PLA2g15 inhibitors

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] PLA2G15 inhibitors

[0002] Field

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

[0007] Description of the invention

[0008] Inhibitors

[0009] In a first aspect, the invention provides a PLA2G15 inhibitor represented by formula (I), or a salt or solvate thereof: wherein L is -NH-, a -N(CI- alkyl)-, a -C1-4 alkylene-NH- wherein N is bound to S, or a C1-4 alkylene; wherein RS2is a 4- to 6-membered ring, which may be substituted by one or more C1-4 alkyls, O-C1- alkyls, halogens or pseudohalogens; wherein RP2is a C1-4 alkyl, a O-C1-4 alkyl, or a CO-C1-4 alkyl; or wherein RP2is a 3- to 5- membered ring, which may be substituted by one or more C1-4 alkyls, O-C1- alkyls, halogens or pseudohalogens; wherein RN1, RN2, RP1, RP3, RS1and RS3are independently H, a C1-4 alkyl, a O-C1-4 alkyl, a CO-C1-4 alkyl, a halogen, or a pseudohalogen; and wherein each C1-4 alkyl and C1-4 alkylene may be independently substituted with one or more halogens or pseudohalogens.

[0010] 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.

[0011] Inhibitors according to the first or the second 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.

[0012] Wherever embodiments and preferences are disclosed in this application, for example in relation to RP1, RP2, RP3, RN1, RN2, RS1, RS2, RS3, and L, explicit reference is made to both inhibitors according to the first and the second aspect of the invention. In other words, such embodiments and preferences relate to both the inhibitors and the inhibitors for use as a medicament. Relatedly, any embodiment aimed at inhibitors mentioned herein also discloses an embodiment for the inhibitors for use as a medicament. Likewise, such embodiments and preferences may be applied mutatis mutandis to related compositions, uses, therapeutic uses, etc. of the inhibitors according to the invention.

[0013] 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.

[0014] 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.

[0015] A ring is an organic ring consisting of covalently bound ring atoms.

[0016] 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 refer to CH3, CH2OH, CHCh, CF3, etc. In contrast, CH3only refers to an unsubstituted methyl.

[0017] RING RS2

[0018] In embodiments, RS2is a four-, five- or six-membered ring, preferably a five- or a sixmembered ring. Preferably, RS2is a five-membered or a six-membered aromatic ring. In embodiments, RS2is an aromatic ring. Preferably, RS2is a phenyl or a heteroaromatic ring, more preferably a phenyl, or a five-membered or six-membered heteroaromatic ring, most preferably a phenyl of a five-membered heteroaromatic ring.

[0019] In embodiments, RS2is a phenyl, a pyridinyl, an oxazole, an isoxazole, a thiazole, an isothiazole, a pyrrole, a pyrazole or an imidazole. Preferably, the aromatic ring RS2is a phenyl, an oxazole, an isoxazole, a thiazole, an isothiazole, a pyrrole, a pyrazole or an imidazole. More preferably, RS2is a phenyl, a thiazole or an isothiazole. Most preferably, RS2is a phenyl or a thiazole.

[0020] In embodiments, RS2is a carbocyclic ring. A carbocyclic ring is a ring wherein each ring atom is a carbon atom. Preferably, RS2is a phenyl, a cyclobutyl, a cyclopentyl or a cyclohexyl.

[0021] In embodiments, RS2is an aliphatic ring. Preferably, RS2is a cyclobutyl, a cyclopentyl or a cyclohexyl. More preferably, RS2is a cyclopentyl or a cyclohexyl.

[0022] In embodiments, RS2is a saturated or partially saturated ring. Preferably, RS2is a saturated or partially saturated aliphatic ring, more preferably a cyclobutyl, a cyclopentyl or a cyclohexyl.

[0023] In embodiments, RS2is a ring comprising a N, O or S ring atom. Preferably, RS2is a ring comprising 1 or 2 N, O or S ring atoms, more preferably 2 N, O or S ring atoms.

[0024] In embodiments, RS2is an unsubstituted ring, meaning that the only exocyclic non-hydrogen atom attached to the ring is the carbon to which RS1and RS3are attached. Accordingly, n substitutions mean that the ring is attached to n exocyclic non-hydrogen atoms apart from said carbon atom. 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.

[0025] In embodiments, RS2is a ring substituted by one or more C1-4 alkyls, O-C1-4 alkyls, halogens or pseudohalogens, wherein each C1-4 alkyl may be independently substituted with one or more halogens or pseudohalogens.

[0026] In embodiments, RS2is an unsubstituted ring or a ring substituted by one or more C1-4 alkyls, O-C1-4 alkyls or halogens, preferably a substituted ring, wherein each C1-4 alkyl may be independently substituted with one or more halogens. Preferably, the C1-4 alkyl is a C1-3 alkyl. More preferably, the C1-4 alkyl is a C1-2 alkyl. Even more preferably, the C1-4 alkyl is a methyl. Most preferably, the C1-4 alkyl is CH3.

[0027] In embodiments, RS2is an unsubstituted ring or a ring substituted by one or more C1-4 alkyls or halogens, preferably a substituted ring, wherein each C1-4 alkyl may be independently substituted with one or more halogens. Preferably, the C1-4 alkyl is a C1-3 alkyl. More preferably, the C1-4 alkyl is a C1-2 alkyl. Even more preferably, the C1-4 alkyl is a methyl. Most preferably, the C1- alkyl is CH3.

[0028] In embodiments, RS2is an unsubstituted ring or a ring substituted by one or more C1-4 alkyls or fluorine atoms (fluors), preferably a substituted ring, wherein each C1-4 alkyl may be independently substituted with one or fluors. Preferably, the C1-4 alkyl is a C1-3 alkyl. More preferably, the C1-4 alkyl is a C1-2 alkyl. Even more preferably, the C1-4 alkyl is a methyl. Most preferably, the C1-4 alkyl is CH3.

[0029] In embodiments, RS2is a ring substituted with 4, 3, 2 or 1 CH3 or F, preferably with 3, 2 or 1 CH3 or F, more preferably with 2 or 1 CH3 or F.

[0030] In embodiments, RS2is a ring substituted with 4, 3, 2 or 1 F, preferably with 3, 2 or 1 F, more preferably with 2 or 1 F.

[0031] In embodiments, RS2is a ring substituted with 4, 3, 2 or 1 CH3, preferably with 3, 2 or 1 CH3, more preferably with 2 or 1 CH3.

[0032] GROUPS RS1AND RS3

[0033] In embodiments, RS1and RS3are independently H or a C1-4 alkyl, preferably H or a C1-3 alkyl, more preferably H or a C1-2 alkyl, even more preferably H or a methyl, most preferably H or CH3.

[0034] In embodiments, RS1is a C1-4 alkyl and RS3is a C1-4 alkyl. Preferably, both C1-4 alkyls are C1- 3 alkyls. More preferably, both C1-4 alkyls are C1-2 alkyls. Even more preferably, both C1-4 alkyls are a methyl. Most preferably, both C1-4 alkyls are CH3.

[0035] In embodiments, RS1is a C1-4 alkyl and RS3is H. Preferably, the C1-4 alkyl is a C1-3 alkyl. More preferably, the C1-4 alkyl is a C1-2 alkyl. Even more preferably, the C1-4 alkyl is a methyl. Most preferably, the C1-4 alkyl is CH3.

[0036] In embodiments, RS1is H and RS3is a C1-4 alkyl. Preferably, the C1-4 alkyl is a C1-3 alkyl. More preferably, the C1-4 alkyl is a C1-2 alkyl. Even more preferably, the C1-4 alkyl is methyl. Even more preferably, the C1-4 alkyl is a methyl. Most preferably, the C1-4 alkyl is CH3.

[0037] In embodiments, RS1is H and RS3is H.

[0038] GROUPS RN1AND RN2

[0039] In embodiments, RN1and RN2are independently H or a C1-4 alkyl, preferably H or a C1-3 alkyl, more preferably H or a C1-2 alkyl, even more preferably H or methyl, most preferably H or CH3. In embodiments, RN1and RN2are independently a C1-4 alkyl, preferably a C1-3 alkyl, more preferably a C1-2 alkyl, even more preferably methyl, most preferably CH3.

[0040] In embodiments, RN1is a C1-4 alkyl and RN2is a C1-4 alkyl. Preferably, both C1-4 alkyls are C1-3 alkyls. More preferably, both C1-4 alkyls are C1-2 alkyls. Even more preferably, both C1-4 alkyls are a methyl. Most preferably, both C1-4 alkyls are CH3.

[0041] In embodiments, RN1is a C1-4 alkyl and RN2is H. Preferably, the C1-4 alkyl is a C1-3 alkyl. More preferably, the C1-4 alkyl is a C1-2 alkyl. Even more preferably, the C1-4 alkyl is methyl. Most preferably, the C1-4 alkyl is CH3.

[0042] In embodiments, RN1is H and RN2is a C1-4 alkyl. Preferably, the C1-4 alkyl is a C1-3 alkyl. More preferably, the C1-4 alkyl is a C1-2 alkyl. Even more preferably, the C1-4 alkyl is methyl. Most preferably, the C1-4 alkyl is CH3.

[0043] In embodiments, RN1is H and RN2is H.

[0044] LINKER L

[0045] In embodiments, L is -NH-, a -N(CI- alkyl)-, a -C1-4 alkylene-NH- wherein N is bound to S.

[0046] In preferred embodiments, L is -NH-. In some embodiments, L is -NH- wherein N is bound to S. In embodiments, L is -NH- or a -N(CI- alkyl)-, wherein the C1-4 alkyl is preferably a C1-3 alkyl, more preferably a C1-2 alkyl, even more preferably a methyl, most preferably CH3.

[0047] GROUP RP2

[0048] In some embodiments, RP2is a C1-4 alkyl, a O-C1-4 alkyl, or a CO-C1-4 alkyl; or wherein RP2is a 3- to 5-membered heterocyclic or carbocyclic ring, or a benzamide (C6H5CONH2) which may be substituted by one or more C1-4 alkyls, O-C1-4 alkyls, 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 -NO2.

[0049] In embodiments, RP2is a C1-4 alkyl, a O-C1-4 alkyl, or a CO-C1-4 alkyl, wherein C1-4 alkyl may be unsubstituted or independently substituted with one or more halogens or pseudohalogens. Preferably, the C1-4 alkyl is a C1-3 alkyl, more preferably a C1-2 alkyl, most preferably a methyl.

[0050] In embodiments, RP2is a C1-4 alkyl, or a O-C1-4 alkyl, wherein C1-4 alkyl may be unsubstituted or independently substituted with one or more halogens or pseudohalogens. Preferably, the C-i- 4 alkyl is a C1-3 alkyl, more preferably a C1-2 alkyl, most preferably a methyl. In embodiments, RP2is a C1-4 alkyl, a O-C1- alkyl, or a CO-C1-4 alkyl, wherein C1-4 alkyl may be unsubstituted or independently substituted with one or more fluors. Preferably, the C1-4 alkyl is a C1-3 alkyl, more preferably a C1-2 alkyl, most preferably a methyl.

[0051] In embodiments, RP2is a C1-4 alkyl, or a O-C1-4 alkyl, wherein C1-4 alkyl may be unsubstituted or independently substituted with one or more fluors. Preferably, the C1-4 alkyl is a C1-3 alkyl, more preferably a C1-2 alkyl, most preferably a methyl.

[0052] In embodiments, RP2is an aliphatic ring. Preferably, RP2is a cyclopropyl, a cyclobutyl or a cyclopentyl. Preferably, the aliphatic ring is unsubstituted or substituted with one or more fluors or methyl, more preferably with one or more F, CH3 or CF3. Alternatively, the aliphatic ring is unsubstituted.

[0053] In embodiments, RP2is a cyclopropyl which is unsubstituted or substituted with a single fluor.

[0054] In embodiments, RP2is a C1-4 alkyl, an O-C1-4 alkyl or a cyclopropyl, wherein C1-4 alkyl and cyclopropyl may be unsubstituted or independently substituted with one or more halogens or pseudohalogens. Preferably, the C1-4 alkyl is a C1-3 alkyl, more preferably a C1-2 alkyl, even more preferably a methyl.

[0055] In embodiments, RP2is a C1-4 alkyl, an O-C1-4 alkyl or a cyclopropyl, wherein C1-4 alkyl and cyclopropyl may be unsubstituted or independently substituted with one or more halogens. Preferably, the C1-4 alkyl is a C1-3 alkyl, more preferably a C1-2 alkyl, even more preferably a methyl.

[0056] In embodiments, RP2is a C1-4 alkyl, an O-C1-4 alkyl or a cyclopropyl, wherein C1-4 alkyl and cyclopropyl may be unsubstituted or independently substituted with one or more fluors. Preferably, the C1-4 alkyl is a C1-3 alkyl, more preferably a C1-2 alkyl, even more preferably a methyl.

[0057] In embodiments, RP2is a C1-4 alkyl, an O-C1-4 alkyl or a cyclopropyl, wherein C1-4 alkyl and cyclopropyl may be unsubstituted or independently substituted with one or more fluors. Preferably, the C1-4 alkyl is a C1-3 alkyl, more preferably a C1-2 alkyl, even more preferably a methyl.

[0058] In embodiments, RP2is a heteroaromatic ring. Preferably, RP2is an aza-aromatic ring. More preferably, RP2is a pyrazole, an imidazole, a pyrrole, an oxazole, an isoxazole, a thiazole or a isothiazole. Even more preferably, RP2is a pyrazole, an imidazole or a pyrrole. Most preferably, RP2is a pyrazole. Preferably, the heteroaromatic ring is unsubstituted or substituted with one or more fluors or methyl, more preferably with one or more F, CH3 or CF3. In embodiments, RP2is a pyrazole, an imidazole or a pyrrole, which is unsubstituted or substituted with one or more methyls, preferably N-substituted with one or more methyls (“N- methylated). Preferably, each methyl is CH3.

[0059] In embodiments, RP2is a pyrazole, an imidazole or a pyrrole, which is unsubstituted or substituted with one methyl, preferably N-substituted with one methyl. Preferably, the methyl is CH3.

[0060] In embodiments RP2is 1-methyl-3-pyrazolyl.

[0061] In embodiments, RP2is a 1-methyl-4-pyrazolylln embodiments RP2is unsubstituted cyclopropyl, 2-fluoro-cyclopropyl or 1 -methyl-3-pyrazolyl.

[0062] In embodiments RP2is CH3, CHF2, CF3, OCH(CH3)2, OCHF2, unsubstituted cyclopropyl, 2- fluoro-cyclopropyl or 1-methyl-3-pyrazolyl.

[0063] In embodiments, RP2 is a substituted benzamide according to: wherein RPP1, RPP2, and RPP3are individually selected from H, C1-4 alkyl, halogens, one or more pseudohalogens selected from the group consisting of -ON, -CP, -NC, -OH, -SH, - SeH,-TeH, -OCN, -SCN, -NCS, -SeCN, -TeCN, -N3, -NO, or -NO2.

[0064] In some embodiments, RPP1, RPP2, and RPP3are all H.

[0065] In some embodiments, RPP1is selected from a pseudohalogens selected from the group consisting of -CN, -CP, -NC, -OH, -SH, -SeH,-TeH, -OCN, -SCN, -NCS, -SeCN, -TeCN, -N3, - NO, or -NO2, RPP2is selected from a halogen, and RPP3is a C1-4 alkyl.

[0066] In some embodiments, RPP1 is an -OH, RPP2 is a chloride and RPP3 is a tert-butyl group.

[0067] GROUPS RP1AND RP3

[0068] In embodiments, RP1and RP3are independently H, a C1-4 alkyl, a O-C1-4 alkyl, a halogen, or a pseudohalogen. Preferably, the C1-4 alkyl is a C1-3 alkyl, more preferably a C1-2 alkyl, even more preferably a methyl.

[0069] In embodiments, RP1and RP3are independently H, a C1-4 alkyl, a O-C1-4 alkyl, fluor, or a pseudohalogen. Preferably, the C1-4 alkyl is a C1-3 alkyl, more preferably a C1-2 alkyl, even more preferably a methyl. In embodiments, RP1and RP3are independently H, a C1-4 alkyl, a O-C1-4 alkyl, fluor, or a cyano group (CN). Preferably, the C1-4 alkyl is a C1-3 alkyl, more preferably a C1-2 alkyl, even more preferably a methyl.

[0070] In embodiments, RP1and RP3are independently H, a C1-4 alkyl, a O-C1-4 alkyl, fluor, or a cyano group (CN). Preferably, the C1-4 alkyl is CH3, CH2CF3 or CF3.

[0071] In embodiments, RP1and RP3are independently H, CH3, O-CH3, CH2CF3, CF3, F, or CN.

[0072] In embodiments, RP1is F and RP3is H; or RP1is H and RP3is F; or RP1is H and RP3is H; or RP1is F and RP3is F.

[0073] PREFERRED INHIBITORS In embodiments, the PLA2G15 inhibitor is represented by any one of formulae (II) to (LV), wherein RS2is an optional substitution with one or more CH3 or F, preferably with one or two CH3or F:

[0074] (VI) (VII)

[0075] (XXX) (XXXI)

[0076] (XLI I) (XLI 11) In embodiments, the PLA2G15 inhibitor is represented by any one of the following formulae:

[0077] (11) (12)

[0078]

[0079] (23) (24)

[0080]

[0081] (35) (36)

[0082]

[0083] (41) (42).

[0084] In embodiments, the PLA2G15 inhibitor is represented by any one of formulae (1) to (42), or to (41), or to (40), or to (39), or to (38), or to (37), or to (36), or to (35), or to (34), or to (33), or to (32), or to (31), or to (30), or to (29), or to (28), or to (27), or to (26), or to (25), or to (24), or to (23), or to (22), or to (21), or to (20), or to (19), or to (18), or to (17), or to (16), or to (15), or to (14), or to (13), or to (12), or to (11), or to (10), or to (9), or to (8), or to (7), or to (6), or to (5), or to (4), or to (3), or to (2).

[0085] ISOMERY

[0086] 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 h a Ifch air-forms; and combinations thereof, hereinafter collectively referred to as “isomers” (or “isomeric forms”).

[0087] 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.

[0088] 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.

[0089] SALTS

[0090] A PLA2G15 inhibitor according to the invention may be present as a pharmaceutically acceptable salt.

[0091] Examples of pharmaceutically acceptable salts are discussed in Berge et al., 1977, “Pharmaceutically Acceptable Salts,” J. Pharm. Sci., Vol. 66, pp. 1 -19.

[0092] 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., NH+). Examples of suitable organic cations include, but are not limited to substituted ammonium ions (e.g., NHsR*, NH2R2+, NHR3+, NR+), for example, where each R is independently linear or branched saturated Ci-isalkyl, Cs-scycloalkyl, Cs-acycloalkyl-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)+.

[0093] If a PLA2G15 inhibitor is cationic, 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.

[0094] 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+, -NH3+), 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. 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.

[0095] 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.

[0096] SOLVATES AND HYDRATES

[0097] A PLA2G15 inhibitor according to the invention may be present as a pharmaceutically acceptable solvate or hydrate.

[0098] 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.

[0099] PLA2G15

[0100] STRUCTURE OF PLA2G15

[0101] 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.

[0102] All embodiments disclosed below relating to PLA2G15 proteins or genes may be applied accordingly to inhibitors and compositions according to the invention.

[0103] 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).

[0104] 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 .

[0105] 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%,

[0106] 82.5%, 83%, 83.5%, 84%, 84.5%, 85%, 85.5%, 86%, 86.5%, 87%, 87.5%, 88%, 88.5%, 89%,

[0107] 89.5%, 90%, 90.5%, 91 %, 91.5%, 92%, 92.5%, 93%, 93.5%, 94%, 94.5%, 95%, 95.5%, 96%,

[0108] 96.5%, 97%, 97.5%, 98%, 98.5%, 99%, 99.5%, or 100% sequence similarity with SEQ ID NO:

[0109] 1 or 2, preferably with SEQ ID NO:1 .

[0110] 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 .

[0111] 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 .

[0112] 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 31 1 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.

[0113] In embodiments, PLA2G15 has a length of 412 amino acids, or 318 amino acids.

[0114] 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.

[0115] ACTIVITY OF PLA2G15

[0116] 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.

[0117] 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.

[0118] 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.

[0119] 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 (paranitrophenyl 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.

[0120] 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. 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.

[0121] 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.

[0122] 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.

[0123] 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.

[0124] In embodiments, a phospholipid in the aspects above is a phosphatidylcholine, a phosphatidylethanolamine, a phosphatidylglycerol, or a phosphatidylserine.

[0125] 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.

[0126] In embodiments, a phospholipid in the aspects above is not a phosphatidylinositol or a sphingomyelin. 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.

[0127] In embodiments, a catalytic activity of PLA2G15 comprises a transacylase activity, wherein said acceptor compound is N-acetyl-sphingosine.

[0128] 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.

[0129] CLINICAL RELEVANCE OF PLA2G15

[0130] 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.

[0131] 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.

[0132] 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.

[0133] 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).

[0134] 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.

[0135] 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 (16). 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 (33).

[0136] Therefore, increasing BMP levels in a wide class of diseases as outlined above is expected to have therapeutic impact.

[0137] 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.

[0138] Therapy DISEASES

[0139] In an aspect, the invention provides a PLA2G15 inhibitor according to the invention or a composition according to the invention, for use as a medicament. In embodiments, the PLA2G15 inhibitor or the composition is 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.

[0140] 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.

[0141] 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.

[0142] 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.

[0143] 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.

[0144] In a specific embodiment, said disease is a sphingolipidosis. Sphingolipidosis is characterized by a disturbance of the sphingolipid metabolism. 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 (32).

[0145] 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 (38). 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.

[0146] 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.

[0147] In a specific embodiment, said sphingolipidosis is a mucopolysaccharidosis (MPS), including, MPS I (Hurler syndrome, MPS II (Hunter syndrome) , MPS IIIA (Sanfilippo syndrome), MPS IIIB (Sanfilippo syndrome), MPS IIIC (Sanfilippo syndrome), MPS HID (Sanfilippo syndrome), MPS VI (Maroteaux-Lamy syndrome), MPS VII (Sly syndrome).

[0148] In a specific embodiment, said sphingolipidosis is a Mucolipidosis, including Mucolipidosis II (l-cell disease), Mucolipidosis III (pseudo-Hurler polydystrophy) and Mucolipidosis IV.

[0149] In a specific embodiment, said disease is glycoproteinosis, including galactosialidosis, mannosidosis, sialidosis.

[0150] In a specific embodiment, said sphingolipidosis is a Neuronal ceroid lipofuscinosis (NCL), including NCL 3 (Batten disease), NCL 10 and Hereditary spastic paraplegia (HSP).

[0151] In a specific embodiment, said disease is phospholipidosis.

[0152] In a specific embodiment, said disease is Alzheimer disease.

[0153] In a specific embodiment, said disease is Parkinson’s disease. In preferred embodiments said condition is selected from neuronal ceroid lipofuscinosis (NCL), CLN3 Batten, CLN5 Batten, GRN , frontotemporal dementia and Niemann Pick disease, preferably of type C.

[0154] In specific embodiments, said condition is selected from the diseases mentioned in Table 1 .

[0155] In a preferred embodiment, said disease is Niemann Pick type C (NPC).

[0156] 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.

[0157] 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).

[0158] NPC disease is characterized by the secondary accumulation of (glyco)-sphingolipids and therefore considered a member of a group of diseases called shingolipidoses (32).

[0159] 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.

[0160] 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.

[0161] 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.

[0162] 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.

[0163] 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.

[0164] 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.

[0165] 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.

[0166] 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 .

