1-4 alkyl and C3-6 cycloalkyl may be independently substituted with one or more halogens or pseudohalogens.

[0029] In embodiments, the PLA2G15 inhibitor is represented by formula (II), and i is 1 . Preferably, Z is O or NH or NCH3, more preferably Z is O or NH.

[0030] In embodiments, the PLA2G15 inhibitor is represented by formula (II), i is 1 , and Z is NH.

[0031] In embodiments, RAand RBare independently H, a C1-4 alkyl, a halogen, or a pseudohalogen. Preferably, RAand RBare independently H, or a C1-4 alkyl. More preferably, RAand RBare independently H, or a C1-3 alkyl.

[0032] In embodiments, RAand RBare independently H or CH3. Preferably, RAand RBare H.

[0033] In embodiments, the PLA2G15 inhibitor is represented by formula (II), i is 1 , Z is NH, and RAand RBare independently H, a C1-4 alkyl, a halogen, or a pseudohalogen. Preferably RAand RBare independently H, or a C1-4 alkyl. More preferably, RAand RBare independently H, or a C1-3 alkyl. Even more preferably, RAand RBare independently H or CH3. Most preferably, RAand RBare H.

[0034] Wherever RAand RBin formula (II) are not the same, the carbon atom bound to RAand RBis a chiral center. In the particular case wherein RBis H and RAis not H, formula (II) may be depicted as formula (III) or (lll#):

[0035] In embodiments, the PLA2G15 inhibitor is represented by formula (III) or formula (lll#). Preferably, the PLA2G15 inhibitor is represented by formula (III).

[0036] In embodiments, the PLA2G15 inhibitor is represented by formula (III) or formula (lll#), wherein RAis H, a C1-4 alkyl, a halogen, or a pseudohalogen. Preferably, the PLA2G15 inhibitor is represented by formula (III).

[0037] In embodiments, the PLA2G15 inhibitor is represented by formula (III) or formula (lll#), wherein RAis H, or a C1-4 alkyl. Preferably, the PLA2G15 inhibitor is represented by formula (III).

[0038] In embodiments, the PLA2G15 inhibitor is represented by formula (III) or formula (lll#), wherein RAis H, or a C1-4 alkyl. Preferably, the PLA2G15 inhibitor is represented by formula (III).

[0039] In embodiments, the PLA2G15 inhibitor is represented by formula (III) or formula (lll#), wherein RAis H, or a C1-3 alkyl. Preferably, the PLA2G15 inhibitor is represented by formula (III). In embodiments, the PLA2G15 inhibitor is represented by formula (III) or formula (lll#), wherein RAis CH3. Preferably, the PLA2G15 inhibitor is represented by formula (III).

[0040] In embodiments, the linker moiety comprises a five- or six-membered ring, preferably a single five- or six-membered ring. Preferably, the five- or six-membered ring is saturated or partially saturated.

[0041] In embodiments, the linker moiety comprises a cyclopentyl, a cyclohexyl, a pyridinyl or a pyrrolidinyl.

[0042] In embodiments, the linker moiety comprises a C=O, S(=O) or S(=O)2 group, preferably a S(=O)2 group, and the linker moiety comprises a five- or six-membered ring, preferably a single five- or sixmembered ring. Preferably, the five- or six-membered ring is saturated or partially saturated.

[0043] In embodiments, the linker moiety comprises a C=O, S(=O) or S(=O)2 group, preferably a S(=O)2 group, and the linker moiety comprises a cyclopentyl, a cyclohexyl, a pyridinyl or a pyrrolidinyl.

[0044] In embodiments, the PLA2G15 inhibitor is represented by formula (IV): (IV).

[0045] X, Y AND RXYIN THE CENTRAL RING

[0046] In embodiments, X is S and Y is CH; or X is S and Y is N; or X is O and Y is CH; or X is O and Y is N.

[0047] In embodiments, RXYis H, a C1-4 alkyl, or a halogen. Preferably, RXYis H, a C1-4 alkyl, or F.

[0048] In embodiments, RXYis H, a C1-3 alkyl, or a halogen. Preferably, RXYis H, a C1-4 alkyl, or F.

[0049] In embodiments, RXYis H, a methyl, or a halogen. Preferably, RXYis H, a methyl, or F.

[0050] In embodiments, RXYis H or CH3, preferably H.

[0051] In embodiments, X is S, Y is CH, and RXYis H, a C1-4 alkyl, or a halogen. Preferably, RXYis H, a C1- 4 alkyl, or F.

[0052] In embodiments, X is S, Y is CH, and RXYis H, a C1-3 alkyl, or a halogen. Preferably, RXYis H, a C1- 4 alkyl, or F. In embodiments, X is S, Y is CH, and RXYis H, a methyl, or a halogen. Preferably, RXYis H, a methyl, or F.

[0053] In embodiments, X is S, Y is CH, and RXYis H or CH3, preferably H.

[0054] RNBOUND TO THE CENTRAL RING

[0055] In embodiments, RNis H.

[0056] In embodiments, RNis a ring, specifically a C3-6 cycloalkyl, a 5-membered aromatic ring or a 6- membered aromatic ring, wherein the ring is substituted with one or more halogens, pseudohalogens, C1-4 alkyls, O-C1-4 alkyls, C3-6 cycloalkyls, or O-C3-6 cycloalkyls; wherein each C1-4 alkyl, or C3-6 cycloalkyl may be independently substituted with one or more halogens or pseudohalogens.

[0057] In embodiments, RNis a ring, specifically a C3-6 cycloalkyl, a 5-membered aromatic ring or a 6- membered aromatic ring, wherein the ring is substituted with one or more halogens, C1-3 alkyls, or O- C1-3 alkyls; wherein each C1-3 alkyl may be independently substituted with one or more halogens. Preferably, the halogen is F and the C1-3 alkyl is CH3.

[0058] In embodiments, RNis a 5-membered aromatic ring or a 6-membered aromatic ring, wherein the aromatic ring is substituted with one or more halogens, pseudohalogens, C1-4 alkyls, O-C1-4 alkyls, C3-6 cycloalkyls, or O-C3-6 cycloalkyls; wherein each C1-4 alkyl, or C3-6 cycloalkyl may be independently substituted with one or more halogens or pseudohalogens.

[0059] In embodiments, RNis a 5-membered aromatic ring or a 6-membered aromatic ring, wherein the aromatic ring is substituted with one or more halogens, C1-3 alkyls, or O-C1-3 alkyls; wherein each C1-3 alkyl may be independently substituted with one or more halogens. Preferably, the halogen is F and the C1-3 alkyl is CH3.

[0060] In embodiments, RNis a cyclopropyl, a 5-membered aromatic ring or a 6-membered aromatic ring, wherein the ring is substituted with one or more halogens, pseudohalogens, C1-4 alkyls, O-C1-4 alkyls, C3-6 cycloalkyls, or O-C3-6 cycloalkyls; wherein each C1-4 alkyl, or C3-6 cycloalkyl may be independently substituted with one or more halogens or pseudohalogens.

[0061] In embodiments, RNis a cyclopropyl, a 5-membered aromatic ring or a 6-membered aromatic ring, wherein the ring is substituted with one or more halogens, C1-3 alkyls, or O-C1-3 alkyls; wherein each C1- 3 alkyl may be independently substituted with one or more halogens. Preferably, the halogen is F and the C1-3 alkyl is CH3.

[0062] In embodiments, RNis a cyclopropyl, a 5-membered azaheterocycle or a 6-membered azaheterocycle, wherein the ring is substituted with one or more halogens, pseudohalogens, C1-4 alkyls, O-C1-4 alkyls, C3-6 cycloalkyls, or O-C3-6 cycloalkyls; wherein each C1-4 alkyl, or C3-6 cycloalkyl may be independently substituted with one or more halogens or pseudohalogens. Preferably, the azaheterocycle is monosubstituted on a nitrogen atom.

[0063] In embodiments, RNis a cyclopropyl, a 5-membered azaheterocycle or a 6-membered azaheterocycle, wherein the ring is substituted with one or more halogens, C1-3 alkyls, or O-C1-3 alkyls; wherein each C1-3 alkyl may be independently substituted with one or more halogens. Preferably, the halogen is F and the C1-3 alkyl is CH3. Preferably, the azaheterocycle is monosubstituted on a nitrogen atom.

[0064] In embodiments, RNis a 5-membered azaheterocycle or a 6-membered azaheterocycle, wherein the ring is substituted with one or more halogens, pseudohalogens, C1-4 alkyls, O-C1-4 alkyls, C3-6 cycloalkyls, or O-C3-6 cycloalkyls; wherein each C1-4 alkyl, or C3-6 cycloalkyl may be independently substituted with one or more halogens or pseudohalogens. Preferably, the azaheterocycle is monosubstituted on a nitrogen atom.

[0065] In embodiments, RNis a 5-membered azaheterocycle or a 6-membered azaheterocycle, wherein the ring is substituted with one or more halogens, C1-3 alkyls, or O-C1-3 alkyls; wherein each C1-3 alkyl may be independently substituted with one or more halogens. Preferably, the halogen is F and the C1-3 alkyl is CH3. Preferably, the azaheterocycle is monosubstituted on a nitrogen atom.

[0066] Wherever a azaheterocycle is mentioned in the context of RN, an aromatic ring comprising a N ring atom is meant. Wherever the substitution of a ring comprising a N ring atom is mentioned, substitution is preferably on the N (ring) atom. More preferably, it is a monosubstitution, i.e. a substitution corresponding with the replacement of a single nitrogen atom on the naked ring.

[0067] In embodiments, RNis a phenyl, a pyridinyl, an oxazole, an isoxazole, a thiazole, an isothiazole, a pyrazole, an imidazole or a pyrrole. Each of these aromatic rings is preferably substituted with one or more halogens, pseudohalogens, C1-4 alkyls, O-C1-4 alkyls, C3-6 cycloalkyls, or O-C3-6 cycloalkyls; wherein each C1-4 alkyl, or C3-6 cycloalkyl may be independently substituted with one or more halogens or pseudohalogens.

[0068] In embodiments, RNis a phenyl, a pyridinyl, an oxazole, an isoxazole, a thiazole, an isothiazole, a pyrazole, an imidazole or a pyrrole. Each of these aromatic rings is preferably substituted with one or more halogens, C1-3 alkyls, or O-C1-3 alkyls; wherein each C1-3 alkyl may be independently substituted with one or more halogens. Preferably, the halogen is F and the C1-3 alkyl is CH3.

[0069] In embodiments, RNis a phenyl, a pyridinyl, an oxazole or a isoxazole. Each of these aromatic rings is preferably substituted with one or more halogens, pseudohalogens, C1-4 alkyls, O-C1-4 alkyls, C3-6 cycloalkyls, or O-C3-6 cycloalkyls; wherein each C1-4 alkyl, or C3-6 cycloalkyl may be independently substituted with one or more halogens or pseudohalogens. In embodiments, RNis a phenyl, a pyridinyl, an oxazole or a isoxazole. Each of these aromatic rings is preferably substituted with one or more halogens, C1-3 alkyls, or O-C1-3 alkyls; wherein each C1-3 alkyl may be independently substituted with one or more halogens. Preferably, the halogen is F and the C1-3 alkyl is CH3.

[0070] In embodiments, RNis a phenyl substituted with one or more halogens, pseudohalogens, C1-4 alkyls, O-C1-4 alkyls, C3-6 cycloalkyls, or O-C3-6 cycloalkyls; wherein each C1-4 alkyl, or C3-6 cycloalkyl may be independently substituted with one or more halogens or pseudohalogens.

[0071] In embodiments, RNis a phenyl substituted with one or more halogens, C1-3 alkyls, or O-C1-3 alkyls; wherein each C1-3 alkyl may be independently substituted with one or more halogens. Preferably, the halogen is F and the C1-3 alkyl is CH3.

[0072] The ring atom bound to RN which is explicitly shown in formula (I) is a chiral center. Hence, formula (I) may be depicted as formula (VI) or (VI#):

[0073] (VI) (VI#)

[0074] In embodiments, the PLA2G15 inhibitor is represented by formula (VI) or (VI#). Preferably, the PLA2G15 inhibitor is represented by formula (VI).

[0075] RING SYSTEM RL

[0076] RLcomprises a carbocyclic ring. A carbocyclic ring is a ring wherein each ring atom is a carbon atom.

[0077] In embodiments, RLcomprises a 3- to 6-membered, a 4- to 6-membered, a 5- to 6-membered, a 3- to 5-membered, a 4- to 5-membered, a 3- to 4-membered, a 3-membered, a 4-membered, a 5- membered, or a 6-membered carbocyclic ring, or a 4-membered heterocyclic ring.

[0078] In embodiments, RLis covalently bound to L, as depicted in formula (I) via a ring atom comprised in the 3- to 6-membered carbocyclic ring or a 4-membered heterocyclic ring comprised in RL.

[0079] In embodiments, RLcomprises a single ring system. A ring system comprised in a compound is a set of ring atoms, each comprised in the compound, that form a continuous covalent network of endocyclic bonds, not interrupted by any exocyclic bonds, wherein the addition to the set of any other ring atom comprised in the compound would break the continuous covalent network. A ring system can be a monocyclic or multicyclic ring, a group of annulated rings, a spiro compound, etc., but cannot be, for example, a ring substituted with two distinct rings (e.g., 2,2-dicyclopropylphenyl). A moiety or a compound comprising a single ring system means that it contains one and only one ring system. In other words, all ring atoms comprised in such moieties or compounds are comprised in the ring system. Similar definitions can be made for moieties or compounds comprising a number of ring systems.

[0080] In embodiments, RLcomprises or consists of a 3- to 6-membered carbocyclic ring. Preferably, RLcomprises or consists of a cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl or phenyl. More preferably, RLcomprises or consists of a phenyl. More preferably alternatively, RLcomprises or consists of a cyclopropyl, cyclobutyl, cyclopentyl or cyclohexyl. In some embodiments, RLcomprises or consist of an azetidine. In these embodiments, the carbocyclic ring or a 4-membered heterocyclic ring may be substituted with one or more halogens, pseudohalogens, C1-4 alkyls, O-C1-4 alkyls, C3-6 cycloalkyls, O- C3-6 cycloalkyls, or phenyls; wherein each phenyl may be independently substituted with one or more halogens, pseudohalogens, C1-4 alkyls, O-C1-4 alkyls, C3-6 cycloalkyls, or O-C3-6 cycloalkyls; and wherein each C1-4 alkyl, or C3-6 cycloalkyl may be independently substituted with one or more halogens or pseudohalogens.

[0081] In embodiments, RLcomprises or consists of a 3- to 6-membered carbocyclic ring. Preferably, RLcomprises or consists of a cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl or phenyl. More preferably, RLcomprises or consists of a phenyl. More preferably alternatively, RLcomprises or consists of a cyclopropyl, cyclobutyl, cyclopentyl or cyclohexyl. In these embodiments, the carbocyclic ring may be substituted with one or more halogens or C1-4 alkyls; wherein each C1-4 alkyl may be independently substituted with one or more halogens. Preferably, the C1-4 alkyls is a methyl. Preferably, the halogen is F.

[0082] In embodiments, RLcomprises or consists of a phenyl substituted with OH, Rm1and Rm2, wherein Rm1and Rm2are independently a C1-4 alkyl, a C3-6 cycloalkyl, a halogen or a pseudohalogen; wherein each C1-4 alkyl or C3-6 cycloalkyl may be independently substituted with one or more halogens or pseudohalogens; and preferably wherein Rm1is t-butyl, cyclopropyl or CF3, and Rm2is F, Cl or cyclopropyl.

[0083] In embodiments, RLcomprises or consists of a phenyl 2-substituted with OH position, and 3,4- substituted or 3,5-substitued or 4,5-substitued with Rm1and Rm2at , wherein Rm1and Rm2are independently a C1-4 alkyl, a C3-6 cycloalkyl, a halogen or a pseudohalogen, wherein Rm1and Rm2are wherein each C1-4 alkyl or C3-6 cycloalkyl may be independently substituted with one or more halogens or pseudohalogens; and preferably wherein Rm1is t-butyl, cyclopropyl or CF3, and Rm2is F, Cl or cyclopropyl.

[0084] In embodiments, RLconsists of a phenyl, wherein the phenyl is represented by (ph-1), and the PLA2G15 inhibitor is consequently represented by (VII):

[0085] (ph-1) (VII), wherein Rm1and Rm2are independently a C1-4 alkyl, a C3-6 cycloalkyl, a halogen or a pseudohalogen; wherein each C1-4 alkyl or C3-6 cycloalkyl may be independently substituted with one or more halogens or pseudohalogens; and preferably wherein Rm1is t-butyl, cyclopropyl or CF3, and Rm2is F, Cl or cyclopropyl. In embodiments, RLcomprises or consists of two ring systems. Preferably, RLcomprises a cyclopropyl and a phenyl.

[0086] In embodiments wherein RLis bound to an asymmetrically substituted sp3carbon atom, e.g., embodiments wherein RLconsists of an asymmetrically substituted cyclopropyl, cyclobutyl, cyclopentyl or cyclohexyl, the PLA2G15 inhibitor may be depicted as formula (V) or (V#): In formulae (V) and (V#), the endocyclic bonds comprised in the asymmetrically substituted cyclopropyl, cyclobutyl, cyclopentyl or cyclohexyl fall in horizontal plane, i.e. in the plane formally described in the ring comprising X and Y. In other words, the straight lines in the tear-shaped representation of RL represent the two endocyclic bonds.

[0087] In embodiments, the PLA2G15 inhibitor is represented by formula (V) or (V#). Preferably, the PLA2G15 inhibitor is represented by formula (V).

[0088] PREFERRED INHIBITORS

[0089] In embodiments, the PLA2G15 inhibitor is represented by any one of formulae (VIII) to (XXVII), wherein RL, L, Z, i, RA, RB, RN, X, Y and RXYare as defined herein:

[0090] (VIII) (IX)

[0091] In embodiments, the PLA2G15 inhibitor is represented by any one of formulae (1) to (48):

[0092]

[0093] (27) (28)

[0094]

[0095] (37) (38)

[0096]

[0097] In embodiments, the PLA2G15 inhibitor is represented by any one of the following formulae:

[0098]

[0099]

[0100] (53) (54)

[0101]

[0102] ISOMERY

[0103] Certain PLA2G15 inhibitors may exist in one or more particular geometric, optical, enantiomeric, diastereoisomeric, epimeric, stereoisomeric, tautomeric, (de)protonated, isotopomeric, isotopologues, isotopologic, conformational, or anomeric forms, including but not limited to, cis- and trans-forms; E- and Z-forms; c-, t-, and r- forms; endo- and exo-forms; R-, S-, and meso-forms; D- and L-forms; d- and l-forms; (+) and (-) forms; keto-, enol-, and enolate-forms; syn- and anti-forms; synclinal- and anticlinal-forms; a- and p-forms; axial and equatorial forms; boat-, chair-, twist-, envelope-, and halfchair-forms; and combinations thereof, hereinafter collectively referred to as “isomers” (or “isomeric forms”).

[0104] Any reference to a compound or a class of compounds, either by a name or by a formula, is meant as a reference to the set of all isomers falling in that class, unless explicitly mentioned otherwise. This includes both the isomers mentioned above and all structural isomers, unless it is clear that they are explicitly excluded. For example, a C1-4 alkyl may refer to n-butyl and tert-butyl, together with all their isomers as mentioned above. A reference to n-butyl, on the other hand, only refers to n-butyl and its stereoisomers, isotopomers, isotopologues, etc., and not to tert-butyl.

[0105] If a compound or a class of compounds refers to multiple species, reference is made to both the isolated species and to any equimolar or non-equimolar mixture of the species.

[0106] SALTS

[0107] A PLA2G15 inhibitor according to the invention may be present as a pharmaceutically acceptable salt.

[0108] Examples of pharmaceutically acceptable salts are discussed in Berge et al., 1977, “Pharmaceutically Acceptable Salts,” J. Pharm. Sci., Vol. 66, pp. 1 -19.

[0109] For example, if a compound according to the invention is anionic, or has a functional group, which may be anionic (e.g., -COOH may be -COO ), then a salt may be formed with a suitable cation. Examples of suitable inorganic cations include, but are not limited to, alkali metal ions such as Na+and K+, alkaline earth cations such as Ca2+and Mg2+, and other cations such as Al3+as well as the ammonium ion (i.e., NH4+). Examples of suitable organic cations include, but are not limited to substituted ammonium ions (e.g., NHsR+, NH2R2+, NHRs+, NR4+), for example, where each R is independently linear or branched saturated Ci-isalkyl, Cs-scycloalkyl, Cs-scycloalkyl-Ci-ealkyl, and phenyl-Ci-ealkyl, wherein the phenyl group is optionally substituted. Examples of some suitable substituted ammonium ions are those derived from: ethylamine, diethylamine, dicyclohexylamine, triethylamine, butylamine, ethylenediamine, ethanolamine, diethanolamine, piperazine, benzylamine, phenylbenzylamine, choline, meglumine, and tromethamine, as well as amino acids, such as lysine and arginine. An example of a common quaternary ammonium ion is N(CH3)4+.

[0110] If a PLA2G15 inhibitor is cationic, or has a functional group, which upon protonation may become cationic (e.g., -NH2 may become -NFV), then a salt may be formed with a suitable anion.

[0111] For example, if a PLA2G15 inhibitor contains a cationic group (e.g., -NMe2+), or has a functional group, which upon protonation may become cationic (e.g., -NH2 may become -NH3+), then a salt may be formed with a suitable anion. In the case of a quaternary ammonium compound a counter-anion is generally always present in order to balance the positive charge. If, in addition to a cationic group (e.g., -NMe2+, -NFV), the compound also contains a group capable of forming an anion (e.g., -COOH), then an inner salt (also referred to as a zwitterion) may be formed.

[0112] Examples of suitable inorganic anions include, but are not limited to, those derived from the following inorganic acids: hydrochloric, hydrobromic, hydroiodic, sulfuric, sulfurous, nitric, nitrous, phosphoric, and phosphorous.

[0113] Examples of suitable organic anions include, but are not limited to, those derived from the following organic acids: 2-acetyloxybenzoic, acetic, trifluoroacetic, ascorbic, aspartic, benzoic, camphorsulfonic, cinnamic, citric, edetic, 1 ,2-ethanedisulfonic, ethanesulfonic, fumaric, glucoheptonic, gluconic, glutamic, glycolic, hydroxymaleic, hydroxynaphthalene carboxylic, isethionic, lactic, lactobionic, lauric, maleic, malic, methanesulfonic, mucic, oleic, oxalic, palmitic, pamoic, pantothenic, phenylacetic, phenylsulfonic, propionic, pyruvic, salicylic, stearic, succinic, sulfanilic, tartaric, toluenesulfonic, and valeric. Examples of suitable polymeric organic anions include, but are not limited to, those derived from the following polymeric acids: tannic acid, carboxymethyl cellulose.

[0114] SOLVATES AND HYDRATES

[0115] A PLA2G15 inhibitor according to the invention may be present as a pharmaceutically acceptable solvate or hydrate.

[0116] The term solvate is used herein in the conventional sense to refer to a complex of solute (e.g., compound, salt of compound) and solvent. If the solvent is water, the solvate may be conveniently referred to as a hydrate, for example, a mono-hydrate, a di-hydrate, a tri-hydrate, etc.

[0117] PLA2G15 STRUCTURE OF PLA2G15

[0118] PLA2G15, which may also be called a PLA2G15 protein, a phospholipase A2 group XV (protein), a lysophospholipase 3 (protein) or LYPLA3, is a protein encoded by a PLA2G15 gene. Herein, all human variants and isoforms, and species homologues and their variants and isoforms are encompassed. In the context of this application, a PLA2G15 or a LYPLA3 refer to a PLA2G15 protein, unless explicitly mentioned otherwise.

[0119] All embodiments disclosed below relating to PLA2G15 proteins or genes may be applied accordingly to inhibitors and compositions according to the invention.

[0120] In embodiments, an PLA2G15 gene is located at open reading frame UNQ341 / PRO540. In this context, a PLA2G15 (protein) may also be called a UNQ341 / PRO540 (protein).

[0121] In embodiments, PLA2G15 is represented by an amino acid sequence having at least 75%, 75.5%, 76%, 76.5%, 77%, 77.5%, 78%, 78.5%, 79%, 79.5%, 80%, 80.5%, 81 %, 81.5%, 82%, 82.5%, 83%, 83.5%, 84%, 84.5%, 85%, 85.5%, 86%, 86.5%, 87%, 87.5%, 88%, 88.5%, 89%, 89.5%, 90%, 90.5%, 91 %, 91.5%, 92%, 92.5%, 93%, 93.5%, 94%, 94.5%, 95%, 95.5%, 96%, 96.5%, 97%, 97.5%, 98%, 98.5%, 99%, 99.5%, or 100% sequence identity with SEQ ID NO: 1 or 2, preferably with SEQ ID NO:1 .

[0122] In embodiments, PLA2G15 is represented by an amino acid sequence having at least 75%, 75.5%, 76%, 76.5%, 77%, 77.5%, 78%, 78.5%, 79%, 79.5%, 80%, 80.5%, 81 %, 81.5%, 82%, 82.5%, 83%, 83.5%, 84%, 84.5%, 85%, 85.5%, 86%, 86.5%, 87%, 87.5%, 88%, 88.5%, 89%, 89.5%, 90%, 90.5%, 91 %, 91.5%, 92%, 92.5%, 93%, 93.5%, 94%, 94.5%, 95%, 95.5%, 96%, 96.5%, 97%, 97.5%, 98%, 98.5%, 99%, 99.5%, or 100% sequence similarity with SEQ ID NO: 1 or 2, preferably with SEQ ID NO:1 .

[0123] In embodiments, PLA2G15 comprises an amino acid sequence represented by SEQ ID NO: 1 or 2, preferably SEQ ID NO: 1 . In more specific aspects, PLA2G15 consists of an amino acid sequence represented by SEQ ID NO: 1 or 2, preferably SEQ ID NO: 1 .

[0124] In embodiments, PLA2G15 has a length from 322 up to 502 amino acids, or from 332 up to 492 amino acids, or from 342 up to 482 amino acids, or from 352 up to 472 amino acids, or from 362 up to 462 amino acids, or from 372 up to 452 amino acids, or from 382 up to 442 amino acids, or from 392 up to 432 amino acids, or from 402 up to 422 amino acids, or from 403 up to 421 amino acids, or from 404 up to 420 amino acids, or from 405 up to 419 amino acids, or from 406 up to 418 amino acids, or from 407 up to 417 amino acids, or from 408 up to 416 amino acids, or from 409 up to 415 amino acids, or from 410 up to 414 amino acids, or from 411 up to 413 amino acids, preferably wherein PLA2G15 is represented by an amino acid sequence having at least 90% sequence identity with SEQ ID NO: 1 , more preferably wherein PLA2G15 is represented by an amino acid sequence comprising SEQ ID NO: 1. In embodiments, PLA2G15 has a length from 218 up to 418 amino acids, or from 228 up to 408 amino acids, or from 238 up to 398 amino acids, or from 248 up to 388 amino acids, or from 258 up to 378 amino acids, or from 268 up to 368 amino acids, or from 278 up to 358 amino acids, or from 288 up to 348 amino acids, or from 298 up to 338 amino acids, or from 308 up to 328 amino acids, or from 309 up to 327 amino acids, or from 310 up to 326 amino acids, or from 311 up to 325 amino acids, or from 312 up to 324 amino acids, or from 313 up to 323 amino acids, or from 314 up to 322 amino acids, or from 315 up to 321 amino acids, or from 316 up to 320 amino acids, or from 317 up to 319 amino acids, preferably wherein PLA2G15 is represented by an amino acid sequence having at least 90% sequence identity with SEQ ID NO: 2, more preferably wherein PLA2G15 is represented by an amino acid sequence comprising SEQ ID NO: 2.

[0125] In embodiments, PLA2G15 has a length of 412 amino acids, or 318 amino acids.

[0126] In embodiments, PLA2G15 is expressed in or is derived from a vertebrate, more preferably a mammal, even more preferably a rat, a mouse, a rabbit or a human, most preferably a human. In this context, PLA2G15 derived from a specific animal may be a recombinant protein expressed in a host organism.

[0127] ACTIVITY OF PLA2G15

[0128] An inhibitor according to the invention is able to inhibit (i.e. decrease) an activity of PLA2G15. In this context, an inhibitor according to the invention may also be called a compound able to inhibit an activity of PLA2G15. Herein, decreasing an activity may mean inhibiting an activity of PLA2G15 via direct or indirect contact between said inhibitor and PLA2G15.

[0129] In specific aspects, an inhibitor according to the invention is a competitive inhibitor of said PLA2G15 protein, wherein said inhibitor is able to specifically bind an active site of said PLA2G15 protein associated with said activity of said PLA2G15 protein.

[0130] In specific aspects, an inhibitor according to the invention is a non-competitive or allosteric inhibitor of said PLA2G15 protein, wherein said inhibitor is able to specifically bind a part of said PLA2G15 protein which is not an active site associated with said activity of said PLA2G15 protein. In this context, said binding site which is not an active site maybe called an allosteric site.

[0131] A skilled person in the art can test the inhibitory effect of the PLA2G15 inhibitors according to the invention on the PLA2G15 enzymatic activity with an inhibition assay, preferably the 4-nitrophenyl butyrate assay. In this 4-nitrophenyl butyrate assay, the compounds pNPA (para-nitrophenyl acetate) and pNPB (para-nitrophenyl butyrate) are substrates that can be used to detect PLA2G15 enzyme activities. The PLA2G15 enzyme catalyzes the hydrolysis of ester bonds between an acyl moiety and p-nitrophenol (pNP): The release of 4-nitrophenolate anion (yellow at pH values above its pKa of 7.08 at 22 °C) is determined as a strong increase in absorbance at 405 nm. In embodiments is provided an inhibitor according to the invention, wherein said inhibitor is able to decrease a catalytic activity of PLA2G15. In this context, PLA2G15 may be called an enzyme and a catalytic activity may be called a enzymatic activity. It is understood that a catalytic activity of a PLA2G15 protein or enzyme means that PLA2G15 increases the rate of a reaction, preferably by a factor of at least 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 200, 300, 400, 500, 600, 700, 800, 900 or 1000, relative to the rate of a corresponding reaction performed in a corresponding environment (e.g. similar cellular conditions, pH, salt concentrations, enzyme concentrations, etc.) and under corresponding conditions (e.g. same temperature, etc.) wherein PLA2G15 is not present. Preferably, said rate increase of said reaction is defined under physiological conditions. In the context of this application, “a catalytic activity of PLA2G15, wherein said catalytic activity comprises a reaction”, “a reaction catalysed by PLA2G15” or similar phrases mean that the rate of said reaction is increased by PLA2G15, as explained above.

[0132] In embodiments, decreasing a catalytic activity means decreasing the rate of a reaction catalysed by PLA2G15, more preferably by a factor equal to or lower than 0.75, 0.7, 0.65, 0.6, 0.55, 0.5, 0.45, 0.4, 0.35, 0.3, 0.25, 0.2, 0.15, 0.1 , 0.09, 0.08, 0.07, 0.06, 0.05, 0.04, 0.03, 0.02, 0.01 , 0.009, 0.008, 0.007, 0.006, 0.005, 0.004, 0.003, 0.002 or 0.001 , relative to the rate of a corresponding reaction performed in a corresponding environment (e.g. similar cellular conditions, pH, salt concentrations, PLA2G15 concentration, other enzyme concentrations, etc.) and under corresponding conditions (e.g. same temperature, etc.) wherein said inhibitor according to the invention is not present. Preferably, said rate decrease of said reaction is defined under physiological conditions. As explained above, decreasing the rate of a reaction catalysed by PLA2G15 may be the result of inhibiting a catalytic activity of PLA2G15 via direct or indirect contact between said inhibitor and PLA2G15. The rate of a reaction catalysed by PLA2G15 may be measured or assessed by any suitable methods well-known in the art.

[0133] In embodiments, the rate of a reaction catalysed by PLA2G15 is measured or defined at pH 4.5 and under otherwise physiological conditions. More preferably, said reaction takes place in an organelle, a cell fraction, a cell, a tissue, an organ or a subject, most preferably in a vertebrate, mammalian or human cell. Preferably, an organelle is a lysosome or an endosome. Preferably, an endosome is a late endosome.

[0134] Without being limited to any specific explanation, mechanism or hypothesis, an inhibitor as used in the invention is capable, in a suitable assay or model and / or upon administration to a subject, of decreasing in specific aspects a catalytic activity of PLA2G15 that originates from, is caused by or is linked to a catalytic triad comprised in PLA2G15, in particular a catalytic triad consisting of a histidine, a aspartic acid and a serine residue, and more in particular the catalytic triad that consists of His-359, Asp-327 and Ser-165.

[0135] In embodiments, a catalytic activity of PLA2G15 comprises the cleavage of a fatty acid residue from a compound, which may also be called a deacylase activity. More preferably, said compound is a lipid and said catalytic activity may be called a lipase activity. Even more preferably, said lipid is a phospholipid and said catalytic activity may be called a phospholipase activity. Most preferably, said phospholipase activity is a phospholipase A1 activity or a phospholipase A2 activity, preferably a phospholipase A2 activity.

[0136] In embodiments, a phospholipid in the aspects above is a phosphatidylcholine, a phosphatidylethanolamine, a phosphatidylglycerol, or a phosphatidylserine.

[0137] In embodiments, a phospholipid in the aspects above is an oxidized phospholipid, preferably wherein said oxidized phospholipid comprises short fatty acid chains (e.g. comprising less than 10 carbon atoms) and / or a free carboxyl or formyl group at the sn-2 position.

[0138] In embodiments, a phospholipid in the aspects above is not a phosphatidylinositol or a and sphingomyelin.

[0139] In embodiments, a catalytic activity of PLA2G15 comprises a transfer of a fatty acid residue from a donor compound to an acceptor compound, which may also be called a transacylase activity. A transacylase activity thus comprises a deacylase activity, to which the preferences above apply.

[0140] In embodiments, a catalytic activity of PLA2G15 comprises a transacylase activity, wherein said acceptor compound is N-acetyl-sphingosine.

[0141] In embodiments, an activity of PLA2G15 comprises a catalytic activity during lipid metabolism. Herein, it is understood that lipid metabolism comprises lipid anabolism and lipid catabolism.

[0142] CLINICAL RELEVANCE OF PLA2G15

[0143] PLA2G15 inhibitors according to the invention and compositions according to the invention are particularly useful in the treatment of diseases characterized by disease characterized by lysosomal dysregulation. Lysosomal dysregulation or dysfunction is a hallmark of rare and common neurodegenerative diseases, metabolic disease and cancer, including neuronopathic lysosomal storage disorders as well as Alzheimer’s disease, Parkinson’s disease and frontotemporal dementia (FTD) (1). Indeed, heterozygous carriers of well-known LSD causing mutations in genes like NPC1 , GRN or GBA carry increased risk of developing Alzheimer’s disease, Frontotemporal dementia or Parkinson’s disease, highlighting a mechanistic link between rare lysosomal storage diseases and more common neurodegenerative diseases (42, 10, 43). Therefore, counteracting lysosomal dysfunction is considered of potential therapeutic benefit not only to lysosomal storage diseases, but also more common neurodegenerative diseases like Alzheimer’s and Parkinson’s or frontotemporal dementia.

[0144] One of the characteristics of lysosomal dysregulation is that levels of bis(monoacylglycerol)- phosphate (BMP) are dysregulated. BMP is a lysosomal / late endosomal lipid that stimulates key lysosomal functions: it activates enzymatic activity mediating lipid degradation, controls cholesterol distribution to extra-lysosomal compartments and is involved in endosomal / lysosomal trafficking dynamics to allow cellular homeostasis (3). The central and limiting role of BMP in regulating lysosomal function is well understood in (glyco)-sphingolipid breakdown: here it plays a structural role as docking station for activating co-factors like GM2A, Saposin A-D or the heat shock protein HSP-70 in complex with lysosomal enzymes mediating degradation of (glyco)-sphingolipids (34, 37). Some of these BMP dependent enzymes are well known targets of genetic LSDs.

[0145] Dysregulation of BMPs may result in increased or decreased levels of BMP. Increased BMP levels have been linked to several well-studied lysosomal lipid storage diseases including NPC, GM1 Gangliosidosis [3], and Gaucher disease

[0045] . Increased BMP levels are also seen in an Alzheimer disease model carrying the APO4 risk allele (41), while in Batten CLN3 and CLN5 (14, 17) or FTD caused by granulin haploinsufficiency (10) BMP level is low. Neuronal ceroid lipofuscinosis variants caused by mutations in CLN5 shows a near absence of BMP, while mutations in CLN3 reduces BMP levels. Disorders caused by GRN mutations, including frontotemporal dementia and neuronal ceroid lipofuscinosis GRN also show decreased BMP abundance. It has been reported that treatments that raise BMP levels alleviate cellular symptoms.

[0146] Cellular treatments to increase BMP levels have been shown to correlate with reduced intracellular biogenesis of HIV viral particles and may therefore present novel therapeutic strategies in HIV treatment (47).

[0147] In Niemann Pick type C (NPC) disease, defects in lysosomal cholesterol trafficking are accompanied by accumulation of BMP, and further elevation of BMP levels is shown to lessen multiple cellular defects, including cholesterol accumulation and clearance of autophagic materials. The related disorders Niemann Pick type A and B disease are caused by accumulation of sphingomyelin due to mutations in the lysosomal enzyme acid sphingomyelinase (ASM) and also display enhanced levels of BMP. As BMP stimulates ASM activity, a further increase in BMP may be beneficial by enhancing hydrolysis of lysosomal sphingomyelin.

[0148] Increased levels of BMP in lysosomal storage disease are not merely considered a secondary storage phenotype but rather an active cellular response to increase BMP dependent lysosomal functions and counteract lysosomal pathology (17). Thus, limiting rather than increased or reduced levels of BMP are considered a pathological factor and restoring balanced BMP expression in lysosomes is considered an attractive therapeutic approach multiple human disorders. Indeed, therapeutic interventions leading to increased levels of BMP have been shown to correlate with therapeutic efficacy, including in preclinical models of NPC (21 , 22, 7) and Parkinson’s disease (36).

[0149] Therefore, increasing BMP levels in a wide class of diseases as outlined above is expected to have therapeutic impact.

[0150] We demonstrated that BMP and its precursor lysophosphatidylglycerol (LPG) are elevated in multiple tissues of mice that lack expression of Pla2g15 or after in vivo pharmacological inhibition of Pla2g15. Similarly, BMP and LPG levels are elevated in cell lines when the Pla2g15 gene is deleted or after inhibition with Pla2g15 specific inhibitors. Furthermore, we have shown mechanistically that PLA2G15 activity controls efficacy of the BMP biosynthetic pathway by hydrolyzing both the precursor LPG and the product BMP in vitro, consistent with our in vivo and cellular findings in genetic and pharmacological models.

[0151] Therapy

[0152] In aspects, the PLA2G15 inhibitors according to the invention and the compositions according to the invention are for use in the treatment of a disease characterized by lysosomal dysregulation. Below, preferred diseases characterized by lysosomal dysregulation, and other preferred features of the treatment are disclosed.

[0153] DISEASES

[0154] Said diseases include neuronopathic lysosomal storage disorders (LSD), Alzheimer’s disease, Parkinson’s disease, frontotemporal degeneration, neurodegenerative diseases, metabolic diseases, a kidney or a liver diseases and cancer.

[0155] In one embodiment, said disease is an LSD. LSDs are inherited diseases characterized by lysosomal dysfunction and neurodegeneration. The term LSDs defines a group of approximately 70 disorders, typically due to single gene defects: deficiency of specific enzymes that are normally required for the breakdown of lysosomal glycosaminoglycans (GAGs), glycosphingolipids or glycoproteins, which thus accumulate in the lysosomes of the cell. This accumulation disrupts the cell's normal functioning and gives rise to the clinical manifestations of LSDs.

[0156] Neurological impairment and neurodegenerative processes are associated to lysosomal dysfunction and represent a predominant feature in most LSDs. Neuropathology can occur in multiple brain regions (e.g., thalamus, cortex, hippocampus, and cerebellum) and involves unique temporal and spatial changes, which often entail early region-specific neurodegeneration and inflammation. As an example, Purkinje neurons degenerate in many of these diseases leading to cerebellar ataxia.

[0157] We demonstrated a reduction of the ganglioside GM3 and its degradation product lactosylceramide (LacCer) in brain of NPC1 / PLA2G15 dKO mice compared to NPC1 KO mice. Similarly, we showed a reduction of the gangliosides GM1 , GM2 and GM3 as well as its degradation products LacCer, Glucosylceramide (GlcCer) and Sphingosine in liver of NPC1 / PLA2G15 dKO mice compared to NPC1 KO mice. In addition, we detect a reduction in levels of Sphingomyelin (SM) and Sulfatide (SM4) in liver of NPC1 / PLA2G15 dKO mice compared to NPC1 KO mice. These findings indicate that PLA2G15 contributes to the accumulation of sphingomyelin and glycosphingolipids known to occur in primary or secondary sphingolipidoses. PLA2G15 inhibition can therefore be expected to have broad therapeutic benefit across this class of diseases. In a specific embodiment, said disease is a sphingolipidosis. Sphingolipidosis is characterized by a disturbance of the sphingolipid metabolism.

[0158] Errors in sphingolipid metabolism represent a major class of lysosomal storage diseases (2). Mutations in key enzymes mediating lysosomal degradation of (glyco)-sphingolipids have been identified across the degradative pathway of this lipid class and give rise to so-call primary sphingolipidoses, including GM1 Gangliosidosis, Tay-Sachs disease (B variant), Sandhoff disease, GM2AP deficiency, Sialidosis, Fabry disease, Gaucher disease, Niemann-Pick Type A / B, Krabbe disease, Metachromatic Leukodystrophy, Farber disease (35).

[0159] In addition, secondary sphingolipidoses like Niemann Pick type C disease and others occur where no mutations in the catabolic enzymes mediating (Glyco)sphingolipid degradation is detected, yet pathologic accumulation of so-called secondary storage lipids of the (Glyco)sphingolipid class is detected (44). Primary storage products like Cholesterol and Sphingomyelin in Niemann Pick type C and A / B, respectively, are thought to inhibit lysosomal activity by counteracting the stimulatory activity of BMP on the (glyco)-sphingolipid degradation pathway in the lysosome, as exemplified for ganglioside degradation by HexA (45). Therefore, restoring the balance between BMP expression in the late endosomal compartment and primary or secondary storage lipids like cholesterol or sphingomyelin is expected to have a positive therapeutic impact on a wide class of LSDs, including sphingolipidoses.

[0160] In a specific embodiment, said disease is a sphingolipidosis, including Niemann-Pick disease, type A an B, Niemann-Pick disease type C, Gaucher disease, Metachromatic leukodystrophy, Krabbe disease and Farber disease.

[0161] In a specific embodiment, said sphingolipidosis is a mucopolysaccharidosis (MPS), including, MPS I (Hurler syndrome, MPS II (Hunter syndrome) , MPS 11 IA (Sanfilippo syndrome), MPS I II B (Sanfilippo syndrome), MPS IIIC (Sanfilippo syndrome), MPS HID (Sanfilippo syndrome), MPS VI (Maroteaux- Lamy syndrome), MPS VII (Sly syndrome).

