Heteroaryl compounds as ligand directed degraders of IRAK4

Abstract

Provided herein are compounds (I) and compositions thereof for modulating IRAK4. In some embodiments, the compounds and compositions are provided for treatment of inflammatory or autoimmune diseases.

Description

HETEROARYL COMPOUNDS AS LIGAND DIRECTED DEGRADERS OF IRAK4CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims priority to US Provisional Application No. 63 / 619,913, filed on January 11, 2024, the disclosure of which is incorporated herein by reference in its entirety for any purpose.FIELD

[0002] The present disclosure relates generally to compounds, compositions, and methods for their preparation and use of the compounds and compositions for treating inflammatory or autoimmune diseases.BACKGROUND

[0003] The recruitment of immune cells to sites of injury involves the concerted interactions of a large number of soluble mediators. Several cytokines appear to play key roles in these processes, including interleukin- 1 (IL-1). IL-1 produces proinflammatory responses and contributes to the tissue degeneration observed in chronic inflammatory conditions. IL-1 has also been implicated in the process of bone resorption and adipose tissue regulation. Thus, IL-1 plays a key role in a large number of pathological conditions including rheumatoid arthritis, inflammatory bowel disease, multiple sclerosis, diabetes, obesity, cancer, and sepsis.

[0004] IL-1 treatment of cells induces the formation of a complex consisting of the two IL-1 receptor chains, IL-1R1 and IL-lRAcP, and the resulting heterodimer recruits an adaptor molecule designated as MyD88, which binds to IL-1 receptor associated kinase (IRAK) (Wesche et al., J. Biol. Chem. 1999, 274, 19403-19410; O’Neill et al., J. Leukoc. Biol. 1998, 63, 650-657; Auron, Cytokine Growth Factor Rev. 1998, 9:221-237; and O’Neill, Biochem. Soc. Trans. 2000, 28, 557 - 563). Four members of the IRAK family have been identified: IRAKI, IRAK2, IRAK3, and IRAK4. These proteins are characterized by a typical N-terminal death domain that mediates interaction with MyD88-family adaptor proteins and a centrally located kinase domain. Of the four members in the mammalian IRAK family, IRAK-4 is considered to be the “master IRAK.” IRAK- 4 is a serine / threonine kinase that plays an essential role in signal transduction by Toll / IL-1 receptors (TIRs). Under overexpression conditions, all IRAKs can mediate the activation of nuclear factor-kappa B and stress-induced mitogen activated protein kinase (MAPK)-signaling cascades. Studies have shown that IRAK4 kinase activity is essential for cytokine production, activation of MAPKs, and induction of NF -kappa B regulated genes in response to TLR ligands (Koziczak-Holbro M. et al., J. Biol. Chem. 2007, 282, 13552-13560). Given the central role of IRAK4 in Toll-like / IL-lR signaling and immunological protection, compounds that modulate the function of IRAK4 may be useful in treating inflammatory, cell proliferative, and immune-relatedconditions and diseases associated with IRAK-mediated signal transduction such as rheumatoid arthritis, inflammatory bowel disease, multiple sclerosis, diabetes, obesity, allergic disease, psoriasis, asthma, graft rejection, cancer and sepsis.

[0005] Protein degradation is a highly regulated and essential process that maintains cellular homeostasis. Selective identification and removal of damaged, misfolded, or excess proteins is achieved through the ubiquitin-proteasome pathway (UPP). The UPP is central to the regulation of almost all cellular processes. Ubiquitination of the protein is accomplished by an E3 ubiquitin ligase that binds to a protein and adds ubiquitin molecules to the protein, thus marking the protein for proteasome degradation.

[0006] Harnessing the UPP for therapeutic use has received significant interest (Zhou et al., Mol. Cell 2000, 6, 751-756). One promising therapy uses proteolysis targeting chimeras, commonly referred to as PROTACs, to effect removal of unwanted proteins by protein degradation (Scheepstra et al., Comp. Struct. Biotech. J. 2019, 77, 160-176). PROTACS are ligand directed degraders that bring together an E3 ligase and a target protein that is to be degraded. These bivalent molecules usually consist of an E3 ligase ligand connected through a linker moiety to small molecule that binds to the target protein. A PROTAC positions the E3 ligase at the appropriate distance and orientation to the target protein, allowing the latter to be ubiquitinated. The ubiquitinated target protein is subsequently recognized by the proteasome, where it is degraded.

[0007] Accordingly, in one aspect, provided herein are compounds that target IRAK4 for degradation.SUMMARY

[0008] Described herein, in certain embodiments, are compounds and compositions thereof for degrading IRAK4. In various embodiments, the compounds and compositions thereof may be used in treatment of inflammatory or autoimmune diseases.

[0009] The present embodiments can be understood more fully by reference to the detailed description and examples, which are intended to exemplify non-limiting embodiments.

[0010] Embodiment 1 is a compound of Formula (I):or a pharmaceutically acceptable salt thereof, wherein:Ring B is 5- to 6-membered monocyclic heterocyclylene or 8- to 10-membered fused bicyclic heterocyclylene, wherein the heterocyclylene contains 1-3 nitrogen atoms; each R2is independently Ci-Ce alkyl or halo; n is 0-3;R3is H, Ci-Ce alkyl, or halo;Ringattached to two adjacent carbon atoms indicated with *; andX is O, CH2, or N(H).

[0011] Embodiment 2 is the compound of embodiment 1, or a pharmaceutically acceptable salt thereof, wherein the compound is of Formula (I’):(I’)

[0012] Embodiment 3 is the compound of embodiment 1, or a pharmaceutically acceptable salt thereof, wherein the compound is of Formula (II):

[0013] Embodiment 4 is the compound of embodiment 1, or a pharmaceutically acceptable salt thereof, wherein the compound is of Formula (III):

[0014] Embodiment 5 is the compound of any one of embodiments 1-3, or a pharmaceutically acceptable salt thereof, wherein:R1is H.

[0015] Embodiment 6 is the compound of any one of embodiments 1-3, or a pharmaceutically acceptable salt thereof, wherein:R1is -NH2.

[0016] Embodiment 7 is the compound of any one of embodiments 1-6, or a pharmaceutically acceptable salt thereof, wherein:L1is -C(O)N(H)-.

[0017] Embodiment 8 is the compound of any one of embodiments 1-6, or a pharmaceutically acceptable salt thereof, wherein:

[0018] Embodiment 9 is the compound of any one of embodiments 1-8, or a pharmaceutically acceptable salt thereof, wherein:Ring B is 6-membered monocyclic heterocyclylene or 8-membered fused bicyclic heterocyclylene, wherein the heterocyclylene contains 1-2 nitrogen atoms; and each R2is independently C1-C3 alkyl or halo.

[0019] Embodiment 10 is the compound of any one of embodiments 1-9, or a pharmaceutically acceptable salt thereof, wherein:

[0020] Embodiment 11 is the compound of any one of embodiments 1-10, or a pharmaceutically acceptable salt thereof, wherein:R3is H or C1-C3 alkyl.

[0021] Embodiment 12 is the compound of any one of embodiments 1-11, or a pharmaceutically acceptable salt thereof, wherein:X is O.

[0022] Embodiment 13 is the compound of any one of embodiments 1-11, or a pharmaceutically acceptable salt thereof, wherein:X is CH2or N(H).

[0023] Embodiment 14 is the compound of any one of embodiments 1-13, or a pharmaceutically acceptable salt thereof, wherein:

[0024] Embodiment 15 is the compound of any one of embodiments 1-14, or a pharmaceutically acceptable salt thereof, wherein the compound is of Formula (IIA), (IIB), (IIC), (HD), (HE), (HF), (IIIA), (IHB), (IIIC), (IHD), (IIIE), or (IHF):

[0025] Embodiment 16 is the compound of embodiment 15, or a pharmaceutically acceptablesalt thereof, wherein:X is O.

[0026] Embodiment 17 is a compound selected from the compounds of Table 1 and pharmaceutically acceptable salts thereof.

[0027] Embodiment 18 is a pharmaceutical composition comprising the compound of any one of embodiments 1-17, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable excipient.

[0028] Embodiment 19 is a method of modulating interleukin- 1 (IL1) receptor-associated kinase 4 (IRAK4) activity comprising contacting IRAK4 with an effective amount of the compound of any one of embodiments 1-17, or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition of embodiment 18.

[0029] Embodiment 20 is a method of treating an inflammatory or autoimmune disease in a subject in need thereof, comprising administering to the subject an effective amount of the compound of any one of embodiments 1-17, or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition of embodiment 18, optionally wherein the inflammatory or autoimmune disease is atopic dermatitis, asthma, lupus, rheumatoid arthritis, familial mediterranean fever, psoriasis, generalized pustular psoriasis, cryoprin-associated periodic syndrome, hidradenitis suppurativa, Bechet’s syndrome, or familial cold autoinflammatory syndrome.DETAILED DESCRIPTIONDefinitions

[0030] As used herein, the terms “comprising” and “including” can be used interchangeably. The terms “comprising” and “including” are to be interpreted as specifying the presence of the stated features or components as referred to, but does not preclude the presence or addition of one or more features, or components, or groups thereof. Additionally, the terms “comprising” and “including” are intended to include examples encompassed by the term “consisting of’. Consequently, the term “consisting of’ can be used in place of the terms “comprising” and “including” to provide for more specific embodiments of the invention.

[0031] The term “consisting of’ means that a subject-matter has at least 90%, 95%, 97%, 98% or 99% of the stated features or components of which it consists. In another embodiment the term “consisting of’ excludes from the scope of any succeeding recitation any other features or components, excepting those that are not essential to the technical effect to be achieved.

[0032] As used herein, the term “or” is to be interpreted as an inclusive “or” meaning any one or any combination. Therefore, “A, B or C” means any of the following: “A; B; C; A and B; A andC; B and C; A, B and C”. An exception to this definition will occur only when a combination of elements, functions, steps or acts are in some way inherently mutually exclusive.

[0033] In the present description, any concentration range, percentage range, ratio range, or integer range is to be understood to include the value of any integer within the recited range and, when appropriate, fractions thereof (such as one tenth and one hundredth of an integer), unless otherwise indicated. Also, any number range recited herein relating to any physical feature, such as polymer subunits, size, or thickness, are to be understood to include any integer within the recited range, unless otherwise indicated. As used herein, the terms “about” and “approximately” mean ± 20%, ± 10%, ± 5%, or ± 1% of the indicated range, value, or structure, unless otherwise indicated.

[0034] An “alkyl” group is a saturated, partially saturated, or unsaturated straight chain or branched non-cyclic hydrocarbon having from 1 to 10 carbon atoms (Ci-Cio alkyl), typically from 1 to 8 carbons (Ci-Cs alkyl) or, in some embodiments, from 1 to 6 (Ci-Ce alkyl), 1 to 3 (C1-C3 alkyl), or 2 to 6 (C2-C6 alkyl) carbon atoms. In some embodiments, the alkyl group is a saturated alkyl group. Representative saturated alkyl groups include -methyl, -ethyl, -n-propyl, -n-butyl, -n-pentyl and -n-hexyl; while saturated branched alkyls include -isopropyl, -sec-butyl, -isobutyl, tert-butyl, -isopentyl, -neopentyl, tertpentyl, -2-methylpentyl, -3 -methylpentyl, -4- methylpentyl, -2,3 -dimethylbutyl and the like. In some embodiments, an alkyl group is an unsaturated alkyl group, also termed an alkenyl or alkynyl group. An “alkenyl” group is an alkyl group that contains one or more carbon-carbon double bonds. An “alkynyl” group is an alkyl group that contains one or more carbon-carbon triple bonds. Examples of unsaturated alkyl groups include, but are not limited to, vinyl, allyl, CH=CH(CH3), -CH=C(CH3)2, -C(CH3)=CH2, -C(CH3)=CH(CH3), -C(CH2CH3)=CH2, -C=CH, -C=C(CH3), -C=C(CH2CH3), -CH2OCH, CH2OQCHs) and CH2C≡CCH2CH3), among others. An alkyl group can be substituted or unsubstituted. When the alkyl groups described herein are said to be “substituted,” they may be substituted with any substituent or substituents as those found in the exemplary compounds and embodiments disclosed herein, as well as halogen; hydroxy; alkoxy; cycloalkyloxy, aryloxy, heterocyclyloxy, heteroaryloxy, heterocycloalkyloxy, cycloalkylalkyloxy, aralkyloxy, heterocyclylalkyloxy, heteroarylalkyloxy, heterocycloalkylalkyloxy; oxo (=0); amino, alkylamino, cycloalkylamino, arylamino, heterocyclylamino, heteroarylamino, heterocycloalkylamino, cycloalkylalkylamino, aralkylamino, heterocyclylalkylamino, heteroaralkylamino, heterocycloalkylalkylamino; imino; imido; amidino; guanidino; enamino; acylamino; sulfonylamino; urea, nitrourea; oxime; hydroxylamino; alkoxyamino; aralkoxyamino; hydrazino; hydrazido; hydrazono; azido; nitro; thio (-SH), alkylthio; =S; sulfinyl; sulfonyl;aminosulfonyl; phosphonate; phosphinyl; acyl; formyl; carboxy; ester; carbamate; amido; cyano; isocyanato; isothiocyanato; cyanato; thiocyanato; or -B(0H)2. In certain embodiments, when the alkyl groups described herein are said to be “substituted,” they may be substituted with any substituent or substituents as those found in the exemplary compounds and embodiments disclosed herein, as well as halogen (chloro, iodo, bromo, or fluoro); alkyl; hydroxyl; alkoxy; alkoxyalkyl; amino; alkylamino; carboxy; nitro; cyano; thiol; thioether; imine; imide; amidine; guanidine; enamine; aminocarbonyl; acylamino; phosphonate; phosphine; thiocarbonyl; sulfinyl; sulfone; sulfonamide; ketone; aldehyde; ester; urea; urethane; oxime; hydroxyl amine; alkoxyamine; aralkoxyamine; N-oxide; hydrazine; hydrazide; hydrazone; azide; isocyanate; isothiocyanate; cyanate; thiocyanate; B(0H)2, or O(alkyl)aminocarbonyl.

[0035] “Alkyl-OH” refers to an unbranched or branched alkyl group as defined above, wherein one or more hydrogen atoms are replaced by -OH. For example, “Ci-Ce alkyl-OH” refers to a Ci-Ce alkyl which is substituted by one or more -OH groups. An alkyl-OH may contain multiple hydroxy groups that are attached to the same carbon atom or to multiple carbon atoms.

[0036] “Alkyl-CN” refers to an unbranched or branched alkyl group as defined above, wherein one or more hydrogen atoms are replaced by -CN. For example, “Ci-Ce alkyl-CN” refers to a Ci-Ce alkyl which is substituted by one or more -CN groups. An alkyl-CN may contain multiple cyano groups that are attached to the same carbon atom or to multiple carbon atoms.

[0037] An “alkoxy” group is -O-(alkyl), wherein alkyl is defined above.

[0038] A “cycloalkyl” group is a saturated, or partially saturated cyclic alkyl group of from 3 to 10 carbon atoms (C3-C10 cycloalkyl) having a single cyclic ring or multiple condensed or bridged rings that can be optionally substituted. In some embodiments, the cycloalkyl group has 3 to 8 ring carbon atoms (C3-C8 cycloalkyl), whereas in other embodiments the number of ring carbon atoms ranges from 3 to 5 (C3-C5 cycloalkyl), 3 to 6 (C3-C6 cycloalkyl), or 3 to 7 (C3-C7 cycloalkyl). In some embodiments, the cycloalkyl groups are saturated cycloalkyl groups. Such saturated cycloalkyl groups include, by way of example, single ring structures such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, 1 methylcyclopropyl, 2methylcyclopentyl, 2-methylcyclooctyl, and the like, or multiple or bridged ring structures such as l-bicyclo[l.l.l]pentyl, bicyclo[2.1.1]hexyl, bicyclo[2.2.1]heptyl, bicyclo[2.2.2]octyl, adamantyl and the like. In other embodiments, the cycloalkyl groups are unsaturated cycloalkyl groups. Examples of unsaturared cycloalkyl groups include cyclohexenyl, cyclopentenyl, cyclohexadienyl, butadienyl, pentadienyl, hexadienyl, among others. A cycloalkyl group can be substituted or unsubstituted. Such substituted cycloalkyl groups include, by way of example, cyclohexanol and the like.

[0039] An “aryl” group is an aromatic carbocyclic group of from 6 to 14 carbon atoms (Ce-Cu aryl) having a single ring (e.g., phenyl) or multiple condensed rings (e.g., naphthyl or anthryl). In some embodiments, aryl groups contain 6-14 carbons (Ce-Cu aryl), and in others from 6 to 12 (Ce- C12 aryl) or even 6 to 10 carbon atoms (Ce-Cio aryl) in the ring portions of the groups. Particular aryls include phenyl, biphenyl, naphthyl and the like. An aryl group can be substituted or unsubstituted. The phrase “aryl groups” also includes groups containing fused rings, such as fused aromatic-aliphatic ring systems (e.g., indanyl, tetrahydronaphthyl, and the like).

[0040] A “halogen” or “halo” is fluorine, chlorine, bromine or iodine.

[0041] “Haloalkyl” refers to an alkyl radical, as defined above, that is substituted by one or more halo radicals, as defined above, e.g, trifluoromethyl, difluoromethyl, tri chloromethyl, 2,2,2- trifluoroethyl, 1,2-difluoroethyl, 3-bromo-2-fluoropropyl, 1,2-dibromoethyl, and the like. In some embodiments, the haloalkyl group has one to six carbon atoms and is substituted by one or more halo radicals (Ci-Ce haloalkyl), or the haloalkyl group has one to three carbon atoms and is substituted by one or more halo radicals (C1-C3 haloalkyl). The halo radicals may be all the same or the halo radicals may be different. Unless specifically stated otherwise, a haloalkyl group is optionally substituted.

[0042] A “heteroaryl” group is an aromatic ring system having one to four heteroatoms as ring atoms in a heteroaromatic ring system, wherein the remainder of the atoms are carbon atoms. In some embodiments, heteroaryl groups contain 3 to 6 ring atoms, and in others from 6 to 9 or even 6 to 10 atoms in the ring portions of the groups. Suitable heteroatoms include oxygen, sulfur and nitrogen. In certain embodiments, the heteroaryl ring system is monocyclic or bicyclic. Nonlimiting examples include but are not limited to, groups such as pyrrolyl, pyrazolyl, imidazolyl, triazolyl, tetrazolyl, oxazolyl, isoxazolyl, benzisoxazolyl (e.g., benzo[d]isoxazolyl), thiazolyl, pyrolyl, pyridazinyl, pyrimidyl, pyrazinyl, thiophenyl, benzothiophenyl, furanyl, benzofuranyl, indolyl (e.g., indolyl-2-onyl or isoindolin-l-onyl), azaindolyl (pyrrol opyridyl or lHpyrrolo[2,3b]pyridyl), indazolyl, benzimidazolyl (e.g., lHbenzo[d]imidazolyl), imidazopyridyl (e.g., azabenzimidazolyl or lHimidazo[4,5b]pyridyl), pyrazol opyridyl, tri azol opyridyl, benzotri azolyl (e.g., lHbenzo[d][l,2,3]triazolyl), benzoxazolyl (e.g., benzo[d]oxazolyl), benzothiazolyl, benzothiadi azolyl, isoxazolopyridyl, thianaphthal enyl, purinyl, xanthinyl, adeninyl, guaninyl, quinolinyl, isoquinolinyl (e.g., 3,4dihydroisoquinolin-l(2H)-onyl), tetrahydroquinolinyl, quinoxalinyl, and quinazolinyl groups. A heteroaryl group can be substituted or unsubstituted.

[0043] A “heterocyclyl” is a non-aromatic cycloalkyl in which one to four of the ring carbon atoms are independently replaced with a heteroatom selected from O, S and N. In some embodiments, heterocyclyl groups include 3 to 10 ring members, whereas other such groups have 3to 5, 3 to 6, or 3 to 8 ring members. Heterocyclyls can also be bonded to other groups at any ring atom (i.e., at any carbon atom or heteroatom of the heterocyclic ring). A heterocycloalkyl group can be substituted or unsubstituted. Heterocyclyl groups encompass saturated and partially saturated ring systems. Further, the term heterocyclyl is intended to encompass any non-aromatic ring containing at least one heteroatom, which ring may be fused to an aryl or heteroary1ring, regardless of the attachment to the remainder of the molecule. The phrase also includes bridged polycyclic ring systems containing a heteroatom. Representative examples of a heterocyclyl group include, but are not limited to, aziridinyl, azetidinyl, azepanyl, pyrrolidyl, imidazolidinyl (e.g., imidazolidin-4-onyl or imidazolidin-2,4-dionyl), pyrazolidinyl, thiazolidinyl, tetrahydrothiophenyl, tetrahydrofuranyl, piperidyl, piperazinyl (e.g., piperazin-2-onyl), morpholinyl, thiomorpholinyl, tetrahydropyranyl (e.g., tetrahydro-2H-pyranyl), tetrahydrothiopyranyl, oxathianyl, dithianyl, l,4dioxaspiro[4.5]decanyl, homopiperazinyl, quinuclidyl, or tetrahydropyrimidin-2(lH)-one. Representative substituted heterocyclyl groups may be monosubstituted or substituted more than once, such as, but not limited to, pyridyl or morpholinyl groups, which are 2-, 3-, 4-, 5-, or 6substituted, or disubstituted with various substituents such as those listed below.

[0044] When the groups described herein, with the exception of alkyl group, are said to be “substituted,” they may be substituted with any appropriate substituent or substituents. Illustrative examples of substituents are those found in the exemplary compounds and embodiments disclosed herein, as well as halogen (chloro, iodo, bromo, or fluoro); alkyl; hydroxyl; alkoxy; alkoxyalkyl; amino; alkylamino; carboxy; nitro; cyano; thiol; thioether; imine; imide; amidine; guanidine; enamine; aminocarbonyl; acylamino; phosphonate; phosphine; thiocarbonyl; sulfinyl; sulfone; sulfonamide; ketone; aldehyde; ester; urea; urethane; oxime; hydroxyl amine; alkoxyamine; aralkoxyamine; N-oxide; hydrazine; hydrazide; hydrazone; azide; isocyanate; isothiocyanate; cyanate; thiocyanate; oxygen (=0); B(0H)2, O(alkyl)aminocarbonyl; cycloalkyl, which may be monocyclic or fused or non-fused polycyclic (e.g., cyclopropyl, cyclobutyl, cyclopentyl, or cyclohexyl), or a heterocyclyl, which may be monocyclic or fused or non-fused polycyclic (e.g., pyrrolidyl, piperidyl, piperazinyl, morpholinyl, or thiazinyl); monocyclic or fused or nonfused polycyclic aryl or heteroaryl (e.g., phenyl, naphthyl, pyrrolyl, indolyl, furanyl, thiophenyl, imidazolyl, oxazolyl, isoxazolyl, thiazolyl, triazolyl, tetrazolyl, pyrazolyl, pyridyl, quinolinyl, isoquinolinyl, acridinyl, pyrazinyl, pyridazinyl, pyrimidyl, benzimidazolyl, benzothiophenyl, or benzofuranyl) aryloxy; aralkyloxy; heterocyclyloxy; and heterocyclyl alkoxy.

[0045] Certain commonly used alternative chemical names may be used. For example, a divalent group such as a divalent “alkyl” group, a divalent “phenyl” group, a divalent “heteroaryl”group, a divalent “heterocyclyl” group etc., may also be referred to as an “alkylene” group, a “phenylene” group, a “heteroarylene” group, or a “heterocyclylene” group, respectively.

[0046] Embodiments of the disclosure are meant to encompass pharmaceutically acceptable salts, tautomers, isotopologues, and stereoisomers of the compounds provided herein, such as the compounds of Formula (I).

[0047] As used herein, the term “pharmaceutically acceptable salt(s)” refers to a salt prepared from a pharmaceutically acceptable non-toxic acid or base including an inorganic acid and base and an organic acid and base. Suitable pharmaceutically acceptable base addition salts of the compounds of Formula (I) include, but are not limited to metallic salts made from aluminum, calcium, lithium, magnesium, potassium, sodium and zinc or organic salts made from lysine, N,N’- dibenzylethylenediamine, chloroprocaine, choline, diethanolamine, ethylenediamine, meglumine (Nmethyl-glucamine) and procaine. Suitable non-toxic acids include, but are not limited to, inorganic and organic acids such as acetic, alginic, anthranilic, benzenesulfonic, benzoic, camphorsulfonic, citric, ethenesulfonic, formic, fumaric, furoic, galacturonic, gluconic, glucuronic, glutamic, glycolic, hydrobromic, hydrochloric, isethionic, lactic, maleic, malic, mandelic, methanesulfonic, mucic, nitric, pamoic, pantothenic, phenylacetic, phosphoric, propionic, salicylic, stearic, succinic, sulfanilic, sulfuric, tartaric acid, and ptoluenesulfonic acid. Specific non-toxic acids include hydrochloric, hydrobromic, maleic, phosphoric, sulfuric, and methanesulfonic acids. Examples of specific salts thus include hydrochloride, formic, and mesylate salts. Others are well- known in the art, see for example, Remington ’s Pharmaceutical Sciences, 18theds., Mack Publishing, Easton PA (1990) o Remington: The Science and Practice of Pharmacy, 19theds., Mack Publishing, Easton PA (1995).

[0048] As used herein and unless otherwise indicated, the term “stereoisomer” or “stereoisomerically pure” means one stereoisomer of a particular compound that is substantially free of other stereoisomers of that compound. For example, a stereoisomerically pure compound having one chiral center will be substantially free of the opposite enantiomer of the compound. A stereoisomerically pure compound having two chiral centers will be substantially free of other diastereomers of the compound. A typical stereoisomerically pure compound comprises greater than about 80% by weight of one stereoisomer of the compound and less than about 20% by weight of other stereoisomers of the compound, greater than about 90% by weight of one stereoisomer of the compound and less than about 10% by weight of the other stereoisomers of the compound, greater than about 95% by weight of one stereoisomer of the compound and less than about 5% by weight of the other stereoisomers of the compound, or greater than about 97% by weight of one stereoisomer of the compound and less than about 3% by weight of the other stereoisomers of thecompound. The compounds disclosed herein can have chiral centers and can occur as racemates, individual enantiomers or diastereomers, and mixtures thereof. All such isomeric forms are included within the embodiments disclosed herein, including mixtures thereof.

[0049] The use of stereoisomerically pure forms of the compounds disclosed herein, as well as the use of mixtures of those forms, are encompassed by the embodiments disclosed herein. For example, mixtures comprising equal or unequal amounts of the enantiomers of a particular compound may be used in methods and compositions disclosed herein. These isomers may be asymmetrically synthesized or resolved using standard techniques such as chiral columns or chiral resolving agents. See, e.g., Jacques, J., et al., Enantiomers, Racemates and Resolutions (Wileylnterscience, New York, 1981); Wilen, S. H., et al., Tetrahedron 33:2725 (1977); Eliel, E. L., Stereochemistry of Carbon Compounds (McGrawHill, NY, 1962); Wilen, S. EL, Tables of Resolving Agents and Optical Resolutions p. 268 (E.L. Eliel, Ed., Univ, of Notre Dame Press, Notre Dame, IN, 1972); Todd, M., Separation Of Enantiomers : Synthetic Methods (Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim, Germany, 2014); Toda, F., Enantiomer Separation: Fundamentals and Practical Methods (Springer Science & Business Media, 2007); Subramanian, G. Chiral Separation Techniques: A Practical Approach (John Wiley & Sons, 2008); Ahuja, S., Chiral Separation Methods for Pharmaceutical and Biotechnological Products (John Wiley & Sons, 2011).

[0050] It should also be noted the compounds disclosed herein can include E and Z isomers, or a mixture thereof, and cis and trans isomers or a mixture thereof. In certain embodiments, the compounds are isolated as either the E or Z isomer. In other embodiments, the compounds are a mixture of the E and Z isomers.

[0051] Tautomers” refers to isomeric forms of a compound that are in equilibrium with each other. The concentrations of the isomeric forms will depend on the environment the compound is found in and may be different depending upon, for example, whether the compound is a solid or is in an organic or aqueous solution. For example, in aqueous solution, pyrazoles may exhibit the following isomeric forms, which are referred to as tautomers of each other:

[0052] As readily understood by one skilled in the art, a wide variety of functional groups and other structures may exhibit tautomerism and all tautomers of compounds of Formula (I) are within the scope of the present disclosure.