[0167] 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. 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:

[0168] — 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,

[0169] 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

[0170] — 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,

[0171] 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

[0172] — 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,

[0173] 6.5, 7, 7.5, 8, 8.5, 9, 9.5, 10, 15, 20, 25, 30, 35, 40, 45, or 50; and / or

[0174] — 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,

[0175] 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

[0176] — 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

[0177] 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

[0178] — 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, 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.

[0179] 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.

[0180] 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).

[0181] 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).

[0182] 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.

[0183] 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.

[0184] 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).

[0185] TREATMENT OPTIONS

[0186] 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.

[0187] 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.

[0188] A medicament according to the invention, may be administered separately, sequentially or simultaneously in combination with another medicaments.

[0189] 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.

[0190] 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 parameter(s) 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.

[0191] 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.

[0192] 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.

[0193] 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.

[0194] 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.

[0195] 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.

[0196] Compositions

[0197] 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.

[0198] 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.

[0199] 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.

[0200] 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.

[0201] 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.

[0202] 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.

[0203] 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.

[0204] 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.

[0205] 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.

[0206] 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. A composition according to the invention may be formulated for transmucosal ortransdermal 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.

[0207] 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.

[0208] Uses

[0209] 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.

[0210] 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.

[0211] 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.

[0212] 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.

[0213] 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.

[0214] 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.

[0215] 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.

[0216] In embodiments, the uses above are for use in vitro.

[0217] In embodiments, the uses above are for use in vivo.

[0218] 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.

[0219] Definitions

[0220] 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. 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.

[0221] As used herein, the singular forms 'a', 'an', and 'the' include both singular and plural referents unless the context clearly dictates otherwise.

[0222] 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.

[0223] The recitation of numerical ranges by endpoints includes all numbers and fractions subsumed within the respective ranges, as well as the recited endpoints.

[0224] 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.

[0225] A concentration is preferably a molar concentration, preferably a molar concentration per weight or per volume, most preferably measured under physiological conditions.

[0226] 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.

[0227] An organelle is preferably a lysosome or an endosome, more preferably a lysosome. An endosome is preferably a late endosome.

[0228] 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.

[0229] 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. 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.

[0230] 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.

[0231] 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.

[0232] 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.

[0233] 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.

[0234] 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). 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%.

[0235] 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).

[0236] 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.

[0237] 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). 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-EBL The aforementioned parameters are the default parameters for a Global Pairwise Sequence alignment of nucleotide sequences (along with no penalty for end gaps).

[0238] 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 He, 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.

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[0286] Examples

[0287] The following examples are offered for illustrative purposes only, and are not intended to limit the scope of the present invention in any way.

[0288] Example 1: Synthesis of compounds

[0289] EXAMPLE 1.1: SYNTHESIS OF (1)

[0290] Scheme 1 : synthesis of (1)

[0291] Synthesis of compound 3, (scheme 1)

[0292] To a solution of compound 1 (135.61 mg, 649.99 pmol, 1 eq) in DCM (2 mL) was added pyridine (154.24 mg, 1.95 mmol, 157.39 pL, 3 eq) and compound 2 (200 mg, 649.99 pmol, 1 eq, HCI). The mixture was stirred at 25 °C for 1 h. The mixture was concentrated at reduced pressure to give a residue. The residue was purified by silica gel chromatography (100-200 mesh silica gel, Petroleum ether / Ethyl acetate = 5 / 1 , 3 / 1. TLC (Petroleum ether: Ethyl acetate = 1 : 1 ; Rf = 0.43)) to give compound 3 (160 mg, 360.84 pmol, 55.51 % yield) as brown oil. LCMS: RT = 0.512 min, m / z = 444.1 (M+H)+.

[0293] Synthesis of compound 4, (scheme 1)

[0294] To a solution of compound 3 (160 mg, 360.84 pmol, 1 eq) in DMF (2 mL) was added NaH (17.32 mg, 433.00 pmol, 60% purity, 1 .2 eq) under 0 °C. The mixture was stirred at 0 °C for 0.5 h, the SEM-CI (66.17 mg, 396.92 pmol, 70.25 pL, 1.1 eq) was added into the mixture, and stirred at 25 °C for 1 h. The mixture was added into saturated NH4CI (5 mL) slowly at 0 °C with stirring, the resulting was extracted with EtOAc (10 mL*2). The combined organic layer was washed with brine (10 mL*2), dried over Na2SO4, filtered and concentrated at reduced pressure to give a residue. The residue was purified by silica gel chromatography (100-200 mesh silica gel, Petroleum ether / Ethyl acetate = 3 / 1 , 1 / 1. TLC (Petroleum ether: Ethyl acetate = 1 : 1 ; Rf = 0.43)) to give compound 4 (200 mg, crude) as brown oil. LCMS: RT = 0.535 min, m / z = 574.2(M+H)+.

[0295] Synthesis of compound 5, (scheme 1)

[0296] To a solution of compound 4 (200 mg, 348.63 pmol, 1 eq) in DMF (2 mL) was added NaH (15.34 mg, 383.49 pmol, 60% purity, 1.1 eq) under N2 at 0 °C, the mixture was stirred at 0 °C for 0.25 h. Then Mel (49.48 mg, 348.63 pmol, 21.70 pL, 1 eq) was added, the mixture was stirred at 25 °C for 1 h. The reaction mixture was quenched by water (20 mL), the resulting mixture was extracted with ethyl acetate (20 mL*3), and the combined organic phase was washed with brine (20 mL*3), dried with anhydrous Na2SO4, filtered and the filtrate was concentrated in vacuum to give residue. The residue was purified by silica gel chromatography (100-200 mesh silica gel, Petroleum ether / Ethyl acetate = 5 / 1 , 3 / 1 . TLC (Petroleum ether: Ethyl acetate = 1 : 1 ; Rf = 0.66)) to give compound 5 (200 mg, crude) as brown oil. LCMS: RT = 0.654 min, m / z = 588.2 (M+H)+.

[0297] Synthesis of (1), (scheme 1)

[0298] To a solution of compound 5 (200 mg, 340.31 pmol, 1 eq) in DCM (2 mL) was added TFA (2 mL). The mixture was stirred at 25 °C for 1 h. The reaction mixture was diluted with ethyl acetate (20 mL) and poured into H2O (50 mL) slowly in portions. Then the pH of the mixture was adjusted to 6-7 with NaHCOs. The resulting mixture was extracted with ethyl acetate (30 mL*3), the combined organic phase was dried with anhydrous Na2SO4, filtered and concentrated in vacuum to give residue. The residue was purified by reversed -phase HPLC (column: Welch Xtimate C18 150*25 mm*5um; mobile phase: [water (FA) -ACN]; gradient: 43%-63% B over 15 min) to give (1) as white solid (45 mg).

[0299] The white solid was purified by SFC (Rt = 1 .505 min and Rt = 1 .620 min) (column: DAICEL CHIRALPAK AD (250 mm*30 mm, 10um); mobile phase: [CO2-i-PrOH (0.1 %NH3H2O)]; B%: 20%, isocratic elution mode) to give (1)-peak1 (20.35 mg, 44.49 pmol, 13.07% yield, 100% purity) as white solid.1H NMR: (400 MHz, CD3OD) 5 = 7.72 (t, J = 7.5 Hz, 1 H), 7.68 - 7.63 (m, 2H), 7.61 (s, 2H), 7.34 - 7.28 (m, 1 H), 7.18 - 7.12 (m, 1 H), 7.05 (t, J = 9.2 Hz, 1 H), 6.14 (br s, 1 H), 4.97 - 4.85 (m, 1 H), 3.16 (s, 3H), 2.37 (s, 3H), 1 .91 (d, J = 7.1 Hz, 3H); LCMS: RT = 0.525 min, m / z = 458.1 (M+H)+; SFC: RT = 1 .508 min

[0300] And to give (1)-peak2 (22.53 mg, 49.25 pmol, 14.47% yield, 100% purity) as white solid.1H NMR: (400 MHz, CD3OD) 5 = 7.74 (t, J = 7.0 Hz, 1 H), 7.68 - 7.63 (m, 2H), 7.61 (br s, 2H), 7.34 - 7.28 (m, 1 H), 7.20 - 7.14 (m, 1 H), 7.06 (t, J = 9.3 Hz, 1 H), 5.74 (br s, 1 H), 4.98 - 4.86 (m, 1 H), 3.16 (s, 3H), 2.39 (s, 3H), 1 .94 (d, J = 7.1 Hz, 3H); LCMS: RT = 0.526 min, m / z = 458.1 (M+H)+;

[0301] SFC: RT = 1.622 min.

[0302] EXAMPLE 1.2: SYNTHESIS OF (2)

[0303] Scheme 2: synthesis of (2)

[0304] Synthesis of compound 2, (scheme 2)

[0305] A mixture of compound 1 (500 mg, 3.67 mmol, 1 eq), TosCI (840.23 mg, 4.41 mmol, 1.2 eq) in pyridine (0.5 mL) and DCM (0.5 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 reaction mixture was cooled to room temperature. EtOAc (20 mL) and water (20 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 compound 2 (200 mg, crude) as white solid.1H NMR: (400 MHz, CDCh) 6 = 7.80 (d, J = 8.3 Hz, 2H), 7.37 (d, J = 8.4 Hz, 2H), 3.92 (s, 2H), 2.58 - 2.48 (m, 1 H), 2.47 (s, 3H), 2.45 - 2.38 (m, 1 H), 2.31 - 2.22 (m, 2H), 1 .24 (s, 3H).

[0306] Synthesis of compound 4, (scheme 2)

[0307] A mixture of compound 2 (200 mg, 688.88 pmol, 1 eq), compound 3 (210 mg, 1 .84 mmol, 2.67 eq) in DMF (2 mL) was degassed and purged with N2 for 3 times, and then the mixture was stirred at 60 °C for 16 h under N2 atmosphere. The reaction mixture was cooled to room temperature. 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 compound 4 (100 mg, crude) as white solid.1H NMR: (400 MHz, CDCh) 6 = 4.22 (s, 2H), 2.41 - 2.37 (m, 2H), 2.34 (s, 3H), 2.29 - 2.24 (m, 2H), 1 .21 (s, 3H). Synthesis of compound 5, (scheme 2)

[0308] To a mixture of compound 4 (100 mg, 514.82 pmol, 1 eq), NCS (274.98 mg, 2.06 mmol, 4 eq), HCI (2 M, 514.82 pL, 2 eq) in ACN (2 mL) was degassed and purged with N2 for 3 times, and then the mixture was stirred at 0 °C for 2 h under N2 atmosphere. EtOAc (10 mL) and water (10 mL) were added and layers were separated. The aqueous phase was extracted with EtOAc (5 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 column chromatography (silica gel, Petroleum ether / Ethyl acetate = 1 : 0 to 3: 1 , TLC: Petroleum ether: Ethyl acetate = 5: 1 , Rf = 0.47) to give compound 5 (100 mg, crude) as white solid.1H NMR: (400 MHz, CDCI3) 6 = 4.02 (s, 2H), 2.85 - 2.74 (m, 2H), 2.62 - 2.52 (m, 2H), 1 .60 (s, 3H)

[0309] Synthesis of compound 7, (scheme 2)

[0310] To a solution of compound 6 (1 .35 g, 4.33 mmol, 1 eq) and 4, 4, 5, 5-tetramethyl-2-(4, 4, 5, 5-tetramethyl- 1 , 3, 2-dioxaborolan-2-yl)-1 , 3, 2-dioxaborolane (2.20 g, 8.65 mmol, 2 eq) in dioxane (15 mL) was added KOAc (848.98 mg, 8.65 mmol, 2 eq), Pd(dppf)Cl2 (158.24 mg, 216.26 pmol, 0.05 eq) under N2, the mixture was stirred at 80 °C for 16 h under N2 atmosphere . The reaction mixture was diluted with ethyl acetate (100 mL). The organic layer was washed with brine (100 mL), 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, dichloromethane: methanol = 1 : 0 to 10: 1 ; TLC: (dichloromethane: methanol = 10: 1), Rf = 0.41) to give compound 7 (660 mg, crude) as brown gum. LCMS: RT = 0.477 min, m / z = 360.1 (M+H)+.

[0311] Synthesis of compound 8, (scheme 2)

[0312] To a solution of compound 7 (610 mg, 1 .70 mmol, 1 eq) in THF (12 mL) was added NaOH (1 M, 5.09 mL, 3 eq) and H2O2 (2.99 g, 26.37 mmol, 2.53 mL, 30% purity, 15.53 eq) at 0 °C, the mixture was stirred at 0 °C for 1 h. The reaction mixture was diluted with ethyl acetate (100 mL). The organic layer was quenched with saturated Na2SO3 aqueous solution (50 mL). Then the pH of the aqueous phase was adjusted to 6 with 1 M HCI aqueous solution, the resulting mixture was extracted with ethyl acetate (50 mL*2). The combined organic phase was washed with brine (30 mL), dried over Na2SO4, filtered and the filtrate was concentrated under vacuum to give compound 8 (420 mg, crude) as brown gum. LCMS: RT = 0.318 min, m / z = 250.0 (M+H)+.

[0313] Synthesis of compound 9, (scheme 2)

[0314] To a solution of compound 8 (290 mg, 1.16 mmol, 1 eq) in DCM (15 mL) was added KOH (1 .96 g, 6.98 mmol, 20% purity, 6 eq) and [bromo(difluoro) methyl]-trimethyl-silane (472.66 mg, 2.33 mmol, 2 eq) at 0 °C, the mixture was stirred at 20 °C for 16 h. The mixture was concentrated under vacuum to remove DCM. Then 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 a residue. The residue was purified by column chromatography (100 mesh silica gel, petroleum ether: ethyl acetate = 1 : 0 to 0: 1 ; TLC (petroleum ether: ethyl acetate = 0: 1), Rf = 0.48) to give a residue compound 9 (140 mg, crude) as yellow solid. LCMS: RT = 0.405 min, m / z = 300.0 (M+H)+.

[0315] Synthesis of compound 10, (scheme 2)

[0316] A mixture of compound 9 (140 mg, 467.87 pmol, 1 eq), Fe (130.65 mg, 2.34 mmol, 5 eq), NH4CI (250.26 mg, 4.68 mmol, 10 eq) in EtOH (5 mL) and H2O (1 mL) was degassed and purged with N2 for 3 times, and then the mixture was stirred at 80 °C for 2 h under N2 atmosphere. The reaction mixture was cooled to room temperature. 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 compound 10 (100 mg, crude) as white solid. LCMS: RT = 0.405 min, m / z = 300.0 (M+H)+.

[0317] Synthesis of (2), (scheme 2)

[0318] A mixture of compound 5 (100 mg, 457.35 pmol, 3.08 eq), compound 10 (40 mg, 148.56 pmol, 1 eq), pyridine (35.25 mg, 445.69 pmol, 35.97 pL, 3 eq) in DCM (1 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 reaction mixture was cooled to room temperature. EtOAc (40 mL) and water (40 mL) were added and layers were separated. The aqueous phase was extracted with EtOAc (30 mL x 2). Combined extracts were washed with brine (60 mL), dried over Na2SO4, filtered, and concentrated under 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: 40%- 70% B over 10 min) to give residue. Then the residue was purified by prep-HPLC (column: Waters Xbridge 150*25 mm* 5um; mobile phase: [water (ammonia hydroxide v / v) -ACN]; gradient: 23%- 53% B over 10 min) to give (2) (4.64 mg, 9.66 pmol, 6.50% yield, 94% purity) as white solid.1H NMR: (400 MHz, CDCb) 6 = 7.53 - 7.47 (m, 1 H), 7.25 (dd, J = 0.9, 1.9 Hz, 1 H), 7.18 - 7.16 (m, 1 H), 7.13 (dd, J = 1.9, 8.1 Hz, 1 H), 6.59 (t, J = 73.2 Hz, 1 H), 6.04 (s, 1 H), 3.48 (s, 2H), 3.21 (s, 3H), 2.80 - 2.66 (m, 2H), 2.52 - 2.41 (m, 2H), 2.39 (s, 3H), 1 .53 (s, 3H); LCMS: RT = 0.505 min, m / z = 452.1 (M+H)+.

[0319] EXAMPLE 1.3: SYNTHESIS OF (3)

[0320] Scheme 3: synthesis of (3)

[0321] Synthesis of compound 2, (scheme 3)

[0322] A mixture of compound 1 (5 g, 26.73 mmol, 1 eq) and ethyl 3-oxobutanoate (4.17 g, 32.08 mmol, 4.06 mL, 1.2 eq) in AcOH (40 mL) was stirred at 100 °C for 1 h. The reaction mixture was diluted with EtOAc (40 mL) and then the resulting mixture was stirred at 20 °C for 2 h. Some solid formed. The mixture was filtered, the filtrate was discarded, the filter cake was collected and dried under reduced pressure to give white solid by oil pump. To give compound 2 (3 g, 1 1 .38 mmol, 42.57% yield, 96% purity) as white solid.1H NMR: (400 MHz, DMSO-d6) 6 = 7.96 - 7.92 (m, 1 H), 7.76 (td, J = 1 .9, 7.3 Hz, 1 H), 7.45 - 7.35 (m, 3H), 2.17 - 2.10 (m, 3H); LCMS: RT = 0.497 min, m / z = 253.1 (M+H)+.

[0323] Synthesis of compound 3, (scheme 3)

[0324] To a solution of compound 2 (2 g, 7.90 mmol, 1 eq in ACN (5 mL) was added CH3I (3.36 g, 23.71 mmol, 1.48 mL, 3 eq). The mixture was stirred at 120 °C for 2 h under microwave. The compound 2 was remained, CH3I (3.36 g, 23.71 mmol, 1.48 mL, 3 eq) was added and the mixture was stirred at 120 °C for 2 h under microwave. The compound 2 was still remained, CH3I (3.36 g, 23.71 mmol, 1 .48 mL, 3 eq was added and the mixture was stirred at 120 °C for

[0325] 2 h under microwave. The 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, PE / EtOAc = 1 / 1 to 1 / 1 , TLC: PE: EtOAc = 1 : 1 , Rf = 0.1), the eluent was concentrated and under reduced pressure to give compound

[0326] 3 (1.76 g, 4.61 mmol, 58.37% yield, 70% purity) as brown oil. LCMS: RT = 0.428 min, m / z = 269.1 (M+H)+.

[0327] Synthesis of compound 4, (scheme 3) To a solution of compound 3 (1 g, 3.74 mmol, 1 eq) in TFA (10 mL) was added HNO3 (2.8 g, 28.88 mmol, 2.00 mL, 65% purity, 7.72 eq) slowly at -20 °C. The mixture was stirred at -20 °C for 1 h. The reaction mixture was poured into ice water slowly, and the pH of mixture was neutralized by adding saturated NaHCOs solution to 6-7. Then the mixture was extracted with EtOAc (100 mL *3). The combined organic layers were washed with brine (100 mL * 2), dried over Na2SO4, filtered under reduced pressure to give compound 4 (400 mg, 858.65 pmol, 22.94% yield, 67% purity) as brown oil. LCMS: RT = 0.393 min, m / z = 312.0 (M+H)+.

[0328] Synthesis of compound 5, (scheme 3)

[0329] To a solution of compound 4 (200 mg, 640.78 pmol, 1 eq) in dioxane (3 mL) and H2O (0.5 mL) was added Pd(dppf)Cl2 (23.44 mg, 32.04 pmol, 0.05 eq), Na2COs (135.83 mg, 1.28 mmol, 2 eq) and cyclopropylboronic acid (550.41 mg, 6.41 mmol, 10 eq), the mixture was stirred at 80 °C for 16 h. The mixture was diluted with H2O (30 mL) and the resulting mixture was extracted with EtOAc (10 mL * 4). 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, PE / EtOAc = 1 / 1 to 0 / 1 ; TLC: PE / EtOAc = 0:1 , Rf = 0.45), the eluent was concentrated under reduced pressure to give compound 5 (40 mg, crude) as brown solid. LCMS: RT = 0.554 min, m / z = 273.1 (M+H)+.

[0330] Synthesis of compound 6, (scheme 3)

[0331] To a solution of compound 5 (40 mg, 146.37 pmol, 1 eq) in EtOH (2 mL) and H2O (2 mL) was added Fe (49.05 mg, 878.20 pmol, 6 eq) and NH4CI (78.29 mg, 1.46 mmol, 10 eq). The mixture was stirred at 80 °C for 2 h. The mixture was filtered, the filtrate was collected under reduced pressure to give residue. The residue was diluted with H2O (20 mL) and 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 (15 mg, 61.65 pmol, 42.12% yield) as brown gum. LCMS: RT = 0.380 min, m / z = 244.1 (M+H)+.

[0332] Synthesis of (3), (scheme 3)

[0333] To a solution of compound 6 (15 mg, 61.65 pmol, 1 eq) in DCM (1 mL) was added Py (14.63 mg, 184.95 pmol, 14.93 pL, 3 eq) and (2-fluorophenyl) methanesulfonyl chloride (12.86 mg, 61.65 pmol, 1 eq) at 0 °C. The mixture was stirred at 20 °C for 16 h. The 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]; gradient: 43%-73% B over 10 min), the eluent was concentrated and then freeze dried to give (3) (0.78 mg, 1 .86 pmol, 3.01 % yield, 99% purity) as yellow solid.1H NMR: (400 MHz, CDCb) 6 = 7.97 - 7.91 (m, 1 H), 7.42 - 7.32 (m, 2H), 7.18 - 7.05 (m, 5H), 6.00 - 5.93 (m, 1 H), 4.63 - 4.59 (m, 2H), 3.23 - 3.20 (m, 3H), 2.39 (s, 3H), 1 .99 - 1 .94 (m, 1 H), 1 .08 - 0.99 (m, 2H), 0.79 - 0.73 (m, 2H);

[0334] LCMS: RT = 0.512 min, m / z = 416.3 (M+H)+.

[0335] EXAMPLE 1.4: SYNTHESIS OF (4)

[0336] Scheme 4: Synthesis of (4)

[0337] To a mixture of compound 1 (60 mg, 148.24 pmol, 1 eq) and pyridine (1 17.26 mg, 1.48 mmol, 119.65 pL, 10 eq) in DCM (1 mL) was added compound 2 (41 .39 mg, 177.89 pmol, 1 .2 eq) at 0 °C, then the mixture was stirred at 20 °C for 16 h. The reaction mixture was concentrated under reduced pressure 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). (4) (24 mg, 48.05 pmol, 32.41 % yield, 96% purity) was obtained as yellow solid.1H NMR: (400 MHz, CDCb) 6 = 7.79 (s, 1 H), 7.72 (s, 1 H), 7.52 - 7.45 (m, 3H), 7.21 - 7.15 (m, 1 H), 6.72 - 6.06 (m, 1 H), 3.96 (s, 3H), 3.21 (s, 3H), 3.16 (d, J = 6.0 Hz, 2H), 2.40 (s, 3H), 2.12 - 2.02 (m, 4H), 1 .80 - 1 .68 (m, 3H), 1 .48 - 1 .35 (m, 2H); LCMS: RT = 0.520 min, m / z = 480.2 (M+H)+.

[0338] EXAMPLE 1.5: SYNTHESIS OF (5)

[0339]

[0340] Scheme 5: Synthesis of (5)

[0341] Synthesis of compound 2, (scheme 5)

[0342] To a solution of compound 1 (3 g, 16.39 mmol, 1 eq) in DCM (20 mL) was added ethyl 2, 2-dibromo-2-fluoro-acetate (8.65 g, 32.78 mmol, 2 eq), EtONa (2.57 g, 37.70 mmol, 2.3 eq). The mixture was stirred at 20 °C for 14 h. PE (60 mL) and water (40 mL) were added and Layers were separated. The aqueous phase was extracted with PE (50 mL x 2). Combined extracts were washed with brine (40 mL), dried over Na2SO4, filtered, and concentrated under vacuum to give a residue. The residue was purified by flash silica gel chromatography (ISCO®; 80 g SepaFlash® Silica Flash Column, Eluent of 0% Ethyl acetate / Petroleum ether gradient @ 70 mL / min). Compound 2 (3 g, 10.21 mmol, 62.27% yield) was obtained as a colorless liquid.1H NMR: (400 MHz, CDCb) 6 = 7.60-7.44 (m, 2H), 7.32 - 7.21 (m, 2H), 2.91 - 2.77 (m, 1 H), 2.14 (ddd, J = 8.0, 11.6, 16.9 Hz, 1 H), 2.02 - 1.87 (m, 1 H), 1.75 (q, J = 8.0 Hz, 1 H).