[0162] In a specific embodiment, said sphingolipidosis is a Mucolipidosis, including Mucolipidosis II (l-cell disease), Mucolipidosis III (pseudo-Hurler polydystrophy) and Mucolipidosis IV.

[0163] In a specific embodiment, said disease is glycoproteinosis, including galactosialidosis, mannosidosis, sialidosis.

[0164] In a specific embodiment, said sphingolipidosis is a Neuronal ceroid lipofuscinosis (NCL), including NCL 3 (Batten disease), NCL 10 and Hereditary spastic paraplegia (HSP).

[0165] In a specific embodiment, said disease is phospholipidosis.

[0166] In a specific embodiment, said disease is Alzheimer disease.

[0167] In a specific embodiment, said disease is Parkinson’s disease. In preferred embodiments said condition is selected from neuronal ceroid lipofuscinosis (NCL), CLN3

[0168] Batten, CLN5 Batten, GRN , frontotemporal dementia and Niemann Pick disease, preferably of type C.

[0169] In specific embodiments, said condition is selected from the diseases mentioned in Table 1 .

[0170] In a preferred embodiment, said disease is Niemann Pick type C (NPC).

[0171] Niemann-Pick disease type C (NPC) is a rare autosomal recessive, lysosomal storage disorder characterized by neurodegeneration in early childhood and death in adolescence. Classically, children with NPC disease demonstrate neurological dysfunction with cerebellar ataxia (an inability to coordinate balance, gait, extremity and eye movements), dysarthria (difficulty speaking), vertical gaze palsy (ability to move eyes in the vertical direction), motor impairment, dysphagia (trouble swallowing), psychotic episodes, and dementia (preferably progressive dementia). Affected individuals often experience progressive decline in intellectual function and about one-third have seizures. NPC is caused by mutations in the genes NPC1 or NPC2. NPC occurs at a frequency of 1 :100000 live births and is an autosomal recessive disorder. The gene products of NPC1 and NPC2 mediate redistribution of endocytic cholesterol from the late endosomal / lysosomal compartment to other cellular compartments like the endoplasmic reticulum and plasma membrane. Consequently, a hall mark of NPC disease is the cellular storage of cholesterol in the lysosomal compartment (38). While a small subset of early infantile cases will die within the first six months of birth from liver or respiratory failure, most patients will develop progressive and neurological complications and typically die between the ages of 10 to 25. The neurological symptoms typically present as cerebellar ataxia, dysarthria, dysphagia, and progressive dementia, and the majority of cases show a characteristic vertical supranuclear gaze palsy (VSGP) (38). In both human patients and preclinical models of NPC, progressive degeneration of the cerebellum and increased circulation of neurodegeneration biomarkers like Neurofilament light chain can be detected (Agrawal, Estibaliz Santiago-Mujica Helyon).

[0172] NPC disease is characterized by the secondary accumulation of (glyco)-sphingolipids and therefore considered a member of a group of diseases called shingolipidoses (35).

[0173] In specific embodiments, said LSD is characterized by progressive neurological symptoms tied to accumulation of lipid species. In specific embodiments, such condition is characterized by defects in lysosomal cholesterol trafficking. In specific embodiments, such condition is characterized by an increased level of cellular BMP. In other specific embodiments, said condition is characterized by a decreased level of cellular BMP. In preferred said cellular BMP level is elevated in the spleen, liver, brain, skin and / or plasma. In specific embodiments, said condition is characterized by cholesterol accumulation and lack of clearance of autophagic materials.

[0174] In specific embodiments, the treatment according to the invention results in the stabilization of cellular BMP levels. In other specific embodiments, the treatment according to the invention results in the increase of cellular BMP levels. In other specific embodiments, the treatment according to the invention results in enhancing hydrolysis of lysosomal sphingomyelin, glycosphingolipids and / or gangliosides. In preferred said cellular BMP level is elevated in the spleen, liver, brain, skin and / or plasma. In other specific embodiments, the treatment according to the invention results in the formation of free oleic acid from PG, LPG and BMP.

[0175] In specific aspects, Niemann-Pick disease type C is caused by mutation in an NPC1 gene (chromosome location 18q11) or an NPC2 gene (chromosome location 14q24.3), preferably in an NPC1 gene. Niemann-Pick disease type C caused by mutation in an NPC1 gene or an NPC2 gene may be called Niemann-Pick disease type C1 (NPC1) or Niemann-Pick disease type C1 (NPC2), respectively.

[0176] Both the NPC1 gene and the NPC2 are involved in the efflux of lipids, particularly cholesterol, from late endosomes and lysosomes. The NPC1 gene encodes a protein that is located in membranes inside the cell and is involved in the movement of cholesterol and lipids within cells. The NPC2 gene on the other hand encodes a protein that binds and transports cholesterol. Niemann-Pick disease type C is biochemically, genetically and clinically distinct from Niemann-Pick disease types A or and B. In types A and B, there is complete or partial deficiency of the lysosomal enzyme called acid sphingomyelinase. Without being bound to this theory, in Niemann-Pick disease type C, the protein product (i.e. the NPC1 protein) of the NPC1 gene is not an enzyme but appears to function as a transporter in the endosomal-lysosomal system, which moves large water-insoluble molecules through the cell. The protein coded by the NPC2 gene (i.e. the NPC2 protein) more closely resembles an enzyme structurally but seems to act in cooperation with the NPC1 protein in transporting molecules in the cell. The disruption of this transport system results in the accumulation of cholesterol and glycolipids in lysosomes.

[0177] Hence, in Niemann-Pick disease type C, large amounts of free or unesterified cholesterol accumulate in lysosomes, leading to relative deficiency of this molecule in multiple membranes and for steroid synthesis.

[0178] The mutations in the NPC1 gene and / or the NPC2 gene comprised in a subject suffering from Niemann-Pick disease type C syndrome result in a decreased NPC1 protein and / or NPC2 protein expression level, respectively, and / or the expression of a defective decreased NPC1 protein and / or NPC2 protein, respectively.

[0179] In the context of this application, a normal NPC1 or NPC2 protein expression level is defined as the NPC1 or NPC2 protein expression level in a healthy subject. A decreased NPC1 or NPC2 protein expression level means a NPC1 or NPC2 protein expression level lower than a normal NPC1 or NPC2 protein expression level, preferably decreased by a factor equal to or lower than 0.75, 0.7, 0.65, 0.6, 0.55, 0.5, 0.45, 0.4, 0.35, 0.3, 0.25, 0.2, 0.15, 0.1 , 0.09, 0.08, 0.07, 0.06, 0.05, 0.04, 0.03, 0.02 or 0.01 .

[0180] In the context of this application, a defective NPC1 or NPC2 protein is an NPC1 or NPC2 protein whose cellular activity is decreased, preferably decreased by a factor equal to or lower than 0.75, 0.7, 0.65, 0.6, 0.55, 0.5, 0.45, 0.4, 0.35, 0.3, 0.25, 0.2, 0.15, 0.1 , 0.09, 0.08, 0.07, 0.06, 0.05, 0.04, 0.03, 0.02 or 0.01 relative to an NPC1 or NPC2 protein expressed in a healthy subject, or no longer comprises such cellular activity.

[0181] In specific aspects, an inhibitor of the invention is able to induce one or more of the following changes when said inhibitor is introduced in a subject suffering from Niemann-Pick disease type C:

[0182] — a restored or partially restored intralysosomal cholesterol concentration, preferably wherein the intralysosomal cholesterol concentration is decreased after introduction of the inhibitor, more preferably wherein the intralysosomal cholesterol concentration is decreased by a factor equal to or greater than 1 .5, 2, 2.5, 3, 3.5, 4, 4.5, 5, 5.5, 6, 6.5, 7, 7.5, 8, 8.5, 9, 9.5, 10, 15, 20, 25, 30, 35, 40, 45, or 50; and / or

[0183] — a restored or partially restored endosomal, preferably late-endosomal cholesterol concentration, preferably wherein the endosomal, preferably the late-endosomal, cholesterol concentration is decreased after introduction of the inhibitor, more preferably wherein the endosomal, preferably the late-endosomal, cholesterol concentration is decreased by a factor equal to or greater than 1 .5, 2, 2.5, 3, 3.5, 4, 4.5, 5, 5.5, 6, 6.5, 7, 7.5, 8, 8.5, 9, 9.5, 10, 15, 20, 25, 30, 35, 40, 45, or 50; and / or

[0184] — a restored or partially restored intralysosomal glycolipid concentration, preferably wherein the intralysosomal glycolipid concentration is decreased after introduction of the inhibitor, more preferably wherein the intralysosomal glycolipid concentration is decreased by a factor equal to or greater than 1 .5, 2, 2.5, 3, 3.5, 4, 4.5, 5, 5.5, 6, 6.5, 7, 7.5, 8, 8.5, 9, 9.5, 10, 15, 20, 25, 30, 35, 40, 45, or 50; and / or

[0185] — a restored or partially restored endosomal, preferably late-endosomal, glycolipid concentration, preferably wherein the endosomal, preferably the late-endosomal, glycolipid concentration is decreased after introduction of the inhibitor, more preferably wherein the endosomal, preferably the late-endosomal, glycolipid concentration is decreased by a factor equal to or greater than 1 .5, 2, 2.5, 3, 3.5, 4, 4.5, 5, 5.5, 6, 6.5, 7, 7.5, 8, 8.5, 9, 9.5, 10, 15, 20, 25, 30, 35, 40, 45, or 50; and / or

[0186] — a restored or partially restored intralysosomal glycosphingolipid concentration, preferably wherein the intralysosomal glycosphingolipid concentration is decreased after introduction of the inhibitor, more preferably wherein the intralysosomal glycosphingolipid concentration is decreased by a factor equal to or greater than 1 .5, 2, 2.5, 3, 3.5, 4, 4.5, 5, 5.5, 6, 6.5, 7,

[0187] 7.5, 8, 8.5, 9, 9.5, 10, 15, 20, 25, 30, 35, 40, 45, or 50; and / or

[0188] — a restored or partially restored endosomal, preferably late-endosomal, glycosphingolipid concentration, preferably wherein the endosomal, preferably the late-endosomal, glycosphingolipid concentration is decreased after introduction of the inhibitor, more preferably wherein the endosomal, preferably the late-endosomal, glycosphingolipid concentration is decreased by a factor equal to or greater than 1 .5, 2, 2.5, 3, 3.5, 4, 4.5, 5,

[0189] 5.5, 6, 6.5, 7, 7.5, 8, 8.5, 9, 9.5, 10, 15, 20, 25, 30, 35, 40, 45, or 50; wherein a partially restored concentration means that the concentration is significantly closer to that in a corresponding healthy subject after introduction of said inhibitor, preferably by the factors described above, wherein a restored concentration means that the concentration is essentially the same as that in in a corresponding healthy subject.

[0190] In specific aspects, administration of an inhibitor according to the invention to a subject in need thereof results in one of the effects described above, particularly in a reduction of the intralysosomal cholesterol concentration and / or the intralysosomal glycosphingolipid concentration. Niemann-Pick disease type C has a wide clinical spectrum. Affected individuals may have enlargement of the spleen (splenomegaly) and liver (hepatomegaly), or enlarged spleen or liver combined (hepatosplenomegaly).

[0191] Progressive neurological disease is the hallmark of Niemann-Pick type C disease. Classically, children with NPC may initially present with delays in reaching normal developmental milestones skills before manifesting cognitive decline (dementia).

[0192] Neurological signs and symptoms include cerebellar ataxia (unsteady walking with uncoordinated limb movements), dysarthria (slurred speech), dysphagia (difficulty in swallowing), tremor, epilepsy (both partial and generalized), vertical supranuclear palsy (upgaze palsy, downgaze palsy, saccadic palsy or paralysis), sleep inversion, gelastic cataplexy (sudden loss of muscle tone or drop attacks), dystonia (abnormal movements or postures caused by contraction of agonist and antagonist muscles across joints), most commonly begins with inturning of one foot when walking (action dystonia) and may spread to become generalized, spasticity (velocity dependent increase in muscle tone), hypotonia, ptosis (drooping of the upper eyelid), microcephaly (abnormally small head), psychosis, dementia (preferably progressive dementia), progressive hearing loss, bipolar disorder, major and psychotic depression that can include hallucinations, delusions, mutism, or stupor.

[0193] In specific aspects, an inhibitor of the invention is able to alleviate at least of the following symptoms when said inhibitor is introduced in a subject suffering from Niemann-Pick disease type C: splenomegaly, hepatomegaly, hepatosplenomegaly, cerebellar ataxia, dysarthria, dysphagia, tremorepilepsy, vertical supranuclear palsy, sleep inversion, gelastic cataplexy, dystonia, spasticity, hypotonia, ptosis, psychosis, dementia (preferably progressive dementia), progressive hearing loss, bipolar disorder, major and psychotic depression, hallucinations, delusions, mutism, and stupor. The alleviation of a symptom is generally acknowledged by the a skilled person (and in particular, by the treating physician) as an improvement.

[0194] In specific aspects, administration of an inhibitor according to the invention to a subject in need thereof results in the alleviation of at least of the following symptoms: cerebellar ataxia, dysarthria, vertical gaze palsy, motor impairment, dysphagia, psychotic episodes, and dementia (preferably progressive dementia).

[0195] TREATMENT OPTIONS

[0196] Wherever an inhibitor or a composition for use as a medicament is disclosed, a corresponding method for the manufacture or the production of a medicament comprising such an inhibitor or such a composition, a corresponding method of treatment comprising the administration of the inhibitor or composition to a subject in need thereof, and a corresponding use of such an inhibitor or such a composition as a medicament are also disclosed. In all these contexts, the inhibitor according to the invention and the composition according to the invention may be referred to as a medicament according to the invention.

[0197] A medicament according to the invention may be administered orally, nasally, buccally, sublingually, vaginally, parenterally, topically, systemically, intravenously, subcutaneously, intraperitoneally, intramuscularly, intrathecally, by inhalation or epidurally.

[0198] A medicament according to the invention, may be administered separately, sequentially or simultaneously in combination with another medicaments.

[0199] As used herein, the term "simultaneous" therapeutic use refers to the administration of at least two active ingredients by the same route and at the same time or at substantially the same time. The term "separate" therapeutic use refers to an administration of at least two active ingredients at the same time or at substantially the same time by different routes. The term "sequential" therapeutic use refers to administration of at least two active ingredients at different times, the administration route being identical or different. More particularly, sequential use refers to the whole administration of one of the active ingredients before administration of the other or others commences. It is thus possible to administer one of the active ingredients over several minutes, hours, or days before administering the other active ingredient or ingredients. There is no simultaneous treatment in this case.

[0200] In the context of this application, the terms treating" or "treatment" refer to therapeutic treatment, wherein the object is to prevent, reduce, alleviate or slow down (lessen), respectively and as applicable, the targeted pathologic disorder or disease and / or its progression in a subject. In particular, said terms relate to a treatment which has the object of improving one or more symptoms and / or physiological parameters that are caused by, associated with and / or characteristic of the disease or disorder that is to be treated, and / or the object to preventing that such symptom(s) to arise and / or that such symptom(s) or physiological parameters) further deteriorate. Based on his general knowledge and the further disclosure herein, the skilled person (and in particular, the treating physician) will be able to suitably determine and measure said symptom(s) or physiological parameter(s), depending on the specific disease involved.

[0201] In the context of this application, the terms "prevention" or "preventing" of a disorder or disease refers to a compound that, in a statistical sample, reduces the occurrence of symptoms of a disorder or disease in the treated sample relative to an untreated control sample, or delays the onset or reduces the severity of one or more symptoms of the disorder or condition relative to the untreated control sample.

[0202] A medicament according to the invention is administered to a subject in need thereof in an effective amount (i.e., amount that have desired therapeutic effect). Preferably, an effective amount refers to an amount of an inhibitor according to the invention comprised in said medicament. The dose and dosage regimen will depend upon the degree of the infection in the subject, the characteristics of the particular inhibitor according to the invention, e.g., its therapeutic index, the subject, and the subject's history. Certain factors may influence the dosage and timing required to effectively treat a subject, including but not limited to, the severity of the disease or disorder, previous treatments, the general health and / or age of the subject, and other diseases present. Moreover, treatment of a subject with a therapeutically effective amount of the medicaments according to the invention can include a single treatment or a series of treatments.

[0203] The effective amount may be determined during pre-clinical trials and clinical trials by methods familiar to physicians and clinicians. An effective amount of a peptide useful in the methods may be administered to a subject in need thereof by any of a number of well-known methods for administering pharmaceutical compounds.

[0204] Dosage, toxicity and therapeutic efficacy of a medicament according to the invention can be determined by standard pharmaceutical procedures in cell cultures or experimental animals, e.g., for determining the LD50 (the dose lethal to 50% of the population) and the ED50 (the dose therapeutically effective in 50% of the population). The dose ratio between toxic and therapeutic effects is the therapeutic index and it can be expressed as the ratio LD50 / ED50. Medicaments that exhibit high therapeutic indices are preferred.

[0205] The data obtained from the cell culture assays and animal studies can be used in formulating a range of dosage for use in humans. The dosage lies preferably within a range of circulating concentrations that include the ED50 with little or no toxicity. The dosage may vary within this range depending upon the dosage form employed and the route of administration utilized. For any medicament according to the invention, the therapeutically effective dose can be estimated initially from cell culture assays.

[0206] Compositions

[0207] In a further aspect, the invention provides a composition comprising an inhibitor according to the invention and a pharmaceutically acceptable excipient, preferably for use as a medicament, more preferably for use in the treatment of a disease characterized by lysosomal dysregulation. A related aspect pertains to a method of preparing such a composition. Such compositions are referred to in the current application as compositions according to or of the invention.

[0208] All specific embodiments disclosed above for an inhibitor according to the invention may be applied accordingly for an inhibitor according to the invention comprised in a composition according to the invention.

[0209] A composition according to the invention may be presented or formulated as capsules, tablets, powders, granules, solutions, suspensions in aqueous or non-aqueous liquids, edible, oil-in-water liquid emulsions, water-in-oil liquid emulsions, solution, syrups and elixirs, in microencapsulated form, liposome delivery systems, such as small unilamellar vesicles, large unilamellar vesicles and multilamellar vesicles, transdermal patches, ointments, creams, suspensions, lotions, powders, solutions, pastes, gels, drops, sprays, aerosols, oils, lozenges, pastilles, mouth washes, suppositories, enemas, aqueous and non-aqueous sterile injection solutions, and so on. It will be appreciated that the compositions may include other agents conventional in the art having regard to the type of formulation.

[0210] Non-limiting examples of a pharmaceutically acceptable carrier comprised in a composition are saline, solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents, and the like, compatible with pharmaceutical administration. Supplementary active compounds, besides an inhibitor according to the invention, can also be incorporated into the compositions.

[0211] A composition according to the invention formulated as solutions or suspensions used for parenteral, intradermal, or subcutaneous application can include the following components: a sterile diluent such as water for injection, saline solution, fixed oils, polyethylene glycols, glycerine, propylene glycol or other synthetic solvents; antibacterial agents such as benzyl alcohol or methyl parabens; antioxidants such as ascorbic acid or sodium bisulfite; chelating agents such as ethylenediaminetetraacetic acid; buffers such as acetates, citrates or phosphates and agents for the adjustment of tonicity such as sodium chloride or dextrose. pH can be adjusted with acids or bases, such as hydrochloric acid or sodium hydroxide. The parenteral preparation can be enclosed in ampoules, disposable syringes or multiple dose vials made of glass or plastic.

[0212] A composition according to the invention formulated as compositions suitable for injectable use can include sterile aqueous solutions (where water soluble) or dispersions and sterile powders for the extemporaneous preparation of sterile injectable solutions or dispersion. For intravenous administration, suitable carriers include physiological saline, bacteriostatic water, Cremophor EL™ (BASF, Parsippany, N.J.) or phosphate buffered saline (PBS). In all cases, a composition for parenteral administration must be sterile and should be fluid to the extent that easy syringability exists. It should be stable under the conditions of manufacture and storage and must be preserved against the contaminating action of microorganisms such as bacteria and fungi.

[0213] In a composition according to the invention prevention of the action of microorganisms can be achieved by various antibacterial and antifungal agents, for example, parabens, chlorobutanol, phenol, ascorbic acid, thiomerasol, and the like. Glutathione and other antioxidants can be included to prevent oxidation. In many cases, it will be preferable to include isotonic agents, for example, sugars, polyalcohols such as mannitol, sorbitol, or sodium chloride in the composition. Prolonged absorption of the injectable compositions can be brought about by including in the composition an agent that delays absorption, for example, aluminum monostearate or gelatin.

[0214] A composition according to the invention formulated as oral compositions generally include an inert diluent or an edible carrier. For the purpose of oral therapeutic administration, the inhibitor according to the invention can be incorporated with excipients and used in the form of tablets, troches, or capsules, e.g., gelatin capsules. Oral compositions can also be prepared using a fluid carrier for use as a mouthwash. Pharmaceutically compatible binding agents, and / or adjuvant materials can be included as part of the composition. The tablets, pills, capsules, troches and the like can contain any of the following ingredients, or compounds of a similar nature: a binder such as microcrystalline cellulose, gum tragacanth or gelatin; an excipient such as starch or lactose, a disintegrating agent such as alginic acid, Primogel, or corn starch; a lubricant such as magnesium stearate or Sterotes; a glidant such as colloidal silicon dioxide; a sweetening agent such as sucrose or saccharin; or a flavoring agent such as peppermint, methyl salicylate, or orange flavoring.

[0215] A composition according to the invention may be formulated for administration by inhalation, the inhibitor according to the invention can be delivered in the form of an aerosol spray from a pressurized container or dispenser, which contains a suitable propellant, e.g., a gas such as carbon dioxide, or a nebulizer.

[0216] A composition according to the invention may be formulated for transmucosal or transdermal administration, penetrants appropriate to the barrier to be permeated are used in the formulation. Such penetrants are generally known in the art, and include, for example, for transmucosal administration, detergents, bile salts, and fusidic acid derivatives. Transmucosal administration can be accomplished through the use of nasal sprays. For transdermal administration, the active compounds are formulated into ointments, salves, gels, or creams as generally known in the art. In one aspect, transdermal administration may be performed by iontophoresis.

[0217] A composition according to the invention may comprise a carrier system such as a colloidal system. The colloidal system can be a liposome, a phospholipid bilayer vehicle. In one aspect, the inhibitor according to the invention is encapsulated in a liposome. An inhibitor according to the invention can also be loaded into a particle prepared from pharmaceutically acceptable ingredients including, but not limited to, soluble, insoluble, permeable, impermeable, biodegradable or gastroretentive polymers or liposomes. Such particles include, but are not limited to, nanoparticles, biodegradable nanoparticles, microparticles, biodegradable microparticles, nanospheres, biodegradable nanospheres, microspheres, biodegradable microspheres, capsules, emulsions, liposomes, micelles and viral vector systems.

[0218] Uses

[0219] In a further aspect, the invention provides the use of a PLA2G15 inhibitor according to the invention or a composition according to the invention for binding and / or inhibiting PLA2G15. Preferably, the binding of PLA2G15 results in in the inhibition of PLA2G15. The inhibition of PLA2G15 is described in more detail above.

[0220] All specific embodiments disclosed above for an inhibitor according to the invention and a composition according to the invention may be applied accordingly for the uses and therapeutic uses described below. In embodiments, the use of a PLA2G15 inhibitor according to the invention or a composition according to the invention is for specifically binding and / or specifically inhibiting PLA2G15. Specifically means that the binding or inhibition stems from the amino acid sequence dependent molecular interaction between the inhibitor, or the inhibitor comprised in the composition, and PLA2G15. As such, the inhibitor or composition is not able to significantly bind or inhibit other enzymes that may share a similar function but a different amino acid structure.

[0221] In embodiments, the use of a PLA2G15 inhibitor according to the invention or a composition according to the invention for binding and / or inhibiting PLA2G15 comprises a PLA2G15 inhibitor which does not specifically bind and / or specifically inhibit other phospholipases than PLA2G15. In other words, whereas the inhibitor, or the inhibitor comprised in the composition, is able to specifically bind or inhibit PLA2G15, is not able to do so with other phospholipases. Other phospholipases include group I phospholipases (PLA2G1 B), group II phospholipases (PLA2G2A, PLA2G2C, PLA2G2D, PLA2G2E, PLA2G2F), group III phospholipases (PLA2G3), group IV phospholipases (PLA2G4A, PLA2G4B, PLA2G4C, PLA2G4D, PLA2G4E, PLA2G4F), group V phospholipases (PLA2G5), group VI phospholipases (PLA2G6), group VII phospholipases (PLA2G7), group X phospholipases (PLA2G10) and group XII phospholipases (PLA2G12A, PLA2G12B). Preferably, the PLA2G15 inhibitor or composition does not significantly bind or inhibit the phospholipases in this list.

[0222] The inhibition of PLA2G15 can be expressed as the half maximal inhibitory concentration (IC50), as known to the skilled person. Lower IC50 values correspond with higher potencies to inhibit PLA2G15. IC50 can be determined using the protocol of Example 2.

[0223] In embodiments, the use of a PLA2G15 inhibitor according to the invention or a composition according to the invention for (specifically) inhibiting PLA2G15 comprises a PLA2G15 inhibitor having an IC50 for PLA2G15 of less than 50, 45, 40, 35, 30, 25, 20, 15, 10, 9, 8, 7, 6, 5,4, 3, 2, 1 , 0.9, 0.8, 0.7, 0.6, 0.5, 0.4, 0.3, 0.2, 0.1 , 0.09, 0.08, 0.07, 0.06, 0.05, 0.04, 0.03, 0.02, 0.01 pmol / L, preferably as measured by the 4-nitrophenyl butyrate activity assay as described herein.

[0224] In embodiments, the use of a PLA2G15 inhibitor according to the invention or a composition according to the invention for (specifically) inhibiting PLA2G15 comprises a PLA2G15 inhibitor having a ratio between its IC50 for PLA2G15 and its IC50 for other phospholipases, as provided above, equal to or higher than 1000, 950, 900, 850, 800, 750, 700, 650, 600, 550, 500, 450, 400, 350, 300, 250, 200, 150, 100, 95, 90, 85, 80, 75, 70, 65, 60, 55, 50, 45, 40, 35, 30, 25, 20, 15, 10, 9, 8, 7, 6, 5, 4, 3, or 2 pmol / L.

[0225] In embodiments, the uses above are for use in vitro.

[0226] In embodiments, the uses above are for use in vivo.

[0227] In embodiments, (the use of) a PLA2G15 inhibitor according to the invention or a composition according to the invention does not induce an accumulation of phospholipids. Definitions

[0228] All documents cited in the present specification are hereby incorporated by reference in their entirety. Unless otherwise defined, all terms used in disclosing the invention, including technical and scientific terms, have the meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. By means of further guidance, term definitions are included to better appreciate the teaching of the present invention.

[0229] Unless indicated otherwise, all methods, steps, techniques and manipulations that are not specifically described in detail can be performed and have been performed in a manner known per se, as will be clear to the skilled person. Reference is made to the standard handbooks, to the general background art referred to above and to the further references cited therein.

[0230] As used herein, the singular forms 'a', 'an', and 'the' include both singular and plural referents unless the context clearly dictates otherwise.

[0231] The terms 'comprising', 'comprises' and 'comprised of as used herein are synonymous with 'including', 'includes' or 'containing', 'contains', and are inclusive or open-ended and do not exclude additional, non-recited members, elements or method steps.

[0232] The recitation of numerical ranges by endpoints includes all numbers and fractions subsumed within the respective ranges, as well as the recited endpoints.

[0233] Physiological conditions are defined in the context of this application as typical environmental conditions in a vertebrate, mammalian or human cell or tissue that is in homeostasis and is not subject to extraordinary external stress. Preferably, physiological conditions mean a temperature from 25°C up to 45°C, more preferably from 30°C up to 40°C.

[0234] A concentration is preferably a molar concentration, preferably a molar concentration per weight or per volume, most preferably measured under physiological conditions.

[0235] A subject is defined in the context of this application as a (living) organism, unless explicitly stated otherwise. A subject may be any organism, including invertebrates and vertebrates. Preferably, a subject is a vertebrate. More preferably, a vertebrate is a starfish or a mammal. Even more preferably, a mammal is a rat, a mouse, a rabbit or a human. Most preferably, a mammal is a human. In an alternative specific aspect, a subject is a non-human animal, more preferably a non-human vertebrate, most preferably a non-human mammal.

[0236] An organelle is preferably a lysosome or an endosome, more preferably a lysosome. An endosome is preferably a late endosome. An increase of a parameter by a factor equal to or higher than X is defined in the context of this application as a change of said parameter from its initial value A to a value equal to or higher than A*X.

[0237] An increase of a parameter by a factor equal to or lower than X is defined in the context of this application as a change of said parameter from its initial value A to a value equal to or lower than A*X.

[0238] A decrease of a parameter by a factor equal to or lower than X is defined in the context of this application as a change of said parameter from its initial value A to a value equal to or lower than A*X.

[0239] A decrease of a parameter by a factor equal to or higher than X is defined in the context of this application as a change of said parameter from its initial value A to a value equal to or higher than A*X.

[0240] A compound or moiety comprising n submoieties, such as atoms, functional groups or ring systems, means that the number of submoieties contained in the compound or moiety is exactly n.

[0241] Specifically named C1-4 alkyls and C1-4 alkylenes, such as methyl, ethyl, propyl, etc., may be unsubstituted (apart from their relevant attachment to the rest of the PLA2G15 inhibitor) or substituted with one or more of halogens or pseudohalogens. For example, methyl may referto CH3, CH2OH, CHCh, CF3, etc. In contrast, CH3 only refers to an unsubstituted methyl.

[0242] Wherever a substitution with “n of A and B” or “n of A or B” is mentioned, a total of n substitutions independently selected from A and B is meant. In other words, n exocyclic hydrogen atoms are independently replaced by moieties selected from A and B.

[0243] A parameter that is essentially the same as in a corresponding composition, organelle, cell fraction, cell, membrane, tissue or organ derived from a healthy subject or as in a corresponding healthy subject, preferably means that the value of said parameter cannot be distinguished by a skilled person from the value of a corresponding parameter in a corresponding composition, organelle, cell fraction, cell, membrane, tissue or organ derived from a healthy subject or in a corresponding healthy subject, and / or that the value of said parameter would be interpreted by a skilled person as measured in a corresponding composition, organelle, cell fraction, cell, membrane, tissue or organ derived from a healthy subject or in a corresponding healthy subject.

[0244] An alteration of a parameter which is significantly smaller after introduction of an inhibitor in a composition, organelle, cell fraction, cell, membrane, tissue, organ or subject preferably means that the absolute difference between the value of said parameter and the value of a corresponding parameter in a corresponding composition, organelle, cell fraction, cell, membrane, tissue or organ derived from a healthy subject or in a corresponding healthy subject is decreased by a factor equal to or lower than 0.95, 0.9, 0.85, 0.8, 0.75, 0.7, 0.65, 0.6, 0.55, 0.5, 0.45, 0.4, 0.35, 0.3, 0.25, 0.2, 0.15, 0.1 , 0.09, 0.08, 0.07, 0.06, 0.05, 0.04, 0.03, 0.02 or 0.01 after said introduction. Any parameter referred to herein is preferably determined using the specific method, assay or methodology described herein. Where the present specification does not mention or describe a specific method, assay or methodology for determining said parameter, said parameter can be measured in a manner suitable per se, as will be clear to the skilled person based upon reading the present disclosure.

[0245] Each amino acid sequence described herein by virtue of its identity or similarity percentage (at least 60%) with a given amino acid sequence respectively has in a further specific aspect an identity or a similarity of at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 97%, 98%, 99% or more identity or similarity with the given amino acid sequence respectively. In a specific aspect, sequence identity or similarity is determined by comparing the whole length of the sequences as identified herein. Unless otherwise indicated herein, identity or similarity with a given SEQ ID NO means identity or similarity based on the full length of said sequence ( / .e. over its whole length or as a whole).

[0246] Sequence identity is defined in the context of this application as a relationship between two or more amino acid (polypeptide or protein) sequences or two or more nucleic acid (polynucleotide) sequences, as determined by comparing the sequences. The identity between two amino acid sequences is preferably defined by assessing their identity within a whole SEQ ID NO as identified herein or part thereof. Part thereof may mean at least 50% of the length of the SEQ ID NO, or at least 60%, or at least 70%, or at least 80%, or at least 90%.

[0247] In the art, sequence identity also means the degree of sequence relatedness between amino acid sequences, as the case may be, as determined by the match between strings of such sequences. Sequence similarity between two amino acid sequences is determined by comparing the amino acid sequence and its conserved amino acid substitutes of one polypeptide to the sequence of a second polypeptide. Sequence identity and similarity can be readily calculated by known methods, including but not limited to those described in Computational Molecular Biology, Lesk, A. M., ed., Oxford University Press, New York, 1988; Biocomputing: Informatics and Genome Projects, Smith, D. W., ed., Academic Press, New York, 1993; Computer Analysis of Sequence Data, Part I, Griffin, A. M., and Griffin, H. G., eds., Humana Press, New Jersey, 1994; Sequence Analysis in Molecular Biology, von Heine, G., Academic Press, 1987; and Sequence Analysis Primer, Gribskov, M. and Devereux, J., eds., M Stockton Press, New York, 1991 ; and Carillo, H., and Lipman, D., SIAM J. Applied Math., 48:1073 (1988).

[0248] Preferred methods to determine sequence identity are designed to give the largest match between the sequences tested. Methods to determine sequence identity and similarity are codified in publicly available computer programs. Preferred computer program methods to determine sequence identity and similarity between two sequences include e.g. the GCG program package (Devereux, J., et al., Nucleic Acids Research 12 (1): 387 (1984)), BestFit, FASTA, BLASTN, and BLASTP (Altschul, S. F. et al., J. Mol. Biol. 215:403-410 (1990)), EMBOSS Needle (Madeira, F., et al., Nucleic Acids Research 47(W1): W636-W641 (2019)). The BLAST program is publicly available from NCBI and other sources (BLAST Manual, Altschul, S„ et al., NCBI NLM NIH Bethesda, MD 20894; Altschul, S„ et al., J. Mol. Biol. 215:403-410 (1990)). The EMOSS program is publicly available from EMBL-EBI. The well-known Smith Waterman algorithm may also be used to determine identity. The EMBOSS Needle program is the preferred program used.

[0249] Preferred parameters for polypeptide sequence comparison include the following: Algorithm: Needleman and Wunsch, J. Mol. Biol. 48 (3):443-453 (1970); Comparison matrix: BLOSUM62 from Henikoff and Henikoff, Proc. Natl. Acad. Sci. USA. 89:10915-10919 (1992); Gap Open Penalty: 10; and Gap Extend Penalty: 0.5. A program useful with these parameters is publicly available as the EMBOSS Needle program from EMBL-EBI. The aforementioned parameters are the default parameters for a Global Pairwise Sequence alignment of proteins (along with no penalty for end gaps).

[0250] Preferred parameters for nucleic acid comparison include the following: Algorithm: Needleman and Wunsch, J. Mol. Biol. 48:443-453 (1970); Comparison matrix: DNAfull; Gap Open Penalty: 10; Gap Extend Penalty: 0.5. A program useful with these parameters is publicly available as the EMBOSS Needle program from EMBL-EBI. The aforementioned parameters are the default parameters for a Global Pairwise Sequence alignment of nucleotide sequences (along with no penalty for end gaps).

[0251] Optionally, in determining the degree of amino acid (sequence) similarity, the skilled person may also take into account so-called "conservative" amino acid substitutions, as will be clear to the skilled person. Conservative amino acid substitutions refer to the interchangeability of residues having similar side chains. For example, a group of amino acids having aliphatic side chains is glycine, alanine, valine, leucine, and isoleucine; a group of amino acids having aliphatic-hydroxyl side chains is serine and threonine; a group of amino acids having amide-containing side chains is asparagine and glutamine; a group of amino acids having aromatic side chains is phenylalanine, tyrosine, and tryptophan; a group of amino acids having basic side chains is lysine, arginine, and histidine; a group of amino acids having acidic side chains is aspartate and glutamate; and a group of amino acids having sulphur-containing side chains is cysteine and methionine. Preferred conservative substitutions for each of the naturally occurring amino acids are as follows: Ala to Ser; Arg to Lys or Gin; Asn to Asp, His or Ser; Asp to Glu or Asn; Gin to Glu, Lys or Arg; Glu to Lys, Asp, Gin; His to Tyr or Asn; lie to Leu, Vai, or Met; Leu to lie, Met or Vai; Lys to Arg, Gin or Glu; Met to Vai, Leu or lie; Phe to Trp or Tyr; Ser to Thr, Ala or Asn; Thr to Ser; Trp to Tyr or Phe; Tyr to His, Trp or Phe; and Vai to He, Leu or Met. Substitutional variants of the amino acid sequence disclosed herein are those in which at least one residue in the disclosed sequences has been removed and a different residue inserted in its place. Preferably, the amino acid change is conservative. References

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[0297] 47. S. Chapuy-Regaud Biochimie 95 (2013) 1677el688 Examples

[0298] The following examples are offered for illustrative purposes only, and are not intended to limit the scope of the present invention in any way.

[0299] Example 1: Synthesis of compounds

[0300] EXAMPLE 1.1: SYNTHESIS OF (1)

[0301] Synthesis of compound 2

[0302] A mixture of compound 1 (10 g, 78.61 mmol, 9.17 mL, 1 eq), benzoic acid (9.60 g, 78.61 mmol, 12.00 mL, 1 eq), HOBt (12.75 g, 94.33 mmol, 1.2 eq), EDCI (18.08 g, 94.33 mmol, 1 .2 eq) and DIEA (25.40 g, 196.53 mmol, 34.23 mL, 2.5 eq) in DMF (100 mL) was stirred at 20 °C for 16 h. The mixture was concentrated under reduced pressure to give a residue. The residue was purified by reversed- phase HPLC (0.1% FA condition), the eluent was concentrated and then freeze dried. Compound 2 (16 g, 69.17 mmol, 87.99% yield) was obtained as a yellow oil.1H NMR: (400 MHz, CDCI3) δ = 7.73 (d, J = 7.3 Hz, 2H), 7.53 - 7.47 (m, 1 H), 7.46 - 7.39 (m, 2H), 7.19 (d, J = 5.1 Hz, 1 H), 7.00 - 6.95 (m, 1 H), 6.89 (d, J = 3.3 Hz, 1 H), 6.34 (br s, 1 H), 3.74 (q, J = 6.2 Hz, 2H), 3.17 (t, J = 6.5 Hz, 2H).

[0303] Synthesis of compound 3

[0304] To a solution of compound 2 (16 g, 69.17 mmol, 1 eq) in Tol. (160 mL) was added POCh (15.91 g, 103.76 mmol, 9.64 mL, 1 .5 eq) under 0 °C, then the mixture was stirred at 110 °C for 3 h. The mixture was added into water (500 mL) slowly, the pH was adjusted to about 7 with K2CO3, then the mixture was extracted with EtOAc (100 mL x 3), the organic phase was dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. Compound 3 (15 g, crude) was obtained as a brown oil. Synthesis of compound 4

[0305] To a solution of compound 3 (6 g, 28.13 mmol, 1 eq) in MeOH (60 mL) was added NaBFL (2.17 g, 57.36 mmol, 2.04 eq) at 0 °C under N2, and the mixture was stirred at 20 °C under N2for 2 h. To the mixture was added into aqueous NH4CI solution (20 mL) slowly and the resulting mixture was extracted with EtOAc (20 mL x 3), the organic phase was dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. Compound 4 (5.8 g, 26.94 mmol, 95.76% yield) was obtained as a yellow solid. LCMS: RT = 0.253 min, m / z = 216.1 (M+H)+.

[0306] Synthesis of compound 5, compound 5A and 5B

[0307] A mixture of compound 4 (1 g, 4.64 mmol, 1 eq), 3-(tert-butoxycarbonylamino) propanoic acid (878.76 mg, 4.64 mmol, 1 eq), DIEA (1 .20 g, 9.29 mmol, 1 .62 mL, 2 eq), HOBt (753.06 mg, 5.57 mmol, 1.2 eq) and EDCI (1.07 g, 5.57 mmol, 1.2 eq) in DMF (10 mL) was stirred at 20 °C for 16 h. To the mixture was added water (20 mL) and extracted with EtOAc (20 mL x 3), the combined organic phase was washed with brine (50 mL), dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by flash silica gel chromatography (ISCO®; 20 g SepaFlash® Silica Flash Column, Eluent of 45-55% Ethyl acetate / Petroleum ethergradient @ 60 mL / min; TLC (Petroleum ether : Ethyl acetate = 1 : 1 ; Rf = 0.53) ), the eluent was concentrated.

[0308] A part of residue was separated by SFC (column: DAICEL CHIRALCEL OJ (250 mm*30 mm, 10um); mobile phase: [0.1 %NH3H2O IRA]; B%: 25%- 25%, 3.2 min).

[0309] Peak 1 (Rt = 0.892 min) was collected to give compound 5A (100 mg, 258.73 μmol, 5.57% yield) was obtained as a yellow oil. LCMS: RT = 0.985 min, m / z = 387.1 (M+H)+. SFC: RT = 0.873 min.

[0310] Peak 2 (Rt = 1.183 min) was collected to give compound 5B (100 mg, 258.73 μmol, 5.57% yield) was obtained as a yellow oil. LCMS: RT = 0.970 min, m / z = 387.1 (M+H)+. SFC: RT =1 .175 min.

[0311] The rest of compound 5 (1 .2 g, 3.10 mmol, 66.85% yield) was obtained as a yellow oil.

[0312] LCMS: RT = 0.983 min, m / z = 387.1 (M+H)+.

[0313] Synthesis of compound 6A A mixture of compound 5A (0.1 g, 258.73 μmol, 1 eq) and TFA (462.00 mg, 4.05 mmol, 0.3 mL, 15.66 eq) in DCM (1 mL) was stirred at 20 °C for 2 h. The mixture was concentrated under reduced pressure to give a residue.. Compound 6A (100 mg, 249.74 μmol, 96.53% yield, TFA) was obtained as a yellow oil. LCMS: RT = 0.764 min, m / z = 287.0 (M+H)+.

[0314] Synthesis of (1)

[0315] A mixture of compound 6A (50 mg, 174.59 μmol, 1 eq), 1-(trifluoromethyl)cyclopentane- carboxylic acid (31.80 mg, 174.59 μmol, 1 eq), HOBt (28.31 mg, 209.50 μmol, 1.2 eq), EDCI (40.16 mg, 209.50 μmol, 1 .2 eq) and DIEA (45.13 mg, 349.17 μmol, 60.82 μL, 2 eq) in DMF (1 mL) was stirred at 20 °C for 16 h. The reaction mixture was concentrated under vacuum to give a residue. The residue was purified by prep-HPLC (column: Unisil 3 - 100 C18 Ultra 150*50 mm*3 um;mobile phase : [water (FA) -ACN] ;B% : 55%- 85%, 7 min) to give (1) (30.83 mg, 67.75 μmol, 38.81 % yield, 99% purity) as white gum.1H NMR: (400 MHz, CDCI3) δ = 7.28 (br s, 3H), 7.25 - 7.20 (m, 1 H), 7.17 (br d, J = 4.9 Hz, 1 H), 6.93 - 6.85 (m, 2H), 6.76 - 6.71 (m, 1 H), 3.83 (br dd, J = 4.4, 14.5 Hz, 1 H), 3.70 - 3.59 (m, 2H), 3.38 - 3.29 (m, 1 H), 3.03 - 2.91 (m, 2H), 2.70 - 2.51 (m, 2H), 2.28 - 2.18 (m, 2H), 2.02 - 1.91 (m, 2H), 1 .75 - 1.64 (m, 4H); LCMS: RT = 0.651 min, m / z = 451 .2 (M+H)+.

