Ligand directed degrader of IRAK4

Abstract

Provided herein is a compound of the following structural formula: or a pharmaceutically acceptable salt thereof, and compositions thereof. In some embodiments, the compound and compositions are provided for modulating IRAK4 and / or treating inflammatory or autoimmune diseases.

Description

LIGAND DIRECTED DEGRADER OF IRAK4CROSS-REFERENCE TO RELATED APPLICATION

[0001] This application claims the benefit of US Provisional Application No. 63 / 620,227, filed January 12, 2024, which is incorporated by reference herein in its entirety for any purpose.FIELD

[0002] The present disclosure relates generally to compounds, compositions, and methods for the 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-related conditions and diseases associated with IRAK-mediatedsignal 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, and degraded.

[0007] Accordingly, there is a need for 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. The present embodiments can be understood more fully by reference to the detailed description and examples, which are intended to exemplify non-limiting embodiments. Provided herein is a compound of the following structural formula:or a pharmaceutically acceptable salt thereof.

[0009] Also provided herein is a compound of the following structural formula:or a pharmaceutically acceptable salt thereof.

[0010] Also provided herein is a compound of the following structural formula:or a pharmaceutically acceptable salt thereof.

[0011] Also provided herein is a pharmaceutical composition comprising a compound of Formula I, la, or P-1, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable excipient.

[0012] Also provided herein 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 Formula I, la, or P-1 (e.g., an effective amount of a compound of Formula I, la, or P-1), or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition comprising the compound of Formula I, la, or P-1, or a pharmaceutically acceptable salt thereof.

[0013] Also provided herein 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 Formula I, la, or P-1, or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition comprising the compound of Formula I, la, or P-1, or a pharmaceutically acceptable salt thereof, 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.

[0014] Also provided herein is a compound of Formula I, la, or P-1, or a pharmaceutically acceptable salt thereof, or a composition (e.g., pharmaceutical composition) of any of the foregoing, for a use described herein (e.g., modulating IRAK4 activity; treating an inflammatory or autoimmune disease). Also provided herein is use of a compound of Formula I, la, or P-1, ora pharmaceutically acceptable salt thereof, or a composition (e.g., pharmaceutical composition) of either of the foregoing, in a method described herein (e.g., modulating IRAK4 activity; treating an inflammatory or autoimmune disease). Also provided herein is use of a compound of Formula I, la, or P-1, or a pharmaceutically acceptable salt thereof, or a composition (e.g., pharmaceutical composition) of either of the foregoing, in the manufacture of a medicament for modulating IRAK4 activity and / or treating an inflammatory or autoimmune disease.DETAILED DESCRIPTIONDefinitions

[0015] 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 thFereof. 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.

[0016] 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.

[0017] 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 and C; 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.

[0018] 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.

[0019] 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, la, or P-1.

[0020] 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’) or (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) or Remington: The Science and Practice of Pharmacy, 19theds., Mack Publishing, Easton PA (1995).

[0021] 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 the compound. 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.

[0022] 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).

[0023] 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.

[0024] 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:

[0025] As readily understood by one skilled in the art, a wide variety of functional groups and other stuctures may exhibit tautomerism, and all tautomers of compounds provided herein, such as compounds of Formula I, e.g., Formula la or P-1, are within the scope of the present disclosure.

[0026] 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 be radiolabeled 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 encriched 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.

[0027] 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.

[0028] 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.

[0029] “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 (e.g., spread) 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.

[0030] “Preventing” as used herein, means a method of delaying and / or precluding the onset or recurrence, 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.

[0031] 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.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 S1P5 mediated disease, or a symptom thereof.

[0032] Although various features of the disclosure may be described in the context of a single embodiment, the features may also be provided separately or in any suitable combination. Conversely, although various features of the disclosure may be described herein in the context of separate embodiments for clarity, the various features of the disclosure may also be implemented in a single embodiment.Compounds

[0033] In one aspect, provided herein is a compound of Formula I:or a pharmaceutically acceptable salt thereof.