[0053] It should also be noted the compounds disclosed herein can contain unnatural proportions of atomic isotopes at one or more of the atoms. For example, the compounds may beradiolabeled with radioactive isotopes, such as for example tritium (3H), iodine-125 (125I), sulfur35 (35S), or carbon-14 (14C), or may be isotopically enriched, such as with deuterium (2H), carbon-13 (13C), or nitrogen- 15 (15N). As used herein, an “isotopologue” is an isotopically enriched compound. The term “isotopically enriched” refers to an atom having an isotopic composition other than the natural isotopic composition of that atom. “Isotopically enriched” may also refer to a compound containing at least one atom having an isotopic composition other than the natural isotopic composition of that atom. The term “isotopic composition” refers to the amount of each isotope present for a given atom. Radiolabeled and isotopically enriched compounds are useful as therapeutic agents, e.g., cancer therapeutic agents, research reagents, e.g., binding assay reagents, and diagnostic agents, e.g., in vivo imaging agents. All isotopic variations of the compounds as described herein, whether radioactive or not, are intended to be encompassed within the scope of the embodiments provided herein. In some embodiments, there are provided isotopologues of the compounds disclosed herein, for example, the isotopologues are deuterium, carbon-13, and / or nitrogen-15 enriched compounds. As used herein, “deuterated”, means a compound wherein at least one hydrogen (H) has been replaced by deuterium (indicated by D or2H), that is, the compound is enriched in deuterium in at least one position.

[0054] It is understood that, independently of stereoisomerical or isotopic composition, each compound disclosed herein can be provided in the form of any of the pharmaceutically acceptable salts discussed herein. Equally, it is understood that the isotopic composition may vary independently from the stereoisomerical composition of each compound referred to herein.Further, the isotopic composition, while being restricted to those elements present in the respective compound or salt thereof disclosed herein, may otherwise vary independently from the selection of the pharmaceutically acceptable salt of the respective compound.

[0055] It should be noted that if there is a discrepancy between a depicted structure and a name for that structure, the depicted structure is to be accorded more weight.

[0056] “Treating” as used herein, means an alleviation, in whole or in part, of a disorder, disease or condition, or one or more of the symptoms associated with a disorder, disease, or condition, or slowing or halting of further progression or worsening of those symptoms, or alleviating or eradicating the cause(s) of the disorder, disease, or condition itself. In one embodiment, the disorder is a neurodegenerative disease, as described herein, or a symptom thereof.

[0057] “Preventing” as used herein, means a method of delaying and / or precluding the onset, recurrence or spread, in whole or in part, of a disorder, disease or condition; barring a subject from acquiring a disorder, disease, or condition; or reducing a subject’s risk of acquiring a disorder,disease, or condition. In one embodiment, the disorder is a neurodegenerative disease, as described herein, or symptoms thereof.

[0058] The term “effective amount” in connection with a compound disclosed herein means an amount capable of treating or preventing a disorder, disease or condition, or symptoms thereof, disclosed herein.

[0059] The term “subject” or “patient” as used herein include an animal, including, but not limited to, an animal such a cow, monkey, horse, sheep, pig, chicken, turkey, quail, cat, dog, mouse, rat, rabbit or guinea pig, in one embodiment a mammal, in another embodiment a human. In one embodiment, a subject is a human having or at risk for having an IRAK4 mediated disease, or a symptom thereof.

[0060] Although various features of the invention may be described in the context of a single embodiment, the features may also be provided separately or in any suitable combination. Conversely, although the invention may be described herein in the context of separate embodiments for clarity, the invention may also be implemented in a single embodiment.Compounds

[0061] In one aspect, provided herein is a compound of Formula (I):or a pharmaceutically acceptable salt thereof, wherein:Ring B is 5- to 6-membered monocyclic heterocyclylene or 8- to 10-membered fused bicyclicheterocyclylene, wherein the heterocyclylene contains 1-3 nitrogen atoms; each R2is independently Ci-Ce alkyl or halo; n is 0-3;R3is H, Ci-Ce alkyl, or halo;Ringattached to two adjacent carbon atoms indicated with *; andX is O, CH2, or N(H).embodiments,some embodiments, A is Cl

[0063] In some embodiments, R1is H or -NH2. In some embodiments, R1is H. In some embodiments, R1is -NH2.

[0064] In some embodiments, L1is -C(O)N(H)-. In some embodiments, L1is -C(O)N(H)-. In some embodiments,

[0065] In some embodiments, Ring B is 5- to 6-membered monocyclic heterocyclylene or 8- to 10-membered fused bicyclic heterocyclylene, wherein the heterocyclylene contains 1-3 nitrogen atoms. In some embodiments, Ring B is 6-membered monocyclic heterocyclylene or 8-membered fused bicyclic heterocyclylene, wherein the heterocyclylene contains 1-2 nitrogen atoms.

[0066] In some embodiments, Ring B is 5- to 6-membered monocyclic heterocyclylene containing 1-3 nitrogen atoms. In some embodiments, Ring B is 5-membered monocyclic heterocyclylene containing 1-3 nitrogen atoms. In some embodiments, Ring B is 5-membered monocyclic heterocyclylene containing 1-2 nitrogen atoms. In some embodiments, Ring B is 5- membered monocyclic heterocyclylene containing one nitrogen atom. In some embodiments, RingB is 5-membered monocyclic heterocyclylene containing 2 nitrogen atoms. In some embodiments, Ring B is 6-membered monocyclic heterocyclylene containing 1-3 nitrogen atoms. In some embodiments, Ring B is 6-membered monocyclic heterocyclylene containing 1-2 nitrogen atoms. In some embodiments, Ring B is 6-membered monocyclic heterocyclylene containing one nitrogen atom. In some embodiments, Ring B is 6-membered monocyclic heterocyclylene containing 2 nitrogen atoms. In some embodiments, Ring B is pyrrolidinylene, piperidinylene, or piperazinylene.

[0067] In some embodiments, Ring B is 8- to 10-membered fused bicyclic heterocyclylene containing 1-3 nitrogen atoms. In some embodiments, Ring B is 8- to 10-membered fused bicyclic heterocyclylene containing 1-2 nitrogen atoms. In some embodiments, Ring B is 8- to 10- membered fused bicyclic heterocyclylene containing one nitrogen atom. In some embodiments, Ring B is 8- to 10-membered fused bicyclic heterocyclylene containing 2 nitrogen atoms. In some embodiments, Ring B is 8-membered fused bicyclic heterocyclylene containing 1-2 nitrogen atoms. In some embodiments, Ring B is 8-membered fused bicyclic heterocyclylene containing one nitrogen atom. In some embodiments, Ring B is 8-membered fused bicyclic heterocyclylene containing 2 nitrogen atoms. In some embodiments, Ring B is 9-membered fused bicyclic heterocyclylene containing 1-2 nitrogen atoms. In some embodiments, Ring B is 9-membered fused bicyclic heterocyclylene containing one nitrogen atom. In some embodiments, Ring B is 9- membered fused bicyclic heterocyclylene containing 2 nitrogen atoms. In some embodiments, Ring B is 10-membered fused bicyclic heterocyclylene containing 1-2 nitrogen atoms. In some embodiments, Ring B is 10-membered fused bicyclic heterocyclylene containing one nitrogen atom. In some embodiments, Ring B is 10-membered fused bicyclic heterocyclylene containing 2 nitrogen atoms.

[0068] In some embodiments, Ring B is

[0069] In some embodiments, each R2is independently Ci-Ce alkyl or halo. In some embodiments, each R2is independently C1-C3 alkyl or halo. In some embodiments, each R2is independently -CH3or -CH2CH3.

[0070] In some embodiments, R2is Ci-Ce alkyl. In some embodiments, R2is C1-C3 alkyl. In some embodiments, R2is methyl, ethyl, w-propyl, or isopropyl. In some embodiments, R2is methyl. In some embodiments, R2is ethyl. In some embodiments, R2is w-propyl. In some embodiments, R2is isopropyl.

[0071] In some embodiments, R2is halo. In some embodiments, R2is Cl, F, or Br. In someembodiments, R2is Cl. In some embodiments, R2is F. In some embodiments, R2is Br.

[0072] In some embodiments, n is 0-3. In some embodiments, n is 0 or 1. In some embodiments, n is 0. In some embodiments, n is 1. In some embodiments, n is 2. In some embodiments, n is 3.

[0074] In some embodiments, R3is H, Ci-Ce alkyl, or halo. In some embodiments, R3is H or C1-C3 alkyl. In some embodiments, R3is H or -CH3.

[0075] In some embodiments, R3is H.

[0076] In some embodiments, R3is Ci-Ce alkyl. In some embodiments, R3is C1-C3 alkyl. In some embodiments, R3is methyl, ethyl, w-propyl, or isopropyl. In some embodiments, R3is methyl. In some embodiments, R3is ethyl. In some embodiments, R3is w-propyl. In some embodiments, R3is isopropyl.

[0077] In some embodiments, R3is halo. In some embodiments, R3is Cl, F, or Br. In some embodiments, R3is Cl. In some embodiments, R3is F. In some embodiments, R3is Br.

[0078] In some embodiments, Ringattached to two adjacent carbon atoms indicated with *, and X is O, CH2, or N(H). In some embodiments, X is O. In some embodiments, X is CH2or N(H). In some embodiments, X is CH2. In some embodiments, X is N(H).

[0080] In some embodiments, the compound of Formula (I) is a compound of Formula (I’):wherein A, L1, Ring B, Ring C, R2, R3, and n are as described for Formula (I).

[0081] In some embodiments, the compound of Formula (I) is a compound of Formula (II):wherein R1, L1, Ring B, Ring C, R2, R3, and n are as described for Formula (I).

[0082] In some embodiments, the compound of Formula (I) is a compound of Formula (IIA), (IIB), (IIC), (IID), (HE), or (IIF):ĨIIB)wherein R1, R2, R3, X, and n are as described for Formula (I). In some variations, X is O. In some variations, X is CH2. In some variations, X is NH.

[0083] In some embodiments, the compound of Formula (I) is a compound of Formula (IIA-1),(IIB-1), (IIC-1), (IID-1), (HE-1), or (IIF-I):(IIF-1) wherein R1, R2, R3, and n are as described for Formula (I).

[0084] In some embodiments, the compound of Formula (I) is a compound of Formula (III):wherein L1, Ring B, Ring C, R2, R3, and n are as described for Formula (I).

[0085] In some embodiments, the compound of Formula (I) is a compound of Formula (IIIA),(IIIB), (IIIC), (IIID), (IIIE), or (IIIF):wherein R2, R3, X, and n are as described for Formula (I). In some variations, X is O. In some variations, X is CH2. In some variations, X is NH.

[0086] In some embodiments, the compound of Formula (I) is a compound of Formula (IIIA- 1), (IIIB-1), (IIIC-1), (IIID-1), (IIIE-1), or (IIIF-1):LZ(IIIF-1) wherein R2, R3, and n are as described for Formula (I).

[0087] In the descriptions herein, it is understood that every description, variation, embodiment, or aspect of a moiety may be combined with every description, variation, embodiment, or aspect of other moieties the same as if each and every combination of descriptions is specifically and individually listed. For example, every description, variation, embodiment, or aspect provided herein with respect to A of Formula (I) may be combined with every description, variation, embodiment, or aspect of R1, R2, R3, n, L1, Ring B, Ring C, and X the same as if each and every combination were specifically and individually listed. It is also understood that all descriptions, variations, embodiments, or aspects of Formula (I), where applicable, apply equally to other formulae detailed herein, and are equally described, the same as if each and every description, variation, embodiment, or aspect were separately and individually listed for all formulae. For example, all descriptions, variations, embodiments, or aspects of Formula (I), where applicable, apply equally to any of the formulae as detailed herein, such as Formulae (F), (II), (IIA), (IIB), (IIC), (HD), (HE), (HF), (IIA-1), (HB-1), (IIC-1), (IID-1), (HE-1), (HF-1), (III), (IIIA), (IHB),(IIIC), (IIID), (IIIE), (IIIF), (IDA-1), (IIIB-1), (IIIC-1), (IIID-1), (IIIE-1), and (IIIF-1), and are equally described, the same as if each and every description, variation, embodiment, or aspect were separately and individually listed for all formulae.

[0088] In some embodiments, provided is a compound selected from the compounds in Table 1 or a pharmaceutically acceptable salt thereof. Although certain compounds described in the present disclosure, including in Table 1, are presented as specific stereoisomers and / or in a nonstereochemical form, it is understood that any or all stereochemical forms, including any enantiomeric or diastereomeric forms, and any tautomers or other forms of any of the compounds of the present disclosure, including in Table 1, are herein described.Table 1.“orl” and “or2” indicates that the absolute stereochemistry was not determined and the stereochemistry may be as drawn or the opposite stereochemistry drawn.

[0089] It is understood that in the present description, combinations of substituents and / or variables of the depicted formulae are permissible only if such contributions result in stable compounds.

[0090] Furthermore, all compounds of Formula (I) that exist in free base or acid form can be converted to their pharmaceutically acceptable salts by treatment with the appropriate inorganic or organic base or acid by methods known to one skilled in the art. Salts of the compounds of Formula (I) can be converted to their free base or acid form by standard techniques.Methods of Synthesis

[0091] The compounds described herein can be made using conventional organic syntheses and commercially available starting materials, or the methods provided herein. By way of example and not limitation, compounds of Formula (I) can be prepared as outlined in Schemes 1 and 2, as well as in the examples set forth herein. It should be noted that one skilled in the art would know how to modify the procedures set forth in the illustrative schemes and examples to arrive at the desired products.

[0092] Target Binding Moieties (TBMs) can be prepared as outlined in Scheme 1. In one synthesis, copper-catalyzed azide alkyne cycloaddition of a and b affords intermediate c, which can be deprotected using, for example, an acid such as p-toluenesulfonic acid, to form d, and subsequently oxidized (for example, using IBX) to form TBM e. In a second synthesis, intermediates f and g are coupled using, for example, HATU, to provide h, which is then oxidized (for example, using IBX) to give TBM i.Scheme 1.wherein A is as described for Formula (I); and Pg is a protecting group such as tetrahydropyran (THP).

[0093] TBMs A (such as e and i from Scheme 1) can be coupled with Cereblon Binding Moieties (CBMs) B under reductive amination conditions to provide compounds of Formula (I) as shown in Scheme 2.Scheme 2.wherein A, L1, Ring B, R2, R3, Ring C, and n are as described for Formula (I).Methods of Use

[0094] Embodiments of the present disclosure provide a method for modulating IRAK4 in a subject in need thereof, the method comprising administering to the subject an effective amount of a compound of Formula (I). Modulation (e.g., inhibition or activation) of IRAK4 can be assessed and demonstrated by a wide variety of ways known in the art. Kits and commercially available assays can be utilized for determining whether and to what degree IRAK4 has been modulated e.g., inhibited or activated).

[0095] In one aspect, provided herein is a method of modulating IRAK4 comprising contacting IRAK4 with an effective amount of a compound of Formula (I) or any embodiment or variation thereof. In some embodiments, the compound of Formula (I) inhibits IRAK4. In other embodiments, the compound of Formula (I) activates IRAK4. In some embodiments, the compound of Formula (I) is an agonist of IRAK4. In some embodiments, the compound of Formula (I) is an antagonist of IRAK4.

[0096] In some embodiments, provided herein is a method for targeting IRAK4 for degradation comprising contacting IRAK4 with an effective amount of a compound of Formula (I) or any embodiment or variation thereof.

[0097] In some embodiments, a compound of Formula (I) modulates the activity of IRAK4 by about 1%, 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100%. In some embodiments, a compound of Formula (I) modulates the activity of IRAK4 by about 1-100%, 5-100%, 10-100%, 15-100%, 20-100%, 25-100%, 30-100%, 35-100%, 40-100%, 45-100%, 50-100%, 55-100%, 60-100%, 65-100%, 70-100%, 75-100%, 80- 100%, 85-100%, 90-100%, 95-100%, 5-95%, 5-90%, 5-85%, 5-80%, 5-75%, 5-70%, 5-65%, 5- 60%, 5-55%, 5-50%, 5-45%, 5-40%, 5-35%, 5-30%, 5-25%, 5-20%, 5-15%, 5-10%, 10-90%, 20- 80%, 30-70%, or 40-60%.

[0098] Also provided in certain embodiments of the present disclosure is a method for degrading IRAK4 in a subject in need thereof, the method comprising administering to the subject an effective amount of a compound of Formula (I). Degradation of IRAK4 can be assessed and demonstrated by a wide variety of ways known in the art. Kits and commercially available assays, including cell -based assays, can be utilized for determining whether and to what degree IRAK4 has been degraded.

[0099] In one aspect, provided herein is a method of degrading IRAK4 comprising contacting IRAK4 with an effective amount of a compound of Formula (I) or any embodiment or variation thereof. In some embodiments, the compound of Formula (I) partially degrades IRAK3. In some embodiments, the compound of Formula (I) fully degrades IRAK4.

[0100] In some embodiments, a compound of Formula (I) degrades IRAK4 by about 1%, 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100%. In some embodiments, a compound of Formula (I) degrades IRAK4 by about 1- 100%, 5-100%, 10-100%, 15-100%, 20-100%, 25-100%, 30-100%, 35-100%, 40-100%, 45-100%, 50-100%, 55-100%, 60-100%, 65-100%, 70-100%, 75-100%, 80-100%, 85-100%, 90-100%, 95- 100%, 5-95%, 5-90%, 5-85%, 5-80%, 5-75%, 5-70%, 5-65%, 5-60%, 5-55%, 5-50%, 5-45%, 5- 40%, 5-35%, 5-30%, 5-25%, 5-20%, 5-15%, 5-10%, 10-90%, 20-80%, 30-70%, or 40-60%.

[0101] In another aspect, provided herein is a method for treating an inflammatory or autoimmune disease in a subject in need thereof, comprising administering to the subject an effective amount of a compound of Formula (I). In some embodiments, provided herein is a method for treating an inflammatory disease in a subject in need thereof, comprising administering to the subject an effective amount of a compound of Formula (I). In some embodiments, provided herein is a method for treating an autoimmune disease in a subject in need thereof, comprising administering to the subject an effective amount of a compound of Formula (I). In some embodiments, provided herein is a method for preventing an inflammatory or autoimmune disease in a subject in need thereof, comprising administering to the subject an effective amount of a compound of Formula (I). In some embodiments, provided herein is a method for preventing an inflammatory disease in a subject in need thereof, comprising administering to the subject an effective amount of a compound of Formula (I). In some embodiments, provided herein is a method for preventing an autoimmune disease in a subject in need thereof, comprising administering to the subject an effective amount of a compound of Formula (I). Non-limiting examples of an inflammatory or autoimmune disease include atopic dermatitis, asthma, lupus, rheumatoid arthritis, familial mediterranean fever, psoriasis, generalized pustular psoriasis,cryoprin-associated periodic syndrome, hidradenitis suppurativa, Bechet’s syndrome, or familial cold autoinflammatory syndrome.

[0102] In some embodiments, administering a compound of Formula (I) to a subject that is predisposed to an inflammatory or autoimmune disease prevents the subject from developing any symptoms of the inflammatory or autoimmune disease. In some embodiments, administering a compound of Formula (I) to a subject that does not yet display symptoms of an inflammatory or autoimmune disease prevents the subject from developing any symptoms of the inflammatory or autoimmune disease. In some embodiments, administering a compound of Formula (I) to a subject in need thereof diminishes the extent of the inflammatory or autoimmune disease in the subject. In some embodiments, administering a compound of Formula (I) to a subject in need thereof stabilizes the inflammatory or autoimmune disease (prevents or delays the worsening of the inflammatory or autoimmune disease). In some embodiments, administering a compound of Formula (I) to a subject in need thereof delays the occurrence or recurrence of the inflammatory or autoimmune disease. In some embodiments, administering a compound of Formula (I) to a subject in need thereof slows the progression of the inflammatory or autoimmune disease. In some embodiments, administering a compound of Formula (I) to a subject in need thereof provides a partial remission of the inflammatory or autoimmune disease. In some embodiments, administering a compound of Formula (I) to a subject in need thereof provides a total remission of the inflammatory or autoimmune disease. In some embodiments, administering a compound of Formula (I) to a subject in need thereof decreases the dose of one or more other medications required to treat the inflammatory or autoimmune disease. In some embodiments, administering a compound of Formula (I) to a subject in need thereof enhances the effect of another medication used to treat the inflammatory or autoimmune disease. In some embodiments, administering a compound of Formula (I) to a subject in need thereof delays the progression of the inflammatory or autoimmune disease. In some embodiments, administering a compound of Formula (I) to a subject in need thereof increases the quality of life of the subject having an inflammatory or autoimmune disease. In some embodiments, administering a compound of Formula (I) to a subject in need thereof prolongs survival of a subject having an inflammatory or autoimmune disease.

[0103] In one aspect, provided herein is method of preventing a subject that is predisposed to an inflammatory or autoimmune disease from developing any symptoms of the inflammatory or autoimmune disease, the method comprising administering a compound of Formula (I) to the subject. In some embodiments, provided herein is a method of preventing a subject that does not yet display symptoms of an inflammatory or autoimmune disease from developing any symptomsof the inflammatory or autoimmune disease, the method comprising administering a compound of Formula (I) to the subject.

[0104] In some aspects, provided herein is a method of diminishing the extent of an inflammatory or autoimmune disease in a subject, the method comprising administering a compound of Formula (I) to the subject. In some embodiments, provided herein is a method of stabilizing an inflammatory or autoimmune disease in a subject, the method comprising administering a compound of Formula (I) to the subject. In some embodiments, the method prevents the worsening of the inflammatory or autoimmune disease. In some embodiments, the method delays the worsening of the inflammatory or autoimmune disease.

[0105] In another aspect, provided herein is a method of delaying the occurrence or recurrence of an inflammatory or autoimmune disease in a subject, the method comprising administering a compound of Formula (I) to the subject.

[0106] In some embodiments, provided herein is a method of slowing the progression of an inflammatory or autoimmune disease in a subject, the method comprising administering a compound of Formula (I) to the subject. In some embodiments, the method provides a partial remission of the inflammatory or autoimmune disease. In some embodiments, the method provides a total remission of the inflammatory or autoimmune disease.

[0107] In further aspects, provided herein is a method of decreasing the dose of one or more other medications required to treat an inflammatory or autoimmune disease in a subject, the method comprising administering a compound of Formula (I) to the subject. In some embodiments, provided herein is a method of enhancing the effect of another medication used to treat an inflammatory or autoimmune disease in a subject, the method comprising administering a compound of Formula (I) to the subject.

[0108] Also provided here is a method of delaying the progression of an inflammatory or autoimmune disease in a subject, the method comprising administering a compound of Formula (I) to the subject. In some embodiments, the method increases the quality of life of the subject having an inflammatory or autoimmune disease. In some embodiments, the method prolongs survival of the subject having an inflammatory or autoimmune disease.

[0109] In another aspect, provided herein is a method for treating inflammatory or autoimmune symptoms caused by a disease in a subject in need thereof, comprising administering to the subject an effective amount of a compound of Formula (I). In some embodiments, provided herein is a method for preventing inflammatory or autoimmune symptoms caused by a disease in a subject in need thereof, comprising administering to the subject an effective amount of a compound of Formula (I). In some embodiments, administering a compound of Formula (I) to a subject that ispredisposed to a disease which causes inflammatory or autoimmune symptoms prevents the subject from developing any inflammatory or autoimmune symptoms. In some embodiments, administering a compound of Formula (I) to a subject that does not yet display inflammatory or autoimmune symptoms of a disease which causes inflammatory or autoimmune symptoms prevents the subject from developing any inflammatory or autoimmune symptoms. In some embodiments, administering a compound of Formula (I) to a subject in need thereof diminishes the extent of the inflammatory or autoimmune symptoms caused by the disease in the subject. In some embodiments, administering a compound of Formula (I) to a subject in need thereof stabilizes the inflammatory or autoimmune symptoms of the disease (prevents or delays the worsening of the inflammatory or autoimmune symptoms). In some embodiments, administering a compound of Formula (I) to a subject in need thereof delays the occurrence or recurrence of the inflammatory or autoimmune symptoms caused by the disease. In some embodiments, administering a compound of Formula (I) to a subject in need thereof slows the progression of the inflammatory or autoimmune symptoms caused by the disease. In some embodiments, administering a compound of Formula (I) to a subject in need thereof provides a partial remission of the disease which causes inflammatory or autoimmune symptoms. In some embodiments, administering a compound of Formula (I) to a subject in need thereof provides a total remission of the disease which causes inflammatory or autoimmune symptoms. In some embodiments, administering a compound of Formula (I) to a subject in need thereof decreases the dose of one or more other medications required to treat the disease which causes inflammatory or autoimmune symptoms. In some embodiments, administering a compound of Formula (I) to a subject in need thereof enhances the effect of another medication used to treat the inflammatory or autoimmune symptoms of the disease. In some embodiments, administering a compound of Formula (I) to a subject in need thereof delays the progression of the disease which causes inflammatory or autoimmune symptoms. In some embodiments, administering a compound of Formula (I) to a subject in need thereof increases the quality of life of the subject having a disease which causes inflammatory or autoimmune symptoms. In some embodiments, administering a compound of Formula (I) to a subject in need thereof prolongs survival of a subject having a disease which causes inflammatory or autoimmune symptoms. In some embodiments, the disease is atopic dermatitis, asthma, lupus, rheumatoid arthritis, familial mediterranean fever, psoriasis, generalized pustular psoriasis, cryoprin-associated periodic syndrome, hidradenitis suppurativa, Bechet’s syndrome, or familial cold autoinflammatory syndrome.

[0110] In some embodiments, compounds of Formula (I) are useful for treating a disorder selected from atopic dermatitis, asthma, lupus, rheumatoid arthritis, familial mediterranean fever,psoriasis, generalized pustular psoriasis, cryoprin-associated periodic syndrome, hidradenitis suppurativa, Bechet’s syndrome, and familial cold autoinflammatory syndrome.

[0111] In some embodiments, compounds of Formula (I) are useful for treating a cancer. In some embodiments, the cancer is a solid tumor, skin cancer, or lymphoma. In some embodiments, the cancer is squamous-cell carcinoma, basal cell carcinoma, adenocarcinoma, hepatocellular carcinoma, renal cell carcinoma, cancer of the bladder, bowel, breast, cervix, colon, esophagus, head, kidney, liver, lung, neck, ovary, pancreas, prostate, or stomach, leukemia, benign and malignant lymphomas, Burkitt's lymphoma, Non-Hodgkin’s lymphoma, benign and malignant melanomas, myeloproliferative diseases, sarcomas, Ewing’s sarcoma, hemangiosarcoma, Kaposi’s sarcoma, liposarcoma, myosarcomas, peripheral neuroepithelioma, synovial sarcoma, gliomas, astrocytomas, oligodendrogliomas, ependymomas, gliobastomas, neuroblastomas, ganglioneuromas, gangliogliomas, medulloblastomas, pineal cell tumors, meningiomas, meningeal sarcomas, neurofibromas, Schwannomas, bowel cancer, breast cancer, prostate cancer, cervical cancer, uterine cancer, lung cancer, ovarian cancer, testicular cancer, thyroid cancer, astrocytoma, esophageal cancer, pancreatic cancer, stomach cancer, liver cancer, colon cancer, melanoma; carcinosarcoma, Hodgkin’s disease, Wilms’ tumor, or teratocarcinomas. Additional cancers which may be treated using compounds of Formula (I) include, for example, T-lineage Acute lymphoblastic Leukemia (T-ALL), T-lineage lymphoblastic Lymphoma (T-LL), Peripheral T-cell lymphoma, Adult T-cell Leukemia, Pre-B ALL, Pre-B Lymphomas, Large B-cell Lymphoma, B- cell ALL, Philadelphia chromosome positive ALL and Philadelphia chromosome positive CML.

[0112] In some embodiments, the cancer is breast cancer, colorectal cancer, non-small cell lung cancer, ovarian, renal, sarcoma, melanoma, head and neck, hepatocellular, thyroid, multidrugresistant leukemia, lymphoma, multiple myeloma, esophageal, large bowel, pancreatic, mesothelioma, carcinoma (e.g., adenocarcinoma, including esophageal adenocarcinoma), sarcoma (e.g., spindle cell sarcoma, liposarcoma, leiomyosarcoma, abdominal leiomyosarcoma, sclerosing epithelioid sarcoma) and melanoma (e.g., metastatic malignant melanoma).

[0113] In some embodiments, the compounds of Formula (I) are useful for treating fibrosis, such as interstitial lung fibrosis, cystic fibrosis, progressive pulmonary fibrosis, and idiopathic pulmonary fibrosis.Pharmaceutical Compositions and Routes of Administration

[0114] The compounds provided herein can be administered to a subject orally, topically or parenterally in the conventional form of preparations, such as capsules, microcapsules, tablets, granules, powder, troches, pills, suppositories, injections, suspensions, syrups, patches, creams, lotions, ointments, gels, sprays, solutions and emulsions.