[0316] EXAMPLE 1.2: SYNTHESIS OF (2)

[0317] Synthesis of compound 3

[0318] A mixture of compound 2 (1 .08 g, 5.95 mmol, 1 eq, HCI), compound 1 (1 g, 5.95 mmol, 1 eq), DIEA (2.31 g, 17.85 mmol, 3.1 1 mL, 3 eq), HOBt (964.51 mg, 7.14 mmol, 1.2 eq) and EDCI (1.37 g, 7.14 mmol, 1 .2 eq) in DMF (10 mL) was stirred at 25 °C for 16 h. The mixture was purified by reversed-phase HPLC (0.1 % FA condition), the eluent was concentrated and then freeze dried. Compound 3 (1.3 g, 4.40 mmol, 74.01 % yield) was obtained as a yellow solid.1H NMR: (400 MHz, CDCI3) δ = 6.32 (br s, 1 H), 3.53 (q, J = 6.0 Hz, 2H), 2.63 - 2.51 (m, 2H), 2.50 - 2.38 (m, 4H), 2.11 - 1 .86 (m, 2H), 1 .45 (s, 9H).

[0319] Synthesis of compound 4 A mixture of compound 3 (1.3 g, 4.40 mmol, 1 eq) in DCM (10 mL) and TFA (10 mL) was stirred at 25 °C for 2 h. The mixture was concentrated at reduced pressure to give compound 4 (1 .3 g, crude) as a yellow oil. LCMS: RT =0.425 min, m / z =240.0 (M+H)+.

[0320] Synthesis of compound (2) A mixture of compound 5 (50 mg, 192.78 μmol, 1 eq), compound 4 (46.11 mg, 192.78 μmol, 1 eq), DIEA (74.74 mg, 578.34 μmol, 100.73 μL, 3 eq), HOBt (31 .26 mg, 231 .34 μmol, 1 .2 eq) and EDCI (44.35 mg, 231 .34 μmol, 1 .2 eq) in DMF (1 mL) was stirred at 20 °C for 16 h. The mixture was purified by prep- HPLC (column: Phenomenex luna C18 150*25 mm* 10um; mobile phase: [water (FA) -ACN]; B%: 48%- 78%, 10 min). Compound (2) (49.13 mg, 100.19 μmol, 51.97% yield, 98% purity) was obtained as a yellow gum.1H NMR: (400 MHz, CDCI3) δ = 7.21 (t, J = 8.1 Hz, 1 H), 6.87 - 6.79 (m, 3H), 6.73 (s, 2H), 6.43 - 6.33 (m, 1 H), 3.85 - 3.74 (m, 4H), 3.73 - 3.55 (m, 2H), 3.42 - 3.26 (m, 1 H), 3.09 - 2.69 (m, 3H), 2.66 - 2.47 (m, 4H), 2.46-2.35 (m, 4H), 2.08 - 1 .83 (m, 2H); LCMS: RT =0.834 min, m / z =481 .2 (M+H)+. SFC: RT =1.410 min

[0321] EXAMPLE 1.3: SYNTHESIS OF (3)

[0322] Synthesis of compound 2A, compound 2B A mixture of compound 1 (900 mg, 6.37 mmol, 1 eq) and 3-methoxybenzaldehyde (867.59 mg, 6.37 mmol, 774.63 μL, 1 eq) in Tol. (10 mL) was stirred at 110 °C for 16 h. The mixture was concentrated under reduced pressure to remove the solution, then to the mixture was added TFA (10 mL) and the resulting mixture was stirred at 25 °C for 16 h. The mixture was concentrated at reduced pressure to give a residue and the residue was purified by prep-HPLC (0.1 % FA condition), the eluent was concentrated and then freeze dried to give a solid. The solid was separated by SFC (column : DAICEL CHIRALPAK AD (250 mm*30 mm, 10um) ;mobile phase : [0.1 %NH3H2O MEOH] ;B% : 20%- 20%, 9.1 min) . Peak 1 (Rt = 1 .313 min) was collected to give compound 2A (610 mg, 2.35 mmol, 36.91 % yield, 100% purity) as yellow oil.1H NMR: (400 MHz, CDCI3) δ = 7.27 - 7.22 (m, 1 H), 6.93 - 6.79 (m, 3H), 6.14 (s, 1 H), 4.97 (s, 1 H), 3.80 (s, 3H), 3.35 - 3.24 (m, 1 H), 3.14 - 3.03 (m, 1 H), 3.00 - 2.89 (m, 1 H), 2.84 - 2.73 (m, 1 H), 2.36 (s, 3H);

[0323] Peak 2 (Rt = 1.731 min) was collected to give compound 2B (640 mg, 2.44 mmol, 38.34% yield, 99% purity) as yellow oil.1H NMR: (400 MHz, CDCI3) δ = 7.26 - 7.22 (m, 1 H) , 6.93 - 6.80 (m, 3H) , 6.13 (s, 1 H) , 5.00 (s, 1 H) , 3.79 (s, 3H) , 3.33 - 3.23 (m, 1 H) , 3.10 - 3.06 (m, 1 H) , 3.02 - 2.92 (m, 1 H) , 2.84 - 2.74 (m, 1 H) , 2.35 (s, 3H);

[0324] Synthesis of compound 4

[0325] A mixture of tert-butyl 3 -aminopropanoate (302.52 mg, 1.67 mmol, 1 eq, HCI) , compound 3 (250 mg, 1 .67 mmol, 1 eq), DIEA (860.91 mg, 6.66 mmol, 1 .16 mL, 4 eq), HOBt (270.02 mg, 2.00 mmol, 1 .2 eq) and EDCI (383.09 mg, 2.00 mmol, 1 .2 eq) in DMF (2 mL) was stirred at 25 °C for 16 h. The mixture was concentrated at reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, Petroleum ether / Ethyl acetate = 1 : 0 to 2 : 1 , TLC : Petroleum ether / Ethyl acetate = 3 : 1 , RF = 0.25). Compound 4 (430 mg, 1 .55 mmol, 93.11 % yield) was obtained as a colorless oil.1H NMR: (400 MHz, DMSO-d6) δ = 8.00 (br t, J = 5.2 Hz, 1 H), 3.28 - 3.16 (m, 2H), 2.89 - 2.75 (m, 1 H), 2.35 (t, J = 6.8 Hz, 2H), 2.19 (br dd, J = 8.9, 17.9 Hz, 3H), 2.08 - 1 .87 (m, 2H), 1 .83 - 1 .70 (m, 1 H), 1.39 (s, 9H);

[0326] Synthesis of compound 5

[0327] A mixture of compound 4 (430 mg, 1.55 mmol, 1 eq) in DCM (5 mL) and TFA (10 mL) was stirred at 25 °C for 2 h. The mixture was concentrated at reduced pressure to give a residue. Compound 5 (440 mg, 1.31 mmol, 84.65% yield, TFA) was obtained as a yellow solid.1H NMR: (400 MHz, DMSO- d6) δ = 8.01 (br t, J = 5.1 Hz, 1 H), 3.24 (q, J = 6.5 Hz, 2H), 2.84 (quin, J = 8.5 Hz, 1 H), 2.37 (t, J = 6.9 Hz, 2H), 2.28 - 2.07 (m, 3H), 2.05 - 1 .87 (m, 2H), 1 .83 - 1 .71 (m, 1 H);

[0328] Synthesis of compound (3)_peak1

[0329] A mixture of compound 2A (80 mg, 308.44 μmol, 1 eq), compound 5 (103.40 mg, 308.44 μmol, 1 eq, TFA), DIEA (119.59 mg, 925.33 μmol, 161.18 μL, 3 eq), HOBt (50.01 mg, 370.13 μmol, 1.2 eq) and EDCI (70.96 mg, 370.13 μmol, 1.2 eq) in DMF (1 mL) was stirred at 25 °C for 16 h. The mixture was purified by prep-HPLC (column : Phenomenex luna C18 150*25 mm* 10um;mobile phase : [water (FA) -ACN] ;B% : 43%- 73%, 10 min) , the eluent was concentrated and then freeze dried. Compound (3)_peak1 (65.49 mg, 135.92 μmol, 44.07% yield, 96% purity) was obtained as a white solid.1H NMR: (400 MHz, CDCI3) δ = 7.23 (dt, J = 1 .7, 8.1 Hz, 1 H), 6.91 - 6.80 (m, 3H), 6.77 - 6.69 (m, 1 H), 6.53 - 6.44 (m, 1 H), 6.42 - 6.35 (m, 1 H), 3.85 - 3.72 (m, 4H), 3.68 - 3.52 (m, 2H), 3.42 - 3.30 (m, 1 H), 2.97 - 2.86 (m, 1 H), 2.85 - 2.48 (m, 4H), 2.42 (s, 3H), 2.38 - 2.12 (m, 3H), 2.10 - 1.88 (m, 3H); LCMS: RT =0.754 min, m / z =463.3 (M+H)+.

[0330] Synthesis of compound (3)_peak2

[0331] A mixture of compound 2B (80 mg, 308.44 μmol, 1 eq), compound 5 (103.40 mg, 308.44 μmol, 1 .00 eq, TFA), DIEA (119.59 mg, 925.33 μmol, 161 .18 μL, 3 eq), HOBt (50.01 mg, 370.13 μmol, 1 .2 eq) and EDCI (70.96 mg, 370.13 μmol, 1.2 eq) in DMF (1 mL) was stirred at 25 °C for 16 h. The mixture was purified by prep-HPLC (column : Phenomenex luna C18 150*25 mm* 10um;mobile phase : [water (FA) -ACN] ;B% : 43%- 73%, 10 min), the eluent was concentrated and then freeze dried. Compound (3)_peak2 (65.49 mg, 55.09 μmol, 17.86% yield, 98% purity) was obtained as a white solid.1H NMR: (400 MHz, CDCI3) δ = 7.26 - 7.19 (m, 1 H), 6.89 - 6.79 (m, 3H), 6.77 - 6.69 (m, 1 H), 6.53 - 6.45 (m, 1 H), 6.42 - 6.34 (m, 1 H), 3.84 - 3.74 (m, 4H), 3.67 - 3.51 (m, 2H), 3.42 - 3.28 (m, 1 H), 3.08 - 2.85 (m, 1 H), 2.84 - 2.47 (m, 4H), 2.42 (s, 3H), 2.39 - 2.14 (m, 3H), 2.13 - 1.88 (m, 3H); LCMS: RT =0.759 min, m / z =463.2 (M+H)+.

[0332] EXAMPLE 1.4: SYNTHESIS OF (4)

[0333] Synthesis of compound 3

[0334] To a mixture of compound 1 (300 mg, 1.83 mmol, 1 eq) and compound 2 (265.37 mg, 1.83 mmol, 1 eq) in DMF (2 mL) was added EDCI (420.43 mg, 2.19 mmol, 1.2 eq), HOBt (296.34 mg, 2.19 mmol, 1 .2 eq) and DIEA (708.61 mg, 5.48 mmol, 955.01 μL, 3 eq). The mixture was stirred at 60 °C for 24 h. The reaction mixture was quenched by addition H2O (30 mL), and the resulting mixture was extracted with EtOAc (10 mL * 3). The combined organic layers were washed with brine (10 mL), dried over Na2SO4, filtered and concentrated under reduced pressure to give compound 3 (650 mg, crude) as a yellow solid.

[0335] Synthesis of compound 4

[0336] To a solution of compound 3 (650 mg, 2.23 mmol, 1 eq) in DCM (5 mL) was added TFA (254.39 mg, 2.23 mmol, 165.19 μL, 1 eq). The mixture was stirred at 25 °C for 16 h. The mixture was concentrated in vacuum to give compound 4 (600 mg, crude) as a yellow solid.

[0337] Synthesis of compound (4)

[0338] To a solution of compound 4 (90.69 mg, 385.56 μmol, 2 eq) and compound 5 (50 mg, 192.78 μmol, 1 eq) in DMF (1 mL) was added HOBt (31 .26 mg, 231 .33 μmol, 1 .2 eq), EDCI (44.35 mg, 231 .33 μmol, 1.2 eq) and DIEA (74.75 mg, 578.33 μmol, 100.74 μL, 3 eq). The mixture was stirred at 25 °C for 8 h. The reaction mixture was filtered and the filtrate was purified by prep-HPLC (column: Phenomenex Luna C18 150*25 mm* 10um;mobile phase: [water (FA) -ACN];B%: 46%-76%, 10 min), followed by lyophilization. Compound (4) (30 mg, 61.06 μmol, 31 .67% yield, 97% purity) was obtained as a white solid.1H NMR (400 MHz, CDCI3) δ = 7.25 - 7.20 (m, 1 H), 6.89 - 6.80 (m, 3H), 6.72 (s, 1 H), 6.58 - 6.45 (m, 1 H), 6.37 (s, 1 H), 3.84 - 3.75 (m, 4H), 3.64 - 3.55 (m, 2H), 3.42 - 3.30 (m, 1 H), 2.97 - 2.53 (m, 4H), 2.42 (s, 3H), 2.19 - 2.05 (m, 3H), 1.91 - 1 .73 (m, 6H); LCMS: RT = 2.504 min, m / z = 477.3 (M+H)+.

[0339] EXAMPLE 1.5: SYNTHESIS OF (5)

[0340] Synthesis of compound 2

[0341] To a solution of compound 1 (55 mg, 123.71 μmol, 1 eq) in DCM (1 mL) was added TFA (141.06 mg, 1.24 mmol, 91.60 μL, 10 eq). The mixture was stirred at 25 °C for 2 h. The reaction mixture was concentrated under reduced pressure to give compound 2 (60 mg, crude) as red oil. LCMS: RT = 0.441 min, m / z = 345.1 (M+H)+.

[0342] Synthesis of (5)

[0343] To a solution of compound 2 (60 mg, 174.18 μmol, 1 eq) and 3, 3-difluorocyclobutanecarboxylic acid (23.71 mg, 174.18 μmol, 1 eq) in DMF (1 mL) was added DIEA (135.07 mg, 1 .05 mmol, 182.03 μL, 6 eq), HOBt (28.24 mg, 209.02 μmol, 1 .2 eq) and EDCI (40.07 mg, 209.02 μmol, 1.2 eq). The mixture was stirred at 25 °C for 16 h. The reaction mixture was concentrated under reduced pressure to give residue. The residue was purified by prep-HPLC (column: YMC-Actus Triart C18 150*30 mm*7um; mobile phase: [water (FA) -ACN]; B%: 55%-5%, 10 min) to give (5) (30.88 mg, 64.76 μmol, 37.18% yield, 97% purity) as white solid.1H NMR: (400 MHz, DMSO-d6) δ = 8.19 - 8.12 (m, 1 H), 7.26 - 7.20 (m, 1 H), 6.91 - 6.83 (m, 1 H), 6.76 - 6.68 (m, 2H), 6.57 (d, J = 11.6 Hz, 1 H), 6.50 (d, J = 5.4 Hz, 1 H), 4.16 - 3.99 (m, 1 H), 3.74 - 3.67 (m, 3H), 3.27 - 2.93 (m, 4H), 2.90 - 2.71 (m, 3H), 2.65 - 2.53 (m, 4H), 2.39 - 2.35 (m, 3H), 1 .03 - 0.88 (m, 3H); LCMS: RT = 0.578 min, m / z = 463.3 (M+H)+; SFC: RT = 1 .669 min, RT = 3.824 min.

[0344] EXAMPLE 1.6: SYNTHESIS OF (6)

[0345] A mixture of compound 6A (50 mg, 174.59 μmol, 1 eq) , 1 -phenylcyclobutanecarboxylic acid (30.76 mg, 174.59 μmol, 1 eq), HOBt (28.31 mg, 209.50 μmol, 1 .2 eq), EDCI (40.16 mg, 209.50 μmol, 1 .2 eq) and DIEA (45.13 mg, 349.17 μmol, 60.82 μL, 2 eq) in DMF (1 mL) was stirred at 20 °C for 16 h. The reaction mixture was concentrated under vacuum to give a residue. The residue was purified by prep- HPLC (column: Unisil 3 - 100 C18 Ultra 150*50 mm*3 um;mobile phase : [water (FA) -ACN] ;B% : 54%- 84%, 7 min) to give (6) (19.66 mg, 43.78 μmol, 25.08% yield, 99% purity) as yellow gum.1H NMR: (400 MHz, CDCI3) δ = 7.42 - 7.30 (m, 4H), 7.27 (s, 5H), 7.16 (tdd, J = 2.7, 5.7, 8.7 Hz, 1 H), 6.82 - 6.76 (m, 2H), 6.24 - 6.12 (m, 1 H), 3.78 (br dd, J = 4.0, 14.1 Hz, 1 H), 3.61 (br s, 2H), 3.30 (ddd, J = 4.9, 11 .4, 14.2 Hz, 1 H), 2.95 - 2.67 (m, 4H), 2.58 - 2.41 (m, 4H), 2.15 - 2.05 (m, 1 H), 1.91 - 1 .82 (m, 1 H); LCMS: RT = 0.643 min, m / z = 445.3 (M+H)+.

[0346] EXAMPLE 1.7: SYNTHESIS OF (7)

[0347] Petroleum ether = 1 : 0).

[0348] (7)_peak1 (24.51 mg, 53.71 μmol, 15.46% yield, 95% purity) was obtained as a white solid.1H NMR: (400 MHz, CDCI3) δ = 8.78 - 8.70 (m, 1 H), 7.25 - 7.17 (m, 1 H), 7.14 - 6.96 (m, 4H), 6.30 (br s, 1 H), 4.93 - 3.92 (m, 1 H), 3.60 - 3.37 (m, 3H), 3.17 - 2.93 (m, 3H), 2.88 - 2.79 (m, 1 H), 2.75 - 2.57 (m, 4H), 2.36 - 2.29 (m, 3H), 1 .17 - 1 .07 (m, 3H); LCMS: RT =0.505 min, m / z = 434.1 (M+H)+. SFC: RT = 0.977 min.

[0349] (7)_peak2 (28.93 mg, 62.73 μmol, 18.06% yield, 94% purity) was obtained as a white solid.1H NMR: (400 MHz, CDCI3) δ = 8.72 (s, 1 H), 7.22 - 7.17 (m, 1 H), 7.15 - 6.98 (m, 4H), 6.19 (br s, 1 H), 4.99 - 4.03 (m, 1 H), 3.56 - 3.33 (m, 3H), 3.19 - 2.99 (m, 3H), 2.83 - 2.72 (m, 1 H), 2.68 - 2.41 (m, 4H), 2.35 - 2.29

[0350] (m, 3H), 1 .21 - 1 .10 (m, 3H); LCMS: RT = 0.503min, m / z = 434(M+H)+. SFC: RT = 1 .172 min.

[0351] EXAMPLE 1.8: SYNTHESIS OF (8)

[0352] Synthesis of compound 4

[0353] To a solution of compound 1 (380 mg, 3.16 mmol, 1 eq) in THF (4 mL) was added compound 2 (358.76 mg, 3.80 mmol, 293.35 μL, 1.2 eq) at 0 °C. The mixture was stirred at 20 °C for 30 min. The iminium salt solution was cooled to 0 °C, and the solution of compound 3 (1 M, 6.33 mL, 2 eq) was added. The reaction mixture was allowed to warm to 20 °C and stirred for 14 h. The reaction mixture was poured into saturated NH4CI aqueous solution (10 mL) and the resulting mixture was extracted with EtOAc (15 mL*3). The combined organic phase was washed with brine (15 mL), dried over Na2SO4, filtered and concentrated at reduced pressure to give a residue. The residue was purified by flash silica gel chromatography (ISCO®; 12 g SepaFlash® Silica Flash Column, Eluent of 0-20% Ethyl acetate / Petroleum ether gradient @ 50 mL / min). Compound 4 (450 mg, 1 .66 mmol, 52.62% yield) was obtained as a yellow oil.

[0354] Synthesis of compound 5

[0355] To a solution of compound 4 (450 mg, 1 .66 mmol, 1 eq) in EtOH (15 mL) was added Pd / C (200 mg, 187.93 μmol, 166.49 μL, 10% purity, 1.13e- 1 eq) under N2. The suspension was degassed under vacuum and purged with H2 several times. The mixture was stirred under H2 (30 psi) at 30 °C for 3 h. The mixture was filtered and the filtrate was concentrated under vacuum to give a residue. The residue was purified by prep-TLC (SiO2, Petroleum ether: MeOH = 3: 1). Compound 5 (310 mg, 1.10 mmol, 66.33% yield, 97% purity) was obtained as a yellow oil. LCMS: RT =0.507 min, m / z =273.1 (M+H)+.

[0356] Synthesis of compound 6

[0357] To a solution of compound 5 (310 mg, 1 .14 mmol, 1 eq) in EtOH (15 mL) and H2O (3 mL) was added KOH (638.79 mg, 11.38 mmol, 10 eq) at 25 °C. The mixture was stirred at 90 °C for 14 h. The mixture was filtered and the filtrate was concentrated under vacuum to give a residue. The residue was purified by flash silica gel chromatography (ISCO®; 12 g SepaFlash® Silica Flash Column, Eluent of 60-100% Ethyl acetate / Petroleum ether gradient @ 35 mL / min). Compound 6 (80 mg, 291.23 μmol, 25.58% yield, 78% purity) was obtained as a yellow oil.

[0358] Synthesis of compound (8)_peak1 & (8)_peak2

[0359] To a solution compound 7 (20.13 mg, 91 .01 μmol, 1 .3 eq) in Py (1 mL) was added EDCI (20.13 mg, 105.01 μmol, 1 .5 eq) and compound 6 (15 mg, 70.01 μmol, 1 eq). The mixture was stirred at 25 °C for 2 h. The mixture was diluted with H2O (20 mL) and the resulting mixture was extracted with Ethyl acetate (20 mL * 3). The combined organic layers were dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue (Batch 1).

[0360] To the mixture of compound 7 (82.59 mg, 373.37 μmol, 2 eq) in Py (1 mL) was added EDCI (53.68 mg, 280.03 μmol, 1.5 eq) and compound 6 (40 mg, 186.69 μmol, 1 eq). The mixture was stirred at 25 °C for 16 h. The mixture was diluted with H2O (20 mL) and the resulting mixture was extracted with Ethyl acetate (20 mL * 3). The combined organic layers were dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue (Batch 2). Two batches were combined and purified by prep- HPLC (column: Waters Xbridge 150*25 mm* 5um; mobile phase: [water (NH4HCO3) -ACN]; gradient: 32%- 62% B over 9 min) to give (8) (30 mg, purity 92%) as a white solid, which was further separated by SFC ( (RT = 1 .041 min and 1 .284 min, column: DAICEL CHIRALPAK AS (250 mm*30 mm, 10um); mobile phase: [CO2 -i-PrOH (0.1 %NH3H2O)]; B%: 25%, isocratic elution mode). Compound (8)_peak1 (9.48 mg, 22.26 μmol, 9.54% yield, 98% purity) was obtained as a yellow solid. Compound (8)_peak2 (10.56 mg, 24.03 μmol, 10.30% yield, 95% purity) was obtained as a yellow solid.

[0361] (8)_peak1 :1H NMR: (400 MHz, CDCI3) δ = 7.90 - 7.84 (m, 1 H), 7.25 - 7.18 (m, 1 H), 7.18 - 7.03 (m, 3H), 6.80-5.92 (m, 2H), 4.99-3.96 (m, 1 H), 3.60 - 3.50 (m, 1 H), 3.49 - 3.30 (m, 2H), 3.19 - 3.08 (m, 1 H), 3.01 - 2.56 (m, 7H), 2.39 - 2.28 (m, 3H), 1.21 - 1.05 (m, 3H); LCMS: RT = 0.570 min, m / z = 418.2 (M+H)+; SFC: RT = 1.023 min.

[0362] (8)_peak2: 1 H NMR: (400 MHz, CDCI3) δ = 7.86 (s, 1 H), 7.26 - 7.18 (m, 1 H), 7.15 - 7.04 (m, 3H), 6.81 - 6.06 (m, 2H), 5.07 - 4.09 (m, 1 H), 3.61 - 3.46 (m, 1 H), 3.46 - 3.30 (m, 2H), 3.28 - 3.11 (m, 1 H), 2.97 - 2.44 (m, 7H), 2.40 - 2.24 (m, 3H), 1.24 - 1.06 (m, 3H); LCMS: RT = 0.566 min, m / z = 418.2 (M+H)+. SFC: RT = 1.280 min.

[0363] EXAMPLE 1.9: SYNTHESIS OF (9)

[0364] Synthesis of compound 2

[0365] A mixture of compound 1 (400 mg, 1.05 mmol, 1 eq) in EtOH (5 mL) was added KOH (118.23 mg, 2.11 mmol, 2 eq), the mixture was stirred at 80 °C for 4 h. The reaction mixture was concentrated in vacuum to give a residue. The residue was dissolved with H2O (10 mL) and extracted with ethyl acetate (5 mL*3), the combined organic phase was discarded, the aqueous phase was acidified to pH = 2 - 3 with 1 N HCI, the resulting mixture was extracted with ethyl acetate (5 mL*3), the combined organic phase was collected, dried with anhydrous Na2SO4, filtered and concentrated in vacuum to give compound 2 (240 mg, 858.63 μmol, 81 .49% yield) as colorless oil. LCMS: RT = 0.350 min, m / z = 278.9 (M+H)+.

[0366] Synthesis of compound 3

[0367] To a mixture of compound 2 (240 mg, 858.63 μmol, 1 eq) in MeOH (5 mL) was added SOCI2 (204.30 mg, 1 .72 mmol, 124.73 μL, 2 eq) dropwise at 0 °C under N2, the mixture was stirred at 25 °C for 16 h. The reaction mixture was concentrated in vacuum to give a residue. The residue was dissolved with ethyl acetate (20 mL) and washed with saturated NaHCOs solution (8 mL*3), the organic phase was dried with anhydrous Na2SO4, filtered and concentrated in vacuum to give compound 3 (250 mg, 851 .67 μmol, 99.19% yield) as colorless oil.1H NMR: (400 MHz, CDCI3) δ = 7.47 (d, J = 2.4 Hz, 1 H), 7.30 (d, J = 2.4 Hz, 1 H), 3.85 (s, 3H), 3.73 (s, 3H), 3.66 (s, 2H).

[0368] Synthesis of compound 5

[0369] A mixture of compound 3 (250 mg, 851 .67 μmol, 1 eq), compound 4 (174.75 mg, 1 .02 mmol, 1 .2 eq), lr[dF(CF3) ppy]2(dtbpy)(PFe) (9.55 mg, 8.52 μmol, 0.01 eq), NiCL’dtbbpy (1.69 mg, 4.26 μmol, 0.005 eq), TTMSS (211.78 mg, 851.67 μmol, 262.75 μL, 1 eq) and Na2CO3(180.54 mg, 1 .70 mmol, 2 eq) in DME (2 mL) was degassed and purged with N2 and the mixture was stirred at 25 °C for 16 h with a 455 nm blue LED. The reaction mixture was diluted with ethyl acetate (30 mL) and washed with H2O (10 mL*3), the organic phase was dried with anhydrous Na2SO4, filtered and concentrated in vacuum to give compound 5 (370 mg, crude) as orange oil. LCMS: RT = 0.565 min, m / z = 305.0 (M+H)+.

[0370] Synthesis of compound 6

[0371] To a mixture of compound 5 (150 mg, 492.26 μmol, 1 eq) in THF (1 mL) and H2O (0.3 mL) was added LiOH’FW (41.31 mg, 984.52 μmol, 2 eq), the mixture was stirred at 25 °C for 1 h. The reaction mixture was concentrated in vacuum to remove THF, then the mixture was acidified to pH = 3 - 4 with 1 N HCI and the resulting mixture was extracted with ethyl acetate (5 mL*3), the combined organic phase was dried with anhydrous Na2SO4, filtered and concentrated in vacuum to give a residue. The residue was purified by prep-HPLC (column: Waters Xbridge 150*25 mm* 5um; mobile phase: [water (NH4HCO3) -ACN]; gradient: 10%- 40% B over 9 min), the eluent was concentrated in vacuum to remove MeCN. The aqueous solution was acidified to pH = 2 - 3 with 1 N HCI and the resulting aqueous solution was extracted with ethyl acetate (10 mL*3), the organic phase was collected, dried with anhydrous Na2SO4, filtered and concentrated in vacuum to give a compound 6 (46 mg, 158.24 μmol, 32.15% yield) as white solid. LCMS: RT = 0.407 min, m / z = 289.1 (M-H)-.

[0372] Synthesis of compound 8

[0373] To a mixture of compound 6 (46 mg, 158.24 μmol, 1 eq) and compound 7 (34.86 mg, 158.24 μmol, 1 eq) in pyridine (1 mL) was added EDCI (45.50 mg, 237.37 μmol, 1 .5 eq), the mixture was stirred at 25 °C for 2 h. The reaction mixture was diluted with ethyl acetate (15 mL) and washed with H2O (5 mL*), the organic phase was collected, dried with anhydrous Na2SO4, filtered and concentrated in vacuum to give a residue. The residue was purified by column chromatography (SiO2, Petroleum ether / Ethyl acetate = 1 / 0 to 0 / 1 , TLC (SiO2, Petroleum ether: Ethyl acetate = 0: 1 , Rf = 0.24)) to give compound 8 (55 mg, 111 .57 μmol, 70.50% yield) as colorless gum. LCMS: RT = 0.536 min, m / z = 493.1 (M+H)+.

[0374] Synthesis of (9)

[0375] To a solution of compound 8 (10 mg, 20.29 μmol, 1 eq) in DCM (1 mL) was added four drops of the solution of BBrs (2 M, 20.29 μL, 2 eq) in DCM (0.02 mL) dropwise at - 30 °C under N2, the mixture was stirred at 25 °C for 1 h. LCMS showed 58% of compound 8 remained and -35% of peak with desired mass. Then four drops of the solution of BBrs (2 M, 20.29 μL, 2 eq) in DCM (0.02 mL) was added to the reaction mixture dropwise at -30 °C under N2, the mixture was stirred at 25 °C for 1 h. The reaction mixture was diluted with ethyl acetate (5 mL) and the resulting mixture was added into MeOH (10 mL) slowly, the mixture was concentrated in vacuum to give a residue. The residue was purified by prep- HPLC (column: Waters Xbridge 150*25 mm* 5um; mobile phase: [water (NH4HCO3) -ACN]; gradient: 36%- 66% B over 9 min) to give an aqueous solution.

[0376] To a solution of compound 8 (30 mg, 60.86 μmol, 1 eq) in DCM (3 mL) was added BBrs (2 M, 60.86 μL, 2 eq) diluted in DCM (0.06 mL) dropwise at - 30 °C under N2, the mixture was stirred at 25 °C for 2 h. The reaction mixture was diluted with ethyl acetate (5 mL) and then added into MeOH (10 mL) slowly, the resulting mixture was concentrated in vacuum to give a residue. The residue was purified by prep- HPLC (column: Waters Xbridge 150*25 mm* 5um; mobile phase: [water (NH4HCO3) -ACN]; gradient: 36%- 66% B over 9 min) to give an aqueous solution. Two batches of the above aqueous solution were combined, dried by lyophilization to give (9) (3.75 mg, 7.83 μmol, 9.90% yield, 100% purity) as off-white solid.1H NMR: (400 MHz, CDCI3) δ = 9.77 (br s, 1 H), 8.75 - 8.64 (m, 1 H), 7.31 (d, J = 2.1 Hz, 1 H), 7.13 (d, J = 2.1 Hz, 1 H), 6.94 - 6.44 (m, 1 H), 6.89 - 6.34 (d, J = 2.0 Hz, 1 H), 6.88 - 6.04 (d, J = 2.3 Hz, 1 H), 5.03 - 4.24 (m, 1 H), 4.20 - 3.85 (m, 2H), 3.92 - 3.83 (m, 3H), 3.35 - 3.20 (m, 1 H), 3.15 - 3.03 (m, 1 H), 3.03 - 2.84 (m, 4H), 2.68 - 2.48 (m, 2H); LCMS: RT = 0.463 min, m / z = 479.1 (M+H)+.

[0377] EXAMPLE 1.10: SYNTHESIS OF (10)

[0378] To a solution of 4, 4-difluorocyclohexanecarboxylic acid (28.66 mg, 174.59 μmol, 1 eq) in DMF (1 mL) was added HOBt (23.59 mg, 174.59 μmol, 1 eq) and DIEA (67.69 mg, 523.76 μmol, 91.23 μL, 3 eq), compound 6A (50.00 mg, 174.59 μmol, 1 eq), EDCI (33.47 mg, 174.59 μmol, 1 eq). The mixture was stirred at 25 °C for 16 h. The mixture was purified by prep-HPLC(column: Unisil 3 - 100 C18 Ultra 150*50 mm*3 um; mobile phase: [water (FA) -ACN]; B%: 45%- 75%, 7 min), the eluent was concentrated and then freeze dried. (10) (13.60 mg, 31 .13 μmol, 17.83% yield, 99% purity) was obtained as an off- white solid.1H NMR: (400 MHz, DMSO-d6) δ = 7.86 - 7.79 (m, 1 H), 7.38 (d, J = 5.3 Hz, 1 H), 7.32 - 7.24 (m, 3H), 7.17 (br d, J = 6.8 Hz, 2H), 6.81 (d, J = 5.3 Hz, 1 H), 6.70 (s, 1 H), 4.01 - 3.92 (m, 1 H), 3.26 - 3.08 (m, 2H), 3.00 - 2.76 (m, 3H), 2.64 - 2.52 (m, 3H), 2.23 - 2.14 (m, 1 H), 2.03 - 1.91 (m, 2H), 1 .77 - 1 .64 (m, 3H), 1 .59 - 1 .47 (m, 2H); LCMS: RT = 0.740 min, m / z = 433.1 (M+H)+. SFC: RT = 1 .541 min.

[0379] EXAMPLE 1.11: SYNTHESIS OF (11) Synthesis of compound 2

[0380] To a solution of compound 1 (2 g, 6.44 mmol, 1 eq) in DMF (20 mL) was added K2CO3 (1.78 g, 12.88 mmol, 2 eq), bromomethylbenzene (1.65 g, 9.66 mmol, 1 .15 mL, 1.5 eq) was added and the mixture was stirred at 20 °C for 3 h. The reaction mixture was diluted with H2O (50 mL), and then the resulting mixture was extracted with Ethyl acetate (50 mL * 3). The combined organic layers were washed with brine (50 mL * 3), dried over Na2SO4, filtered and concentrated under reduced pressure to give residue. The residue was purified by column chromatography (SiO2, Petroleum ether / Ethyl acetate = 1 / 0 to 10 / 1 , TLC, Petroleum ether / Ethyl acetate = 10 / 1 , Rf = 0.7), the eluent was concentrated under reduced pressure to compound 2 (2.5 g, 6.24 mmol, 96.88% yield) as brown oil.1H NMR: (400 MHz, CD3SOCD3) δ = 7.77 (d, J = 2.1 Hz, 1 H), 7.58 (d, J = 7.1 Hz, 2H), 7.53 (d, J = 2.3 Hz, 1 H), 7.47 - 7.40 (m, 3H), 4.92 (s, 2H), 1.27 (s, 9H).

[0381] Synthesis of compound 3

[0382] To a solution of compound 2 (500 mg, 1.25 mmol, 1 eq) in DMSO (5 mL) was added K2CO3 (1.29 g, 9.36 mmol, 7.5 eq) and Cui (47.53 mg, 249.58 μmol, 0.2 eq) under N2. Then ethyl 3-oxobutanoate (812.00 mg, 6.24 mmol, 789.89 μL, 5 eq) was added under N2 and the mixture was stirred at 100 °C for 16 h. The reaction mixture was diluted with H2O (30 mL), and the resulting mixture was extracted with Ethyl acetate (30 mL *3). The combined organic layers were washed with brine (30 mL *2), dried over Na2SO4, filtered and concentrated under reduced pressure to give residue. The residue was purified by column chromatography (SiO2, Petroleum ether / Ethyl acetate = 10 / 1 to 5 / 1 , TLC, Petroleum ether / Ethyl acetate = 10:1 , Rf = 0.6), the eluent was concentrated under reduced pressure to give compound 3 (50 mg, crude) as colorless gum.

[0383] Synthesis of compound 4

[0384] To a solution of compound 3 (50 mg, 138.55 μmol, 1 eq) in THF (2 mL) and H2O (1 mL) was added LiOH»H2O (29.07 mg, 692.76 μmol, 5 eq). The mixture was stirred at 20 °C for 2 h. Then NaOH (11 .08 mg, 277.11 μmol, 2 eq) was added and the mixture was stirred at 20 °C for 2 h. The mixture was diluted with H2O (10 mL), the pH of mixture was adjusted to 5-6 by adding HCI (1 M) aqueous solution. Then the resulting mixture was extracted with Ethyl acetate (10 mL *3). The combined organic layers were washed with brine (10 mL *2), dried over Na2SO4, filtered and concentrated under reduced pressure to give compound 4 (60 mg, crude) as colorless gum. LCMS: RT = 0.403 min, m / z = 331.0 (M+H)+.

[0385] Synthesis of compound 6

[0386] To a solution of compound 5 (29.25 mg, 135.21 μmol, 1 eq) in DMF (1 mL) was added DIEA (34.95 mg, 270.42 μmol, 47.10 μL, 2 eq), HOBt (21.92 mg, 162.25 μmol, 1.2 eq), EDCI (31.10 mg, 162.25 μmol, 1 .2 eq) and compound 4 (45 mg, 135.21 μmol, 1 eq). The mixture was stirred at 20 °C for 16 h. The mixture was diluted with H2O (10 mL), then the resulting mixture was extracted with Ethyl acetate (10 mL *3). The combined organic layers were washed with brine (10 mL *2), dried over Na2SO4, filtered and concentrated under reduced pressure to give residue. The residue was purified by column chromatography (SiO2, Petroleum ether / Ethyl acetate = 3 / 1 to 1 / 1 , TLC, Petroleum ether / Ethyl acetate = 1 : 1 , Rf = 0.8), the eluent was concentrated under reduced pressure to give compound 6 (40 mg, crude) as white solid. LCMS: RT = 0.718 min, m / z = 531 .3 (M+H)+.

[0387] Synthesis of (11)

[0388] To a solution of compound 6 (0.04 g, 75.31 μmol, 1 eq) in MeOH (1 mL) was added Pd / C (16.03 mg, 15.06 μmol, 10% purity, 0.2 eq) under N2. The suspension was degassed under vacuum and purged with H2 (20 psi) 3 times. The mixture was stirred under H2 (20 psi) at 25 °C for 16 h. Compound 6 was remained. The reaction mixture was filtered. Then Pd / C (16.03 mg, 15.06 μmol, 10% purity, 0.2 eq) was added into the mixture under N2, The suspension was degassed under vacuum and purged with H2 (20 psi) 3 times. The mixture was stirred under H2 (20 psi) at 25 °C for another 16 h. The reaction mixture was filtered and concentrated in vacuum to give residue. The residue was purified by prep-HPLC (column: YMC-Actus Triart C18 150*30 mm*7um; mobile phase: [water (FA) -ACN]; gradient: 58%-88% B over 10 min) to give (11) (1.06 mg, 2.38 μmol, 3.50% yield, 99% purity) as off-white solid.1H NMR: (400 MHz, CDCI3) δ = 9.12 (s, 1 H), 8.73 (d, J = 7.7 Hz, 1 H), 8.22 (s, 1 H), 7.39 - 7.28 (m, 4H), 7.26 - 7.22 (m, 1 H), 7.11-6.48 (m, 2H), 4.98 - 4.10 (m, 1 H), 3.94 (d, J = 7.1 Hz, 1 H), 3.86 (s, 1 H), 3.54 - 2.83 (m, 3H), 1 .25 - 1 .17 (m, 9H); LCMS: RT = 0.639 min, m / z = 441 .3 (M+H)+.

[0389] EXAMPLE 1.12: SYNTHESIS OF (12)

[0390] Synthesis of compound 2

[0391] To a solution of compound 1 (4 g, 21 .66 mmol, 1 eq) in ACN (40 mL) was added CS2CO3 (14.12 g, 43.32 mmol, 2 eq) and CH3I (6.15 g, 43.32 mmol, 2.70 mL, 2 eq). The mixture was stirred at 80 °C for 2 h. The mixture was diluted with H2O (50 mL) and the resulting mixture was extracted with Ethyl acetate (50 mL *3). The combined organic layers were washed with brine (50 mL *2), dried over Na2SO4, filtered and concentrated under reduced pressure to give compound 2 (4.28 g, crude) as colorless oil.1H NMR: (400 MHz, CDCI3) δ = 7.39 (d, J = 2.3 Hz, 1 H), 7.23 (dd, J = 2.3, 8.6 Hz, 1 H), 6.87 (d, J = 8.6 Hz, 1 H), 3.89 (s, 3H), 1.30 (s, 9H).

[0392] Synthesis of compound 3 To a solution of compound 2 (2 g, 10.07 mmol, 1 eq) in ACN (15 mL) was added TsOH»H2O (1.91 g, 10.07 mmol, 1 eq) at 0 °C, the mixture was stirred at 0 °C for 0.5 h. NIS (2.49 g, 11 .07 mmol, 1 .1 eq) was added and the mixture was stirred at 20 °C for 16 h. The reaction mixture was quenched by adding saturated Na2S20s solution (20 mL) and then the resulting mixture was extracted with EtOAc (50 mL * 3). The combined organic layers were washed with brine (50 mL * 3), dried over Na2SO4, filtered and concentrated under reduced pressure to give residue. The residue was purified by column chromatography (SiO2, Petroleum ether / Ethyl acetate = 1 / 0 to 10 / 1 , TLC, Petroleum ether / Ethyl acetate = 10 / 1 , Rf = 0.8), the eluent was concentrated under reduced pressure to give compound 3 (2.44 g, 7.50 mmol, 74.53% yield) as colorless oil.1H NMR: (400 MHz, CDCI3) δ = 7.65 (d, J = 2.3 Hz, 1 H), 7.35 (d, J = 2.2 Hz, 1 H), 3.86 (s, 3H), 1.29 (s, 9H).

[0393] Synthesis of compound 4

[0394] To a solution of compound 3 (7.02 g, 53.92 mmol, 6.83 mL, 5 eq) in DMSO (30 mL) was added Cui (410.72 mg, 2.16 mmol, 0.2 eq) and K2CO3 (11.18 g, 80.87 mmol, 7.5 eq) under N2. Ethyl 3- oxobutanoate (7.02 g, 53.92 mmol, 6.83 mL, 5 eq) was added and the mixture was stirred at 100 °C for 16 h. The mixture was diluted with H2O (50 mL) and the resulting mixture was extracted with Ethyl acetate (50 mL *3). The combined organic layers were washed with brine (50 mL *2), dried over Na2SO4, filtered and concentrated under reduced pressure to give residue. The residue was purified by column chromatography (SiO2, Petroleum ether / Ethyl acetate = 1 / 0 to 10 / 1 , TLC, Petroleum ether / Ethyl acetate = 10 / 1 , Rf = 0.6), the eluent was concentrated under reduced pressure to give compound 4 (810 mg, crude) as colorless oil.1H NMR: (400 MHz, CDCI3) δ = 7.38 (d, J = 2.3 Hz, 1 H), 7.15 (d, J = 2.3 Hz, 1 H), 4.19 (q, J = 7.1 Hz, 2H), 3.85 (s, 3H), 3.67 (s, 2H), 1 .30 (s, 9H), 1 .29 - 1 .25 (m, 3H).