[0343] Synthesis of compound 3, (scheme 5)

[0344] A solution of compound 2 (2.4 g, 8.16 mmol, 1 eq) and AIBN (268.13 mg, 1.63 mmol, 0.2 eq) in toluene (20 mL) was degassed with N2. To this mixture was added a solution of tributylstannane (8.32 g, 28.58 mmol, 7.57 mL, 3.5 eq) in toluene (1 mL). The reaction mixture was stirred at 80 °C for 14 h. EtOAc (40 mL) and KF (60 mL, 1 M) 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®; 40 g SepaFlash® Silica Flash Column, Eluent of 0% Ethyl acetate / Petroleum ether gradient @ 50 mL / min). Compound 3 (310 mg, 1 .44 mmol, 17.66% yield) was obtained as a colorless liquid.1H NMR: (400 MHz, CDCb) 6 = 7.48 - 7.34 (m, 2H), 7.23 - 7.15 (m, 2H), 4.93 - 4.66 (m, 1 H), 2.14 - 1 .97 (m, 1 H), 1 .39 - 1 .18 (m, 2H).

[0345] Synthesis of compound 4, (scheme 5)

[0346] To a solution of compound 3 (190 mg, 883.47 pmol, 1 eq) in dioxane (4 mL) was added tert-butyl A / -aminocarbamate (233.52 mg, 1.77 mmol, 2 eq), ditert-butyl-[2-(2, 4, 6- triisopropylphenyl) phenyl]phosphane (75.03 mg, 176.69 pmol, 0.2 eq), Pd2 (dba)3 (80.90 mg, 88.35 pmol, 0.1 eq) and CS2CO3 (575.70 mg, 1 .77 mmol, 2 eq). The mixture was stirred at 100 °C for 14 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 flash silica gel chromatography (ISCO®; 12 g SepaFlash® Silica Flash Column, Eluent of 0-20% Ethyl acetate / Petroleum ether gradient @ 35 mL / min). Compound 4 (200 mg, 533.21 pmol, 60.35% yield, 71 % purity) (LCMS: EW33193-717-P1 B) was obtained as a yellow oil. LCMS: RT = 0.522 min, m / z = 211 .1 (M+H-tBu)+.

[0347] Synthesis of compound 5, (scheme 5)

[0348] To a solution of compound 4 (200 mg, 751.00 pmol, 1 eq) in EtOAc (8 mL) was added HCI / EtOAc (4 M, 4 mL). The mixture was stirred at 20 °C for 4 h. The mixture was concentrated under vacuum to give a residue. Compound 5 (120 mg, 592.14 pmol, 78.85% yield, HCI) was obtained as a yellow solid.1H NMR: (400 MHz, DMSO-d6) 6 = 10.20 (br s, 3H), 7.20 (t, J = 7.8 Hz, 1 H), 6.95 - 6.78 (m, 3H), 5.05 - 4.76 (m, 1 H), 2.16 - 2.03 (m, 1 H), 1 .37 - 1 .14 (m, 2H).

[0349] Synthesis of compound 7, (scheme 5)

[0350] A solution of compound 5 (50 mg, 246.72 pmol, 1 eq, HCI) and ethyl compound 6 (64.22 mg, 493.45 pmol, 62.47 pL, 2 eq) in AcOH (0.5 mL) was stirred under an N2 atmosphere at 100 °C for 1 h. Most of AcOH was removed after concentration to give a residue. The residue was poured into NaHCOs (15 mL, 2 M). The aqueous Layer was extracted with EtOAc (20 mL). The organic layer was washed with brine (15 mL), dried over Na2SO4 and filtered. The filtrate was concentrated. The residue was purified by prep-TLC (SiO2, PE: EtOAc = 1 : 1). Compound 7 (28 mg, 1 13.32 pmol, 45.93% yield, 94% purity) was obtained as a yellow oil. LCMS: RT = 0.443 min, m / z = 233.1 (M+H)+.

[0351] Synthesis of compound 8, (scheme 5)

[0352] Compound 7 (53 mg, 228.20 pmol, 1 eq) and Mel (291 .51 mg, 2.05 mmol, 127.86 pL, 9 eq) were taken up into a microwave tube in ACN (1 mL). The sealed tube was heated at 120 °C for 3 h under microwave. Most of ACN was removed after concentration under reduced pressure to give a residue. The residue was poured into H2O (15 mL). The aqueous Layer was extracted with EtOAc (15 mL). The organic Layer was washed with brine (10 mL), dried over Na2SO4 and filtered. The filtrate was concentrated under reduced pressure. Compound 8 (56 mg, 163.72 pmol, 71.74% yield, 72% purity) was obtained as a yellow oil. LCMS: RT = 0.425 min, m / z =

[0353] 247.1 (M+H)+.

[0354] Synthesis of compound 9, (scheme 5)

[0355] To a solution of compound 8 (56 mg, 227.38 pmol, 1 eq) in TFA (1 mL) and DCM (1 mL) was added HNO3 (63.21 mg, 682.15 pmol, 45.15 pL, 68% purity, 3 eq) at 0 °C. The mixture was stirred at 20 °C for 1 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. Compound 9 (70 mg, 218.69 pmol, 96.18% yield, 91 % purity) was obtained as a yellow solid. LCMS: RT = 0.410 min, m / z = 292.1 (M+H)+.

[0356] Synthesis of compound 10, (scheme 5)

[0357] To a solution of compound 9 (70 mg, 240.32 pmol, 1 eq) in EtOH (4 mL) and H2O (1 mL) was added Fe (67.10 mg, 1.20 mmol, 5 eq) and NH4CI (102.84 mg, 1.92 mmol, 8 eq). The mixture was stirred at 85 °C for 2 h. The mixture was filtered and the filtrate was concentrated undervacuum to give a residue. Compound 10 (53 mg, 178.50 pmol, 74.27% yield, 88% purity) (LCMS: EW33193-730-P1 B) was obtained as a yellow solid. LCMS: RT = 0.360 min, m / z =

[0358] 262.2 (M+H)+.

[0359] Synthesis of compound (5), (scheme 5)

[0360] To a solution of compound 10 (53 mg, 202.84 pmol, 1 eq) in DCM (1 mL) was added Py (320.89 mg, 4.06 mmol, 327.44 pL, 20 eq) and (2-fluorophenyl) methanesulfonyl chloride (50.78 mg, 243.40 pmol, 1 .2 eq). The mixture was stirred at 20 °C for 14 h. EtOAc (10 mL) and water (10 mL) were added and Layers were separated. The aqueous phase was extracted with EtOAc (10 mL * 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: YMC-Actus Triart C18 150*30 mm*7um; mobile phase: [water (FA) -ACN]; gradient: 38%-68% B over 10 min). Compound (5) (19.02 mg, 43.00 pmol, 21.20% yield, 98% purity) was obtained as a brown solid.1H NMR: (400 MHz, CDCb) 6 = 7.96 - 7.88 (m, 1 H), 7.49 - 7.40 (m, 1 H), 7.37 - 7.28 (m, 2H), 121 (d, J = 1 .4 Hz, 2H), 7.17 - 7.05 (m, 2H), 6.22 (br s, 1 H), 4.95 - 4.71 (m, 1 H), 4.59 (s, 2H), 3.20 (s, 3H), 2.38 (s, 3H), 2.20 - 2.09 (m, 1 H), 1 .38 - 1 .24 (m, 2H); LCMS: RT = 0.500 min, m / z = 434.1 (M+H)+.

[0361] Synthesis of Compound (5)-peak1 and (5)-peak2, (scheme 5)

[0362] (5) was purified by SFC (RT = 2.224 min and 2.722 min; column: DAICEL CHIRALPAK AD (250mm*30mm,10um);mobile phase: [CO2-MeOH (0.1 % NH3 H2O)];B%:40%, isocratic elution mode) to give (5)-peak1 (8.21 mg, 18.37 pmol, 41.87% yield, 97% purity) as yellow solid.1H NMR: (400 MHz, CDCb) 6 = 7.91 (br t, J = 7.3 Hz, 1 H), 7.49 - 7.42 (m, 1 H), 7.35 - 121 (m, 4H), 7.17 - 7.05 (m, 2H), 4.93 - 4.74 (m, 1 H), 4.59 (s, 2H), 3.21 (s, 3H), 2.38 (s, 3H), 2.23 - 2.08 (m, 1 H), 1 .31 - 1 .21 (m, 2H); LCMS: RT = 0.507 min, m / z = 434.2 (M+H)+; SFC: RT = 2.232 min.

[0363] (5)-peak2 (7.95 mg, 15.77 pmol, 35.95% yield, 86% purity) as yellow solid.1H NMR: (400 MHz, CDCb) 6 = 7.92 (br t, J = 7.1 Hz, 1 H), 7.51 - 7.40 (m, 1 H), 7.37 - 7.26 (m, 4H), 7.19 - 7.03 (m, 2H), 4.95 - 4.70 (m, 1 H), 4.59 (s, 2H), 3.20 (s, 3H), 2.38 (s, 3H), 2.17-2.12 (m, 1 H), 1.34 - 1 .25 (m, 2H); LCMS: RT = 0.488 min, m / z = 434.2 (M+H)+; SFC: RT = 2.693 min.

[0364] EXAMPLE 1.6: SYNTHESIS OF (6) AND (28)

[0365] Scheme 6: Synthesis of (6) and (28)

[0366] Synthesis of compound 2, (scheme 6)

[0367] A mixture of compound 1 (1 g, 4.14 mmol, 1 eq, HCI), ethyl 3-oxobutanoate (538.91 mg, 4.14 mmol, 524.24 pL, 1 eq) in AcOH (5 mL) was degassed and purged with N2 for 3 times, and then the mixture was stirred at 100 °C for 1 h under N2 atmosphere. The mixture was concentrated under reduced pressure to give compound 2 (1 g, crude) as white solid. LCMS: RT = 0.495 min, m / z = 271 .0 (M+H)+.

[0368] Synthesis of compound 3, (scheme 6)

[0369] A mixture of compound 2 (1 g, 3.69 mmol, 1 eq), Mel (1.57 g, 11.07 mmol, 688.94 pL, 3 eq) in ACN (5 mL) was degassed and purged with N2 for 3 times, and then the mixture was stirred at 120 °C for 3 h under microwave reactor. The mixture was concentrated under reduced pressure to give a residue. The residue was purified by reversed-phase HPLC (0.1 % NH3 H2O condition), the eluent was concentrated and then freeze dried to give compound 3 (1 g, crude) as white solid. LCMS: RT = 0.436 min, m / z = 287.0 (M+H)+. Synthesis of compound 4, (scheme 6)

[0370] To a mixture of compound 3 (2 g, 7.01 mmol, 1 eq) in TFA (5 mL) was added HNO3 (1 .5 g, 23.80 mmol, 1 .07 mL, 3.39 eq) at 0 °C. Then the mixture was stirred at 0 °C for 2 h under N2 atmosphere. The reaction mixture was poured into ice cold water (50 mL), and the pH of mixture was neutralized by adding sodium bicarbonate to about 7. Then the resulting mixture was extracted with EtOAc (50 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 reversed-phase HPLC (0.1 % NH3 H2O condition), the eluent was concentrated and then freeze dried to give a compound 4 (2.2 g, 6.46 mmol, 92.16% yield, 97% purity) as white solid. LCMS: RT = 0.416 min, m / z = 332.0 (M+H)+.

[0371] Synthesis of compound 6, (scheme 6)

[0372] A mixture of compound 4 (1 g, 3.03 mmol, 1 eq), compound 5 (780.62 mg, 9.09 mmol, 3 eq), Pd(dppf)Cl2 (221.66 mg, 302.93 pmol, 0.1 eq), CsF (920.32 mg, 6.06 mmol, 223.65 pL, 2 eq) in dioxane (10 mL) was degassed and purged with N2 for 3 times, and then the mixture was stirred at 80 °C for 16 h under N2 atmosphere. The reaction mixture was cooled to room temperature. EtOAc (20 mL) and water (20 mL) were added and layers were separated. The aqueous phase was extracted with EtOAc (10 mL x 2). Combined extracts were washed with brine (20 mL), dried over Na2SO4, filtered and concentrated under vacuum to give a residue. The residue was purified by reversed-phase HPLC (0.1 % FA condition), the eluent was concentrated and then freeze dried to give compound 6 (300 mg, 1.01 mmol, 33.32% yield, 98% purity) as white solid. LCMS: RT = 0.448 min, m / z = 292.2 (M+H)+.

[0373] Synthesis of compound 7, (scheme 6)

[0374] A mixture of compound 6 (300 mg, 1 .03 mmol, 1 eq), Fe (287.59 mg, 5.15 mmol, 5 eq), NH4CI (550.93 mg, 10.30 mmol, 10 eq) in MeOH (8 mL) and H2O (2 mL) was degassed and purged with N2 for 3 times, and then the mixture was stirred at 80 °C for 2 h under N2 atmosphere. The mixture was concentrated under reduced pressure to give a residue. EtOAc (10 mL) and water (10 mL) were added and layers were separated. The aqueous phase was extracted with EtOAc (6 mL x 2). Combined extracts were washed with brine (6 mL), dried over Na2SO4, filtered, and concentrated under vacuum to give compound 7 (200 mg, crude) as white solid. LCMS: RT = 0.393 min, m / z = 262.2 (M+H)+.

[0375] Synthesis of (6), (scheme 6)

[0376] A mixture of compound 7 (150 mg, 574.07 pmol, 1 eq), compound 8 (119.77 mg, 574.07 pmol, 1 eq), pyridine (454.09 mg, 5.74 mmol, 463.35 pL, 10 eq) in DCM (1 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 reaction mixture was cooled to room temperature. EtOAc (10 mL) and water (10 mL) were added and layers were separated. The aqueous phase was extracted with EtOAc (5 mL x 2). Combined extracts were washed with brine (6 mL), dried over Na2SO4, filtered, and concentrated under 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: 40%- 70% B over 10 min) to give (6) (11.23 mg, 25.91 pmol, 4.51 % yield, 100% purity) as white solid.1H NMR: (400 MHz, CDCb) 6 = 7.93 (br t, J = 7.1 Hz, 1 H), 7.37 - 7.29 (m, 1 H), 7.16 - 7.07 (m, 4H), 6.90 (br d, J = 5.9 Hz, 1 H), 6.05 - 5.93 (m, 1 H), 4.59 (s, 2H), 3.18 (br s, 3H), 2.37 (s, 3H), 2.17 - 2.08 (m, 1 H), 1.08 - 1.01 (m, 2H), 0.79 (q, J = 5.2 Hz, 2H); LCMS: RT = 0.512 min, m / z = 434.2 (M+H)+.

[0377] Synthesis of (28), (scheme 7)

[0378] Scheme 7: Synthesis of (28)

[0379] To the mixture of (6) (30 mg, 69.21 pmol, 1 eq) in DMF (1 mL) was added NaH (3.32 mg, 83.05 pmol, 60% purity, 1 .2 eq) at 0 °C, the mixture was stirred at 20 °C for 0.5 h. To the mixture was added Mel (9.82 mg, 69.21 pmol, 4.31 pL, 1 eq) at 0 °C. The mixture was stirred at 20 °C for 1 .5 h. To the mixture was added saturated NH4CI solution (10 mL) and the resulting mixture was extracted with EtOAc (10 mL x 3), the combined organic phase was 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: 43%- 73% B over 10 min) to give (28) (17.63 mg, 37.03 pmol, 53.51 % yield, 94% purity) as brown solid.1H NMR: (400 MHz, CDCb) 6 = 7.76 (br t, J = 7.4 Hz, 1 H), 7.37 - 7.29 (m, 1 H), 7.18 - 7.04 (m, 4H), 6.92 (br d, J = 5.4 Hz, 1 H), 4.67 - 4.46 (m, 2H), 3.25 (s, 3H), 3.17 (s, 3H), 2.33 (s, 3H), 2.18 - 2.08 (m, 1 H), 1.11 - 0.99 (m, 2H), 0.80 (br d, J = 3.9 Hz, 2H); LCMS: RT = 0.537 min, m / z = 448.2 (M+H)+.

[0380] EXAMPLE 1.7: SYNTHESIS OF (7)

[0381] Scheme 8: Synthesis of (7)

[0382] A mixture of compound 1 (44.52 mg, 191.36 pmol, 1 eq), compound 2 (50 mg, 191.36 pmol, 1 eq), pyridine (151 .36 mg, 1 .91 mmol, 154.45 pL, 10 eq) in DCM (1 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 reaction mixture was cooled to room temperature. EtOAc (10 mL) and water (10 mL) were added and layers were separated. The aqueous phase was extracted with EtOAc (5 mL x 2). Combined extracts were washed with brine (6 mL), dried over Na2SO4, filtered, and concentrated under 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: 45%- 75% B over 10 min) to give (7) (34.66 mg, 72.73 pmol, 38.01 % yield, 96% purity) as white solid.1H NMR: (400 MHz, CDCI3) 6 = 7.15 - 7.07 (m, 2H), 6.86 (dd, J = 1 .8, 6.6 Hz, 1 H), 6.10 (br s, 1 H), 3.16 (d, J = 6.5 Hz, 2H), 3.15 (s, 3H), 2.37 (s, 3H), 2.16 - 2.05 (m, 5H), 1 .81 - 1 .64 (m, 3H), 1 .50 - 1 .36 (m, 2H), 1 .08 - 1 .01 (m, 2H), 0.80 - 0.74 (m, 2H); LCMS: RT = 0.541 min, m / z = 458.2 (M+H)+.

[0383] EXAMPLE 1.8: SYNTHESIS OF (8)

[0384] Scheme 9: Synthesis of (8)

[0385] A mixture of compound 1 (57.88 mg, 264.71 pmol, 1 eq), compound 2 (75 mg, 264.71 pmol, 1 eq), pyridine (62.82 mg, 794.13 pmol, 64.10 pL, 3 eq) in DCM (2 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 prep-HPLC (column: YMC-Actus Triart C18 150*30 mm*7um; mobile phase: [water (FA) -ACN]; gradient: 33%- 63% B over 10 min) to give (8) (18.89 mg, 40.58 pmol, 15.33% yield, 100% purity) as white solid.1H NMR: (400 MHz, CDCb) 6 = 7.80 (s, 1 H), 7.73 (s, 1 H), 7.54 - 7.44 (m, 3H), 7.22 - 7.16 (m, 1 H), 6.67 (s, 1 H), 3.96 (s, 3H), 3.48 (s, 2H), 3.22 (s, 3H), 2.72 (q, J = 13.6 Hz, 2H), 2.48 - 2.40 (m, 2H), 2.39 (s, 3H), 1.51 (s, 3H); LCMS: RT = 0.461 min, m / z = 466.1 (M+H)+.

[0386] EXAMPLE 1.9: SYNTHESIS OF (9)

[0387] Scheme 10: Synthesis of (9)

[0388] Synthesis of compound 2, (scheme 10)

[0389] To the mixture of compound 1 (1 g, 4.83 mmol, 1 eq) in DMF (10 mL) was added acetylsulfanylpotassium (0.89 g, 7.79 mmol, 1 .61 eq) at 0 °C. The mixture was stirred at 25 °C for 16 h. To the mixture was added water (50 mL) and the resulting mixture was extracted with EtOAc (50 mL x 3), the combined organic phase was dried over Na2SO4, filtered and the filtrate was concentrated under reduced pressure to give compound 2 (2 g, crude) as brown oil.

[0390] Synthesis of compound 3, (scheme 10)

[0391] To a solution of NCS (2.64 g, 19.78 mmol, 4 eq and HCI (2 M, 4.95 mL, 2 eq in ACN (10 mL) was added compound 2 (1 g, 4.95 mmol, 1 eq) at 0 °C under N2. The mixture was stirred at 0 °C for 1 h. The reaction mixture was concentrated in vacuum to give residue. The residue was purified by silica gel chromatography (100-200 mesh silica gel, Petroleum ether / Ethyl acetate = 1 / 0, 10 / 1) to give compound 3 (470 mg, 2.07 mmol, 41.94% yield) as colorless oil.1H NMR: (400MHz, CDCI3) 6 =7.59 - 7.49 (m, 1 H), 7.07 - 6.90 (m, 2H), 4.91 (s, 2H).

[0392] Synthesis of (9), (scheme 10)

[0393] To a solution of compound 3 (73.65 mg, 324.99 pmol, 1 eq) in DCM (0.5 mL) was added pyridine (77.12 mg, 974.98 pmol, 78.69 pL, 3 eq and compound 4 (100 mg, 324.99 pmol, 1 eq, HCI). The mixture was stirred at 25 °C for 1 h. The mixture was concentrated at reduced pressure to give a residue. The reaction mixture was purified by prep-HPLC (column: Welch Xtimate C18 150*25 mm*5um; mobile phase: [water (FA) -ACN]; gradient: 38%-68% B over 15 min) to give (9) (21.76 mg, 47.16 pmol, 14.51 % yield, 100% purity) as an off-white solid.1H NMR: (400MHz, CDCb) 6 = 8.00 - 7.92 (m, 1 H), 7.69 - 7.58 (m, 4H), 6.94 - 6.81 (m, 2H), 5.85 (s, 1 H), 4.54 (s, 2H), 3.22 (s, 3H), 2.41 (s, 3H); LCMS: RT = 0.523 min, m / z = 462.1 (M+H)+

[0394] EXAMPLE 1.10: SYNTHESIS OF (10)

[0395] Scheme 11 : Synthesis of (10)

[0396] Synthesis of compound 3, (Scheme 11)

[0397] A mixture of compound 1 (500 mg, 4.09 mmol, 1 eq), 4-methylbenzenesulfonyl chloride (936.74 mg, 4.91 mmol, 1.2 eq), DMAP (50.02 mg, 409.46 pmol, 0.1 eq), DIEA (1 .59 g, 12.28 mmol, 2.14 mL, 3 eq) in DCM (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. EtOAc (20 mL) and water (20 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 column chromatography (silica gel, Petroleum ether / Ethyl acetate = 1 : 0 to 3: 1 , TLC: Petroleum ether: Ethyl acetate = 5: 1 , Rf = 0.46) to give compound 3 (600 mg, crude) as white solid.1H NMR: (400 MHz, CDCb) 6 = 7.80 (d, J = 8.3 Hz, 2H), 7.37 (d, J = 8.0 Hz, 2H), 4.07 (d, J = 6.6 Hz, 2H), 2.70 - 2.58 (m, 2H), 2.47 (s, 3H), 2.36 - 2.23 (m, 2H);

[0398] Synthesis of compound 4, (Scheme 11)

[0399] A mixture of compound 3 (600 mg, 2.17 mmol, 1 eq), potassium; ethanethioate (496.02 mg, 4.34 mmol, 2 eq) in DMF (5 mL) was degassed and purged with N2 for 3 times, and then the mixture was stirred at 60 °C for 16 h under N2 atmosphere. The reaction mixture was cooled to room temperature. EtOAc (20 mL) and water (15 mL) were added and layers were separated. The aqueous phase was extracted with EtOAc (5 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 column chromatography (silica gel, Petroleum ether / Ethyl acetate = 1 : 0 to 3: 1 , TLC: Petroleum ether: Ethyl acetate = 5: 1 , Rf = 0.43) to give compound 4 (400 mg, crude) as white solid.1H NMR: (400 MHz, CDCb) 6 = 3.04 (d, J = 7.4 Hz, 2H), 2.74 - 2.61 (m, 2H), 2.43 - 2.36 (m, 1 H), 2.35 (s, 3H), 2.31 - 2.17 (m, 2H).