[0034] In another aspect, provided herein is a compound of Formula la:or a pharmaceutically acceptable salt thereof.

[0035] In yet another aspect, provided herein is a compound of Formula P-1 :or a pharmaceutically acceptable salt thereof. Used herein, “orl” indicates that the absolute stereochemistry at the indicated atom was not determined and the stereochemistry may be as drawn or the opposite of the stereochemistry as drawn. Thus, for example, Formula P-1 means

[0036] In another aspect, provided herein is a compound of Formula P-la:la), or a pharmaceutically acceptable salt thereof.

[0037] In another aspect, provided herein is a compound of Formula P-lb:-lb), or a pharmaceutically acceptable salt thereof.

[0038] 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.

[0039] Furthermore, all compounds, e.g., 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 compounds, e.g., of Formula I, can be converted to their free base or acid form by standard techniques.Methods of Synthesis

[0040] 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 Scheme 1, 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.

[0041] Compounds of Formula L-l can be prepared as outlined in Scheme 1. Protection of an alcohol group of intermediate Sl-1’ with a silyl protecting group, such as TBDPS, forms intermediate Sl-2’, which is then activated, e.g., with mesyl chloride, to form intermediate SIS’. The activated alcohol of intermediate Sl-3’ can be replaced with a cyano to form intermediate Sl-4’, and then reduced to an aldehyde to form intermediate Sl-5’. Further reduction affords intermediate Sl-6'. The alcohol group of intermediate Sl-6' can be protected, e.g., with THP, to form intermediate Sl-7’, and subsequent deprotection under acidic conditions forms intermediate Sl-8’. Intermediate Sl-8’ is then oxidized to form intermediate Sl-9’. The aldehyde of intermediate Sl-9’ can be converted to an alkyne via Seyferth-Gilbert homologation to form intermediate L-l.Scheme 1.Seyferth-Gilbert HomologationStep S1-9'57 %

[0042] Compounds of Formula A-l can be prepared as outlined in Scheme 2. Addition of S2-2’ to S2-1’ led to intermediate S2-3’. Dehydration of S2-3’ produced nitrile S2-4’. Cross coupling of S2-4’ with S2-5’ yielded S2-6’, which was reduced to produce S2-7’. Azide formation led to intermediate S2-8’ and cycloadditon of S2-8’ with L-1 produced intermediate S2-9’. Deprotection of S2-9’ followed by oxidation yielded intermediate A-l, comprising the target binding moiety (TBM).Scheme 2.

[0043] Compounds of Formula B-l can be prepared as outlined in Scheme 3. Methylation of 1’ under basic conditions yielded 2’. Conjugate addition of 3’ with 2’ produced intermediate 4’. Intermediate 6’ was formed by cross couping of 4’ with 5’. Subsequent deprotection and cyclization produced 7’. Boc protection and SFC seperation of the diastereomers yielded 9a’ (peak one) and 9b’ (peak two). Deprotection of the protecting group on 9a’ provided Cl.Scheme 3,4N HCI dioxane

[0044] Compounds of Formula I, such as compounds of Formula P-1, can be formed by reductive amination of a compound of Formula Al and a compound of Formula Cl, as outlined in Scheme 4.Attorney Docket No. 01277-0057-00PCScheme 4,Methods of Use

[0045] 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 disclosedcompound disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof. 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).

[0046] In one aspect, provided herein is a method of modulating IRAK4 comprising contacting IRAK4 with a compound disclosedcompound disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof (e.g., an effective amount of a compound described herein). In some embodiments, the compound inhibits IRAK4. In other embodiments, the compound activates IRAK4. In some embodiments, the compound is an agonist of IRAK4. In some embodiments, the compound is an antagonist of IRAK4.

[0047] In some embodiments, provided herein is a method for targeting IRAK4 for degradation comprising contacting IRAK4 with a compound disclosedcompound disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof (e.g., an effective amount of a compound disclosed herein).