[0395] Synthesis of compound 5

[0396] To a solution of compound 4 (810 mg, 2.84 mmol, 1 eq) in DCM (10 mL) was added BBrs (1.07 g, 4.27 mmol, 411.10 μL, 1 .5 eq) dropwise at 0 °C under N2. The mixture was stirred at 20 °C for 1 h. The reaction mixture was quenched by MeOH (10 mL), and then the pH of the resulting mixture of was adjusted to 7-8 by adding saturated NaHCOs aqueous solution, the resulting mixture was extracted with EtOAc (30 mL * 3). The combined organic layers were washed with brine (30 mL * 2), dried over Na2SO4, filtered and concentrated under reduced pressure to give residue. The residue was purified by prep- HPLC (column: Waters Xbridge Prep OBD C18 150*40 mm*10um; mobile phase: [water (NH4HCO3) - ACN]; B%: 43%, isocratic elution mode), the eluent was concentrated and then freeze dried to give compound 5 (200 mg, 738.69 μmol, 25.97% yield) as yellow gum. LCMS: RT = 0.650 min, m / z = 269.1 (M-H)+.

[0397] Synthesis of (12)

[0398] To a solution of compound 6 (30 mg, 136.18 μmol, 1 eq) and compound 5 (36.87 mg, 136.18 μmol, 1 eq) in THF (1 mL) was added AIMes (2 M, 102.14 μL, 1 .5 eq) under N2. The mixture was stirred at 60 °C for 4 h. AIMes (9.82 mg, 136.18 μmol, 1 eq) was added, the mixture was stirred at 60 °C for 3 h. The mixture was quenched by adding Na2SO4*10H2O (40 mg, 124.15 μmol, 28.57 μL, 2.46e-1 eq), and then the resulting mixture was diluted with H2O (10 mL) and the mixture was extracted with EtOAc (10 mL *3). The combined organic layers were washed with brine (10 mL *2), dried over Na2SO4, filtered and concentrated under reduced pressure to give residue. The residue was purified by prep-HPLC (column: YMC-Actus Triart C18 150*30 mm*7um; mobile phase: [water (FA) -ACN]; gradient: 348%-78% B over 10 min), the eluent was concentrated and then freeze dried to give (12) (6.57 mg, 14.47 μmol, 10.63% yield, 98% purity) as white solid.1H NMR: (400 MHz, CDCI3) δ = 9.54 (br s, 1 H), 8.73 - 8.65 (m, 1 H), 7.30 - 7.27 (m, 1 H), 7.26 - 7.24 (m, 1 H), 7.02 - 6.98 (m, 1 H), 6.95 (s, 1 H), 6.46 (s, 1 H), 6.35 (d, J = 2.1 Hz, 1 H), 6.05 (d, J = 2.1 Hz, 1 H), 4.97 (dd, J = 4.2, 12.8 Hz, 1 H), 4.29 - 4.21 (m, 1 H), 4.17 - 3.97 (m, 1 H), 3.90 (s, 2H), 3.86 (s, 1 H), 3.82 (s, 1 H), 3.17 - 2.82 (m, 3H), 1 .26 - 1 .22 (m, 9H); LCMS: RT = 0.560 min, m / z = 445.3 (M+H)+; SFC: RT = 1 .797 min.

[0399] Synthesis of compound 2A, 2B

[0400] A mixture of 4 -fluorobenzaldehyde (175.75 mg, 1 .42 mmol, 148.94 μL, 1 eq) and compound 1 (200 mg, 1 .42 mmol, 1 eq) in toluene (2 mL) was stirred at 110 °C for 16 h. The mixture was concentrated at reduced pressure to give a residue. The residue was dissolved with TFA (4 mL) and the mixture was stirred at 25 °C for 16 h. The mixture was concentrated at reduced pressure to give a residue. The residue was purified by prep-HPLC (column: YMC-Actus Triart C18 150*30 mm*7um; mobile phase: [water (FA) -ACN]; B%: 10%- 40%, 10 min) to give a solid. The solid was separated by SFC (column: DAICEL CHIRALPAK IG (250 mm*30 mm, 10um); mobile phase: [0.1 %NH3H2O MEOH]; B%: 20%- 20%, 5.2 min). Peak 1 (RT = 1 .383 min) was collected to give compound 2A as off white solid.1H NMR: (400 MHz, CDCI3) δ = 7.28 (br s, 1 H), 7.25 (d, J = 5.5 Hz, 1 H), 7.06 - 6.95 (m, 2H), 6.09 (s, 1 H), 4.93 (s, 1 H), 3.28 (td, J = 4.7, 12.2 Hz, 1 H), 3.14 - 3.05 (m, 1 H), 2.97 - 2.86 (m, 1 H), 2.80 - 2.70 (m, 1 H), 2.36 (s, 3H); SFC: RT = 1.383 min. Peak 2 (RT = 1.623 min) was collected to give compound 2B as off yellow gum. 1 H NMR: (400 MHz, CDCI3) δ = 7.30 (dd, J = 5.2, 8.7 Hz, 2H), 7.05 (t, J = 8.6 Hz, 2H), 6.04 (s, 1 H), 5.29 (s, 1 H), 3.25 - 3.06 (m, 3H), 2.89 (br d, J = 14.3 Hz, 1 H), 2.36 (s, 3H); SFC: RT = 1.623 min.

[0401] Synthesis of compound (13)_peak1

[0402] A mixture of compound 2A (30 mg, 121.30 μmol, 1 eq), compound 4 (29.01 mg, 121.30 μmol, 1 eq), DIEA (47.03 mg, 363.89 μmol, 63.38 μL, 3 eq), HOBt (19.67 mg, 145.55 μmol, 1.2 eq) and EDCI (27.90 mg, 145.55 μmol, 1 .2 eq) in DMF (1 mL) was stirred at 20 °C for 16 h. The mixture was purified by prep-HPLC (column: YMC-Actus Triart C18 150*30 mm*7um; mobile phase: [water (FA) -ACN]; B%: 60%- 90%, 10 min). Compound (13)_peak1 (21.59 mg, 45.62 μmol, 37.61 % yield, 99% purity) was obtained as an off white solid.1H NMR: (400 MHz, CDCI3) δ = 7.26 - 7.15 (m, 2H), 6.97 (t, J = 8.7 Hz, 2H), 6.73 (s, 1 H), 6.71 - 6.63 (m, 1 H), 6.33 (s, 1 H), 3.86 - 3.75 (m, 1 H), 3.72 - 3.55 (m, 2H), 3.29 (ddd, J = 4.1 , 11 .9, 14.1 Hz, 1 H), 2.95 - 2.74 (m, 2H), 2.67 - 2.47 (m, 4H), 2.47 - 2.34 (m, 5H), 2.07 - 1 .85 (m, 2H); LCMS: RT =0.619 min, m / z =469.2 (M+H)+; SFC: RT =1.811 min

[0403] Synthesis of compound (13)_peak2

[0404] A mixture of compound 2B (30 mg, 121.30 μmol, 1 eq), compound 4 (29.01 mg, 121.30 μmol, 1 eq), DIEA (47.03 mg, 363.89 μmol, 63.38 μL, 3 eq), HOBt (19.67 mg, 145.55 μmol, 1.2 eq) and EDCI (27.90 mg, 145.55 μmol, 1 .2 eq) in DMF (1 mL) was stirred at 20 °C for 16 h. The mixture was purified by prep-HPLC (column: YMC-Actus Triart C18 150*30 mm*7um; mobile phase: [water (FA) -ACN]; B%: 60%- 90%, 10 min). Compound (13)_peak2 (13.51 mg, 28.55 μmol, 23.54% yield, 99% purity) was obtained as a off white solid.1H NMR: (400 MHz, CDCI3) δ = 7.27 - 7.16 (m, 2H), 7.07 - 6.91 (m, 2H), 6.77 - 6.62 (m, 2H), 6.40 - 6.30 (m, 1 H), 3.80 (dd, J = 4.9, 14.1 Hz, 1 H), 3.73 - 3.57 (m, 2H), 3.29 (ddd, J = 4.3, 12.0, 14.0 Hz, 1 H), 2.97 - 2.74 (m, 2H), 2.67 - 2.47 (m, 4H), 2.65-2.42 (m, 4H), 2.09 - 1.83 (m, 2H); LCMS: RT =0.617 min, m / z =469.2 (M+H)+; SFC: RT =1 .487 min.

[0405] EXAMPLE 1.14: SYNTHESIS OF (14) Synthesis of compound 3

[0406] A mixture of compound 1 (100 mg, 549.03 μmol, 1 eq), compound 2 (64.32 mg, 418.71 μmol, 7.63e- 1 eq, HCI), HOBt (89.02 mg, 658.83 μmol, 1.2 eq), EDCI (126.30 mg, 658.83 μmol, 1.2 eq) and DIEA (177.40 mg, 1.37 mmol, 239.08 μL, 2.5 eq) in DMF (1.5 mL) was stirred at 25 °C for 16 h. To the reaction mixture was added water (6 mL), the resulting mixture was extracted with ethyl acetate (2 mL * 4), and the combined organic phase was washed with brine (10 mL * 1), dried with anhydrous Na2SO4, filtered and the filtrate was concentrated in vacuum to give a residue. Compound 3 (120 mg, 426.64 μmol, 77.71 % yield) was obtained as a yellow oil.1H NMR: (400 MHz, CDCI3) δ = 6.44 (br s, 1 H), 3.71 (s, 3H), 3.54 (ddd, J = 4.5, 6.1 , 13.5 Hz, 1 H), 3.38 (ddd, J = 5.9, 8.0, 13.7 Hz, 1 H), 2.79 - 2.70 (m, 1 H), 2.32 - 2.22 (m, 2H), 2.04 - 1 .94 (m, 2H), 1 .78 - 1 .67 (m, 4H), 1 .19 (d, J = 7.3 Hz, 3H).

[0407] Synthesis of compound 4

[0408] A mixture of compound 3 (120 mg, 426.64 μmol, 1 eq) and LiOH»H2O (35.81 mg, 853.27 μmol, 2 eq) in THF (2 mL) and H2O (0.2 mL) was stirred at 25 °C for 16 h. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was diluted with H2O (6 mL) and acidified to pH = 5 - 6 with 1 M HCI aqueous solution, the resulting mixture was extracted with EtOAc (3 mL *5). The combined organic layers were dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. Compound 4 (96 mg, 359.22 μmol, 84.20% yield) was obtained as a yellow oil.1H NMR: (400 MHz, CDCI3) δ = 6.50 - 6.38 (m, 1 H), 3.61 - 3.53 (m, 1 H), 3.41 (ddd, J = 5.9, 7.9, 13.7 Hz, 1 H), 2.85 - 2.75 (m, 1 H), 2.38 - 2.19 (m, 2H), 2.05 - 1 .94 (m, 2H), 1 .73 (br s, 4H), 1 .28 - 1 .23 (m, 3H).

[0409] Synthesis of (14)

[0410] A mixture of compound 5 (30 mg, 136.18 μmol, 1 eq), compound 4 (36.39 mg, 136.18 μmol, 1 eq) and EDCI (52.21 mg, 272.36 μmol, 2 eq) in pyridine (1 mL) was stirred at 25 °C for 2 h. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep- HPLC (column: YMC-Actus Triart C18 150*30 mm*7um; mobile phase: [water (FA) -ACN]; gradient: 38%- 68% B over 10 min). (14) (23.3 mg, 47.14 μmol, 34.62% yield, 95% purity) was obtained as a white solid.1H NMR: (400 MHz, CDCI3) δ = 8.74 - 8.61 (m, 1 H), 7.29 (d, J = 3.3 Hz, 1 H), 6.94 (br s, 1 H), 6.83 - 6.58 (m, 1 H), 6.29 (br s, 1 H), 5.02 - 4.09 (m, 1 H), 3.93 - 3.69 (m, 4H), 3.54 - 3.44 (m, 2H), 3.31 - 3.09 (m, 1 H), 3.08 - 2.86 (m, 2H), 2.29 - 1.85 (m, 4H), 1.66 - 1.46 (m, 4H), 1 .22 - 1.13 (m, 3H); LCMS: RT =0.490 min, m / z = 470.2 (M+H)+.

[0411] EXAMPLE 1.15: SYNTHESIS OF (15)

[0412] A mixture of compound 1 (50 mg, 174.59 μmol, 1 eq), compound 2 (22.38 mg, 174.59 μmol, 21 .72 μL, 1 eq), HOBt (28.31 mg, 209.50 μmol, 1.2 eq), EDCI (40.16 mg, 209.50 μmol, 1.2 eq) and DIEA (45.13 mg, 349.17 μmol, 60.82 μL, 2 eq) in DMF (1 mL) was stirred at 20 °C for 16 h. The mixture was purified by prep-HPLC (column: Phenomenex luna C18 150*25 mm* 10um; mobile phase: [water (FA) -ACN]; B%: 47%- 77%, 10 min), the eluent was concentrated and then freeze dried. Compound (15) (17.65 mg, 43.62 μmol, 24.98% yield, 98% purity) was obtained as a white solid.1H NMR: (400 MHz, CDCI3) δ = 7.35 - 7.27 (m, 4H), 7.17 (d, J = 5.0 Hz, 1 H), 6.89 (s, 1 H), 6.77 - 6.69 (m, 1 H), 6.39 (br s, 1 H), 3.85 (br dd, J = 4.5, 14.2 Hz, 1 H), 3.66 - 3.50 (m, 2H), 3.34 (ddd, J = 5.0, 11.5, 14.0 Hz, 1 H), 3.04 - 2.86 (m, 2H), 2.85 - 2.50 (m, 2H), 2.07 - 1 .96 (m, 1 H), 1 .84 - 1 .68 (m, 5H), 1 .42 - 1 .13 (m, 5H); LCMS: RT = 0.856 min, m / z = 397.2 (M+H)+; SFC: RT = 0.842 min.

[0413] EXAMPLE 1.16: SYNTHESIS OF (16)

[0414] Synthesis of compound 3

[0415] To a solution of compound 1 (500 mg, 3.67 mmol, 1 eq) and compound 2 (667.39 mg, 3.67 mmol, 1 eq, HCI) in DMF (10 mL) was added HOBt (595.71 mg, 4.41 mmol, 1.2 eq) and EDCI (845.14 mg, 4.41 mmol, 1 .2 eq) and DIEA (1 .42 g, 11 .02 mmol, 1 .92 mL, 3 eq). The mixture was stirred at 25 °C for 16 h. The reaction mixture was purified by reversed-phase HPLC(0.1 % FA condition) followed by lyophilization to give compound 3 (800 mg, 3.04 mmol, 82.71 % yield) as white solid. LCMS: RT =0.729 min, m / z =208.1 (M+H-f-Bu)+.

[0416] Synthesis of compound 4 To a solution of compound 3 (800 mg, 3.04 mmol, 1 eq) in DCM (9 mL) was added TFA (4.62 g, 40.52 mmol, 3 mL, 13.33 eq). The mixture was stirred at 25 °C for 2 h. The reaction mixture was concentrated in vacuum to give compound 4 (965 mg, crude) as yellow oil.

[0417] Synthesis of (16)

[0418] To a solution of compound 4 (50 mg, 241 .34 μmol, 1 eq) and compound 5 (62.60 mg, 241 .34 μmol, 1 eq) in DMF (1 mL) was added HOBt (39.13 mg, 289.61 μmol, 1.2 eq) and EDCI (55.52 mg, 289.61 μmol, 1.2 eq) and DIEA (124.77 mg, 965.37 μmol, 168.15 μL, 4 eq). The mixture was stirred at 25 °C for 16 h. The reaction mixture was purified by prep-HPLC(column: Phenomenex luna C18 150*25 mm* 10um; mobile phase: [water (FA) -ACN]; B%: 43%- 73%, 10 min) followed by lyophilization to give (16) (18.1 mg, 40.35 μmol, 16.72% yield, 100% purity) as white solid.1H NMR: (400 MHz, CD3OD) δ = 7.31 - 7.17 (m, 1 H), 6.90 - 6.76 (m, 3H), 6.64 (s, 1 H), 6.47 - 6.00 (m, 1 H), 4.59 (s, 1 H), 3.99 (br dd, J = 5.3, 14.1 Hz, 1 H), 3.78 - 3.73 (m, 3H), 3.55 - 3.44 (m, 2H), 3.03 - 2.75 (m, 4H), 2.74 - 2.55 (m, 6H), 2.41 (s, 3H); LCMS: RT =0.877 min, m / z =449.3 (M+H)+; SFC: RT=1 .377 min.

[0419] EXAMPLE 1.17: SYNTHESIS OF (17)

[0420] Synthesis of compound 3

[0421] To a solution of compound 1 (100 mg, 385.56 μmol, 1 eq) and compound 2 (78.36 mg, 385.56 μmol, 1 eq) in DMF (1 mL) was added DIEA (99.66 mg, 771.11 μmol, 134.31 μL, 2 eq), HOBt (62.52 mg, 462.67 μmol, 1 .2 eq) and EDCI (88.69 mg, 462.67 μmol, 1 .2 eq). The mixture was stirred at 25 °C for 16 h. The reaction mixture was diluted with H2O (10 mL) and the resulting mixture was extracted with EtOAc (10 mL * 3). The combined organic layers were washed with brine (10 mL), dried over Na2SO4, filtered and filtrate was concentrated under reduced pressure to give residue. The residue was purified by column chromatography (SiO2, PEr / EtOAc = 1 / 1 to 1 / 1 , TLC: PE: EtOAc = 1 : 1 , Rf = 0.8). The eluent was concentrated and reduced pressure to give compound 3 (110 mg, 215.26 μmol, 55.83% yield, 87% purity) as yellow oil. LCMS: RT = 0.694 min, m / z = 445.4 (M+H)+.

[0422] Synthesis of compound 4 To a solution of compound 3 (55 mg, 123.71 μmol, 1 eq) in DCM (2 mL) was added TFA (154.00 mg, 1.35 mmol, 0.1 mL, 10.92 eq). The mixture was stirred at 25 °C for 1 h. The reaction mixture was concentrated and reduced pressure to give compound 4 (40 mg, crude) as red oil. LCMS: RT = 0.658min, m / z = 343.3(M+H)+.

[0423] Synthesis of (17)

[0424] To a solution of compound 4 (40 mg, 116.12 μmol, 1 eq) and compound 5 (13.25 mg, 116.12 μmol, 12.62 μL, 1 eq) in DMF (1 mL) was added DIEA (90.05 mg, 696.72 μmol, 121.36 μL, 6 eq), HOBt (18.83 mg, 139.34 μmol, 1.2 eq) and EDCI (26.71 mg, 139.34 μmol, 1.2 eq). The mixture was stirred at 25 °C for 16 h. The reaction mixture was concentrated under reduced pressure to give residue. The residue was purified by prep-HPLC (column: Phenomenex luna C18 150*25 mm* 10um; mobile phase: [water (FA) -ACN]; B%: 52%-82%, 10 min), the eluent was concentrated and freeze dried. To give (17) (26.24 mg, 58.96 μmol, 50.77% yield, 99% purity) as yellow gum.

[0425] 1H NMR: (400 MHz, DMSO-d6) δ = 7.90 - 7.78 (m, 1 H), 7.23 (q, J = 8.1 Hz, 1 H), 6.85 (br d, J = 9.5 Hz, 1 H), 6.78 - 6.73 (m, 1 H), 6.73 - 6.69 (m, 1 H), 6.61 - 6.55 (m, 1 H), 6.51 (d, J = 5.4 Hz, 1 H), 4.24 - 3.95 (m, 1 H), 3.75 - 3.67 (m, 3H), 3.18 - 3.05 (m, 3H), 2.96 - 2.68 (m, 3H), 2.38 (s, 3H), 2.35-2.30 (m, 1 H), 1 .64 - 1 .42 (m, 8H), 1 .03 - 0.94 (m, 3H); LCMS: RT = 0.667 min, m / z = 441 .4 (M+H)+; SFC: RT = 2.146 min, RT = 2.545 min.

[0426] VT-1H NMR: (400 MHz, DMSO-d6) δ = 7.52 - 7.41 (m, 1 H), 121 - 7.18 (m, 1 H), 6.88 - 6.72 (m, 3H), 6.52 - 6.46 (m, 1 H), 3.72 (s, 3H), 3.27 - 3.17 (m, 2H), 3.14 (br dd, J = 2.9, 4.9 Hz, 1 H), 3.14-3.05 (m, 1 H), 3.03 - 2.94 (m, 2H), 2.85 - 2.75 (m, 2H), 2.43-2.40 (m, 1 H), 2.39 (s, 3H), 1 .70 - 1 .43 (m, 8H), 0.98 (br d, J = 6.0 Hz, 3H);

[0427] EXAMPLE 1.18: SYNTHESIS OF (18)

[0428] To a solution of compound 1 (5 g, 27.08 mmol, 1 eq) in ACN (50 mL) was added 4- methylbenzenesulfonic acid; hydrate (5.15 g, 27.08 mmol, 1 eq) at 0 °C, the mixture was stirred at 0 °C for 0.25 h. To the mixture was added NIS (6.70 g, 29.78 mmol, 1.1 eq), the mixture was stirred at 10 °C for 16 h. The reaction mixture was quenched by adding saturated Na2S20s solution (100 mL), and then the resulting mixture was extracted with EtOAc (50 mL * 3). The combined organic layers were washed with brine (50 mL * 3), dried over Na2SO4, filtered and concentrated under reduced pressure to give residue. The residue was purified by column chromatography (SiO2, Petroleum ether / Ethyl acetate = 1 / 0 to 20 / 1 , TLC, Petroleum ether / Ethyl acetate = 10 / 1 , Rf = 0.5), the eluent was concentrated under reduced pressure to compound 2 (7 g, 22.54 mmol, 83.25% yield) as pink oil.1H NMR: (400 MHz, DMSO-de) δ = 9.79 (s, 1 H), 7.61 (s, 1 H), 7.34 (s, 1 H), 1.22 (s, 9H).

[0429] Synthesis of compound 3

[0430] To a solution of compound 2 (4 g, 12.88 mmol, 1 eq) in ACN (30 mL) was added CS2CO3 (8.39 g, 25.76 mmol, 2 eq) and CH3I (3.66 g, 25.76 mmol, 1 .60 mL, 2 eq). The mixture was stirred at 80 °C for 16 h. The reaction mixture was diluted with H2O (100 mL) and the resulting mixture was extracted with EtOAc (50 mL * 3). The combined organic layers were washed with brine (50 mL * 2), dried over Na2SO4, filtered and concentrated under reduced pressure to give residue. The residue was purified by column chromatography (SiO2, Petroleum ether / Ethyl acetate = 1 / 0 to 20 / 1 , TLC, Petroleum ether / Ethyl acetate = 10 / 1 , Rf = 0.8), the eluent was concentrated under reduced pressure to give compound 3 (4 g, 12.32 mmol, 95.68% yield) as colorless oil.1H NMR: (400 MHz, DMSO-d6) δ = 7.71 (s, 1 H), 7.48 (s, 1 H), 3.75 (s, 3H), 1.24 (s, 9H).

[0431] Synthesis of compound 4

[0432] To a solution of compound 3 (1 g, 3.08 mmol, 1 eq) in DMF (5 mL) was added DIEA (796.36 mg, 6.16 mmol, 1.07 mL, 2 eq), TBAC (941.85 mg, 3.39 mmol, 947.53 μL, 1.1 eq), 3, 3-dimethoxyprop-1- ene (471.98 mg, 4.62 mmol, 1.5 eq) and Pd(OAc)2 (69.17 mg, 308.09 μmol, 0.1 eq) under N2. The mixture was stirred at 120 °C for 0.5 h under microwave. The reaction mixture was quenched by addition of HCI (1 M, 30 mL) solution, and then the resulting mixture was diluted with H2O (50 mL) and extracted with (50 mL * 3). The combined organic layers were washed with brine (50 mL * 3), dried over Na2SO4, filtered and concentrated under reduced pressure to give residue. The residue was purified by column chromatography (SiO2, Petroleum ether / Ethyl acetate = 10 / 1 to 5 / 1 , TLC, Petroleum ether / Ethyl acetate = 5: 1), the eluent was concentrated under reduced pressure to give compound 4 (650 mg, 2.28 mmol, 74.09% yield) as yellow oil.1H NMR: (400 MHz, CD3OD) δ = 7.25 (s, 1 H), 7.17 (s, 1 H), 3.81 (s, 3H), 3.65 (s, 3H), 2.95 (t, J = 7.6 Hz, 2H), 2.62 (t, J = 7.6 Hz, 2H), 1 .28 (s, 9H).

[0433] Synthesis of compound 5

[0434] To a solution of compound 4 (300 mg, 1.05 mmol, 1 eq) in DCM (10 mL) was added BBrs (395.87 mg, 1 .58 mmol, 210.69 μL, 1 .5 eq) at 0 °C. The mixture was stirred at 20 °C for 1 h. The reaction mixture was quenched by adding MeOH (10 mL) slowly, and then the pH resulting mixture was adjusted to 7-8, the resulting mixture was extracted with EtOAc (30 mL * 3). The combined organic layers were washed with brine (30 mL * 2), dried over Na2SO4, filtered and concentrated under reduced pressure to give residue. The residue was purified by column chromatography (SiO2, Petroleum ether / Ethyl acetate = 10 / 1 to 5 / 1 , TLC, Petroleum ether / Ethyl acetate = 5 / 1 , Rf = 0.75), the eluent was concentrated under reduced pressure to give compound 5 (180 mg, 591 .69 μmol, 56.17% yield, 89% purity) as yellow oil. LCMS: RT = 0.393 min, m / z = 271 .3 (M+H)+.

[0435] Synthesis of compound 6

[0436] To a solution of compound 5 (180 mg, 664.82 μmol, 1 eq) in THF (2 mL) and H2O (0.5 mL) was added LiOH’FW (139.49 mg, 3.32 mmol, 5 eq). The mixture was stirred at 20 °C for 2 h. The reaction mixture was diluted with H2O (10 mL), then HCI (1 M, 20 mL) was added. The resulting mixture was extracted with EtOAc (10 mL*3), The combined organic layers were washed with brine (10 mL *2), dried over Na2SO4, filtered and concentrated under reduced pressure to give compound 6 (100 mg, 389.52 μmol, 58.59% yield) as yellow solid.1H NMR: (400 MHz, CD3OD) δ = 7.15 (d, J = 2.3 Hz, 1 H), 7.10 (d, J = 2.3 Hz, 1 H), 2.91 (t, J = 7.6 Hz, 2H), 2.59 (t, J = 7.6 Hz, 2H), 1 .27 (s, 9H).

[0437] Synthesis of (18)

[0438] To a solution of compound 7 (30 mg, 138.70 μmol, 1 eq) and compound 6 (35.61 mg, 138.70 μmol, 1 eq) in pyridine (1 mL) was added EDCI (31 .91 mg, 166.43 μmol, 1.2 eq). The mixture was stirred at 20 °C for 16 h. The reaction mixture was diluted with EtOAc (10 mL), then the resulting mixture was washed with H2O (10 mL*3), dried over Na2SO4, filtered and concentrated under reduced pressure to give residue. The residue was purified by prep-HPLC (column: Welch Xtimate C18 150*25 mm*5um; mobile phase: [water (FA) -ACN]; gradient: 60%-80% B over 10 min), the eluent was concentrated and then freeze dried to give (18) (6.1 mg, 13.27 μmol, 9.57% yield, 99% purity) as white solid.1H NMR: (400 MHz, CD3OD) δ = 8.91 - 8.84 (m, 1 H), 7.35 - 7.24 (m, 3H), 7.21 - 7.00 (m, 4H), 6.95 (s, 1 H), 4.11 (dd, J = 5.5, 14.4 Hz, 1 H), 3.35-3.17 (m, 1 H), 3.08 - 2.83 (m, 4H), 2.78 - 2.65 (m, 2H), 1.24 - 1.16 (m, 1 H), 1 .17 (s, 6H); LCMS: RT = 0.651 min, m / z = 455.3 (M+H)+.

[0439] EXAMPLE 1.19: SYNTHESIS OF (19)

[0440] Synthesis of compound 2 To a solution of compound 1 (300 mg, 2.91 mmol, 1 eq) in MeOH (3 mL) added SOCh (692.23 mg, 5.82 mmol, 422.60 μL, 2 eq) dropwise under 0 °C. The mixture was stirred at 25 °C for 16 h.The reaction mixture was concentrated to give a residue. Compound 2 (439 mg, 2.86 mmol, 98.24% yield, HCI) was obtained as a white solid.

[0441] Synthesis of compound 4

[0442] A mixture of compound 2(439 mg, 2.86 mmol, 1 eq, HCI), compound 3(388.96 mg, 2.86 mmol, 1 eq), DIEA (923.42 mg, 7.14 mmol, 1.24 mL, 2.5 eq), HOBt (463.41 mg, 3.43 mmol, 1.2 eq) and EDCI (657.45 mg, 3.43 mmol, 1 .2 eq) in DMF (4 mL) was stirred at 25 °C for 16 h. The reaction mixture was diluted with H2O (15 mL) and the resulting mixture was extracted with EtOAc (60 mL * 3). The combined organic layers were washed with brine (50 mL *1), dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. Compound 4 (600 mg, 1.99 mmol, 69.62% yield, 78% purity) was obtained as a brown solid. LCMS: RT = 0.41 Omin, m / z = 204.1 (M+H)+.

[0443] Synthesis of compound 5

[0444] A mixture compound 4 (600 mg, 2.55 mmol, 1 eq) and LiOH H2O (214.08 mg, 5.10 mmol, 2 eq) in THF (6 mL) and MeOH (0.6 mL) and H2O (1.5 mL) was stirred at 20 °C for 32 h. To the mixture was added HCI (2M) and the pH of the mixture was adjusted to about 4. The mixture was diluted with H2O (10 mL) and the resulting mixture was extracted with EtOAc (12 mL* 3). The combined organic layers were concentrated at reduced pressure to give compound 5 (328 mg, 1.47 mmol, 57.55% yield, 99% purity) as a white solid. LCMS: RT =0.334 min, m / z = 204(M+H)+.

[0445] Synthesis of (19)

[0446] A mixture of compound 6 (28 mg, 119.51 μmol, 1 eq), compound 5 (42.30 mg, 191.21 μmol, 1 .6 eq), HOBt (19.38 mg, 143.41 μmol, 1.2 eq), EDCI (27.49 mg, 143.41 μmol, 1.2 eq) and DIEA (38.61 mg, 298.77 μmol, 52.04 μL, 2.5 eq) in DMF (1 mL) was stirred at 20 °C for 16 h. The reaction mixture was diluted with H2O (6 mL) and the resulting mixture was extracted with EtOAc (5 mL * 3). The combined organic layers were washed with brine (15 mL * 1), dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (column: Phenomenex luna C18 150*25 mm* 10um; mobile phase: [water (FA) -ACN]; gradient: 38%- 68% B over 7 min).

[0447] (19)_peak1 (6.78 mg, 15.19 μmol, 12.71 % yield, 98% purity) was obtained as a white solid.1H NMR: (400 MHz, CDCI3) δ = 8.73 (s, 1 H), 7.24 - 7.21 (m, 1 H), 7.08 - 6.95 (m, 3H), 6.32 - 6.05 (m, 1 H), 4.14 - 4.06 (m, 1 H), 3.52 - 3.33 (m, 3H), 3.19 - 3.09 (m, 1 H), 3.09 - 2.99 (m, 2H), 2.92 - 2.74 (m, 1 H), 2.73 - 2.48 (m, 4H), 1.21 - 1.10 (m, 3H); LCMS: RT =0.502 min, m / z = 438.1 (M+H)+; SFC: RT = 1.115 min.

[0448] (19)_peak2 (6.60 mg, 15.09 μmol, 12.62% yield, 100% purity) was obtained as a white solid.1H NMR:(400 MHz, CDCI3) δ = 8.76 - 8.72 (m, 1 H), 7.25-7.20 (m, 1 H), 7.06 - 6.95 (m, 3H), 6.33 - 6.19 (m, 1 H), 4.07 - 3.91 (m, 1 H), 3.59 - 3.30 (m, 3H), 3.29 - 3.07 (m, 1 H), 3.05 - 2.90 (m, 2H), 2.89 - 2.78 (m, 1 H), 2.77 - 2.51 (m, 4H), 1.21 - 1 .06 (m, 3H); LCMS: RT = 0.495min, m / z = 438.1 (M+H)+; SFC: RT = 1 .253 min.

[0449] EXAMPLE 1.20: SYNTHESIS OF (20)

[0450] (20), peak 1 (20), peak 2

[0451] Synthesis of compound 3

[0452] A mixture of compound 1 (500 mg, 2.49 mmol, 1 eq, 2HCI), compound 2 (416.08 mg, 2.73 mmol, 1 .1 eq), HOBt (403.12 mg, 2.98 mmol, 1 .2 eq), EDCI (571 .91 mg, 2.98 mmol, 1 .2 eq) and DIEA (803.28 mg, 6.22 mmol, 1.08 mL, 2.5 eq) in DMF (10 mL) was stirred at 20 °C for 16 h. The reaction mixture was diluted with H2O (60 mL) and the resulting mixture was extracted with EtOAc (60 mL). The organic layer was concentrated at reduced pressure to give a residue. The residue was purified by flash silica gel chromatography (ISCO®; 12 g SepaFlash® Silica Flash Column, Eluent of 45-50% Ethyl acetate / Petroleum ethergradient @ 40 mL / min). Compound 3 (534 mg, 2.04 mmol, 81 .88% yield) was obtained as colorless oil.1H NMR: (400 MHz, CDCI3) δ = 8.71 (s, 1 H), 7.68 (s, 1 H), 7.38 - 7.28 (m, 3H), 7.06 (td, J = 1 .2, 8.0 Hz, 1 H), 6.69 (br s, 1 H), 3.86 (s, 3H), 3.73 (q, J = 6.5 Hz, 2H), 3.23 (t, J = 6.6 Hz, 2H).

[0453] Synthesis of compound4

[0454] A mixture of compound 3 (241 mg, 918.70 μmol, 1 eq) in POCI3 (4 mL) was stirred at 105 °C for 16 h. The reaction mixture was added into H2O (30 mL) slowly at 20 °C to quenched the reaction, and then the resulting mixture was extracted with EtOAc (20 mL*3). The combined organic layers were dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by flash silica gel chromatography (ISCO®; 4 g SepaFlash® Silica Flash Column, Eluent of 30-45% Ethyl acetate / Petroleum ethergradient @ 8 mL / min). Compound 4 (30 mg, 110.51 μmol, 12.03% yield, 90% purity) as a brown oil.1H NMR: (400 MHz, CDCI3) δ = 8.68 (s, 1 H), 7.60 - 7.54 (m, 2H), 7.36 (t, J = 7.9 Hz, 1 H), 7.01 (dd, J = 1 .8, 8.1 Hz, 1 H), 4.04 (t, J = 8.2 Hz, 2H), 3.86 (s, 3H), 3.09 (t, J = 8.2 Hz, 2H).

[0455] Synthesis of compound 5

[0456] To a solution of compound 4 (50 mg, 204.66 μmol, 1 eq) in MeOH (2 mL) was added NaBH4 (40 mg, 1.06 mmol, 5.17 eq) and the resulting mixture was stirred at 20 °C for 2 h. The mixture was quenched with aqueous NH4CI solution (4 mL) and the resulting mixture was extracted with EtOAc (6 mL x 3), the combined organic phase was dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. Compound 5 (36 mg, 146.15 μmol, 71.41 % yield) was obtained as a white solid.

[0457] Synthesis of (20)_peak1 , (20)_peak2

[0458] A mixture of compound 5 (36 mg, 146.15 μmol, 1 eq), compound 6 (30.28 mg, 146.15 μmol, 1 eq), HOBt (23.70 mg, 175.38 μmol, 1.2 eq), EDCI (33.62 mg, 175.38 μmol, 1.2 eq) and DIEA (47.22 mg, 365.37 μmol, 63.64 μL, 2.5 eq) in DMF (1 mL) was stirred at 20 °C for 16 h. The reaction mixture was diluted with H2O (8 mL) and the resulting mixture was extracted with EtOAc (10 mL * 3). The combined organic layers were dried over Na2SO4, filtered and concentrated at reduced pressure to give a residue. The residue was purified by prep-HPLC (column: YMC-Actus Triart C18 150*30 mm*7um; mobile phase: [water (FA) -ACN]; B%: 35%- 65%, 10 min) to give a white solid. The solid was separated by SFC (column: DAICEL CHIRALPAK AS (250 mm*30 mm, 10um); mobile phase: [CO2-ACN / i-PrOH (0.1 %IPAm)]; 55% B isocratic elution mode).

[0459] Peakl (RT = 1 .473 min) was collected to give (20)_peak1 as a white solid.

[0460] Peak2 (RT = 2.332 min) was collected to give (20)_peak2 as a white solid.

[0461] (20)_peak1 (7.91 mg, 17.20 μmol, 11.77% yield, 94.7% purity) was obtained as a white solid.1H NMR (400 MHz, CDCI3) δ = 8.73 (s, 1 H), 7.24 (dd, J = 7.3, 9.1 Hz, 1 H), 7.00 (s, 1 H), 6.96 - 6.91 (m,

[0462] 1 H), 6.86 - 6.79 (m, 2H), 6.56 - 6.47 (m, 1 H), 3.97 - 3.86 (m, 1 H), 3.82 - 3.75 (m, 3H), 3.67 - 3.58 (m,

[0463] 2H), 3.50 - 3.40 (m, 1 H), 3.09 - 2.94 (m, 2H), 2.90 - 2.77 (m, 2H), 2.75 - 2.58 (m, 5H); LCMS: RT = 0.481 min, m / z =436.4 (M+H)+; SFC: RT = 1 .489 min.

[0464] (20)_peak2 (7.77 mg, 17.31 μmol, 11 .84% yield, 97% purity) was obtained as a white solid.1H NMR: (400 MHz, CDCI3) δ = 8.74 - 8.70 (m, 1 H), 7.24 (br dd, J = 7.3, 9.2 Hz, 1 H), 7.00 (s, 1 H), 6.96 - 6.91 (m, 1 H), 6.86 - 6.79 (m, 2H), 6.56 - 6.45 (m, 1 H), 3.92 (br dd, J = 3.4, 12.7 Hz, 1 H), 3.81 - 3.76 (m, 3H), 3.66 - 3.59 (m, 2H), 3.50 - 3.40 (m, 1 H), 3.07 - 2.97 (m, 2H), 2.89 - 2.80 (m, 2H), 2.75 - 2.62 (m, 5H); LCMS:

[0465] RT = 0.474 min, m / z = 436.3(M+H)+; SFC: RT = 2.372 min.

[0466] EXAMPLE 1.21: SYNTHESIS OF (21)

[0467] A mixture of compound 6A (50.00 mg, 174.59 μmol, 1 eq), 1 -(3-fluorophenyl)cyclopropane- carboxylic acid (31.46 mg, 174.59 μmol, 1 eq), HOBt (28.31 mg, 209.50 μmol, 1.2 eq) , EDCI (40.16 mg, 209.50 μmol, 1 .2 eq) and DIEA (45.13 mg, 349.17 μmol, 60.82 μL, 2 eq) in DMF (1 mL) was stirred at 20 °C for 16 h. The reaction mixture was concentrated under vacuum to give a residue. The residue was purified by prep-HPLC (column: Unisil 3 - 100 C18 Ultra 150*50 mm*3 um;mobile phase : [water (FA) -AON] ;B% : 54%- 84%, 7 min) to give (21) (28.06 mg, 61.93 μmol, 35.47% yield, 99% purity) as white solid.1H NMR: (400 MHz, DMSO-d6) δ = 7.40 - 7.26 (m, 5H), 7.17 - 7.01 (m, 5H), 6.85 - 6.74 (m, 2H), 6.64 (s, 1 H), 3.97 - 3.89 (m, 1 H), 3.30 - 3.20 (m, 2H), 3.10 (ddd, J = 5.3, 11 .1 , 14.2 Hz, 1 H), 2.98 - 2.70 (m, 3H), 2.57 (br s, 1 H), 1 .35 - 1 .27 (m, 2H), 1 .01 - 0.91 (m, 2H); LCMS: RT = 0.958 min, m / z = 449.1 (M+H)+

[0468] EXAMPLE 1.22: SYNTHESIS OF (22)

[0469] Synthesis of compound 2

[0470] To the mixture of compound 1 (300 mg, 2.91 mmol, 1 eq) in MeOH (10 mL) was added SOCh (1 .04 g, 8.73 mmol, 633.91 μL, 3 eq). The mixture was stirred at 25 °C for 16 hr. The reaction mixture was concentrated under reduced pressure to give a residue. Compound 2 (400 mg, crude, HCI) was obtained as a white solid.1H NMR: (400 MHz, DMSO-d6) δ = 8.25 (br s, 3H), 3.64 (s, 3H), 3.07 - 2.98 (m, 1 H), 2.91 - 2.78 (m, 2H), 1.16 (d, J = 6.9 Hz, 3H).

[0471] Synthesis of compound 4

[0472] To the mixture of compound 2 (400 mg, 3.41 mmol, HCI salt) in DMF (4 mL) was added HOBt (507.52 mg, 3.76 mmol, 1.1 eq), EDCI (720.03 mg, 3.76 mmol, 1.1 eq), DIEA (2.21 g, 17.07 mmol, 2.97 mL, 5 eq) and compound 3 (464.71 mg, 3.41 mmol, 1 eq). The mixture was stirred at 25 °C for 16 h. The residue was diluted with H2O (30 mL) and the resulting mixture was extracted with Ethyl acetate (40 mL * 3). The combined organic layers were dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (column: YMC-Actus Triart C18 150*30 mm*7um; mobile phase: [water (FA) -ACN]; gradient: 23%- 53% B over 10 min). Compound 4 (353 mg, crude) was obtained as a yellow oil.1H NMR: (400 MHz, CDCI3) δ = 6.26 (br s, 1 H), 3.67 (s, 3H), 3.49 (ddd, J = 4.5, 6.6, 13.7 Hz, 1 H), 3.30 (ddd, J = 5.8, 8.2, 13.7 Hz, 1 H), 2.89 - 2.59 (m, 6H), 1.15 (d, J = 7.3 Hz, 3H)

[0473] Synthesis of compound 5

[0474] To the mixture of compound 4 (353 mg, 1.50 mmol, 1 eq) in THF (4 mL) was added MeOH (0.7 mL), H2O (0.4 mL) and LiOH (71 .88 mg, 3.00 mmol, 2 eq). The mixture was stirred at 25 °C for 2 hr. To the mixture was added aqueous HCI solution and the pH of the mixture was adjusted to 4~5. To the mixture was added water (40 mL) and the resulting mixture was extracted with EtOAc (40 mL * 3), the combined organic phase was dried over Na2SO4, filtered and the filtrate was concentrated under reduced pressure to give a residue. Compound 5 (276 mg, crude) was obtained as a white solid.1H NMR: (400 MHz, CDCI3) δ = 6.30 (br t, J = 5.6 Hz, 1 H), 3.55 (ddd, J = 4.3, 6.7, 13.7 Hz, 1 H), 3.36 (ddd, J = 5.9, 8.3, 13.9 Hz, 1 H), 2.99 - 2.61 (m, 5H), 1 .23 (d, J = 7.3 Hz, 3H).