[0400] Synthesis of compound 5, (Scheme 11)

[0401] To a mixture of NCS (592.77 mg, 4.44 mmol, 4 eq), HCI (2 M, 1 .11 mL, 2 eq) in ACN (3 mL) was added compound 4 (200 mg, 1.11 mmol, 1 eq) at 0 °C. Then the mixture stirred at 0 °C for 2 h. 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 : 0 to 3: 1 , TLC: Petroleum ether: Ethyl acetate = 5: 1 , Rf = 0.47) to give compound 5 (200 mg, crude) as white solid.1H NMR: (400 MHz, CDCb) 6 = 3.87 (d, J = 6.9 Hz, 2H), 3.04 - 2.87 (m, 3H), 2.63 - 2.48 (m, 2H).

[0402] Synthesis of (10), (Scheme 11)

[0403] A mixture of compound 5 (50 mg, 244.35 pmol, 1 eq), compound 6 (65.40 mg, 268.79 pmol, 1.1 eq), Py (57.98 mg, 733.06 pmol, 59.17 pL, 3 eq) in DCM (2 mL) was degassed and purged with N2 for 3 times, and then the mixture was stirred at 25 °C for 16 h under N2 atmosphere. The 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: 43%- 73% B over 10 min) to give (10) (3.61 mg, 8.77 pmol, 3.59% yield, 100% purity) as white solid.1H NMR: (400 MHz, CDCb) 6 = 7.37 (t, J = 7.8 Hz, 1 H), 7.11 (br d, J = 8.3 Hz, 1 H), 7.08 - 7.03 (m, 2H), 5.98 (br s, 1 H), 3.40 (br d, J = 6.1 Hz, 2H), 3.19 (s, 3H), 2.88 - 2.78 (m, 3H), 2.55 - 2.42 (m, 2H), 2.37 (s, 3H), 2.00 - 1 .91 (m, 1 H), 1 .07 - 0.99 (m, 2H), 0.78 - 0.72 (m, 2H); LCMS: RT = 0.497 min, m / z = 412.2 (M+H)+.

[0404] EXAMPLE 1.11: SYNTHESIS OF (11)

[0405] Scheme 12: Synthesis of (11)

[0406] To the mixture of compound 1 (17 mg, 37.16 pmol, 1 eq) in DMF (0.5 mL) was added NaH (1.93 mg, 48.31 pmol, 60% purity, 1.3 eq) at 0 °C under N2. The mixture was stirred at 25 °C for 0.5 h. Then to the mixture was added Mel (6.86 mg, 48.31 pmol, 3.01 pL, 1.3 eq) at 0 °C. The mixture was stirred at 25 °C for 1 .5 h under N2. To the mixture was added saturated NH4CI solution (5 mL) and the resulting mixture was extracted with EtOAc (5 mL * 3), the combined organic phase was 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: 48%- 78% B over 10 min) to give compound (11) (5.02 mg, 10.54 pmol, 28.36% yield, 99% purity) as yellow solid.1H NMR: (400 MHz, CDCb) 6 = 7.71 (br t, J = 7.3 Hz, 1 H), 7.67 - 7.58 (m, 4H), 7.29 (br d, J = 7.1 Hz, 1 H), 7.19 - 7.13 (m, 1 H), 7.06 (br t, J = 9.2 Hz, 1 H), 4.98 (q, J = 7.0 Hz, 1 H), 3.24 (br s, 3H), 3.18 (s, 3H), 2.35 (s, 3H), 1 .90 (br d, J = 7.0 Hz, 3H); LCMS: RT = 0.557 min, m / z = 472.1 (M+H)+.

[0407] EXAMPLE 1.12: SYNTHESIS OF (12)

[0408] Scheme 13: Synthesis of (12)

[0409] To a solution of compound 2 (22.23 mg, 95.53 pmol, 1.29 eq) in DCM (1 mL) was added compound 1 (20 mg, 74.28 pmol, 1 eq) and pyridine (22.23 mg, 281.00 pmol, 22.68 pL, 3.78 eq) at 0 °C, then the resulting mixture was stirred at 20 °C for 12 h. The reaction mixture was quenched by addition H2O (30 mL), and then the resulting mixture was extracted with EtOAc (15 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: Phenomenex luna C18 150*25 mm* 10um; mobile phase: [water (FA) -ACN]; gradient: 38%- 68% B over 10 min) to give compound (12) (14.35 mg, 29.78 pmol, 40.09% yield, 96.6% purity) as a white solid.1H NMR: (400 MHz, CDCb) 6 = 7.55 - 7.48 (m, 1 H), 7.26 - 7.23 (m, 1 H), 7.16 (br s, 2H), 6.60 (t, J = 32.8 Hz, 1 H), 6.02 (br s, 1 H), 3.22 (s, 3H), 3.20 - 3.15 (m, 2H), 2.40 (s, 3H), 2.14 - 2.02 (m, 5H), 1 .83 - 1 .70 (m, 2H), 1.51 - 1 .38 (m, 2H); LCMS: RT = 0.562 min, m / z = 464.2 (M-H)+.

[0410] EXAMPLE 1.13: SYNTHESIS OF (13)

[0411] Scheme 14: Synthesis of (13)

[0412] A mixture of compound 1 (67.07 mg, 334.27 pmol, 1.5 eq), compound 2 (60 mg, 222.84 pmol, 1 eq), pyridine (52.88 mg, 668.53 pmol, 53.96 pL, 3 eq) in DCM (2 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 prep-MPLC (column: Welch Ultimate XB-CN 250*50*1 Oum; mobile phase: [Hexane-EtOH]; gradient: 10%- 50% B over 15 min) 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) to give (13) (17.02 mg, 37.30 pmol, 16.74% yield, 95% purity) as white solid.1H NMR: (400 MHz, CDCb) 6 = 7.49 (t, J = 8.1 Hz, 1 H), 7.27 - 7.23 (m, 1 H), 7.18 - 7.10 (m, 2H), 6.80 - 6.39 (m, 1 H), 6.06 (s, 1 H), 3.37 - 3.30 (m, 2H), 3.19 (s, 3H), 3.11 (td, J = 8.1 , 16.2 Hz, 1 H), 2.65 - 2.41 (m, 2H), 2.38 (s, 3H), 2.28 - 2.00 (m, 2H), 1 .49 - 1 .39 (m, 3H); LCMS: RT = 0.485 min, m / z = 434.1 (M+H)+.

[0413] EXAMPLE 1.14: SYNTHESIS OF (14)

[0414] Scheme 15: Synthesis of (14)

[0415] Synthesis of compound 2, (Scheme 15)

[0416] A mixture of compound 1 (200 mg, 640.78 pmol, 1 eq), AcOK (125.78 mg, 1.28 mmol, 2 eq), Pd(dppf)Cl2 (46.89 mg, 64.08 pmol, 0.1 eq) and BPin2 (195.26 mg, 768.94 pmol, 1.2 eq) in dioxane (2 mL) was stirred at 80 °C for 16 h. To the mixture was added water (2 mL) and EtOAc (2 mL), then the mixture was filtered and the filter cake was concentrated. The filtrate was extracted with EtOAc (2 mL x 3), the organic phase was washed with brine (5 mL), dried over Na2SO4, filtered and the filtrate was concentrated under reduced pressure to give a residue. The filter cake and the residue were combined. Compound 2 (250 mg, crude) was obtained as a brown solid. LCMS: RT = 0.483 min, m / z = 260.2 (M+H)+.

[0417] Synthesis of compound 3, (Scheme 15) A mixture of compound 2 (120 mg, 433.15 pmol, 1 eq), m-CPBA (149.49 mg, 866.30 pmol, 2 eq) in EtOH (0.5 mL) and H2O (0.5 mL) was stirred at 0 °C for 2 h. To the mixture was added water (2 mL) and the resulting mixture was filtered, the filter cake was dried under reduced pressure. Compound 3 (100 mg, crude) was obtained as a white solid. LCMS: RT = 0.322 min, m / z = 250.1 (M+H)+.

[0418] Synthesis of compound 4, (Scheme 15)

[0419] To a solution of compound 3 (100 mg, 401.25 pmol, 1 eq) in DCM (1.5 mL) and H2O (0.5 mL) was added KOH (112.56 mg, 2.01 mmol, 5 eq) and [bromo (difluoro) methyl] -trimethyl- silane (122.24 mg, 601 .87 pmol, 1 .5 eq) at 0 °C. The mixture was stirred at 0 °C for 2 hr. LCMS showed compound 3 remained. To the mixture was added [bromo (difluoro) methyl] -trimethyl- silane (162.99 mg, 802.50 pmol, 2 eq) and the mixture was stirred at 20 °C for another 2 h. To the mixture was added water (5 mL) and the resulting mixture was extracted with EtOAc (5 mL x 3), the organic phase was dried over Na2SO4, filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by reverse flash (0.1 % FA condition), the eluent was concentrated and then freeze dried. Compound 4 (15 mg, 50.13 pmol, 12.49% yield) was obtained as a red solid. LCMS: RT = 0.410 min, m / z = 300.1 (M+H)+.

[0420] Synthesis of compound 5, (Scheme 15)

[0421] A mixture of compound 4 (15 mg, 50.13 pmol, 1 eq), Fe (14 mg, 250.64 pmol, 5 eq) and NH4CI (13.41 mg, 250.64 pmol, 5 eq) in EtOH (0.5 mL) and H2O (0.5 mL) was stirred at 80 °C for 2 h. The mixture was filtered and the filtrate was concentrated under reduced pressure to give a residue. Compound 5 (14 mg, crude) was obtained as a yellow solid. LCMS: RT = 0.363 min, m / z = 270.1 (M+H)+.

[0422] Synthesis of compound (14), (Scheme 15)

[0423] A mixture of compound 5 (14 mg, 52.00 pmol, 1 eq), compound 6 (10.85 mg, 52.00 pmol, 1 eq) and Py (16.45 mg, 207.99 pmol, 16.79 pL, 4 eq) in DCM (1 mL) was stirred at 20 °C for 16 h. To the mixture was added water (5 mL) and the 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-HPLC (column: Waters Xbridge 150*25 mm* 5um; mobile phase: [water (NH4HCO3) -ACN]; gradient: 34%- 64% B over 9 min), the eluent was concentrated and then freeze dried. Compound (14) (7.19 mg, 15.96 pmol, 30.70% yield, 98% purity) was obtained as a white solid.1H NMR: (400 MHz, CDCI3) 6 = 7.90 (t, J = 7.2 Hz, 1 H), 7.51 (t, J = 8.3 Hz, 1 H), 7.38 - 7.29 (m, 2H), 7.21 - 7.08 (m, 4H), 6.59 (t, J = 73.3 Hz, 1 H), 5.90 (br s, 1 H), 4.60 (s, 2H), 3.23 (s, 3H), 2.40 (s, 3H); LCMS: RT = 0.497 min, m / z = 442.1 (M+H)+.

[0424] EXAMPLE 1.15: SYNTHESIS OF (15)

[0425] Scheme 16: Synthesis of (15)

[0426] Synthesis of compound 3, (Scheme 16)

[0427] A mixture of compound 1 (500 mg, 3.33 mmol, 1 eq), 4-methylbenzenesulfonyl chloride (761.75 mg, 4.00 mmol, 1.2 eq), DMAP (40.68 mg, 332.97 pmol, 0.1 eq), DIEA (1.29 g, 9.99 mmol, 1 .74 mL, 3 eq) in DCM (5 mL) was stirred at 20 °C for 16 h. EtOAc (20 mL) and water (20 mL) were added and layers were separated. The aqueous phase was extracted with EtOAc (10 mL x 2). Combined extracts were washed with brine (20 mL), dried over Na2SO4, filtered, and concentrated under vacuum to give a residue. The residue was purified by column chromatography (silica gel, Petroleum ether / Ethyl acetate = 1 : 0 to 3: 1 , TLC: Petroleum ether: Ethyl acetate = 5: 1 , Rf = 0.46) to give compound 3 (600 mg, crude) as white solid.1H NMR: (400 MHz, CDCb) 6 = 7.79 (d, J = 8.3 Hz, 2H), 7.36 (d, J = 8.1 Hz, 2H), 3.90 - 3.85 (m, 2H), 2.47 (s, 3H), 2.13 - 2.03 (m, 2H), 1 .84 - 1 .59 (m, 5H), 1 .34 - 1 .22 (m, 2H)

[0428] Synthesis of compound 4, (Scheme 16)

[0429] A mixture of compound 3 (600 mg, 1.97 mmol, 1 eq), potassium; ethanethioate (450.30 mg, 3.94 mmol, 2 eq) in DMF (5 mL) was degassed and purged with N2 for 3 times, and then the mixture was stirred at 60 °C for 16 h under N2 atmosphere. The reaction mixture was cooled to room temperature. EtOAc (10 mL) and water (15 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 compound 4 (400 mg, crude) as white solid.1H NMR: (400 MHz, CDCb) 6 = 2.84 (d, J = 6.8 Hz, 2H), 2.35 (s, 3H), 2.15 - 2.03 (m, 2H), 1.91 - 1.81 (m, 2H), 1.79 - 1.71 (m, 1 H), 1 .68 - 1.64 (m, 1 H), 1 .60 - 1 .51 (m, 1 H), 1 .40 - 1 .31 (m, 2H).

[0430] Synthesis of compound 5, (Scheme 16)

[0431] To a mixture of NCS (512.93 mg, 3.84 mmol, 4 eq), HCI (2 M, 960.30 pL, 2 eq) in ACN (3 mL) was added compound 4 (200 mg, 960.30 pmol, 1 eq) at 0 °C. Then the mixture was stirred at 0 °C for 2 h. 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 : 0 to 3: 1 , TLC: Petroleum ether: Ethyl acetate = 5: 1 , Rf = 0.47) to give compound 4 (200 mg, crude) as white solid.1H NMR: (400 MHz, CDCb) 6 = 3.68 (d, J = 6.4 Hz, 2H), 2.37 - 2.24 (m, 1 H), 2.21 - 2.07 (m, 4H), 1 .92 - 1 .73 (m, 2H), 1 .59 - 1 .52 (m, 2H)

[0432] Synthesis of (15), (Scheme 16)

[0433] A mixture of compound 5 (50 mg, 214.89 pmol, 1 eq), compound 6 (57.51 mg, 236.38 pmol, 1.1 eq) and Py (17.00 mg, 214.89 pmol, 17.34 pL, 1 eq) in DCM (2 mL) was degassed and purged with N2 for 3 times, and then the mixture was stirred at 25 °C for 16 h under N2 atmosphere. The mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (column: Waters Xbridge 150*25 mm* 5um; mobile phase: [water (ammonia hydroxide v / v) -ACN]; gradient: 33%- 63% B over min) to give (15) (3.25 mg, 7.10 pmol, 3.30% yield, 96% purity) as white solid.1H NMR: (400 MHz, CDCb) 6 = 7.40 - 7.34 (m, 1 H), 7.13 - 7.09 (m, 1 H), 7.08 - 7.04 (m, 2H), 5.88 (br s, 1 H), 3.20 (d, J = 6.3 Hz, 2H), 3.18 (s, 3H), 2.38 (s, 3H), 2.19 - 2.05 (m, 5H), 1.99 - 1.91 (m, 1 H), 1.83 - 1.69 (m, 2H), 1.50 - 1.39 (m, 2H), 1 .06 - 1 .00 (m, 2H), 0.77 - 0.72 (m, 2H); LCMS: RT = 0.526 min, m / z = 440.2 (M+H)+.

[0434] EXAMPLE 1.16: SYNTHESIS OF (16)

[0435] Scheme 17: Synthesis of (16)

[0436] To a solution of compound 3 (38.46 mg, 184.34 pmol, 1 eq) in DCM (0.5 mL) was added pyridine (43.74 mg, 553.02 pmol, 44.64 pL, 3 eq) and compound 1 (100 mg, 368.68 pmol, 2 eq). The mixture was stirred at 25 °C for 1 h. The mixture was concentrated at reduced pressure to give a residue. The reaction mixture was purified by prep-HPLC (column: Welch Xtimate C18 150*25 mm*5um; mobile phase: [water (FA) -ACN]; gradient: 35%-65% B over 15 min) to give (16) (20.18 mg, 45.51 pmol, 24.69% yield, 100% purity) as off-white solid.1H NMR: (400 MHz, CDCb) 6 = 7.68 - 7.61 (m, 4H), 7.47 - 7.42 (m, 2H), 7.36 - 7.30 (m, 1 H), 7.10 - 7.01 (m, 1 H), 5.79 (br s, 1 H), 4.50 (s, 2H), 3.19 (s, 3H), 2.39 (s, 3H); LCMS: RT = 0.518 min, m / z = 444.0 (M+H)+.

[0437] EXAMPLE 1.17: SYNTHESIS OF (17)

[0438] Scheme 18: Synthesis of (17)

[0439] Synthesis of compound 2, (Scheme 18)

[0440] To a mixture of compound 1 (500 mg, 3.78 mmol, 1 eq) in THF (10 mL) was added borane; tetrahydrofuran (1 M, 7.57 mL, 2 eq) at 0 °C under N2 atmosphere. Then the mixture stirred at 20 °C for 3 h. The reaction mixture was quenched with H2O (10 mL) and the resulting mixture was extracted with EtOAc (10 mL*2). The combined organic layer was washed with brine (10 mL), dried over Na2SO4, filtered, and concentrated under vacuum to give compound 2 (370 mg, crude) as colorless oil.1H NMR: (400 MHz, CDCb) 6 = 3.61 (d, J = 6.8 Hz, 2H), 2.63 - 2.53 (m, 1 H), 2.50 - 2.36 (m, 2H), 1 .97 - 1 .87 (m, 2H), 1 .47 - 1 .41 (m, 3H).

[0441] Synthesis of compound 3, (Scheme 18)

[0442] A mixture of compound 2 (370 mg, 3.13 mmol, 1 eq), TosCI (895.6 mg, 4.70 mmol, 1 .5 eq), DMAP (38.3 mg, 313.16 pmol, 0.1 eq), DIEA (1 .62 g, 12.53 mmol, 2.18 mL, 4 eq) in DCM (3 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 reaction mixture was cooled to room temperature. 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 column chromatography (silica gel, Petroleum ether / Ethyl acetate = 1 : 0 to 3: 1 , TLC: Petroleum ether: Ethyl acetate = 5: 1 , Rf = 0.51) to give compound 3 (450 mg, crude) as colorless oil.1H NMR: (400 MHz, CDCb) 6 = 7.79 (d, J = 8.3 Hz, 2H), 7.36 (d, J = 8.1 Hz, 2H), 4.01 (d, J = 6.4 Hz, 2H), 2.78 - 2.64 (m, 1 H), 2.46 (s, 3H), 2.44 - 2.29 (m, 2H), 2.03 - 1 .84 (m, 2H), 1.45 - 1.33 (m, 3H).

[0443] Synthesis of compound 5, (Scheme 18)

[0444] To a mixture of compound 3 (450 mg, 1.65 mmol, 1 eq) in DMF (3 mL) was added compound 4 (1 .31 g, 11 .47 mmol, 6.94 eq). Then the mixture was stirred at 60 °C for 2 h under N2 atmosphere. The reaction mixture was cooled to room temperature. EtOAc (20 mL) and water (20 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 compound 5 (250 mg, crude) as white solid.1H NMR: (400 MHz, CDCb) 6 = 2.97 (d, J = 7.6 Hz, 2H), 2.59 (td, J = 8.1 , 16.0 Hz, 1 H), 2.51 - 2.35 (m, 3H), 2.32 (s, 3H), 2.02 - 1 .94 (m, 1 H), 1 .47 - 1 .40 (m, 3H).

[0445] Synthesis of compound 6, (Scheme 18)

[0446] To a mixture of NCS (757.61 mg, 5.67 mmol, 4 eq), HCI (2 M, 1 .42 mL, 2 eq) in ACN (3 mL) was added compound 5 (250 mg, 1 .42 mmol, 1 eq) at 0 °C under N2 atmosphere. Then the mixture stirred at 0 °C for 2 h. 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 : 0 to 3: 1 , TLC: Petroleum ether: Ethyl acetate = 5: 1 , Rf = 0.47) to give compound 6 (130 mg, crude) as white solid.1H NMR: (400 MHz, CDCb) 6 = 3.80 (d, J = 7.5 Hz, 2H), 3.32 - 3.15 (m, 1 H), 2.79 - 2.63 (m, 2H), 2.18 - 2.02 (m, 2H), 1 .52 - 1 .45 (m, 3H).

[0447] Synthesis of (17), (Scheme 18)

[0448] To a mixture of compound 7 (70.60 mg, 249.18 pmol, 1 eq), pyridine (98.55 mg, 1 .25 mmol, 100.56 pL, 5 eq) in DCM (2 mL) was added compound 6 (50 mg, 249.18 pmol, 1 eq) 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 prep-MPLC (column: Welch Ultimate XB-CN 250*50*1 Oum; mobile phase: [Hexane-EtOH]; gradient: 15%- 55% B over 15 min) to give (17) (57.74 mg, 122.57 pmol, 49.19% yield, 95% purity) as white solid.1H NMR: (400 MHz, CDCb) 6 = 7.82 - 7.73 (m, 2H), 7.56 - 7.45 (m, 3H), 7.17 (dt, J = 2.1 , 4.4 Hz, 1 H), 3.96 (s, 3H), 3.37 - 3.26 (m, 2H), 3.21 (s, 3H), 3.18 - 3.03 (m, 1 H), 2.61 - 2.41 (m, 2H), 2.39 (s, 3H), 2.26 - 1 .96 (m, 2H), 1.41 - 1 .33 (m, 3H); LCMS: RT = 0.449 min, m / z = 448.1 (M+H)+.

[0449] EXAMPLE 1.18: SYNTHESIS OF (18)

[0450] Scheme 19: Synthesis of (18) Synthesis of compound 3, (Scheme 19)

[0451] To the mixture of compound 1 (2 g, 9.41 mmol, 1 .48 mL, 1 eq, HCI) in AcOH (15 mL) was added compound 2 (1 .47 g, 11 .29 mmol, 1 .43 mL, 1 .2 eq). The mixture was stirred at 100 °C for 1 hr. The reaction 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: Phenomenex luna C18 150*40 mm* 15um; mobile phase: [water (FA) -ACN]; B%: 32%- 62%, 11 min). Compound 3 (900 mg, crude) was obtained as a yellow solid.1H NMR: (400 MHz, CDCI3) 6 = 8.20 - 8.14 (m, 2H), 7.51 (t, J = 7.9 Hz, 1 H), 7.43 (d, J = 7.9 Hz, 1 H), 3.48 (s, 2H), 2.23 (s, 3H).

[0452] Synthesis of compound 4, (Scheme 19)

[0453] To the mixture of compound 3 (450 mg, 1 .86 mmol, 1 eq) in ACN (10 mL) was added Mel (791.16 mg, 5.57 mmol, 347.00 pL, 3 eq). The mixture was stirred at 120 °C for 2 h under microwave. Two batches were combined together and the 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: 30%- 60% B over 10 min). Compound 4 (545 mg, crude) was obtained as a yellow oil.1H NMR: (400 MHz, CDCI3) 6 = 7.74 - 7.68 (m, 1 H), 7.68 - 7.56 (m, 3H), 5.53 (s, 1 H), 3.22 (s, 3H), 2.33 (s, 3H).