[0048] In some embodiments, a compound disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, 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 disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, 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%.

[0049] 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 disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof,. Degradation of IRAK4 can be assessed and demonstrated by a wide variety of ways known in the art. Kits and commercially availableassays, including cell-based assays, can be utilized for determining whether and to what degree IRAK4 has been degraded.

[0050] In one aspect, provided herein is a method of degrading IRAK4 comprising contacting IRAK4 with a compound disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof (e.g., an effective amount of a compound described herein). In some embodiments, the compound partially degrades IRAK3. In some embodiments, the compound fully degrades IRAK4.

[0051] In some embodiments, a compound disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, 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 disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, 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%.

[0052] 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 disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof,. 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 disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof,. 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 disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof,. 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 disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof,. 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 disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof,. 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 compounddisclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof,. 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, hi dradenitis suppurativa, Bechet’s syndrome, or familial cold autoinflammatory syndrome.

[0053] In some embodiments, administering a compound disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, 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 disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, 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 disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, to a subject in need thereof diminishes the extent of the inflammatory or autoimmune disease in the subject. In some embodiments, administering a compound disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, 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 disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, to a subject in need thereof delays the occurrence or recurrence of the inflammatory or autoimmune disease. In some embodiments, administering a compound disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, to a subject in need thereof slows the progression of the inflammatory or autoimmune disease. In some embodiments, administering a compound disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, to a subject in need thereof provides a partial remission of the inflammatory or autoimmune disease. In some embodiments, administering a compound disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, to a subject in need thereof provides a total remission of the inflammatory or autoimmune disease. In some embodiments, administering a compound disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, 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 disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, to a subject in need thereof enhances the effect ofanother medication used to treat the inflammatory or autoimmune disease. In some embodiments, administering a compound disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, to a subject in need thereof delays the progression of the inflammatory or autoimmune disease. In some embodiments, administering a compound disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, 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 disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, to a subject in need thereof prolongs survival of a subject having an inflammatory or autoimmune disease.

[0054] 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 disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, 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 symptoms of the inflammatory or autoimmune disease, the method comprising administering a compound disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, to the subject.

[0055] 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 disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, 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 disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, 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.

[0056] 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 disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, to the subject.

[0057] 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 disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptablesalt thereof, 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.

[0058] 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 disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, 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 disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, to the subject.

[0059] 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 disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, 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.

[0060] 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 disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof,. 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 disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof,. In some embodiments, administering a compound disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, to a subject that is predisposed 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 disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, 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 disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, to a subject in need thereof diminishes the extent ofthe inflammatory or autoimmune symptoms caused by the disease in the subject. In some embodiments, administering a compound disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, 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 disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, 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 disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, 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 disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, 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 disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, 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 disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, 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 disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, 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 disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, to a subject in need thereof delays the progression of the disease which causes inflammatory or autoimmune symptoms. In some embodiments, administering a compound disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, 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 disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, 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 pustularpsoriasis, cryoprin-associated periodic syndrome, hi dradenitis suppurativa, Bechet’s syndrome, or familial cold autoinflammatory syndrome.

[0061] In some embodiments, compounds described herein 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, hi dradenitis suppurativa, Bechet’s syndrome, and familial cold autoinflammatory syndrome.

[0062] In some embodiments, compounds described herein 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’) or (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.

[0063] In some embodiments, the cancer is breast cancer, colorectal cancer, non-small cell lung cancer, ovarian, renal, sarcoma, melanoma, head and neck, hepatocellular, thyroid, multidrug-resistant 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).

[0064] In some embodiments, the compounds described herein 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

[0065] 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.