[0475] Synthesis of compound (22)

[0476] To the mixture of compound 5 (44.90 mg, 202.98 μmol, 1 eq) in DMF (1 mL) was added HOBt (32.91 mg, 243.58 μmol, 1 .2 eq), EDCI (46.69 mg, 243.58 μmol, 1 .2 eq), DIEA (78.70 mg, 608.95 μmol, 106.07 μL, 3 eq) and compound 6 (50 mg, 202.98 μmol, 1 eq). The mixture was stirred at 25 °C for 16 h. The mixture was diluted with H2O (20 mL) and the resulting mixture was extracted with Ethyl acetate (20 mL * 3). The combined organic layers were dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (column: YMC-Actus Triart C18 150*30 mm*7um; mobile phase: [water (FA) -ACN]; gradient: 38%- 68% B over 10 min). Compound (22) (19.46 mg, 42.86 μmol, 21.11 % yield, 99% purity) was obtained as a white solid.1H NMR: (400 MHz, CDCI3) δ = 8.75 - 8.71 (m, 1 H), 7.26 - 7.19 (m, 1 H), 7.04 (br d, J = 4.0 Hz, 1 H), 6.90 - 6.75 (m, 3H), 6.32 - 6.10 (m, 1 H), 4.15 - 3.91 (m, 1 H), 3.82 - 3.72 (m, 3H), 3.63 - 3.50 (m, 1 H), 3.48 - 3.28 (m, 2H), 3.25 - 3.07 (m, 1 H), 3.06 - 2.99 (m, 1 H), 2.98 - 2.76 (m, 2H), 2.74 - 2.39 (m, 4H), 1 .22 - 1 .05 (m, 3H); LCMS: RT =0.493 min, m / z = 450.2(M+H)+. EXAMPLE 1.23: SYNTHESIS OF (23)

[0477] Synthesis of compound 2

[0478] A mixture of compound 1 (2 g, 15.72 mmol, 1.83 mL, 1 eq), cyclopropanecarboxylic acid (1.35 g, 15.72 mmol, 1.24 mL, 1 eq), HOBt (2.55 g, 18.87 mmol, 1.2 eq), DIEA (6.10 g, 47.17 mmol, 8.22 mL, 3 eq) and EDCI (3.62 g, 18.87 mmol, 1.2 eq) in DMF (20 mL), and then the mixture was stirred at 20 °C for 16 h. The reaction mixture was concentrated under vacuum. The reaction mixture was diluted with ethyl acetate (50 mL). The organic layer was washed with HCI (1 M) (20 mL*3), dried over Na2SO4, filtered and concentrated. Without purification to give compound 2 (2.7 g, 13.83 mmol, 87.94% yield) as yellow solid.1H NMR: (400 MHz, CHLOROFORM-d) δ = 7.17 (dd, J = 1 .1 , 5.1 Hz, 1 H), 6.96 (dd, J = 3.4, 5.1 Hz, 1 H), 6.85 (dd, J = 0.8, 3.4 Hz, 1 H), 5.91 (br s, 1 H), 3.55 (q, J = 6.6 Hz, 2H), 3.04 (t, J = 6.7 Hz, 2H), 1 .35 - 1 .28 (m, 1 H), 1 .01 - 0.93 (m, 2H), 0.77 - 0.68 (m, 2H)

[0479] Synthesis of compound 3

[0480] A mixture of compound 2 (500 mg, 2.56 mmol, 1 eq) in ACN (5 mL) was added POCh (1 .57 g, 10.24 mmol, 951.73 μL, 4 eq), then the mixture was stirred at 60 °C for 16 h. The mixture was quenched by water (10 mL), To the mixture was added aqueous NaHCOs solution and the pH was adjusted to 7, the mixture was extracted with EtOAc (15 mL x 3), dried over Na2SO4, filtered and the filtrate was concentrated under reduced pressure to give a residue. Without further purification and the products was used for next step directly to give compound 3 (180 mg, 1 .02 mmol, 39.66% yield) as yellow gum. LCMS: RT = 0.313 min, m / z = 178.1 (M+H)+.

[0481] Synthesis of compound 4

[0482] To the mixture of compound 3 (180 mg, 1 .02 mmol, 1 eq) in MeOH (3 mL) was added NaBH4 (190 mg, 5.02 mmol, 4.95 eq) at 0 °C. The mixture was stirred at 20 °C for 2 h. To the mixture was added water (10 mL) and extracted with EtOAc (10 mL x 3), the organic phase was dried over Na2SO4, filtered and the filtrate was concentrated under reduced pressure to give a residue. LCMS: EW31975-592-P10. The pH of the aqueous phase was adjusted to 7 by hydrochloric acid (1 M), and then pour it into the waste liquid barrel to give compound 4 (170 mg, crude) was obtained as yellow oil. LCMS: RT = 0.527 min, m / z = 180.2 (M+H)+. Synthesis of (23)

[0483] To a solution of compound 4 (50 mg, 278.89 μmol, 1 eq), HOBt (45.22 mg, 334.67 μmol, 1.2 eq), EDCI (64.16 mg, 334.67 μmol, 1.2 eq) and DIEA (108.13 mg, 836.67 μmol, 145.73 μL, 3 eq) in DMF (2 mL) was addedi - (2 -chlorophenyl) sulfonylpiperidine- 4 -carboxylic acid (84.72 mg, 278.89 μmol, 1 eq). The mixture was stirred at 20 °C for 16 h. The reaction mixture was concentrated under vacuum. The residue was purified by prep-HPLC (column: YMC-Actus Triart C18 150*30 mm*7um; mobile phase: [water (FA) -ACN]; B%: 50%- 80%, 10 min) to give (23) (23.08 mg, 49.14 μmol, 17.62% yield, 99% purity) as off-white solid.1H NMR: (400 MHz, CDCI3) δ = 8.08 (br d, J = 7.6 Hz, 1 H), 7.56 - 7.46 (m, 2H), 7.43 - 7.36 (m, 1 H), 7.11 (d, J = 5.0 Hz, 1 H), 6.91 - 6.79 (m, 1 H), 5.07 (d, J = 8.9 Hz, 1 H), 4.05 (br d, J = 14.4 Hz, 1 H), 3.97 - 3.56 (m, 3H), 3.05 - 2.61 (m, 5H), 2.07 - 1 .68 (m, 4H), 1 .24 - 1 .02 (m, 1 H), 0.72 - 0.30 (m, 4H); LCMS: RT = 0.598 min, m / z = 465.1 (M+H)+.

[0484] Synthesis of (23)_peak 1 and (23)_peak 2

[0485] The 21.65 mg off-white solid was purified by SFC (column: DAICEL CHIRALPAK AD (250 mm*30 mm, 10um); mobile phase: [ACN / IPA (0.1 %NH3H2O)]; B%: 45%- 45%, 2.9 min) to give (23)_peak 1 (crude) and (23)_peak 2 (crude). The (23)_peak 1 (crude) was purified by prep-HPLC (column: Phenomenex luna C18 150*25 mm* 10um; mobile phase: [water (FA) -ACN]; B%: 60%- 80%, 10 min) to give (23)_peak 1 (5.73 mg, 12.08 μmol, 4.33% yield, 98% purity) as white solid.1H NMR: (400 MHz, CDCI3) δ = 8.08 (br d, J = 6.9 Hz, 1 H), 7.56 - 7.46 (m, 2H), 7.43 - 7.36 (m, 1 H), 7.11 (br d, J = 4.9 Hz, 1 H), 6.92 - 6.80 (m, 1 H), 5.13 - 4.85 (m, 1 H), 4.48 - 3.99 (m, 1 H), 3.97 - 3.57 (m, 3H), 3.01 - 2.61 (m, 5H), 2.00 - 1 .67 (m, 4H), 1 .23 - 1 .04 (m, 1 H), 0.68 - 0.30 (m, 4H); LCMS: RT = 0.594 min, m / z = 464.9 (M+H)+; SFC: RT = 1.932 min.

[0486] The (23)_peak 2 (crude) was purified by prep-HPLC (column: Phenomenex luna C18 150*25 mm* 10um; mobile phase: [water (FA) -ACN]; B%: 60%- 80%, 10 min) to give (23)_peak 2 (6.1 1 mg, 12.74 μmol, 4.57% yield, 97% purity) as white solid.1H NMR: (400 MHz, CDCI3) δ = 8.11 - 8.03 (m, 1 H), 7.55 - 7.46 (m, 2H), 7.43 - 7.36 (m, 1 H), 7.11 (br d, J = 5.0 Hz, 1 H), 6.92 - 6.79 (m, 1 H), 5.11 - 4.85 (m, 1 H), 4.47 - 3.99 (m, 1 H), 3.97 - 3.58 (m, 3H), 3.01 - 2.63 (m, 5H), 1 .97 - 1 .68 (m, 4H), 1 .22 - 1 .05 (m, 1 H), 0.69 - 0.32 (m, 4H); LCMS: RT = 0.595 min, m / z = 465.2 (M+H)+; SFC: RT = 1 .447 min .

[0487] EXAMPLE 1.24: SYNTHESIS OF (24)

[0488] 1 3 (24)

[0489] Synthesis of compound 3

[0490] A mixture of compound 2 (96.40 mg, 708.05 μmol, 86.07 μL, 1 eq) and compound 1 (100 mg, 708.05 μmol, 1 eq) in Tol. (2 mL) was stirred at 1 10 °C for 16 h. The mixture was concentrated at reduced pressure to remove the solution, then to the mixture was added TFA (5 mL) and stirred at 25 °C for 16h . The mixture was concentrated at reduced pressure to give residue. The residue was purified by reversed-phase HPLC (0.1 % FA condition), the eluent was concentrated and then freeze dried. Compound 5 (160 mg, 616.89 μmol, 87.13% yield) was obtained as a yellow gum.1H NMR: (400 MHz, CDCI3) δ = 7.27 (s, 2H), 6.94 - 6.83 (m, 3H), 6.11 (s, 1 H), 5.27 (s, 1 H), 3.76 (s, 3H), 3.29 - 3.09 (m, 3H), 2.95 - 2.84 (m, 1 H), 2.36 (s, 3H).

[0491] Synthesis of compound (24)

[0492] A mixture of compound 4 (57.13 mg, 308.44 μmol, 1 eq) , compound 3 (80 mg, 308.44 μmol, 1 eq) , DIEA (119.59 mg, 925.33 μmol, 161.18 μL, 3 eq) , HOBt (50.01 mg, 370.13 μmol, 1.2 eq) and EDCI (70.96 mg, 370.13 μmol, 1 .2 eq) in DMF (1 mL) was stirred at 20 °C for 16 h. The reaction mixture was purified by prep-HPLC (column : Phenomenex luna C18 150*25 mm* 10um;mobile phase : [water (FA) -ACN] ;B% : 61 %- 91 %, 10 min) , the eluent was concentrated and then freeze dried. Compound (24) (33.58 mg, 76.36 μmol, 24.76% yield, 97% purity) was obtained as a white solid.1H NMR: (400 MHz, CDCI3) δ = 7.22 (t, J = 8.0 Hz, 1 H), 6.89 - 6.79 (m, 3H), 6.78 - 6.70 (m, 1 H), 6.43 - 6.30 (m, 2H), 3.85 - 3.76 (m, 4H), 3.65 - 3.53 (m, 2H), 3.34 (ddd, J = 4.3, 12.1 , 14.0 Hz, 1 H), 2.96 - 2.85 (m, 1 H), 2.83 - 2.74 (m, 1 H), 2.73 - 2.49 (m, 2H), 2.49 - 2.43 (m, 1 H), 2.41 (s, 3H), 1.88 - 1.61 (m, 8H); LCMS: RT =0.973 min, m / z =427.0 (M+H)+.

[0493] Synthesis of compound (24)_peak1 and (24)_peak2

[0494] The (24) (32 mg) was separated by SFC (column : Daicel ChiralPak IG (250*30 mm, 10um) ;mobile phase : [0.1 %NH3H2O ETCH] ;B% : 60%- 60%, 4.1 min) .

[0495] Peak 1 (SFC: RT =1.149 min) was collected to give (24)_peak1 (13.25 mg, 29.82 μmol, 39.75% yield, 96% purity) as yellow solid.1H NMR: (400 MHz, CDCI3) δ = 7.15 (br t, J = 8.1 Hz, 1 H), 6.82 - 6.71 (m, 3H), 6.69 - 6.62 (m, 1 H), 6.41 - 6.23 (m, 2H), 3.88 - 3.61 (m, 4H), 3.51 (br s, 2H), 3.33 - 3.20 (m, 1 H), 2.99 - 2.77 (m, 1 H), 2.76 - 2.66 (m, 1 H), 2.64 - 2.44 (m, 2H), 2.43 - 2.31 (m, 4H), 1 .78 - 1 .59 (m, 8H); LCMS: RT =0.772 min, m / z =427.2 (M+H)+; SFC: RT =1 .149 min.

[0496] Peak 2 (SFC: RT =1.776 min) was collected to give (24)_peak2 (13.1 1 mg, 29.81 μmol, 39.74% yield, 97% purity) as off white solid.1H NMR: (400 MHz, CDCI3) δ = 7.22 (t, J = 8.0 Hz, 1 H) , 6.89 - 6.79 (m, 3H) , 6.77 - 6.69 (m, 1 H) , 6.44 - 6.33 (m, 2H) , 3.86 - 3.73 (m, 4H) , 3.67 - 3.50 (m, 2H) , 3.40 - 3.27 (m, 1 H) , 3.06 - 2.84 (m, 1 H) , 2.83 - 2.70 (m, 1 H) , 2.67 - 2.50 (m, 2H) , 2.46 (br t, J = 7.8 Hz, 1 H) , 2.41 (s, 3H) , 1 .90 - 1 .66 (m, 8H); LCMS: RT =0.765 min, m / z =427.2 (M+H)+; SFC: RT =1 .776 min.

[0497] EXAMPLE 1.25: SYNTHESIS OF (25)

[0498] Synthesis of compound 2A and compound 2B

[0499] The 560 mg of compound 1 was separated by SFC (column: DAICEL CHIRALPAK AY-H (250 mm • 30 mm, 10um); mobile phase: [0.1 %NH3H2O ETCH]; B%: 20%- 20%, 2.7 min). Peak 1 (SFC: 1.204 min) was collected to give compound 2A (200 mg, 1.07 mmol, 34.29% yield, 96% purity) as a yellow gum.1H NMR: (400 MHz, CDCI3) δ = 7.13 - 7.09 (m, 1 H), 7.08 - 7.04 (m, 1 H), 3.50 - 3.42 (m, 1 H), 3.14 (br d, J = 9.6 Hz, 1 H), 3.09 - 2.96 (m, 2H), 2.89 - 2.79 (m, 1 H), 1 .08 (dddd, J = 3.1 , 4.8, 7.9, 9.6 Hz, 1 H), 0.82 - 0.71 (m, 1 H), 0.67 - 0.59 (m, 1 H), 0.58 - 0.52 (m, 1 H), 0.48 (td, J = 4.6, 9.3 Hz, 1 H);

[0500] Peak 2 (SFC: 1 .428 min) was collected to give compound 2B (240 mg, 1.27 mmol, 40.71 % yield, 95% purity) as a yellow gum.1H NMR: (400 MHz, CDCI3) δ = 7.14 (d, J = 5.3 Hz, 1 H), 7.05 (d, J = 5.1 Hz, 1 H), 3.61 - 3.51 (m, 1 H), 3.33 (br d, J = 9.8 Hz, 1 H), 3.22 - 3.09 (m, 2H), 2.99 - 2.87 (m, 1 H), 1 .27 - 1 .09 (m, 1 H), 0.89 - 0.78 (m, 1 H), 0.75 - 0.63 (m, 2H), 0.62 - 0.53 (m, 1 H).

[0501] Synthesis of compound 5

[0502] To a solution of compound 4 (367.87 mg, 2.57 mmol, 1 eq) and Py (609.68 mg, 7.71 mmol, 622.12 μL, 3 eq) in DCM (5 mL) was added compound 3 (500 mg, 2.57 mmol, 342.47 μL, 1 eq). The mixture was stirred at 20 °C for 16 h. The mixture was concentrated at reduced pressure to give a residue. Compound 5 (1 .3 g, crude) was obtained as a yellow solid. LCMS: RT =0.524 min, m / z =302.1 (M+H)+.

[0503] Synthesis of compound 6

[0504] A mixture of compound 5 (1.3 g, 4.31 mmol, 1 eq) and LiOH. H2O (362.08 mg, 8.63 mmol, 2 eq) in THF (13 mL), MeOH (6 mL) and H2O (4 mL) was stirred at 20 °C for 2 h. To the mixture was added aqueous HCI solution and the pH was adjusted to about 3~4. EtOAc (25 mL) were added and Layers were separated. The aqueous phase was extracted with EtOAc (25 mL x 2). The combined organic phase was dried over Na2SO4, filtered, and concentrated under vacuum to give a residue. Compound 6 (800 mg, crude) was obtained as a white solid.1H NMR: (400 MHz, DMSO-de) δ = 12.26 (br s, 1 H), 7.76 - 7.67 (m, 1 H), 7.64 - 7.53 (m, 3H), 3.50 (br d, J = 11 .9 Hz, 2H), 2.50-2.47 (m, 2H), 2.35 - 2.24 (m, 1 H), 1 .93 - 1 .83 (m, 2H), 1.61 - 1 .46 (m, 2H).

[0505] Synthesis of compound (25)

[0506] A mixture of compound 2A (50 mg, 278.89 μmol, 1 eq), compound 6 (96.15 mg, 334.67 μmol, 1.2 eq), DIEA (108.13 mg, 836.67 μmol, 145.73 μL, 3 eq), HOBt (45.22 mg, 334.67 μmol, 1.2 eq) and EDCI (64.16 mg, 334.67 μmol, 1 .2 eq) in DMF (1 mL) was stirred at 20 °C for 16 h. The mixture was purified by prep-HPLC (column: YMC-Actus Triart C18 150*30 mm*7um; mobile phase: [water (FA) -ACN]; B%: 55%- 85%, 10 min). Compound (25)_peak1 (52.26 mg, 115.34 μmol, 41 .36% yield, 99% purity) was obtained as an off-white solid.1H NMR: (400 MHz, CDCI3) δ = 7.61 - 7.52 (m, 2H), 7.52 - 7.46 (m, 1 H), 7.35 - 7.29 (m, 1 H), 7.11 (d, J = 5.1 Hz, 1 H), 6.89 (d, J = 5.1 Hz, 1 H), 5.10 - 4.82 (m, 1 H), 4.43 - 3.93 (m, 1 H), 3.90 - 3.09 (m, 3H), 2.94 - 2.73 (m, 2H), 2.67 - 2.38 (m, 3H), 2.18 - 1.68 (m, 4H), 1.25 - 1.02 (m, 1 H), 0.74 - 0.23 (m, 4H); LCMS: RT =0.581 min, m / z =449.2 (M+H)+; SFC: RT =1 .609 min.

[0507] Synthesis of compound (25)_peak2

[0508] A mixture of compound 2B (50 mg, 278.89 μmol, 1 eq), compound 6 (96.15 mg, 334.67 μmol, 1 .2 eq), DIEA (108.13 mg, 836.67 μmol, 145.73 μL, 3 eq), HOBt (45.22 mg, 334.67 μmol, 1.2 eq) and EDCI (64.16 mg, 334.67 μmol, 1.2 eq) in DMF (1 mL) was stirred at 20 °C for 16 h. the mixture was purified by prep-HPLC (column: YMC-Actus Triart C18 150*30 mm*7um; mobile phase: [water (FA) -ACN]; B%: 55%- 85%, 10 min. Compound (25)_peak2 (34.52 mg, 76.96 μmol, 27.59% yield, 100% purity) was obtained as an off-white solid.1H NMR: (400 MHz, CDCI3) δ = 7.61 - 7.46 (m, 3H), 7.35 - 7.29 (m, 1 H), 7.11 (d, J = 5.1 Hz, 1 H), 6.89 (d, J = 5.1 Hz, 1 H), 5.05 (d, J = 8.8 Hz, 1 H), 4.41 - 3.90 (m, 1 H), 3.90 - 3.09 (m, 3H), 2.93 - 2.71 (m, 2H), 2.67 - 2.39 (m, 3H), 2.18 - 1.67 (m, 4H), 1.24 - 1.03 (m, 1 H), 0.74 - 0.20 (m, 4H); LCMS: RT =0.583 min, m / z =449.3 (M+H)+; SFC: RT =0.675 min

[0509] EXAMPLE 1.26: SYNTHESIS OF (26)

[0510] Ethyl acetate = 3: 1 ; Rf = 0.26)) to give compound 3 (1 g, 2.85 mmol, 77.54% yield) as white solid. Synthesis of compound 4

[0511] To the mixture of compound 3 (550 mg, 1.57 mmol, 1 eq) in toluene (8 mL) was added Pd(OAc)2 (70.31 mg, 313.20 μmol, 0.2 eq), PCys (43.91 mg, 156.60 μmol, 50.77 μL, 0.1 eq), cyclopropylboronic acid (672.57 mg, 7.83 mmol, 5 eq), K3PO4 (997.21 mg, 4.70 mmol, 3 eq) and H2O (0.8 mL). The mixture was degassed and purged with N2 for 3 times, and then the mixture was stirred at 100 °C for 2 h under N2 atmosphere. The mixture was filtered and the filtrate was concentrated under vacuum to give a residue. The residue was purified by flash silica gel chromatography (ISCO®; 12 g SepaFlash® Silica Flash Column, Eluent of 20-30% Ethyl acetate / Petroleum ether gradient @ 40 mL / min). Compound 4 (440 mg, 1.32 mmol, 84.55% yield, 94% purity) was obtained as a yellow oil. LCMS: RT = 0.566 min, m / z = 313.3 (M+H)+.

[0512] Synthesis of compound 5

[0513] To a solution of compound 4 (440 mg, 1.41 mmol, 1 eq) in EtOH (10 mL) was added Pd(OH)2 / C (395.61 mg, 10% purity) under N2. The suspension was degassed under vacuum and purged with H2 several times. The mixture was stirred under H2 (50 psi) at 60 °C for 14 h. The mixture was filtered and the filtrate was concentrated under vacuum to give a residue. Compound 5 (400 mg, 1.27 mmol, 90.33% yield) was obtained as a yellow oil. LCMS: RT = 0.553min, m / z = 315.1 (M+H)+.

[0514] Synthesis of compound 6

[0515] To a solution of compound 5 (320 mg, 1 .02 mmol, 1 eq) in EtOH (10 mL) and H2O (2 mL) was added KOH (285.52 mg, 5.09 mmol, 5 eq) at 25 °C. The mixture was stirred at 90 °C for 14 h. The mixture was filtered and the filtrate was concentrated under vacuum to give a residue. The residue was purified by flash silica gel chromatography (ISCO®; 12 g SepaFlash® Silica Flash Column, Eluent of 70-100% Ethyl acetate / Petroleum ether gradient @ 40 mL / min). Compound 6 (110 mg, 420.49 μmol, 41.31 % yield, 98% purity) was obtained as a yellow oil. LCMS: RT = 0.366 min, m / z = 257.1 (M+H)+.

[0516] Synthesis of compound (26)_peak1& (26)_peak2

[0517] To a solution of compound 6 (52 mg, 202.84 μmol, 1 eq) in DCM (2 mL) was added compound 7 (54.63 mg, 263.69 μmol, 1 .3 eq), HOBt (35.63 mg, 263.69 μmol, 1 .3 eq), EDCI (50.55 mg, 263.69 μmol, 1.3 eq) and DIEA (78.65 mg, 608.51 μmol, 105.99 μL, 3 eq) at 20 °C. The mixture was stirred at 20 °C for 16 h. EtOAc (30 mL) and water (30 mL) were added and Layers were separated. The aqueous phase was extracted with EtOAc (30 mL x 2). Combined extracts were washed with brine (30 mL), dried over Na2SO4, filtered, and concentrated under vacuum to give a residue. The residue was purified by prep- TLC (SiO2, PE: EtOAc = 1 : 1) to give desired compound (26) (80 mg, purity 93%) as a white solid, which was further separated by SFC (RT= 1 .495 min and 1 .721 min, column: DAICEL CHIRALPAK AS (250 mm*30 mm, 10um) ; mobile phase: [CO2-EtOH (0.1 %NH3H2O) ]; B%: 35%, isocratic elution mode). Compound (26)_peak1 (48.61 mg, 102.56 μmol, 50.56% yield, 94% purity) was obtained as a white solid.1H NMR: (400 MHz, CDCI3) δ = 8.93 (s, 1 H), 7.53 - 7.10 (m, 5H), 6.73 (br s, 1 H), 4.12 (br d, J = 14.1 Hz, 1 H), 3.84 (q, J = 5.6 Hz, 2H), 3.37 - 2.73 (m, 10H), 2.14 - 1.99 (m, 1 H), 1.24 - 1.10 (m, 2H), 0.87 (br t, J = 4.8 Hz, 2H); LCMS: RT = 0.517 min, m / z = 446.2 (M+H)+; SFC: RT = 1 .480 min.

[0518] Compound (26)_peak2 (34.92 mg, 77.60 μmol, 38.26% yield, 99% purity) was obtained as a white solid.1H NMR: (400 MHz, CDCI3) δ = 8.73 (s, 1 H), 7.26 - 6.90 (m, 5H), 6.50 (br s, 1 H), 3.91 (br d, J = 14.3 Hz, 1 H), 3.63 (q, J = 5.5 Hz, 2H), 3.17 - 2.56 (m, 10H), 1.94 - 1.85 (m, 1 H), 1.03 - 0.89 (m, 2H), 0.66 (br t, J = 4.6 Hz, 2H); LCMS: RT = 0.518 min, m / z = 446.2 (M+H)+; SFC: RT = 1 .708 min.

[0519] EXAMPLE 1.27: SYNTHESIS OF (27)

[0520] Synthesis of compound 3

[0521] A mixture of compound 1 (230 mg, 1.50 mmol, 1 eq,HCI), compound 2 (347.37 mg, 2.55 mmol, 1.70 eq), DIEA (483.80 mg, 3.74 mmol, 652.02 μL, 2.5 eq), HOBt (242.79 mg, 1 .80 mmol, 1.2 eq) and EDCI (344.45 mg, 1 .80 mmol, 1 .2 eq) in DMF (3 mL) was stireed at 25 °C for 16 h. The reaction mixture was purified by prep-HPLC (column: YMC-Actus Triart C18 150*30 mm*7um; mobile phase: [water (FA) -ACN]; B%: 20%- 50%, 10 min). Compound 3 (155 mg, 658.94 μmol, 44.01 % yield) was obtained as colorless oil.1H NMR: (400 MHz, CDCI3) δ = 6.06 (br s, 1 H), 3.72 (s, 3H), 3.55 (ddd, J = 4.3, 6.7, 13.7 Hz, 1 H), 3.32 (ddd, J = 5.8, 8.3, 13.8 Hz, 1 H), 2.96 - 2.79 (m, 2H), 2.78 - 2.63 (m, 4H), 1.19 (d, J = 7.3 Hz, 3H).

[0522] Synthesis of compound4

[0523] A mixture of compound 3 (50 mg, 212.56 μmol, 1 eq) and LiOH (10.18 mg, 425.12 μmol, 2 eq) in H2O (0.1 mL) and THF (1 mL) was stirred at 25 °C for 16 h. The reaction mixture was diluted with H2O (5 mL) and the pH of the mixture was adjusted to about 5 with HCI (1 M). The resulting mixture was extracted with EtOAc (3 mL*2). The combined organic phase was concentrated at reduced pressure to give compound 4 (60 mg, crude) as white solid. Synthesis of (27)

[0524] A mixture of compound 4 (40.00 mg, 161.73 μmol, 1 eq), compound 5 (60 mg, 271.25 μmol, 1.68 eq), DIEA (52.25 mg, 404.32 μmol, 70.42 μL, 2.5 eq), HOBt (26.22 mg, 194.07 μmol, 1 .2 eq) and EDCI (37.20 mg, 194.07 μmol, 1.2 eq) in DMF (1 mL) was stirred at 25 °C for 16 h. The mixture was diluted with H2O (30 mL) and extracted with EtOAc (30 mL). The organic layers was concentrated at reduced pressure to give a residue. The residue was purified by prep-HPLC (column: Phenomenex luna C18 150*25 mm* 10um; mobile phase: [water (FA) -ACN]; B%: 50%- 80%, 10 min). (27) (3.77 mg, 7.70 μmol, 4.76% yield, 92% purity) was obtained as a white solid.1H NMR: (400 MHz, CD3CN) δ = 7.30 - 7.22 (m, 2H), 7.10 - 7.01 (m, 2H), 6.71 - 6.62 (m, 1 H), 6.39 (br s, 1 H), 4.11 - 3.95 (m, 1 H), 3.36 - 2.97 (m, 4H), 2.90 - 2.42 (m, 7H), 2.39 (s, 3H), 1 .08 - 0.95 (m, 3H); LCMS: RT = 0.590 min, m / z = 451 ,2(M+H)+.

[0525] EXAMPLE 1.28: SYNTHESIS OF (28)

[0526] (400 MHz, CDCI3) δ = 9.16 (s, 1 H), 8.64 (d, J = 5.6 Hz, 1 H), 8.16 (s, 1 H), 7.58 (d, J = 5.0 Hz, 1 H).

[0527] Synthesis of compound 3

[0528] To a solution of compound 2 (380 mg, 3.16 mmol, 1 eq) in THF (4 mL) was added methyl carbonochloridate (358.76 mg, 3.80 mmol, 293.35 μL, 1 .2 eq) at 0 °C. The mixture was stirred at 25 °C for 1 h. The iminium salt solution wascooled to 0 °C, and the solution of bromo-(4-fluorophenyl) magnesium (2 M, 3.16 mL, 2 eq) was added. The reaction allowed to warm to 25 °C and stirred for 14 h. The reaction mixture was poured into saturated NH4CI aqueous solution (10 mL) and the resulting mixture was extracted with EtOAc (15 mL*3). The combined organic layer was washed with brine (15 mL), dried over Na2SO4, filtered and concentrated at reduced pressure to give a residue. The residue was purified by flash silica gel chromatography (ISCO®; 12 g SepaFlash® Silica Flash Column, Eluent of 0-20% Ethyl acetate / Petroleum ether gradient @ 50 mL / min). Compound 3 (650 mg, 2.37 mmol, 74.91 % yield) was obtained as a yellow oil. LCMS: RT = 0.525 min, m / z = 275.1 (M+H)+.

[0529] Synthesis of compound 4

[0530] To a solution of compound 3 (650 mg, 2.37 mmol, 1 eq) in EtOH (15 mL) was added Pd / C (300 mg, 10% purity) under N2. The suspension was degassed under vacuum and purged with H2 several times. The mixture was stirred under H2 (30 psi) at 30 °C for 14 h. The mixture was filtered and the filtrate was concentrated under vacuum to give a residue. Compound 4 (650 mg, 2.35 mmol, 99.27% yield) was obtained as a yellow oil. LCMS: RT = 0.492 min, m / z = 277.1 (M+H)+.

[0531] Synthesis of compound 5

[0532] To a solution of compound 4 (650 mg, 2.35 mmol, 1 eq) in EtOH (15 mL) and H2O (3 mL) was added KOH (1 .32 g, 23.53 mmol, 10 eq) at 25 °C. The mixture was stirred at 90 °C for 14 h. The mixture was filtered and the filtrate was concentrated under vacuum to give a residue. The residue was purified by flash silica gel chromatography (ISCO®; 12 g SepaFlash® Silica Flash Column, Eluent of 60-100% Ethyl acetate / Petroleum ether gradient @ 35 mL / min). Compound 5 (180 mg, 593.88 μmol, 25.24% yield, 72% purity) was obtained as a yellow oil. LCMS: RT = 0.272 min, m / z = 219.2 (M+H)+.

[0533] Synthesis of compound (28)_peak1& (28)_peak2

[0534] To a solution of compound 5 (50 mg, 229.12 μmol, 1 eq) in pyridine (1 mL) and DCM (1 mL) was added EDCI (65.88 mg, 343.68 μmol, 1 .5 eq) and compound 6 (50.68 mg, 229.12 μmol, 1 eq). The mixture was stirred at 20 °C for 16 h. EtOAc (10 mL) and water (10 mL) were added and Layers were separated. The aqueous phase was extracted with EtOAc (10 mL x 2). Combined extracts were washed with brine (10 mL), dried over Na2SO4, filtered, and concentrated under vacuum to give a residue. The residue was purified by prep-HPLC (column: C18 150x30 mm; mobile phase: [water (FA) -ACN]; gradient: 38%-68% B over 7 min) to give desired compound (28) (60 mg, purity 100%) as a white solid, which was further separated by SFC (RT= 1 .921 min and 2.298 min , column: DAICEL CHIRALPAK IG (250 mm*30 mm, 10um) ; mobile phase: [CO2-EtOH (0.1 %NH3H2O) ]; B%: 43%, isocratic elution mode).

[0535] Compound (28)_peak1 (24.01 mg, 56.98 μmol, 24.87% yield, 100% purity) was obtained as an off- white solid.1H NMR: (400 MHz, CDCI3) δ = 7.93 - 7.84 (m, 1 H), 7.32 - 121 (m, 1 H), 7.27-7.25 (m, 1 H), 7.11 - 6.96 (m, 2H), 6.80 (s, 1 H), 6.41 - 6.16 (m, 1 H), 3.99 (dd, J = 5.4, 14.4 Hz, 1 H), 3.60 - 3.25 (m, 3H), 3.21 - 3.05 (m, 1 H), 3.01 - 2.55 (m, 7H), 1 .20 - 1 .06 (m, 3H); LCMS: RT = 0.492 min, m / z = 422.2 (M+H)+; SFC: RT = 1.926 min.

[0536] Compound (28)_peak2 (23.32 mg, 55.34 μmol, 24.15% yield, 100% purity) was obtained as an off- white solid.1H NMR: (400 MHz, CDCI3) δ = 7.93 - 7.81 (m, 1 H), 7.45 - 121 (m, 2H), 7.13 - 6.94 (m, 2H), 6.80 (s, 1 H), 6.30 - 6.03 (m, 1 H), 4.11 (dd, J = 5.1 , 14.5 Hz, 1 H), 3.56 - 3.27 (m, 3H), 3.25 - 3.10 (m, 1 H), 3.05 - 2.45 (m, 7H), 1.22 - 1.07 (m, 3H); LCMS: RT = 0.483 min, m / z = 422 (M+H)+; SFC: RT =

[0537] 2.298 min.

[0538] EXAMPLE 1.29: SYNTHESIS OF (29)

[0539] Synthesis of compound 2

[0540] A mixture of compound 1 (200 mg, 850.24 μmol, 1 eq) and Lawesson reagent (206.34 mg, 510.15 μmol, 0.6 eq) in THF (2 mL) was stirred at 25 °C for 2 h. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, Petroleum ether / Ethyl acetate = 1 / 0 to 3 / 1 , TLC (SiO2, Petroleum ether: Ethyl acetate = 5: 1 , Rf = 0.40)). Compound 2 (210 mg, 835.68 μmol, 98.29% yield) was obtained as a colorless oil.

[0541] Synthesis of compound 3

[0542] A mixture of compound 2 (210 mg, 835.68 μmol, 1 eq) and LiOH»H2O (70.14 mg, 1.67 mmol, 2 eq) in THF (2 mL) and H2O (0.2 mL) was stirred at 25 °C for 2 h. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was diluted with H2O (5 mL) and acidified to pH = 5 - 6 with 1 M HCI aqueous solution, the resulting mixture was extracted with EtOAc (10 mL * 3). The combined organic layers were dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. Compound 3 (196 mg, 826.08 μmol, 98.85% yield) was obtained as a colorless oil.

[0543] Synthesis of (29)

[0544] A mixture of compound 4 (20 mg, 90.79 μmol, 1 eq), compound 3 (32.31 mg, 136.18 μmol, 1 .5 eq) and EDCI (26.11 mg, 136.18 μmol, 1.5 eq) in pyridine (0.5 mL) was stirred at 25-40 °C for 2 h. To the reaction mixture was added water (5 mL), the resulting mixture was extracted with ethyl acetate (2 mL * 4), and the combined organic phase was dried with anhydrous Na2SO4, filtered and the filtrate was concentrated in vacuum to give a residue. The residue was purified by prep-HPLC (column: YMC-Actus Triart C18 150*30 mm*7um; mobile phase: [water (FA) -ACN]; gradient: 35%- 65% B over 10 min). (29) (8.11 mg, 18.45 μmol, 20.32% yield, 100% purity) was obtained as a yellow gum.1H NMR: (400 MHz, CDCI3) δ = 8.93 - 8.38 (m, 2H), 7.30 (d, J = 2.0 Hz, 1 H), 6.96 - 6.04 (m, 2H), 5.43 - 4.12 (m, 2H), 3.92 - 3.83 (m, 3H), 3.69 - 3.45 (m, 2H), 3.26 - 2.79 (m, 6H), 2.77 - 2.46 (m, 2H), 1 .26 - 1 .21 (m, 3H); LCMS:

[0545] RT = 0.500min, m / z =440.1 (M+H)+.

[0546] EXAMPLE 1.30: SYNTHESIS OF (30)

[0547] Synthesis of compound 3

[0548] To the mixture of compound 1 (367.87 mg, 2.57 mmol, 1 eq) in DCM (5 mL) was added Py (609.68 mg, 7.71 mmol, 622.12 μL, 3 eq) and compound 2 (500 mg, 2.57 mmol, 1 eq) at 0 °C. The mixture was stirred at 20 °C for 16 h. The mixture was concentrated at reduced pressure to give a residue. Compound 3 (1 .19 g, crude) was obtained as a yellow solid.

[0549] Synthesis of compound 4

[0550] To the mixture of compound 3 (1.1 g, 3.65 mmol, 1 eq) in THF (11 mL) was added MeOH (5 mL), H2O (3 mL) and LiOH (174.84 mg, 7.30 mmol, 2 eq). The mixture was stirred at 25 °C for 2 h. To the mixture was added aqueous HCI solution and the pH was adjusted to 4~5. To the mixture was added water (40 mL) and the mixture was extracted with EtOAc (25 mL x 3), the combined organic phase was dried over Na2SO4, filtered and the filtrate was concentrated under reduced pressure to give a residue. Compound 4 (500 mg, crude) was obtained as a yellow solid.1H NMR: (400 MHz, CDCI3) 6 = 7.83 - 7.74 (m, 2H), 7.22 (t, J = 8.6 Hz, 2H), 3.64 (td, J = 3.9, 11 .9 Hz, 2H), 2.52 (dt, J = 2.8, 11 .3 Hz, 2H), 2.33 (ft, J = 4.0, 10.5 Hz, 1 H), 2.07 - 1 .96 (m, 2H), 1.91 - 1 .77 (m, 2H).

[0551] Synthesis of compound (30)

[0552] To the mixture of compound 4 (50 mg, 278.89 μmol, 1 eq) in DMF (1 mL) was added HOBt (41 .45 mg, 306.78 μmol, 1.1 eq) and EDCI (58.81 mg, 306.78 μmol, 1.1 eq) and DIEA (108.13 mg, 836.67 μmol, 145.73 μL, 3 eq) and compound 5 (80.13 mg, 278.89 μmol, 1 eq). The mixture was stirred at 25 °C for 16 h. The mixture was purified by prep-HPLC (column: YMC-Actus Triart C18 150*30 mm*7um; mobile phase: [water (FA) -ACN]; B%: 55%- 85%, 10 min). Compound (30) (34.89 mg, 75.45 μmol, 27.05% yield, 97% purity) was obtained as a white solid.1H NMR: (400 MHz, CDCI3) δ = 7.81 (br dd, J = 5.2, 8.6 Hz, 2H), 7.26 - 7.20 (m, 2H), 7.11 (d, J = 5.3 Hz, 1 H), 6.89 (d, J = 5.3 Hz, 1 H), 5.20 - 4.77 (m, 1 H), 4.38 - 3.54 (m, 4H), 3.19 - 2.79 (m, 2H), 2.65 - 2.37 (m, 3H), 2.08 - 1.70 (m, 4H), 1.25 - 1.01 (m, 1 H), 0.71 - 0.49 (m, 3H), 0.46 - 0.24 (m, 1 H); LCMS: RT =0.582 min, m / z = 449.1 (M+H)+; SFC: RT =1 .674 min.

[0553] EXAMPLE 1.31: SYNTHESIS OF (31)

[0554] Synthesis of compound 3

[0555] A mixture of compound 2 (1 g, 8.06 mmol, 847.46 μL, 1 eq) and compound 1 (1.02 g, 8.06 mmol, 940.31 μL, 1 eq) in Tol. (15 mL) was stirred at 110 °C for 16 h. Then the mixture was concentrated, to the residue was added TFA (10 mL), the mixture was stirred at 20 °C for 16 h. The mixture was concentrated, the residue was purified by reversed-phase HPLC (0.1 % FA condition) , the eluent was concentrated and then freeze dried. Compound 3 (350 mg, 1 .50 mmol, 18.62% yield, 100% purity) was obtained as a yellow solid.1H NMR: (400 MHz, CHLOROFORM-d) δ = 8.24 - 8.13 (m, 1 H), 7.30 (dd, J = 5.3, 8.4 Hz, 2H), 7.15 (d, J = 5.3 Hz, 1 H), 7.05 (t, J = 8.5 Hz, 2H), 6.43 (d, J = 5.1 Hz, 1 H), 5.35 (s, 1 H), 3.30 - 3.10 (m, 3H), 3.05 - 2.94 (m, 1 H); LCMS: RT =0.951 min, m / z =234.0 (M+H)+.

[0556] Synthesis of (31)

[0557] A mixture of compound 4 (39.69 mg, 214.31 μmol, 1 eq) , compound 3 (50 mg, 214.31 μmol, 1 eq) , DIEA (83.10 mg, 642.94 μmol, 111 .99 μL, 3 eq) , HOBt (34.75 mg, 257.18 μmol, 1.2 eq) and EDCI (49.30 mg, 257.18 μmol, 1.2 eq) in DMF (1 mL) was stirred at 20 °C for 16 h. The mixture was concentrated, the residue was purified by reversed-phase HPLC (column : Phenomenex luna C18 150*25 mm* 10um;mobile phase : [water (FA) -ACN] ;B% : 46%- 76%, 2 min), the eluent was concentrated and then freeze dried. Compound (31) (28.5 mg, 69.02 μmol, 32.21 % yield, 97% purity) was obtained as off white solid.1H NMR: (400 MHz, DMSO-d6) δ = 7.79 - 7.70 (m, 1 H), 7.40 (d, J = 5.1 Hz, 1 H), 7.32 - 7.11 (m, 4H), 6.81 (d, J = 5.1 Hz, 1 H), 6.69 (s, 1 H), 3.98 (br dd, J = 3.3, 13.4 Hz, 1 H), 3.28 (br s, 1 H), 3.19 - 3.08 (m, 1 H), 3.02 - 2.71 (m, 3H), 2.65 - 2.52 (m, 3H), 1 .68 - 1 .40 (m, 8H); LCMS: RT =0.793 min, m / z =401 .0 (M+H)+.