[0454] Synthesis of compound 5, (Scheme 19)

[0455] To the mixture of compound 4 (270 mg, 1 .05 mmol, 1 eq) in TFA (3 mL) was added HNO3 (220 mg, 2.27 mmol, 157.14 pL, 65% purity, 2.15 eq) at - 10 °C. The mixture was stirred at - 10 °C for 1 h. The mixture was added into ice-cold water (20 mL) slowly 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. Compound 5 (320 mg, crude) was obtained as yellow oil.

[0456] Synthesis of compound 6, (Scheme 19)

[0457] To the mixture of compound 5 (317 mg, 1 .05 mmol, 1 eq) in EtOH (3 mL) and H2O (3 mL) was added Fe (235.08 mg, 4.21 mmol, 4 eq) and NH4CI (281.47 mg, 5.26 mmol, 5 eq). The mixture was stirred at 80 °C for 2 h. The reaction mixture was filtered, the filtrate was concentrated at reduced pressure to give a residue. The residue 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. Compound 6 (258 mg, crude) was obtained as a brown oil. Synthesis of compound (18), (Scheme 19)

[0458] To the mixture of compound 6 (97.50 mg, 359.48 pmol, 1 .5 eq) in DCM (2 mL) was added Py (56.87 mg, 718.95 pmol, 58.03 pL, 3 eq) and compound 7 (50 mg, 239.65 pmol, 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: 45%- 75% B over 10 min). Compound (18) (32.11 mg, 70.97 pmol, 29.61 % yield, 98% purity) was obtained as a gray solid.1H NMR: (400 MHz, CDCh) 6 = 7.84 (dt, J = 1 .4, 7.5 Hz, 1 H), 7.70 - 7.60 (m, 4H), 7.37 - 7.29 (m, 1 H), 7.17 - 7.06 (m, 2H), 6.27 (s, 1 H), 4.57 (s, 2H), 3.22 (s, 3H), 2.40 (s, 3H); LCMS: RT = 0.520min, m / z = 444.1 (M+H)+.

[0459] EXAMPLE 1.19: SYNTHESIS OF (19)

[0460] Scheme 20: Synthesis of (19)

[0461] To a solution of compound 1 (30 mg, 104.44 pmol, 1 eq) in DCM (0.5 mL) was added pyridine (24.78 mg, 313.33 pmol, 25.29 pL, 3 eq) and compound 2 (21 .79 mg, 104.44 pmol, 1 eq). The mixture was stirred at 20 °C for 2 h. EtOAc (5 mL) and water (8 mL) were added and Layers were separated. The aqueous phase was extracted with EtOAc (5 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-HPLC (column: YMC-Actus Triart C18 150*30 mm*7um; mobile phase: [water (FA) -ACN]; gradient: 38%- 68% B over 10 min). The eluent was concentrated and then freeze dried. Compound (19) (13.85 mg, 29.85 pmol, 28.58% yield, 99% purity) was obtained as an off-white solid.1H NMR: (400 MHz, CDCh) 6 = 7.87 - 7.77 (m, 1 H), 7.36 - 7.28 (m, 4H), 7.15 - 7.03 (m, 2H), 6.87 - 6.45 (m, 2H), 4.53 (s, 2H), 3.22 (s, 3H), 2.39 (s, 3H); LCMS: RT =0.491 min, m / z =460.0 (M+H)+.

[0462] EXAMPLE 1.20: SYNTHESIS OF (20)

[0463] Scheme 21 : Synthesis of (20)

[0464] Synthesis of compound 3, (Scheme 21)

[0465] To a solution of compound 1 (2 g, 9.41 mmol, 1.48 mL, 1 eq, HCI) in AcOH (20 mL) was added compound 2 (1 .20 g, 10.35 mmol, 1 .12 mL, 1.1 eq). The mixture was stirred at 100 °C for 1 h. The mixture was concentrated at reduced pressure to give a residue. The residue was purified by silica gel chromatography (100-200 mesh silica gel, Petroleum ether / Ethyl acetate = 3 / 1 , 1 / 1 . TLC (Petroleum ether: Ethyl acetate = 1 : 1 ; Rf = 0.43)) to give compound 3 (2 g, crude) as yellow solid. LCMS: RT = 0.499 min, m / z = 243.1 (M+H)+.

[0466] Synthesis of compound 4, (Scheme 21)

[0467] To a solution of compound 3 (1 g, 4.13 mmol, 1 eq) in ACN (5 mL) was added Mel (2.93 g, 20.64 mmol, 1.29 mL, 5 eq). The sealed tube was heated at 120 °C for 3 h under microwave. LCMS showed compound 3 was remained, then Mel (1.76 g, 12.39 mmol, 771 .1 1 pL, 3 eq) was added into the mixture and heated at 120 °C for another 3 h under microwave. The mixture was concentrated under reduced pressure to give compound 4 (3.6 g, crude) as brown oil.

[0468] Synthesis of compound 5, (Scheme 21)

[0469] To a solution of compound 4 (3.6 g, 14.05 mmol, 1 eq) in TFA (20 mL) was added HNO3 (4.49 g, 46.32 mmol, 3.21 mL, 65% purity, 3.30 eq) slowly under 0 °C. The mixture was stirred at 0 °C for 1 h. To the reaction mixture was added water (100 mL), then the pH of the mixture was adjusted to 6-7 with NaHCOs. The resulting mixture was filtered. The filtrate was discarded and the filter cake was collected and dried by oil pump to give compound 5 (2.2 g, crude) as brown solid. LCMS: RT = 0.430 min, m / z = 302.0(M+H)+.

[0470] Synthesis of compound 6, (Scheme 21) To a solution of compound 5 (2.2 g, 7.30 mmol, 1 eq) in EtOH (20 mL) and H2O (5 mL) was added NH4CI (3.91 g, 73.04 mmol, 10 eq) and Fe (2.04 g, 36.52 mmol, 5 eq). The mixture was stirred at 80 °C for 2 h. The mixture was filtered and the filtrate was concentrated at reduced pressure to give a residue. The residue was purified by silica gel chromatography (100-200 mesh silica gel, Petroleum ether / Ethyl acetate = 3 / 1 , 0 / 1. TLC (Petroleum ether: Ethyl acetate = 0: 1 ; Rf = 0.7)). The fractions were collected and HCI (12 M, 0.3 mL) was added, the resulting fraction was concentrated at reduced pressure to give compound 6 (1.7 g, crude, HCI) as yellow oil. LCMS: RT = 0.387 min, m / z = 272.0 (M+H)+.

[0471] Synthesis of compound 7, (Scheme 21)

[0472] To a solution of NCS (2.96 g, 22.20 mmol, 4 eq) and HCI (2 M, 5.55 mL, 2 eq) in ACN (10 mL) was added compound 8 (1 g, 5.55 mmol, 1 eq) at 0 °C under N2. The mixture was stirred at 0 °C for 1 h. The reaction mixture was concentrated in vacuum to give residue. The residue was purified by silica gel chromatography (100-200 mesh silica gel, Petroleum ether / Ethyl acetate = 1 / 0, 10 / 1) to give compound 7 (1 g, 4.89 mmol, 88.07% yield) as colorless oil.1H NMR: (400MHz, CDCI3) 6 = 3.87 (d, J = 6.8 Hz, 2H), 3.06 - 2.85 (m, 3H), 2.63 - 2.46 (m, 2H).

[0473] Synthesis of (20), (Scheme 21)

[0474] To a solution of compound 6 (50 mg, 184.34 pmol, 1 eq) in DCM (0.5 mL) was added pyridine (43.74 mg, 553.02 pmol, 44.64 pL, 3 eq) and compound 7 (37.72 mg, 184.34 pmol, 1 eq). The mixture was stirred at 25 °C for 1 h. The mixture was concentrated at reduced pressure to give a residue. The reaction mixture was purified by prep-HPLC (column: Welch Xtimate C18 150*25 mm*5um; mobile phase: [water (FA) -ACN]; gradient: 35%-65% B over 15 min) to give (20) (22.73 mg, 51.73 pmol, 28.06% yield, 100% purity) as an off-white solid.1H NMR: (400MHz, CDCI3) 6 = 7.67 - 7.58 (m, 4H), 6.05 (s, 1 H), 3.38 (d, J = 6.3 Hz, 2H), 3.20 (s, 3H), 2.91 - 2.74 (m, 3H), 2.58 - 2.44 (m, 2H), 2.40 (s, 3H); LCMS: RT = 0.502 min, m / z = 440.0 (M+H)+.

[0475] EXAMPLE 1.21: SYNTHESIS OF (21), (25) AND (38)

[0476] 5 (25)

[0477] Scheme 22: Synthesis of (25), (21) and (38)

[0478] Synthesis of compound 2, (Scheme 22)

[0479] To a solution of AlCh (4.69 g, 35.20 mmol, 1.92 mL, 6 eq) in DCM (15 mL) was added dropwise the solution of compound 1 (1.65 g, 5.87 mmol, 1 eq) in DCM (15 mL). The mixture was stirred at 40 °C for 16 h. The reaction mixture was poured into 1 M HCI (20 mL) slowly under 0 °C, some solid were separated out. Then to the resulting mixture was added MeOH (10 mL). The resulting mixture was filtered and the filter cake was collected and dried under reduced pressure. The crude product was triturated with MeOH at 20 °C for 16 h. The mixture was filtered and the filter cake was collected and dried under reduced pressure. Compound 2 (1 .5 g, crude) was obtained as a brown solid. LCMS: RT = 0.332 min, m / z = 268.0 (M+H)+.

[0480] Synthesis of compound 4, (Scheme 22)

[0481] To the mixture of compound 2 (1 .4 g, 5.24 mmol, 1 eq) in MeCN (10 mL) was added KOH (6 M, 10.06 mL, 11 .52 eq). Then to the mixture was added compound 3 (3.14 g, 15.72 mmol, 3 eq). The resulting mixture was stirred at 25 °C for 2 h. The mixture was diluted with water (20 mL) and the resulting mixture was extracted with Ethyl acetate (20 mL * 3). The combined organic layers were washed with brine (10 mL * 2), dried overNa2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (column: Phenomenex luna C18 150*40 mm* 15um; mobile phase: [water (FA) -AON]; gradient: 20%- 50% B over 15 min) Compound 4 (720 mg, 2.27 mmol, 43.32% yield, 100% purity) was obtained as off-white solid. LCMS: RT = 0.416 min, m / z = 318.0 (M+H)+.

[0482] Synthesis of compound 5, (Scheme 22)

[0483] To the mixture of compound 4 (200 mg, 630.48 pmol, 1 eq) in EtOH (3 mL) was added

[0484] Pd / C (50 mg, 10% purity) under Ar atmosphere. The mixture was stirred at 25 °C for 2 h under H2 (30 psi). The mixture was filtered. 0.1 mL of HCI (1 M) was added and the combined filtrate was concentrated at reduced pressure to give compound 5 (176 mg, 516.53 pmol, 81.93% yield, 95% purity, HCI) as a yellow oil.1H NMR: (400 MHz, CDCb) 6 = 7.41 - 7.28 (m, 2H), 7.26 - 7.20 (m, 1 H), 6.67 (t, J = 73.2 Hz, 2H), 2.97 (s, 3H), 2.27 (s, 3H).

[0485] Synthesis of compound (25), (Scheme 22)

[0486] To the mixture of compound 5 (55 mg, 169.91 pmol, 1 eq, HCI) in DCM (1 mL) was added pyridine (40.32 mg, 509.73 pmol, 41.14 pL, 3 eq) and compound 6 (38.25 mg, 169.91 pmol, 1 eq). The mixture was stirred at 25 °C for 18 h. LCMS showed compound 5 was remained, then to the mixture was added compound 6 (35 mg, 155.49 pmol, 9.15e-1 eq), pyridine (40 mg). The mixture was stirred at 25 °C for 3 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: 40%- 70% B over 10 min). HNMR showed the purity was not enough. The residue was purified by prep-HPLC (column: Welch Ultimate XB-SiOH 250*50*1 Oum; mobile phase: [Hexane-EtOH]; gradient: 5%- 45% B over 15 min). Compound (25) (31 .68 mg, 66.57 pmol, 39.18% yield, 100% purity) was obtained as an off-white solid.1H NMR: (400 MHz, CDCb) 6 = 7.83 (dd, J = 1 .8, 7.4 Hz, 1 H), 7.40 (dd, J = 1.3, 7.8 Hz, 1 H), 7.35 - 7.28 (m, 3H), 7.27 - 7.19 (m, 2H), 6.93 (s, 1 H), 6.67 (t, J = 72.9 Hz, 1 H), 4.65 (s, 2H), 3.22 (s, 3H), 2.39 (s, 3H); LCMS: RT = 0.507 min, m / z = 476.0 (M+H)+.

[0487] Synthesis of compound (21), (Scheme 22)

[0488] To the mixture of compound 5 (55 mg, 169.91 pmol, 1 eq, HCI) in DCM (1 .5 mL) was added pyridine (40.32 mg, 509.73 pmol, 41.14 pL, 3 eq) and compound 7 (35.45 mg, 169.91 pmol, 1 eq). The mixture was stirred at 25 °C for 18 h. LCMS showed compound 5 was remained, then to the mixture was added compound 7 (35 mg, 167.76 pmol, 9.87e-1 eq), pyridine (40 mg). The mixture was stirred at 25 °C for 3 h. The reaction mixture 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: 30%- 60% B over 10 min). Compound (21) (45.84 mg, 99.78 pmol, 58.72% yield, 100% purity) was obtained as a white solid.1H NMR: (400 MHz, CDCb) 6 = 7.47 - 7.42 (m, 2H), 7.36 - 7.28 (m, 4H), 7.08 - 7.00 (m, 1 H), 6.85 - 6.45 (m, 1 H), 5.91 (s, 1 H), 4.48 (s, 2H), 3.18 (s, 3H), 2.37 (s, 3H); LCMS: RT = 0.500 min, m / z = 460 .0 (M+H)+.

[0489] Synthesis of compound (38), (Scheme 22)

[0490] To the mixture of compound 5 (55 mg, 169.91 pmol, 1 eq, HCI) in DCM (1 .5 mL) was added pyridine (40.32 mg, 509.73 pmol, 41.14 pL, 3 eq) and compound 8 (35.45 mg, 169.91 pmol, 1 eq). The mixture was stirred at 25 °C for 18 h. LCMS showed compound 5 was remained, then to the mixture was added compound 8 (35 mg, 167.76 pmol, 9.87e-1 eq) and pyridine (40 mg). The mixture was stirred at 25 °C for 3 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: 40%- 70% B over 10 min). Compound (38) (50.05 mg, 107.85 pmol, 63.48% yield, 99% purity) was obtained as a white solid.1H NMR: (400 MHz, CDCb) 6 = 7.71 - 7.61 (m, 2H), 7.36 - 7.27 (m, 2H), 7.27 - 7.24 (m, 1 H), 7.08 - 6.98 (m, 2H), 6.89 - 6.45 (m, 1 H), 6.18 (s, 1 H), 4.41 (s, 2H), 3.19 (s, 3H), 2.37 (s, 3H); LCMS: RT = 0.498 min, m / z = 460.0 (M+H)+.

[0491] EXAMPLE 1.22: SYNTHESIS OF (22)

[0492] Scheme 23: Synthesis of (22)

[0493] To a solution of compound 2 (20 mg, 97.74 pmol, 1.32 eq) in DCM (1 mL) was added compound 1 (20 mg, 74.28 pmol, 1 eq) and pyridine (20 mg, 252.84 pmol, 20.41 pL, 3.40 eq) at 0 °C, then the resulting mixture was stirred at 20 °C for 12 h. The reaction mixture was quenched by addition of H2O (30 mL), and then the resulting mixture was extracted with EtOAc (15 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: Phenomenex luna C18 150*25 mm* 10um; mobile phase: [water (FA) -ACN]; gradient: 35%- 65% B over 10 min) to give compound (22) (14.96 mg, 31 .98 pmol, 43.05% yield, 93.5% purity) as a white solid.1H NMR :(400 MHz, CDCb) 6 = 7.50 (t, J = 8.2 Hz, 1 H), 7.25 (br d, J = 8.9 Hz, 1 H), 7.18 (br s, 1 H), 7.13 (br d, J = 8.4 Hz, 1 H), 6.87 - 6.62 (m, 1 H), 6.44 (br s, 1 H), 3.35 (br d, J = 5.4 Hz, 2H), 3.22 (s, 3H), 2.86 - 2.74 (m, 3H), 2.53 - 2.41 (m, 2H), 2.39 (s, 3H); LCMS: RT = 0.499 min, m / z = 436.2 (M-H)+.

[0494] EXAMPLE 1.23: SYNTHESIS OF (23) Scheme 24: Synthesis of (23)

[0495] Synthesis of compound 2, (Scheme 24)

[0496] A mixture of compound 1 (300 mg, 908.79 pmol, 1 eq), Fe (253.76 mg, 4.54 mmol, 5 eq), NH4CI (486.12 mg, 9.09 mmol, 10 eq) in MeOH (8 mL) and H2O (2 mL) was degassed and purged with N2 for 3 times, and then the mixture was stirred at 80 °C for 2 h under N2 atmosphere. The mixture was cooled to room temperature and concentrated under reduced pressure to give a residue. The residue was diluted with water (10 mL) and stirred for 5 min. The aqueous phase was extracted with EtOAc (10 mL x 2). The combined organic phase was washed with brine (10 mL), dried with anhydrous Na2SO4, filtered and concentrated in vacuum to give compound 2 (20 mg, crude) as white solid. LCMS: RT = 0.386 min, m / z = 302.1 (M+H)+.

[0497] Synthesis of compound 4, (Scheme 24)

[0498] A mixture of compound 2 (120 mg, 399.83 pmol, 1 eq), compound 3 (98.18 mg, 479.80 pmol, 1 .2 eq), pyridine (94.88 mg, 1.20 mmol, 96.82 pL, 3 eq) in DCM (1 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 cooled to room temperature and concentrated under reduced pressure to give a residue. The residue was diluted with water (10 mL) and stirred for 5 min. The aqueous phase was extracted with EtOAc (10 mL x 2). The combined organic phase was washed with brine (10 mL), dried with anhydrous Na2SO4, filtered and concentrated in vacuum to give a residue. The residue was purified by reversed-phase HPLC (0.1 % NH3 H2O condition), the eluent was concentrated and then freeze dried to give compound 4 (50 mg, crude) as white solid. LCMS: RT = 0.498 min, m / z = 468.1 (M+H)+.

[0499] Synthesis of (23), (Scheme 24)

[0500] A mixture of compound 4 (50 mg, 106.77 pmol, 1 eq), compound 6 (27.51 mg, 320.32 pmol, 3 eq), CsF (32.44 mg, 213.54 pmol, 7.88 pL, 2 eq), Pd(dppf)Cl2 (7.81 mg, 10.68 pmol, 0.1 eq) in dioxane (1 mL) was degassed and purged with N2 for 3 times, and then the mixture was stirred at 80 °C for 16 h under N2 atmosphere. The mixture was cooled to room temperature and concentrated under reduced pressure to give a residue. The residue was diluted with water (10 mL) and stirred for 5 min. The aqueous phase was extracted with EtOAc (5 mL x 2). The combined organic phase was washed with brine (10 mL), dried with anhydrous Na2SO4, filtered and 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: 40%- 70% B over 10 min) to give (23) (13.65 mg, 31 .47 pmol, 29.47% yield, 99% purity) as white solid.1H NMR: (400 MHz, CDCI3) 6 = 7.17 - 7.08 (m, 2H), 6.91 - 6.85 (m, 1 H), 6.37 (s, 1 H), 3.35 (br d, J = 6.1 Hz, 2H), 3.16 (s, 3H), 2.87 - 2.74 (m, 3H), 2.54 - 2.40 (m, 2H), 2.36 (s, 3H), 2.16 - 2.08 (m, 1 H), 1 .08 - 1 .02 (m, 2H), 0.82 - 0.76 (m, 2H); LCMS: RT = 0.514 min, m / z =430.2 (M+H)+. EXAMPLE 1.24: SYNTHESIS OF (24)

[0501] Scheme 25: Synthesis of (24)

[0502] Synthesis of compound 3, (Scheme 25)

[0503] A mixture of compound 2 (11 .18 g, 85.90 mmol, 10.87 mL, 1 eq), compound 1 (15 g, 85.90 mmol, 1 eq) in AcOH (20 mL) was degassed and purged with N2 for 3 times, and then the mixture was stirred at 100 °C for 1 h under N2 atmosphere. 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 to give compound 3 (4.4 g, 20.68 mmol, 24.08% yield, 96% purity) as white solid. LCMS: RT = 0.426 min, m / z = 205.2 (M+H)+.

[0504] Synthesis of compound 4, (Scheme 25)

[0505] A mixture of compound 3 (1 .4 g, 6.86 mmol, 1 eq), Mel (973.02 mg, 6.86 mmol, 426.76 pL, 1 eq) in ACN (5 mL) was degassed and purged with N2 for 3 times, and then the mixture was stirred at 120 °C for 1 h under microwave reactor. The mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (silica gel, DCM / MeOH = 1 : 0 to 5: 1 , TLC: DCM: MeOH = 10: 1 , Rf = 0.43) to give compound 4 (3 g, crude) as white solid. LCMS: RT = 0.408 min, m / z = 219.0 (M+H)+.

[0506] Synthesis of compound 5, (Scheme 25)

[0507] To a mixture of compound 4 (3 g, 13.75 mmol, 1 eq) in TFA (5 mL) was added HNO3 (2.27 g, 23.42 mmol, 1 .62 mL, 65% purity, 1 .70 eq) slowly at - 20 °C under N2 atmosphere. Then the mixture was stirred at - 20 °C for 1 h. The reaction mixture was poured into ice water (50 mL), and the pH of mixture was neutralized by adding sodium bicarbonate to about 7. The mixture was filtrate and the filter cake was collected and dried by oil pump. Then the mixture was filtrate extracted with EtOAc (40 mL * 3). The combined organic layers were washed with brine (20 mL * 2), dried over Na2SO4, filtered and concentrated under reduced pressure to give crude product. The crude product (combined with filter cake) was triturated with Ethyl acetate (15 mL) at 20 °C for 5 min to give compound 5 (2.1 g, 6.94 mmol, 50.49% yield, 87% purity) as white solid. LCMS: RT = 0.381 min, m / z = 264.1 (M+H)+.

[0508] Synthesis of compound 6, (Scheme 25)

[0509] A mixture of compound 5 (2.1 g, 7.98 mmol, 1 eq) in DCM (2 mL) was added BBrs (1 M, 9.57 mL, 1.2 eq) (in DCM) at 0 °C. Then the mixture was stirred at 20 °C for 16 h under N2 atmosphere. The reaction mixture was quenched by MeOH (10 mL), and then the pH of the resulting mixture was adjusted to about 7 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 residuet was triturated with Ethyl acetate (15 mL) at 20 °C for 5 min to give compound 6 (1.3 g, 4.49 mmol, 56.23% yield, 86% purity) as white solid. LCMS: RT = 0.319 min, m / z = 250.0 (M+H)+.

[0510] Synthesis of compound 8, (Scheme 25)

[0511] A mixture of compound 6 (1 .2 g, 4.81 mmol, 1 eq), compound 7 (1 .96 g, 9.63 mmol, 2 eq), KOH (5.40 g, 28.89 mmol, 30% purity, 6 eq) in DCM (3 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 reaction mixture was poured into ice water (20 mL), and the pH of mixture was neutralized by adding sodium bicarbonate to about 7. Then the mixture was extracted with EtOAc (20 mL * 3). The combined organic layers were washed with brine (10 mL * 2), dried over Na2SO4, filtered and 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: Petroleum ether: Ethyl acetate = 0: 1 , Rf = 0.23) to give compound 8 (750 mg, 1.38 mmol, 28.63% yield, 55% purity) as white solid. LCMS: RT = 0.406 min, m / z = 300.0 (M+H)+.