[0115] The compounds disclosed herein can be administered to a subject orally, topically or parenterally in the conventional form of preparations, such as capsules, microcapsules, tablets, granules, powder, troches, pills, suppositories, injections, suspensions, syrups, patches, creams, lotions, ointments, gels, sprays, solutions and emulsions. Suitable formulations can be prepared by methods commonly employed using conventional, organic or inorganic additives, such as an excipient (e.g., sucrose, starch, mannitol, sorbitol, lactose, glucose, cellulose, talc, calcium phosphate or calcium carbonate), a binder (e.g., cellulose, methylcellulose, hydroxymethylcellulose, polypropylpyrrolidone, polyvinylpyrrolidone, gelatin, gum arabic, polyethyleneglycol, sucrose or starch), a disintegrator (e.g., starch, carboxymethylcellulose, hydroxypropyl starch, low substituted hydroxypropylcellulose, sodium bicarbonate, calcium phosphate or calcium citrate), a lubricant (e.g., magnesium stearate, light anhydrous silicic acid, talc or sodium lauryl sulfate), a flavoring agent (e.g., citric acid, menthol, glycine or orange powder), a preservative (e.g, sodium benzoate, sodium bisulfite, methylparaben or propylparaben), a stabilizer (e.g., citric acid, sodium citrate or acetic acid), a suspending agent (e.g., methylcellulose, polyvinyl pyrrolidone or aluminum stearate), a dispersing agent (e.g., hydroxypropylmethylcellulose), a diluent (e.g., water), and base wax (e.g., cocoa butter, white petrolatum or polyethylene glycol). The effective amount of the compounds of Formula (I) in the pharmaceutical composition may be at a level that will exercise the desired effect.

[0116] A compound of Formula (I) can be administered orally for reasons of convenience. In one embodiment, when administered orally, a compound of Formula (I) is administered with a meal and water. In another embodiment, the compound of Formula (I) is dispersed in water or juice (e.g., apple juice or orange juice) or any other liquid and administered orally as a solution or a suspension.

[0117] The compounds disclosed herein can also be administered intradermally, intramuscularly, intraperitoneally, percutaneously, intravenously, subcutaneously, intranasally, epidurally, sublingually, intracerebrally, intravaginally, transdermally, rectally, mucosally, by inhalation, or topically to the ears, nose, eyes, or skin. The mode of administration is left to the discretion of the health-care practitioner, and can depend inpart upon the site of the medical condition.

[0118] In one embodiment, provided herein are capsules containing a compound of Formula (I) without an additional carrier, excipient or vehicle.

[0119] In another embodiment, provided herein are compositions comprising an effective amount of a compound of Formula (I) and a pharmaceutically acceptable carrier or vehicle,wherein a pharmaceutically acceptable carrier or vehicle can comprise an excipient, diluent, or a mixture thereof. In one embodiment, the composition is a pharmaceutical composition.

[0120] The compositions can be in the form of tablets, chewable tablets, capsules, solutions, parenteral solutions, troches, suppositories and suspensions and the like. Compositions can be formulated to contain a daily dose, or a convenient fraction of a daily dose, in a dosage unit, which may be a single tablet or capsule or convenient volume of a liquid. In one embodiment, the solutions are prepared from water-soluble salts, such as the hydrochloride salt. In general, all of the compositions are prepared according to known methods in pharmaceutical chemistry. Capsules can be prepared by mixing a compound of Formula (I) with a suitable carrier or diluent and filling the proper amount of the mixture in capsules. The usual carriers and diluents include, but are not limited to, inert powdered substances such as starch of many different kinds, powdered cellulose, especially crystalline and microcrystalline cellulose, sugars such as fructose, mannitol and sucrose, grain flours and similar edible powders.

[0121] Tablets can be prepared by direct compression, by wet granulation, or by dry granulation. Their formulations usually incorporate diluents, binders, lubricants and disintegrators as well as the compound. Typical diluents include, for example, various types of starch, lactose, mannitol, kaolin, calcium phosphate or sulfate, inorganic salts such as sodium chloride and powdered sugar. Powdered cellulose derivatives are also useful. Typical tablet binders are substances such as starch, gelatin and sugars such as lactose, fructose, glucose and the like. Natural and synthetic gums are also convenient, including acacia, alginates, methylcellulose, polyvinylpyrrolidine and the like. Polyethylene glycol, ethylcellulose and waxes can also serve as binders.

[0122] A lubricant might be necessary in a tablet formulation to prevent the tablet and punches from sticking in the dye. The lubricant can be chosen from such slippery solids as talc, magnesium and calcium stearate, stearic acid and hydrogenated vegetable oils. Tablet disintegrators are substances that swell when wetted to break up the tablet and release the compound. They include starches, clays, celluloses, algins and gums. More particularly, com and potato starches, methylcellulose, agar, bentonite, wood cellulose, powdered natural sponge, cation-exchange resins, alginic acid, guar gum, citrus pulp and carboxymethyl cellulose, for example, can be used as well as sodium lauryl sulfate. Tablets can be coated with sugar as a flavor and sealant, or with filmforming protecting agents to modify the dissolution properties of the tablet. The compositions can also be formulated as chewable tablets, for example, by using substances such as mannitol in the formulation.

[0123] When it is desired to administer a compound of Formula (I) as a suppository, typical bases can be used. Cocoa butter is a traditional suppository base, which can be modified by addition of waxes to raise its melting point slightly. Water-miscible suppository bases comprising, particularly, polyethylene glycols of various molecular weights are in wide use.

[0124] The effect of the compound of Formula (I) can be delayed or prolonged by proper formulation. For example, a slowly soluble pellet of the compound of Formula (I) can be prepared and incorporated in a tablet or capsule, or as a slow-release implantable device. The technique also includes making pellets of several different dissolution rates and filling capsules with a mixture of the pellets. Tablets or capsules can be coated with a film that resists dissolution for a predictable period of time. Even the parenteral preparations can be made long-acting, by dissolving or suspending the compound of Formula (I) in oily or emulsified vehicles that allow it to disperse slowly in the serum.Exemplary Embodiments

[0125] The present disclosure is further described by the following embodiments.

[0126] Embodiment Pl. A compound of Formula (I)or a pharmaceutically acceptable salt thereof, wherein:Ring B is 5- to 6-membered monocyclic heterocyclylene or 8- to 10-membered fused bicyclic heterocyclylene, wherein the heterocyclylene contains 1-3 nitrogen atoms; each R2is independently Ci-Ce alkyl or halo;n is 0-3;R3is H, Ci-Ce alkyl, or halo;Ringattached to two adjacent carbon atoms indicated with *; andX is O, CH2, or N(H).

[0127] Embodiment P2. The compound of embodiment Pl, or a pharmaceutically acceptable salt thereof, wherein the compound is of Formula (I’):

[0128] Embodiment P3. The compound of embodiment P 1 , or a pharmaceutically acceptable salt thereof, wherein the compound is of Formula (II):

[0129] Embodiment P4. The compound of embodiment Pl, or a pharmaceutically acceptable salt thereof, wherein the compound is of Formula (III):

[0130] Embodiment P5. The compound of any one of embodiments P1-P3, or a pharmaceutically acceptable salt thereof, wherein: R1is H.

[0131] Embodiment P6. The compound of any one of embodiments P1-P3, or a pharmaceutically acceptable salt thereof, wherein: R1is -NH2.

[0132] Embodiment P7. The compound of any one of embodiments P1-P6, or a pharmaceutically acceptable salt thereof, wherein:L1is -C(O)N(H)-.

[0133] Embodiment P8. The compound of any one of embodiments P1-P6, or a pharmaceutically acceptable salt thereof, wherein:

[0134] Embodiment P9. The compound of any one of embodiments P1-P8, or a pharmaceutically acceptable salt thereof, wherein:Ring B is 6-membered monocyclic heterocyclylene or 8-membered fused bicyclic heterocyclylene, wherein the heterocyclylene contains 1-2 nitrogen atoms.

[0135] Embodiment P10. The compound of embodiment P9, or a pharmaceutically acceptable salt thereof, wherein:Ring

[0136] Embodiment Pl 1. The compound of any one of embodiments Pl -PIO, or a pharmaceutically acceptable salt thereof, wherein: n is 0 or 1.

[0137] Embodiment P12. The compound of any one of embodiments Pl-Pl 1, or apharmaceutically acceptable salt thereof, wherein: each R2is independently C1-C3 alkyl or halo.

[0138] Embodiment P13. The compound of embodiment Pl 2, or a pharmaceutically acceptable salt thereof, wherein: each R2is independently -CH3or -CH2CH3.

[0139] Embodiment P14. The compounds of any one of embodiments P1-P13, or a pharmaceutically acceptable salt thereof, wherein:

[0140] Embodiment Pl 5. The compound of any one of embodiments Pl -Pl 4, or a pharmaceutically acceptable salt thereof, wherein:R3is H or C1-C3 alkyl.

[0141] Embodiment Pl 6. The compound of embodiment Pl 5, or a pharmaceutically acceptable salt thereof, wherein:R3is H or -CH3.

[0142] Embodiment Pl 7. The compound of any one of embodiments Pl -Pl 6, or a pharmaceutically acceptable salt thereof, wherein:X is O.

[0143] Embodiment Pl 8. The compound of any one of embodiments Pl -Pl 6, or a pharmaceutically acceptable salt thereof, wherein:X is CH2or N(H).

[0144] Embodiment Pl 9. The compound of any one of embodiments Pl -Pl 8, or a pharmaceutically acceptable salt thereof, wherein:

[0145] Embodiment P20. The compound of any one of embodiments P1-P3, P5, P6, P8-P17, and P19, or a pharmaceutically acceptable salt thereof, wherein the compound is of Formula (IIA),(IIB), (IIC), (HD), (HE), or (IIF):

[0146] Embodiment P21. The compound of any one of embodiments Pl, P2, P4-P6, P8-P17, and P19, or a pharmaceutically acceptable salt thereof, wherein the compound is of Formula (IIIA),(IIIB), (IIIC), (IIID), (IIIE), or (IIIF):

[0147] Embodiment P22. The compound of embodiment P20 or P21, or a pharmaceuticallyacceptable salt thereof, wherein:X is O.

[0148] Embodiment P23. A compound selected from the compounds of Table 1 and pharmaceutically acceptable salts thereof.

[0149] Embodiment P24. A pharmaceutical composition comprising the compound of any one of embodiments P1-P23, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable excipient.

[0150] Embodiment P25. A method of modulating interleukin- 1 (IL1) receptor-associated kinase 4 (IRAK4) activity comprising contacting IRAK4 with an effective amount of the compound of any one of embodiments P1-P23, or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition of embodiment P24.

[0151] Embodiment P26. A method of treating an inflammatory or autoimmune disease in a subject in need thereof, comprising administering to the subject an effective amount of the compound of any one of embodiments P1-P23, or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition of embodiment P24.

[0152] Embodiment P27. The method of embodiment P26, wherein the inflammatory or autoimmune disease is atopic dermatitis, asthma, lupus, rheumatoid arthritis, familial mediterranean fever, psoriasis, generalized pustular psoriasis, cryoprin-associated periodic syndrome, hidradenitis suppurativa, Bechet’s syndrome, or familial cold autoinflammatory syndrome.EXAMPLES

[0153] The following Examples are presented by way of illustration, not limitation.Compounds are named using the automatic name generating tool provided in ChemBiodraw Ultra (Cambridgesoft), which generates systematic names for chemical structures, with support for the Cahn-Ingold-Prelog rules for stereochemistry. One skilled in the art can modify the procedures set forth in the illustrative examples to arrive at the desired products.

[0154] Salts of the compounds described herein can be prepared by standard methods, such as inclusion of an acid (for example TFA, formic acid, or HC1) in the mobile phases during chromatography purification, or stirring of the products after chromatography purification, with a solution of an acid (for example, aqueous HC1).

[0155] As used in certain of the chemical structures provided in the following Examples, designation of a particular atom with “orl” or “or2” indicates that the absolute stereochemistry of the indicated atom was not determined.

[0156] The following abbreviations may be relevant for the application.AbbreviationsACN or MeCN: acetonitrileAcOH: acetic acidADMP: 2-azido-l,3-dimethylimidazolinium hexafluorophosphate aq: aqueousBoc: tert-butyloxycarbonylCAM: ceric ammonium molybdate cat.: catalyticCBM: cereblon binding moiety cone.: concentratedC V : column volumeDCM: dichloromethaneDHP: dihydropyranDIBAL-H: diisobutylaluminum hydrideDIPEA: N,N-diisopropylethylamineDMAP: 4-dimethylaminopyridineDMF: dimethylformamideDMP: Dess-Martin PeriodinaneDMSO: dimethyl sulfoxide eq.: equivalentsEtOAc: ethyl acetate h: hour(s)HATU: l-[bis(dimethylamino)methylene]-lH-l,2,3-triazolo[4,5-b]pyridinium 3-oxide hexafluorophosphate hept.: heptanesIBX: 2-iodoxybenzoic acidMeOH: methanol min: minute(s)MTBE: methyl tert-butyl etherOAc: acetatePdCh(dppf): bis(diphenylphosphino)ferrocene)palladium(II) dichloridePd2(dba)s: tris(dibenzylideneacetone)dipalladium(0) pet ether: petroleum etherPhMe: toluenePPTS: pyridinium p-toluenesulfonatePrep-HPLC: preparative High Performance Liquid Chromatography.PTSA or / >TSA: p-toluenesulfonic acidRT : room temperatureRuPhos Pd G3: (2-dicyclohexylphosphino-2',6'-diisopropoxy-l,l'-biphenyl)[2-(2'-amino-l,l'- biphenyl)]palladium(II) methanesulfonateRuPhos Pd G4: [dicyclohexyl(2',6'-diisopropoxy-2-biphenylyl)phosphine-kP](methanesulfonatato- KO )[2'-(methylamino-kN )-2-biphenylyl-kC2]palladium sat.: saturatedSFC: supercritical-fluid chromatographyTBAF: tetrabutylammonium fluorideTBM: target binding moietyTFA: trifluoroacetic acidTFAA: trifluoroacetic anhydrideTHF: tetrahydrofuranTLC: thin layer chromatographyUPLC: ultra-performance liquid chromatographyXantphos:(SP-4-3)-[[5-(diphenylphosphino)-9,9-dimethyl-9H -xanthen-4-yl]diphenylphosphine- KP](methanesulfonato-kO )[2'-(methylamino-kN )[l,r-biphenyl]-2-yl-kC]- PalladiumSynthetic ExamplesProcedures for Linker Synthesis

[0157] The numbering of the intermediate compounds referred to in each example is limited to each example only. For instance, intermediate 1’ in Example SI and intermediate 1’ in Example S2 are described in different examples and, as such, refer to different compounds.Example SI: Preparation of intermediate alkyne 2-(2-((1r,4r)-4- ethynylcyclohexyl)ethoxy)tetrahydro-2H-pyran (L-l) and 2-((1r,4r)-4- ethynylcyclohexyl)ethan-1-ol (L-2)

[0158] Step 1’. Preparation of [4-[[tert- butyl(diphenyl)silyl]oxymethyl]cyclohexyl]methanol 2’: To a solution of [4-(hydroxymethyl)cyclohexyl]methanol 1’ (3 g, 20.8 mmol, 1 eq.) in dry DMF (83.2 mL, 0.25 M) was added imidazole (0.85 g, 12.48 mmol, 0.6 eq.), followed by tert-butylchlorodiphenylsilane (2.7 mL, 10.4 mmol, 0.5 eq.). After stirring for 18 h at room temperature, TLC (25 % EtOAc in Heptanes, revealing UV + KMnCh) showed full conversion. To the reaction mixture was added half-brine solution (40 mL). The aqueous phase was extracted with EtOAc (2 x 50 mL). The organics were washed with half-brine (4 x 25 mL), dried over Na2SO4 and concentrated to dryness. The residue was dry-packed and purified by normal phase flash chromatography (80 g silica column, elution: 0 to 30 % EtOAc / Heptane over 10 CV, product came out around 20 % EtOAc). Fractions were combined and concentrated to give 2’ (6.48 g, 43 % yield) as a colorless oil.

[0159] 1H NMR (400 MHz, chloroform-d): 5 ppm 0.82 - 1.02 (m, 4 H), 1.03 - 1.07 (m, 9 H), 1.38 - 1.55 (m, 2 H), 1.66 - 1.88 (m, 4 H), 2.04 (br d, J= 3.2 Hz, 2 H), 3.42 - 3.49 (m, 3 H), 7.32 - 7.44 (m, 6 H), 7.65 (d, J= 6.6 Hz, 3 H), 7.70 - 7.75 (m, 1 H).

[0160] Step 2’. Preparation of [4-[[tert-butyl(diphenyl)silyl]oxymethyl]cyclohexyl]methyl methanesulfonate 3’: To a round bottom flask was added [4-[[tert- butyl(diphenyl)silyl]oxymethyl]cyclohexyl]methanol 2’ (6.48 g, 8.93 mmol, 1 eq.) in DCM (44.65 mL, 0.2 M). The flask was then cooled to 0 °C, then methanesulfonyl chloride (0.76 mL, 9.82 mmol, 1.1 eq.) and triethylamine (1.62 mL, 11.61 mmol, 1.3 eq.) were added. After 2 h, TLC(Heptanes / EtOAc; 10 / 90) showed complete conversion of the starting material into the desired product (KMnO4 revelator). The mixture was partitioned between EtOAc and water. The organic phase was washed once with water and once with 1 N HC1, dried over magnesium sulfate, filtered and evaporated under reduced pressure, affording 3’ (3.65 g, 89 % yield) as a colorless solid. The crude was used directly as is for the next reaction.

[0161] 1H NMR (400 MHz, chloroform-d): 5 ppm 1.03 (br s, 3 H), 1.07 (br d, J= 12.7 Hz, 9 H), 1.45 - 1.56 (m, 1 H), 1.64 - 1.77 (m, 1 H), 1.83 - 1.90 (m, 3 H), 3.01 (s, 3 H), 3.48 (d, J= 6.1 Hz, 2 H), 4.05 (d, J= 6.6 Hz, 2 H), 7.35 - 7.47 (m, 6 H), 7.64 - 7.69 (m, 3 H), 7.71 - 7.79 (m, 1 H). Two protons were not apparent by1H NMR.

[0162] Step 3’. Preparation of 2-[4-[[tert- butyl(diphenyl)silyl]oxymethyl]cyclohexyl]acetonitrile 4’: To a round bottom flask were added [4-[[tert-butyl(diphenyl)silyl]oxymethyl]cyclohexyl]methyl methanesulfonate 3’ (3.65 g, 7.92 mmol, 1 eq.) and NaCN (1.19 g, 18.21 mmol, 2.3 eq.) in DMSO (12.2 mL, 0.65 M). The reaction mixture was then stirred at 50 °C. After an overnight period, TLC (Heptanes / EtOAc; 50 / 50) showed complete conversion of the starting material into compound 4’ (KMnO4 revelator). The reaction mixture was poured into an Erlenmeyer flask of crushed ice and stirred until all ice was melted, and the resulting solid was isolated by filtration and dissolved in a minimum of ethyl acetate, washed with NaHCOs and brine, and evaporated to dryness. The residue was dry-packed and purified by normal phase flash chromatography (80 g silica column, elution: 5 to 100 % EtOAc / Heptane over 15 CV). Fractions were combined and concentrated to give 4’ (2.35 g, 76 % yield) as a colorless oil.

[0163] 1H NMR (400 MHz, chloroform-d): 5 ppm 0.83 - 0.91 (m, 1 H), 0.98 - 1.05 (m, 2 H),I.05 - 1.09 (m, 9 H), 1.09 - 1.18 (m, 2 H), 1.46 - 1.56 (m, 1 H), 1.58 - 1.65 (m, 1 H), 1.89 (br t, J =I I.9 Hz, 3 H), 2.27 (d, J= 6.6 Hz, 2 H), 3.48 (d, J = 5.9 Hz, 2 H), 7.37 - 7.46 (m, 6 H), 7.64 - 7.69 (m, 3 H), 7.71 - 7.75 (m, 1 H).

[0164] Step 4’. Preparation of 2-[4-[[tert- butyl(diphenyl)silyl]oxymethyl]cyclohexyl]acetaldehyde 5’: To a round bottom flask was added 2-[4-[[tert-butyl(diphenyl)silyl]oxymethyl]cyclohexyl]acetonitrile 4’ (2.35 g, 6 mmol, 1 eq.) in DCM (10.7 mL, 0.56 M), then the reaction mixture was cooled at -78 °C. To the cooled reaction mixture was then added DIBAL-H (1 M solution in DCM) (17.99 mL, 17.99 mmol, 3 eq.) and the reaction stirred at -78 °C. After 4 h, 40 mL of 4 M HC1 were added carefully and the solution was slowly warmed to RT. Stirring was continued at RT for 10 min, and the substance was partitioned between ethyl acetate and 1 M HC1. The organic phases were washed with water, dried over magnesium sulfate and concentrated under reduced pressure. The residue was dry-packed andpurified by normal phase flash chromatography (40 g silica column, elution: 1 :3 Hept.:EtOAc). Fractions were combined and concentrated to give 5’ (803 mg, 34 % yield) as a colorless oil that was used as is for the next reaction.

[0165] Step 5’. Preparation of 2-[4-[[tert- butyl(diphenyl)silyl]oxymethyl]cyclohexyl]ethanol 6’: To a round bottom flask was added 2-[4- [[tert-butyl(diphenyl)silyl]oxymethyl]cyclohexyl]acetaldehyde 5’ (800 mg, 2.03 mmol, 1 eq.) in EtOH (10.2 mL, 0.2 M) at 0 °C. To the reaction mixture was added NaBEU (192.26 mg, 5.08 mmol, 2.5 eq.) and the reaction stirred at 0 °C. After 4 h, Rochelle salt solution was added at 0°C, then the solution was heated up to room temperature. Organic layers were extracted twice with EtOAc and the latter was washed with brine twice and dried over MgSO4. The residue was dry-packed and purified by normal phase flash chromatography (40 g silica column, elution: 0 to 30 % EtOAc / Heptane over 15 CV, product came out around 20 % EtOAc). Fractions were combined and concentrated to give 6’ (353.7 mg, 44 % yield) as a colorless oil.

[0166] 1H NMR (400 MHz, chloroform-d): 5 ppm 0.93 - 1.03 (m, 4 H), 1.06 (s, 9 H), 1.13 - 1.20 (m, 1 H), 1.31 - 1.41 (m, 1 H), 1.50 (q, J= 6.8 Hz, 3 H), 1.74 - 1.86 (m, 4 H), 3.47 (d, J= 6.1 Hz, 2 H), 3.67 - 3.73 (m, 2 H), 7.36 - 7.45 (m, 6 H), 7.67 (dd, J= 7.8, 1.5 Hz, 4 H).

[0167] Step 6’. Preparation of tert-butyl-diphenyl-[[4-(2-tetrahydropyran-2- yloxyethyl)cyclohexyl] methoxy] silane 7’: To a solution of 2-[4-[[tert- butyl(diphenyl)silyl]oxymethyl]cyclohexyl]ethanol 6’ (1.62 g, 4.08 mmol, 1 eq.) in DCM (25.06 mL, 0.16 M) was added PPTS (205.28 mg, 0.82 mmol, 0.2 eq.) and DHP (0.93 mL, 10.21 mmol, 2.5 eq.). The reaction was stirred at room temperature. After 24 h, TLC showed complete conversion of the starting material 6’. The reaction mixture was concentrated to dryness, and the residue was dry-packed and purified by normal phase flash chromatography (80 g silica column, elution: 0 to 30 % EtOAc / Heptane over 15 CV, product came out around 20 % EtOAc). Fractions were combined and concentrated to give 7’ (1.79 g, 91 % yield) as a colorless oil.

[0168] 1H NMR (400 MHz, chloroform-d): 5 ppm 0.84 - 0.91 (m, 3 H), 0.93 - 1.00 (m, 3 H), 1.03 - 1.11 (m, 9 H), 1.29 - 1.39 (m, 2 H), 1.50 - 1.55 (m, 3 H), 1.56 - 1.63 (m, 2 H), 1.70 (br s, 5 H), 3.40 - 3.45 (m, 1 H), 3.47 (d, J= 6.1 Hz, 2 H), 3.48 - 3.55 (m, 1 H), 3.76 - 3.84 (m, J= 9.7, 7.2, 7.2 Hz, 1 H), 3.84 - 3.92 (m, 1 H), 4.55 - 4.61 (m, 1 H), 7.35 - 7.45 (m, 6 H), 7.64 - 7.76 (m, 4 H).

[0169] Step 7’. Preparation of [4-(2-tetrahydropyran-2-yloxyethyl)cyclohexyl]methanol8’: To a stirred solution of tert-butyl-diphenyl-[[4-(2-tetrahydropyran-2- yloxyethyl)cyclohexyl]methoxy]silane 7’ (1.79 g, 3.72 mmol, 1 eq.) in THF (4.65 mL, 0.8 M) was added 1 M TBAF solution in THF (14.89 mL, 14.89 mmol, 4 eq.) at room temperature. After 5 h, TLC showed complete conversion of the starting material 7’. Solvents were removed under reducedpressure and the residue was purified by normal phase flash chromatography (80 g gold column, solid deposit, elution 0 to 40 % EtOAc / Heptane over 10 CV) (CAM was used as the TLC stain). Fractions were combined and concentrated to give 8’ (577.5 mg, 58 % yield) as a colorless oil.

[0170] 1H NMR (400 MHz, chloroform-d): 5 ppm 0.90 - 1.01 (m, 4 H), 1.33 - 1.46 (m, 2 H), 1.48 - 1.63 (m, 7 H), 1.68 - 1.76 (m, 1 H), 1.77 - 1.89 (m, 5 H), 3.39 - 3.54 (m, 4 H), 3.77 - 3.91 (m, 2 H), 4.56 - 4.60 (m, 1 H).

[0171] Step 8’. Preparation of 4-(2-tetrahydropyran-2- yloxyethyl)cyclohexanecarbaldehyde 9’: To a stirred solution of [4-(2-tetrahydropyran-2- yloxyethyl)cyclohexyl]methanol 8’ (704.5 mg, 2.91 mmol, 1 eq.) in DCM (36.34 mL, 0.04 M) and THF (36.34 mL, 0.04 M) was added DMP (2.47 g, 5.81 mmol, 2 eq.) and water (3 drops) at 0 °C. After 2.5 h, TLC showed complete conversion of the starting material 8’. Solvents were removed under reduced pressure without a heating bath, taken up with EtOAc, washed with sat. NaHCCL, extracted with EtOAc (3x), and then washed with brine. The combined organic layers were dried over Na2SO4, filtered and concentrated under reduced pressure. The residue was then purified by normal phase flash chromatography (24 g column, solid deposit, elution 0 to 30 % EtOAc / Heptane over 10 CV). Fractions were combined and concentrated to give 9’ (459.5 mg, 66 % yield) as a colorless solid.

[0172] 1H NMR (400 MHz, DMSO-d6): 5 ppm 0.92 - 1.01 (m, 2 H), 1.08 - 1.22 (m, 2 H), 1.26 - 1.36 (m, 1 H), 1.38 - 1.53 (m, 6 H), 1.55 - 1.64 (m, 1 H), 1.65 - 1.83 (m, 3 H), 1.84 - 1.94 (m, 2 H), 2.15 - 2.25 (m, 1 H), 3.32 - 3.45 (m, 2 H), 3.62 - 3.77 (m, 2 H), 4.49 - 4.56 (m, 1 H), 9.55 (d, J = 1.0 Hz, 1 H).

[0173] Step 9’. Preparation of 2-[2-(4-ethynylcyclohexyl)ethoxy]tetrahydropyran L-l: To a flame-dried round bottom flask were added 4-(2-tetrahydropyran-2- yloxyethyl)cyclohexanecarbaldehyde 9’ (459.5 mg, 1.91 mmol, 1 eq.) and K2CO3 (494.18 mg, 3.82 mmol, 2 eq.). Vacuum was applied and the flask was then filled with nitrogen (repeated three times). Then, MeOH (9.56 mL, 0.2 M) was added and the reaction mixture stirred at room temperature under nitrogen for 20 minutes. Then, Bestmann-Ohira reagent (4.4 mL, 2.29mmol, 1.2 eq.) (10% in acetonitrile) was added to the reaction mixture. The resulting mixture was stirred at room temperature under a nitrogen atmosphere. After 4 h, TLC (eluting 10% EtOAc in Heptanes) showed complete conversion of starting material 9’. The residue was dry-packed and then purified by normal phase flash chromatography (24 g column, solid deposit, elution 0 to 10 % EtOAc / Heptane over 10 CV). Fractions were combined and concentrated to give L-l (275.7 mg, 57 % yield) as a colorless oil.