[0066] 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 in the pharmaceutical composition may be at a level that will exercise the desired effect; for example, about 0.005 mg / kg of a subject’s body weight to about 10 mg / kg of a subject’s body weight in unit dosage for both oral and parenteral administration.

[0067] The dose of a compound disclosed herein to be administered to a subject is rather widely variable and can be subject to the judgment of a health-care practitioner. In general, the compounds disclosed herein can be administered one to four times a day in a dose of about 0.001 mg / kg of a subject’s body weight to about 10 mg / kg of a subject’s body weight, but the above dosage may be properly varied depending on the age, body weight and medical condition of the subject and the type of administration. In one embodiment, the dose is about 0.001 mg / kgof a subject’s body weight to about 5 mg / kg of a subject’s body weight, about 0.01 mg / kg of a subject’s body weight to about 5 mg / kg of a subject’s body weight, about 0.05 mg / kg of a subject’s body weight to about 1 mg / kg of a subject’s body weight, about 0.1 mg / kg of a subject’s body weight to about 0.75 mg / kg of a subject’s body weight or about 0.25 mg / kg of a subject’s body weight to about 0.5 mg / kg of a subject’s body weight. In one embodiment, one dose is given per day. In any given case, the amount of the compound administered will depend on such factors as the solubility of the active component, the formulation used and the route of administration.

[0068] In some embodiments, a compound disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, is administered to a subject at a dose of about 0.01 mg / day to about 750 mg / day, about 0.1 mg / day to about 375 mg / day, about 0.1 mg / day to about 150 mg / day, about 0.1 mg / day to about 75 mg / day, about 0.1 mg / day to about 50 mg / day, about 0.1 mg / day to about 25 mg / day, or about 0.1 mg / day to about 10 mg / day.

[0069] In another embodiment, provided herein are unit dosage formulations that comprise between about 0.1 mg and 500 mg, about 1 mg and 250 mg, about 1 mg and about 100 mg, about 1 mg and about 50 mg, about 1 mg and about 25 mg, or between about 1 mg and about 10 mg of a compound disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof.

[0070] In a particular embodiment, provided herein are unit dosage formulations comprising about 0.1 mg or 100 mg of a compound disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof.

[0071] In another embodiment, provided herein are unit dosage formulations that comprise 0.5 mg, 1 mg, 5 mg, 10 mg, 15 mg, 20 mg, 30 mg, 35 mg, 50 mg, 70 mg, 100 mg, 125 mg, 140 mg, 175 mg, 200 mg, 250 mg, 280 mg, 350 mg, 500 mg, 560 mg, 700 mg, 750 mg, 1000 mg or 1400 mg of a compound disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof.

[0072] A compound disclosed herein, e.g. a compound of Formula I, or a pharmaceutically acceptable salt thereof, can be administered once, twice, three, four or more times daily. In a particular embodiment, doses of 100 mg or less are administered as a once daily dose and doses of more than 100 mg are administered twice daily in an amount equal to one half of the total daily dose.

[0073] A compound disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof can be administered orally for reasons of convenience. In one embodiment, when administered orally, a compound disclosed herein, e.g., a compound ofFormula I, or a pharmaceutically acceptable salt thereof, is administered with a meal and water. In another embodiment, a compound disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, 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.

[0074] 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.

[0075] In one embodiment, provided herein are capsules containing a compound disclosed herein without an additional carrier, excipient or vehicle.

[0076] In another embodiment, provided herein are compositions comprising an effective amount of a compound disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, 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.

[0077] 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 disclosed herein 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.

[0078] 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 andthe 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.

[0079] 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 film-forming 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.

[0080] When it is desired to administer a compound disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof 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.

[0081] The effect of the compound a compound disclosed herein, e.g., a compound of Formula I, or a pharmaceutically acceptable salt thereof, can be delayed or prolonged by proper formulation. For example, a slowly soluble pellet of the compound 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 in oily or emulsified vehicles that allow it to disperse slowly in the serum.EXAMPLES

[0082] The following Examples are presented by way of illustration, not limitation. Compounds are named using the automatic name generating tool provided in ChemBiodrawUltra (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.