[0512] Synthesis of compound 9, (Scheme 25)

[0513] To a mixture of compound 8 (750 mg, 2.51 mmol, 1 eq) in EtOH (5 mL) was added Pd / C (673.08 mg, 632.47 pmol, 10% purity, 2.52e- 1 eq) at 20 °C. The mixture was degassed under vacuum and purged with H2 several times. The mixture was stirred under H2 (15 psi) at 20 °C for 4 h. The mixture was filtered and the filtrate was concentrated under vacuum to give compound 9 (500 mg, 1.32 mmol, 52.60% yield, 71 % purity) as colorless oil. LCMS: RT = 0.359 min, m / z = 270.1 (M+H)+.

[0514] Synthesis of (24), (Scheme 25) A mixture of compound 10 (85.27 mg, 389.98 pmol, 1.5 eq), compound 9 (70 mg, 259.98 pmol, 1 eq), pyridine (61.69 mg, 779.95 pmol, 62.95 pL, 3 eq) in DCM (2 mL) was degassed and purged with N2for 3 times, and then the mixture was stirred at 20 °C for 16 h under N2atmosphere. The 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) to give (24) (27.68 mg, 60.09 pmol, 23.11% yield, 98% purity) as white solid.1H NMR: (400 MHz, CDCb) 6 = 7.54 - 7.45 (m, 1 H), 7.24 (d, J = 7.2 Hz, 1 H), 7.19 - 7.09 (m, 2H), 6.60 (t, J = 73.3 Hz, 1 H), 6.1 1 (br s, 1 H), 3.28 - 3.22 (m, 1 H), 3.20 (s, 3H), 2.82 - 2.55 (m, 4H), 2.39 (s, 3H), 2.38 - 2.26 (m, 1 H), 1 .48 (d, J = 6.8 Hz, 3H); LCMS: RT = 0.496 min, m / z = 452.1 (M+H)+.

[0515] Synthesis of (24)-peak1 & (24)-peak2, (Scheme 25)

[0516] The compound (24) was purified by SFC (RT=1.571 min and 1.817 min; column: DAICEL CHIRALPAK AD (250 mm*30 mm, 10um); mobile phase: [CO2-EtOH (0.1%NH3H2O)]; B%: 21 %, isocratic elution mode) to give (24)-peak1 (11.13 mg, 23.67 pmol, 42.74% yield, 96% purity) as white solid.1H NMR: (400 MHz, CDCb) 6 = 7.52 - 7.46 (m, 1 H), 7.26 - 7.23 (m, 1 H), 7.18 - 7.10 (m, 2H), 6.59 (t, J = 73.2 Hz, 1 H), 3.27 (br t, J = 6.9 Hz, 1 H), 3.19 (s, 3H), 2.78 - 2.63 (m, 4H), 2.39 (s, 3H), 2.38 - 2.32 (m, 1 H), 1 .50 (d, J = 6.8 Hz, 3H); LCMS: RT = 0.557 min, m / z = 452.1 (M+H)+; SFC: RT = 1 .591 min. and (24)-peak2 (11.94 mg, 25.39 pmol, 45.85% yield, 96% purity) as white solid.1H NMR: (400 MHz, CDCb) 6 = 7.52 - 7.46 (m, 1 H), 7.26 - 7.23 (m, 1 H), 7.17 - 7.09 (m, 2H), 6.79 - 6.40 (m, 1 H), 3.29 - 3.22 (m, 1 H), 3.18 (s, 3H), 2.77 - 2.62 (m, 4H), 2.38 (s, 3H), 2.37 - 2.30 (m, 1 H), 1 .49 (d, J = 6.8 Hz, 3H); LCMS: RT = 0.552 min, m / z = 452.1 (M+H)+; SFC: RT = 1 .811 min.

[0517] EXAMPLE 1.25: SYNTHESIS OF (26) Scheme 26: Synthesis of (26)

[0518] Synthesis of compound 3, (Scheme 26)

[0519] A mixture of compound 1 (1 g, 5.19 mmol, 1 eq), compound 2 (675.64 mg, 5.19 mmol, 657.24 pL, 1 eq) in AcOH (5 mL) was degassed and purged with N2 for 3 times, and then the mixture was stirred at 100 °C for 1 h under N2 atmosphere. The mixture was concentrated under reduced pressure to give compound 3 (1.1 g, crude) as white solid. LCMS: RT = 0.435 min, m / z = 223.1 (M+H)+.

[0520] Synthesis of compound 4, (Scheme 26)

[0521] A mixture of compound 3 (1.1 g, 4.95 mmol, 1 eq), Mel (2.11 g, 14.85 mmol, 924.50 pL, 3 eq) in ACN (5 mL) was degassed and purged with N2 for 3 times, and then the mixture was stirred at 120 °C for 3 h under microwave reactor. The mixture was concentrated under reduced pressure to give a residue. The residue was purified by reversed-phase HPLC (0.1 % NHs’FW condition), the eluent was concentrated and then freeze dried to give compound 4 (500 mg, crude) as white solid. LCMS: RT = 0.408 min, m / z = 237.2 (M+H)+.

[0522] Synthesis of compound 5, (Scheme 26)

[0523] To a mixture of compound 4 (400 mg, 1.69 mmol, 1 eq) in TFA (2 mL) was added HNO3 (550 mg, 8.73 mmol, 392.86 pL, 5.16 eq) slowly at - 20 °C under N2 atmosphere. Then the mixture was stirred at - 20 °C for 1 h. The reaction mixture was poured into ice water (20 mL), and the pH of mixture was neutralized to about 7 by adding sodium bicarbonate. The mixture was filtrated and the filter cake was collected and dried by oil pump. Then the mixture was filtrate 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 (combined with the filter cake) was purified by column chromatography (silica gel, Dichloromethane / Methanol = 1 : 0 to 10: 1 , TLC: Petroleum ether: Ethyl acetate = 10: 1 , Rf = 0.51) to give compound 5 (200 mg, crude) as a white solid. LCMS: RT = 0.390 min, m / z = 282.2 (M+H)+.

[0524] Synthesis of compound 6, (Scheme 26)

[0525] To a mixture of compound 5 (170 mg, 604.47 pmol, 1 eq) in DCM (1 mL) was added BBrs (1 M, 725.36 pL, 1.2 eq) (in DCM) at 0 °C. Then the mixture was stirred at 20 °C for 1 h. The reaction mixture was quenched by MeOH (5 mL), and then the pH of the resulting mixture was adjusted to about 7 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 compound 6 (120 mg, crude) as white solid. Synthesis of compound 8, (Scheme 26)

[0526] To a mixture of compound 6 (110 mg, 411 .66 pmol, 1 eq) in DCM (1 mL) was added KOH (692.89 mg, 2.47 mmol, 20% purity, 6 eq) and compound 7 (250.82 mg, 1 .23 mmol, 3 eq) at 0 °C, then the mixture was stirred at 20 °C for 16 h. The pH of mixture was neutralized to about 7 by adding 1 M HCI. EtOAc (10 mL) and water (10 mL) were added and layers were separated. The aqueous phase was extracted with EtOAc (5 mL x 2). Combined extracts were washed with brine (10 mL), dried over Na2SO4, filtered, and concentrated under vacuum to give compound 8 (80 mg, crude) as white solid.

[0527] Synthesis of compound 9, (Scheme 26)

[0528] A mixture of compound 8 (80 mg, 252.19 pmol, 1 eq), Fe (70.42 mg, 1.26 mmol, 5 eq), NH4CI (134.90 mg, 2.52 mmol, 10 eq) in MeOH (5 mL) and H2O (1 mL) was degassed and purged with N2 for 3 times, and then the mixture was stirred at 80 °C for 2 h under N2 atmosphere. The reaction mixture was cooled to room temperature. EtOAc (10 mL) and water (10 mL) were added and layers were separated. The aqueous phase was extracted with EtOAc (5 mL x 2). Combined extracts were washed with brine (10 mL), dried over Na2SO4, filtered, and concentrated under vacuum to give compound 9 (50 mg, crude) as white solid.

[0529] Synthesis of (26), (Scheme 26)

[0530] A mixture of compound 10 (53.43 mg, 261.11 pmol, 1.5 eq), compound 9 (50 mg, 174.07 pmol, 1 eq), pyridine (68.85 mg, 870.36 pmol, 70.25 pL, 5 eq) in DCM (1 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 prep-HPLC (column: Welch Xtimate C18 150*25 mm*5um; mobile phase: [water (FA) -ACN]; gradient: 30%- 60% B over 10 min) to give a residue. Then residue was purified by prep-HPLC (column: Welch Ultimate XB-CN 250*50*1 Oum; mobile phase: [Hexane-EtOH]; gradient: 10%- 50% B over 15 min) to give (26) (3.46 mg, 7.14 pmol, 4.10% yield, 94% purity) as white solid.1H NMR: (400 MHz, CDCb) 6 = 7.35 - 7.28 (m, 2H), 7.27 - 7.23 (m, 1 H), 6.64 (t, J = 72.8 Hz, 1 H), 3.37 (d, J = 6.4 Hz, 2H), 3.19 (s, 3H), 2.90 - 2.75 (m, 3H), 2.57 - 2.44 (m, 2H), 2.38 (s, 3H); LCMS: RT = 0.482 min, m / z = 456.1 (M+H)+.

[0531] EXAMPLE 1.26: SYNTHESIS OF (27)

[0532] Scheme 27: Synthesis of (27)

[0533] Synthesis of compound 2, (Scheme 27)

[0534] To a solution of compound 1 (20 g, 109.26 mmol, 1 eq) in DCM (200 mL) at 0 °C was added triethylamine trihydrofluoride (35.23 g, 218.53 mmol, 35.62 mL, 2 eq), followed by NBS (21.39 g, 120.19 mmol, 1 .1 eq). After stirring at 20 °C for 14 h. EtOAc (70 mL) and water (60 mL) were added and Layers were separated. The aqueous phase was extracted with EtOAc (50 mL x 2). Combined extracts were washed with brine (40 mL), dried over Na2SO4, filtered, and concentrated under vacuum to give a residue. The residue was purified by flash silica gel chromatography (ISCO®; 120 g SepaFlash® Silica Flash Column, Eluent of 0% Ethyl acetate / Petroleum ethergradient @ 100 mL / min). Compound 2 (23.9 g, 84.77 mmol, 77.58% yield) was obtained as a colorless liquid.1H NMR: (400 MHz, CDCh) 6 = 7.57 - 7.48 (m, 2H), 7.30 (d, J = 5.4 Hz, 2H), 5.75 - 5.46 (m, 1 H), 3.81 - 3.34 (m, 2H).

[0535] Synthesis of compound 3, (Scheme 27)

[0536] To a solution of compound 2(23.9 g, 84.77 mmol, 1 eq) in THF (110 mL) was added ABuOK (11 .89 g, 105.96 mmol, 1 .25 eq) at 0 °C. The mixture was stirred at 20 °C for 16 h. Petroleum ether (30 mL) and water (80 mL) were added and Layers were separated. The aqueous phase was extracted with Petroleum ether (80 mL x 2). Combined extracts were washed with brine (70 mL), dried over Na2SO4, filtered, and concentrated under vacuum to give a residue. The residue was purified by flash silica gel chromatography (ISCO®; 120 g SepaFlash® Silica Flash Column, Eluent of 0% Ethyl acetate / Petroleum ether gradient @ 100 mL / min). Compound 3 (14.4 g, 71.63 mmol, 84.50% yield) was obtained as a colorless liquid.1H NMR: (400 MHz, CDCh) 6 = 7.71 (t, J = 1 .7 Hz, 1 H), 7.54 - 7.47 (m, 2H), 7.27 (s, 1 H), 5.16 - 4.98 (m, 1 H), 4.91 (dd, J = 3.8, 17.6 Hz, 1 H).

[0537] Synthesis of compound 4, (Scheme 27) A mixture of compound 3 (14.4 g, 71.63 mmol, 1 eq), NaOH (20 M, 140 mL, 39.09 eq) and benzyl(triethyl) ammonium; bromide (1.95 g, 7.16 mmol, 3.90 mL, 0.1 eq) in CHBrs (288.90 g, 1.14 mol, 100 mL, 15.96 eq) was stirred at 0 °C for 3 h. DCM (100 mL) and water (300 mL) were added and Layers were separated. The aqueous phase was extracted with DCM (100 mL x 2). Combined extracts were washed with brine (100 mL), dried over Na2SO4, filtered, and 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 0% Ethyl acetate / Petroleum ether gradient @ 100 mL / min). Compound 4 (10.8 g, 28.97 mmol, 40.44% yield) was obtained as a yellow oil.1H NMR: (400 MHz, CDCb) 6 = 7.64 (s, 1 H), 7.59 (dd, J = 0.8, 7.9 Hz, 1 H), 7.46 - 7.38 (m, 1 H), 7.37 - 7.29 (m, 1 H), 2.51 - 2.35 (m, 2H).

[0538] Synthesis of compound 5, (Scheme 27)

[0539] The mixture of compound 4 (10.8 g, 28.97 mmol, 1 eq), Zn (14.02 g, 214.35 mmol, 7.4 eq), NH4CI (13.94 g, 260.69 mmol, 9 eq) and MeOH (200 mL) was warmed at 70 °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®; 120 g SepaFlash® Silica Flash Column, Eluent of 0% Ethyl acetate / Petroleum ether gradient @ 100 mL / min). Compound 5 (2.48 g, 11.53 mmol, 39.81 % yield) was obtained as a colorless liquid.1H NMR: (400 MHz, CDCb) 6 = 7.50 - 7.40 (m, 2H), 7.33 - 7.14 (m, 2H), 1 .60 - 1 .47 (m, 2H), 1 .17 - 1 .05 (m, 2H).

[0540] Synthesis of compound 6, (Scheme 27)

[0541] To a solution of compound 5 (1.1 g, 5.11 mmol, 1 eq) in dioxane (30 mL) was added tertbutyl A / -aminocarbamate (743.57 mg, 5.63 mmol, 1.1 eq), ditert-butyl-[2-(2, 4, 6- triisopropylphenyl) phenyl]phosphane (217.20 mg, 511.48 pmol, 0.1 eq) Pd2(dba)3 (234.19 mg, 255.74 pmol, 0.05 eq) and CS2CO3 (3.33 g, 10.23 mmol, 2 eq). The mixture was stirred at 80 °C for 2 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 flash silica gel chromatography (ISCO®; 12 g SepaFlash® Silica Flash Column, Eluent of 0~20%Ethylacetate / Petroleum ether gradient @ 35 mL / min). Compound 6 (360 mg, 986.82 pmol, 19.29% yield, 73% purity) was obtained as a yellow oil. LCMS: RT = 0.542 min, m / z = 211 .2 (M-tBu+H)+.

[0542] Synthesis of compound 7, (Scheme 27)

[0543] To a solution of compound 6 (360 mg, 1 .35 mmol, 1 eq) in EtOAc (12 mL) was added HCI / EtOAc (4 M, 4 mL). The mixture was stirred at 20 °C for 14 h. The mixture was concentrated under vacuum to give a residue. Compound 7 (300 mg, crude, HCI) was obtained as a yellow oil. LCMS: RT = 0.361 min, m / z = 167.2 (M+H)+. Synthesis of compound 9, (Scheme 27)

[0544] A solution of compound 7 (300 mg, 1.81 mmol, 1 eq) and compound 8 (469.84 mg, 3.61 mmol, 457.04 pL, 2 eq) in AcOH (5 mL) was stirred under an argon atmosphere at 100 °C for 1 h. Most of AcOH was removed after concentration under reduced pressure to give a residue. The residue was poured into NaHCOs (2 M, 15 mL). The aqueous Layer was extracted with EtOAc (20 mL). The organic Layer was washed with brine (15 mL), dried over Na2SO4 and filtered. The filtrate 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 50-70% Ethyl acetate / Petroleum ether gradient @ 45 mL / min). Compound 9 (130 mg, 537.35 pmol, 29.77% yield, 96% purity) was obtained as a yellow oil. LCMS: RT = 0.477 min, m / z = 233.2 (M+H)+.

[0545] Synthesis of compound 10, (Scheme 27)

[0546] Compound 9 (104 mg, 447.79 pmol, 1 eq) and Mel (912.00 mg, 6.43 mmol, 400.00 pL, 14.35 eq) were taken up into a microwave tube in ACN (0.5 mL). The sealed tube was heated at 120 °C for 3 h under microwave. DCM (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 flash silica gel chromatography (ISCO®; 12 g SepaFlash® Silica Flash Column, Eluent of 60-80% Ethyl acetate / Petroleum ethergradient @ 40 mL / min). Compound 10 (150 mg, 395.89 pmol, 88.41 % yield, 65% purity) was obtained as a yellow oil. LCMS: RT = 0.438 min, m / z = 247.3 (M+H)+.

[0547] Synthesis of compound 11 , (Scheme 27)

[0548] To a solution of compound 10 (150 mg, 609.06 pmol, 1 eq) in TFA (1 mL) and DCM (1 mL) was added HNO3 (169.32 mg, 1.83 mmol, 120.94 pL, 68% purity, 3 eq) at 0 °C. The mixture was stirred at 0 °C for 1 h. Most of TFA was removed after concentration under reduced pressure to give a residue. The residue was poured into NaHCOs (2 M, 15 mL). The aqueous Layer was extracted with EtOAc (20 mL). The organic Layer was washed with brine (15 mL), dried over Na2SO4 and filtered. The filtrate was concentrated under reduced pressure to give a residue. The residue was purified by prep-TLC (SiO2, PE: EtOAc = 0: 1 , TLC: PE: EtOAc = 0: 1 , Rf = 0.26). Compound 11 (43 mg, 110.72 pmol, 18.18% yield, 75% purity) was obtained as a yellow solid. LCMS: RT = 0.431 min, m / z = 292.1 (M+H)+.

[0549] Synthesis of compound 12, (Scheme 27)

[0550] To a solution of compound 11 (35 mg, 120.16 pmol, 1 eq) in EtOH (2 mL) and H2O (0.5 mL) was added Fe (67.10 mg, 1.20 mmol, 10 eq) and NH4CI (102.84 mg, 1.92 mmol, 16 eq). The mixture was stirred at 80 °C for 2 h. The filtrate was concentrated in vacuum to give a residue. The residue was diluted with H2O (5 mL) and the resulting mixture was extracted by EtOAc (6 mL x 2). The combined organic phase was washed with brine, dried over anhydrous Na2SO4 and concentrated in vacuum. Compound 12 (34 mg, crude) was obtained as a yellow solid. LCMS: RT = 0.382 min, m / z = 262.2 (M+H)+.

[0551] Synthesis of compound (27), (Scheme 27)

[0552] To a solution of compound 12 (20 mg, 76.54 pmol, 1 eq) in DCM (1 mL) was added pyridine (121 .09 mg, 1 .53 mmol, 123.56 pL, 20 eq) and (2 -fluorophenyl) methanesulfonyl chloride (31 .94 mg, 153.08 pmol, 2 eq). The mixture was stirred at 20 °C for 4 h. DCM (5 mL) and water (6 mL) were added and Layers were separated. The aqueous phase was extracted with DCM (5 mL x 2). Combined extracts were washed with brine (6 mL), dried over Na2SO4, filtered, and concentrated under 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: 40%-70% B over 10 min). Compound (27) (9.35 mg, 20.92 pmol, 27.34% yield, 97% purity) was obtained as a yellow solid.1H NMR: (400 MHz, CDCb) 6 = 7.91 (dt, J = 1 .4, 7.5 Hz, 1 H), 7.55 - 7.47 (m, 1 H), 7.36 - 7.28 (m, 3H), 7.21 (d, J = 7.9 Hz, 1 H), 7.16 - 7.06 (m, 2H), 6.12 (br s, 1 H), 4.59 (s, 2H), 3.22 (s, 3H), 2.39 (s, 3H), 1 .58 - 1 .50 (m, 2H), 1 .20 - 1 .10 (m, 2H); LCMS: RT = 0.503 min, m / z = 434.1 (M+H)+.

[0553] EXAMPLE 1.27: SYNTHESIS OF (29)

[0554] Scheme 28: Synthesis of (29)

[0555] To a solution of compound 2 (41.49 mg, 184.34 pmol, 1 eq) in DCM (0.5 mL) was added pyridine (43.74 mg, 553.02 pmol, 44.64 pL, 3 eq) and compound 1 (50 mg, 184.34 pmol, 1 eq). The mixture was stirred at 25 °C for 1 h. The mixture was concentrated at reduced pressure to give a residue. The reaction mixture was purified by prep-HPLC (column: Welch Xtimate C18 150*25 mm*5um; mobile phase: [water (FA) -ACN]; gradient: 35%-65% B over 15 min) to give (29) (16.51 mg, 35.90 pmol, 19.48% yield, 100% purity) as an off-white solid.1H NMR: (400 MHz, CDCb) 6 = 7.98 - 7.93 (m, 1 H), 7.69 - 7.60 (m, 4H), 7.46 - 7.41 (m, 1 H), 7.32 - 7.27 (m, 2H), 5.91 - 5.80 (m, 1 H), 4.75 (s, 2H), 3.22 (s, 3H), 2.41 (s, 3H); LCMS: RT = 0.519 min, m / z = 460.1 (M+H)+.

[0556] EXAMPLE 1.28: SYNTHESIS OF (30)

[0557] Scheme 29: Synthesis of (30)

[0558] To the mixture of compound 1 (35 mg, 84.24 pmol, 1 eq) in DMF (1 mL) was added NaH (4.04 mg, 101.09 pmol, 60% purity, 1.2 eq) at 0 °C, the mixture was stirred at 20 °C for 0.5 h. To the mixture was added Mel (13.15 mg, 92.66 pmol, 5.77 pL, 1.1 eq) at 0 °C. The mixture was stirred at 20 °C for 1 h. To the mixture was added saturated NH4CI solution (10 mL) and the resulting mixture was extracted with EtOAc (10 mL x 3), the combined organic phase was 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: 45%-75% B over 10 min) to give compound (30) (12.41 mg, 28.60 pmol, 33.96% yield, 99% purity) as white solid.1H NMR: (400 MHz, CDCb) 6 = 7.79 (dt, J = 1 .4, 7.4 Hz, 1 H), 7.41 - 7.35 (m, 1 H), 7.35 - 7.29 (m, 1 H), 7.18 - 7.09 (m, 4H), 7.08 - 7.03 (m, 1 H), 4.67 - 4.52 (m, 2H), 3.25 (s, 3H), 3.20 (s, 3H), 2.34 (s, 3H), 2.02 - 1.91 (m, 1 H), 1 .06 - 0.98 (m, 2H), 0.81 - 0.71 (m, 2H); LCMS: RT = 0.532 min, m / z = 430.2 (M+H)+.

[0559] EXAMPLE 1.29: SYNTHESIS OF (31)

[0560] Scheme 30: Synthesis of (31)

[0561] Synthesis of compound 7, (Scheme 30)

[0562] To a solution of compound 4 (200 mg, 640.78 pmol, 1 eq) in dioxane (2 mL) and H2O (0.5 mL) was added Pd(dppf)Cl2 (23.44 mg, 32.04 pmol, 0.05 eq), Na2COs (135.83 mg, 1.28 mmol, 2 eq) and methylboronic acid (383.57 mg, 6.41 mmol, 10 eq). The mixture was diluted with H2O (30 mL) and the resulting mixture was extracted with EtOAc (10 mL * 4). 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, PE / EtOAc = 1 / 1 to 0 / 1 ; TLC: PE / EtOAc = 0: 1, Rf = 0.35), the eluent was concentrated and then under reduced pressure to give compound 7 (40 mg, crude) as brown solid. LCMS: RT = 0.386 min, m / z = 248.1 (M+H)+.