[0174] 1H NMR (400 MHz, chloroform-d): 5 ppm 0.88 - 0.98 (m, 2 H), 1.33 - 1.45 (m, 3 H),1.48 - 1.62 (m, 7 H), 1.68 - 1.76 (m, 1 H), 1.77 - 1.88 (m, 3 H), 1.95 - 2.03 (m, 2 H), 2.14 - 2.24 (m, 1 H), 3.36 - 3.56 (m, 2 H), 3.75 - 3.90 (m, 2 H), 4.55 - 4.59 (m, 1 H).

[0175] Step 10’. Preparation of 4-(2-tetrahydropyran-2- yloxyethyl)cyclohexanecarbaldehyde L-2: To a stirred solution of 2-[2-(4- ethynylcyclohexyl)ethoxy]tetrahydropyran L-l (50 mg, 0.2 mmol, 1 eq.) in MeOH (1.97 mL, 0.1 M) was added PTSA (3.39 mg, 0.02 mmol, 0.1 eq.) at room temperature. After 1 h, TLC showed complete conversion of the starting material L-l. The reaction mixture was quenched with triethylamine and the mixture was concentrated under reduced pressure. The residue was then purified by normal phase flash chromatography (24 g gold column, solid deposit, elution 0 to 30 % EtOAc / Heptane over 10 CV) (CAM was used as a TLC revelator). Fractions were combined and concentrated to give L-2 (18 mg, 60 % yield) as a colorless oil.

[0176] 1H NMR (400 MHz, chloroform-d): 5 ppm 0.88 - 1.01 (m, 2 H), 1.30 - 1.50 (m, 6 H), 1.73 - 1.82 (m, 2 H), 1.95 - 2.03 (m, 2 H), 2.03 - 2.07 (m, 1 H), 2.13 - 2.24 (m, 1 H), 3.68 (t, J= 6.6 Hz, 2 H).Example S2: Preparation of intermediate 2-((1r,4r)-4-aminocyclohexyl)ethan-l-ol (L-3) HCI in

[0177] LCMS Method 1. Kinetex XB- C18 75 x 3.0 mm 2.6 p, Temperature: RT, Flow: 1.5 mL / min, Run Time: 5.5 minutes, Mobile Phase Conditions: Mobile Phase-A: 0.1% TFA in H2O, Mobile Phase-B: 0.1% TFA in ACN, Gradient: Initial 95% Mobile Phase A and 5% Mobile Phase B linear gradient to 5% Mobile phase A and 95% Mobile Phase B for 4.0 min. MSD positive.

[0178] Step 1’. Preparation of tert-butyl ((1r,4r)-4-(2-hydroxyethyl)cyclohexyl)carbamate 2’: A solution of 2-((1r,4r)-4-((tert-butoxycarbonyl)amino)cyclohexyl)acetic acid 1’ (10 g, 38.1 mmol) in THF (250 mL) was stirred at 0 °C for 15 minutes under nitrogen atmosphere. To the stirred solution BHs’DMS (7.23 mL, 76 mmol) was added portion wise at 0 °C under nitrogen atmosphere. The reaction mixture was then stirred at 0 °C for 15 minutes and further stirred for 2 hours at room temperature under nitrogen atmosphere. The reaction mixture was concentrated under reduced pressure to afford crude product, which was diluted with cold water and extracted with EtOAc. The combined organic layers were washed with brine solution, dried over Na2SC>4, filtered and concentrated under reduced pressure to afford crude product. The crude product was purified by flash column chromatography using silica gel (230 - 400 mesh) with 40 - 50% ethyl acetate in pet ether to obtain tert-butyl ((1r,4r)-4-(2-hydroxyethyl)cyclohexyl)carbamate 2’ (7.21 g, 78% yield) as a white solid.

[0179] LCMS method 1 : retention time: 2.315 min, [M+H-56]+= 188.2.

[0180] Step 2’. Preparation of 2-((1r,4r)-4-aminocyclohexyl)ethan-l-ol 3’: To a stirred solution of tert-butyl ((1r,4r)-4-(2-hydroxyethyl)cyclohexyl)carbamate 2’ (7.0 g, 28.8 mmol) in DCM (100 mL) was added 4.0 M HC1 in dioxane (36.0 mL, 144 mmol) and the reaction mixture stirred at 25 °C for 30 minutes under nitrogen atmosphere. The solvent was evaporated under reduced pressure to obtain 2-((1r,4r)-4-aminocyclohexyl)ethan-l-ol HC1 3’ (5.465 g) as a white solid, which was used for the next step without further purification.

[0181] LCMS method 1 : retention time 0.493 min, [M+H]+= 144.2.

[0182] 1H NMR (400 MHz, DMSO-d6): 8 ppm 0.88 - 1.00 (m, 2H), 1.25 - 1.38 (m, 5H), 1.69 - 1.80 (m, 2H), 1.90 - 1.99 (m, 2H), 2.83 - 2.93 (m, 1H), 3.41 (t, J= 6.4 Hz, 2H), 8.08 (s, 2H).Procedures for TBM SynthesisExample S3: Preparation of l-(4-(((R)-l-cyanoethyl)amino)-5-(4-((1r,4R)-4-(2- oxoethyl)cyclohexyl)-lH-l,2,3-triazol-l-yl)pyridin-2-yl)-lH-pyrazolo[3,4-b]pyridine-5- carbonitrile (A-l)

[0183] LCMS Method 1. Column: Kinetex XB - C18 75 x 3.0 mm, 2.6 pm. Temperature: 45°C, Flow: 1.0 mL / min, run time: 5.0 min. Mobile Phase Conditions: Mobile Phase-A: 5.0 mm Ammonium formate pH 3.3:CHsCN (98:02), Mobile Phase-B: CH3CN: Buffer (98:02), Gradient:Initial 80% Mobile Phase A and 20% Mobile Phase B linear gradient to 100% Mobile Phase B for 4.0 min then hold for 0.6 minute. MSD positive.

[0184] UPLC Method 2, Column: Zorbax SB - C18 50 x 2.1 mm, 1.8 pm. Temperature: 45 °C, Flow: 0.7 mL / min, run time: 3.0 min. Mobile Phase Conditions: Mobile Phase-A: 5.0 mM Ammonium acetate :CHsCN (95:05), Mobile Phase-B: CH3CN: 5.0 mM Ammonium acetate (95:05), Gradient: Initial 80% Mobile Phase A and 20% Mobile Phase B linear gradient to 100% Mobile Phase B for 2.0 min then hold for 0.3 minute. MSD positive.

[0185] Step 1’. Preparation of (R )-2-((2-chloro-5-nitropyridin-4-yl)amino)propanamide 3’: To a stirred solution of 2, 4-di chi oro-5 -nitropyridine 1’ (20.0 g, 104 mmol, 1.0 eq.) in acetonitrile (200 mL) was added (A)-2-aminopropanamide hydrochloride 2’ (15.49 g, 124 mmol, 1.2 eq.) followed by DIPEA (54.3 mL, 311 mmol, 3.0 eq.) under N2 atmosphere at RT. Then, the reaction was heated to 40 °C for 6 h. The reaction was cooled down to RT, poured into ice cold water (500 mL) and extracted with ethyl acetate (3 X 700 mL). The combined organic layer was washed with brine (2 X 250 mL), dried over sodium sulphate and concentrated under reduced pressure to afford (A)-2-((2-chl oro-5 -nitropyridin-4-yl)amino)propanamide 3’ (27 g, 97% yield) as a pale yellow solid.

[0186] LCMS method 1 : retention time: 0.876 min, 91.45% purity at 220 nm, [M+H]+= 245.0.

[0187] 1H NMR (300 MHz, DMSO-d6): 5 ppm 1.42 (d, J= 8.8 Hz, 3H), 4.34 - 4.39 (m, 1H),6.90 (s, 1H), 7.46 (s, 1H), 7.69 (s, 1H), 8.66 - 8.68 (m, 1H), 8.93 (s, 1H).

[0188] Step 2’. Preparation of (R )-2-((2-chloro-5-nitropyridin-4-yl)amino)propanenitrile 4’: To a stirred solution of (A)-2-((2-chloro-5-nitropyridin-4-yl)amino)propanamide 3’ (27.0 g, 100 mmol, 1.0 eq.) in DCM (270 mL) at 0 °C was added pyridine (81 mL, 1004 mmol, 10.0 eq.) followed by trifluoroacetic anhydride (70.9 mL, 502 mmol, 5.0 eq.) over a period of 10 min. Then, the reaction mixture was slowly warmed to room temperature and stirring was continued for 2 h. The reaction mixture was slowly quenched with ice cold water (300 mL) and extracted with DCM (2 X 300 mL). The combined organic layer was washed with 1.5 N HC1 solution (2 X 200 mL) and brine (2 X 250 mL), dried over sodium sulphate and concentrated under reduced pressure to afford (A)-2-((2-chloro-5-nitropyridin-4-yl)amino)propanenitrile 4’ (21 g, 89% yield) as a pale yellow solid.

[0189] LCMS method 1 : retention time: 1.583 min, 96.3% purity at 220 nm, [M+H]+= 227.0.

[0190] 1H NMR (400 MHz, DMSO-d6): 8 ppm 1.68 (d, J = 7.2 Hz, 3H), 5.18 - 5.22 (m, 1H), 7.35 (s, 1H), 8.42 (d, J= 8.4 Hz, 1H), 8.95 (s, 1H).

[0191] Step 3’. Preparation of (R )-l-(4-((l-cyanoethyl)amino)-5-nitropyridin-2-yl)-lH- pyrazolo[3,4-b]pyridine-5-carbonitrile 6’: To a stirred solution of (A)-2-((2-chloro-5-nitropyridin-4-yl)amino)propanenitrile 4’ (1.965 g, 8.33 mmol, 1.0 eq.) and U / -pyrazolo[3,4- b ]pyridine-5-carbonitrile 5’ (1.2 g, 8.33 mmol, 1.0 eq.) in 1,4-dioxane (70 mL) was added zinc acetate (0.917 g, 5.00 mmol, 0.6 eq.) and K2CO3 (2.301 g, 16.65 mmol, 2.0 eq.) under N2 atmosphere and the purging was continued for 10 min. Then, Xantphos (0.482 g, 0.833 mmol, 0.1 eq.) and Pd2(dba)s (0.381 g, 0.416 mmol, 0.05 eq.) was added under N2atmosphere and the purging was continued for another 5 min. Then, the reaction mixture was stirred at 100 °C for 16 h. The reaction was cooled to RT and filtered through a Celite bed, and the residue (filter cake) was washed with DCM (500 mL). The filtrate was concentrated under reduced pressure to give the crude product. The crude product was purified by column chromatography using silica gel (230- 400 mesh) with 80% ethyl acetate in pet ether to give (R )-l-(4-((l-cyanoethyl)amino)-5- nitropyridin-2-yl)-1H -pyrazolo[3,4-b ]pyridine-5 -carbonitrile 6’ (1.5 g, 53.9% yield) as a yellow solid.

[0192] 1H NMR (400 MHz, DMSO-d6): 5 ppm 1.77 (d, J= 7.2 Hz, 3H), 5.21 - 5.25 (m, 1H), 7.86 (s, 1H), 8.53 (d, J= 7.6 Hz, 1H), 8.78 (s, 1H), 9.03 - 9.11 (m, 2H), 9.22 (s, 1H).

[0193] Step 4’. Preparation of ( R )- 1 -( 5-a m ino-4-(( 1 -cy a noct by 1 )a in in 0 )py rid in-2-y 1 )- 1 H - pyrazolo[3,4-b ]pyridine-5-carbonitrile 7’: To a stirred solution of (R)- 1 -(4-((l - cyanoethyl)amino)-5-nitropyridin-2-yl)-U / -pyrazolo[3,4-b ]pyridine-5-carbonitrile 6’ (1.5 g, 4.49 mmol) in 100 mL of ethanokwater (7:3) at RT, iron (1.253 g, 22.44 mmol) and ammonium chloride (0.600 g, 11.22 mmol) were added and stirred for 6 h at 85 °C. The reaction mixture was cooled to RT and filtered through celite bed, the bed was washed with DCM (500 mL) and concentrated under reduced pressure to give crude product. The crude product was dissolved in water and stirred for 10 min at RT. Then, the crude product was filtered through Buchner funnel, washed with water, and dried to afford (R )-l-(5-amino-4-((l-cyanoethyl)amino)pyridin-2-yl)-U / -pyrazolo[3,4- b ]pyridine-5-carbonitrile 7’ (1.2 g, 61.5% yield) as a brown solid, which was used in the next step without further purification.

[0194] UPLC method 2: retention time: 0.561 min, [M+H]+= 305.0.

[0195] Step 5’. Preparation of ( R )-l-(5-azido-4-(( 1-cyanoet hyl)am ino)pyr idin-2-yl )-1H - pyrazolo[3,4-b ]pyridine-5-carbonitrile 8’: To a stirred solution of (R)-l-(5-amino-4-((l- cyanoethyl)amino)pyridin-2-yl)-U7-pyrazolo[3,4-b ]pyridine-5-carbonitrile 7’ (1.2 g, 3.94 mmol, 1.0 eq.) in acetonitrile (20 mL), ADMP (2.81 g, 9.86 mmol, 2.5 eq.) and DMAP (1.204 g, 9.86 mmol, 2.5 eq.) were added at room temperature and stirred for 16 h at RT under nitrogen atmosphere. The reaction mixture was treated with ice-cold water (100 mL) and extracted with ethyl acetate (3 X 150 mL). The combined organic layer was dried over sodium sulphate, and then concentrated under reduced pressure to give ( R )-l-(5-azido-4-((l-cyanoethyl)amino)pyridin-2-yl)-1H -pyrazolo[3,4-b ]pyridine-5-carbonitrile 8’ (1.3 g, 56.8% yield) as a gummy liquid, which was used in the next step without further purification.

[0196] LCMS method 1 : retention time: 1.484 min, [M+H]+= 331.0.

[0197] Step 6’. Preparation of l-(4-(((R)-l-cyanoethyl)amino)-5-(4-((1r,4R)-4-(2- ((tetrahydro-2H-pyran-2-yl)oxy)ethyl)cyclohexyl)-lH-l,2,3-triazol-l-yl)pyridin-2-yl)-lH- pyrazolo[3,4-b]pyridine-5- carbonitrile 9’: To a stirred solution of (R)-l-(5-azido-4-((l- cyanoethyl)amino)pyridin-2-yl)-lH-pyrazolo[3,4-b]pyridine-5-carbonitrile 8’ (430 mg, 1.302 mmol) in acetone (6.0 mL) was added 2-(2-((1r,4r)-4-ethynylcyclohexyl)ethoxy)tetrahydro-2H- pyran L-l (338 mg, 1.432 mmol) followed by sodium ascorbate (129 mg, 0.651 mmol) and then a solution of copper(II) sulphate pentahydrate (65 mg, 0.26 mmol) in H2O (1.0 mL) was added at room temperature. The reaction mixture was stirred at room temperature for 16 h. The reaction mixture was treated with water (100 mL) and extracted with ethyl acetate (3 x 75 mL). The combined organic layer was washed with brine (10 mL), dried over sodium sulphate and concentrated under reduced pressure to give the crude product. The crude compound was purified by column chromatography using silica gel (230-400 mesh) with 80-90% ethyl acetate / pet ether to obtain l-(4-(((R)-l-cyanoethyl)amino)-5-(4-((1r,4R)-4-(2-((tetrahydro-2H-pyran-2- yl)oxy)ethyl)cyclohexyl)-lH-l,2,3-triazol-l-yl)pyridin-2-yl)-lH-pyrazolo[3,4-b]pyridine-5- carbonitrile 9’ (220 mg, 20.2% yield).

[0198] LCMS method 1 : retention time 3.078 min, 67.93% purity at 220 nm, [M+ H]+= 567.0.

[0199] Step 7’. Preparation of l-(4-(((R)-l-cyanoethyl)amino)-5-(4-((1r,4R)-4-(2- hydroxyethyl)cyclohexyl)-lH-l,2,3-triazol-l-yl)pyridin-2-yl)-lH-pyrazolo[3,4-b]pyridine-5- carbonitrile 10’: To a stirred solution of l-(4-(((R)-l-cyanoethyl)amino)-5-(4-((1r,4R)-4-(2- ((tetrahydro-2H-pyran-2-yl)oxy)ethyl)cyclohexyl)-lH- 1 ,2,3-triazol- 1 -yl)pyri din-2 -yl)- 1H- pyrazolo[3,4-b]pyridine-5-carbonitrile 9’ (220 mg, 0.388 mmol) in MeOH (4.0 mL), / ?TSA (73.8 mg, 0.388 mmol) was added at RT and stirred for 2 h. The reaction mixture was treated saturated sodium bicarbonate solution (30 mL) and extracted with DCM (2 x 40 mL). The organic layer was washed with brine (25.0 mL), dried over sodium sulphate, and concentrated under reduced pressure to give crude product l-(4-(((R)-l-cyanoethyl)amino)-5-(4-((1r,4R)-4-(2- hydroxyethyl)cyclohexyl)-lH-l,2,3-triazol-l-yl)pyridin-2-yl)-lH-pyrazolo[3,4-b]pyridine-5- carbonitrile 10’ (130 mg) as a brown solid, which was used in the next step without further pufi cation.

[0200] LCMS method 1 : retention time 1.994 min, [M+ H]+= 483.2.

[0201] Step 8’. Preparation of l-(4-(((R)-l-cyanoethyl) amino)-5-(4-((1r,4R)-4-(2-oxoethyl) cyclohexyl)-lH-l,2,3-triazol-l-yl) pyridin-2-yl)-lH-pyrazolo[3,4-b] pyridine-5-carbonitrile A-1: IBX (151 mg, 0.539 mmol, 2.0 eq.) was added to a solution of l-(4-(((R)-l -cyanoethyl) amino)- 5-(4-((1r,4R)-4-(2-hydroxyethyl) cyclohexyl)-lH-l,2,3-triazol-l-yl) pyridin-2-yl)-lH-pyrazolo[3,4- b] pyridine-5-carbonitrile 10’ (130 mg, 0.269 mmol, 1.0 eq.) in DMSO (3 mL) at RT. The resulting solution was stirred for 2 h at RT. The reaction mixture was diluted ethyl acetate (15 mL) and washed with ice-cold water, aqueous sodium bicarbonate solution followed by brine solution, dried over sodium sulphate, and concentrated under reduced pressure to give the crude product 1 -(4- (((R)-l -cyanoethyl) amino)-5-(4-((1r,4R)-4-(2-oxoethyl) cyclohexyl)-lH-l,2,3-triazol-l-yl) pyridin-2-yl)-lH-pyrazolo[3,4-b] pyridine-5-carbonitrile A-l (90 mg) as a brown solid, which was used in the next step without further purification.

[0202] LCMS method 1 : retention time: 2.297 min, [M+H]+= 481.0.Example S4: Preparation of 6-amino-l-(4-(((R)-l-cyanoethyl)amino)-5-(4-((1r,4R)-4-(2- oxoethyl)cyclohexyl)-lH-l,2,3-ti'iazol-l-yl)pyridin-2-yl)-lH-pyrazolo[3,4-b]pyridine-5- carbonitrile (A-2)

[0203] LCMS Method 1. Kinetex XB - C18, 75 x 3.0 mm, 2.6 pm, Temperature: RT, Flow: 1.0 mL / min, Run Time: 5 minutes, Mobile Phase Conditions: Mobile Phase-A: 5.0 mm Ammonium formate pH 3.3:CHsCN (98:02), Mobile Phase-B: CH3CN: 5.0 mm Ammonium formate pH 3.3 (98:02), Gradient: Initial 80% Mobile Phase A and 20% Mobile Phase B linear gradient to 100% Mobile Phase B for 4.0 min. MSD positive.

[0204] LCMS Method 2, Aquity Uplc BEH- C18, 50 x 3.0 mm, 1.7 pm, Temperature: RT, Flow: 1.0 mL / min, Run Time: 5 minutes, Mobile Phase Conditions: Mobile Phase-A: 0.1% TFA in H2O, Mobile Phase-B: 0.1% TFA in ACN, Gradient: Initial 80% Mobile Phase A and 20% Mobile Phase B linear gradient to 100% Mobile Phase B for 3.0 min. MSD positive.

[0205] Step 1’. Preparation of ( R )-2-((2-chloro-5-nitropyridin-4-yl)amino)propenamide 3’: To a stirred solution of 2,4-dichloro-5-nitropyridine 1’ (50.0 g, 259 mmol) in acetonitrile (500 mL)was added (A)-2-aminopropanamide hydrochloride 2’ (38.7 g, 311 mmol) and DIPEA (113.0 mL, 648 mmol) at RT under nitrogen atmosphere and the resulting solution was stirred for 2 h at 45 °C. The solution was then cooled to room temperature and concentrated under reduced pressure (70% volume). Finally, cold water (500 mL) was added to the reaction mixture and stirred for 10 min to afford a pale yellow solid. The solid was filtered, washed with water and dried to give the pure compound (R)-2-((2-chloro-5-nitropyridin-4-yl)amino)propanamide 3’ (55.0 g, 86% yield) as a pale yellow solid.

[0206] LCMS method 1 : retention time 0.88 min, 99.29% purity at 220 nm, [M+H]+= 245.0.

[0207] Step 2’. Preparation of (R )-2-((2-chloro-5-nitropyridin-4-yl)amino)propanenitrile 4’: To a stirred solution of (A)-2-((2-chloro-5-nitropyridin-4-yl) amino) propanamide 3’ (54.0 g, 221 mmol) in DCM (500 mL) at 0 °C was added pyridine (179.0 mL, 2207 mmol) slowly over 10 min. Then, TFAA (156.0 mL, 1104 mmol) was added in a drop-wise manner (over 30 min) at 0 °C and stirred at RT for 3 h. The reaction mixture was cooled to 0 °C and treated with ice-cold water (500 mL). The reaction mixture was extracted with DCM (3 x 300 mL), the combined organic layer was washed with 1.0 N HC1 (200 mL), followed by brine solution (500 mL), dried over sodium sulphate, and concentrated under reduced pressure to give the crude product. The crude product was washed with MTBE (3 x 200 mL) to afford a pale yellow solid and dried to give pure compound (R)-2-((2-chloro-5-nitropyridin-4-yl)amino)propanenitrile 4’ (46.0 g, 92% yield) as pale-yellow solid.

[0208] LCMS method 1 : retention time 1.75 min, 99.94% purity at 220 nm, [M- H]+= 225.0.

[0209] Step 3’. Preparation of ( R )-2-( ( 5-n it ro-2-( 1 H -py razolo |3.4-d ] pyrini id in- 1 - yl)pyridin-4-yl)amino)propanenitrile 6’: To a stirred solution of (A)-2-((2-chloro-5-nitropyridin- 4-yl) amino) propanenitrile 4’ (10.0 g, 44.1 mmol) in dioxane (80 mL) was added U / -pyrazolo[3,4- d\ pyrimidine 5’ (5.83 g, 48.5 mmol), zinc acetate (8.10 g, 44.1 mmol), and DIPEA (46.2 mL, 265 mmol) at RT under nitrogen atmosphere. Pd2(dba)s (2.02 g, 2.206 mmol) was added and the reaction mixture was degassed for another 15 min. The reaction mixture was stirred at 85 °C for 16 h. The reaction mixture was cooled to RT, filtered through a Celite bed, and concentrated under reduced pressure to give the crude product. The crude product was purified by flash column chromatography on silica with 10% methanol / DCM to afford compound (R)-2-((5-nitro-2-(lH- pyrazolo[3,4-d]pyrimidin-l-yl)pyridin-4-yl)amino)propanenitrile 6’ (7.0 g, 48.3% yield) as a yellow solid.

[0210] LCMS method 2: retention time 2.33 min, 94.01% purity at 220 nm, [M+ H]+= 311.

[0211] Step 4’. Preparation of (R )-2-((5-amino-2-( lH-pyrazolo[3,4-J]pyrimidin-l- yl)pyridin-4-yl)amino)propanenitrile 7’: To a stirred solution of (A)-2-((5-nitro-2-(U7-pyrazolo[3,4-d]pyrimidin-l-yl)pyridin-4-yl)amino)propanenitrile 6’ (7.0 g, 22.56 mmol) in 150 mL of ethanokwater (4: 1) at RT, iron (3.15 g, 56.4 mmol) and ammonium chloride (3.02 g, 56.4 mmol) were added and stirred for 4 h at 85 °C. The reaction mixture was cooled to RT, treated with ice- cold saturated sodium bicarbonate (50 mL) solution and extracted with 20% methanol in DCM (3 x 200 mL). The combined organic layer was dried over sodium sulphate and concentrated under reduced pressure to give the crude compound (R)-2-((5-amino-2-(lH-pyrazolo[3,4-d]pyrimidin-l- yl)pyridin-4-yl)amino)propanenitrile 7’ (3.0 g), which was used in the next step without further purification.

[0212] LCMS method 1 : retention time 2.20 min, [M+ H]+= 281.2

[0213] Step 5’. Preparation of (R )-2-((5-:izido-2-(1H -pyrazolo|3.4-7|pyrimidin-l- yl)pyridin-4-yl)amino)propanenitrile 8’: To a stirred solution of (R)-2-((5-amino-2-(177- pyrazolo[3,4-d] pyrimidin-l-yl) pyridin-4-yl) amino) propane nitrile 7’ (3.0 g, 10.70 mmol) in acetonitrile (30 mL), ADMP (2.195 g, 26.8 mmol) and DMAP (1.961 g, 16.05 mmol) were added at room temperature and stirred for 16 h under nitrogen atmosphere. The reaction mixture was treated with ice-cold water and extracted with ethyl acetate (3 x 100 mL). The combined organic layer was washed with brine, dried over sodium sulphate and then concentrated under reduced pressure to give compound (R )-2-((5-azido-2-(lJ / -pyrazolo[3,4-J]pyrimidin-l-yl)pyridin-4- yl)amino)propanenitrile 8’ (4.0 g), which was directly used in the next step without further purification.

[0214] LCMS method 1 : retention time 1.15 min, [M+ H]+= 305.

[0215] Step 6’. Preparation of (2R )-2-((2-(1H -pyrazolo|3.4-7|pyrimidin-l-yl)-5-(4- ((1r,4R )-4-(2-((tetrahydro-2Z / -pyran-2-yl)oxy)ethyl)cyclohexyl)-lZ / -l,2,3-triazol-l-yl)pyridin- 4-yl)amino)propanenitrile 9’: To a stirred solution of (R )-2-((5-azido-2-(lJ / -pyrazolo[3,4-J] pyrimidin-l-yl) pyridin-4-yl) amino) propane nitrile 8’ (4.0 g, 13.06 mmol) in 55.0 mL of acetone:water (11 : 1), 2-(2-((1r,4r)-4-ethynylcyclohexyl)ethoxy)tetrahydro-2H-pyran L-l (3.40 g, 14.37 mmol), sodium ascorbate (0.536 g, 6.53 mmol) and copper(II) sulphate pentahydrate (1.630 g, 6.53 mmol) were added at room temperature. The reaction mixture was stirred at room temperature for 16 h. The reaction mixture was treated with water and extracted with ethyl acetate (3 x 40 mL). The combined organic layer was dried over sodium sulphate and concentrated under reduced pressure to give the crude product. The crude product was purified by column chromatography using silica gel (230-400 mesh) and eluted with 70-90% ethyl acetate in pet ether to give the compound (2R)-2-((2-(lH-pyrazolo[3,4-d]pyrimidin-l-yl)-5-(4-((1r,4R)-4-(2- ((tetrahydro-2H-pyran-2-yl)oxy)ethyl)cyclohexyl)-lH-l,2,3-triazol-l-yl)pyridin-4- yl)amino)propanenitrile 9’ (1.6 g, 18.69% yield) as a yellow solid.

[0216] LCMS method 1 : retention time 2.72 min, 82.87% purity at 220 nm, [M+ H]+= 543.2.

[0217] Step 7’. Preparation of 6-amino-l-(4-(((R )-l-cyanoethyl)amino)-5-(4-((1r,4R )-4-(2- ((tetrahydro-2H -pyran-2-yl)oxy)ethyl)cyclohexyl)-lH / -l .2.3-triazol-l-yl)pyridin-2-yl)-1H - pyrazolo[3,4-b]pyridine-5-carbonitrile 10’: To a stirred solution of (2A)-2-((2-(lH-pyrazolo[3,4- d\ pyrimidin-l-yl)-5-(4-((1r,4A)-4-(2-((tetrahydro-2J / -pyran-2-yl) oxy) ethyl) cyclohexyl)- 177- 1,2, 3 -triazol- 1-yl) pyridin-4-yl) amino) propane nitrile 9’ (1.1 g, 2.027 mmol) in 2-propanol (10 mL), DIPEA (2.83 mL, 16.22 mmol) was added at RT. The reaction mixture was warmed up to 80 °C and stirred for 10 min, and then malononitrile (0.453 mL, 10.14 mmol) was added at the same temperature and stirred for another for 6 h at 80 °C. The reaction mixture was cooled to RT, filtered, the solid was washed with 2-propanol (2 x 10 mL) and dried to give the crude compound 6-amino-l-(4-(((R)-l-cyanoethyl)amino)-5-(4-((1r,4R)-4-(2-((tetrahydro-2H-pyran-2- yl)oxy)ethyl)cyclohexyl)-lH-l,2,3-triazol-l-yl)pyridin-2-yl)-lH-pyrazolo[3,4-b]pyridine-5- carbonitrile 10’ (1.0 g), which was directly used in the next step without further purification.