[0558] EXAMPLE 1.32: SYNTHESIS OF (32)

[0559] A mixture of compound 6A, 2-(4 -fluorophenyl) cyclopropanecarboxylic acid (31.46 mg, 174.59 μmol, 1 eq), DIEA (45.13 mg, 349.17 μmol, 60.82 μL, 2 eq), HOBt (28.31 mg, 209.50 μmol, 1 .2 eq) and EDCI (40.16 mg, 209.50 μmol, 1 .2 eq) in DMF (1 mL) was stirred at 20 °C for 16 h. The reaction mixture was diluted with Ethyl acetate (50 mL). The organic layer was washed with brine (20 mL*2), dried over Na2SO4, filtered and concentrated at reduced pressure to give a residue. The residue was purified by prep-HPLC (column: Unisil 3 - 100 C18 Ultra 150*50 mm*3 um;mobile phase : [water (FA) -ACN] ;B% : 51 %- 81 %, 7 min) to give (32) (22.36 mg, 46.36 μmol, 26.55% yield, 93% purity) as white solid.1H NMR: (400 MHz, CD3OD) δ = 7.30 - 7.20 (m, 6H), 7.12 - 7.06 (m, 2H), 7.01 - 6.95 (m, 2H), 6.84 - 6.78 (m, 1 H), 6.71 (t, J = 4.3 Hz, 1 H), 4.04 (br dd, J = 4.8, 14.1 Hz, 1 H), 3.52 (td, J = 6.6, 13.2 Hz, 2H), 3.40 - 3.34 (m, 1 H), 3.40 - 3.32 (m, 1 H), 3.03 - 2.90 (m, 2H), 2.82 - 2.63 (m, 2H), 2.38 - 2.29 (m, 1 H), 1.81 - 1 .72 (m, 1 H), 1.48 - 1.40 (m, 1 H), 1.21 - 1.15 (m, 1 H); LCMS: RT = 0.612 min, m / z = 449.2 (M+H)+.

[0560] EXAMPLE 1.33: SYNTHESIS OF (33)

[0561] A mixture of compound 6A (50.00 mg, 174.59 μmol, 1 eq), 3,3-difluorocyclobutane- carboxylic acid (23.76 mg, 174.59 μmol, 1 eq), DIEA (45.13 mg, 349.18 μmol, 60.82 μL, 2 eq), HOBt (28.31 mg, 209.51 μmol, 1 .2 eq) and EDCI (40.16 mg, 209.51 μmol, 1 .2 eq) in DMF (1 mL) , was stirred at 20 °C for 16 h. The reaction mixture was diluted with Ethyl acetate (50 mL). The organic layer was washed with brine (20 mL*2), dried over Na2SO4, filtered and concentrated at reduced pressure to give a residue. The residue was purified by prep-HPLC (column: Unisil 3 - 100 C18 Ultra 150*50 mm*3 um;mobile phase : [water (FA) -ACN] ;B% : 42%- 72%, 7 min) to give (33) (20.07 mg, 49.12 μmol, 28.14% yield, 99% purity) as white solid.1H NMR: (400 MHz, CD3OD) δ = 7.37 - 7.21 (m, 6H), 6.80 (s, 1 H), 6.71 (d, J = 5.3 Hz, 1 H), 4.03 (br dd, J = 5.0, 14.1 Hz, 1 H), 3.50 (br t, J = 6.8 Hz, 2H), 3.39 - 3.34 (m, 1 H), 3.07 - 2.82 (m, 3H), 2.77 - 2.55 (m, 6H); LCMS: RT = 0.580 min, m / z = 405.2 (M+H)+.

[0562] EXAMPLE 1.34: SYNTHESIS OF (34)

[0563]

[0564] Synthesis of compound 2

[0565] A mixture of compound 1 (400 mg, 2.79 mmol, 1 eq), methylsulfonyl methanesulfonate (973.12 mg, 5.59 mmol, 2 eq) , DIEA (1 .44 g, 1 1.17 mmol, 1 .95 mL, 4 eq) in DCM (5 mL) was stirred at 0 °C for 2 h. The mixture was concentrated under reduced pressure to give a residue. The residue was poured into water (50 mL) and stirred for 5 min. The aqueous phase was extracted with EtOAc (50 mL * 2). The combined organic phase was washed with brine (50 mL), dried with anhydrous Na2SO4, filtered and concentrated in vacuum to give compound 2 (560 mg, crude) as colorless oil.1H NMR: 5 = 7.40 (s, 1 H), 4.38 (t, J = 6.4 Hz, 2H), 3.23 (t, J = 6.4 Hz, 2H), 2.99 (s, 3H), 2.67 (s, 3H);

[0566] Synthesis of compound 3

[0567] To the mixture of compound 2 (560 mg, 2.53 mmol, 1 eq) in DMF (5 mL) was added NaN3 (246.77 mg, 3.80 mmol, 1 .5 eq), then the mixture was stirred at 80 °C for 2 h. To the mixture was added NaOH (2 M) to about pH>10, the resulting mixture was extracted with EtOAc (30 mL * 3). The combined organic layers were dried over Na2SO4, filtered and concentrated under reduced pressure to give compound 3 (630 mg, crude) as yellow oil.1H NMR: (400 MHz, CDCI3) δ = 7.36 (s, 1 H), 3.50 (t, J = 6.7 Hz, 2H), 3.02 (t, J = 6.8 Hz, 2H), 2.65 (s, 3H);

[0568] Synthesis of compound 4

[0569] A mixture of compound 3 (550 mg, 3.27 mmol, 1 eq), PPh3 (1 .72 g, 6.54 mmol, 2 eq), H2O (294.51 mg, 16.35 mmol, 294.51 uL, 5 eq) in THF (5 mL) was degassed and purged with N2 for 3 times, and then the mixture was stirred at 20 °C for 16 h under N2 atmosphere. The mixture was concentrated at reduced pressure to give a residue. To the residue was added 1 N HCI (20 mL), the resulting mixture was washed with EA (30 mL * 4). The water phase was lyophilized to give compound 4 (450 mg, crude, HCI) as white solid.1H NMR: (400 MHz, DMSO) δ = 8.34 (br s, 3H), 7.76 (s, 1 H), 3.20 - 3.13 (m, 2H), 3.08 - 3.01 (m, 2H), 2.74 (s, 3H);

[0570] Synthesis of compound 6 A mixture of compound 4 (400 mg, 2.81 mmol, 1 eq), compound 5 (470.71 mg, 3.09 mmol, 1.1 eq), HOBt (418.04 mg, 3.09 mmol, 1.1 eq), EDCI (593.08 mg, 3.09 mmol, 1.1 eq) and DIEA (1.45 g, 11 .25 mmol, 1 .96 mL, 4 eq) in DMF (5 mL) was degassed and purged with N2 for 3 times, and then the mixture was stirred at 20 °C for 16 h under N2 atmosphere. The mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (silica gel, Petroleum ether / Ethyl acetate=1 :1 to 0:1 , TLC : PE:EA=0:1 , Rf=0.67) to give compound 6 (400 mg, 1 .40 mmol, 49.92% yield, 97% purity) as white solid. LCMS: RT = 0.460 min, m / z = 277.1 (M+H)+.

[0571] Synthesis of compound 7

[0572] A mixture of compound 6 (50 mg, 180.93 umol, 1 eq) in POCh (3 mL) was stirred at 110 °C for 16 h. The mixture was cooled to room temperature and concentrated under reduced pressure to give a residue. The residue was added into water (50 mL) slowly and stirred for 5 min. The aqueous phase was extracted with EtOAc (50 mL * 2). The combined organic phase was washed with brine (50 mL), dried with anhydrous Na2SO4, filtered and concentrated in vacuum to give compound 7 (70 mg, crude) as white solid. LCMS: RT = 0.385 min, m / z = 259.2 (M+H)+.

[0573] Synthesis of compound 8

[0574] To a mixture of compound 7 (70 mg, 270.96 umol, 1 eq) in MeOH (2 mL) was added NaBFL (25.63 mg, 677.41 umol, 2.5 eq) at 0 °C. The mixture was stirred at 20 °C for 2 h. The mixture was poured into sat. NH4CI (5 mL) and the resulting mixture was extracted with EtOAc (10 mL * 2). The combined organic layers were washed with brine (10 mL x 2), dried over Na2SO4, filtered and concentrated under reduced pressure to give compound 8 (50 mg, crude) as white solid.

[0575] Synthesis of compound (34)

[0576] A mixture of compound 8 (50 mg, 192.05 umol, 1 eq), compound 9 (39.79 mg, 192.05 umol, 1 eq), HOBt (31.14 mg, 230.46 umol, 1.2 eq), EDCI (44.18 mg, 230.46 umol, 1.2 eq) and DIEA (99.28 mg, 768.18 umol, 133.80 uL, 4 eq) in DMF (2 mL) was stirred at 20 °C for 16 h under N2 atmosphere. The mixture was purified by prep-HPLC (column: Waters Xbridge 150*25mm* 5um;mobile phase: [water( NH4HCO3)-ACN];B%: 32%-62%,9min) to give (34) (13.93 mg, 30.99 umol, 16.14% yield, 100% purity) as white solid.1H NMR: (400 MHz, CDCI3) δ = 7.30 - 7.28 (m, 1 H), 7.26 - 7.20 (m, 1 H), 6.94 - 6.80 (m, 3H), 6.49 (br d, J = 6.4 Hz, 1 H), 4.05 - 4.04 (m, 1 H), 3.85 (br dd, J = 5.1 , 14.0 Hz, 1 H), 3.81 - 3.77 (m, 3H), 3.62 (q, J = 5.8 Hz, 2H), 3.41 (ddd, J = 4.8, 1 1 .5, 14.3 Hz, 1 H), 3.08 - 2.93 (m, 1 H), 2.93 - 2.84 (m, 2H), 2.84 - 2.74 (m, 2H), 2.74 - 2.69 (m, 2H), 2.69 - 2.67 (m, 3H), 2.66 - 2.61 (m, 1 H), 2.61 - 2.49 (m, 1 H); LCMS: RT = 0.496 min, m / z = 449.9 (M+H)+.

[0577] Synthesis of (34)_peak1 & (34)_peak2

[0578] (34) was purified by SFC (RT = 0.981 min and RT = 2.389 min; DAICEL CHIRALPAK AS (250 mm*30 mm, 10um) ; mobile phase: [CO2-MeOH (0.1 %NH3H2O) ]; B%: 75%-75%, 3.44 min) to give (34)_peak1 (4.7 mg, 10.46 μmol, 36.15% yield, 100% purity) as off-white solid.1H NMR: (400 MHz, CDCI3) 6 = 7.26

[0579] - 7.21 (m, 1 H), 6.94 - 6.90 (m, 1 H), 6.85 - 6.81 (m, 2H), 6.50 (br d, J = 5.4 Hz, 1 H), 3.88 - 3.82 (m, 1 H), 3.80 - 3.77 (m, 3H), 3.62 (q, J = 5.5 Hz, 2H), 3.52 - 3.33 (m, 1 H), 3.09 - 2.94 (m, 1 H), 2.93 - 2.85 (m, 2H), 2.85 - 2.75 (m, 2H), 2.72 (br s, 2H), 2.69 - 2.68 (m, 3H), 2.65 - 2.61 (m, 1 H), 2.61 - 2.50 (m, 1 H); LCMS: RT = 0.502 min, m / z = 450.1 (M+H)+; SFC: RT = 0.997 min.

[0580] (34)_peak2 (4.07 mg, 9.05 μmol, 31.31 % yield, 100% purity) as off-white solid.1H NMR: (400 MHz, CDCI3) δ = 7.26 - 7.21 (m, 1 H), 6.94 - 6.90 (m, 1 H), 6.85 - 6.82 (m, 2H), 6.49 (br d, J = 6.6 Hz, 1 H), 3.85 (br dd, J = 4.6, 14.1 Hz, 1 H), 3.80 - 3.77 (m, 3H), 3.62 (q, J = 5.8 Hz, 2H), 3.50 - 3.34 (m, 1 H), 3.08

[0581] - 2.93 (m, 1 H), 2.93 - 2.84 (m, 2H), 2.84 - 2.75 (m, 2H), 2.71 (br d, J = 5.9 Hz, 2H), 2.69 - 2.67 (m, 3H), 2.66 - 2.61 (m, 1 H), 2.60 - 2.51 (m, 1 H); LCMS: RT = 0.492 min, m / z = 450.1 (M+H)+; SFC: RT = 2.390 min.

[0582] EXAMPLE 1.35: SYNTHESIS OF (35)

[0583] To a solution of compound 4A (50 mg, 174.59 μmol, 1 eq) in DMF (1 mL) was added HOBt (28.31 mg, 209.50 μmol, 1.2 eq) and DIEA (67.69 mg, 523.76 μmol, 91.23 μL, 3 eq), (cis) 4- (trifluoromethyl)cyclohexanecarboxylic acid (34.25 mg, 174.59 μmol, 1 eq), EDCI (40.16 mg, 209.50 μmol, 1 .2 eq). The mixture was stirred at 25 °C for 16 h. The mixture was purified by prep-HPLC(column: Phenomenex luna C18 150*25 mm* 10um; mobile phase: [water (FA) -ACN]; B%: 52%- 82%, 10 min), the eluent was concentrated and then freeze dried. (35) (32.79 mg, 69.88 μmol, 40.03% yield, 99% purity) was obtained as a white solid.1H NMR: (400 MHz, CDCI3) δ = 7.38 - 7.28 (m, 3H), 7.26 - 7.19 (m, 2H), 7.17 (d, J = 5.0 Hz, 1 H), 6.88 (s, 1 H), 6.77 - 6.70 (m, 1 H), 6.56 (br s, 1 H), 3.85 (br dd, J = 4.3, 13.8 Hz, 1 H), 3.67 - 3.57 (m, 2H), 3.35 (ddd, J = 5.1 , 11 .2, 14.2 Hz, 1 H), 3.05 - 2.92 (m, 2H), 2.91 - 2.71 (m, 1 H), 2.68 - 2.52 (m, 2H), 2.40 - 2.32 (m, 1 H), 2.07 - 1 .95 (m, 3H), 1 .70 (br dd, J = 4.8, 6.9 Hz, 3H), 1 .58 - 1 .49 (m, 2H); LCMS: RT = 0.787 min, m / z = 465.1 (M+H)+; SFC: RT = 1 .531 min

[0584] EXAMPLE 1.36: SYNTHESIS OF (36)

[0585] Synthesis of compound 2

[0586] A mixture of compound 1 (1 g, 6.51 mmol, 1 eq, HCI) and H2SO4 (6.39 g, 65.10 mmol, 3.47 mL, 10 eq) in EtOH (10 mL) was stirred at 80 °C for 16 h. The mixture was added into H2O (40 mL) slowly. The pH of the mixture was adjusted to about 7 and the resulting mixture was concentrated at reduced pressure to give a residue. Compound 2 (900 mg, 6.20 mmol, crude) was obtained as a white solid. LCMS: RT =0.267 min, m / z =146.2 (M+H)+.

[0587] Synthesis of compound 4

[0588] A mixture of compound 2 (900 mg, 6.20 mmol, 1 eq), compound 3 (843.58 mg, 6.20 mmol, 1 eq), DIEA (4.01 g, 30.99 mmol, 5.40 mL, 5 eq), HOBt (1.01 g, 7.44 mmol, 1.2 eq) and EDCI (1 .43 g, 7.44 mmol, 1 .2 eq) in DMF (10 mL) was stirred at 25 °C for 16 h. To the mixture was added water (30 mL) and the mixture was extracted with EtOAc (25 mL x 3), the combined organic phase was washed with brine (20 mL), dried over Na2SO4, filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, Petroleum ether: Ethyl acetate = 1 : 0 to 2: 1 , TLC: Petroleum ether: Ethyl acetate = 2: 1 , Rf = 0.61). Compound 4 (600 mg, 2.28 mmol, 36.77% yield) was obtained as a colorless oil.1H NMR: (400 MHz, CDCI3) δ = 6.12 (br s, 1 H), 4.20 - 4.09 (m, 2H), 3.38 (d, J = 6.4 Hz, 2H), 2.96 - 2.67 (m, 5H), 1 .30 - 1 .26 (m, 3H), 1 .19 (s, 6H).

[0589] Synthesis of compound 5

[0590] A mixture of compound 4 (550 mg, 2.09 mmol, 1 eq) and LiOH. H2O (175.33 mg, 4.18 mmol, 2 eq) in THF (6 mL) and H2O (2 mL) was stirred at 25 °C for 16 h. To the mixture was added aqueous HCI solution and the pH was adjusted to 7. The pH of the mixture was adjusted to about 5 and the resulting mixture was extracted with EtOAc (8 mL x 3), the combined organic phase was dried over Na2SO4, filtered and the filtrate was concentrated under reduced pressure to give a residue. Compound 5 (350 mg, 1 .49 mmol, 58.33% yield) was obtained as a white solid.1H NMR: (400 MHz, CDCI3) 6 = 6.09 (br s, 1 H), 3.42 (d, J = 6.4 Hz, 2H), 2.96 - 2.65 (m, 5H), 1 .31 - 1 .20 (m, 6H).

[0591] Synthesis of (36) A mixture of compound 5 (29.02 mg, 123.38 μmol, 1.6 eq), DIEA (29.90 mg, 231.33 μmol, 40.29 μL, 3 eq) and HATU (35.18 mg, 92.53 μmol, 1 .2 eq) in DMF (0.5 mL) was stirred at 25 °C for 0.5 h. Then m compound 7 (20 mg, 77.11 μmol, 1 eq) was added and the mixture was stirred at 25 °C for 16 h. The mixture was purified by prep-HPLC (column: YMC-Actus Triart C18 150*30 mm*7um; mobile phase: [water (FA) -ACN]; B%: 63%- 93%, 10 min). Compound (36) (14.83 mg, 29.87 μmol, 38.74% yield, 96% purity) was obtained as a off white solid.1H NMR: (400 MHz, CDCI3) δ = 7.23 (br t, J = 7.8 Hz, 1 H), 6.85 - 6.79 (m, 3H), 6.79 - 6.72 (m, 1 H), 6.62 - 6.54 (m, 1 H), 6.42 (br s, 1 H), 4.22 - 4.10 (m, 1 H), 3.78 (s, 3H), 3.52 - 3.30 (m, 3H), 3.02 - 2.76 (m, 4H), 2.76 - 2.65 (m, 3H), 2.44 (s, 3H), 1 .35 (s, 3H), 1 .23 (s, 3H); LCMS: RT =0.618 min, m / z =477.2 (M+H)+; SFC: RT =1.112 min

[0592] EXAMPLE 1.37: SYNTHESIS OF (37)

[0593] Synthesis of compound 3

[0594] A mixture of compound 1 (500 mg, 3.44 mmol, 1 eq), CDI (614.21 mg, 3.79 mmol, 1 .1 eq) and TEA (696.90 mg, 6.89 mmol, 958.60 μL, 2 eq) in THF (5 mL) was stirred at 25 °C for 2 h. Then to the mixture was added compound 2 (320.51 mg, 2.47 mmol, 7.19e- 1 eq, HCI), DMAP (84.14 mg, 688.71 μmol, 0.2 eq) and TEA (1.05 g, 10.33 mmol, 1.44 mL, 3 eq), the resulting mixture was stirred at 25 °C for 16 h. The mixture was filtered and the filtrate was concentrated at reduced pressure to give a residue. The residue was purified by prep-HPLC (column: YMC-Actus Triart C18 150*30 mm*7um; mobile phase: [water (FA) -ACN]; B%: 33%- 63%, 10 min). Compound 3 (400 mg, 1.51 mmol, 43.95% yield) was obtained as a white solid.1H NMR: (400 MHz, CDCI3) δ = 5.02 (br s, 1 H), 4.26 (t, J = 12.1 Hz, 4H), 3.45 (q, J = 6.0 Hz, 2H), 2.49 - 2.43 (m, 2H), 1 .46 (s, 9H).

[0595] Synthesis of compound 4

[0596] A mixture of compound 3 (400 mg, 1 .51 mmol, 1 eq) in DCM (4 mL) and TFA (4 mL) was stirred at 25 °C for 2 h. The mixture was concentrated at reduced pressure to give a residue. Compound 4 (470 mg, crude) was obtained as a white solid.1H NMR: (400 MHz, CDCI3) δ = 4.33 (t, J = 11 .9 Hz, 4H), 3.54 (t, J = 5.7 Hz, 2H), 2.66 (t, J = 5.8 Hz, 2H). Synthesis of compound (37)

[0597] A mixture of compound 4 (40.13 mg, 192.78 μmol, 1 eq), compound 5 (50.00 mg, 192.78 μmol, 1 eq), DIEA (74.75 mg, 578.33 μmol, 100.74 μL, 3 eq), HOBt (31 .26 mg, 231 .33 μmol, 1 .2 eq) and EDCI (44.35 mg, 231 .33 μmol, 1 .2 eq) in DMF (1 mL) was stirred at 25 °C for 16 h. The mixture was purified by prep-HPLC (column: YMC-Actus Triart C18 150*30 mm*7um; mobile phase: [water (FA) -ACN]; B%: 50%- 80%, 10 min). Compound (37) (20.48 mg, 44.65 μmol, 23.16% yield, 98% purity) was obtained as an off white solid.1H NMR: (400 MHz, CDCI3) δ = 7.23 (t, J = 7.9 Hz, 1 H), 6.89 - 6.79 (m, 3H), 6.78 - 6.68 (m, 1 H), 6.42 - 6.34 (m, 1 H), 5.37 - 5.21 (m, 1 H), 4.30 - 4.14 (m, 4H), 3.84 - 3.72 (m, 4H), 3.62 - 3.48 (m, 2H), 3.41 - 3.30 (m, 1 H), 2.98 - 2.86 (m, 1 H), 2.85 - 2.74 (m, 1 H), 2.72 - 2.47 (m, 2H), 2.42 (s, 3H); LCMS: RT =0.552 min, m / z =450.1 (M+H)+; SFC: RT =1 .354 min.

[0598] EXAMPLE 1.38: SYNTHESIS OF (38)

[0599] A mixture of compound 2B (50 mg, 202.16 μmol, 1 eq), compound 4 (41.88 mg, 202.16 μmol, 1 eq), DIEA (78.38 mg, 606.48 μmol, 105.64 μL, 3 eq), HOBt (32.78 mg, 242.59 μmol, 1.2 eq) and EDCI (46.51 mg, 242.59 μmol, 1.2 eq) in DMF (1 mL) was stirred at 25 °C for 16 h. The mixture was concentrated, the crude was purified by prep-HPLC (column: YMC-Actus Triart C18 150*30 mm*7um; mobile phase: [water (FA) -ACN]; B%: 55%- 85%, 10 min). Compound 3, 3 -difluoro-N- [3 - [4 - (4 - fluorophenyl) - 2 -methyl- 6, 7 -dihydro- 4H-thieno [3, 2 -c] pyridin- 5 -yl] - 3 -oxo-propyl] cyclobutanecarboxamide (26.93 mg, 59.23 μmol, 29.30% yield, 96% purity) was obtained as a white solid.1H NMR: (400 MHz, CDCI3) δ = 7.28 (br s, 1 H), 7.25 (br d, J = 5.5 Hz, 1 H), 7.07 - 6.95 (m, 2H), 6.71 (s, 1 H), 6.50 (br s, 1 H), 6.37 - 6.31 (m, 1 H), 3.79 (dd, J = 5.1 , 14.1 Hz, 1 H), 3.71 - 3.50 (m, 2H), 3.31 (ddd, J = 4.4, 12.0, 14.1 Hz, 1 H), 2.98 - 2.77 (m, 4H), 2.75 - 2.54 (m, 5H), 2.42 (s, 3H); LCMS: RT =0.571 min, m / z =437.2 (M+H)+; SFC: RT =0.894 min

[0600] EXAMPLE 1.39: SYNTHESIS OF (39)

[0601] Synthesis of compound 3

[0602] To a solution of compound 1 (0.3 g, 2.10 mmol, 1 eq) in DCM (3 mL) was added DIEA (812.36 mg, 6.29 mmol, 1.09 mL, 3 eq) and compound 2 (399.45 mg, 2.10 mmol, 302.61 μL, 1 eq) at 0 °C. The mixture was stirred at 20 °C for 16 h. The reaction mixture was diluted with H2O (5 mL) and the resulting mixture was extracted with EtOAc (2 mL *3). The combined organic layers were dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, Petroleum ether / Ethyl acetate = 1 / 0 to 0 / 1 , TLC, Petroleum ether: Ethyl acetate = 3: 1 , Rf = 0.37) to give compound 3 (499 mg, 1 .68 mmol, 80.09% yield) as a brown oil. LCMS: RT = 0.519 min, m / z = 298.1 (M+H)+.

[0603] Synthesis of compound 4

[0604] To a solution of compound 3 (0.499 g, 1 .68 mmol, 1 eq) in THF (4 mL) and H2O (2 mL) was added LiOH’FW (281 .67 mg, 6.71 mmol, 4 eq). The mixture was stirred at 20 °C for 2 h. The reaction mixture was diluted with H2O (5 mL), acidified with HCI (1 M) to pH = 5~6, and then the resulting mixture was extracted with EtOAc (2 mL * 3). The combined organic layers were dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure to give compound 4 (512 mg, crude) as a yellow oil.

[0605] Synthesis of compound (39)

[0606] To a solution of compound 4 (20.00 mg, 110.95 μmol, 1 eq) in DMF (1 mL) was added compound 5 (31 .44 mg, 110.95 μmol, 1 eq), HOBt (17.99 mg, 133.13 μmol, 1 .2 eq), EDCI (25.52 mg, 133.13 μmol, 1.2 eq) and DIEA (57.36 mg, 443.78 μmol, 77.30 μL, 4 eq). The mixture was stirred at 20 °C for 16 h. The reaction mixture was diluted with H2O (10 mL) and the resulting mixture was extracted with EtOAc (3 mL *3). The combined organic layers were dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (FA condition; column: Welch Xtimate C18 150*25 mm*5um; mobile phase: [water (FA) -ACN]; gradient: 45%- 65% B over 10 min) to give the (39) (25.35 mg, 56.32 μmol, 50.76% yield, 99% purity) as a white solid.1H NMR: (400 MHz, CDCI3) δ = 8.69 - 8.60 (m, 1 H), 7.99 - 7.89 (m, 1 H), 7.69 - 7.44 (m, 1 H), 7.34-7.32 (m, 2H), 5.38 - 4.88 (m, 1 H), 4.80 - 3.96 (m, 1 H), 3.84 - 3.18 (m, 2H), 3.28 - 3.17 (m, 1 H), 2.96 - 2.84 (m, 2H), 2.82 - 2.68 (m, 1 H), 2.64 (br d, J = 4.2 Hz, 3H), 2.60 - 2.24 (m, 1 H), 2.12 - 1 .67 (m, 4H), 1 .27 - 1 .09 (m, 1 H), 0.73 - 0.63 (m, 2H), 0.59 - 0.36 (m, 2H); LCMS: RT = 0.535 min, m / z = 446.2 (M+H)+.

[0607] EXAMPLE 1.40: SYNTHESIS OF (40)

[0608] A mixture of compound 2 (81.99 mg, 334.18 μmol, 1 eq), compound 1 (100 mg, 334.18 μmol, 1 eq, TFA) , DIEA (129.57 mg, 1.00 mmol, 174.62 μL, 3 eq) , HOBt (54.19 mg, 401.01 μmol, 1.2 eq) and EDCI (76.87 mg, 401 .01 μmol, 1.2 eq) in DMF (1 mL) was stirred at 20 °C for 16h. the mixture was concentrated. The residue was purified by reversed-phase HPLC (column : Phenomenex luna C18 150*25 mm* 10um;mobile phase : [water (FA) -ACN] ;B% : 45%- 75%, 10 min) , the eluent was concentrated and then freeze dried. Compound (40) (64.62 mg, 155.07 μmol, 46.40% yield, 99% purity) was obtained as a off white solid.1H NMR: (400 MHz, T=80°C, DMSO-d6) δ = 7.51 - 7.31 (m, 2H) , 7.23 (br t, J = 7.8 Hz, 1 H) , 6.93 - 6.54 (m, 5H) , 4.20 - 3.89 (m, 1 H) , 3.72 (s, 3H) , 3.33 (td, J = 6.6, 13.0 Hz, 2H) , 3.05 (br s, 1 H) , 2.91 (br s, 2H) , 2.74 - 2.56 (m, 3H) , 1 .74 - 1 .45 (m, 8H); LCMS: RT =0.916 min, m / z =413.1 (M+H)+.

[0609] EXAMPLE 1.41: SYNTHESIS OF (41)

[0610] Synthesis of compound 3

[0611] A mixture of compound 2 (87.88 mg, 708.05 μmol, 74.47 μL, 1 eq) and compound 1 (100.00 mg, 708.05 μmol, 1 eq) in Tol. (1 mL) was stirred at 110 °C for 16 h. The mixture was concentrated at reduced pressure to remove the solution, then to the mixture was added TFA (2 mL) and the mixture was stirred at 25 °C for 16 h. The mixture was concentrated at reduced pressure to give a residue. The residue was purified by reversed-phase HPLC (0.1 % FA condition), the eluent was concentrated and then freeze dried. Compound 3 (140 mg, 543.40 μmol, 76.75% yield, 96% purity) was obtained as a white gum.1H NMR: (400 MHz, CDCI3) δ = 7.30 (br d, J = 7.7 Hz, 2H), 7.04 (br t, J = 8.3 Hz, 2H), 6.06 (s, 1 H), 5.26 (s, 1 H), 3.24 - 3.11 (m, 3H), 2.95 - 2.83 (m, 1 H), 2.37 (s, 3H). Synthesis of compound (41)

[0612] A mixture of compound 4 (37.44 mg, 125.13 μmol, 6.19e- 1 eq, TFA) , compound 3 (50 mg, 202.16 μmol, 1 eq), DIEA (78.38 mg, 606.48 μmol, 105.64 μL, 3 eq), HOBt (32.78 mg, 242.59 μmol, 1.2 eq) and EDCI (46.51 mg, 242.59 μmol, 1.2 eq) in DMF (1 mL) was stirred at 20 °C for 16 h. The reaction mixture was purified by prep-HPLC (column : Phenomenex luna C18 150*25 mm* 10um;mobile phase : [water (FA) -ACN] ;B% : 51 %- 81 %, 10 min), the eluent was concentrated and then freeze dried. Compound (41 ) (17 mg, 40.19 μmol, 19.88% yield, 98% purity) was obtained as a white solid.1H NMR: (400 MHz, CDCI3) δ = 7.27 - 7.17 (m, 1 H), 6.98 (t, J = 8.6 Hz, 2H), 6.73 (s, 1 H), 6.37 - 6.30 (m, 2H), 3.82 (br dd, J = 4.8, 13.9 Hz, 1 H), 3.67 - 3.51 (m, 2H), 3.35 - 3.23 (m, 1 H), 2.97 - 2.86 (m, 1 H), 2.84 - 2.74 (m, 1 H), 2.64 - 2.52 (m, 2H), 2.49 (br s, 1 H), 2.42 (s, 3H), 1 .87 - 1 .59 (m, 8H); LCMS: RT =0.898 min, m / z =415.2 (M+H)+.

[0613] Synthesis of compound (41)_peak1 and (41)_peak2

[0614] The solid was separated by SFC (column : DAICEL CHIRALCEL OD (250 mm*30 mm, 10um) ;mobile phase : [0.1 %NH3H2O ETCH] ;B% : 25%- 25%, 2.7 min) .

[0615] Peak 1 (SFC: RT =1 .294 min) was collected to give (41 )_peak1 (4.77 mg, 11 .39 μmol, 30.47% yield, 99% purity) as white solid.1H NMR: (400 MHz, CDCI3) δ = 7.28 (br d, J = 1.9 Hz, 1 H), 7.26 - 7.16 (m, 1 H), 7.03 - 6.95 (m, 2H), 6.73 (s, 1 H), 6.39 - 6.29 (m, 2H), 3.82 (br dd, J = 4.9, 14.1 Hz, 1 H), 3.67 - 3.49 (m, 2H), 3.36 - 3.24 (m, 1 H), 2.99 - 2.87 (m, 1 H), 2.84 - 2.71 (m, 1 H), 2.65 - 2.51 (m, 2H), 2.49 - 2.44 (m, 1 H), 2.42 (s, 3H), 1 .87 - 1 .64 (m, 8H); LCMS: RT =0.872 min, m / z =415.3 (M+H)+; SFC: RT =1 .294 min

[0616] Peak 2 (RT =1 .468 min) was collected to give (41)_peak2 (5.42 mg, 12.94 μmol, 34.62% yield, 99% purity) as white solid.1H NMR: (400 MHz, CDCI3) δ = 7.28 (d, J = 2.0 Hz, 1 H) , 7.26 - 7.17 (m, 1 H) , 6.98 (t, J = 8.7 Hz, 2H) , 6.73 (s, 1 H) , 6.38 - 6.30 (m, 2H) , 3.82 (br dd, J = 5.0, 13.8 Hz, 1 H) , 3.67 - 3.49 (m, 2H) , 3.36 - 3.22 (m, 1 H) , 2.98 - 2.86 (m, 1 H) , 2.85 - 2.71 (m, 1 H) , 2.65 - 2.50 (m, 2H) , 2.49 - 2.44 (m, 1 H) , 2.42 (s, 3H) , 1.87 - 1 .67 (m, 6H) , 1.59 - 1.50 (m, 2H); LCMS: RT =0.874 min, m / z =415.3 (M+H)+; SFC: RT =1.468 min.

[0617] EXAMPLE 1.42: SYNTHESIS OF (42)

[0618]

[0619] Synthesis of Compound 3

[0620] To a solution of compound 2 (1 g, 6.53 mmol, 1 eq) in THF (20 mL) was added into LiAIH4 (500.00 mg, 13.17 mmol, 2.02 eq) at 0 °C. The reaction solution was gradually heated followed by heating at 65 °C for 4 h. The reaction was quenched by ice slowly and then the resulting mixture was extracted with DCM (20 mL*3). The combined organic phase was washed with brine (10 mL*3), dried over anhydrous Na2SO4, filtered and concentrated in vacuum. Compound 3 (700 mg, crude) was obtained as a yellow oil.

[0621] Synthesis of Compound 4

[0622] To a solution of compound 3 (700 mg, 5.59 mmol, 1 eq) in DMF (15 mL) was added 3- methoxybenzoic acid (850.88 mg, 5.59 mmol, 1.95 mL, 1 eq), HOBt (906.79 mg, 6.71 mmol, 1.2 eq), EDCI (1 .29 g, 6.71 mmol, 1 .2 eq) and DIEA (1 .81 g, 13.98 mmol, 2.44 mL, 2.5 eq) at 25 °C. The mixture was stirred at 25 °C for 14 h. EtOAc (30 mL) and water (30 mL) were added and Layers were separated. The aqueous phase was extracted with EtOAc (30 mL * 2). Combined extracts were washed with brine (30 mL), dried over Na2SO4, filtered, and concentrated under vacuum to give a residue. The residue was purified by flash silica gel chromatography (ISCO®; 20 g SepaFlash® Silica Flash Column, Eluent of 40-60% Ethyl acetate / Petroleum ethergradient @ 50 mL / min). Compound 4 (570 mg, 1.58 mmol, 28.30% yield, 72% purity) was obtained as a yellow oil. LCMS: RT = 0.565 min, m / z = 260.0 (M+H)+.

[0623] Synthesis of Compound 5

[0624] A mixture of compound 4 (400 mg, 1.54 mmol, 1 eq) and POCh (1.42 g, 9.26 mmol, 860.11 μL, 6 eq) in Tol. (8 mL) was degassed and purged with N2 for 3 times, and then the mixture was stirred at 100 °C for 14 h under N2 atmosphere. The pH of reaction mixture was adjusted to about 8 with NH3. H2O (25%). The mixture was filtered and the filtrate was concentrated under vacuum to give a residue. The residue was purified by flash silica gel chromatography (ISCO®; 4 g SepaFlash® Silica Flash Column, Eluent of 0-80% Ethyl acetate / Petroleum ethergradient @ 30 mL / min). Compound 5 (260 mg, 937.48 μmol, 60.77% yield, 87% purity) was obtained as a yellow oil. LCMS: RT = 0.623 min, m / z = 242.1 (M+H)+.

[0625] Synthesis of Compound 6

[0626] To a solution of compound 5 (260 mg, 1 .08 mmol, 1 eq) in MeOH (6 mL) was added NaBFU (81 .53 mg, 2.16 mmol, 2 eq) at 25 °C. The mixture was stirred at 25 °C for 2 h. The mixture was quenched with H2O (1 mL). The resulting mixture was filtered and the filtrate was concentrated under vacuum to give a residue. Compound 6 (220 mg, 786.68 μmol, 73.01 % yield, 87% purity) was obtained as a yellow oil. LCMS: RT = 0.942 min, m / z = 244.1 (M+H)+.

[0627] Synthesis of (42), (42)_peak1 & (42)_peak2

[0628] A mixture of compound 6 (100 mg, 411 .01 μmol, 1 eq), compound 7 (93.67 mg, 452.12 μmol, 1.1 eq), HOBt (72.20 mg, 534.32 μmol, 1.3 eq), EDCI (102.43 mg, 534.32 μmol, 1.3 eq) and DIEA (159.36 mg, 1 .23 mmol, 214.77 μL, 3 eq) in DMF (2 mL) was degassed and purged with N2 for 3 times, and then the mixture was stirred at 25 °C for 14 h under N2 atmosphere. The mixture was filtered and the filtrate was concentrated under vacuum to give a residue. The residue was purified by prep-HPLC (column: Waters Xbridge 150*25 mm* 5um; mobile phase: [water (NH4HCO3) -ACN]; B%: 43%-73%, 8 min). Compound (42) (20.13 mg, 46.55 μmol, 11.33% yield, 100% purity) was obtained as a yellow solid. (42) was further separated by SFC (RT = 1 .073 min and RT = 1 .765 min; column: DAICEL CHIRALPAK AD (250mm*30mm,10um);mobile phase: [0.1 %NH3H2O ETOH];B%: 50%-50%,3.7min). Compound (42)_peak1 (5.79 mg, 13.12 umol, 3.19% yield, 98% purity) was obtained as an off-white solid. Compound (42)_peak2 (5.69 mg, 13.16 umol, 3.20% yield, 100% purity) was obtained as an off-white solid.

[0629] Compound (42)1H NMR: (400 MHz, DMSO-d6) δ = 8.11 - 8.00 (m, 1 H), 7.32 - 7.19 (m, 1 H), 6.92 - 6.74 (m, 3H), 6.45 (s, 1 H), 6.13 - 5.85 (m, 1 H), 4.03 - 3.89 (m, 1 H), 3.79 - 3.66 (m, 3H), 3.35-3.20 (m, 2H), 3.17 - 3.03 (m, 1 H), 2.93 - 2.73 (m, 2H), 2.72 - 2.53 (m, 7H), 2.30 - 2.19 (m, 3H); LCMS: RT = 0.967 min, m / z = 433.2 (M+H)+.

[0630] Compound (42)_peak1 :1H NMR: (400 MHz, CDCI3) δ = 7.24 (t, J = 7.9 Hz, 1 H), 6.96 - 6.76 (m, 3H), 6.65 (s, 1 H), 6.53 (br s, 1 H), 5.95 - 5.66 (m, 1 H), 3.85 - 3.70 (m, 4H), 3.68 - 3.52 (m, 2H), 3.37 - 2.94 (m, 1 H), 2.92 - 2.75 (m, 3H), 2.74 - 2.51 (m, 6H), 2.35 - 2.23 (m, 3H); LCMS: RT = 0.61 1 min, m / z = 433.4 (M+H)+; SFC: RT = 1 .053 min

[0631] Compound (42)_peak2:1H NMR: (400 MHz, CDCI3) δ = 7.24 (s, 1 H), 6.98 - 6.77 (m, 3H), 6.65 (s, 1 H), 6.57 - 6.44 (m, 1 H), 5.95 - 5.65 (m, 1 H), 3.92 - 3.70 (m, 4H), 3.68 - 3.54 (m, 2H), 3.36 - 2.95 (m, 1 H), 2.93 - 2.75 (m, 3H), 2.75 - 2.47 (m, 6H), 2.29 (s, 3H); LCMS: RT = 0.620 min, m / z = 433.2 (M+H)+;

[0632] SFC: RT = 1.729 min

[0633] EXAMPLE 1.43: SYNTHESIS OF (43) To a solution of Compound 2 (33.67 mg, 161 .73 μmol, 1 eq) in DMF (1 mL) was added DIEA (62.71 mg, 485.18 μmol, 84.51 μL, 3 eq) and EDCI (37.20 mg, 194.07 μmol, 1 .2 eq), Compound 1 (40.00 mg, 161 .73 μmol, 1 eq), HOBt (26.22 mg, 194.07 μmol, 1 .2 eq). The mixture was stirred at 25 °C for 16 hr. The mixture was purified by prep-HPLC (column: YMC-Actus Triart C18 150*30 mm*7um; mobile phase: [water (FA) -ACN]; B%: 50%- 80%, 10 min). (43) (25.68 mg, 57.53 μmol, 35.57% yield, 98% purity) was obtained as a yellow gum.1H NMR: (400 MHz, CDCI3) δ = 121 - 7.15 (m, 2H), 6.99 (br t, J = 8.5 Hz, 2H), 6.71 (s, 1 H), 6.39 - 6.29 (m, 1 H), 5.28 (br d, J = 1.6 Hz, 1 H), 4.37 - 4.12 (m, 4H), 3.88 - 3.75 (m, 1 H), 3.69 - 3.46 (m, 2H), 3.38 - 3.25 (m, 1 H), 3.01 - 2.87 (m, 1 H), 2.86 - 2.78 (m, 1 H), 2.58 (br d, J = 1 .9 Hz, 2H), 2.43 (s, 3H); LCMS: RT = 0.558 min, m / z = 438.2 (M+H)+; SFC: RT =1 .347 min.

[0634] EXAMPLE 1.44: SYNTHESIS OF (44)

[0635] Synthesis of compound 2 To a solution of compound 1 (500 mg, 3.67 mmol, 1 eq) in THF (5 mL) was added methyl carbonochloridate (620 mg, 6.56 mmol, 506.95 μL, 1.79 eq) at 0 °C and the mixture was stirred at 20 °C for 1 hr. To the mixture was added a solution of bromo (m-tolyl) magnesium (1 M, 7.34 mL, 2 eq) in THF slowly at 0 °C and the mixture was stirred at 20 °C for 16 hr. To the mixture was added water (10 mL) and the resulting mixture was extracted with EtOAc (10 mL x 3), the combined organic phase was dried over Na2SO4, filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by flash silica gel chromatography (ISCO®; 20 g SepaFlash® Silica Flash Column, Eluent of 70-95% Ethyl acetate / Petroleum ethergradient @ 60 mL / min; TLC (Petroleum ether: Ethyl acetate = 1 : 1 ; Rf = 0.23)), the eluent was concentrated. Compound 2 (930 mg, crude) was obtained as a white solid. LCMS: RT = 0.550 min, m / z = 287.1 (M+H)+.