[0083] 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).

[0084] 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.

[0085] The following abbreviations may be relevant for the application.AbbreviationsAc: acetateACN or MeCN: acetonitrileADMP: 2-azido-l,3-dimethylimidazolinium hexafluorophosphate aq: aqueousCBM: cereblon binding moietyC V : column volume d: day(s) dba: dibenzylideneacetoneDCM: dichloromethaneDHP: dihydropyranDiBAl: diisobutylaluminumDIPEA: N, N-diisopropylethylamineDMAP: 4-dimethylaminopyridineDMP: Dess-Martin PeriodinaneDMSO: Dimethyl sulfoxideEquiv or eq. : equivalentsESI: electrospray ionizationEt: ethylEtOAc: ethyl acetateFA: formic acidFBS: fetal bovine serumFC: flash chromatography h: hour(s)HPLC: high-performance liquid chromatographyIBX: 2-iodoxybenzoic acidLCMS: liquid chromatography mass spectrometryMeOH: methanol min: minute(s)Ms: mesylMSD: mass selective detectorPet ether: petroleum etherPS: penicillin-streptomycinPPTS: pyridinium p-toluenesulfonatePTSA: p-toluenesulfonic acid prep: preparative quant.: quantitativeRT or r.t.: room temperature rt: retention time sat.: saturatedSFC: supercritical-fluid chromatographyTBAF : tetrabutylammonium fluorideTBDPS: tert-butyldiphenylsilylTBM: target binding moietyTEA: triethylamineTFA: trifluoroacetic acidTHF : tetrahydrofuranTHP: tetrahydropyranTLC: thin layer chromatographyUPLC: ultra-performance liquid chromatographySynthetic ExamplesExample SI: Preparation of 2-(2-((lr,4r)-4-ethynylcyclohexyl)ethoxy)tetrahydro-2H-pyran

[0086] Step Sl-1’. Preparation of ((lr,4r)-4-(((tertbutyldiphenylsilyl)oxy)methyl)cyclohexyl)methanol Sl-2’: To a solution of [4-(hydroxymethyl)cyclohexyl]methanol Sl-1’ (3 g, 20.8 mmol, 1 eq.) in dry DMF (83.2 mL, 0.25M) was added imidazole (0.85 g, 12.48 mmol, 0.6 eq.), followed by tertbutylchlorodiphenylsilane (2.7 mL, 10.4 mmol, 0.5 eq.). After stirring for 18 h at room temperature, TLC (25 % EtOAc in heptanes, revealing UV + KMnO4) 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 Sl-2’(6.48 g, 43 % yield) as a colorless oil. 'H 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).

[0087] Step Sl-2’. Preparation of ((lr,4r)-4-(((tertbutyldiphenylsilyl)oxy)methyl)cyclohexyl)methyl methanesulfonate Sl-3’: To a round bottom flask was added ((lr,4r)-4-(((tert-butyldiphenylsilyl)oxy)methyl)cyclohexyl)methanolSl-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 (KMnCU 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 Sl-3’ (3.65 g, 89 % yield) as a colorless solid. The crude was used directly as is for the next reaction.JH 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 by 'H NMR.

[0088] Step Sl-3’. Preparation 2-((lr,4r)-4-(((tert- butyldiphenylsilyl)oxy)methyl)cyclohexyl)acetonitrile Sl-4’: To a round bottom flask were added (( l r,4r)-4-((( / c / 7-butyldiphenylsilyl)oxy)methyl)cyclohexyl)methyl methanesulfonate SIS’ (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 Sl-4’ (KMnO4 revelator). The reacting mixture was poured into an Ehrlenmeyer of crushed ice and stirred until all ice was melted, the resulting solid was isolated by filtration and dissolved in a minimum of ethyl acetate, washed with NaHCCh 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 Sl-4’ (2.35 g, 76 % yield) as a colorless oil. 'H NMR (400 MHz, chloroform-d): 5 ppm 0.83 - 0.91 (m, 1 H), 0.98 - 1.05 (m, 2 H), 1.05 - 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= 11.9 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).