[0218] LCMS method 1 : retention time 2.23 min, [M+H]+= 582.2.

[0219] Step 8’. Preparation of 6-amino-l-(4-(((R )-l-cyanoethyl)amino)-5-(4-((1r,4R )-4-(2- hydroxyethyl)cyclohexyl)-1H -l .2.3-tri:izol-l-yl)pyridin-2-yl)-1H -pyra zolo|3.4-b |pyridine-5- carbonitrile 11’: To a stirred solution of 6-amino-l-(4-(((R )-l -cyanoethyl) amino)-5-(4-((1r,4R )- 4-(2-((tetrahydro-2H -pyran-2-yl) oxy) ethyl) cyclohexyl)-1H -l,2,3-triazol-l-yl) pyridin-2-yl)-17T- pyrazolo[3,4-b ] pyridine-5-carbonitrile 10’ (800 mg, 1.375 mmol) in MeOH, p TSA (131 mg, 0.688 mmol) was added at RT and stirred for 1 h. The reaction mixture was diluted with 20% methanol in DCM (50 mL) and treated with saturated sodium bicarbonate solution (20 mL). The organic layers were collected, washed with brine, dried over sodium sulphate, and concentrated under reduced pressure to give the crude product. The crude was washed with MTBE (3 x 20 mL) to give the compound 6-amino-l-(4-(((R )-l-cyanoethyl)amino)-5-(4-((1r,4R )-4-(2- hydroxyethyl)cyclohexyl)-1H -l,2,3-triazol-l-yl)pyridin-2-yl)-1H -pyrazolo[3,4-b ]pyridine-5- carbonitrile 11’ (450 mg, 61.1% yield).

[0220] Step 9’. Preparation of 6-amino-l-(4-(((R )-l-cyanoethyl)amino)-5-(4-((1r,4R )-4-(2- oxoethyl)cyclohexyl)-l H-l ,2,3-triazol-l-yl)pyridin-2-yl)-lH-pyrazolo[3,4-b]pyridine-5- carbonitrile A-2: To a stirred solution of 6-amino-l-(4-(((R )-l-cyanoethyl)amino)-5-(4-((1r,4R )- 4-(2-hydroxyethyl)cyclohexyl)-1H -l,2,3-triazol-l-yl)pyridin-2-yl)-1H -pyrazolo[3,4-b ]pyridine-5- carbonitrile 11’ (450 mg, 0.904 mmol) in DMSO (5 mL), IBX (760 mg, 2.71 mmol) was added at RT and stirred for 2 h. The reaction mixture was quenched with sodium bicarbonate solution and extracted with ethyl acetate (3x 20 mL). The combined organic layer was washed with brine, dried over sodium sulphate, and concentrated under reduced pressure to give the compound 6-amino-l-(4-(((R )- 1 -cyanoethyl)amino)-5-(4-((1r,4R )-4-(2-oxoethyl)cyclohexyl)- 1H- 1 ,2,3 -tri azol- 1 - yl)pyridin-2-yl)-lJ7-pyrazolo[3,4-b ]pyridine-5-carbonitrile A-2 (400 mg), which was used in the next step without further purification.Example S5: Preparation of 6-(5-cyano-lH-pyrazolo[3,4-b]pyridin-l-yl)-4-(((R)-l- cyanoethyl)amino)-N-((1r,4R)-4-(2-oxoethyl)cyclohexyl)nicotinamide (A-3)

[0221] LCMS Method 1. Column: Luna C18 (2) 50 X 3 mm, 3 um. Temperature: 45 °C, Flow: 1.5 mL / min, run time: 2.5 min. Mobile phase conditions: Initial 95 % H2O 0.1 % FA / 5 % MeCN 0.1 % FA, linear gradient to 95 % MeCN 0.1 % FA over 1.3 min then hold for 1.2 minute at 95 % MeCN 0.1 % FA. MSD: ESI Positive.

[0222] LCMS Method 2, Kinetex XB - C18, 75 x 3.0 mm, 2.6 pm. Temperature: RT, Flow: 1.0 mL / min, run time: 5.0 min. Mobile Phase Conditions: Mobile Phase-A: 5.0 mm ammonium formate pH 3.3 in CH3CN (98:02), Mobile Phase-B: CH3CN in 5.0 mm ammonium formate (98:02), Gradient: Initial 80% Mobile Phase A and 20% Mobile Phase B linear gradient to 100% Mobile Phase B for 4.6 min. MSD positive.

[0223] Step 1’. Preparation of Tert-butyl 6-chloro-4-[[(lR)-2-amino-l-methyl-2-oxo- ethyl]amino]pyridine-3-carboxylate 3’: To a solution of (2R)-2-aminopropanamide hydrochloride 2’ (1.5 g, 12.091 mmol, 3.0 eq.) and tert-butyl 4,6-dichloropyridine-3-carboxylate 1’ (1.0 g, 4.030 mmol, 1.0 eq.) in MeCN (20 mL, 0.2M) under nitrogen was added DIPEA (2.81 mL, 16.122 mmol, 4.0 eq.). The reaction was stirred at 65 °C for 96 hours. The reaction mixture was cooled down to room temperature. Acetonitrile was removed under high vacuum. DCM and water were added, then the phases were separated. The aqueous phase was extracted 3 times with DCM.The combined organic phases were washed once with brine, dried over magnesium sulfate, filtered and concentrated. The residue was purified by normal phase flash chromatography (40 g silica column, elution: 0 to 10 % CH2CI2 / CH3OH over 15 CV, product exited at 5 % CH3OH). Fractions were combined and concentrated to give tert-butyl 6-chloro-4-[[(lR)-2-amino-l-methyl-2-oxo- ethyl]amino]pyridine-3-carboxylate 3’ (1 g, 83 % yield) as a white solid.

[0224] LCMS method 1 : retention time: 1.601 min, 99.9 % purity at 215 nm, [M+H]+= 300.2.

[0225] 1H NMR (400 MHz, DMSO-d6): 8 ppm 1.36 (d, J= 6.8 Hz, 3 H), 1.55 (s, 9 H), 4.15 (t, J= 6.8 Hz, 1 H), 6.57 (s, 1 H), 7.31 (br s, 1 H), 7.62 (br s, 1 H), 8.44 (d, J= 6.8 Hz, 1 H), 8.50 (s, 1 H).

[0226] Step 2’. Preparation of tert-butyl 6-chloro-4-[[(lR)-l-cyanoethyl]amino]pyridine-3- carboxylate 4’: To a solution of tert-butyl 6-chloro-4-[[(lR)-2-amino-l-methyl-2-oxo- ethyl]amino]pyridine-3-carboxylate 3’ (1.0 g, 3.336 mmol, 1.0 eq.) and pyridine (2.7 mL, 33.361 mmol, 10.0 eq.) in DCM (19.0 mL) was added TFAA (0.9 mL, 6.672 mmol, 2.0 eq.) in DCM (11.4 mL, 0.1 M) dropwise at -15 °C. The resulting mixture was allowed to warm to room temperature and stirred for 2.5 hours. The mixture was washed with 1 M HC1 (3x30 mL), saturated sodium bicarbonate (1x40 mL) and brine (1x40 mL). The organic layer was dried over lSfeSCL, filtered and concentrated. The residue was purified by normal phase flash chromatography (40 g silica column, elution: 0 to 5 % CH2Q2 / CH3OH over 15 CV, product exited at 1.0 % CH3OH). Fractions were combined and concentrated to give tert-butyl 6-chloro-4-[[(lR)-l-cyanoethyl]amino]pyridine-3- carboxylate 4’ (0.76 g, 84 % yield) as a white solid.

[0227] LCMS method 1 : retention time: 1.833 min, 98.7 % purity at 215 nm, [M+H]+= 282.2.

[0228] 1H NMR (400 MHz, DMSO-d6): 6 ppm 1.56 (s, 9 H), 1.66 (d, J= 7.1 Hz, 3 H), 5.01(quin, J= 7.1 Hz, 1 H), 7.06 (s, 1 H), 8.19 (d, J= 7.6 Hz, 1 H), 8.59 (s, 1 H).

[0229] Step 3’. Preparation of tert-butyl 6-(5-cyanopyrazolo[3,4-b]pyridin-l-yl)-4-[[(lR)- 1-cyanoethyl] amino] pyridine-3-carboxylate 6’: To a dried sealed tube was added tert-butyl 6- chloro-4-[[(lR)-l-cyanoethyl]amino]pyridine-3-carboxylate 4’ (760.0 mg, 2.697 mmol, 1.0 eq.), lH-pyrazolo[3,4-b]pyridine-5-carbonitrile 5’ (388.8 mg, 2.697 mmol, 1.0 eq.), XantPhos (0.234 g, 0.404 mmol, 0.15 eq.), Pd2(dba)3'CHC13 (418.8 mg, 0.404 mmol, 0.15 eq.), and Zn(OAc)2 (494.9 mg, 2.697 mmol, 1.0 eq.) in 1,4-dioxane (9.0 mL, 0.3 M). Then nitrogen was sparged through the mixture for 15 minutes. The tube was sealed and the mixture was stirred at 105 °C for 16 hours.The reaction was filtered over Celite, washed with DCM and the filtrate was evaporated. The residue was purified by normal phase flash chromatography (40 g silica column, elution: 0 to 10 % DCM / MeOH over 15 CV, product exited at 5 % MeOH). Fractions from the major peak were combined and concentrated under reduced pressure to afford tert-butyl 6-(5-cyanopyrazolo[3,4-b]pyridin-l-yl)-4-[[(lR)-l-cyanoethyl]amino]pyridine-3-carboxylate 6’ (906 mg, 77 % yield) as an orange solid. Contaminated with XantPhosCh.

[0230] LCMS method 1 : retention time: 1.781 min, 88 % purity at 215 nm, [M+H]+= 390.2.

[0231] 1H NMR (400 MHz, DMSO-d6): 8 ppm 1.60 (s, 9 H), 1.72 (br d, J= 6.8 Hz, 3 H), 4.99 -5.10 (m, 1 H), 7.60 (s, 1 H), 8.30 (br d, J= 7.1 Hz, 1 H), 8.71 (s, 1 H), 8.87 (s, 1 H), 9.05 (dd, J = 12.1, 1.8 Hz, 1 H), 11.96 (br s, 1 H).

[0232] Step 4’. Preparation of 6-(5-Cyanopyrazolo[3,4-6]pyridin-l-yl)-4-[[(lR )-l- cyanoethyl] amino] pyridine-3-carboxylic acid 7’: To a solution of tert-butyl 6-(5- cyanopyrazolo[3,4-b ]pyridin-l-yl)-4-[[(lA)-l-cyanoethyl]amino]pyridine-3-carboxylate 6’ (1.17 g, 3.025 mmol, 1.0 eq.) in DCM (8.9 mL, 0.3 M) was added TFA (11.6 mL, 45.376 mmol, 15 eq.). The reaction was stirred at room temperature for 16 hours. The solvent was removed in vacuo and the residue was co-evaporated with PhMe (3 times) followed by MeCN (2 times), which afforded a yellow semi-solid to which was added H2O (10 mL) / MeCN (2 mL) / EtOAc (2 mL) and the mixture was sonicated 3-5 minutes. The solvent was evaporated almost until dry, a precipitate formed which was filtered and washed with MTBE, which afforded 182 mg of 7’. The analysis of the remaining solution showed a mixture of 7’ and by-products. The product was purified by reverse-phase chromatography column using a 50G Cl 8 column (elution: 5 % MeOH / 0.1 % HCOOH over 4 CV, then 5 % to 75 % MeOH / 0.1 % HCOOH over 15 CV, product exited at 48 % MeOH). The pure fractions were combined and concentrated under reduced pressure to afford 6-(5- cyanopyrazolo[3,4-b ]pyridin-l-yl)-4-[[(lA)-l-cyanoethyl]amino]pyridine-3-carboxylic acid 7’ (315 mg, 29 % yield) as a white solid.

[0233] LCMS method 1 : retention time: 1.372 min, 99.9 % purity at 215 nm, [M+H]+= 334.2.

[0234] 1H NMR (400 MHz, DMSO-d6): 6 ppm 1.71 (d, J= 6.8 Hz, 3 H), 4.99 - 5.10 (m, 1 H),7.59 (s, 1 H), 8.55 (br d, J= 7.6 Hz, 1 H), 8.72 (s, 1 H), 8.88 (s, 1 H), 9.04 (d, J= 2.0 Hz, 1 H), 9.07 (d, J= 2.0 Hz, 1 H), 13.63 (br s, 1 H).

[0235] Step 5’. Preparation of 6-(5-cyano-lH-pyrazolo[3,4-b]pyridin-l-yl)-4-(((R)-l- cyanoethyl)amino)-N-((1r,4R)-4-(2-hydroxyethyl)cyclohexyl)nicotinamide 8’: To a stirred solution of (R)-6-(5-cyano-lH-pyrazolo[3,4-b] pyridin-l-yl)-4-((l -cyanoethyl) amino) nicotinic acid 7’ (3.50 g, 10.50 mmol) in DCM (30 mL) at room temperature under nitrogen atmosphere was added HATU (5.99 g, 15.75 mmol) and DIPEA (9.17 mL, 52.5 mmol). The reaction mixture was stirred for 10 minutes under nitrogen atmosphere at room temperature and 2-((1r,4r)-4- aminocyclohexyl)ethan-l-ol L-3 (2.256 g, 15.75 mmol) was added and the reaction mixture was stirred at RT for 8 h. The reaction mixture was concentrated under reduced pressure and the crude product was treated with cold water (250 mL) and extracted with ethyl acetate (2 x 300 mL). Theorganic layer was washed with brine solution, and the combined organic layers were dried over anhydrous sodium sulphate, filtered and concentrated under reduced pressure. The crude product was purified by 60-120 mesh silica gel column chromatography and the desired product was eluted by 40% of ethyl acetate / pet ether to obtain 6-(5-cyano-lH-pyrazolo[3,4-b] pyridin-1- yl)-4-(((R)-l- cyanoethyl) amino)-N-((1r,4R)-4-(2-hydroxy ethyl) cyclohexyl)nicotinamide 8’ (4.62 g).

[0236] LCMS method 1 : retention time: 2.309 min, [M+H]+= 459.2

[0237] Step 6’. Preparation of 6-(5-cyano-lH-pyrazolo[3,4-b]pyridin-l-yl)-4-(((R)-l- cyanoethyl)amino)-N-((1r,4R)-4-(2-oxoethyl)cyclohexyl)nicotinamide A-3: To a stirred solution of 6-(5-cyano-lH-pyrazolo[3,4-b]70yridine-l-yl)-4-(((R)-l-cyanoethyl)amino)-N-((1r,4R)-4-(2- hydroxyethyl)cyclohexyl)nicotinamide 8’ (500 mg, 0.916 mmol) in DMSO (10 mL) was added IBX (1710 mg, 2.75 mmol) portion wise at room temperature under nitrogen atmosphere, then the reaction mixture was stirred at room temperature for 4 h. The reaction mixture was quenched with saturated sodium bicarbonate solution and extracted with ethyl acetate (3 x 100 mL). The combined organic layers were dried over sodium sulphate, filtered, and concentrated under reduced pressure to obtain 6-(5-cyano-lH-pyrazolo[3,4-b]pyridin-l-yl)-4-((R)-l-cyanoethyl)amino)-N-((1r,4R)-4-(2- oxoethyl)cyclohexyl)nicotinamide A-3 (330.0 mg) as a pale yellow solid. The crude product was used in the next step without further purification.

[0238] LCMS method 2: retention time: 1.940 min, [M+H]+= 457.2

[0239] 1H NMR (400 MHz, CDC13): 8 ppm 1.06 - 1.19 (m, 2H), 1.33 - 1.47 (m, 2H), 1.67 (d, J = 7.2 Hz, 3H), 1.72 - 1.97 (m, 4H), 2.32 - 2.39 (m, 2H), 3.70 - 3.81 (m, 1H), 4.90 - 5.00 (m, 1H), 7.48 (s, 1H), 8.52 - 8.68 (m, 1H), 8.69 - 8.85 (m, 3H), 9.01 - 9.11 (m, 2H), 9.67 - 9.72 (m, 1H).Example S6: Preparation of 5-chloro-6-((4-(((R)-l-cyanoethyl)amino)-5-(4-((1r,4R)-4-(2- oxoethyl)cyclohexyl)-lH-l,2,3-triazol-l-yl)pyridin-2-yl)amino)nicotinonitrile (A-4)

[0240] LCMS Method 1. Kinetex XB - C18, 50 x 4.6 mm, 5.0 pm. Temperature: RT, Flow: 1.0mL / min, run time: 5.5 min. Mobile Phase Conditions: Mobile Phase-A: 0.1% TFA in H2O, Mobile Phase-B: 0.1% TFA in ACN, Gradient: Initial 95% Mobile Phase A and 5% Mobile Phase B linear gradient to 95% Mobile Phase B for 2.5 min. MSD positive.

[0241] LCMS Method 2, Kinetex XB - C18, 75 x 3.0 mm, 2.6 pm. Temperature: RT, Flow: 1.0 mL / min, run time: 5.0 min. Mobile Phase Conditions: Mobile Phase-A: 5.0 mm Ammonium formate pH 3.3:CH3CN (98:02), Mobile Phase-B: CH3CN: 5.0 mm Ammonium formate pH 3.3 (98:02), Gradient: Initial 98% Mobile Phase A and 2% Mobile Phase B linear gradient to 100% Mobile Phase B for 4 min. MSD positive.

[0242] Step 12 Preparation of (R)-5-chloro-6-((4-((l-cyanoethyl)amino)-5-nitropyridin-2- yl)amino)nicotinonitrile 3’: To a stirred solution of (R)-2-((2-chloro-5-nitropyridin-4- yl)amino)propanenitrile 1’ (2.5 g, 8.60 mmol) and 6-amino-5-chloronicotinonitrile 2’ (1.32 g, 8.60 mmol) in 1,4-dioxane (25 mL) was added Pd2(dba)3 (0.315 g, 0.344 mmol) and purged with nitrogen for 5 min. Xantphos (0.398 g, 0.688 mmol) followed by zinc acetate (0.474 g, 2.58 mmol) and K2CO3 (2.97 g, 21.5 mmol) were added to the reaction mixture and purged with nitrogen for 2 min. The reaction mixture was stirred at 100 °C for 16 h under nitrogen atmosphere. Then, the reaction mixture was filtered through a Celite bed, and the Celite bed was washed with 5% MeOH / DCM (500 mL). The filtrate was then transferred into a separating funnel containing water (400 mL) and extracted using 5% MeOH / DCM (2 X 250 mL). The combined organic extracts were washed with brine (100 mL), dried over sodium sulfate, filtered, and concentrated under reduced pressure. The obtained solid was washed with ethyl acetate (50 mL), filtered, and dried under vacuum to obtain (R)-5-chloro-6-((4-((l-cyanoethyl)amino)-5-nitropyridin-2- yl)amino)nicotinonitrile 3’ (2.13 g, 60.6% yield) as a brown solid. It was used in the next step without further purification.

[0243] LCMS method 1 : retention time: 2.59 min, [M+H]+= 344.1.

[0244] 1H NMR (400 MHz, DMSO-d6): 8 ppm 1.74 (d, J= 6.8 Hz, 3H), 4.96 - 5.04 (m, 1H), 7.78 (s, 1H), 8.34 (d, J= 7.2 Hz, 1H), 8.54 (d, J= 1.6 Hz, 1H), 8.71 (d, J= 1.6 Hz, 1H), 9.00 (s, 1H), 9.58 (brs, 1H).

[0245] Step 2’. Preparation of (R)-6-((5-amino-4-((l-cyanoethyl)amino)pyridin-2- yl)amino)-5-chloronicotinonitrile 4’: To a stirred solution of (A)-5-chloro-6-((4-((l- cyanoethyl)amino)-5-nitropyridin-2-yl)amino)nicotinonitrile 3’ (2.10 g, 5.14 mmol) in 1,4-dioxane (20 mL) was added 10% palladium on carbon (0.547 g). The resulting reaction mixture was stirred under hydrogen atmosphere at room temperature for 72 h. The reaction mixture was filtered through a Celite bed, and the Celite bed was washed with 5% MeOH / DCM (150 mL). The filtrate was evaporated under reduced pressure to obtain (R)-6-((5-amino-4-((l-cyanoethyl)amino)pyridin-2-yl)amino)-5-chloronicotinonitrile 4’ (1.8 g, 76% yield) as a brown solid, which was used in the next step without further purification.

[0246] LCMS method 1 : retention time: 1.92 min, [M+H]+= 314.1.

[0247] Step 32 Preparation of (R)-6-((5-azido-4-((l-cyanoethyl)amino)pyridin-2- yl)amino)-5-chloronicotinonitrile 5’: To a stirred solution of (R)-6-((5-amino-4-((l- cyanoethyl)amino)pyridin-2-yl)amino)-5-chloronicotinonitrile 4’ (500 mg, 1.08 mmol) and ADMP (649 mg, 2.16 mmol) in acetonitrile (10 mL) was added DMF (2.5 mL). Then, DMAP (264 mg, 2.161 mmol) was added to the reaction mixture, and the reaction mixture was stirred at room temperature for 16 h. Then, the reaction mixture was transferred into a conical flask containing ice, and the solution was stirred at room temperature for 30 min, which produced a brown solid. The solid was filtered, washed with ice-cold water, and evaporated to dryness to obtain (R)-6-((5-azido- 4-((l-cyanoethyl)amino)pyri din-2- yl)amino)-5-chloronicotinonitrile 5’ (513 mg, 0.907 mmol, 84% yield) as a brown solid. It was used in the next step without further purification.

[0248] LCMS method 2: retention time: 1.742 min, [M+H]+= 340.0.

[0249] Step 4’. Preparation of l-(4-(((R)-l-cyanoethyl)amino)-5-(4-((1r,4R)-4-(2- ((tetrahydro-2H-pyran-2-yl)oxy)ethyl)cyclohexyl)-lH-l,2,3-triazol-l-yl)72yridine-2-yl)-lH- pyrazolo[3,4-b]pyridine-5-carboni-trile 6’: To a stirred solution of ((R)-6-((5-azido-4-((l- cyanoethyl)amino)pyridin-2-yl)amino)-5-chloronicotinonitrile 5’ (360 mg, 0.721 mmol) in acetone (5.0 mL) was added 2-((1r,4r)-4-ethynylcyclohexyl)ethan-l-ol 2’ (110 mg, 0.721 mmol), followed by sodium ascorbate (71.4 mg, 0.360 mmol). A solution of copper(II) sulphate pentahydrate (65.4 mg 0.360 mmol) in H2O (0.556 mL) was added to the reaction mixture at room temperature. After being stirred at room temperature for 2 h, the reaction mixture was concentrated using a rota evaporator and the residue was diluted with water (10 mL) and stirred at room temperature for 15 min. The brown solid was filtered, washed with water, and evaporated to dryness to obtain 5- chloro-6-((4-(((R)-l-cyanoethyl)amino)-5-(4-((1r,4R)-4-(2-hydroxyethyl)cyclohexyl)-lH-l,2,3- triazol-l-yl)pyri din-2 -yl)amino)nicotinonitrile 6’ (350 mg) as a brown solid. The crude product was used in the next step without further purification.

[0250] LCMS method 1 : retention time 2.037 min, [M+ H]+= 492.3.

[0251] Step 5’. 5-chloro-6-((4-(((R)-l-cyanoethyl)amino)-5-(4-((1r,4R)-4-(2- oxoethyl)cyclohexyl)-lH-l,2,3-triazol-l-yl)pyridin-2-yl)amino)nicotinonitrile A-4: To a stirred solution of 5-chloro-6-((4-(((R)-l -cyanoethyl)amino)-5-(4-((1r,4R)-4-(2 -hydroxy ethyl)cy cl ohexyl)- lH-l,2,3-triazol-l-yl)pyridin-2-yl)amino)nicotinonitrile 6’ (600 mg, 0.549 mmol) in DMSO (4 mL) under nitrogen atmosphere at room temperature was added IBX (384 mg, 1.372 mmol) and the reaction mixture was stirred for 4 h at room temperature. The reaction mass was treated with 10%NaHCCL solution (30 mL) and stirred at room temperature for 15 min. The brown solid was filtered, washed with water, and evaporated to dryness to obtain 5 -chloro-6-((4-(((R)-l -cyanoethyl) amino)-5-(4-((1r,4R)-4-(2-oxoethyl)cyclohexyl)-lH-l,2,3-triazol-l-yl)pyridin-2- yl)amino)nicotinonitrile A-4 (600 mg) as a brown solid. The crude product was used in the next step without further purification.

[0252] LCMS method 2: retention time: 2.748 min, [M+H]+= 490.0.Procedures for CBM SynthesisExample S7: Preparation of 6-(piperazin-l-yl)-2H-spiro[benzofuran-3,3'-piperidine]-2',6'-

[0253] LCMS Method 1. Kinetex XB - C18, 75 x 3.0 mm, 2.6 pm. Temperature: RT, Flow: 1.0 mL / min, run time: 4.7 min. Mobile Phase Conditions: Mobile Phase-A: 5.0 mm ammonium foramte pH 3.3 in CH3CN (98:02), Mobile Phase-B: CH3CN in 5.0 mm ammonium foramte (98:02), Gradient: Initial 80% Mobile Phase A and 20% Mobile Phase B linear gradient to 100% Mobile Phase B for 4.6 min. MSD positive.

[0254] LCMS Method 2, Kinetex XB - C18, 50 x 4.6 mm, 5.0 pm. Temperature: RT, Flow: 1.0 mL / min, run time: 5.5 min. Mobile Phase Conditions: Mobile Phase-A: 0.1% TFA in H2O, Mobile Phase-B: 0.1% TFA in ACN, Gradient: Initial 95% Mobile Phase A and 5% Mobile Phase B linear gradient to 95% Mobile Phase B for 2.5 min. MSD positive.

[0255] Step 1’. Preparation of ethyl 6-bromobenzofuran-3-carboxylate 2’: To a stirred solution of 4-bromo-2-hydroxybenzaldehyde 1’ (8.5 g, 42.3 mmol) in CH2CI2 (1.0 mL) was added tetrafluoroboric acid diethyl ether complex (0.572 mL, 4.23 mmol), followed by ethyl diazoacetatesolution in toluene (27.0 mL, 42.3 mmol) dropwise to the reaction mixture, and the reaction was not allowed to go above 38 °C. Once gas evolution ceased, the reaction mixture was concentrated using a rotary evaporator and H2SO4 (0.3 to 0.5 mL) was added to the mixture while stirring. After 5 to 10 min, the mixture was diluted with CH2Q2 (5 to 10 mL) and the H2SO4 was quenched with saturated NaHCOs solution. The reaction mixture was extracted with DCM, the combined organic layers dried over sodium sulphate, filtered, and concentrated, and the crude compound was purified by column chromatography using silica gel (230-400 mesh) with 0 - 5% ethyl acetate / pet ether to obtain ethyl 6-bromobenzofuran-3 -carboxylate 2’ (5.5 g, 48.3% yield) as a pale yellow solid.

[0256] 1H NMR (400 MHz, CDCI3): 8 ppm 1.44 (t, J= 7.2 Hz, 3H), 4.43 (q, J= 7.2 Hz, 2H), 7.50 (dd, J= 8.2 Hz and 1.6 Hz, 1H), 7.74 (d, J= 1.6 Hz, 1H), 7.95 (d, J= 8.4 Hz, 1H), 8.24 (s, 1H).