[0636] Synthesis of compound 3

[0637] The mixture of compound 2 (930 mg, 3.25 mmol, 1 eq) and Pd(OH)2 / C (100 mg, 10% purity) in EtOH (10 mL) was stirred at 60 °C for 84 h under H2 (50 psi). The mixture was filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (column: Phenomenex Luna C18 150*25 mm*10um; mobile phase: [water (FA) -ACN]; gradient: 38%- 68% B over 10 min), the eluent was concentrated and then freeze dried. Compound 3 (200 mg, 693.57 μmol, 21.36% yield) was obtained as a colorless oil.1H NMR: (400 MHz, CDCI3) δ = 8.69 (s, 1 H), 7.23 - 7.18 (m, 1 H), 7.14 - 7.04 (m, 3H), 6.63 - 6.35 (m, 1 H), 4.55 - 4.21 (m, 1 H), 3.79 (br s, 3H), 3.28 - 2.96 (m, 2H), 2.95 - 2.84 (m, 1 H), 2.33 (s, 3H).

[0638] Synthesis of compound 4

[0639] A mixture of compound 3 (200 mg, 693.57 μmol, 1 eq) and KOH (466.96 mg, 8.32 mmol, 12 eq) in EtOH (4 mL) and H2O (1 mL) was stirred at 90 °C for 48 hr. The mixture was concentrated under reduced pressure to give a residue. The residue was purified by flash silica gel chromatography (ISCO®; 12 g SepaFlash® Silica Flash Column, Eluent of 100% Ethyl acetate / Petroleum ethergradient @ 60 mL / min; TLC (Petroleum ether: Ethyl acetate = 0: 1 ; Rf = 0.06)), the eluent was concentrated. Compound 4 (140 mg, 607.83 μmol, 87.64% yield) was obtained as a colorless oil. LCMS: RT = 0.366 min, m / z = 231.1 (M+H)+.

[0640] Synthesis of compound (44) Peak 1 and Peak 2

[0641] A mixture of compound 4 (70 mg, 303.91 μmol, 1 eq), compound 5 (75.56 mg, 364.70 μmol, 1 .2 eq), HOBt (49.28 mg, 364.70 μmol, 1.2 eq), EDCI (69.91 mg, 364.70 μmol, 1.2 eq) and DIEA (78.56 mg, 607.83 μmol, 105.87 μL, 2 eq) in DCM (1 mL) was stirred at 20 °C for 16 h. To the mixture was added water (5 mL) and the resulting mixture was extracted with EtOAc (5 mL x 3), the combined organic phase was dried over Na2SO4, filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by prep-TLC (SiO2, Petroleum ether: Ethyl acetate = 0: 1), the eluent was concentrated to give (44). (44) was separated by SFC (column: DAICEL CHIRALPAK AS (250 mm*30 mm, 10um); mobile phase: [CO2 -EtOH (0.1 %NH3H2O)]; B%: 20%, isocratic elution mode). Peak 1 (RT = 1 .168 min) was collected to give (44)_peak1 as white solid.

[0642] Peak 2 (RT = 1 .351 min) was collected to give (44)_peak2 as white solid.

[0643] Compound (44)_peak1 (19.66 mg, 46.87 μmol, 15.42% yield, 100% purity) was obtained as a white solid.1H NMR: (400 MHz, CDCI3) δ = 8.72 (s, 1 H), 7.26 - 6.98 (m, 5H), 6.53 - 6.46 (m, 1 H), 3.91 (br d, J = 13.6 Hz, 1 H), 3.68 - 3.58 (m, 2H), 3.51 - 3.40 (m, 1 H), 3.11 - 2.94 (m, 2H), 2.92 - 2.77 (m, 2H), 2.76

[0644] - 2.53 (m, 5H), 2.37 - 2.29 (m, 3H); LCMS: RT = 0.498 min, m / z = 420.2 (M+H)+; SFC: RT= 1 .171 min.

[0645] Compound (44)_peak2 (20.43 mg, 48.70 μmol, 16.03% yield, 100% purity) was obtained as a white solid. 1 H NMR: (400 MHz, CDCI3) δ = 8.73 - 8.71 (m, 1 H), 7.26 - 6.97 (m, 5H), 6.50 (br s, 1 H), 3.95 - 3.87 (m, 1 H), 3.68 - 3.56 (m, 2H), 3.51 - 3.41 (m, 1 H), 3.12 - 2.94 (m, 2H), 2.91 - 2.77 (m, 2H), 2.75 - 2.58 (m, 5H), 2.36 - 2.29 (m, 3H); LCMS: RT = 0.493 min, m / z = 420.2 (M+H)+; SFC: RT= 1 .356 min.

[0646] EXAMPLE 1.45: SYNTHESIS OF (45)

[0647] Synthesis of compound 2B

[0648] To a solution of compound 2A (2 g, 9.09 mmol, 1 eq) and 2-iodopropane (1 .85 g, 10.91 mmol, 1 .09 mL, 1 .2 eq) in DMF (20 mL) was added K2CO3 (1 .63 g, 11 .82 mmol, 1 .3 eq). The reaction was heated to 55 °C for 5 h. Petroleum ether (30 mL) was added and layers were separated. The aqueous phase was extracted with Petroleum ether (30 mL x 2). Combined extracts were washed with brine (30 mL x 3), dried over Na2SO4, filtered, and concentrated under vacuum to give a residue. Compound 2B (2.1 g, 8.01 mmol, 88.14% yield) was obtained as a colorless liquid.1H NMR: (400 MHz, CDCI3) δ = 7.21 - 7.14 (m, 2H), 6.90 (t, J = 7.5 Hz, 1 H), 6.77 (d, J = 8.5 Hz, 1 H), 4.43 (spt, J = 6.0 Hz, 1 H), 1.25 (d, J =

[0649] 6.0 Hz, 6H).

[0650] Synthesis of compound 2 A solution of compound 2B (1 g, 3.82 mmol, 1 eq) in THF (3 mL) was added over 0.5 hour to a solution of Lithium; chloro (isopropyl) magnesium; chloride (1.3 M, 2.94 mL, 1 eq) at 0 ° C. Stirring of the mixture was continued at 25 ° C for 2.5 h. Reaction mixture was used directly.

[0651] Synthesis of compound 3

[0652] To a solution of compound 1 (260 mg, 1.91 mmol, 1 eq) in THF (4 mL) was added methyl carbonochloridate (216.51 mg, 2.29 mmol, 177.03 μL, 1 .2 eq) at 0 °C. The mixture was stirred at 20 °C for 30 min. The iminium salt solution was recooled to 0 °C, and the mixture of compound 2 (in THF, 2 eq) was added slowly. The reaction allowed to warm to 20 °C and stirred for 14 h. The reaction mixture was poured into saturated NH4CI aqueous solution (10 mL) and the resulting mixture was extracted with EtOAc (15 mL*3). The combined organic layer was washed with brine (15 mL), dried over Na2SO4, filtered and concentrated at reduced pressure to give a residue. The residue was purified by flash silica gel chromatography (ISCO®; 12 g SepaFlash® Silica Flash Column, Eluent of 0-30% Ethyl acetate / Petroleum ether gradient @ 45 mL / min). Compound 3 (400 mg, 1 .14 mmol, 59.60% yield, 94% purity) was obtained as a yellow oil. LCMS: RT = 0.569 min, m / z = 331 .2 (M+H)+.

[0653] Synthesis of compound 4

[0654] To a solution of compound 3 (400 mg, 1.21 mmol, 1 eq) in EtOH (15 mL) was added Pd(OH)2 / C (510.05 mg, 10% purity) under N2. The suspension was degassed under vacuum and purged with H2 several times. The mixture was stirred under H2 (50 psi) at 60 °C for 62 h. The mixture was filtered and the filtrate was concentrated under vacuum to give a residue. Compound 4 (380 mg, 1.14 mmol, 94.42% yield) was obtained as a yellow oil. LCMS: RT = 0.549 min, m / z = 333.2 (M+H)+.

[0655] Synthesis of compound 5

[0656] To a solution of compound 4 (380 mg, 1 .14 mmol, 1 eq) in EtOH (10 mL) and H2O (2 mL) was added KOH (641.37 mg, 11.43 mmol, 10 eq) at 25 °C. The mixture was stirred at 90 °C for 14 h. The mixture was filtered and the filtrate was concentrated under vacuum to give a residue. The residue was purified by flash silica gel chromatography (ISCO®; 12 g SepaFlash® Silica Flash Column, Eluent of 70-100% Ethyl acetate / Petroleum ethergradient @ 40 mL / min). Compound 5 (150 mg, 475.62 μmol, 41.61 % yield, 87% purity) was obtained as a yellow oil. LCMS: RT = 0.382 min, m / z = 275.2 (M+H)+.

[0657] Synthesis of compound (45)_peak1 & (45)_peak2

[0658] To a solution of compound 5 (75 mg, 273.34 μmol, 1 eq) in DCM (2 mL) was added compound 6 (62.29 mg, 300.68 μmol, 1.1 eq), HOBt (44.32 mg, 328.01 μmol, 1 .2 eq), EDCI (62.88 mg, 328.01 μmol, 1 .2 eq) and DIEA (105.98 mg, 820.03 μmol, 142.83 μL, 3 eq) at 25 °C. The mixture was stirred at 20 °C for 16 h. The mixture was filtered and the filtrate was concentrated under vacuum to give a residue. The residue was purified by prep-TLC (SiO2, PE: EtOAc = 1 : 2) to give desired compound 7 (100 mg, purity 98%) as a white solid, which was further separated by SFC (RT = 0.568 min and 0.812 min, column: DAICEL CHIRALPAK IC (250 mm*30 mm, 10um) ; mobile phase: [CO2-EtOH (0.1%NH3H2O) ]; B%: 37.5%, isocratic elution mode).

[0659] Compound (45)_peak1 (38.49 mg, 82.20 μmol, 30.07% yield, 99% purity) was obtained as an off- white solid.1H NMR: (400 MHz, CDCI3) δ = 8.78 -8.68 (m, 1H), 7.26-7.15 (m, 1H), 6.99 (s, 1H), 6.93 -6.75 (m, 3H), 6.53 (td, J = 5.5, 10.5 Hz, 1 H), 4.62 - 4.43 (m, 1H), 3.90 (brdd, J = 3.1, 12.8 Hz, 1H), 3.71 - 3.55 (m, 2H), 3.51 - 3.32 (m, 1H), 3.17-2.96 (m, 2H), 2.93-2.61 (m, 7H), 1.39 - 1.25 (m, 6H); LCMS: RT = 0.516 min, m / z = 464.2 (M+H)+; SFC: RT = 0.567 min.

[0660] Compound (45)_peak2 (33.18 mg, 70.86 μmol, 25.92% yield, 99% purity) was obtained as a white solid.1H NMR: (400 MHz, CDCI3) δ = 8.78-8.67 (m, 1H), 7.26-7.17 (m, 1H), 6.99 (s, 1H), 6.93-6.73 (m, 3H), 6.61 -6.46 (m, 1H), 4.52 (sxt, J = 6.1 Hz, 1H), 3.90 (brdd, J = 3.1, 12.7 Hz, 1H), 3.73 - 3.55 (m, 2H), 3.45 (ddd, J= 6.5, 9.7, 14.4 Hz, 1H), 3.14-2.95 (m, 2H), 2.90 -2.57 (m, 7H), 1.36 - 1.28 (m, 6H); LCMS: RT = 0.514 min, m / z = 464.2 (M+H)+; SFC: RT = 0.807 min.

[0661] EXAMPLE 1.46: SYNTHESIS OF (46)

[0662] Synthesis of compound 3

[0663] To a mixture of compound 1 (4 g, 31.44 mmol, 3.67 mL, 1 eq) and TEA (3.18 g, 31.44 mmol, 4.38 mL, 1 eq) in DCM (40 mL) was added compound 2 (2.91 g, 31.44 mmol, 2.91 mL, 1 eq) at 0 °C, the mixture was stirred at 20 °C for 2 h. To the mixture was added water (40 mL) and extracted with EtOAc (40 mL x 3), the organic phase was dried over Na2SO4, filtered and the filtrate was concentrated under reduced pressure to give a residue. Without further purification and the products was used for next step directly. Compound 3 (5.7 g, 31.10 mmol, 98.91% yield) was obtained as a yellow oil.1H NMR: (400 MHz, CDCI3) δ = 7.17 (br d, J = 5.1 Hz, 1 H), 6.95 (dd, J = 3.6, 4.8 Hz, 1 H), 6.85 - 6.81 (m, 1 H), 5.65 (br s, 1 H), 3.53 (q, J = 6.4 Hz, 2H), 3.04 (br t, J = 6.6 Hz, 2H), 2.19 (q, J = 7.6 Hz, 2H), 1 .18 - 1 .09 (m, 3H).

[0664] Synthesis of compound 4

[0665] A mixure of compound 3 (5.7 g, 31.10 mmol, 1 eq) and POCh (14.31 g, 93.30 mmol, 8.67 mL, 3 eq) in ACN (60 mL) was stirred at 60 °C for 16 h. The mixture was added into water (80 mL) and the pH was adjusted to 10 by K2CO3, then the mixture was extracted with EtOAc (50 mL x 3), the organic phase was dried over Na2SO4, filtered and the filtrate was concentrated under reduced pressure to give a residue. Without further purification and the products was used for next step directly. Compound 4 (3.1 g, 18.76 mmol, 60.31 % yield) was obtained as a yellow oil.1H NMR: (400 MHz, CDCI3) δ = 7.12 - 7.06 (m, 2H), 3.81 - 3.74 (m, 2H), 2.85 - 2.78 (m, 2H), 2.65 (tq, J = 1 .3, 7.5 Hz, 2H), 1 .21 (t, J = 7.4 Hz, 3H).

[0666] Synthesis of compound 5

[0667] To a solution of compound 4 (3.1 g, 18.76 mmol, 1 eq) in MeOH (20 mL) was added NaBH4 (1.55 g, 41 .08 mmol, 2.19 eq) at 0 °C under N2, and the mixture was stirred at 25 °C under N2 for 2 h. To the mixture was added saturated NH4CI aqeous solution (20 mL) and extracted with EtOAc (20 mL x 3), the organic phase was dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. Without further purification and the products was used for next step directly. Compound 5 (1 .5 g, 8.97 mmol, 47.80% yield) was obtained as a white solid.

[0668] Synthesis of compound 7

[0669] A mixture of compound 5 (1.4 g, 8.37 mmol, 1 eq), compound 6 (1.92 g, 8.37 mmol, 1 eq), TEA (1 .69 g, 16.74 mmol, 2.33 mL, 2 eq), HOBt (1 .36 g, 10.04 mmol, 1 .2 eq) and EDCI (1 .93 g, 10.04 mmol, 1.2 eq) inDCM (10 mL) was stirred at 20 °C for 16 h. The mixture was concentrated under reduced pressure to give a residue. The residue was purified by flash silica gel chromatography (ISCO®; 40 g SepaFlash® Silica Flash Column, Eluent of 45-55% Ethyl acetate / Petroleum ethergradient @ 60 mL / min; TLC(Petroleum ether : Ethyl acetate = 1 : 1 ; Rf = 0.69) ), the eluent was concentrated to give a residue. The residue was purified by reversed-phase HPLC(0.1% FA condition), the eluent was concentrated and then freeze dried. Compound 7 (1.69 g, 4.46 mmol, 53.34% yield) was obtained as a yellow oil. LCMS: RT = 0.976 min, m / z = 323.1 (M + H -tBu)+.

[0670] Synthesis of compound 8

[0671] A mixture of compound 7 (1.69 g, 4.46 mmol, 1 eq) and TFA (32.53 g, 285.32 mmol, 21.12 mL, 63.91 eq) in DCM (20 mL) was stirred at 20 °C for 2 h. The mixture was concentrated under reduced pressure to give a residue. Without further purification and the products was used for next step directly. Compound 8 (1.7 g, 4.33 mmol, 97.03% yield, TFA) was obtained as a yellow oil. LCMS: RT = 0.648 min, m / z = 279.1 (M+H)+.

[0672] Synthesis of (46), (46)_peak1 & (46)_peak2 To a solution of 2-chlorobenzenesulfonyl chloride (53.78 mg, 254.82 μmol, 34.70 μL, 1 eq) (53.78 mg, 254.82 μmol, 34.70 μL, 1 eq) and compound 8 (100 mg, 254.82 μmol, 1 eq, TFA) in DCM (2 mL) was added TEA (103.14 mg, 1.02 mmol, 141.87 μL, 4 eq). The mixture was stirred at 20 °C or 16 h. The reaction mixture was concentrated in vacuum. The residue was purified by prep-HPLC (column: Phenomenex Luna C18 150*25 mm* 10um; mobile phase: [water (FA) -ACN]; B%: 51 %-81 %, 10 min), the eluent was concentrated and then freeze dried. Compound (46) (34.57 mg, 76.31 μmol, 29.95% yield, 100% purity) was obtained as off-white solid.1H NMR: (400 MHz, T=80°C, DMSO-de) 6 = 7.99 (d, J = 7.9 Hz, 1 H), 7.70 - 7.62 (m, 2H), 7.59 - 7.52 (m, 1 H), 7.27 (d, J = 5.1 Hz, 1 H), 6.89 (d, J = 5.1 Hz, 1 H), 5.42 (br d, J = 2.5 Hz, 1 H), 4.26 - 4.02 (m, 1 H), 3.79 - 3.60 (m, 2H), 3.43 - 3.23 (m, 1 H), 2.97 - 2.84 (m, 3H), 2.80 (br s, 2H), 1.91 - 1 .72 (m, 2H), 1 .70 - 1 .46 (m, 4H), 0.86 (br s, 3H); LCMS: RT =0.964 min, m / z =453.2 (M+H)+.

[0673] The solid was separated by SFC (column: DAICEL CHIRALCEL OX (250 mm*30 mm, 10um); mobile phase: [0.1 %NH3H2O ETOH]; B%: 55%- 55%, 5.2 min). Peak 1 was collected to give (46)_peak1 (9.83 mg, 21 .26 μmol, 29.19% yield, 98% purity) as off white solid.1H NMR: (400 MHz, CDCI3) 6 = 8.12 - 8.03 (m, 1 H), 7.57 - 7.46 (m, 2H), 7.45 - 7.35 (m, 1 H), 7.16 - 7.07 (m, 1 H), 6.84 - 6.74 (m, 1 H), 5.57 (br dd, J = 5.3, 8.7 Hz, 1 H), 4.91 - 4.66 (m, 1 H), 4.04 - 3.36 (m, 3H), 3.03 - 2.55 (m, 5H), 2.14 - 1.61 (m, 6H), 1 .07 - 0.86 (m, 3H); LCMS: RT =0.822 min, m / z =453.1 (M+H)+; SFC: RT =1 .024 min.

[0674] Peak 2 was collected to give (46)_peak2 (18.07 mg, 39.49 μmol, 54.21 % yield, 99% purity) as off white solid.1H NMR: (400 MHz, CDCI3) δ = 8.12 - 8.01 (m, 1 H), 7.56 - 7.45 (m, 2H), 7.44 - 7.34 (m, 1 H), 7.16 - 7.07 (m, 1 H), 6.84 - 6.74 (m, 1 H), 5.57 (dd, J = 5.2, 8.9 Hz, 1 H), 4.92 - 4.67 (m, 1 H), 4.06 - 3.39 (m, 3H), 3.02 - 2.61 (m, 5H), 2.13 - 1 .61 (m, 6H), 1 .05 - 0.87 (m, 3H); LCMS: RT =0.814 min, m / z =453.2 (M+H)+; SFC: RT =1.625 min.

[0675] EXAMPLE 1.47: SYNTHESIS OF (47)

[0676]

[0677] Synthesis of compound 3

[0678] To a mixture of compound 1 (4 g, 31 .44 mmol, 3.67 mL, 1 eq) and TEA (3.18 g, 31 .44 mmol, 4.38 mL, 1 eq) in DCM (40 mL) was added compound 2 (2.91 g, 31.44 mmol, 2.91 mL, 1 eq) at 0 °C, the mixture was stirred at 20 °C for 2 h. To the mixture was added water (40 mL) and extracted with EtOAc (40 mL x 3), the organic phase was dried over Na2SO4, filtered and the filtrate was concentrated under reduced pressure to give a residue. Without further purification and the products was used for next step directly. Compound 3 (5.7 g, 31.10 mmol, 98.91% yield) was obtained as a yellow oil.1H NMR: (400 MHz, CDCI3) δ = 7.17 (br d, J = 5.1 Hz, 1 H), 6.95 (dd, J = 3.6, 4.8 Hz, 1 H), 6.85 - 6.81 (m, 1 H), 5.65 (br s, 1 H), 3.53 (q, J = 6.4 Hz, 2H), 3.04 (br t, J = 6.6 Hz, 2H), 2.19 (q, J = 7.6 Hz, 2H), 1 .18 - 1 .09 (m, 3H).

[0679] Synthesis of compound 4

[0680] A mixure of compound 3 (5.7 g, 31.10 mmol, 1 eq) and POCb (14.31 g, 93.30 mmol, 8.67 mL, 3 eq) in ACN (60 mL) was stirred at 60 °C for 16 h. The mixture was added into water (80 mL) and the pH was adjusted to 10 by K2CO3, then the mixture was extracted with EtOAc (50 mL x 3), the organic phase was dried over Na2SO4, filtered and the filtrate was concentrated under reduced pressure to give a residue. Without further purification and the products was used for next step directly. Compound 4 (3.1 g, 18.76 mmol, 60.31 % yield) was obtained as a yellow oil.1H NMR: (400 MHz, CDCI3) δ = 7.12 - 7.06 (m, 2H), 3.81 - 3.74 (m, 2H), 2.85 - 2.78 (m, 2H), 2.65 (tq, J = 1 .3, 7.5 Hz, 2H), 1 .21 (t, J = 7.4 Hz, 3H).

[0681] Synthesis of compound 5

[0682] To a solution of compound 4 (3.1 g, 18.76 mmol, 1 eq) in MeOH (20 mL) was added NaBH4 (1.55 g, 41 .08 mmol, 2.19 eq) at 0 °C under N2, and the mixture was stirred at 25 °C under N2 for 2 h. To the mixture was added saturated NH4CI aqeous solution (20 mL) and extracted with EtOAc (20 mL x 3), the organic phase was dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. Without further purification and the products was used for next step directly. Compound 5 (1 .5 g, 8.97 mmol, 47.80% yield) was obtained as a white solid.

[0683] Synthesis of compound 7

[0684] A mixture of compound 5 (1.4 g, 8.37 mmol, 1 eq), compound 6 (1.92 g, 8.37 mmol, 1 eq), TEA (1 .69 g, 16.74 mmol, 2.33 mL, 2 eq), HOBt (1 .36 g, 10.04 mmol, 1 .2 eq) and EDCI (1 .93 g, 10.04 mmol, 1.2 eq) inDCM (10 mL) was stirred at 20 °C for 16 h. The mixture was concentrated under reduced pressure to give a residue. The residue was purified by flash silica gel chromatography (ISCO®; 40 g SepaFlash® Silica Flash Column, Eluent of 45-55% Ethyl acetate / Petroleum ethergradient @ 60 mL / min; TLC(Petroleum ether : Ethyl acetate = 1 : 1 ; Rf = 0.69) ), the eluent was concentrated to give a residue. The residue was purified by reversed-phase HPLC(0.1% FA condition), the eluent was concentrated and then freeze dried. Compound 7 (1.69 g, 4.46 mmol, 53.34% yield) was obtained as a yellow oil. LCMS: RT = 0.976 min, m / z = 323.1 (M + H -tBu)+.

[0685] Synthesis of compound 8

[0686] A mixture of compound 7 (1.69 g, 4.46 mmol, 1 eq) and TFA (32.53 g, 285.32 mmol, 21.12 mL, 63.91 eq) in DCM (20 mL) was stirred at 20 °C for 2 h. The mixture was concentrated under reduced pressure to give a residue. Without further purification and the products was used for next step directly. Compound 8 (1.7 g, 4.33 mmol, 97.03% yield, TFA) was obtained as a yellow oil. LCMS: RT = 0.648 min, m / z = 279.1 (M+H)+.

[0687] Synthesis of compound (47)

[0688] To a mixture of Compound 8 (80 mg, 203.86 μmol, 1 eq, TFA) and TEA (61.88 mg, 61 1.57 μmol, 85.12 μL, 3 eq) in DCM (1 mL) was added 2,6-dichlorobenzenesulfonyl chloride (50.05 mg, 203.86 μmol, 1 eq) at 0 °C, then the mixture was stirred at 20 °C for 16 h. The mixture was concentrated under reduced pressure to give a residue. The residue was purified by reversed-phase HPLC(column: Unisil 3 - 100 C18 Ultra 150*50 mm*3 um; mobile phase: [water (FA) -ACN]; B%: 41 %- 71 %, 7 min), the eluent was concentrated and then freeze dried. Compound (47) (19.53 mg, 39.26 μmol, 19.26% yield, 98% purity) was obtained as a white solid.1H NMR: (400 MHz, DMSO-de) δ = 7.67 - 7.63 (m, 2H), 7.59 - 7.54 (m, 1 H), 7.27 (d, J = 5.1 Hz, 1 H), 6.89 (d, J = 5.3 Hz, 1 H), 5.51 - 5.29 (m, 1 H), 4.28 - 4.04 (m, 1 H), 3.87 - 3.71 (m, 2H), 3.45 - 3.24 (m, 1 H), 3.02 - 2.90 (m, 3H), 2.81 (br d, J = 2.4 Hz, 2H), 1 .87 - 1 .60 (m, 6H), 0.88 (br d, J = 2.1 Hz, 3H). LCMS: RT = 0.995 min, m / z = 487.0 (M+H)+.

[0689] EXAMPLE 1.48: SYNTHESIS OF (48)

[0690] Synthesis of compound 3

[0691] A mixture of compound 1 (1 g, 7.86 mmol, 917.43 uL, 1 eq), compound 2 (1.07 g, 8.65 mmol, 909.52 uL, 1.1 eq) in Tol. (10 mL) was stirred at 110 °C for 16 h. The mixture was concentrated, to the residue was TFA (12.55 g, 110.06 mmol, 8.15 mL, 14 eq) and stirred at 25 °C for 16 h. The mixture was concentrated under reduced pressure to give a residue. The residue was purified by reversed-phase HPLC (0.1 % NH3 H2O), the eluent was concentrated and then freeze dried to give compound 3 (640 mg, 2.74 mmol, 34.90% yield, 100% purity) as a red solid.1H NMR: (400 MHz, DMSO-d6) 6 = 7.40 - 7.31 (m, 1 H), 7.28 - 6.97 (m, 4H), 6.43 (br d, J = 5.0 Hz, 1 H), 4.94 (s, 1 H), 3.14 - 3.07 (m, 1 H), 2.99 - 2.79 (m, 3H), 2.77 - 2.69 (m, 1 H).

[0692] Synthesis of (48)

[0693] A mixture of compound 4 (39.69 mg, 214.31 umol, 1 eq), compound 3 (50 mg, 214.31 umol, 1 eq) , HOBt (34.75 mg, 257.18 umol, 1.2 eq) , DIEA (166.19 mg, 1 .29 mmol, 223.98 uL, 6 eq) and EDCI (49.30 mg, 257.18 umol, 1.2 eq) in DMF (2 mL) was stirred at 25 °C for 16 h. The mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (column: Phenomenex luna C18 150*25mm* 10um;mobile phase: [water(FA)-ACN];B%: 44%-74%,10min) to give (48) (57.09 mg, 139.69 umol, 65.18% yield, 98% purity) as a yellow solid.1H NMR: (400 MHz, DMSO- d6) δ = 7.81 - 7.70 (m, 1 H), 7.43 - 7.32 (m, 2H), 7.09 (br d, J = 2.4 Hz, 1 H), 7.06 (br s, 1 H), 6.95 (br d, J = 10.1 Hz, 1 H), 6.86 (d, J = 5.1 Hz, 1 H), 6.69 (s, 1 H), 4.05 - 3.96 (m, 1 H), 3.29 (br s, 1 H), 3.20 - 3.13 (m, 1 H), 3.01 - 2.84 (m, 2H), 2.83 - 2.71 (m, 1 H), 2.55 (br s, 3H), 1 .77 - 1 .36 (m, 8H) LCMS: RT = 0.981 min, m / z = 401 .0 (M + H)+.

[0694] EXAMPLE 1.49: SYNTHESIS OF (49)

[0695]

[0696] Synthesis of compound 2

[0697] To a solution of compound 1 (500 mg, 4.27 mmol, 1 eq) in EtOH (5 mL) was added TEA (1.30 g, 12.81 mmol, 1.78 mL, 3 eq) and hydroxylamine; hydrochloride (890.17 mg, 12.81 mmol, 3 eq), the mixture was stirred at 80 °C for 16 h. The reaction mixture was diluted with ethyl acetate (50 mL). The organic layer was washed with brine (30 mL), dried over Na2SO4, filtered and the filtrate was concentrated under vacuum to give compound 2 (620 mg, 4.13 mmol, 96.72% yield) as colorless gum.1H NMR: (400 MHz, DMSO-d6) δ = 9.09 (s, 1 H), 5.45 (br s, 2H), 2.77 - 2.63 (m, 5H).

[0698] Synthesis of compound 3

[0699] To a solution of compound 2 (300 mg, 2.00 mmol, 1 eq) and 3-methoxy-2 -methyl-3-oxo-propanoic acid (290.41 mg, 2.20 mmol, 1.1 eq) in DMF (4 mL) was added DIEA (774.79 mg, 5.99 mmol, 1 .04 mL, 3 eq), HOBt (405.02 mg, 3.00 mmol, 1 .5 eq) and EDCI (574.62 mg, 3.00 mmol, 1 .5 eq), the mixture was stirred at 20 °C for 16 h. The reaction mixture was diluted with ethyl acetate (30 mL). The organic layer was washed with brine (20 mL*2), dried over Na2SO4, filtered and the filtrate was concentrated under vacuum to give compound 3 (600 mg, crude) as colorless gum. LCMS: RT =0.418 min, m / z =265.1 (M+H)+.

[0700] Synthesis of compound 4

[0701] To a solution of compound 3 (200 mg, 756.93 μmol, 1 eq) in toluene (2 mL) was added AcOH (136.37 mg, 2.27 mmol, 130.00 μL, 3 eq), the mixture was stirred at 110 °C for 2 h. The reaction mixture was diluted with ethyl acetate (50 mL). The organic layer was washed with brine (30 mL), dried over Na2SO4, filtered and the filtrate was concentrated under vacuum to give compound 4 (130 mg, 528.00 μmol, 69.76% yield) as yellow gum. LCMS: RT =0.489 min, m / z =247.1 (M+H)+.

[0702] Synthesis of compound 5

[0703] To a solution of compound 4 (240 mg, 974.78 μmol, 1 eq) in THF (3 mL) and H2O (1 mL) was added LiOH H2<D (81.81 mg, 1.95 mmol, 2 eq), then the mixture was stirred at 25 °C for 16 h. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was diluted with H2O (5 mL) and acidified to pH = 5 - 6 with 1 M HCI aqueous solution, the resulting mixture was extracted with ethyl acetate (10 mL * 3). The combined organic layers were dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. Compound 5 (210 mg, 904.46 μmol, 92.79% yield) was obtained as a light yellow oil.

[0704] Synthesis of (49)_peak1 and (49)_peak2

[0705] To a mixture of compound 2 (32.70 mg, 140.85 μmol, 1 .1 eq) and compound 3 (30.00 mg, 128.05 μmol, 1 eq) in pyridine (1 mL) was added EDCI (36.82 mg, 192.07 μmol, 1.5 eq), then the mixture was stirred at 25 °C for 16 h. The mixture was concentrated in vacuum to give a residue. The residue was purified by prep-HPLC (column: YMC-Actus Triart C18 150*30 mm*7um; mobile phase: [water (FA) - ACN]; gradient: 48%- 78% B over 10 min) to give (49).

[0706] Then compound (49) was further separated by SFC (RT=1 .825 min &2.040 min; column: DAICEL CHIRALPAK IG (250 mm*30 mm, 10um); mobile phase: [CO2 -EtOH]; B%: 35%, isocratic elution mode). Peak 1 was collected to give (49)_peak1 (4.11 mg, 8.98 μmol, 7.01 % yield, 98% purity) as a yellow solid.1H NMR: (400 MHz, CDCI3) δ = 8.78 - 8.69 (m, 1 H), 7.43 - 7.28 (m, 1 H), 7.24 (br d, J = 5.5 Hz, 1 H), 7.12 - 6.29 (m, 3H), 4.96 - 4.00 (m, 2H), 3.55 - 2.65 (m, 8H), 1 .73-1 .69 (m, 3H). LCMS: RT = 0.546 min, m / z = 449.1 (M+H)+. SFC: RT = 1 .837 min.

[0707] Peak 2 was collected to give (49)_peak2 (8.09 mg, 17.68 μmol, 13.81 % yield, 98% purity) was obtained as a yellow gum.1H NMR: (400 MHz, CDCI3) δ = 8.78 - 8.69 (m, 1 H), 7.43 - 7.28 (m, 1 H), 7.24 (br d, J = 5.5 Hz, 1 H), 7.12 - 6.29 (m, 3H), 4.96 - 4.00 (m, 2H), 3.55 - 2.65 (m, 8H), 1 .63-1 .61 (m, 3H). LCMS: RT = 0.542 min, m / z = 449.1 (M+H)+. SFC: RT = 2.069 min.

[0708] EXAMPLE 1.50: SYNTHESIS OF (50)

[0709] Synthesis of compound 2 To a solution of compound 1 (2 g, 17.99 mmol, 1 eq) in / -PrOH (15 mL) was added isobutyl nitrite (2.78 g, 26.99 mmol, 1.5 eq) and diiodomethane (8.43 g, 31.49 mmol, 2.54 mL, 1.75 eq), the mixture was stirred at 55 °C for 16 h. The reaction mixture was diluted with ethyl acetate (100 mL). The organic layer was washed with brine (50 mL*2), dried over Na2SO4, filtered and the filtrate was concentrated under vacuum to give a residue. The residue was purified by column chromatography (100 mesh silica gel, petroleum ether: ethyl acetate = 1 : 0 to 3: 1) to give compound 2 (2.35 g, 10.58 mmol, 58.82% yield) as colorless oil.1H NMR: (400 MHz, CDCI3) δ = 6.17 (s, 1 H), 3.79 (s, 3H), 2.26 (s, 3H).

[0710] Synthesis of compound 3

[0711] To a solution of compound 2 (2.35 g, 10.58 mmol, 1 eq) in THF (10 mL) was added / -PrMgChLiCI (1 .3 M, 8.14 mL, 1 eq) at - 30 °C under N2, the mixture was stirred at - 30 °C for 3 h under N2. Without work-up and the reaction mixture was used to next step directly. To give compound 3 (1.64 g, crude) (in 18.14 mL THF) as yellow liquid.

[0712] Synthesis of compound 5

[0713] To a mixture of compound 4 (500 mg, 3.67 mmol, 1 eq) in THF (10 mL) was added methyl carbonochloridate (0.346 g, 3.66 mmol, 282.91 μL, 9.97e- 1 eq) at 0 °C under N2 atmosphere. The mixture was stirred at 20 °C for 0.5 h. Then compound 3 (1 .62 g, 10.46 mmol, 2.85 eq) (in 18.14 mL THF) was added slowly at 0 °C. The reaction was allowed to warm to 20 °C and stirred for 16 h. To the reaction mixture was added water (10 mL), the resulting mixture was extracted with ethyl acetate (20 mL*2), and the combined organic phase was washed with brine (10 mL), dried with anhydrous Na2SO4, filtered and the filtrate was concentrated in vacuum to give residue. The residue was purified by column chromatography (100 mesh silica gel, petroleum ether: ethyl acetate = 1 : 0 to 0: 1), Rf = 0.20) to give compound 5 (490 mg, 1.69 mmol, 45.96% yield) as yellow gum.1H NMR: (400 MHz, CDCI3) 6 = 8.74 - 8.50 (m, 1 H), 7.22 - 6.93 (m, 1 H), 6.91 - 6.64 (m, 1 H), 6.07 - 5.79 (m, 2H), 3.85 (s, 3H), 3.69 (s, 3H), 2.23 - 2.14 (m, 3H).

[0714] Synthesis of compound 6

[0715] To a solution of compound 5 (490 mg, 1.69 mmol, 1 eq) in EtOH (10 mL) was added Pd(OH)2 / C (49.00 mg, 348.92 μmol, 2.07e-1 eq), the suspension was degassed under vacuum and purged with H2 for 3 times. The reaction mixture was stirred under H2 (50 psi) at 70 °C for 96 h. The reaction mixture was cooled to room temperature. EtOH (150 mL) was added, the mixture was filtered through a pad of silica, the filtrate was concentrated under vacuum to give compound 6 (380 mg, crude) as yellow gum.

[0716] Synthesis of compound 7

[0717] To a solution of compound 6 (380 mg, 1 .30 mmol, 1 eq) in EtOH (4 mL) and H2O (1 mL) was added KOH (218.78 mg, 3.90 mmol, 3 eq), the mixture was stirred at 90 °C for 16 h under N2. The reaction mixture was concentrated under vacuum to removed EtOH, then diluted with H2O (5 mL), the resulting mixture was extracted with ethyl acetate (10 mL*2), the combined organic layers were dried over Na2SO4, filtered and the filtrate was concentrated under vacuum to give compound 7 (270 mg, crude) as yellow solid.

[0718] Synthesis of compound (50)

[0719] To a solution of compound 8 (50 mg, 215.35 μmol, 1 eq) and compound 7 (50.46 mg, 215.35 μmol, 1 eq) in pyridine (1 mL) was added EDCI (61 .92 mg, 323.02 μmol, 1 .5 eq), the mixture was stirred at 20 °C for 16 h. The reaction mixture was diluted with ethyl acetate (20 mL). The organic layer was washed with brine (10 mL), dried over Na2SO4, filtered and the filtrate was concentrated under vacuum to give a residue. The residue was purified by prep-HPLC (column: Welch Xtimate C18 150*25 mm*5um; mobile phase: [water (FA) -ACN]; gradient: 28%- 58% B over 10 min) to give (50) (13.13 mg, 28.98 μmol, 13.46% yield, 99% purity) as yellow solid.1H NMR: (400 MHz, CDCI3) δ = 8.74 - 8.62 (m, 1 H), 6.84 - 6.18 (m, 1 H), 6.18 - 5.76 (m, 1 H), 4.98 - 4.87 (m, 1 H), 4.48 - 3.82 (m, 1 H), 3.76 - 3.63 (m, 3H), 3.57 - 3.33 (m, 1 H), 3.25 - 2.63 (m, 7H), 2.25 - 2.16 (m, 3H), 1.76 - 1 .67 (m, 3H). LCMS: RT = 0.483 min, m / z = 449.0 (M+H)+.

[0720] 30 mg of mixture of (50) was purified by SFC (column: DAICEL CHIRALPAK IC (250 mm*30 mm, 10um); mobile phase: [CO2 -i-PrOH / ACN]; B%: 55%, isocratic elution mode).

[0721] Peak 1 was collected to give (50)_peak1 (14.48 mg, 32.29 μmol, 20.69% yield, 100% purity) as yellow gum.1H NMR: (400 MHz, CDCI3) δ = 8.79 - 8.62 (m, 1 H), 6.84 - 6.18 (m, 1 H), 6.18 - 5.72 (m, 1 H), 5.00 - 4.12 (m, 2H) , 4.04 - 3.12 (m, 1 H), 3.76 - 3.62 (m, 3H), 3.55 - 3.41 (m, 1 H), 3.08 - 2.78 (m, 6H), 2.21 (d, J = 8.3 Hz, 3H), 1.74 - 1.67 (m, 3H). LCMS: RT = 0.477 min, m / z = 449.1 (M+H)+. SFC: RT = 0.807 min.

[0722] Peak 2 was collected to give (50)_peak2 (13.69 mg, 30.52 μmol, 19.56% yield, 100% purity) as yellow solid.1H NMR: (400 MHz, CDCI3) δ = 8.67 (s, 1 H), 6.86 - 6.30 (m, 1 H), 6.17 - 5.73 (m, 1 H), 4.96

[0723] - 4.02 (m, 1 H), 4.88 - 4.33 (m, 1 H), 3.96 - 3.19 (m, 1 H), 3.80 - 3.62 (m, 3H), 3.54 - 3.33 (m, 1 H), 3.08 - 2.63 (m, 6H), 2.26 - 2.15 (m, 3H), 1.75 - 1.66 (m, 3H). LCMS: RT = 0.481 min, m / z = 449.1 (M+H)+. SFC: RT = 0.867 min.

[0724] Peak 3 was collected to give (50)_peak3 (11 .97 mg, 26.69 μmol, 17.10% yield, 100% purity) as off- white.1H NMR:(400 MHz, CDCI3) δ = 8.78 - 8.62 (m, 1 H), 6.85 - 6.19 (m, 1 H), 6.18 - 5.72 (m, 1 H), 4.98

[0725] - 4.12 (m, 2H), 4.01 - 3.18 (m, 1 H), 3.69 (d, J = 6.7 Hz, 3H), 3.56 - 3.43 (m, 1 H), 3.08 - 2.79 (m, 6H), 2.21 (d, J = 8.7 Hz, 3H), 1 .71 (t, J = 6.6 Hz, 3H). LCMS: RT = 0.476 min, m / z = 449.1 (M+H)+. SFC: RT = 1.078 min.

[0726] Peak 4 was collected to give (50)_peak4 (13.01 mg, 29.01 μmol, 18.59% yield, 100% purity) as off- white solid.1H NMR:(400 MHz, CDCI3) δ = 8.73 - 8.61 (m, 1 H), 6.85 - 6.28 (m, 1 H), 6.17 - 5.73 (m, 1 H), 4.96 - 4.03 (m, 1 H), 4.91 - 4.34 (m, 1 H), 3.80 - 3.60 (m, 3H), 3.40 (dt, J = 2.3, 8.4 Hz, 1 H), 3.19 (dt, J = 3.5, 12.3 Hz, 1 H), 3.09 - 2.66 (m, 6H), 2.26 - 2.17 (m, 3H), 1 .75 - 1 .67 (m, 3H). LCMS: RT = 0.475 min, m / z = 449.1 (M+H)+. SFC: RT = 1 .253 min.

[0727] EXAMPLE 1.51: SYNTHESIS OF (51)

[0728] Synthesis of compound 2

[0729] To the mixture of compound 1 (1 g, 4.29 mmol, 1 eq), compound 2 (552.91 mg, 6.44 mmol, 1.5 eq) in dioxane (10 mL) was added CsF (1 .30 g, 8.58 mmol, 2 eq) and Pd(dppf)Cl2 (313.99 mg, 429.12 μmol, 0.1 eq). The mixture was stirred at 100 °C for 16 h under N2. The mixture was concentrated at reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, Petroleum ether / Ethyl acetate = 1 : 0 to 15: 1 , RF = 0.7). Compound 3 (740 mg, 3.81 mmol, 88.80% yield) was obtained as colorless oil.1H NMR: (400 MHz, CDCI3) δ = 7.86 (dd, J = 6.2, 8.7 Hz, 1 H), 6.88 (dt, J = 2.6, 8.3 Hz, 1 H), 6.67 (dd, J = 2.6, 10.5 Hz, 1 H), 3.91 (s, 3H), 2.82 - 2.71 (m, 1 H), 1 .09 - 1 .00 (m, 2H), 0.73 - 0.64 (m, 2H).