[0563] Synthesis of compound 7, (Scheme 30)

[0564] To a solution of compound 7 (40 mg, 161.78 pmol, 1 eq) in EtOH (1 mL) and H2O (1 mL) was added Fe (54.21 mg, 970.68 pmol, 6 eq) and NH4CI (86.54 mg, 1.62 mmol, 10 eq). The mixture was stirred at 80 °C for 2 h. The mixture was filtered, the filtrate was collected under reduced pressure to give residue. The residue was diluted with H2O (20 mL) and 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 8 (50 mg, crude) as brown gum. LCMS: RT = 0.357 min, m / z = 218.3 (M+H)+.

[0565] Synthesis of (31), (Scheme 30)

[0566] To a solution of compound 8 (50 mg, 230.13 pmol, 1 eq) and (2-fluorophenyl) methanesulfonyl chloride (48.01 mg, 230.13 pmol, 1 eq) in DCM (1 mL) was added Py (54.61 mg, 690.40 pmol, 55.72 pL, 3 eq) at 0 °C. The mixture was stirred at 20 °C for 16 h. The 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]; gradient: 38%-68% B over 10 min), the eluent was concentrated and then freeze dried to give (31) (14.62 mg, 36.41 pmol, 15.82% yield, 97% purity) as off-white solid.1H NMR: (400 MHz, CDCI3) 6 = 7.94 (t, J = 7.5 Hz, 1 H), 7.43 - 7.37 (m, 1 H), 7.36 - 7.29 (m, 1 H), 7.24 - 7.06 (m, 5H), 6.09 (br s, 1 H), 4.61 (s, 2H), 3.22 (s, 3H), 2.43 (s, 3H), 2.38 (s, 3H); LCMS: RT = 0.481 min, m / z = 390.2 (M+H)+.

[0567] EXAMPLE 1.30: SYNTHESIS OF (32)

[0568] Scheme 31 : Synthesis of (32)

[0569] Synthesis of compound 2, (Scheme 31)

[0570] To the mixture of compound 1 (50 mg, 367.27 pmol, 1 eq), DIEA (189.87 mg, 1 .47 mmol, 255.89 pL, 4 eq) in DCM (1 mL) was added MsCI (110 mg, 960.27 pmol, 74.32 pL, 2.61 eq) at 0 °C. The mixture was stirred at 0 °C for 1 h under N2. The mixture was quenched by saturated NH4CI (20 mL), 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 compound 2 (80 mg, crude) as brown oil.1H NMR: (400 MHz, CDCh) 6 = 4.82 (quin, J = 6.3 Hz, 1 H), 3.04 (s, 3H), 2.73 - 2.61 (m, 2H), 2.59 - 2.50 (m, 1 H), 2.47 - 2.32 (m, 2H), 1 .41 (s, 3H).

[0571] Synthesis of compound 3, (Scheme 31)

[0572] To the mixture of compound 2 (80 mg, 373.43 pmol, 1 eq) in DMF (1 mL) was added acetylsulfanylpotassium (110 mg, 963.16 pmol, 2.58 eq). The mixture was stirred at 60 °C for 2 h. The reaction mixture was cooled to room temperature and diluted with water (10 mL). The resulting mixture was extracted with EtOAc (10 mL*3). The combined organic phase were washed with brine (10 mL), dried over Na2SO4, filtered, and concentrated under vacuum to compound 3 (60 mg, crude) as brown oil.1H NMR: (400 MHz, CDCh) 6 = 3.64 - 3.51 (m, 1 H), 2.72 - 2.54 (m, 2H), 2.34 (s, 3H), 2.32 - 2.18 (m, 3H), 1 .28 (d, J = 6.9 Hz, 3H).

[0573] Synthesis of compound 4, (Scheme 31)

[0574] A mixture of compound 3 (60 mg, 308.89 pmol, 1 eq), NCS (164.99 mg, 1.24 mmol, 4 eq), HCI (2 M, 308.89 pL, 2 eq) in ACN (1 mL) was degassed and purged with N2 for 3 times, and then the mixture was stirred at 0 °C for 1 h under N2 atmosphere. The reaction mixture was cooled to room temperature. EtOAc (10 mL) and water (10 mL) were added and layers were separated. The aqueous phase was extracted with EtOAc (5 mL x 2). Combined extracts were washed with brine (10 mL), dried over Na2SO4, filtered, and concentrated under vacuum to give compound 4 (60 mg, crude) as yellow oil.1H NMR: (400 MHz, CDCh) 6 = 3.70 - 3.61 (m, 1 H), 2.87 - 2.78 (m, 2H), 2.71 - 2.63 (m, 2H), 2.48 - 2.36 (m, 1 H), 1 .58 (d, J = 6.8 Hz, 3H).

[0575] Synthesis of (32), (Scheme 31)

[0576] To a mixture of compound 5 (103.66 mg, 365.88 pmol, 1 eq), pyridine (144.71 mg, 1.83 mmol, 147.66 pL, 5 eq) in DCM (2 mL) was added compound 4 (80 mg, 365.88 pmol, 1 eq) at 20 °C. Then the mixture was stirred at 20 °C for 16 h under N2 atmosphere. The reaction mixture was cooled to room temperature. EtOAc (40 mL) and water (40 mL) were added and layers were separated. The aqueous phase was extracted with EtOAc (30 mL x 2). Combined extracts were washed with brine (60 mL), dried over Na2SO4, filtered, and concentrated under 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: 33%- 63% B over 10 min) to give (32) (25 mg, 49.94 pmol, 13.65% yield, 93% purity) as white solid.

[0577] Synthesis of (32)-peak1 & (32)-peak2 , (Scheme 31) The (32) was purified by SFC (RT= 2.191 min and 2.61 min; column: DAICEL CHIRALPAK IC (250 mm*30 mm, 10um); mobile phase: [CO2 -EtOH (0.1 %NH3H2O)]; B%: 50%, isocratic elution mode) to give (32)-peak1 (8.15 mg, 16.81 pmol, 31.30% yield, 96% purity) as white solid.1H NMR: (400 MHz, CDCh) 6 = 7.79 (s, 1 H), 7.70 (s, 1 H), 7.50 - 7.43 (m, 3H), 7.18 (ddd, J = 2.1 , 3.4, 5.5 Hz, 1 H), 3.96 (s, 3H), 3.34 - 3.25 (m, 1 H), 3.21 (s, 3H), 2.76 - 2.62 (m, 4H), 2.40 (s, 3H), 2.39 - 2.30 (m, 1 H), 1 .51 (d, J = 6.8 Hz, 3H); LCMS: RT = 0.464 min, m / z = 466.2 (M+H)+; SFC: RT = 2.196 min.

[0578] (32)-peak2 (9.53 mg, 19.04 pmol, 35.45% yield, 93% purity) as white solid.1H NMR: (400 MHz, CDCh) 6 = 7.79 (s, 1 H), 7.71 (s, 1 H), 7.51 - 7.44 (m, 3H), 7.20 - 7.16 (m, 1 H), 3.96 (s, 3H), 3.33 - 3.25 (m, 1 H), 3.21 (s, 3H), 2.77 - 2.63 (m, 4H), 2.39 (s, 3H), 2.38 - 2.31 (m, 1 H), 1 .50 (d, J = 6.8 Hz, 3H); LCMS: RT = 0.487 min, m / z = 466.2 (M+H)+; SFC: RT = 2.611 min.

[0579] EXAMPLE 1.31: SYNTHESIS OF (33)

[0580] Scheme 32: Synthesis of (33)

[0581] A mixture of compound 1 (49.45 mg, 212.53 pmol, 1 eq), compound 2 (50 mg, 212.53 pmol, 1 eq), pyridine (50.43 mg, 637.60 pmol, 51.46 pL, 3 eq) in DCM (1 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 reaction mixture was cooled to room temperature. EtOAc (10 mL) and water (10 mL) were added and layers were separated. The aqueous phase was extracted with EtOAc (5 mL x 2). Combined extracts were washed with brine (6 mL), dried over Na2SO4, filtered, and concentrated under 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: 40%- 70% B over 10 min) to give (33) (5.95 mg, 13.38 pmol, 6.29% yield, 97% purity) as white solid.1H NMR: (400 MHz, CDCh) 6 = 7.20 (br d, J = 6.6 Hz, 1 H), 7.15 - 7.09 (m, 2H), 6.09 (br s, 1 H), 3.17 (s, 3H), 3.15 (s, 2H), 2.37 (s, 3H), 2.33 (d, J = 1 .8 Hz, 3H), 2.14 - 2.01 (m, 5H), 1 .79 - 1 .65 (m, 2H), 1 .48 - 1 .38 (m, 2H); LCMS: RT = 0.502 min, m / z = 432.2 (M+H)+.

[0582] EXAMPLE 1.32: SYNTHESIS OF (34)

[0583] Scheme 33: Synthesis of (34)

[0584] Synthesis of compound 3, (Scheme 33)

[0585] To the mixture of compound 1 (25 g, 111 .86 mmol, 1 eq, HCI) in AcOH (60 mL) was added compound 2 (14.56 g, 111 .86 mmol, 14.16 mL, 1 eq). The mixture was stirred at 100 °C for 1 h. The reaction mixture diluted with EtOAc (60 mL), the resulting mixture was stirred at 25 °C for 2 h. The mixture was filtered, the filter cake was collected and dried under reduced pressure to give compound 3 (22 g, crude) as a white solid, the filtrate was discarded.

[0586] Synthesis of compound 4, (Scheme 33)

[0587] To the mixture of compound 3 (1 g, 3.95 mmol, 1 eq) in ACN (120 mL) was added Mel (3.36 g, 23.71 mmol, 1 .48 mL, 6 eq). The mixture was stirred at 120 °C for 4 h under microwave. Twelve batches of the reactions were combined together. The mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (column: Phenomenex luna C18 (250*70 mm, 10 um); mobile phase: [water (FA) -ACN]; gradient: 20%- 50% B over 20 min). Compound 4 (5.45 g, 19.19 mmol, 40.48% yield, 94% purity) was obtained as a yellow solid.1H NMR: (400 MHz, DMSO-d6) 6 = 7.57 - 7.53 (m, 1 H), 7.52 - 7.41 (m, 2H), 7.35 - 7.28 (m, 1 H), 5.35 (s, 1 H), 3.07 (s, 3H), 2.24 (s, 3H).

[0588] Synthesis of compound 5, (Scheme 33)

[0589] To the mixture of compound 4 (5.45 g, 20.40 mmol, 1 eq) in TFA (20 mL) was added dropwise HNO3 (6.91 g, 71 .28 mmol, 4.94 mL, 65% purity, 3.49 eq) at - 20 °C. The mixture was stirred at 25 °C for 1 h. The reaction mixture was poured into ice water (50 mL), and the pH of mixture was neutralized by adding sodium bicarbonate to about 7. The mixture was filtrate. The filter cake was collected and dried by oil pump to give compound 5 (5.3 g, 16.81 mmol, 82.40% yield, 99% purity) as a white solid. Then the filtrate was extracted with EtOAc (50 mL * 3). The combined organic layers were washed with brine (30 mL * 2), dried over Na2SO4, filtered and concentrated under reduced pressure to give compound 5 (1 g, 2.27 mmol, 11.15% yield, 71 % purity) as a yellow oil.1H NMR: (400 MHz, DMSO-d6) 5 = 7.78 - 7.64 (m, 2H), 7.54 (br t, J = 7.8 Hz, 1 H), 7.44 (br d, J = 7.4 Hz, 1 H), 3.42 (s, 3H), 2.69 (s, 3H).

[0590] Synthesis of compound 7, (Scheme 33)

[0591] To the mixture of compound 5 (100 mg, 320.39 pmol, 1 eq) in dioxane (3 mL) was added compound 6 (48.41 mg, 384.47 pmol, 1 .2 eq) and Na2COs (67.92 mg, 640.78 pmol, 2 eq) and Pd (dppf)Cl2 (11 .72 mg, 16.02 pmol, 0.05 eq) and H2O (0.5 mL). The mixture was degassed and purged with N2 for 3 times, and then the mixture was stirred at 80 °C for 16 h under N2 atmosphere. The mixture was diluted with H2O (20 mL) and the resulting mixture was extracted with Ethyl acetate (30 mL * 3). The combined organic layers were dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by prep- TLC (SiO2, Ethyl acetate: MeOH = 20: 1 , RF = 0.08). Compound 7 (35 mg, 69.26 pmol, 21 .62% yield, 62% purity) was obtained as a brown oil. LCMS: RT =0.379min, m / z =314.1 (M+H)+.

[0592] Synthesis of compound, (Scheme 33)

[0593] To the mixture of compound 7 (35 mg, 11 1 .71 pmol, 1 eq) in EtOH (2 mL) and H2O (2 mL) was added Fe (24.95 mg, 446.84 pmol, 4 eq) and NH4CI (29.88 mg, 558.55 pmol, 5 eq). The mixture was stirred at 80 °C for 2 h. The reaction mixture was filtered, the filtrate was concentrated at reduced pressure to give a residue. The residue was triturated with Ethyl acetate (200 mL) and MeOH (20 mL), the resulting mixture was filtered. The filtrate was concentrated at reduced pressure to give compound 8 (40 mg, crude) as a brown oil.

[0594] Synthesis of compound (34), (Scheme 33)

[0595] To the mixture of compound 8 (40 mg, 141 .18 pmol, 1 eq) in DCM (2 mL) was added Py (33.50 mg, 423.54 pmol, 34.19 pL, 3 eq) and compound 9 (29.46 mg, 141 .18 pmol, 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. 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) to give crude product. The crude was purified by prep-HPLC (column: Waters Xbridge 150*25 mm* 5um; mobile phase: [water (NH4HCO3) -ACN]; gradient: 24%- 54% B over 9 min). Compound (34) (1.28 mg, 2.78 pmol, 1 .97% yield, 99% purity) was obtained as a white solid.1H NMR: (400 MHz, CDCh) 6 = 7.97 - 7.91 (m, 1 H), 7.80 (s, 1 H), 7.71 (s, 1 H), 7.52 - 7.45 (m, 3H), 7.36 - 7.28 (m, 1 H), 7.25 - 7.20 (m, 1 H), 7.16 - 7.05 (m, 2H), 6.19 (s, 1 H), 4.60 (s, 2H), 3.96 (s, 3H), 3.25 (s, 3H), 2.41 (s, 3H);

[0596] LCMS: RT = 0.454 min, m / z = 456.4(M+H)+.

[0597] EXAMPLE 1.33: SYNTHESIS OF (35)

[0598] Scheme 34: Synthesis of (35)

[0599] To a solution of compound 4 (38.46 mg, 184.34 pmol, 1 eq) in DCM (0.5 mL) was added pyridine (43.74 mg, 553.02 pmol, 44.64 pL, 3 eq) and compound 1 (50 mg, 184.34 pmol, 1 eq). The mixture was stirred at 25 °C for 1 h. The mixture was concentrated at reduced pressure to give a residue. The reaction mixture was purified by prep-HPLC (column: Phenomenex luna

[0600] 10 C18 150*25 mm* 10um; mobile phase: [water (FA) -ACN]; gradient: 38%-68% B over 1 min) to give (35) (5.25 mg, 1 1 .84 pmol, 6.42% yield) as an off-white solid.1H NMR: (400 MHz, CDCb) 5 = 7.72 - 7.62 (m, 6H), 7.09 - 7.01 (m, 2H), 5.67 (s, 1 H), 4.47 (s, 2H), 3.20 (s, 3H), 2.39 (s, 3H); LCMS: RT = 0.509 min, m / z = 444.1 (M+H)+.

[0601] EXAMPLE 1.34: SYNTHESIS OF (36)

[0602] -| (36), peak 1 (36), peak 2

[0603] Scheme 35: Synthesis of (36)

[0604] Synthesis of compound 3, (Scheme 35)

[0605] To a solution of compound 1 (100 mg, 312.71 pmol, 1 eq, HCI) in DCM (1 mL) was added pyridine (74.21 mg, 938.12 pmol, 75.72 pL, 3 eq) and compound 2 (68.37 mg, 312.71 pmol, 1 eq) at 0 °C. The mixture was stirred at 25 °C for 2 h. EtOAc (8 mL) and water (10 mL) were added and Layers were separated. The aqueous phase was extracted with EtOAc (8 mL x 2). Combined extracts were washed with brine (8 mL), dried over Na2SO4, filtered, and concentrated under vacuum 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 = 0: 1 , RF = 0.24). Compound 3 (57 mg, 122.44 pmol, 39.16% yield) was obtained as a yellow gum. LCMS: RT =0.459 min, m / z =466.2 (M+H)+.

[0606] Synthesis of compound (36)-peak1 , (36)-peak2, (Scheme 35)

[0607] To a solution of compound 3 (57 mg, 122.44 pmol, 1 eq) in DMF (1 mL) was added NaH (11.02 mg, 183.67 pmol, 60% purity, 1.5 eq) at 0 °C under N2 atmosphere. The mixture was stirred at 25 °C for 0.5 h. Then to the mixture was added iodomethane (34.76 mg, 244.89 pmol, 15.25 pL, 2 eq) at 0 °C. The mixture was stirred at 25 °C for 15.5 h. The mixture was quenched with aqueous NH4CI solution (5 mL) and the resulting mixture was extracted with EtOAc (5 mL x 3), the combined organic phase was 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: 35%- 65% B over 10 min). The eluent was concentrated and then freeze dried to give (36). Then (36) was purified by SFC (RT=1.075 min & 1.307 min; column: DAICEL CHIRALCEL OJ (250 mm*30 mm, 10um); mobile phase: [CO2-MeOH (0.1 %NH3H2O)]; B%: 20%, isocratic elution mode). The eluent was concentrated and then freeze dried. (36)-peak1 (12.6 mg, 26.28 pmol, 21 .46% yield, 100% purity) was obtained as an orange solid.1H NMR: (400 MHz, CDCh) 6 = 7.79 (s, 1 H), 7.67 (s, 1 H), 7.51 - 7.42 (m, 3H), 7.20 (td, J = 2.0, 6.8 Hz, 1 H), 3.96 (s, 3H), 3.45-3.32 (m, 4H), 3.21 (s, 3H), 2.84 - 2.48 (m, 4H)„ 2.45-2.35 (m, 4H), 1.58 (br s, 3H); LCMS: RT =0.486 min, m / z =480.2 (M+H)+; SFC: RT =1 .072 min.

[0608] (36)-peak2 (12.22 mg, 25.48 pmol, 20.81 % yield, 100% purity) was obtained as a brown gum. 1 H NMR: (400 MHz, CDCh) 6 = 7.79 (s, 1 H), 7.67 (s, 1 H), 7.51 - 7.42 (m, 3H), 7.23 - 7.15 (m, 1 H), 3.96 (s, 3H), 3.45-3.32 (m, 4H), 3.21 (s, 3H), 2.84 - 2.48 (m, 4H), 2.46 - 2.28 (m, 4H), 1 .59 (br s, 3H); LCMS: RT =0.483 min, m / z =480.1 (M+H)+; SFC: RT =1 .294 min.

[0609] EXAMPLE 1.35: SYNTHESIS OF (37)

[0610] Scheme 36: Synthesis of (37)

[0611] Synthesis of compound 2, (Scheme 36)

[0612] A mixture of compound 1 (100 mg, 401 .25 pmol, 1 eq), 2-iodopropane (102.31 mg, 601 .87 pmol, 60.08 pL, 1 .5 eq) and K2CO3 (110.91 mg, 802.50 pmol, 2 eq) in ACN (2 mL) was stirred at 80 °C for 16 h. The mixture was filtered and the filtrate was concentrated under reduced pressure to give a residue. Compound 2 (120 mg, crude) was obtained as a black solid. LCMS: RT = 0.438 min, m / z = 292.2 (M+H)+.

[0613] Synthesis of compound 3, (Scheme 36)

[0614] A mixture of compound 2 (120 mg, 411 .94 pmol, 1 eq), Fe (115.03 mg, 2.06 mmol, 5 eq) and NH4CI (1 10.18 mg, 2.06 mmol, 5 eq) in EtOH (2 mL) and H2O (2 mL) was stirred at 80 °C for 2 h. The mixture was filtered and the filtrate was concentrated under reduced pressure to give a residue. To the residue was added water (5 mL) and the 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. Compound 3 (50 mg, 191.34 pmol, 46.45% yield) was obtained as a brown solid. LCMS: RT = 0.380 min, m / z = 262.2 (M+H)+.

[0615] Synthesis of compound (37), (Scheme 36)

[0616] A mixture of compound 3 (50 mg, 191.34 pmol, 1 eq), (2 -fluorophenyl) methanesulfonyl chloride (47.90 mg, 229.60 pmol, 1.2 eq) and Py (60.54 mg, 765.35 pmol, 61.77 pL, 4 eq) in DCM (1 mL) was stirred at 20 °C for 16 h. To the mixture was added water (5 mL) and the 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-HPLC (column: Waters Xbridge 150*25 mm* 5um; mobile phase: [water (NH4HCO3) -ACN]; gradient: 35%- 65% B over 9 min), the eluent was concentrated and then freeze dried. Compound (37) (19.12 mg, 43.22 pmol, 22.59% yield, 98% purity) was obtained as a white solid.1H NMR: (400 MHz, CDCh) 6 = 7.92 (br t, J = 7.1 Hz, 1 H), 7.42 - 7.29 (m, 2H), 7.17 - 7.06 (m, 2H), 6.96 - 6.88 (m, 3H), 6.19 - 6.08 (m, 1 H), 4.64 - 4.56 (m, 3H), 3.23 (s, 3H), 2.38 (s, 3H), 1 .37 (d, J = 5.9 Hz, 6H); LCMS: RT = 0.528 min, m / z = 434.2 (M+H)+.

[0617] EXAMPLE 1.36: SYNTHESIS OF (39)

[0618] Scheme 37: Synthesis of (39)

[0619] Synthesis of compound 3, (Scheme 37)

[0620] A mixture of compound 1 (15 g, 67.1 1 mmol, 1 eq) and compound 2 (10.48 g, 80.54 mmol, 10.20 mL, 1.2 eq) in AcOH (100 mL) was stirred at 100 °C for 1 h. The reaction mixture was concentrated in vacuum to give a residue. The residue was triturated with ethyl acetate (300 mL) at 25 °C for 30 min. The mixture was filtered, the filter cake was washed with ethyl acetate (50 mL), dried under reduced pressure to give compound 3 (14 g, 50.89 mmol, 75.83% yield, 92% purity) as light yellow solid. LCMS: RT = 0.505 min, m / z = 252.9 (M+H)+.

[0621] Synthesis of compound 4, (Scheme 37) Compound 3 (14 g, 55.32 mmol, 1 eq) and Mel (23.55 g, 165.95 mmol, 10.33 mL, 3 eq) in NMP (180 mL) were pumped (10 mL / min) to flow reactor (FLRi, SS, Coils reactor, 3.175(1 / 8”) mm, 60.009 mL, 180 °C). The cooling coils was SS, Coils reactor, 3.175(1 / 8") mm, 5.013 mL, 25 °C. The residence time of flow reactor was 6 min. The backpressure regulator was adjusted to 6 MPa and stop collecting the reaction mixture after 30 minutes. The reaction mixture was diluted with H2O (300 mL) and the resulting mixture was extracted with ethyl acetate (100 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 prep-HPLC (column: Phenomenex luna C18 (250*70 mm, 10 um); mobile phase: [H2O (0.1 %TFA) -ACN]; gradient: 20%- 50% B over 22.0 min) followed by lyophilization to give compound 4 (10.4 g, 38.93 mmol, 70.38% yield) as green solid.1H NMR: (400 MHz, CDCb) 6 = 7.53 (s, 1 H), 7.51 - 7.47 (m, 1 H), 7.36 (d, J = 5.3 Hz, 2H), 5.58 (s, 1 H), 3.16 (s, 3H), 2.28 (s, 3H).