[0257] Step 2’. Preparation of tert-butyl 4-(3-(ethoxycarbonyl)benzofuran-6- yl)piperazine-l-carboxylate 4’: To a stirred solution of a mixture of ethyl 6-bromobenzofuran-3- carboxylate, 2’ (3.0 g, 11.15 mmol) in DMF (90 mL) was added tert-butyl piperazine- 1 -carboxylate 3’ (2.492 g, 13.38 mmol) followed by CS2CO3 (7.26 g, 22.30 mmol). The resulting reaction mixture was degassed for 5 min with nitrogen gas, and then RuPhos Pd G3 (0.466 g, 0.557 mmol) was added. The reaction mixture was heated to 90 °C and stirred for 6 h in a sealed glass vial. The reaction mixture was cooled to room temperature and concentrated under reduced pressure to obtain the crude product. The crude product was then purified by silica gel column chromatography with 15 - 20% ethyl acetate in petroleum ether to give tert-butyl 4-(3-(ethoxycarbonyl)benzofuran- 6-yl)piperazine-l -carboxylate, 4’ (1.8 g, 4.52 mmol, 40.5% yield) as a pale yellow solid.

[0258] LCMS Method 2: Retention time: 2.837 min, [M+H]+= 374.4 .

[0259] Step 3’. Preparation of tert-butyl 4-(3-(methoxycarbonyl)-2,3-dihydrobenzofuran- 6-yl)piperazine-l-Carboxylate 5’: To a stirred solution of tert-butyl 4-(3- (ethoxycarbonyl)benzofuran-6-yl)piperazine-l -carboxylate, 4’ (600 mg, 1.506 mmol) in MeOH (40 mL) was added magnesium (366 mg, 15.06 mmol) portion wise at 0 - 20°C under Nitrogen atmosphere. The reaction mixture was then allowed to warm at RT and stirred for 48 h. The reaction mixture was quenched with aqueous ammonium chloride (100 mL) and concentrated to remove methanol. The obtained crude product was then extracted with ethyl acetate (3 x 100 mL). The separated organic layer was washed with brine (40 mL), dried over anhydrous sodium sulfate, and filtered. The filtrate was concentrated under reduced pressure to give the crude product. The obtained crude product was purified by column chromatography with 13 - 15% ethyl acetate in petroleum ether to give tert-butyl 4-(3 -(methoxy carbonyl)-2, 3 -dihy drobenzofuran-6-yl)piperazine- 1 -carboxylate, 5’ (1.45 g, 3.93 mmol, 87% yield) as a white solid.

[0260] LCMS Method 2: Retention time: 2.55 min, [M+H]+= 363.2.

[0261] Step 4’. Preparation of tert-butyl 4-(3-(2-cyanoethyl)-3-(methoxycarbonyl)-2,3- dihydrobenzofuran-6-yl)piperazine-l-carboxylate 6’: To a stirred solution of tert-butyl 4-(3- (methoxycarbonyl)-2,3-dihydrobenzofuran-6-yl)piperazine-l-carboxylate, 5’ (1.450 g, 3.92 mmol) and K2CO3 (1.084 g, 7.84 mmol) in toluene (40 mL) was added benzyltriethylammonium chloride (0.089 g, 0.392 mmol) followed by acrylonitrile (0.416 g, 7.84 mmol) at RT. The reaction mixture was then heated to 90 °C and stirred for 3 h. The reaction mixture was quenched with water (50 mL) and extracted with ethyl acetate (3 x 50.0 mL). The combined organic layer was then dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to give the compound tert-butyl 4-(3-(2-cyanoethyl)-3-(methoxycarbonyl)-2,3-dihydrobenzofuran-6- yl)piperazine-l -carboxylate, 6’ (1.7 g, 3.56 mmol, 91 % yield) as a thick brown liquid. The product was used in the next step without further purification.

[0262] LCMS Method 2: Retention time: 2.615 min, [M+H]+= 416.3.

[0263] Step 5’. Preparation of 6-(piperazin-l-yl)-2H-spiro[benzofuran-3,3'-piperidine]- 2',6'-dione, 2 AcOH 7’: To a stirred solution of tert-butyl 4-(3-(2-cyanoethyl)-3- (methoxycarbonyl)-2,3-dihydrobenzofuran-6-yl)piperazine-l-carboxylate, 6’ (1.7 g, 3.31 mmol) in AcOH (40.0 mL) was added H2SO4 (0.353 mL, 6.63 mmol). The resulting reaction mixture was heated to 120 °C and stirred for 12 h. The reaction mixture was concentrated under reduced pressure to obtain crude 6-(piperazin-l-yl)-2H-spiro[benzofuran-3,3'-piperidine]-2',6'-dione, 2 AcOH, 7’ (2.0 g) as a thick brown liquid, which was used in the next step without further purification.

[0264] LCMS Method 2: retention time: 1.262 min, [M+H]+= 302.2.

[0265] Step 6’. Preparation of tert-butyl 4-(2',6'-dioxo-2H-spiro[benzofuran-3,3'- piperidin]-6-yl)piperazine-l-Carboxylate 8’: To a stirred solution of 6-(piperazin-l-yl)-2H- spiro[benzofuran-3,3'-piperidine]-2',6'-dione, 2 AcOH, 7’ (2.0 g) in acetonitrile (30.0 mL) at RT was added DIPEA (8.0 mL, 45.8 mmol) and stirred for 5 min. Then di -tert-butyl dicarbonate (3.36 mL, 4.47 mmol) was added to the reaction mixture, and the resulting reaction mixture was stirred at RT for 4 h. The reaction mixture was diluted with water (50 mL) and extracted with DCM (3 x 100 mL). The combined organic layer was washed with brine (50 mL), dried over anhydrous sodium sulfate, and filtered, and the filtrate was concentrated under reduced pressure to obtain the crude product. The crude product was purified by column chromatography using silica gel (230-400 mesh) with 60% ethyl acetate / pet ether to obtain tert-butyl 4-(2',6'-dioxo-2H-spiro[benzofuran-3,3'- piperidin]-6-yl)piperazine-l -carboxylate 8’ (650 mg, 1.587 mmol, 54.8% yield) as a white solid.

[0266] LCMS Method 2: retention time: 2.319 min, [M+H]+= 402.2.

[0267] Step 7’. Chiral SFC separation of tert-butyl 4-(2',6'-dioxo-2H-spiro[benzofuran- 3,3'-piperidin]-6-yl)piperazine-l-carboxylate 9a' (Peak-1) and tert-butyl 4-(2',6'-dioxo-2H- spiro[benzofuran-3,3'-piperidin]-6-yl)piperazine-l-carboxylate 9b' (Peak-2): 750 mg of the racemic compound was purified by chiral SFC to obtain tert-butyl 4-(2',6'-dioxo-2H- spiro[benzofuran-3,3'-piperidin]-6-yl)piperazine-l-carboxylate 9a’ (240 mg, Peak-1) and tert-butyl 4-(2',6'-dioxo-2H-spiro[benzofuran-3,3'-piperidin]-6-yl)piperazine-l-carboxylate 9b’ (260 mg, Peak-2).

[0268] SFC Method: YMC Cellulose SC, 250 X 4.6 mm, 5 pm, Flow: 3.0 mL / min. Co-Solvent: 50.0% IPA.

[0269] Peak-1 (RT 4.843 min, ee 99.95%): tert-butyl4-(2',6'-dioxo-2H-spiro[benzofuran-3,3'- piperidin]-6-yl)piperazine- 1 -carboxylate, 9a’ .

[0270] 1H NMR (400 MHz, DMSO-d6): 5 ppm 1.41 (s, 9H), 2.08 - 2.14 (m, 2H), 2.50 - 2.74 (m, 2H), 3.05 - 3.08 (m, 4H), 3.35 - 3.43 (m, 4H), 4.39 (d, J= 9.2 Hz, 1H), 4.81 (d, J= 9.2 Hz, 1H), 6.43 - 6.46 (m, 2H), 7.04 (d, J= 8.8 Hz, 1H), 10.95 (s, 1H).

[0271] LCMS method 2: retention time: 2.321 min, 96.06% purity at 220 nm, [M+H]+= 402.2.

[0272] Peak-2 (RT 5.995 min, ee 99.82%): tert-butyl4-(2',6'-dioxo-2H-spiro[benzofuran-3,3'- piperidin]-6-yl)piperazine- 1 -carboxylate, 9b’ .

[0273] 1H NMR (400 MHz, DMSO-d6): 5 ppm 1.42 (s, 9H), 2.06 - 2.16 (m, 2H), 2.60 - 2.68 (m, 2H), 3.06 - 3.09 (m, 4H), 3.41 - 3.44 (m, 4H), 4.40 (d, J= 9.2 Hz, 1H), 4.81 (d, J= 9.2 Hz, 1H), 6.44 - 6.46 (m, 2H), 7.04 - 7.06 (m, 1H), 10.96 (s, 1H).

[0274] LCMS method 2: retention time: 2.322 min, 96.59% purity at 220 nm, [M+H]+= 402.2.

[0275] Step 8a’. Preparation of 6-(piperazin-l-yl)-2Hspiro[benzofuran-3,3'-piperidine]- 2',6'-dione.2 HC1 C-l: To a stirred solution of tert-butyl 4-(2',6'-dioxo-2H-spiro[benzofuran-3,3'- piperidin]-6-yl)piperazine-l -carboxylate, 9a’ (Peak-1) (230 mg, 0.55 mmol) in DCM (5 mL) at RT was added 4.0 N HC1 in 1,4-dioxane (4.0 mL, 16 mmol). The resulting reaction mixture was stirred at RT for 6 h. The reaction mixture was concentrated under reduced pressure to obtain crude 6- (piperazin-l-yl)-2Hspiro[benzofuran-3,3'-piperidine]-2',6'-dione.2HCl, C-l (Peak-1) (200 mg) as an off-white solid, which was used in the next step without further purification.

[0276] LCMS method 2: retention time: 1.284 min, [M+H]+= 302.2.

[0277] Step 8b’. Preparation of 6-(piperazin-l-yl)-2Hspiro[benzofuran-3,3'-piperidine]- 2',6'-dione.2HCl C-2: To a stirred solution of tert-butyl4-(2',6'-dioxo-2H-spiro[benzofuran-3,3'- piperidin]-6-yl)piperazine-l -carboxylate, 9b’ (Peak-2) (250 mg, 0.601 mmol) in DCM (5 mL) at RT was added 4.0 N HC1 in 1,4-dioxane (4.0 mL, 16 mmol). The resulting reaction mixture was stirred at RT for 6 h. The reaction mixture was concentrated under reduced pressure to obtain crude6-(piperazin-l-yl)-2H-spiro[benzofuran-3,3'-piperidine]-2',6'-dione, 2 HC1, C-2 (Peak-2) (210 mg) as an off-white solid, which was used in the next step without further purification.

[0278] LCMS method 2: retention time: 1.305 min, [M+H]+= 302.1.

[0279] CBM intermediates prepared via the procedure described in Example S7 are summarized in Table 2.Table 2.Example S8: Preparation of 6-(piperidin-4-yl)-2H-spiro[benzofuran-3,3'-piperidine]-2',6'- dione (C-ll and C-12)

[0280] LCMS Method 1. Kinetex XB - C18, 50 x 4.6 mm, 5.0 pm. Temperature: RT, Flow: 1.0 mL / min, run time: 5.5 min. Mobile Phase Conditions: Mobile Phase-A: 0.1% TFA in H2O, Mobile Phase-B: 0.1% TFA in ACN, Gradient: Initial 95% Mobile Phase A and 5% Mobile Phase B linear gradient to 95% Mobile Phase B for 2.5 min. MSD positive.

[0281] LCMS Method 2, Aquity Uplc BEH Cl 8, 50 x 2.1 mm, 1.7 pm. Temperature: RT, Flow: 0.7 mL / min, run time: 2.5 min. Mobile Phase Conditions: Mobile Phase-A: 0.1% TFA in H2O, Mobile Phase-B: 0.1% TFA in ACN, Gradient: Initial 95% Mobile Phase A and 5% Mobile Phase B linear gradient to 98% Mobile Phase B for 1.5 min and then hold for 0.5 min. MSD positive.

[0282] LCMS Method 3 , Column: Kinetex XB - C18 75 x 3.0 mm, 2.6 pm. Temperature: RT Flow: 1.0 mL / min, run time: 5.0 min. Mobile Phase Conditions: Mobile Phase-A: 5.0 mm Ammonium formate pH 3.3:CHsCN (98:02), Mobile Phase-B: CH3CN: Buffer (98:02), Gradient: Initial 40% Mobile Phase A and 60% Mobile Phase B linear gradient to 100% Mobile Phase B for 3.5 min then hold for 1.5 minute. MSD positive.

[0283] LCMS Method 4, Column: Kinetex XB - C18 75 x 3.0 mm, 2.6 pm. Temperature: RT, Flow: 1.0 mL / min, run time: 5.0 min. Mobile Phase Conditions: Mobile Phase-A: 5.0 mm Ammonium formate pH 3.3:CHsCN (98:02), Mobile Phase-B: CH3CN: Buffer (98:02), Gradient: Initial 20% Mobile Phase A and 80% Mobile Phase B linear gradient to 100% Mobile Phase B for 4.0 min then hold for 0.6 minute. MSD positive.

[0284] Step 1’. Preparation of tert-butyl 4-(3-(ethoxycarbonyl)benzofuran-6-yl)-3,6- dihydropyridine-l(2H)-carboxylate 3’: To a stirred solution of ethyl 6-bromobenzofuran-3- carboxylate 1’ (650 mg, 2.416 mmol) in 1,4-dioxane (10 mL) and water (1.11 mL) was added K2CO3 (835 mg, 6.04 mmol) and tert-butyl 4-(3-(ethoxycarbonyl)benzofuran-6-yl)-3,6- dihydropyridine-l(2H)-carboxylate 2’ (822 mg, 2.66 mmol) at RT, and the reaction mixture was degassed for 10 min. Then PdC12(dppf) (177 mg, 0.242 mmol) was added and stirred at 100 °C for 16 h. The reaction mixture was filtered through a Celite bed and washed with ethyl acetate (3 x 20 mL). The combined organic layer was washed with saturated brine solution (20 mL), dried over Na2SO4, filtered, and then concentrated under reduced pressure to give the crude product. The crude product was purified by column chromatography using silica gel (230-400 mesh) and eluted with 25% ethyl acetate in pet ether to give tert-butyl 4-(3-(ethoxycarbonyl)benzofuran-6-yl)-3,6- dihydropyridine-l(2H)-carboxylate 3’ (600 mg, 63.3% yield) as a pale yellow liquid.

[0285] LCMS method 3 : retention time: 2.96 min, [M+H-100]+= 272.1.

[0286] 1H NMR (400 MHz, DMSO-d6): 5 ppm 1.35 (t, J= 7.2 Hz, 3H), 1.44 (s, 9H), 2.50 -2.54 (m, 2H), 3.57 (t, J = 5.6 Hz, 2H), 4.03 - 4.04 (m, 2H), 4.32 - 4.38 (m, 2H), 6.27 (s, 1H), 7.53 -7.55 (m, 1H), 7.75 (s, 1H), 7.91 (d, J= 8.4 Hz, 1H), 8.74 (s, 1H).

[0287] Step 2’. Preparation of tert-butyl 4-(3-(ethoxycarbonyl)-2,3-dihydrobenzofuran-6- yl)piperidine-l-carboxylate 4’: To a stirred solution of tert-butyl 4-(3- (ethoxycarbonyl)benzofuran-6-yl)-3,6-dihydropyridine-l(2H)-carboxylate 3’ (600 mg, 1.615 mmol) in ethyl acetate (10 mL) was added 10% Pd / C (344 mg) under hydrogen atmosphere (balloon pressure) and stirred at 25 °C for 16 h. The reaction mixture was filtered through a Celite bed and washed with ethyl acetate (3 x 20 mL). The combined organic layer was concentrated under reduced pressure to give crude tert-butyl 4-(3-(ethoxycarbonyl)-2,3-dihydrobenzofuran-6- yl)piperidine-l -carboxylate 4’ (650 mg, 100% yield), which was used in the next step without further purification.

[0288] LCMS method 4: retention time: 3.50 min, [M+H-100]+= 276.0.

[0289] Step 3’. Preparation of tert-butyl 4-(3-(2-cyanoethyl)-3-(ethoxycarbonyl)-2,3- dihydrobenzofuran-6-yl)piperidine-l-carboxylate 5’: To a stirred solution of tert-butyl 4-(3- (ethoxycarbonyl)-2,3-dihydrobenzofuran-6-yl)piperidine-l-carboxylate 4’ (600 mg, 1.598 mmol) in toluene (15 mL) was added acrylonitrile (170 mg, 3.20 mmol) and benzyltri ethylammonium chloride (72.8 mg, 0.320 mmol), followed by K2CO3 (442 mg, 3.20 mmol). The reaction mixture was stirred at 80 °C for 16 h. The reaction mixture was quenched with ice cold water (10 mL) and extracted with ethyl acetate (3 x 10 mL). The combined organic layer was washed with brine solution, dried over sodium sulphate and concentrated under reduced pressure to give crude tert-butyl 4-(3-(2-cyanoethyl)-3 -(ethoxycarbonyl)-2, 3 -dihydrobenzofuran-6-yl)piperi dine- 1 -carboxylate 5’ (700 mg, 84% yield), which was used in the next step without further purification.

[0290] LCMS method 4: retention time: 2.48 min [M+H-56]+= 373.2.

[0291] Step 4’. Preparation of 6-(piperidin-4-yl)-2H-spiro[benzofuran-3,3'-piperidine]- 2',6'-dione 6’: To a stirred solution of tert-butyl 4-(3-(2-cyanoethyl)-3-(ethoxycarbonyl)-2,3- dihydrobenzofuran-6-yl)piperidine-l -carboxylate 5’ (600 mg, 1.400 mmol) in acetic acid (5 mL) was added H2SO4 (0.373 mL, 7.00 mmol) and stirred at 120 °C for 16 h. The reaction mixture was concentrated under reduced pressure and the residual amount of acid was removed by codistillation with ACN (10 mL) and toluene (10 mL) to afford (crude) 6-(piperidin-4-yl)-2H- spiro[benzofuran-3,3'-piperidine]-2',6'-dione 6’ (500 mg, 81 % yield) as a pale yellow liquid, which was used in the next step without further purification.

[0292] LCMS method 2: retention time 0.85 min, [M+H]+= 301.0.

[0293] Step 5’. Preparation of tert-butyl 4-(2',6'-dioxo-2H-spiro[benzofuran-3,3'- piperidin]-6-yl)piperidine-l-carboxylate 7’: At 0 °C, to a stirred solution of 6-(piperidin-4-yl)- 2H-spiro[benzofuran-3,3'-piperidine]-2',6'-dione 6’ (600 mg, 1.998 mmol) in acetonitrile (10 mL) was added DIPEA (3.49 mL, 19.98 mmol) and Boc-anhydride (0.928 mL, 4.00 mmol), and stirred for 2 h. The reaction mixture was diluted with ethyl acetate (20 mL), cold water was added (20 mL) and extracted with ethyl acetate (3 x 20 mL). The combined organic layers were washed with brine solution, dried over sodium sulfate and then concentrated under reduced pressure to give the crude product. The crude product was purified by column chromatography using silica gel (230 -400 mesh) and eluted with 65% ethyl acetate in pet ether to give tert-butyl 4-(2',6'-dioxo-2H- spiro[benzofuran-3,3'-piperidin]-6-yl)piperidine-l-carboxylate 7’ (300 mg) as a pale yellow solid (mixture of enantiomers).

[0294] LCMS method 4: retention time 2.63 min, [M+H-56]+= 345.0.

[0295] Step 6’. Chiral SFC separation of tert-butyl 4-(2',6'-dioxo-2H-spiro[benzofuran- 3,3'-piperidin]-6-yl)piperidine-l-carboxylate 8a’ (Peak-01) and tert-butyl 4-(2',6'-dioxo-2H- spiro[benzofuran-3,3'-piperidin]-6-yl)piperidine-l-carboxylate 8b' (Peak- 02): The enantiomers of tert-butyl 4-(2',6'-dioxo-2H-spiro[benzofuran-3,3'-piperidin]-6-yl)piperidine-l- carboxylate 7’ ( 500 mg) were purified using SFC to give pure 8a’ (Peak-01) (100 mg, 12.27% yield) as an off-white solid, and 8b’(Peak-02) (200 mg, 24.16 % yield) as an off-white solid.

[0296] SFC Separation: YMC Cellulose-SC, 250 mm x 4.6 x 5 pm, Flowrate: 3.0 mL / min, Co-Solvent: 40.0%.

[0297] Peak 01: 8a’ (Rt 4.21 min, ee 100%).

[0298] LCMS method 4: retention time: 2.58 min, [M+H-56]+= 345.2.

[0299] 1H NMR (400 MHz, DMSO-cU): 6 ppm 1.44 - 1.47 (m, 11H), 1.70 - 1.73 (m, 2H), 2.13 - 2.17 (m, 2H), 2.60 - 2.77 (m, 5H), 4.03 - 4.06 (m, 2H), 4.43 (d, J= 92 Hz, 1H), 4.80 (d, J= 92 Hz, 1H), 6.75 - 6.77 (m, 2H), 7.14 (d, J= 8.4 Hz, 1H), 11.00 (s, 1H).

[0300] Peak 02: 8b’ (Rt 4.81 min, ee 98.74%).

[0301] LCMS method 4: retention time: 2.64 min, [M+H-100]+= 301.2.

[0302] 1H NMR (400 MHz, DMSO-d6): 8 ppm 1.38 - 1.47 (m, 11H), 1.70 - 1.73 (m, 2H), 2.13 - 2.17 (m, 2H), 2.60 - 2.82 (m, 5H), 4.03 - 4.06 (m, 2H), 4.43 (d, J= 9.2 Hz, 1H), 4.80 (d, J= 92 Hz, 1H), 6.75 - 6.77 (m, 2H), 7.14 (d, J= 8.4 Hz, 1H), 11.00 (s, 1H).

[0303] Step 7’. Preparation of 6-(piperidin-4-yl)-2H-spiro[benzofuran-3,3'-piperidine]- 2',6'-dione,HCl, C-ll: To a stirred solution of tert-butyl 4-(2',6'-dioxo-2H-spiro[benzofuran-3,3'- piperidin]-6-yl)piperidine-l -carboxylate 8a’ (100 mg, 0.250 mmol) in DCM (2 mL) was added 4.0 M HC1 in 1,4-dioxane (0.624 mL, 2.497 mmol) at RT and stirred for 1 h. The reaction mixture was concentrated under reduced pressure and the residual amount of acid was removed by codistillation with ACN (10 mL) and toluene (10 mL) to afford crude 6-(piperidin-4-yl)-2H- spiro[benzofuran-3,3'-piperidine]-2',6'-dione, HC1 C-ll (100 mg), which was used in the next step without further purification.

[0304] LCMS method 1 : retention time 1.61 min, [M+H]+= 301.3.

[0305] Step 8’. Preparation of 6-(piperidin-4-yl)-2H-spiro[benzofuran-3,3'-piperidine]- 2',6'-dione,HCl C-12: To a stirred solution of tert-butyl 4-(2',6'-dioxo-2H-spiro[benzofuran-3,3'- piperidin]-6-yl)piperidine-l -carboxylate 8b’ (140 mg, 0.350 mmol) in DCM (5 mL) was added 4.0 M HC1 in 1,4-dioxane (0.437 mL, 1.748 mmol) at RT and stirred for 1 h. The reaction mixture was concentrated under reduced pressure and the residual amount of acid was removed by codistillation with ACN (10 mL) and toluene (10 mL) to afford crude 6-(piperidin-4-yl)-2H- spiro[benzofuran-3,3'-piperidine]-2',6'-dione, HC1 C-12 (Peak -02) (131 mg), which was used in the next step without further purification.

[0306] LCMS method 1 : retention time 1.56 min, [M+H]+= 301.1.Example S9: Preparation of 5-methyl-6-(piperazin-l-yl)-2H-spiro[benzofuran-3,3'- piperidine]-2',6'-dione (C-13 and C-14)

[0307] LCMS Method 1. Kinetex XB - Cl 8, 75 x 3.0 mm, 2.6 m. Temperature: RT, Flow: 1.0 mL / min, run time: 5.5 min. Mobile Phase Conditions: Mobile Phase-A: 0.1% TFA in H2O, Mobile Phase-B: 0.1% TFA in ACN, Gradient: Initial 95% Mobile Phase A and 5% Mobile Phase B linear gradient to 95% Mobile Phase B for 4.5 min. MSD positive.

[0308] LCMS Method 2, Kinetex XB - C18, 75 x 3.0 mm, 2.6 pm. Temperature: RT, Flow: 1.0 mL / min, run time: 5.0 min. Mobile Phase Conditions: Mobile Phase-A: 5.0 mm ammonium foramte pH 3.3 in CH3CN (98:02), Mobile Phase-B: CH3CN in 5.0 mm ammonium foramte (98:02), Gradient: Initial 80% Mobile Phase A and 20% Mobile Phase B linear gradient to 100% Mobile Phase B for 4.06 min. MSD positive.

[0309] LCMS Method 3 , Aquity Uplc BEH C18 50 x 2.1 mm, 1.7pm. Temperature: RT, Flow:0.7 mL / min, run time: 2.0 min. Mobile Phase Conditions: Mobile Phase-A: 0.1% TFA in H2O, Mobile Phase-B: 0.1% TFA in ACN, Gradient: Initial 95% Mobile Phase A and 40% Mobile Phase B linear gradient to 98% Mobile Phase B for 2.0 min. MSD positive.

[0310] Step 1’. Preparation of ethyl 6-bromo-5-methylbenzofuran-3-carboxylate 2’: To a stirred solution of 4-bromo-2-hydroxy-5-methylbenzaldehyde 1’ (1.0 g, 4.65 mmol) in DCM (1.0 mL) was added tetrafluoroboric acid diethyl ether complex (0.063 mL, 0.465 mmol) followed by ethyl diazoacetate solution in toluene (7.90 mL, 9.30 mmol), which was added dropwise to the reaction mixture. The reaction was not allowed to go above 38 °C. After being stirred for 1 h at room temperature, tetrafluoroboric acid diethyl ether complex (0.063 mL, 0.465 mmol) and ethyl diazoacetate solution in toluene (7.90 mL, 9.30 mmol) was added to the reaction mixture and stirred for another 1 h. Once gas evolution ceased, the reaction mixture was concentrated using a rotary evaporator and H2SO4 (0.3 to 0.5 mL) was added to the mixture while stirring. After 5 to 10min, the mixture was diluted with CH2CI2 (5 to 10 mL) and the H2SO4 was quenched with saturated NaHCCh solution. The reaction mixture was extracted with DCM, the combined organic layers dried over sodium sulphate, filtered, concentrated and the crude compound was purified by column chromatography using silica gel (230-400 mesh) with 0 - 5% ethyl acetate / pet ether to obtain ethyl 6-bromo-5-methylbenzofuran-3-carboxylate 2’ (900 mg, 68.4% yield) as a pale-yellow solid.

[0311] 1H NMR (400 MHz, CDCI3): 8 ppm 1.44 (t, J= 7.2 Hz, 3H), 2.54 (s, 3H), 4.43 (q, J= 6.8 Hz, 2H), 7.77 (s, 1H), 7.95 (s, 1H), 8.21 (s, 1H).

[0312] Step 2’. Preparation of tert-butyl 4-(3-(ethoxycarbonyl)-5-methylbenzofuran-6- yl)piperazine-l-carboxylate 4’: To a stirred solution of ethyl 6-bromo-5-methylbenzofuran-3- carboxylate 2’ (0.900 g, 3.18 mmol) and tert-butyl piperazine- 1 -carboxylate 3’ (0.987 g, 5.30 mmol) in 1,4-dioxane (10.0 mL) under nitrogen atmosphere at room temperature was added cesium carbonate (1.726 g, 5.30 mmol). The reaction mixture was degassed with nitrogen gas for 10 min and RuPhos Pd G3 (0.222 g, 0.265 mmol) was added under positive flow of nitrogen gas and further stirred at 100 °C for 6 h. The reaction mixture was cooled to room temperature, quenched with saturated ammonium chloride solution and extracted with ethyl acetate. The combined organic layers were dried over anhydrous sodium sulphate, filtered, and concentrated under reduced pressure to give the crude product. The crude product was purified by column chromatography using silica gel (230 - 400 mesh) with 0 - 20% ethyl acetate / pet ether to obtain tert-butyl 4-(3- (ethoxycarbonyl)-5-methylbenzofuran-6-yl)piperazine-l -carboxylate 4’ (600 mg, 53.6% yield) as a yellow liquid.

[0313] LCMS Method 1 : Retention time: 3.589 min, [M+H-56]+= 333.9.