[0730] Synthesis of compound 4

[0731] To the mixture of compound 3 (740 mg, 3.81 mmol, 1 eq) in THF (8 mL) was added LiOH» H2O (319.80 mg, 7.62 mmol, 2 eq) and H2O (2 mL). The mixture was stirred at 25 °C for 2 h. LiOH» H2O (319.8 mg, 7.62 mmol, 2 eq) was added to the mixture. The mixture was stirred at 25 °C for 48 h. The reaction mixture was concentrated under reduced pressure to remove MeOH and THF. The residue was diluted with H2O (2 mL), acidified with HCI (1 M) to pH = 1—2, and the resulting mixture was extracted with EtOAc (5 mL *3). The combined organic layers were dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure to give a residue. Compound 4 (650 mg, crude) was obtained as a white solid. LCMS: RT = 0.472 min, m / z = 181.1 (M+H)+.

[0732] Synthesis of compound (51) To the mixture of compound 4 (21 .98 mg, 122.00 μmol, 1 eq) in pyridine (1 .5 mL) was added EDCI (46.77 mg, 244.00 μmol, 2 eq) and compound 5 (40 mg, 122.00 μmol, 1 eq, HCI). The mixture was stirred at 25 °C for 16 h.The mixture was stirred at 40 °C for 2 h. Compound 4 (25 mg, 138.75 μmol, 1.14 eq) and EDCI (50 mg, 260.82 μmol, 2.14 eq) were added to the mixture. The mixture was stirred at 40 °C for 16 h.The mixture was purified directly. The residue was purified by prep-HPLC (column: Unisil 3 - 100 C18 Ultra 150*50 mm*3 um; mobile phase: [water (FA) -ACN]; gradient: 35%- 65% B over 10 min). Compound (51) (13.41 mg, 28.38 μmol, 23.26% yield, 96% purity) was obtained as a white solid.1H NMR: (400 MHz, CDCI3) δ = 8.70 - 8.61 (m, 1 H), 7.24 - 7.05 (m, 1 H), 6.95 - 6.78 (m, 1 H), 6.61 - 6.45 (m, 1 H), 5.46 - 4.08 (m, 1 H), 5.04 - 4.67 (m, 2H), 3.81 - 3.16 (m, 2H), 3.11 -3.02 (m, 1 H), 3.02 - 2.77 (m, 4H), 2.09 - 1 .72 (m, 5H), 1.18 - 0.94 (m, 3H), 0.84 - 0.66 (m, 3H), 0.65 - 0.35 (m, 3H). LCMS: RT = 0.524 min, m / z = 454.1 (M+H)+.

[0733] EXAMPLE 1.52: SYNTHESIS OF (52)

[0734] To a solution of compound 1 (20 mg, 61.00 μmol, 1 eq, HCI) and DIEA (23.65 mg, 183.00 μmol, 31 .87 μL, 3 eq) in DCM (1 mL) was added compound 2 (12.73 mg, 61 .00 μmol, 1 eq) under 0 °C, then the mixture was stirred at 25 °C for 16 h. LCMS showed -39% of compound 1 remained and -38% of desired mass was detected. To the reaction mixture was added DIEA (1 1 .83 mg, 91 .50 μmol, 15.94 μL, 1 .5 eq) and compound 2 (12.73 mg, 61 .00 μmol, 1 eq) under 0 °C, then the mixture was stirred at 25 °C for 2 h. To the reaction mixture was added H2O (15 mL), the resulting mixture was extracted with ethyl acetate (5 mL * 4), and the combined organic phase was washed with brine (5 mL * 3), dried with anhydrous Na2SO4, filtered and the filtrate was concentrated in vacuum to give a residue. The residue was purified by prep-HPLC (column: Unisil 3 - 100 C18 Ultra 150*50 mm*3 um; mobile phase: [water (FA) -ACN]; gradient: 40%- 60% B over 10 min) to give compound (52) (4.1 mg, 8.84 μmol, 14.50% yield) as a yellow gum.1H NMR: (400 MHz, CDCI3) δ = 8.65 (br d, J = 11 .4 Hz, 1 H), 7.52 (br d, J = 4.9 Hz, 1 H), 7.37 (q, J = 6.7 Hz, 1 H), 7.22 - 7.10 (m, 2H), 5.38-4.80 (m, 1 H), 4.95 - 4.06 (m, 1 H), 4.30 (br d, J = 6.2 Hz, 2H), 3.77-3.20 (m, 3H), 3.72 - 3.63 (m, 2H), 2.99 - 2.81 (m, 3H), 2.78 - 2.53 (m, 2H), 2.05 - 1 .64 (m, 4H), 1 .29-1 .11 (m, 1 H), 0.79 - 0.64 (m, 2H), 0.62 - 0.34 (m, 2H). LCMS: RT = 0.584 min, m / z = 464.2 (M+H)+.

[0735] EXAMPLE 1.53: SYNTHESIS OF (53)

[0736]

[0737] Synthesis of compound 3

[0738] To a solution of compound 2 (100 mg, 516.34 μmol, 1 eq) and DIEA (200.20 mg, 1 .55 mmol, 269.81 μL, 3 eq) in DCM (2 mL) was added compound 1 (118.13 mg, 619.61 μmol, 89.49 μL, 1.2 eq) at 0 °C under N2. The reaction mixture was stirred at 25 °C for 1 h. The reaction mixture was diluted with ethyl acetate (20 mL) and the resulting mixture was washed with H2O (8 mL*3), the organic was dried with anhydrous Na2SO4, filtered and the filtrate was concentrated in vacuum to give compound 3 (145 mg, crude) as yellow oil. LCMS: RT = 0.568 min, m / z = 312.0 (M+H)+.

[0739] Synthesis of compound 4

[0740] To a solution of compound 3 (145 mg, 465.65 μmol, 1 eq) in THF (1.5 mL), H2O (0.5 mL) and MeOH (0.5 mL) was added LiOH • H2O (58.62 mg, 1 .40 mmol, 3 eq). The reaction mixture was stirred at 25 °C for 2 h. LiOH • H2O (39.08 mg, 931 .29 μmol, 2 eq) was added to the reaction mixture. The reaction mixture was stirred at 25°C for another 2 h. The reaction mixture was diluted with H2O (10 mL) and concentrated in vacuum to remove MeOH. The aqueous solution was acidified with 1 N HCI to pH = 5~6, then the resulting mixture was extracted with ethyl acetate (5 mL*3), the combined organic was dried with anhydrous Na2SO4, filtered and the filtrate was concentrated in vacuum to give a residue. The residue was purified by column chromatography (SiO2, Petroleum ether / Ethyl acetate = 1 / 0 to 3 / 1 , TLC (Petroleum ether: Ethyl acetate = 3: 1)) to give compound 4 (70 mg, 235.40 μmol, 50.55% yield) as colorless oil.1H NMR: (400 MHz, CDCI3) δ = 7.93 - 7.61 (m, 1 H), 7.50 - 7.43 (m, 1 H), 7.35 - 7.29 (m, 2H), 3.94-3.75 (m, 1 H), 3.58 - 3.48 (m, 2H), 2.97 - 2.84 (m, 2H), 2.63 - 2.44 (d, 3H), 2.20 - 2.12 (m, 2H), 1 .59 - 1 .51 (m, 2H), 1 .28 - 1 .23 (m, 3H).

[0741] Synthesis of compound (53)

[0742] To a solution of compound 4 (20 mg, 67.26 μmol, 1 eq) and DIEA (26.08 mg, 201 .77 μmol, 35.14 μL, 3 eq) in DMF (0.5 mL) was added HATU (30.69 mg, 80.71 μmol, 1.2 eq), the reaction mixture was stirred at 25 °C for 0.5 h. Then compound 5 (12.12 mg, 67.26 μmol, 1 eq) was added to the reaction mixture. The reaction mixture was stirred at 25 °C for 2 h. The reaction mixture was diluted with ethyl acetate (8 mL) and the resulting mixture was washed with H2O (2 mL*3), the organic was dried with anhydrous Na2SO4, filtered and the filtrate was concentrated in vacuum to give a residue. The residue was purified by prep-HPLC (column: Unisil 3 - 100 C18 Ultra 150*50 mm*3 um; mobile phase: [water (FA) -ACN]; gradient: 48%- 78% B over 2 min) followed by lyophilization to give (53) (5.24 mg, 11.40 μmol, 16.95% yield, 100% purity) as yellow solid.1H NMR: (400 MHz, CDCI3) δ = 8.64 (s, 1 H), 7.89 - 7.59 (m, 1 H), 7.55 - 7.38 (m, 1 H), 7.32 - 7.28 (m, 2H), 5.41 - 5.24 (m, 1 H), 4.58 - 4.35 (m, 1 H), 3.67 - 3.46 (m, 3H), 3.09 - 2.81 (m, 4H), 2.80 - 2.37 (m, 4H), 2.29 (br d, J = 12.5 Hz, 2H), 1 .36 - 1 .24 (m, 4H), 1 .14 - 1 .06 (m, 1 H), 0.72 - 0.47 (m, 3H), 0.40 - 0.27 (m, 1 H). LCMS: RT = 0.531 min, m / z = 460.2 (M+H)

[0743] EXAMPLE 1.54: SYNTHESIS OF (54)

[0744] Synthesis of compound 3

[0745] To a solution of compound 2 (200 mg, 972.38 μmol, 1 eq, HCI) and DIEA (377.02 mg, 2.92 mmol, 508.11 μL, 3 eq) in DCM (3 mL) was added compound 1 (222.46 mg, 1.17 mmol, 168.53 μL, 1.2 eq) at 0 °C under N2. The reaction mixture was stirred at 25 °C for 2 h. LCMS showed compound 2 remained and desired mass detected. Then compound 1 (148.30 mg, 777.90 μmol, 112.35 μL, 0.8 eq) was added to the reaction mixture at 0 °C. The reaction mixture was stirred at 25 °C for 2 h. The reaction mixture was diluted with ethyl acetate (20 mL) and the resulting mixture was washed with H2O (8 mL*3), the organic was dried with anhydrous Na2SO4, filtered and the filtrate was concentrated in vacuum to give a residue. The residue was purified by column chromatography (SiO2, Petroleum ether / Ethyl acetate = 1 / 0 to 3 / 1 , TLC (Petroleum ether: Ethyl acetate = 3: 1)) to give compound 3 (220 mg, 680.26 μmol, 69.96% yield) as colorless oil.1H NMR: (400 MHz, CDCI3) δ = 8.02 (d, J = 7.9 Hz, 1 H), 7.50 - 7.43 (m, 1 H), 7.34 - 7.28 (m, 2H), 4.27 - 4.20 (m, 2H), 3.66 (s, 3H), 2.76 - 2.66 (m, 4H), 2.19 - 2.09 (m, 2H), 1 .90 - 1 .79 (m, 4H), 1 .74 - 1 .66 (m, 2H).

[0746] Synthesis of compound 4

[0747] To a solution of compound 3 (220 mg, 680.26 μmol, 1 eq) in THF (3 mL) and H2O (1 mL) was added UOH H2O (85.64 mg, 2.04 mmol, 3 eq). The reaction mixture was stirred at 25 °C for 2 h. The reaction mixture was diluted with H2O (10 mL) and the mixture was acidified with 1 N HCI to pH = 5~6, then the resulting mixture was extracted with ethyl acetate (4 mL*3), the combined organic was dried with anhydrous Na2SO4, filtered and the filtrate was concentrated in vacuum to give compound 4 (172 mg, 555.95 μmol, 81 .73% yield) as white solid. LCMS: RT = 0.437 min, m / z = 310.0 (M+H)+.

[0748] Synthesis of compound (54)

[0749] To a solution of compound 4 (20 mg, 64.65 μmol, 1 eq) and DIEA (25.06 mg, 193.94 μmol, 33.78 μL, 3 eq) in DMF (0.5 mL) was added HATU (29.50 mg, 77.57 μmol, 1.2 eq). The reaction mixture was stirred at 25 °C for 0.5 h. Then compound 5 (11.65 mg, 64.65 μmol, 1 eq) was added to the reaction mixture. The reaction mixture was stirred at 25 °C for 1 h. The reaction mixture was purified by prep- HPLC (column: Unisil 3 - 100 C18 Ultra 150*50 mm*3 um; mobile phase: [water (FA) -ACN]; gradient: 40%- 70% B over 10 min) followed by lyophilization to give (54) (8.95 mg, 18.98 μmol, 29.35% yield, 100% purity) as white solid.1H NMR: (400 MHz, CDCI3) δ = 8.65 (d, J = 6.8 Hz, 1 H), 8.02 (br dd, J = 3.7, 7.1 Hz, 1 H), 7.50 - 7.42 (m, 1 H), 7.34 - 7.28 (m, 2H), 5.35 - 4.73 (m, 1 H), 4.94 - 4.07 (m, 1 H ), 4.35 - 4.15 (m, 2H), 3.74 - 3.14 (m, 1 H), 3.07 - 2.76 (m, 3H), 2.75 - 2.67 (m, 3H), 2.37 - 2.08 (m, 3H), 2.07 - 1.96 (m, 1 H), 1.87 - 1.66 (m, 4H), 1.50 - 1.39 (m, 1 H), 1.17 - 0.33 (m, 4H). LCMS: RT = 0.553 min, m / z = 472.2 (M+H)+.

[0750] EXAMPLE 1.55: SYNTHESIS OF (55)

[0751]

[0752] Synthesis of compound 3

[0753] To a solution of compound 2 (200 mg, 1.27 mmol, 196.08 μL, 1 eq) and DIEA (493.26 mg, 3.82 mmol, 664.77 μL, 3 eq) in DCM (5 mL) was added compound 1 (417.54 mg, 1.53 mmol, 223.28 μL, 1.2 eq) at 0 °C under N2. The reaction mixture was stirred at 25 °C for 2 h. The reaction mixture was diluted with ethyl acetate (20 mL) and the resulting mixture was washed with H2O (8 mL*3), the organic was dried with anhydrous Na2SO4, filtered and the filtrate was concentrated in vacuum to give a residue. The residue was purified by column chromatography (SiO2, Petroleum ether / Ethyl acetate = 1 / 0 to 3 / 1 , TLC (Petroleum ether: Ethyl acetate = 3: 1)) to give compound 3 (496 mg, 1.26 mmol, 98.89% yield) as colorless oil.1H NMR: (400 MHz, CDCI3) δ = 8.13 (dd, J = 5.9, 8.9 Hz, 1 H), 7.49 (dd, J = 2.6, 8.1 Hz, 1 H), 7.15 (ddd, J = 2.6, 7.5, 8.8 Hz, 1 H), 4.15 (q, J = 7.1 Hz, 2H), 3.73 (td, J = 3.8, 13.0 Hz, 2H), 3.00 - 2.90 (m, 2H), 2.43 (tt, J = 4.0, 10.6 Hz, 1 H), 2.03 - 1 .93 (m, 2H), 1 .86 - 1 .73 (m, 2H), 1 .27 - 1 .24 (m, 3H).

[0754] Synthesis of compound 5

[0755] To a solution of compound 3 (496 mg, 1 .26 mmol, 1 eq) and compound 4 (324.19 mg, 3.77 mmol, 3 eq) in dioxane (10 mL) was added CsF (573.31 mg, 3.77 mmol, 3 eq) and Pd(dppf)Cl2 (92.05 mg, 125.81 μmol, 0.1 eq) under N2. The reaction mixture was stirred at 100 °C for 16 h. The reaction mixture was concentrated in vacuum to give a residue. The residue was purified by column chromatography (SiO2, Petroleum ether / Ethyl acetate = 1 / 0 to 1 / 1 , TLC (Petroleum ether: Ethyl acetate = 3: 1)) to give compound 5 (419 mg, 1.18 mmol, 93.71 % yield) as yellow oil. LCMS: RT = 0.595 min, m / z = 356.1 (M+H)+.

[0756] Synthesis of compound 6

[0757] To a solution of compound 5 (419 mg, 1.18 mmol, 1 eq) in THF (5 mL) and H2O (2 mL) was added LiOH»H2O (98.94 mg, 2.36 mmol, 2 eq). The reaction mixture was stirred at 25 °C for 2 h. The reaction mixture was diluted with H2O (10 mL) and acidified with 1 N HCI to pH = 5~6, then the resulting mixture was extracted with ethyl acetate (5 mL*3), the combined organic phase was dried with anhydrous Na2SO4, filtered and the filtrate was concentrated in vacuum to give compound 6 (330 mg, 1 .01 mmol, 85.51 % yield) as white solid. LCMS: RT = 0.491 min, m / z = 328.0 (M+H)+.

[0758] Synthesis of compound (55)

[0759] To a solution of compound 6 (30 mg, 91.64 μmol, 1 eq) and DIEA (35.53 mg, 274.92 μmol, 47.89 μL, 3 eq) in DMF (1 mL) was added HATU (41.81 mg, 109.97 μmol, 1.2 eq). The reaction mixture was stirred at 25 °C for 0.5 h. Then compound 7 (16.52 mg, 91.64 μmol, 1 eq) was added to the mixture. The reaction mixture was stirred at 25 °C for 2 h. The reaction mixture was diluted with ethyl acetate (8 mL) and the resulting mixture was washed with H2O (3 mL*3), the organic was dried with anhydrous Na2SO4, filtered and the filtrate was concentrated in vacuum to give a residue. The residue was purified by prep-HPLC (column: Unisil 3 - 100 C18 Ultra 150*50 mm*3 um; mobile phase: [water (FA) -ACN]; gradient: 48%- 68% B over 10 min) followed by lyophilization to give (55) (15.75 mg, 32.17 μmol, 35.10% yield, 100% purity) as white solid.1H NMR: (400 MHz, CDCI3) δ = 8.65 (d, J = 12.4 Hz, 1 H), 7.97 (ddd, J = 6.0, 8.9, 1 1 .0 Hz, 1 H), 6.98 - 6.89 (m, 1 H), 6.66 - 6.58 (m, 1 H), 5.39 - 4.75 (m, 1 H), 4.97 - 4.04 (m, 1 H), 3.85 - 3.18 (m, 3H), 2.96 - 2.76 (m, 4H), 2.76 - 2.57 (m, 2H), 2.11 - 1 .92 (m, 1 H), 1 .92 - 1 .76 (m, 2H), 1.75 - 1 .65 (m, 1 H), 1.25 - 1.10 (m, 3H), 0.86 - 0.78 (m, 2H), 0.73 - 0.63 (m, 2H), 0.60 - 0.36 (m, 2H). LCMS: RT = 0.546 min, m / z = 490.1 (M+H)+.

[0760] EXAMPLE 1.56: SYNTHESIS OF (56)

[0761] Synthesis of compound 2

[0762] To a solution of compound 1 (2 g, 16.65 mmol, 1 eq) in THF (20 mL) was added methyl carbonochloridate (2.74 g, 29.00 mmol, 2.24 mL, 1 .74 eq) at 0 °C under N2. The mixture was stirred at 25 °C for 0.5 h. The reaction mixture was cooled to 0 °C, and the solution of bromo(cyclopropyl) magnesium (1 M, 33.30 mL, 2 eq) was added. The reaction allowed to warm to 25 °C and stirred for 16 h under N2. The reaction mixture was quenched with NH4CI aqueous solution (80 mL) and the resulting mixture was extracted with EtOAc (60 mL*3). The combined organic layer was washed with brine (50 mL), dried over Na2SO4, filtered and concentrated at reduced pressure to give a residue. The residue was purified by column chromatography (100 mesh silica gel, petroleum ether: ethyl acetate = 1 : 0 to 5: 1) to give compound 2 (1 .5 g, 6.81 mmol, 40.90% yield) as yellow gum.1H NMR: (400 MHz, CDCI3) 6 = 7.72 (s, 1 H), 7.11 - 6.79 (m, 1 H), 5.76 (br s, 1 H), 5.54 - 5.24 (m, 1 H), 3.86 (s, 3H), 1 .25 - 1 .16 (m, 1 H), 0.63 - 0.32 (m, 4H).

[0763] Synthesis of compound 3

[0764] To Pd(OH)2 / C (200 mg, 10% purity) was added EtOH (20 mL) and compound 2 (1.5 g, 6.81 mmol, 1 eq) under N2. The suspension was degassed under vacuum and purged with H2 several times. The mixture was stirred under H2 (30 psi) at 30 °C for 16 h. The mixture was filtered, and the filtrate was concentrated under vacuum to give compound 3 (1.2 g, 5.40 mmol, 79.27% yield) as yellow gum. LCMS: RT =0.438 min, m / z =223.1 (M+H)+.

[0765] Synthesis of compound 4

[0766] To a solution of compound 3 (1 .2 g, 5.40 mmol, 1 eq) in EtOH (20 mL) and H2O (4 mL) was added KOH (1 .51 g, 27.00 mmol, 5 eq), the mixture was stirred at 90 °C for 16 h. The reaction mixture was concentrated under vacuum to give a residue. The residue was diluted with ethyl acetate (100 mL). The organic layer was washed with brine (50 mL), dried over Na2SO4, filtered and the filtrate was concentrated under vacuum to give a residue. The residue was purified by reversed phase flash (0.1% NH3«H2O) to give compound 4 (220 mg, 1 .34 mmol, 24.81 % yield) as yellow gum.

[0767] Synthesis of compound (56)

[0768] To a solution of compound 4 (20 mg, 121.80 μmol, 1 eq) and compound 5 (39.87 mg, 121 .80 μmol, 1 eq) in pyridine (1 mL) was added EDCI (28.02 mg, 146.16 μmol, 1 .2 eq) , the mixture was stirred at 25 °C for 2 h. The reaction mixture was diluted with ethyl acetate (20 mL). The organic layer was washed with brine (10 mL), dried over Na2SO4, filtered and the filtrate was concentrated under vacuum to give a residue. The residue was purified by prep-HPLC (column: Unisil 3 - 100 C18 Ultra 150*50 mm*3 um; mobile phase: [water (FA) -ACN]; gradient: 40%- 70% B over 10 min) to give (56) (26.89 mg, 56.78 μmol, 46.62% yield, 100% purity) as white solid.1H NMR: (400 MHz, CDCI3) δ = 8.02 - 7.93 (m, 1 H), 7.76 (s, 1 H), 6.98 - 6.90 (m, 1 H), 6.62 (br d, J = 10.3 Hz, 1 H), 5.26 - 4.67(m, 1 H), 4.91 - 4.01 (m, 1 H), 3.88 - 3.15 (m, 3H), 2.91 - 2.54 (m, 6H), 1 .95 - 1 .61 (m, 4H), 1 .22 - 1 .02 (m, 3H), 0.87 - 0.78 (m, 2H), 0.72 - 0.35 (m, 4H). LCMS: RT =0.531 min, m / z =474.1 (M+H)+.

[0769] Example 2: Inhibition of PLA2G15 IC50 of the following compounds for PLA2G15 was determined using the 4-nitrophenyl butyrate assay. Specifically, compounds were prediluted starting from 10mM DMSO stocks to obtain 10-point 3- fold dilution series in neat DMSO. 0.1 pL of all resulting samples was diluted 100x to 10pL assay buffer (35mM aq. sodium acetate, pH 4.5, 0.01 % BSA, 0.01 % pluronic F-127) in a clear 384-well flatbottom plate. The dose responses for each compound were performed in duplicate. To both high and low control wells 10pL 1 % DMSO in assay buffer was added. Then, 5pL assay buffer was added to the low control wells or 5pL 24.2pM PLA2G15 (recombinantly produced in HEK293T) solution in PBS to all other wells. 3.5pL neat 4-nitrophenyl butyrate (Sigma) was diluted to 400pL with neat DMSO to obtain a 50mM stock which was stored at -20°C until use. Directly prior to an experiment, 100pL of this substrate stock was diluted to 10mL with assay buffer, and 15pL was added to all wells. The plate was spun at 1000 rpm for T, and then incubated at RT for exactly 45’, followed by another spin at 1000 rpm for T. After adding 30pL stop solution (50 mM aq. Tris, pH 9.0) to all wells, the plate was spun at WOOrpm for T and the absorbance at 405 nm measured in a plate reader (SpectraMax). All absorbances were normalized to high control (100%) and low control (0%) and fitted to a 4-parameter dose-response model with free plateaus in an unweighted fit, yielding the reported IC50S.

[0770] Example 3

[0771] A clear 96-well high content imaging plate was coated with 0.1 % gelatin for 1 h at 37°C. HMC3 cells or HMC3 cells with constitutive PLA2G15 knockout (EMEM supplemented with 10% FCS and 1 % penstrep was used as medium throughout) were washed with PBS, trypsinized with 0.25% trypsin / EDTA for 5’, resuspended in medium and spun at 1500rpm for 4’. After resuspending in 3mL medium, cells were counted in a BioRad TC20 counter according to vendor protocol, and the cells diluted with medium to 75,000 cells / mL. Of this cell stock, 100uL was plated onto the 96-well plate after aspiration of the gelatin. An 8-point 2-fold serial dilution of the compound as described in Example 8 in DMSO prepared from 10mM stocks so that after adding to the well as below, a final top concentration of 10mM was obtained. For amiodarone a similar serial dilution series was added however starting at 4mM top concentration in well. 12uL LipidTox Green was added to 12mL medium. And pass through a 2um filter. After removing the medium on the plate, to individual wells were added 199.5uL medium with LipidTox Green and 0.5uL compound dilution. A well with medium without LipidTox Green was added as negative control. The plates were then incubated for 48h at 37°C. Lysotracker was diluted 1 :333 in medium, and 50uL was added to each well, followed by further incubation of 1 h at 37°C. Cells were washed with PBS and fixed for 10’ at 37°C in 4% PFA, followed by another wash. A mixture of 1 :10,000 CellMask Far Red (ThermoFisher, C10046) and 1 :1000 Dapi (1 mg / mL stock) in 12mL PBS was prepared, of which 100uL was added to each well, followed by incubation for 1 h at 37°C and 2 PBS washes. Images were taken on a Revvity Operetta confocal microscope employing 4 fluorescence channels (Dapi 350nm ex, 430-500 filter; LipidTox Green 495nm ex, 500-550 filter; Lysotracker 577nm ex, 570-650nm filter; CellMask 650nm ex, 655-760nm filter). Quantification was by spot intensity as mean per well.

[0772] Conclusions: The LipidTox assay effectively distinguishes between compounds that induce phospholipid accumulation and those that do not. Amiodarone, used as a control, clearly causes phospholipid accumulation, validating the assay. In contrast, the specific PLA2G15 inhibitors tested did not show a phospholipidosis phenotype in this assay, making them suitable candidates for further development without the associated risk of inducing this condition.

[0773] Example 4

[0774] Lysosomal storage diseases, in particular Niemann Pick disease type C (NPC) are hallmarked by liver malfunction and progressive neurodegeneration in human. The liver phenotype includes enlargement of the liver (hepatomegaly), macrophage infiltration, foam cell formation and storage of glycosphingolipids, contributing to apoptosis of hepatic cells. The loss of neurons in the central nervous system leads to a multitude of neurological symptoms, and most commonly a loss of motor function is observed. Patients regularly experience ataxia, difficulty walking, swallowing problems, loss of muscle tone and tremors. Both the liver and neurological phenotypes are well recapitulated in animal models.

[0775] We identified PLA2G15 as genetic modifier of NPC in cellular models, and we hypothesized that Pla2g15 knockout would alleviate the liver and neurological phenotype in Npc1 KO mice. In the experiments below, we tested the effect of Pla2g15 knockout on liver damage using established biomarkers aspartate transaminase (AST) and alanine transaminase (ALT). Both enzymes are highly expressed in liver, and during liver injury, damaged cells will release both enzymes into the bloodstream. Thus, elevated levels of AST and ALT are commonly used as biomarkers to monitor liver damage. Furthermore, we quantified neurological function using a neurological composite score, where an observer scores mouse performance on six neurological-driven behavioral phenotypes. We observed significant improvements in both the liver damage and neurological composite scoring tests, indicating that knockout of Pla2g15 lessens liver and neurological phenotypes in NPC.

[0776] NEUROLOGICAL COMPOSITE SCORE

[0777] Pla2g15!Balb / C animals were crossed with Npc1m1N / Jheterozygous (HET) animals, and from the resulting offspring, double heterozygous mice were used to obtain the following genotypes (gene order: Npc1m1N / J / Pla2g15): WT / WT, HOM / WT, HOM / HET, HOM / KO and WT / KO. Neurological composite phenotype score (consisting of ledge test, hindlimb clasping, gait, kyphosis, tremor, and grooming scores) was performed on a weekly basis starting at 6 weeks of age. These tests were performed following an established protocol (Davidson et al 2022, PMID 34407999). Higher composite score means worse neurological performance.

[0778] In the Npc1 disease model (HOM / WT), we observed a progressive worsening in the neurological composite score compared to WT / WT starting from week 7, across all domains tested. In mice that lack both Npc1 and Pla2g15 expression (HOM / KO), the neurological phenotype was significantly improved at week 7 compared to HOM / WT. And although slow worsening of the phenotype was observed after week 7, the neurological composite score remained significantly lower compared to HOM / WT at all time points tested. Improvement was found across all domains tested, and was strongest in the gait, tremor and ledge tests. Heterozygous deletion of Pla2g15 (HOM / HET) did not affect performance in the neurological composite score compared to HOM / WT. In conclusion, these results show that deletion of PLA2G15 expression in a NPC1 disease model slows neurological disease progression.

[0779] AST AND ALT PLASMA CONCENTRATION

[0780] At 8 weeks of age (P56 ± 2 days) animals were terminally anesthetized by intraperitoneal injection of Pentobarbital (600 mg / kg) and blood plasma was obtained. AST and ALT levels were determined with a Kit (AST: Cat# 04467493190, Roche; ALT: Cat No 04467388190, Roche) according to International Federation of Clinical Chemistry and Laboratory Medicine (IFCC) with pyridoxal phosphate activation (Roche). Therefore, a kinetic measurement of the enzyme activity with a redox reaction of NADH was performed, using L-Aspartate and 2-Oxoglutarat as substrate for the AST measurement and L-Alanine and 2-Oxoglutarat as substrate for the ALT determination. The two enzyme levels were measured on a Roche Cobas 6000 / c501 analyzer.

[0781] To measure the effect of Pla2g15 KO on liver damage in the NPC1 mouse disease model, we measured the levels of biomarkers aspartate transaminase (AST) and alanine transaminase (ALT). We observed a profound increase in both AST and ALT in the disease model (Npc1 KO). Removal of Pla2g15 in the disease model (double knockout, DKO) strongly reduced AST and ALT levels, indicating reduced liver damage. Knock out of Pla2g15 alone did not influence the levels of either biomarker. Thus, Pla2g15 KO lessens liver damage in Npc1 KO mice.

[0782] Example 5: HMC3 PFO cholesterol accumulation assay

[0783] HMC3 cells were used to assess the effects of PLA2G15 inhibitors on lysosomal cholesterol accumulation. A total of 200,000 HMC3 cells / well were plated into 6-well cell culture plates and treated with DMSO or compound SC-003863 for 7 days at 37°C in a CO2incubator. The medium (MDEM supplemented with 10% FCS and 1 % PenStrep) was refreshed after 4 days. The experiment utilized wildtype (WT), NPC1 -deficient (NPC1-ko), and NPC1 / PLA2G15 double-knockout (NPC1 / PLA2G15- dko) cells, all derived from the HMC3-WT parent line as polyclonal pools generated by CRISPR / Cas9 modification.

[0784] After 7 days of incubation, the cells reached confluence and were reseeded into individual wells of a Phenoplate-96 plate at 15,000 cells / well. The cells were allowed to attach for at least 6 hours, preferably overnight, in 100 pL medium. Subsequently, 1 :1000 v / v Lysotracker Red reagent was added to each well and incubated for 1 hour at 37°C. After incubation, the medium was aspirated, and the cells were washed with PBS. Fixation was performed using 4% PFA in PBS for 10 minutes at 37°C, followed by washing twice with PBS. The cells were permeabilized with 0.1 % saponin in PBS for 10 minutes at room temperature.

[0785] After washing, the cells were stained with Alexa-647 labelled recombinant perfringolysin O (PFO) to detect cholesterol-rich lysosomal membranes. Staining was performed using 60 pL / well of PFO diluted in PBS (1 :1000 from a 0.3 mg / mL stock). After staining, the cells were washed with PBS and counterstained with CellMask Green and DAPI according to the vendor protocol. Following a final wash with PBS, the plates were imaged using an Operetta CLS Imager, employing appropriate channels for Alexa-647 (PFO), DAPI, CellMask Green, and Lysotracker Red. Viability was assessed by counting DAPI-positive nuclei, and individual cells were segmented based on CellMask Green staining. PFO fluorescence was quantified exclusively in Lysotracker Red-positive puncta.

[0786] Baseline cholesterol accumulation was assessed in WT, NPC1 -ko, and NPC1 / PLA2G15-dko cells using the PFO staining assay. NPC1 -ko cells exhibited significant lysosomal cholesterol accumulation compared to WT cells, consistent with the pathological impact of NPC1 deficiency. In NPC1 / PLA2G15- dko cells, cholesterol accumulation was reduced compared to NPC1 -ko cells but remained elevated relative to WT levels, indicating a partial rescue of cholesterol storage in the absence of PLA2G15.

[0787] Further analysis demonstrated the effect of a PLA2G15 inhibitor on lysosomal cholesterol levels in NPC1-ko and NPC1 / PLA2G15-dko cells. The PLA2G15 inhibitor significantly reduced cholesterol accumulation in NPC1-ko cells while having no effect in NPC1 / PLA2G15-dko cells. This indicates that the observed rescue of lysosomal cholesterol accumulation is specifically dependent on PLA2G15 activity.

[0788] These findings highlight the dual role of PLA2G15 inhibition in restoring lysosomal homeostasis and reducing pathological cholesterol accumulation in NPC1 -deficient cells. Furthermore, they underscore the therapeutic potential of targeting PLA2G15 in NPC, providing robust support for the claims and aligning with the proposed mechanisms of BMP regulation and lysosomal function in NPC pathology.

[0789] Example 6: Evaluation ofPLA2G15 Inhibitors in Restoring BMP Levels in Models of Batten Disease and GRN-Mediated Conditions

[0790] This study evaluated the potential of PLA2G15 inhibitors to restore BMP levels in cellular models of Batten disease and GRN-mediated conditions. Cellular models included CLN3- and CLN5-deficient ARPE19 cells generated using CRISPR-Cas9 technology and GRN-deficient bone marrow-derived macrophages (GRN-KO BMDMs) derived from GRN knockout mice. The models were selected to represent lysosomal storage disorders characterized by BMP deficiency, a hallmark of these conditions.

[0791] Cells were grown to confluency for 4 days and treated with 10 pM of PLA2G15 inhibitors SC4395 or SC3863 for an 11-day experimental duration. During the first 2 hours of this period, cells were exposed to the cell cycle inhibitor mitomycin C to synchronize cell division. DMSO-treated cells served as controls. After treatment, BMP levels were quantified using a validated UPLC-MS / MS-based method.

[0792] Cell pellets containing 2 million cells were resuspended in 100 pL water and lysed by sonication. The lysates were transferred to a 96-well plate and spiked with the internal standard d5-36:2-BMP. In-plate protein precipitation was performed by adding 300 pL acetonitrile supplemented with 1 % formic acid. Samples were processed using positive pressure SPE filtration, employing methanol as the eluent (5 min loading followed by 5 min elution). The filtrate was evaporated completely over 2 hours, and the resulting lipid pellet was reconstituted in 100 pL of 10 mM ammonium formate in methanol. A 10 pL aliquot was injected into an ACE3 C18 reversed-phase column mounted on a Vanquish UPLC system and eluted using a gradient of 10 mM ammonium formate in water to 10 mM ammonium formate in methanol. The eluate was analyzed on an inline TSQ Quantiva MS / MS detector in positive mode electron spray ionization, focusing on a predefined library of BMP species with acyl chain profiles of 36:2, 38:5, 40:7, and 44:12. The levels of multiple species of Bis(monoacylglycerol)phosphate (BMP) are shown in the ARPE19 retinal pigment epithelial cell line with wildtype, CLN3ko, or CLN5ko genotype. The BMP levels were significantly reduced in untreated CLN3ko and CLN5ko cells compared to wildtype cells. T reatment with SC4395 or SC3863 restored BMP levels in CLN3ko cells, with SC4395 also partly restoring BMP levels in CLN5ko cells.

[0793] These results demonstrate that PLA2G15 inhibitors can restore BMP levels in cellular models of Batten disease and GRN-mediated conditions, addressing a key biochemical defect in these disorders. Restoration of BMP levels is associated with improved lysosomal function and supports the therapeutic relevance of targeting PLA2G15 as a disease-modifying strategy for Batten disease and GRN-mediated conditions.

Claims

Claims1. A PLA2G15 inhibitor for use in the treatment of a disease characterized by lysosomal dysregulation which is a lysosomal storage disease, HIV, Alzheimer’s disease, Parkinson’s disease, Niemann Pick type C, or a neuronal ceroid lipofuscinosis such as CLN3 disease or Batten disease, CLN5 disease, or GRN frontotemporal dementia, represented by formula (I), or a salt or solvate thereof:(I), wherein X is S or O; wherein Y is CH or N; wherein L is a single bond or a linker moiety; wherein RXYis H, a C1-4 alkyl, a halogen, or a pseudohalogen selected from the group consisting of -CN, -CP, -NC, -OH, -SH, -SeH,-TeH, -OCN, -SCN, -NCS, -SeCN, -TeCN, - N3, -NO, or -NO; wherein RNis a C1-4 alkyl, a C3-6 cycloalkyl, a 5-membered aromatic ring, or a 6- membered aromatic ring; wherein RLcomprises a 3- to 6-membered carbocyclic ring or a 4-membered heterocyclic ring; wherein each 5-membered aromatic ring and 6-membered aromatic ring may be independently substituted with one or more halogens, one or more pseudohalogens selected from the group consisting of -CN, -CP, -NC, -OH, -SH, -SeH,-TeH, -OCN, -SCN, -NCS, -SeCN, -TeCN, -N3, -NO, or -NO, C1-4 alkyls, O-C1-4 alkyls, C3-6 cycloalkyls, or O- C3-6 cycloalkyls; and wherein each C1-4 alkyl and C3-6 cycloalkyl may be independently substituted with one or more halogens or one or more pseudohalogens selected from the group consisting of - CN, -CP, -NC, -OH, -SH, -SeH,-TeH, -OCN, -SCN, -NCS, -SeCN, -TeCN, -N3, -NO, or - NO.

2. The PLA2G15 inhibitor for use according to claim 1 , represented by formula (II):(II), wherein RAand RBare independently H, a C1-4 alkyl, a halogen, or a pseudohalogenselected from the group consisting of -CN, -CP, -NC, -OH, -SH, -SeH,-TeH, -OCN, -SCN, -NCS, -SeCN, -TeCN, -N3, -NO, or -NO, or wherein RAand RBare fused to form a C3-6 cycloalkyl; wherein Z is O or NR’, wherein R’ is H or a C1-4 alkyl; wherein i is 0 or 1 ; and wherein each C1-4 alkyl and C3-6 cycloalkyl may be independently substituted with one or more halogens or one or more pseudohalogens selected from the group consisting of - CN, -CP, -NC, -OH, -SH, -SeH,-TeH, -OCN, -SCN, -NCS, -SeCN, -TeCN, -N3, -NO, or - NO.

3. The PLA2G15 inhibitor for use according to claim 2, wherein i is 1 and Z is NH.

4. The PLA2G15 inhibitor for use according to claim 2 or 3, wherein RAand RBare independentlyH or CH3, preferably wherein RBis H and the PLA2G15 inhibitor is represented by formula (III):(III).

5. The PLA2G15 inhibitor for use according to claim 1 , represented by formula (IV):(IV).

6. The PLA2G15 inhibitor for use according to any one of claims 1 to 5, wherein X is S and Y is CH.

7. The PLA2G15 inhibitor for use according to any one of claims 1 to 6, wherein RLis a saturated or partially saturated 3- to 6-membered carbocyclic ring, wherein the carbocyclic ring may be substituted with one or more halogens, pseudohalogens, C1-4 alkyls, O-C1-4 alkyls, C3-6 cycloalkyls, O-C3-6 cycloalkyls, or phenyls; wherein each phenyl may be independently substituted with one or more halogens, pseudohalogens, C1-4 alkyls, O-C1-4 alkyls, C3-6 cycloalkyls, or O-C3-6 cycloalkyls; andwherein each C1-4 alkyl, or C3-6 cycloalkyl may be independently substituted with one or more halogens or pseudohalogens.

8. The PLA2G15 inhibitor for use according to claim 7, represented by formula (V):

9. The PLA2G15 inhibitor for use according to any one of claims 1 to 6, wherein RLis a phenyl, wherein the phenyl may be independently substituted with one or more halogens, pseudohalogens, C1-4 alkyls, O-C1-4 alkyls, C3-6 cycloalkyls, or O-C3-6 cycloalkyls; and wherein each C1-4 alkyl, or C3-6 cycloalkyl may be independently substituted with one or more halogens or pseudohalogens.

10. The PLA2G15 inhibitor for use according to claim 9, wherein the phenyl is represented by (ph-1):(Ph-1), wherein Rm1and Rm2are independently a C1-4 alkyl, a C3-6 cycloalkyl, a halogen or a pseudohalogen; wherein each C1-4 alkyl or C3-6 cycloalkyl may be independently substituted with one or more halogens or pseudohalogens; and preferably wherein Rm1is t-butyl, cyclopropyl or CF3, and Rm2is F, Cl or cyclopropyl.11 . The PLA2G15 inhibitor for use according to any one of claims 1 to 10, wherein RXYis H or CH3, preferably H.

12. The PLA2G15 inhibitor for use according to any one of claims 1 to 11 , wherein RNis a phenyl substituted with one or more halogens, pseudohalogens, C1-4 alkyls, O-C1-4 alkyls, C3-6 cycloalkyls, or O-C3-6 cycloalkyls; wherein each C1-4 alkyl, or C3-6 cycloalkyl may be independently substituted with one ormore halogens or pseudohalogens; preferably wherein RNis a phenyl substituted with one F, CH3 or O-CH3.

13. The PLA2G15 inhibitor for use according to any one of claims 1 to 11 , wherein RNis a cyclopropyl, a 5-membered azaheterocycle, or a 6-membered azaheterocycle, wherein the azaheterocycle may be substituted with one or more C1-4 alkyls or C3-6 cycloalkyls, preferably wherein the azaheterocycle is monosubstituted on a nitrogen atom.

14. The PLA2G15 inhibitor for use according to any one of claims 1 to 13, represented by formula (VI):(VI).

15. The PLA2G15 inhibitor as defined in claim 1 , represented by any one of formula (1) to (46) and (48) to (56), preferably for use as a medicament, more preferably for use in treating a disease characterized by lysosomal dysregulation which is a lysosomal storage disease, HIV, Alzheimer’s disease, Parkinson’s disease, Niemann Pick type C, or a neuronal ceroid lipofuscinosis such as CLN3 disease or Batten disease, CLN5 disease, or GRN frontotemporal dementia:(3) (4)16. Use of the PLA2G15 inhibitor as defined in any one of claims 1 to 15 for specifically binding and / or inhibiting PLA2G15 in vitro, preferably wherein the PLA2G15 inhibitor does not specifically bind and / or inhibit other phospholipases than PLA2G15.

17. The PLA2G15 inhibitor as defined in any one of claims 1 to 15 for use as a medicament wherein the use comprises specifically binding and / or inhibiting PLA2G15, preferably wherein the PLA2G15 inhibitor does not specifically bind and / or inhibit other phospholipases than PLA2G15.

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