[0622] Synthesis of compound 5, (Scheme 37)

[0623] To a solution of compound 4 (10.4 g, 38.93 mmol, 1 eq) in TFA (100 mL) was added HNO3 (5.490 g, 56.63 mmol, 3.92 mL, 65% purity, 1 .45 eq) at - 20 °C. The reaction mixture was stirred at 0 °C for 1 h. The reaction mixture was added dropwise into H2O (150 mL) slowly. Lots of solid precipitated and then the resulting mixture was filtered, the filter cake was washed with ethyl acetate (10 mL) and concentrated under reduced pressure to give a compound 5 (7.3 g, 23.39 mmol, 60.07% yield) as light yellow solid. LCMS: RT = 0.394 min, m / z = 311 .9 (M+H)+.

[0624] Synthesis of compound 6, (Scheme 37)

[0625] To a solution of compound 5 (1 g, 3.20 mmol, 1 eq) in MeOH (15 mL) was added TEA (972.60 mg, 9.61 mmol, 1.34 mL, 3 eq) and Pd(dppf)Cl2 (468.86 mg, 640.78 pmol, 0.2 eq), the suspension was degassed under vacuum and purged with CO (50 psi) for 3 times. The reaction mixture stirred under CO (50 psi) at 80 °C for 16 h. The reaction mixture was concentrated under vacuum 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 = 0: 1)) to give compound 6 (270 mg, 1.03 mmol, 32.25% yield) as brown oil.1H NMR: (400 MHz, DMSO-de) 5 = 7.98 (t, J = 1 .8 Hz, 1 H), 7.81 (td, J = 1 .3, 7.6 Hz, 1 H), 7.73 - 7.68 (m, 1 H), 7.64 - 7.59 (m, 1 H), 3.89 (br s, 2H), 3.88 (s, 3H), 2.76 (s, 3H), 2.1 1 (s, 3H).

[0626] Synthesis of compound 8, (Scheme 37)

[0627] To a solution of compound 6 (270 mg, 1.03 mmol, 1 eq) and pyridine (326.96 mg, 4.13 mmol, 333.64 pL, 4 eq) in DCM (5 mL) was added compound 8 (211 .45 mg, 1 .03 mmol, 1 eq) 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), dried with anhydrous Na2SO4, filtered and the filtrate was concentrated in vacuum to give compound 8 (330 mg, 768.45 pmol, 74.36% yield) as red solid. LCMS: RT = 0.471 min, m / z = 430.1 (M+H)+. Synthesis of compound 9, (Scheme 37)

[0628] To a solution of compound 8 (330 mg, 768.45 pmol, 1 eq) in THF (3 mL) and H2O (1 mL) was added UOH H2O (64.49 mg, 1.54 mmol, 2 eq). The reaction mixture was stirred at 25 °C for 2 h. The reaction mixture was diluted with H2O (10 mL) and concentrated in vacuum to remove THF. The aqueous solution 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 phase was dried with anhydrous Na2SO4, filtered and the filtrate was concentrated in vacuum to give compound 9 (310 mg, 746.25 pmol, 97.11 % yield) as brown solid. LCMS: RT = 0.421 min, m / z = 416.1 (M+H)+.

[0629] Synthesis of compound (39), (Scheme 37)

[0630] To a solution of compound 9 (50 mg, 120.36 pmol, 1 eq) and compound 10 (24.03 mg, 120.36 pmol, 1 eq) in DMF (1 mL) was added HOBt (19.52 mg, 144.44 pmol, 1.2 eq), EDCI (27.69 mg, 144.44 pmol, 1 .2 eq) and DIEA (62.22 mg, 481.45 pmol, 83.86 pL, 4 eq). The reaction mixture was stirred at 25 °C for 2 h. The reaction mixture was diluted with ethyl acetate (10 mL) and the resulting mixture was washed with H2O (5 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: [H2O (0.225% FA) -ACN]; gradient: 44%-74% B over 15.0 min) followed by lyophilization to give (39) (13.33 mg, 22.33 pmol, 18.55% yield, 100% purity) as white solid.1H NMR: (400 MHz, DMSO-de) 6 = 10.29 - 10.05 (m, 1 H), 9.78 - 9.34 (m, 1 H), 8.97 - 8.78 (m, 1 H), 7.98 (d, J = 7.5 Hz, 1 H), 7.92 (s, 1 H), 7.71 - 7.64 (m, 1 H), 7.61 - 7.56 (m, 1 H), 7.43 (d, J = 1 .8 Hz, 1 H), 7.25 (d, J = 2.3 Hz, 1 H), 3.36 (br s, 2H), 3.16 (s, 3H), 2.77 - 2.64 (m, 3H), 2.61 - 2.52 (m, 2H), 2.27 (s, 3H), 1 .26 (s, 9H). LCMS: RT = 0.590 min, m / z = 597.2 (M+H)+.

[0631] EXAMPLE 1.37: SYNTHESIS OF (40)

[0632] Scheme 38: Synthesis of (40)

[0633] To a solution of compound 1 (50 mg, 120.36 pmol, 1 eq) and compound 2 (13.45 mg, 144.44 pmol, 13.16 pL, 1.2 eq) in pyridine (1 mL) was added EDCI (27.69 mg, 144.44 pmol, 1 .2 eq). The reaction mixture was stirred at 25 °C for2 h. The reaction mixture 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 urn; mobile phase: [H2O (0.225% FA) -ACN]; gradient: 32%- 62% B over 15.0 min) followed by lyophilization to give (40) (30.92 mg, 63.03 pmol, 52.37% yield, 100% purity) as white solid.1H NMR: (400 MHz, CDCb) 6 = 8.80 (br s, 1 H), 8.38 (br d, J = 3.1 Hz, 1 H), 8.15 - 8.08 (m, 2H), 7.82 (br d, J = 7.8 Hz, 2H), 7.67 (br t, J = 7.6 Hz, 1 H), 7.45 - 7.37 (m, 3H), 7.20 - 7.14 (m, 1 H), 3.29 (s, 3H), 3.25 (br d, J = 6.7 Hz, 2H), 2.80 - 2.64 (m, 3H), 2.43 (s, 3H), 2.40 - 2.29 (m, 2H). LCMS: RT = 0.498 min, m / z = 491 .1 (M+H)+.

[0634] EXAMPLE 1.38: SYNTHESIS OF (41)

[0635] Scheme 39: Synthesis of (41)

[0636] To a mixture of compound 1 (60 mg, 148.24 pmol, 1 eq) and pyridine (117.26 mg, 1.48 mmol, 119.65 pL, 10 eq) in DCM (1 mL) was added compound 2 (36.40 mg, 177.89 pmol, 1 .2 eq) at 0 °C, then the mixture was stirred at 20 °C for 16 h. The reaction mixture 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 (NH4HCO3) -ACN]; gradient: 26%- 56% B over 10 min). (41) (19.53 mg, 41 .09 pmol, 27.72% yield, 95% purity) was obtained as a white solid.1H NMR: (400 MHz, CDCb) 6 = 7.80 (d, J = 10.1 Hz, 2H), 7.54 (s, 1 H), 7.49 (br d, J = 4.5 Hz, 2H), 7.21 - 7.15 (m, 1 H), 7.06 (br s, 1 H), 3.97 (s, 3H), 3.33 (br d, J = 6.4 Hz, 2H), 3.24 (s, 3H), 2.87 - 2.71 (m, 3H), 2.52 - 2.42 (m, 2H), 2.40 (s, 3H). LCMS: RT = 0.450 min, m / z = 452.1 (M+H)+.

[0637] EXAMPLE 1.39: SYNTHESIS OF (42)

[0638]

[0639] Scheme 40: Synthesis of (42)

[0640] Synthesis of compound 2, (Scheme 42)

[0641] A mixture of compound 1 (300 mg, 908.79 pmol, 1 eq), Fe (253.76 mg, 4.54 mmol, 5 eq), NH4CI (486.12 mg, 9.09 mmol, 10 eq) in MeOH (8 mL) and H2O (2 mL) was degassed and purged with N2 for 3 times, and then the mixture was stirred at 80 °C for 2 h under N2 atmosphere. The mixture was cooled to room temperature and concentrated under reduced pressure to give a residue. The residue was diluted with water (10 mL) and stirred for 5 min. The aqueous phase was extracted with EtOAc (10 mL x 2). The combined organic phase was washed with brine (10 mL), dried with anhydrous Na2SO4, filtered and concentrated in vacuum to give compound 2 (20 mg, crude) as white solid. LCMS: RT = 0.386 min, m / z = 302.1 (M+H)+.

[0642] Synthesis of compound 4, (Scheme 42)

[0643] A mixture of compound 2 (120 mg, 399.83 pmol, 1 eq), compound 3 (98.18 mg, 479.80 pmol, 1 .2 eq), pyridine (94.88 mg, 1.20 mmol, 96.82 pL, 3 eq) in DCM (1 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 cooled to room temperature and concentrated under reduced pressure to give a residue. The residue was diluted with water (10 mL) and stirred for 5 min. The aqueous phase was extracted with EtOAc (10 mL x 2). The combined organic phase was washed with brine (10 mL), dried with anhydrous Na2SO4, filtered and concentrated in vacuum to give a residue. The residue was purified by reversed-phase HPLC (0.1 % NH3 H2O condition), the eluent was concentrated and then freeze dried to give compound 4 (50 mg, crude) as white solid. LCMS: RT = 0.498 min, m / z = 468.1 (M+H)+.

[0644] Synthesis of compound (42), (Scheme 42)

[0645] A mixture of compound 4 (45 mg, 96.09 pmol, 1 eq), compound 5 (18.15 mg, 144.14 pmol, 1.5 eq), CsF (29.19 mg, 192.19 pmol, 7.09 pL, 2 eq), Pd(dppf)Cl2 (7.03 mg, 9.61 pmol, 0.1 eq) in dioxane (1 mL) was degassed and purged with N2 for 3 times, and then the mixture was stirred at 80 °C for 16 h under N2 atmosphere. The mixture was cooled to room temperature and concentrated under reduced pressure to give a residue. The residue was diluted with water (10 mL) and stirred for 5 min. The aqueous phase was extracted with EtOAc (5 mL x 2). The combined organic phase was washed with brine (10 mL), dried with anhydrous Na2SO4, filtered and 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: 30%- 660% B over 10 min) to give (42) (10.24 mg, 21.16 pmol, 22.02% yield, 97% purity) as white solid.1H NMR: (400 MHz, CDCI3) 6 = 7.95 - 7.81 (m, 2H), 7.63 - 7.47 (m, 1 H), 7.26 - 7.09 (m, 2H), 6.56 - 6.18 (m, 1 H), 4.01 - 3.95 (m, 3H), 3.37 (br s, 2H), 3.24 (br s, 3H), 2.91 - 2.71 (m,

[0646] 3H), 2.55 - 2.43 (m, 2H), 2.40 (br s, 3H). LCMS: RT = 0.464 min, m / z = 470.2 (M+H)+.

[0647] Synthesis of SC-003863 (E)

[0648] 1a 3a 4a 5a

[0649] Synthetic Scheme 1: Synthesis of Intermediate 5a

[0650] SC-003863

[0651] Synthetic Scheme 2: Synthesis of SC-003863 Experimental Procedure: Synthesis of compound 3a

[0652] 1a 3a

[0653] A mixture of compound la (500 mg, 4.09 mmol, 1 eq), 4-methylbenzenesulfonyl chloride (936.74 mg, 4.91 mmol, 1.2 eq), DMAP (50.02 mg, 409.46 pmol, 0.1 eq), DIEA (1.59 g, 12.28 mmol, 2.14 mL, 3 eq) in DCM (5 mL) was degassed and purged with N? for 3 times, and then the mixture was stirred at 20 °C for 16 h under N? atmosphere. EtOAc (20 mL) and water (20 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 NajSCU, filtered, and concentrated under vacuum to give a residue. The residue was purified by column chromatography (silica gel, Petroleum ether / Ethyl acetate = 1: 0 to 3: 1, TLC: Petroleum ether: Ethyl acetate = 5: 1, Rf = 0.46) to give compound 3 (600 mg, crude) as white solid.

[0654] 2H NMR: (400 MHz, CDCI3) 6 = 7.80 (d, J = 8.3 Hz, 2H), 7.37 (d, J = 8.0 Hz, 2H), 4.07 (d, J =

[0655] 6.6 Hz, 2H), 2.70 - 2.58 (m, 2H), 2.47 (s, 3H), 2.36 - 2.23 (m, 2H).

[0656] Synthesis of compound 4a , ,

[0657] 3a 4a

[0658] A mixture of compound 3a (600 mg, 2.17 mmol, 1 eq), potassium; ethanethioate (496.02 mg, 4.34 mmol, 2 eq) in DMF (5 mL) was degassed and purged with N? for 3 times, and then the mixture was stirred at 60 °C for 16 h under N? atmosphere. The reaction mixture was cooled to room temperature. EtOAc (20 mL) and water (15 mL) were added and layers were separated. The aqueous phase was extracted with EtOAc (5 mL x 2). Combined extracts were washed with brine (10 mL), dried over NajSCU, filtered, and concentrated under vacuum to give a residue. The residue was purified by column chromatography (silica gel, Petroleum ether / Ethyl acetate = 1: 0 to 3: 1, TLC: Petroleum ether: Ethyl acetate = 5: 1, Rf = 0.43) to give compound 4a (400 mg, crude) as white solid.1H NMR: (400 MHz, CDCI3) 6 = 3.04 (d, J = 7.4 Hz, 2H), 2.74 - 2.61 (m, 2H), 2.43 - 2.36 (m, 1H), 2.35 (s, 3H), 2.31 - 2.17 (m, 2H).

[0659] Synthesis of compound 5a

[0660] 4a 5a

[0661] To a mixture of NCS (592.77 mg, 4.44 mmol, 4 eq), HCI (2 M, 1.11 mL, 2 eq) in ACN (3 mL) was added compound 4a (200 mg, 1.11 mmol, 1 eq) at 0 °C. Then the mixture stirred at 0 °C for 2 h. 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: 0 to 3: 1, TLC: Petroleum ether: Ethyl acetate = 5: 1, Rf = 0.47) to give compound 5a (200 mg, crude) as white solid. NMR: (400 MHz, CDCI3) 6 = 3.87 (d, J = 6.9 Hz, 2H), 3.04 - 2.87 (m, 3H), 2.63 - 2.48 (m, 2H).

[0662] Synthesis of compound 3

[0663] To the mixture of compound 1 (25 g, 111.86 mmol, 1 eq, HCI) in AcOH (60 mL) was added compound 2 (14.56 g, 111.86 mmol, 14.16 mL, 1 eq). The mixture was stirred at 100 °C for 1 h. The reaction mixture diluted with EtOAc (60 mL), the resulting mixture was stirred at 25 °C for 2 h. The mixture was filtered, the filter cake was collected and dried under reduced pressure to give compound 3 (22 g, crude) as a white solid, the filtrate was discarded.

[0664] Synthesis of compound 4

[0665] 3 4

[0666] To the mixture of compound 3 (1 g, 3.95 mmol, 1 eq) in ACN (120 mL) was added Mel (3.36 g, 23.71 mmol, 1.48 mL, 6 eq). The mixture was stirred at 120 °C for4 h under microwave. Twelve batches of the reactions were combined together. The mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (column: Phenomenex luna C18 (250*70 mm, 10 um); mobile phase: [water (FA) -ACN]; gradient: 20%- 50% B over 20 min). Compound 4 (5.45 g, 19.19 mmol, 40.48% yield, 94% purity) was obtained as a yellow solid. NMR: (400 MHz, DMSO-d6) 6 = 7.57 - 7.53 (m, 1H), 7.52 - 7.41 (m, 2H), 7.35 - 7.28 (m, 1H), 5.35 (s, 1H), 3.07 (s, 3H), 2.24 (s, 3H).

[0667] Synthesis of compound 5

[0668] To the mixture of compound 4 (5.45 g, 20.40 mmol, 1 eq) in TFA (20 mL) was added dropwise HNO3 (6.91 g, 71.28 mmol, 4.94 mL, 65% purity, 3.49 eq) at - 20 °C. The mixture was stirred at 25 °C for 1 h. The reaction mixture was poured into ice water (50 mL), and the pH of mixture was neutralized by adding sodium bicarbonate to about 7. The mixture was filtrate. The filter cake was collected and dried by oil pump to give compound 5 (5.3 g, 16.81 mmol, 82.40% yield, 99% purity) as a white solid. Then the filtrate was extracted with EtOAc (50 mL * 3). The combined organic layers were washed with brine (30 mL * 2), dried over NajSC , filtered and concentrated under reduced pressure to give compound 5 (1 g, 1. 1 mmol, 11.15% yield, 71% purity) as a yellow oil.2H NMR: (400 MHz, DMSO-d6) 6 = 7.78 - 7.64 (m, 2H), 7.54 (br t, J = 7.8 Hz, 1H), 7.44 (br d, J = 7.4 Hz, 1H), 3.42 (s, 3H), 2.69 (s, 3H).

[0669] Synthesis of compound 7

[0670] 5 7

[0671] To the mixture of compound 5 (100 mg, 320.39 pmol, 1 eq) in dioxane (3 mL) was added compound 6 (48.41 mg, 384.47 pmol, 1.2 eq) and NajCOs (67.92 mg, 640.78 pmol, 2 eq) and Pd (dppf)CL (11.72 mg, 16.02 pmol, 0.05 eq) and H?O (0.5 mL). The mixture was degassed and purged with N2 for 3 times, and then the mixture was stirred at 80 °C for 16 h under N2 atmosphere. The mixture was diluted with H2O (20 mL) and the resulting mixture was extracted with Ethyl acetate (30 mL * 3). The combined organic layers were dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by prep-TLC (SiC>2, Ethyl acetate: MeOH = 20: 1, RF = 0.08). Compound 7 (35 mg, 69.26 pmol, 21.62% yield, 62% purity) was obtained as a brown oil. LCMS: RT =0.379min, m / z =314.1(M+H)+.

[0672] Synthesis of compound 8

[0673] To the mixture of compound 7 (35 mg, 111.71 pmol, 1 eq) in EtOH (2 mL) and H2O (2 mL) was added Fe (24.95 mg, 446.84 pmol, 4 eq) and NH4CI (29.88 mg, 558.55 pmol, 5 eq). The mixture was stirred at 80 °C for 2 h. The reaction mixture was filtered, the filtrate was concentrated at reduced pressure to give a residue. The residue was triturated with Ethyl acetate (200 mL) and MeOH (20 mL), the resulting mixture was filtered. The filtrate was concentrated at reduced pressure to give compound 8 (40 mg, crude) as a brown oil.

[0674] Synthesis of SC-003863

[0675] To a mixture of compound 8 (60 mg, 148.24 pmol, 1 eq) and pyridine (117.26 mg, 1.48 mmol, 119.65 pL, 10 eq) in DCM (1 mL) was added compound 5a (36.40 mg, 177.89 pmol, 1.2 eq) at 0 °C, then the mixture was stirred at 20 °C for 16 h. The reaction mixture 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 (NH4HCO3) - ACN]; gradient: 26%- 56% B over 10 min). SC-003863 (19.53 mg, 41.09 tmol, 27.72% yield, 95% purity) was obtained as a white solid.1H NMR: (400 MHz, CDCU) 6 = 7.80 (d, J = 10.1 Hz, 2H), 7.54 (s, 1H), 7.49 (br d, J = 4.5 Hz, 2H), 7.21 - 7.15 (m, 1H), 7.06 (br s, 1H), 3.97 (s, 3H), 3.33 (br d, J = 6.4 Hz, 2H), 3.24 (s, 3H), 2.87 - 2.71 (m, 3H), 2.52 - 2.42 (m, 2H), 2.40 (s, 3H); LCMS: RT = 0.450 min, m / z = 452.1 (M+H)+.

[0676] Example 2: Inhibition of PLA2G15

[0677] 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 OOrpm 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.

[0678] Example 3

[0679] A clear 96-well high content imaging plate was coated with 0.1% gelatin for 1 h at 37°C.

[0680] 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 SC-003863 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.

[0681] 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.

[0682] Example 4

[0683] 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.

[0684] We identified PLA2G 15 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.

[0685] NEUROLOGICAL COMPOSITE SCORE

[0686] 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.

[0687] 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.

[0688] AST AND ALT PLASMA CONCENTRATION

[0689] 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. 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.

[0690] Example 5: HMC3 PFO cholesterol accumulation assay

[0691] 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), N PC 1 -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.

[0692] 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.

[0693] 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.

[0694] 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. 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.

[0695] 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.

[0696] Example 6: Evaluation ofPLA2G15 Inhibitors in Restoring BMP Levels in Models of Batten Disease and GRN-Mediated Conditions

[0697] 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.

[0698] 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.

[0699] 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. Treatment with SC4395 or SC3863 restored BMP levels in CLN3ko cells, with SC4395 also partly restoring BMP levels in CLN5ko cells.

[0700] 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 represented by formula (I), or a salt or solvate thereof:wherein L is -NH-, a -N(CI- alkyl)-, a -C1-4 alkylene-NH- wherein N is bound to S, or a C1-4 alkylene; wherein RS2is a 4- to 6-membered ring, which may be substituted by one or more C1-4 alkyls, O-C1-4 alkyls, halogens or pseudohalogens; wherein RP2is a C1-4 alkyl, a O-C1-4 alkyl, or a CO-C1-4 alkyl; or wherein RP2is a 3- to 5-membered heterocyclic or carbocyclic ring, or a benzamide (C6H5CONH2) which may be substituted by one or more C1-4 alkyls, O-C1-4 alkyls, 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 - NO2;wherein RN1, RN2, RP1, RP3, RS1and RS3are independently H, a C1-4 alkyl, a O-C1-4 alkyl, a CO-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 -NO2; and wherein each C1-4 alkyl and C1-4 alkylene 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 -NO2.

2. The PLA2G15 inhibitor according to claim 1 , wherein RS2is an aromatic ring.

3. The PLA2G15 inhibitor according to claim 2, wherein the aromatic ring is a phenyl or a thiazole.

4. The PLA2G15 inhibitor according to claim 1 , wherein RS2is a cyclobutyl, cyclopentyl or cyclohexyl.

5. The PLA2G15 inhibitor according to any one of claims 1 to 4, wherein the ring comprised in RS2is unsubstituted or substituted with one or more CH3 or F.

6. The PLA2G15 inhibitor according to any one of claims 1 to 5, wherein RS1and RS3are independently H or CH3.

7. The PLA2G15 inhibitor according to any one of claims 1 to 6, wherein RN1and RN2are independently a C1-4 alkyl, preferably methyl or ethyl, more preferably wherein RN1and RN2are methyl.

8. The PLA2G15 inhibitor according to any one of claims 1 to 7, wherein L is NH or - N(CH3)-.

9. The PLA2G15 inhibitor according to any one of claims 1 to 8, wherein RP2is a cyclopropyl, a methyl or an O-methyl, preferably wherein the cyclopropyl, methyl or O- methyl is unsubstituted or substituted with one or more fluorines.

10. The PLA2G15 inhibitor according to any one of claims 1 to 8, wherein RP2is a heteroaromatic ring, preferably a pyrrole, a pyrazole or an imidazole, preferably wherein the heteroaromatic ring is unsubstituted or N-methylated.

11. The PLA2G15 inhibitor according to any one of claims 1 to 10, wherein RP1and RP3are independently H, CH3, O-CH3, CH2CF3, CF3, F, or CN.

12. The PLA2G15 inhibitor according to claim 1 , represented by any one of formula (1) to (42):(3) (4)(15) (16)13. Use of the PLA2G15 inhibitor according to any one of claims 1 to 12 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.

14. The PLA2G15 inhibitor according to any one of claims 1 to 12, for use as a medicament.

15. The PLA2G15 inhibitor according to claim 13, 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 thanPLA2G15.

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