[0314] Step 3’. Preparation of tert-butyl 4-(3-(methoxycarbonyl)-5-methyl-2,3- dihydrobenzofuran-6-yl)piperazine-l-carboxylate 5’: To a stirred solution of tert-butyl 4-(3- (ethoxycarbonyl)-5-methylbenzofuran-6-yl)piperazine-l-carboxylate 4’ (600 mg, 1.545 mmol) in MeOH (15.0 mL) was added magnesium (563 mg, 23.17 mmol) at room temperature under nitrogen atmosphere and the reaction mixture was stirred for 16 h at room temperature. The reaction mixture was quenched with aqueous ammonium chloride (100 mL) and concentrated to remove methanol. The crude product was extracted with ethyl acetate (3 x 100 mL) and the organic layer was washed with brine (40 mL). The combined organic layers were dried over anhydrous sodium sulfate, and filtered, and concentrated under reduced pressure to obtain the crude product tert-butyl 4-(3-(methoxycarbonyl)-5-methyl-2,3-dihydrobenzofuran-6-yl)piperazine-l-carboxylate 5’ (585 mg, 91% yield) as a yellow liquid. The crude product was used in the next step without further purification.

[0315] LCMS Method 2: Retention time: 3.281 min, [M+H]+= 377.4.

[0316] Step 4’. Preparation of tert-butyl 4-(3-(2-cyanoethyl)-3-(methoxycarbonyl)-5- methyl-2,3-dihydro benzofuran-6-yl)piperazine-l-carboxylate 6’: To a stirring solution of tertbutyl 4-(3-(methoxycarbonyl)-5-methyl-2,3-dihydrobenzofuran-6-yl)piperazine-l -carboxylate 5’ (581 mg, 1.543 mmol), acrylonitrile (0.203 mL, 3.09 mmol) and K2CO3 (427 mg, 3.09 mmol) in toluene (10.0 mL) was added benzyltriethylammonium chloride (35.2 mg, 0.154 mmol) at room temperature and the reaction mixture was then stirred at 90 °C for 16 h. The reaction mixture was treated with water (50 mL) and extracted with ethyl acetate (3 x 50.0 mL). The combined organic layer was then dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give the crude product. The crude product was purified by column chromatography using silica gel (230 - 400 mesh) with 0 - 20% ethyl acetate / pet ether to obtain tert-butyl 4-(3-(2- cyanoethyl)-3-(methoxycarbonyl)-5-methyl-2,3-dihydrobenzofuran-6-yl)piperazine-l-carboxylate 6’ (450 mg, 59.1% yield) as a yellow liquid.

[0317] LCMS Method 2: Retention time: 3.193 min, [M+H]+= 430.4.

[0318] Step 5’. Preparation of 5-methyl-6-(piperazin-l-yl)-2H-spiro[benzofuran-3,3'- piperidine]-2',6'-dione 7’: To a stirred solution of tert-butyl 4-(3-(2-cyanoethyl)-3- (methoxycarbonyl)-5-methyl-2,3-dihydrobenzofuran-6-yl) piperazine- 1 -carboxylate 6’ (450 mg, 1.048 mmol) in acetic acid (5.0 mL) was added H2SO4 (0.112 mL, 2.095 mmol) dropwise at room temperature and the reaction mixture was stirred at 120 °C for 16 h. The reaction mixture was concentrated under reduced pressure to obtain crude 5-methyl-6-(piperazin-l-yl)-2H- spiro[benzofuran-3,3'-piperidine]-2',6'-dione, 2 sulfuric acid salt 7’ (600 mg) as a thick brown liquid. The crude product was used in the next step without further purification.

[0319] LCMS Method 3: retention time: 0.388 min, [M+H]+= 316.0 .

[0320] Step 6’. Preparation of tert-butyl 4-(5-methyl-2',6'-dioxo-2H-spiro[benzofuran- 3,3'-piperidin]-6-yl)piperazine-l-carboxylate 8’: To a stirred solution of 5-methyl-6-(piperazin- l-yl)-2H-spiro[benzofuran-3,3'-piperidine]-2',6'-dione, 2 sulfuric acid salt 7’ (600 mg, 1.173 mmol) in acetonitrile (5.0 mL) was added DIPEA (3.07 mL, 17.59 mmol) until the solution became basic, followed by Boc-anhydride (1.362 mL, 5.86 mmol) dropwise at room temperature under the positive flow of nitrogen gas and the reaction mixture was stirred at room temperature for 4 h. The reaction mixture was treated with water (50 mL) and extracted with ethyl acetate (3 x 100 mL). The combined organic layers were washed with brine (50 mL), dried over anhydrous sodium sulphate, filtered, and concentrated under reduced pressure to obtain the crude product. The crude product was purified by column chromatography using silica gel (230 - 400 mesh) with 0 - 60% ethyl acetate / pet ether to give tert-butyl 4-(5-methyl-2',6'-dioxo-2H-spiro[benzofuran-3,3'-piperidin]-6- yl)piperazine-l -carboxylate 8’ (340 mg, 64.2% yield) as an off-white solid.

[0321] LCMS Method 3: retention time: 1.345 min, [M+H]+= 416.0.

[0322] Step 7’. Chiral SFC separation of tert-butyl 4-(5-methyl-2',6'-dioxo-2H- spiro[benzofuran-3,3'-piperidin]-6-yl)piperazine-l-carboxylate 9a' (Peak-1) and tert-butyl 4- (5-methyl-2',6'-dioxo-2Hspiro[benzofuran-3,3'-piperidin]-6-yl)piperazine-l-carboxylate 9b' (Peak-2): 340 mg of the racemic compound was purified by chiral SFC to obtain tert-butyl 4-(5- methyl-2',6'-dioxo-2Hspiro[benzofuran-3,3'-piperidin]-6-yl)piperazine-l-carboxylate 9a' (120 mg, Peak-1) and tert-butyl 4-(5-methyl-2',6'-dioxo-2Hspiro[benzofuran-3,3'-piperidin]-6-yl)piperazine- 1-carboxylate 9b' (130 mg, Peak-2).

[0323] SFC method: YMC Cellulose-SC, 250 mm x 4.6 x 5 pm, Flowrate: 3.0 mL / min (1 : 1 ACN and IP A), Co-Solvent: 40.0%, BPR: 100.0 bar, BPR Temperature: 50 °C.

[0324] Peak 01: RT 3.083 min, ee 100%.

[0325] LCMS method 2: retention time: 2.767 min, [M+H]+= 416.2.

[0326] 1H NMR (400 MHz, DMSO-d6): 5 ppm 1.22 - 1.31 (m, 1H), 1.42 (s, 9H), 2.10 - 2.19 (m, 5H), 2.70 - 2.79 (m, 4H), 3.40 - 3.50 (m, 4H), 4.49 (d, J= 92 Hz, 1H), 4.79 (d, J= 92 Hz, 1H), 6.55 (s, 1H), 7.02 (s, 1H), 10.98 (s, 1H).

[0327] Peak 02: RT 3.797 min, ee 99.73%.

[0328] LCMS method 1 : retention time: 2.767 min, [M+H]+= 416.2.

[0329] Step 8’. Preparation of 5-methyl-6-(piperazin-l-yl)-2H-spiro[benzofuran-3,3'- piperidine]-2',6'-dione, 2 HC1 C-13: To a stirred solution of tert-butyl 4-(5-methyl-2',6'-dioxo- 2H-spiro[benzofuran-3,3'-piperidin]-6-yl)piperazine-l-carboxylate 9’ (0.120 g, 0.289 mmol) in DCM (10 mL) at room temperature under nitrogen atmosphere was added 4.0 N hydrogen chloride solution in 1,4-dioxane (2.166 mL, 8.66 mmol) dropwise and the reaction mixture was stirred for 3 h at room temperature. The reaction mixture was concentrated under reduced pressure to obtain crude 5-methyl-6-(piperazin-l-yl)-2H-spiro[benzofuran-3,3'-piperidine]-2',6'-dione, 2 HC1 C13 (120 mg) as an off-white solid. The crude product was used in the next step without further purification.

[0330] LCMS method 2: retention time: 1.993 min, [M+H]+= 316.2.Example S10: Preparation of 7-(piperazin-l-yl)-2H-spiro[benzofuran-3,3'-piperidine]-2',6'- dione (C-15 and C-16)

[0331] LCMS Method 1. Kinetex XB - C18, 75 x 3.0 mm, 2.6 pm. Temperature: RT, Flow: 1.0 mL / min, run time: 4.7 min. Mobile Phase Conditions: Mobile Phase-A: 5.0 mm ammonium foramte pH 3.3 in CH3CN (98:02), Mobile Phase-B: CH3CN in 5.0 mm ammonium foramte (98:02), Gradient: Initial 60% Mobile Phase A and 40% Mobile Phase B linear gradient to 100% Mobile Phase B for 3.5 min and then hold for 1.0 min. MSD positive.

[0332] LCMS Method 2, Kinetex XB - C18, 75 x 3.0 mm, 2.6 pm. Temperature: RT, Flow: 1.0 mL / min, run time: 5.5 min. Mobile Phase Conditions: Mobile Phase-A: 0.1% TFA in H2O, Mobile Phase-B: 0.1% TFA in ACN, Gradient: Initial 95% Mobile Phase A and 5% Mobile Phase B linear gradient to 95% Mobile Phase B for 2.5 min and then hold for 2.0 min. MSD positive.

[0333] LCMS Method 3 , Aquity Uplc BEH Cl 8, 50 x 2.1 mm, 1.7pm. Temperature: RT, Flow: 0.7 mL / min, run time: 2.0 min. Mobile Phase Conditions: Mobile Phase-A: 0.1% TFA in H2O, Mobile Phase-B: 0.1% TFA in ACN, Gradient: Initial 80% Mobile Phase A and 20% Mobile Phase B linear gradient to 98% Mobile Phase B for 1.5 min and then hold for 0.5 min. MSD positive.

[0334] LCMS Method 4, Kinetex XB - C18, 75 x 3.0 mm, 2.6 pm. Temperature: RT, Flow: 1.0 mL / min, run time: 4.7 min. Mobile Phase Conditions: Mobile Phase-A: 5.0 mm ammonium foramte pH 3.3 in CH3CN (98:02), Mobile Phase-B: CH3CN in 5.0 mm ammonium foramte (98:02), Gradient: Initial 80% Mobile Phase A and 20% Mobile Phase B linear gradient to 100% Mobile Phase B for 4.0 min and then hold for 0.6 min. MSD positive.

[0335] Step 1’. Preparation of ethyl 7-bromobenzofuran-3-carboxylate 2’: To a stirred solution of 3-bromo-2-hydroxybenzaldehyde 1’ (3.5 g, 17.41 mmol) in DCM (15 mL) was added tetrafluoro boric acid diethyl ether complex (0.235 mL, 1.741 mmol), followed by ethyl diazo acetate in toluene (29.6 mL, 34.8 mmol, 15% in toluene) dropwise and stirred at 35 °C. Once the effervescence ceased, the reaction mixture was concentrated under reduced pressure and cone. cat. H2SO4 (0.5 mL) was added and stirred for 16 h. The reaction mixture was diluted with ethyl acetate (30 mL) and the H2SO4 was quenched with saturated NaHCCh solution. The reaction mixture was extracted with ethyl acetate (3 x 30 mL), the combined organic layers were collected, dried over sodium sulfate, filtered, concentrated and the crude compound was purified by column chromatography using silica gel (230-400 mesh) with 0 - 5% DCM / pet ether to obtain ethyl 7- bromobenzofuran-3 -carboxylate 2’ (700 mg, 9.83% yield) as a pale-yellow liquid.

[0336] 1H NMR (400 MHz, DMSO-d6): 8 ppm 1.36 (t, J= 7.2 Hz, 3H), 4.36 (q, J= 7.2 Hz, 2H), 7.37 (t, J= 7.2 Hz, 1H), 7.68 (dd, J= 1.2, 8.0 Hz, 1H), 7.98 (dd, J = 1.2 Hz and 8.0 Hz, 1H), 8.88 (s, 1H).

[0337] Step 2’. Preparation of tert-butyl 4-(3-(ethoxycarbonyl)benzofuran-7- yl)piperazine-l-carboxylate 4’: To a stirred solution of ethyl 7-bromobenzofuran-3 -carboxylate 2’ (750 mg, 2.79 mmol) and tert-butyl piperazine- 1 -carboxylate 3’ (519 mg, 2.79 mmol) in 1,4- dioxane (2 mL)) under nitrogen atmosphere at room temperature was added CS2CO3 (1816 mg, 5.57 mmol). The reaction mixture was degassed with nitrogen gas for 10 min and RuPhos-Pd-G3 (233 mg, 0.279 mmol) was added and stirred at 100 °C for 16 h. The reaction mixture was cooled to room temperature and quenched with saturated ammonium chloride solution (10 mL) and extracted with ethyl acetate (3 x 30 mL). The combined organic layers were collected, dried over anhydrous sodium sulphate, filtered, concentrated under reduced pressure to give crude product which was purified by column chromatography using silica gel (230 - 400 mesh) with 30% ethyl acetate / pet ether to obtain tert-butyl 4-(3-(ethoxycarbonyl)benzofuran-7-yl)piperazine-l-carboxylate 4’ (600 mg, 55.2% yield) as a pale yellow liquid.

[0338] LCMS Method 1 : Retention time: 2.83 min, [M+H]+= 375.2.

[0339] Step 3’. Preparation of tert-butyl 4-(3-(methoxycarbonyl)-2,3-dihydrobenzofuran- 7-yl)piperazine-l-carboxylate 5’: To a stirred solution of tert-butyl 4-(3- (ethoxycarbonyl)benzofuran-7-yl)piperazine-l -carboxylate 4’ (600 mg, 1.602 mmol) in methanol (10 mL) was added magnesium (1.93 g, 16.02 mmol) at 0 °C under nitrogen atmosphere and the reaction mixture was stirred at room temperature for 16 h. The reaction mixture was quenched with aqueous ammonium chloride (50 mL) and concentrated to remove methanol. The crude product was extracted with ethyl acetate (3 x 50 mL) and the organic layer was washed with brine (40 mL).The combined organic layer was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain the crude product tert-butyl 4-(3-(methoxycarbonyl)-2,3- dihydrobenzofuran-7-yl)piperazine-l-carboxylate 5’ (500 mg, 76% yields) as a pale yellow liquid. The crude product was used in the next step without further purification.

[0340] LCMS Method 2: Retention time: 3.00 min, [M+H]+= 363.2.

[0341] Step 4’. Preparation of tert-butyl 4-(3-(2-cyanoethyl)-3-(methoxycarbonyl)-2,3- dihydrobenzofuran-7-yl)piperazine-l-carboxylate 6’: To a stirred solution of tert-butyl 4-(3- (methoxycarbonyl)-2,3-dihydrobenzofuran-7-yl)piperazine-l-carboxylate 5’ (600 mg, 1.656 mmol), acrylonitrile (132 mg, 2.483 mmol), and K2CO3 (458 mg, 3.31 mmol) in toluene (10 mL) was added benzyltriethylammonium chloride (75 mg, 0.331 mmol) at room temperature under the positive flow of nitrogen gas and the reaction mixture was then stirred at 100 °C for 16 h. The reaction mixture was cooled to RT, treated with cold water (20 mL) and extracted with ethyl acetate (3 x 30.0 mL). The combined organic layer was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give crude tert-butyl 4-(3-(2-cyanoethyl)-3- (methoxycarbonyl)-2,3-dihydrobenzofuran-7-yl)piperazine-l-carboxylate 6’ (600 mg, 70.9% yield). The crude product was used in the next step without further purification.

[0342] LCMS Method 4: Retention time: 3.00 min, [M+H]+= 416.2.

[0343] Step 5’. Preparation of 7-(piperazin-l-yl)-2H-spiro[benzofuran-3,3'-piperidine]- 2',6'-dione 7’: To a stirred solution of tert-butyl 4-(3-(2-cyanoethyl)-3-(methoxycarbonyl)-2,3- dihydrobenzofuran-7-yl)piperazine-l-carboxylate 6’ (500 mg, 1.203 mmol) in acetic acid (10.0 mL) was added H2SO4 (0.160 mL, 3.01 mmol) dropwise at room temperature. The reaction mixture was stirred at 120 °C for 16 h. The reaction mixture was concentrated under reduced pressure to obtain crude sulfate salt of 7-(piperazin-l-yl)-2H-spiro[benzofuran-3,3'-piperidine]-2',6'-dione 7’ (500 mg) as a dark brown oil, which was used in the next step without further purification.

[0344] LCMS Method 3: retention time: 0.389 min, [M+H]+= 302.0.

[0345] Step 6’. Preparation of tert-butyl 4-(2',6'-dioxo-2H-spiro[benzofuran-3,3'- piperidin]-7-yl)piperazine-l-carboxylate 8’: To a stirred solution of 7-(piperazin-l-yl)-2H- spiro[benzofuran-3,3'-piperidine]-2',6'-dione 7’ (500 mg, 1.518 mmol) in acetonitrile (5 mL) was added DIPEA (2.65 mL, 15.18 mmol) until the solution became basic, then Boc-anhydride (0.705 mL, 3.04 mmol) was added at 0 °C under the positive flow of nitrogen gas and the reaction mixture was stirred at room temperature for 16 h. The reaction mixture was treated with water (20 mL) and extracted with ethyl acetate (3 x 20 mL). The combined organic layers were washed with brine (50 mL), dried over anhydrous sodium sulphate, filtered, and concentrated under reduced pressure to obtain the crude product. The crude product was purified by column chromatography using silicagel (230-400 mesh) with 65% ethyl acetate / pet ether to give the enantiomeric mixture of tert-butyl 4-(2',6'-dioxo-2H-spiro[benzofuran-3,3'-piperidin]-7-yl)piperazine-l-carboxylate 8’ (250 mg, 37.5% yield) as a pale yellow solid.

[0346] 1H NMR (400 MHz, DMSO- d6): b ppm 1.41 (s, 9H), 2.14 - 2.18 (m, 2H), 2.63 - 2.68 (m, 2H), 2.95 - 3.07 (m, 4H), 3.42 - 3.46 (m, 4H), 4.50 (d, J= 92 Hz, 1H), 4.82 (d, J= 8.8 Hz, 1H), 6.76 - 6.83 (m, 3H), 11.02 (s, 1H).

[0347] Step 7’. Chiral SFC separation of tert-butyl 4-(2',6'-dioxo-2H-spiro[benzofuran- 3,3'-piperidin]-7-yl)piperazine-l-carboxylate 9a’ (Peak-1) and tert-butyl 4-(2',6'-dioxo-2H- spiro[benzofuran-3,3'-piperidin]-7-yl)piperazine-l-carboxylate 9b’ (Peak-2): 250 mg of the racemic compound was purified by chiral SFC to obtain tert-butyl 4-(2',6'-dioxo-2H- spiro[benzofuran-3,3'-piperidin]-7-yl)piperazine-l-carboxylate 9a’ (60 mg, Peak-01) and tert-butyl 4-(2',6'-dioxo-2H-spiro[benzofuran-3,3'-piperidin]-7-yl)piperazine-l-carboxylate 9b’ (77 mg, Peak- 02).

[0348] SFC Method: Cellulose-1, 250 mm x 4.6 x 5 pm, Flowrate: 3.0 mL / min, Co-Solvent: 30.0%.

[0349] Peak 01 : Rt: 3.189 min, ee 100%.

[0350] LCMS method 2: retention time: 3.01 min, [M+H]+= 402.9.

[0351] 1H NMR (400 MHz, DMSO- d6): b ppm 1.41 (s, 9H), 2.14 - 2.18 (m, 2H), 2.63 - 2.68 (m, 2H), 2.95 - 3.07 (m, 4H), 3.44 - 3.45 (m, 4H), 4.50 (d, J= 92 Hz, 1H), 4.82 (d, J= 92 Hz, 1H), 6.77 - 6.83 (m, 3H), 11.02 (s, 1H).

[0352] Peak 02: Rt: 4.107 min, ee 99.83%.

[0353] LCMS method 4: retention time: 2.38 min, [M+H]+= 402.2.

[0354] 1H NMR (400 MHz, DMSO- d6): b ppm 1.42 (s, 9H), 2.14 - 2.18 (m, 2H), 2.63 - 2.68 (m, 2H), 2.97 - 3.07 (m, 4H), 3.04 - 3.45 (m, 4H), 4.50 (d, J= 92 Hz, 1H), 4.82 (d, J= 92 Hz, 1H), 6.77 - 6.83 (m, 3H), 11.02 (s, 1H).

[0355] Step 8a’. Preparation of 7-(piperazin-l-yl)-2H-spiro[benzofuran-3,3'-piperidine]-2',6'-dione, HC1 C-15: To a stirred solution of tert-butyl 4-(2',6'-dioxo-2H-spiro[benzofuran-3,3'- piperidin]-7-yl)piperazine-l -carboxylate 9a’ (60 mg, 0.149 mmol) in DCM (3 mL) at room temperature under the positive flow of nitrogen gas, 4.0 M HC1 in dioxane (0.187 mL, 0.747 mmol) was added dropwise and the reaction mixture was stirred for 1 h at room temperature. The reaction mixture was concentrated under reduced pressure to obtain crude 7-(piperazin-l-yl)-2H- spiro[benzofuran-3,3'-piperidine]-2',6'-dione, HC1 C-15 (50 mg, 97 % yield) as a pale yellow solid. The crude product was used in the next step without further purification.

[0356] LCMS method 4: retention time: 0.68 min, [M+H]+= 302.0.

[0357] Step 8b’. Preparation of 7-(piperazin-l-yl)-2H-spiro[benzofuran-3,3'-piperidine]- 2',6'-dione, HC1 C-16: To a stirred solution of tert-butyl 4-(2',6'-dioxo-2H-spiro[benzofuran-3,3'- piperidin]-7-yl)piperazine-l -carboxylate 9b’ (75 mg, 0.187 mmol) in DCM (5 mL) at room temperature under the positive flow of nitrogen gas, 4.0 M HC1 in dioxane (0.46 mL, 1.86 mmol) was added dropwise and the reaction mixture was stirred for 3 h at room temperature. The reaction mixture was concentrated under reduced pressure to obtain crude 7-(piperazin-l-yl)-2H- spiro[benzofuran-3,3'-piperidine]-2',6'-dione, HC1 C-16 (88 mg). The crude product was used in the next step without further purification.

[0358] LCMS method 1 : retention time: 0.76 min, [M+H]+= 302.0.Example Sil: Preparation of 5-((R)-2-methylpiperazin-l-yl)-2,3-dihydrospiro[indene-l,3'- p

[0359] LCMS Method 1. Kinetex XB - Cl 8, 75 x 3.0 mm, 2.6 m. Temperature: RT, Flow: 1.0 mL / min, run time: 5.0 min. Mobile Phase Conditions: Mobile Phase-A: 5.0 mm ammonium foramte pH 3.3 in CH3CN (98:02), Mobile Phase-B: CH3CN in 5.0 mm ammonium foramte (98:02), Gradient: Initial 80% Mobile Phase A and 20% Mobile Phase B linear gradient to 100% Mobile Phase B for 4.06 min. MSD positive.

[0360] LCMS Method 2, Aquity Uplc BEH C18 50 x 3.0 mm, 1.7pm. Temperature: RT, Flow: 0.7 mL / min, run time: 2.0 min. Mobile Phase Conditions: Mobile Phase-A: 0.1% TFA in H2O, Mobile Phase-B: 0.1% TFA in ACN, Gradient: Initial 80% Mobile Phase A and 20% Mobile Phase B linear gradient to 98% Mobile Phase B for 2.0 min. MSD positive.

[0361] LCMS Method 3 , Kinetex XB - C18, 75 x 3.0 mm, 2.6 pm. Temperature: RT, Flow: 1.0 mL / min, run time: 5.0 min. Mobile Phase Conditions: Mobile Phase-A: 0.1% TFA in H2O, MobilePhase-B: 0.1% TFA in ACN, Gradient: Initial 95% Mobile Phase A and 5% Mobile Phase B linear gradient to 95% Mobile Phase B for 4.0 min. MSD positive.

[0362] Step 1’. Preparation of tert-butyl 3-(5-bromo-l-cyano-2,3-dihydro-lH-inden-l- yl)propanoate 2’: To a stirred solution of 5-bromo-2,3-dihydro-lH-indene-l-carbonitrile 1’ (750 mg, 3.38 mmol) and tert-butyl acrylate (0.989 mL, 6.75 mmol) in toluene (2.0 mL) at room temperature was added K2CO3 (933 mg, 6.75 mmol) and benzyltriethylammonium chloride (77 mg, 0.338 mmol), and the reaction mixture was stirred at 90 °C for 20 h. The reaction was treated with water (100 mL) and extracted with ethyl acetate (3 x 50 mL), then the combined organic layers were dried over sodium sulfate, filtered, and concentrated under reduced pressure to obtain crude tert-butyl 3-(5-bromo-l-cyano-2,3-dihydro-lH-inden-l-yl)propanoate 2’ (1.05 g, 72.8% yield). The crude product was used in the next step without further purification.

[0363] LCMS Method 1 : Retention time: 3.640 min, [M+H]+= 350.0 and [M+H+2]+= 352.0.

[0364] Step 2’. Preparation of tert-butyl (3R)-4-(l-(3-(tert-butoxy)-3-oxopropyl)-l-cyano- 2,3-dihydro-lH-inden-5-yl)-3-methylpiperazine-l-carboxylate 4’: To a stirred solution of tertbutyl 3-(5-bromo-l-cyano-2,3-dihydro-lH-inden-l-yl)propanoate 2’ (1.0 g, 2.86 mmol) and (R)-l- Boc-3-methylpiperazine 3’ (0.858 g, 4.28 mmol) in 1,4-dioxane (1.0 mL) was added cesium carbonate (1.860 g, 5.71 mmol), then the reaction mixture was degassed with nitrogen gas for 5 mins, followed by addition of RuPhos Pd G4 (0.243 g, 0.286 mmol) at room temperature under a positive flow of nitrogen gas, and the reaction mixture was stirred at 100 °C for 16 h. The reaction mixture was cooled to room temperature, quenched with saturated ammonium chloride solution, and extracted with ethyl acetate. The combined organic layers were dried over anhydrous sodium sulphate, filtered, and concentrated under reduced pressure to give the crude product. The crude product was purified by column chromatography using silica gel (230 - 400 mesh) with 0 - 20% ethyl acetate / pet ether to obtain tert-butyl (3R)-4-(l-(3-(tert-butoxy)-3-oxopropyl)-l-cyano-2,3- dihydro-lH-inden-5-yl)-3-methylpiperazine-l-carboxylate 4’ (1.00 g, 61.5% yield) as a yellow liquid.

[0365] LCMS Method 1 : Retention time: 3.946 min, [M+H]+= 470.2.

[0366] Step 3’. Preparation of 5-((R)-2-methylpiperazin-l-yl)-2,3-dihydrospiro[indene- l,3'-piperidine]-2',6'-dione 5’: To a stirred solution of tert-butyl (3R)-4-(l-(3-(tert-butoxy)-3- oxopropyl)-l-cyano-2,3-dihydro-lH-inden-5-yl)-3-methylpiperazine-l-carboxylate 4’ (1.0g, 2.129 mmol) in AcOH (10.0 mL) was added sulfuric acid (0.340 mL, 6.39 mmol) at room temperature and the reaction mixture was stirred at 120 °C for 24 h. The reaction mixture was concentrated under reduced pressure to obtain crude 5-((R)-2-methylpiperazin-l-yl)-2,3-dihydrospiro[indene- l,3'-piperidine]-2',6'-dione, 2 sulfate salt 5’ (750 mg, 112% yield) as a brown liquid. The crudeproduct was used in the next step without further purification.

[0367] LCMS Method 2: Retention time: 0.378 min, [M+H]+= 314.0.

[0368] Step 4’. Preparation of tert-butyl (3R)-4-(2',6'-dioxo-2,3-dihydrospiro[indene-l,3'- piperidin]-5-yl)-3-methylpiperazine-l-carboxylate 6’: To stirred solution of 5-((R)-2- methylpiperazin-l-yl)-2,3-dihydrospiro[indene-l,3'-piperidine]-2',6'-dione 2 sulfate salt 5’ (0.670 g, 2.138 mmol) in acetonitrile (2.0 mL) was added DIPEA (5.60 mL, 32.1 mmol) until the solution became basic, then Boc-anhydride (2.98 mL, 12.83 mmol) was added at room temperature and the reaction mixture was stirred at room temperature for 4 h. The reaction mixture was treated with water (50 mL) and extracted with ethyl acetate (3 x 100 mL). The combined organic layers were washed with brine (50 mL), dried over anhydrous sodium sulphate, filtered, and concentrated under reduced pressure to obtain the crude product. The crude product was purified by column chromatography using silica gel (230 - 400 mesh) with 0 - 60% ethyl acetate / pet ether to give tertbutyl (3R)-4-(2',6'-dioxo-2,3-dihydrospiro[indene-l,3'-piperidin]-5-yl)-3-methylpiperazine-l- carboxylate 6’ (370.4 mg, 39.0% yield) as an off-white solid.

[0369] LCMS Method 3 : Retention time: 2.243 min, [M+H]+= 414.1.

[0370] Step 5’. Chiral SFC separation of tert-butyl (3R)-4-(2',6'-dioxo-2,3- dihydrospiro[indene-l,3'-piperidin]-5-yl)-3-methylpiperazine-l-carboxylate 7a’ (Peak-1) and tert-butyl (3R)-4-(2',6'-dioxo-2,3-dihydrospiro[indene-l,3'-piperidin]-5-yl)-3- methylpiperazine-l-carboxylate 7b’ (Peak-2): 370 mg of the racemic compound was purified by chiral SFC to obtain tert-butyl (3R)-4-(2',6'-dioxo-2,3-dihydrospiro[indene-l,3'-piperidin]-5-yl)-3- m ethylpiperazine- 1 -carboxylate 7a’ (170 mg, Peak-1) and tert-butyl (3R)-4-(2',6'-dioxo-2,3- dihydrospiro[indene-l,3'-piperidin]-5-yl)-3-methylpiperazine-l-carboxylate 7b’ (170 mg, Peak-2).

[0371] SFC method: YMC Cellulose-SC, 250 mm x 4.6 x 5 pm, Flowrate: 3.0 mL / min (1 : 1 ACN and IP A), Co-Solvent: 40.0%, BPR: 100.0 bar, BPR Temperature: 50 °C.

[0372] Peak 01: RT 6.240 min, ee 100%.

[0373] LCMS method 3: retention time: 2.245 min, [M+H]+= 414.1.

[0374] 1H NMR (400 MHz, CDC13): 8 ppm 1.03 (d, J = 6.4 Hz, 3H), 1.51 (s, 9H), 2.00 - 2.15 (m, 2H), 2.18 - 2.26 (m, 1H), 2.68 - 2.77 (m, 3H), 2.98 - 3.23 (m, 5H), 3.50 - 3.40 (m, 1H), 3.74 - 3.88 (m, 3H), 6.73 - 6.80 (m, 1H), 6.83 (s, 1H), 6.99 - 7.04 (m, 1H), 7.89 (s, 1H).

[0375] Peak 02: RT 7.488 min, ee 99.667%.

[0376] LCMS method 3: retention time: 2.237 min, [M+H]+= 414.1.

[0377] 1H NMR (400 MHz, CDCI3): 6 ppm 0.89 (d, J= 6.4 Hz, 3H), 1.42 (s, 9H), 1.85 - 1.94 (m, 1H), 2.02 - 2.19 (m, 2H), 2.45 - 2.50 (m, 1H), 2.52 - 2.58 (m, 1H), 2.59 - 2.70 (m, 1H), 2.87 - 2.98 (m, 3H), 2.99 - 3.29 (m, 3H), 3.65 - 3.72 (m, 1H), 3.80 - 3.95 (m, 2H), 6.73 (dd, J= 8.4 and2.0 Hz, 1H), 6.80 - 6.85 (m, 1H), 6.97 (d, J= 8.4 Hz, 1H), 10.77 (s, 1H).

[0378] Step 6’. Preparation of 5-((R)-2-methylpiperazin-l-yl)-2,3-dihydrospiro[indene- l,3'-piperidine]-2',6'-dione C17: To a stirred solution of tert-butyl (3R)-4-(2',6'-dioxo-2,3- dihydrospiro[indene-l,3'-piperidin]-5-yl)-3-methylpiperazine-l-carboxylate 7’ (0.150 g, 0.363 mmol) in DCM (5 mL) at room temperature under nitrogen atmosphere was added 4.0 N hydrogen chloride solution in 1,4-di oxane (2.72 mL, 10.88 mmol) dropwise and the reaction mixture was stirred for 3 h at room temperature. The reaction mixture was concentrated under reduced pressure to obtain crude 5-((R)-2-methylpiperazin-l-yl)-2,3-dihydrospiro[indene-l,3'-piperidine]-2',6'-dione, 2 HC1 C-17 (150.0 mg, 90% yield) as an off-white solid. The crude product was used in the next step without further purification.

[0379] LCMS method 3: retention time: 1.754 min, [M+H]+= 314.2.Example S12: Preparation of 6-(piperazin-l-yl)spiro[indoline-3,3'-piperidine]-2',6'-dione (C-

[0380] LCMS Method 1. Kinetex XB - C18, 75 x 3.0 mm, 2.6 pm. Temperature: RT, Flow: 1.0 mL / min, run time: 5.0 min. Mobile Phase Conditions: Mobile Phase-A: 5.0 mm ammonium formate pH 3.3 in CH3CN (98:02), Mobile Phase-B: CH3CN in 5.0 mm ammonium formate (98:02), Gradient: Initial 80% Mobile Phase A and 20% Mobile Phase B linear gradient to 100% Mobile Phase B for 4.5 min. MSD positive.

[0381] LCMS Method 2, Aquity Uplc BEH C18 50 x 3.0 mm, 1.7pm. Temperature: RT, Flow: 0.7 mL / min, run time: 2.0 min. Mobile Phase Conditions: Mobile Phase-A: 0.1% TFA in H2O, Mobile Phase-B: 0.1% TFA in ACN, Gradient: Initial 80% Mobile Phase A and 20% Mobile Phase B linear gradient to 98% Mobile Phase B for 2.0 min. MSD positive.

[0382] LCMS Method 3 , Aquity Uplc BEH C18 50 x 3.0 mm, 1.7pm. Temperature: RT, Flow: 0.7 mL / min, run time: 2.6 min. Mobile Phase Conditions: Mobile Phase-A: 5.0 mm ammonium formate pH 3.3 in CH3CN (98:02), Mobile Phase-B: CH3CN in 5.0 mm ammonium formate (98:02), Gradient: Initial 80% Mobile Phase A and 20% Mobile Phase B linear gradient to 98% Mobile Phase B for 2.0 min. MSD positive.

[0383] Step 1’. Preparation of l-(tert-butyl) 3-methyl 6-bromo-lH-indole-l,3- dicarboxylate 2’: To a stirred solution of methyl 6-bromo-lH-indole-3-carboxylate 1’ (1.3 g, 5.12 mmol) in acetonitrile (10.23 ml) at room temperature under nitrogen atmosphere, DMAP (0.063 g, 0.512 mmol) followed by Boc-anhydride (1.782 ml, 7.67 mmol) were added and the reaction was stirred at room temperature for 3 h. The reaction mixture was treated with water and extracted with ethyl acetate. The combined organic layers were dried over sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography using silica gel (230 - 400 mesh) with 0 - 30% ethyl acetate / pet ether to obtain 1 -(tert-butyl) 3- methyl 6-bromo-lH-indole-l,3-dicarboxylate 2’ (1.7 g, 91% yield) as an off-white solid.

[0384] LCMS Method 1 : Retention time: 1.177 min, [M+H]+= 254.

[0385] Step 2’. Preparation of l-(tert-butyl) 3-methyl 6-bromoindoline-l,3-dicarboxylate 3’: To a stirred solution of 1 -(tert-butyl) 3-methyl 6-bromo-lH-indole-l,3-dicarboxylate 2’ (1.7 g, 4.80 mmol) in MeOH (20 mL) and DCM (5.00 mL) was added magnesium (0.933 g, 38.4 mmol) at 0 °C under nitrogen atmosphere. The reaction mixture was slowly brought to room temperature and stirred of 6 h. The reaction mixture was quenched with aq. ammonium chloride (100 mL) and extracted with ethyl acetate (3 x 50 mL). The combined organic layers were dried over sodium sulfate, filtered, and concentrated under reduced pressure. The crude product 1 -(tert-butyl) 3- methyl 6-bromoindoline-l,3-dicarboxylate 3’ (1.6 g, 94% yield) was used in the next step without further purification.

[0386] LCMS Method 2: Retention time: 1.379 min, [M+H-100]+= 256.0.

[0387] Step 3’. Preparation of l-(tert-butyl) 3-methyl 6-bromo-3-(2-cyanoethyl)indoline- 1,3-dicarboxylate 4’: To a stirred solution of 1 -(tert-butyl) 3-methyl 6-bromoindoline-l,3- dicarboxylate 3’ (1600 mg, 4.49 mmol), acrylonitrile (0.591 mL, 8.98 mmol), and K2CO3 (1242 mg, 8.98 mmol) in toluene (10 mL) was added benzyl triethyl ammonium chloride (102 mg, 0.449 mmol) at room temperature. The reaction mixture was then stirred at 90 °C for 3 h. The crude reaction mixture was treated with water and extracted by ethyl acetate (3 x 50 mL). The combined organic layers were dried over sodium sulphate, filtered, and concentrated. The crude product 1- (tert-butyl) 3-methyl 6-bromo-3-(2-cyanoethyl)indoline-l,3-dicarboxylate 4’ (1.488 g, 81% yield) was used in the next step without further purification.

[0388] LCMS Method 2: Retention time: 1.327 min, [M+H+2-100]+= 311.2.

[0389] Step 4’. Preparation of tert-butyl (3R)-4-(2',6'-dioxo-2,3-dihydrospiro[indene-l,3'- piperidin]-5-yl)-3-methylpiperazine-l-carboxylate 6’: To a stirred solution of 1 -(tert-butyl) 3- methyl 6-bromo-3-(2-cyanoethyl)indoline- 1,3 -dicarboxylate 4’ (1488 mg, 3.64 mmol) and tertbutyl piperazine- 1 -carboxylate 5’ (1016 mg, 5.45 mmol) in 1,4-dioxane (10.0 mL) was added cesium carbonate (2369 mg, 7.27 mmol)), then the reaction mixture was degassed with nitrogen gas for 5 min, followed by addition of RuPhos Pd G4 (309 mg, 0.364 mmol) at room temperature under a positive flow of nitrogen gas. The reaction mixture was then stirred at 100 °C for 16 h. The reaction mixture was treated with water (50 mL) and extracted with ethyl acetate (3 x 100 mL). The combined organic layers were washed with brine (50 mL), dried over anhydrous sodium sulphate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography using silica gel (230 - 400 mesh) with 0 - 60% ethyl acetate / pet ether to obtain 1- (tert-butyl) 3-methyl 6-(4-(tert-butoxycarbonyl)piperazin-l-yl)-3-(2-cyanoethyl)indoline-l,3- dicarboxylate 6’ (1.176 g, 34.6 % yield) as a brown semi-solid.

[0390] LCMS Method 3: Retention time: 1.669 min, [M+H]+= 515.2.

[0391] Step 5’. Preparation of 6-(piperazin-l-yl)spiro[indoline-3,3'-piperidine]-2',6'-dione sulfuric acid salt 7’: To a stirred solution of 1 -(tert-butyl) 3-methyl 6-(4-(tert-butoxy carbonyl) piperazin-l-yl)-3-(2-cyanoethyl)indoline-l,3-dicarboxylate 6’ (1.17 g, 2.274 mmol) in acetic acid (10.0 mL) was added sulfuric acid (0.242 mL, 4.55 mmol) dropwise at room temperature under nitrogen atmosphere and the reaction mixture was stirred at 120 °C for 3 h. The crude reaction mixture was concentrated under reduced pressure and co-distilled with acetonitrile (3 x 20 mL) to obtain crude 6-(piperazin-l-yl) spiro[indoline-3,3'-piperidine]-2',6'-dione sulfuric acid salt 7’ (1.0 g, 74.0% yield) as a brown thick liquid. The crude product was used in the next step without further purification.

[0392] LCMS Method 2: Retention time: 0.44 min, [M+H]+= 301.2.

[0393] Step 6’. Preparation of tert-butyl 6-(4-(tert-butoxycarbonyl)piperazin-l-yl)-2',6'- dioxospiro[indoline-3,3'-piperidine]-l-carboxylate 8’: To a stirred solution of 6-(piperazin-l- yl)spiro[indoline-3,3'-piperidine]-2',6'-dione sulfuric acid salt 7’ (1.0 g, 1.682 mmol) in acetonitrile (10 mL) was added boc-anhydride (0.390 mL, 1.682 mmol) and DIPEA (1.469 mL, 8.41 mmol) at room temperature under nitrogen atmosphere, and the reaction mixture was stirred for 16 h at room temperature. The reaction mixture was treated with water and extracted with ethyl acetate (3x50 mL). The combined organic layers were dried over sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column purification by using silica gel (230- 400 mesh) with 0-80% ethyl acetate / pet ether to obtain tert-butyl 6-(4-(tertbutoxy carbonyl)piperazin-l-yl)-2',6'-dioxospiro[indoline-3,3'-piperidine]-l-carboxylate 8’ (355 mg, 38.4% yield) as an off-white solid.

[0394] LCMS Method 3 : Retention time: 1.494 min, [M+H]+= 501.2.

[0395] Step 7’. Chiral SFC separation of tert-butyl 6-(4-(tert-butoxycarbonyl)piperazin- l-yl)-2',6'-dioxospiro[indoline-3,3'-piperidine]-l-carboxylate 9a' (Peak-1) and tert-butyl 6-(4- (tert-butoxycarbonyl)piperazin-l-yl)-2',6'-dioxospiro[indoline-3,3'-piperidine]-l-carboxylate 9b' (Peak-2): 355 mg of the racemic compound was purified by chiral SFC to obtain tert-butyl 6- (4-(tert-butoxycarbonyl)piperazin-l-yl)-2',6'-dioxospiro[indoline-3,3'-piperidine]-l-carboxylate 9a' (150 mg, Peak-1) and tert-butyl 6-(4-(tert-butoxycarbonyl)piperazin-l-yl)-2',6'-dioxospiro[indoline- 3,3'-piperidine]-l-carboxylate 9b' (160 mg, Peak-2).

[0396] SFC method: (R,R)-WHELK-01 250 mm x 4.6 x 5 pm, Flowrate: 3.0 mL / min (1 : 1 ACN and IP A), Co-Solvent: 40.0%, BPR: 100.0 bar, BPR Temperature: 50 °C.

[0397] Peak 01: RT 2.304 min, ee 100%.

[0398] LCMS method 1 : retention time: 2.895 min, [M+H]+= 501.3.

[0399] 1H NMR (400 MHz, CDC13): 8 ppm 1.42 (s, 9H), 1.52 (br s, 9H), 2.04 - 2.19 (m, 2H), 2.52 - 2.56 (m, 2H), 3.00 - 3.15 (m, 4H), 3.73 - 3.86 (m, 2H), 4.26 (br d, J= 11.01 Hz, 1H), 6.57 (dd, J= 8.25, 2.25 Hz, 1H), 7.04 (d, J= 8.50 Hz, 1H), 7.39 - 7.49 (m, 1H), 10.96 (s, 1H).

[0400] Peak 02: RT 3.723 min, ee 99.8%.

[0401] LCMS method 1 : retention time: 2.899 min, [M+H]+= 501.3.

[0402] 1H NMR (400 MHz, CDCI3): 6 ppm 1.42 (s, 9H), 1.52 (br s, 9H), 2.10 - 2.15 (m, 2H), 2.62 - 2.68 (m, 2H), 3.01 - 3.10 (m, 4H), 3.43 - 3.50 (m, 4H), 3.84 (br d, J= 11.6 Hz, 1H), 4.27 (br d, J = 11.6 Hz, 1H), 6.57 (dd, J= 8.25, 2.25 Hz, 1H), 7.04 (d, J= 8.50 Hz, 1H), 7.35 - 7.50 (m, 1H), 10.96 (s, 1H).

[0403] Step 8’. Preparation of 5-((R)-2-methylpiperazin-l-yl)-2,3-dihydrospiro[indene- l,3'-piperidine]-2',6'-dione C-19: To a stirred solution of tert-butyl 6-(4-(tert- butoxycarbonyl)piperazin-l-yl)-2',6'-dioxospiro[indoline-3,3'-piperidine]-l-carboxylate 9a’ (150 mg, 0.300 mmol) in DCM (10 mL) was added hydrochloric acid in dioxane (0.375 mL, 1.498 mmol) and stirred for 3 h at room temperature. The reaction mixture was concentrated under reduced pressure to obtain crude 6-(piperazin-l-yl)spiro[indoline-3,3'-piperidine]-2',6'-dione HC1 10’ (145 mg, 116% yield) as off-white solid. The crude product was used in the next step without further purification.

[0404] LCMS method 1 : retention time: 0.993 min, [M+H]+= 301.2.Synthesis of the final compound with the TBM and CBM moleculesExample S13: Preparation of l-(4-(((R)-l-cyanoethyl)amino)-5-(4-((1r,4R)-4-(2-(4-(2',6'- dioxo-2Hspiro[benzofuran-3,3'-piperidin]-6-yl)piperazin-l-yl)ethyl)cyclohexyl)-lH-l,2,3- triazol-l-yl)pyrid ine-2-yl)-lH-pyrazolo[3,4-b]pyridine-5-carbonitrile P-1.

[0405] LCMS Method 1. Kinetex XB - C18, 75 x 3.0 mm, 2.6 pm. Temperature: RT, Flow: 1.0 mL / min, run time: 5.5 min. Mobile Phase Conditions: Mobile Phase-A: 0.1% TFA in H2O, Mobile Phase-B: 0.1% TFA in ACN, Gradient: Initial 95% Mobile Phase A and 5% Mobile Phase B linear gradient to 95% Mobile Phase B for 4.0 min. MSD positive.

[0406] Step 1’. Preparation of l-(4-(((R)-l-cyanoethyl)amino)-5-(4-((1r,4R)-4-(2-(4-(2',6'- dioxo-2Hspiro[benzofuran-3,3'-piperidin]-6-yl)piperazin-l-yl)ethyl)cyclohexyl)-lH-l,2,3- triazol-l-yl)pyrid ine-2-yl)-lH-pyrazolo[3,4-b]pyridine-5-carbonitrile P-1: To a stirred solution of l-(4-(((R)-l-cyanoethyl)amino)-5-(4-((1r,4R)-4-(2-oxoethyl)cyclohexyl)-lH-l,2,3-triazol-l- yl)pyridin-2-yl)-lH-pyrazolo[3,4-b]pyridine-5-carbonitrile A-l (100 mg, 0.169 mmol) in DMSO (3.0 mL) was added 6-(piperazin-l-yl)-2H-spiro[benzofuran-3,3'-piperidine]-2',6'-dione, 2 HC1 C-l (67.6 mg, 0.177 mmol). The reaction mixture was stirred at room temperature for 30 min and sodium triacetoxyborohydride (107 mg, 0.506 mmol) was added under nitrogen atmosphere, then the reaction mixture was stirred at room temperature for 1 h. The reaction mixture was treated with water and extracted with ethyl acetate, and the combined organic layers were dried over sodium sulphate, filtered, and concentrated under reduced pressure. The crude compound was purified by Prep-HPLC to obtain l-(4-(((R)-l-cyanoethyl)amino)-5-(4-((1r,4R)-4-(2-(4-(2',6'-dioxo- 2Hspiro[benzofuran-3,3'-piperidin]-6-yl)piperazin-l-yl)ethyl)cyclohexyl)-lH-l,2,3-triazol-l- yl)pyridin-2-yl)-lH-pyrazolo[3,4-b]pyridine-5-carbonitrile P-1 (30 mg, 17.28% yield) as an off- white solid.

[0407] Prep-HPLC method: X-Bridge C18, 150 x 19 mm, 5 pm, Mobile Phase-A: 0.1% TFA in H2O, Mobile Phase-B: ACN, Flow rate: 15.0 mL / min.

[0408] LCMS method 1 : retention time: 2.123 min, [M+H]+= 766.2.

[0409] 1H NMR (400 MHz, DMSO-t / e): 8 ppm 1.12-1.28 (m, 2H), 1.48 (s, 1H), 1.45-1.59 (m, 2H), 1.61 (d, 6.80 Hz, 3H), 1.65 (m, 2H), 1.84-1.90 (m, 2H), 2.17-2.13 (m, 4H), 2.66-2.68 (m,1H), 2.73-2.81 (m, 2H), 2.97 (t, J= 8.8 Hz, 2H), 3.12 (q, J= 8.0 Hz, 2H), 3.25-3.23 (m, 2H), 3.58 (d, J= 11.60 Hz, 2H), 3.84 (d, J= 12.80 Hz, 2H), 4.43 (d, J= 9.20 Hz, 1H), 4.82 (d, J= 9.20 Hz, 1H), 5.02 (t, J= 7.2 Hz, 1H), 6.57-6.50 (m, 2H), 7.20-7.08 (m, 2H), 7.67 (s, 1H), 8.36 (s, 1H), 8.41(s, 1H), 8.72 (s, 1H), 9.06 (d, J= 2.00 Hz, 2H), 10.98 (s, 1H).

[0410] Table 3 summarizes the compounds prepared using the procedure described in Example S13.able 3. Final Compounds Prepared via General Procedure Shown in Example S13Biological ExamplesExample Bl. Reagent Preparations

[0411] Cell culture media was prepared in a tissue culture hood in a sterile environment by adding 10% FBS and 1% Penicillin Streptomycin to 500 mL no phenol red RPMI 1640 media. The media was filtered through a Nalgene Bottle Top Filter and stored at 4 °C.

[0412] The Cell titer Gio (CTG) buffer and substrate (CellTiter-Glo Luminescent Cell Viability Assay, Promega Ref.# G7573) were stored in -20°C. The CTG buffer (100 mL) was warmed in a bead bath and added to the CTG substrate bottle in a tissue culture hood. The solution was mixed with a pipette until it became homogenous. CTG reagent were aliquoted into 15 mL falcon tubes and stored at -20 °C.

[0413] For Homogenous Time Resolved Fluorescence (HTRF) assays, a Cisbio HTRF kit was used, which included: Lysis Buffer #1 4X, Blocking Reagent #3 100X, 20X Antibody 1 (Anti-IRAK4 d2), 20X Antibody 2 (Anti-IRAK4 k), and Detection Buffer.

[0414] 4X Lysis Buffer was stored at 4 °C. For use as IX Lysis buffer, the 4X solution was diluted with de-ionized water (distilled water, Gibco Cat.# 15230279) and 100X Blocking Reagent in a 1 :3 :0.04 volume ratio.

[0415] 20X Antibody Solution aliquots were stored in -80 °C and the Detection Buffer was stored in 4 °C. For use as a IX Antibody Solution, the 20X Antibody Solution aliquot was diluted with Detection Buffer in a 1 : 19 volume ratio.Example B2. THP1 Homogeneous Time Resolved Fluorescence (THP1 HTRF) Procedure

[0416] The THP1 plates were prepared by the following method. For each dosing plate, a duplicate for CTG assay was prepared. Two THP-1 T-175 flasks could be used to plate one 384 well plate. The cells were collected from two T-175 flasks into 200 mL centrifuge tubes and spun down at 1200 rpm for 10 min. The supernatant was removed completely and cells were resuspended in 7.5 mL cell culture media. To 5 pL of the cell suspension was added 25 pL of media and 30 pL of Trypan Blue stain. The cells were counted using countess (automated cell counter, Thermo Fisher Scientific) twice and the viability and live cell count was recorded. The actual cell count was 6X the live cell count. Based on the count, a cell suspension was prepared in colorless RPMI media for a final concentration of 7.5e6 cells / mL. Using Standard cassette multidrop combi, 20 pL of the 7.5e6 cells / mL cell stock was added to entire 384 well plate but for top half of column 1 (negative control) The multidrop combi was always primed to achieve a steady flow and washed after use with 20 mL de-ionized water followed by 20 mL alcohol. The cells were incubated in 5% CO2 incubator at 37 °C

[0417] Dosing was conducted using the following method. To dose assay plates and CTGplate was added 20 nL of compounds in DMSO using Echo (Beckman Coulter), and the plates were incubated at 37 °C for 18 h. The compound plates were spun at 1200 rpm for 1 min before dosing. The compound plate was sealed and properly stored during incubation.

[0418] After dosing, an HTRF assay was conducted using the following method. The treatment plates (IRAK4 HTRF) were centrifuged at 600 g for 10 min. Using CyBio Felix , Automated liquid handler (Analytikjena), 13 pL cell culture media was removed. To all columns, 7.5 pL lysis buffer was added, and an additional 7.5 pL of lysis buffer was added to column l(top half, negative control). The plate was incubated on a shaker at room temperature for 2 h. The plate was spun down at 600 g for 5 min. Custom 20X IRAK4 Antibody obtained from CISBIO Anti-IRAK4-d2 (acceptor Ab) and Anti-IRAK4-K (donor Ab) were diluted to IXantibody mix Using a multichannel repeater, 4 pL of IX antibody mix was added to each wellThe plate was spun down at 600 g for 5 min and incubated in the dark at room temperature for 18 h The next day (after 18hrs), HTRF assay plate was spun down at 600 g for 5 min and read at 665 / 615 nm using a Envision plate reader.

[0419] Next, a CTG assay was run to assess cell death using the following method. To all wells 20 pL of CTG reagent was added, and the plate was covered The plate was placed on a shaker for 1-2 min and was let to sit at room temperature in the dark for 20 min (no more than 30 min). The plate was spun down at 1200 rpm for 1 min and the luminescence was analyzed using a plate reader (Envision, PerkinElmer).

[0420] A summary of the THP1 HTRF data for the tested compounds is provided in Table 4 below.Table 4. THP1 HTRF Results of the CompoundsN.D. = not determined

[0421] Although the present invention has been described in some detail by way of illustration and example for purposes of clarity of understanding, the descriptions and examples should not be construed as limiting the scope of the invention. The disclosures of all patent and scientific literature cited herein are expressly incorporated herein in their entirety by reference.

Claims

CLAIMS1. A compound of F ormula (I)or a pharmaceutically acceptable salt thereof, wherein:Ring B is 5- to 6-membered monocyclic heterocyclylene or 8- to 10-membered fused bicyclic heterocyclylene, wherein the heterocyclylene contains 1-3 nitrogen atoms; each R2is independently Ci-Ce alkyl or halo; n is 0-3;R3is H, Ci-Ce alkyl, or halo;Ringattached to two adjacent carbon atoms indicated with *; andX is O, CH2, or N(H).

2. The compound of claim 1, or a pharmaceutically acceptable salt thereof, wherein the compound is of Formula (I’):3 The compound of claim 1, or a pharmaceutically acceptable salt thereof, wherein the compound is of Formula (II):4 The compound of claim 1, or a pharmaceutically acceptable salt thereof, wherein the compound is of Formula (III):

5. The compound of any one of claims 1-3, or a pharmaceutically acceptable salt thereof, wherein:R1is H.

6. The compound of any one of claims 1-3, or a pharmaceutically acceptable salt thereof, wherein:R1is -NH2.7 The compound of any one of claims 1-6, or a pharmaceutically acceptable salt thereof, wherein:L1is -C(O)N(H)-.8 The compound of any one of claims 1-6, or a pharmaceutically acceptable salt thereof, wherein:9 The compound of any one of claims 1-8, or a pharmaceutically acceptable salt thereof, wherein:Ring B is 6-membered monocyclic heterocyclylene or 8-membered fused bicyclic heterocyclylene, wherein the heterocyclylene contains 1-2 nitrogen atoms; and each R2is independently C1-C3 alkyl or halo.10 The compound of any one of claims 1-9, or a pharmaceutically acceptable salt thereof, wherein:

11. The compound of any one of claims 1-10, or a pharmaceutically acceptable salt thereof, wherein:R3is H or C1-C3 alkyl.

12. The compound of any one of claims 1-11, or a pharmaceutically acceptable salt thereof, wherein:X is O.

13. The compound of any one of claims 1-11, or a pharmaceutically acceptable salt thereof, wherein:X is CH2or N(H).

14. The compound of any one of claims 1-13, or a pharmaceutically acceptable salt thereof, wherein:

15. The compound of any one of claims 1-14, or a pharmaceutically acceptable salt thereof, wherein the compound is of Formula (IIA), (IIB), (IIC), (IID), (HE), (HF), (IIIA), (IHB), (IIIC), (HID), (IIIE), or (IHF):The compound of claim 15, or a pharmaceutically acceptable salt thereof, wherein: is O.

17. A compound selected from the compounds of Table 1 and pharmaceutically acceptable salts thereof.

18. A pharmaceutical composition comprising the compound of any one of claims 1-17, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable excipient.

19. A method of modulating interleukin-1 (IL1) receptor-associated kinase 4 (IRAK4) activity comprising contacting IRAK4 with an effective amount of the compound of any one of claims 1-17, or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition of claim 18.

20. A method of treating an inflammatory or autoimmune disease in a subject in need thereof, comprising administering to the subject an effective amount of the compound of any one of claims 1-17, or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition of claim 18, optionally wherein the inflammatory or autoimmune disease is atopic dermatitis, asthma, lupus, rheumatoid arthritis, familial mediterranean fever, psoriasis, generalized pustular psoriasis, cryoprin-associated periodic syndrome, hidradenitis suppurativa, Bechet’s syndrome, or familial cold autoinflammatory syndrome.

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