[0089] Step Sl-4’. Preparation of 2-((lr,4r)-4-(((tert- butyldiphenylsilyl)oxy)methyl)cyclohexyl)acetaldehyde Sl-5’: To a round bottom flask was added 2-((lr,4r)-4-(((tert-butyldiphenylsilyl)oxy)methyl)cyclohexyl)acetonitrile Sl-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 werewashed with water, dried over magnesium sulfate and concentrated under reduced pressure. The residue was dry-packed and purified by normal phase flash chromatography (40 g silica column, elution: 1 :3 Hept.:EtOAc). Fractions were combined and concentrated to give Sl-5’ (803 mg, 34 % yield) as a colorless oil that was used as is for the next reaction.

[0090] Step Sl-5’. Preparation of 2-((lr,4r)-4-(((tert- butyldiphenylsilyl)oxy)methyl)cyclohexyl)ethan-l-ol Sl-6’: To a round bottom flask was added 2-((lr,4r)-4-(((tert-butyldiphenylsilyl)oxy)methyl)cyclohexyl)acetaldehyde Sl-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 NaBEE (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 Sl-6’ (353.7 mg, 44 % yield) as a colorless oil. 'H 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).

[0091] Step Sl-6’. Preparation of tert-butyldiphenyl(((lr,4r)-4-(2-((tetrahydro-2H- pyran-2-yl)oxy)ethyl)cyclohexyl)methoxy)silane Sl-7’: To a solution of 2-((lr,4r)-4-(((tert- butyldiphenylsilyl)oxy)methyl)cyclohexyl)ethan-l-ol Sl-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 Sl-7’ (1.79 g, 91 % yield) as a colorless oil. ‘HNMR (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).

[0092] Step Sl-7’. Preparation of ((lr,4r)-4-(2-((tetrahydro-2H-pyran-2- yl)oxy)ethyl)cyclohexyl)methanol Sl-8’: To a stirred solution of tert-butyldiphenyl(((lr,4r)-4- (2-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)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, 4eq.) at room temperature. After 5 h, TLC showed complete conversion of the starting material Sl-7’. Solvents were removed under reduced pressure 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 Sl-8’ (577.5 mg, 58 % yield) as a colorless oil.JH 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).

[0093] Step Sl-8’. Preparation of (lr,4r)-4-(2-((tetrahydro-2H-pyran-2- yl)oxy)ethyl)cyclohexane-l-carbaldehyde Sl-9’: To a stirred solution of ((lr,4r)-4-(2- ((tetrahydro-2H-pyran-2-yl)oxy)ethyl)cyclohexyl)methanol Sl-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 Sl-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 Sl-9’ (459.5 mg, 66 % yield) as a colorless solid. 'H NMR (400 MHz, DMSO-de): 8 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).

[0094] Step Sl-9’. Preparation of 2-(2-((lr,4r)-4-ethynylcyclohexyl)ethoxy)tetrahydro- 2H-pyran (L-l): To a flame-dried round bottom flask were added (lr,4r)-4-(2-((tetrahydro-2H- pyran-2-yl)oxy)ethyl)cyclohexane-l-carbaldehyde Sl-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 Sl-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. 'H 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).Example S2: Preparation of l-(4-(((R)-l-cyanoethyl)amino)-5-(4-((lr,4R)-4-(2- oxoethyl)cyclohexyl)-l H-l ,2,3-triazol-l-yl)pyridin-2-yl)-l H-pyrazolo[3,4-b]pyridine-5- carbonitrile (A-l)

[0095] LCMS Method S2-1. 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.

[0096] UPLC Method S2-2. 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.

[0097] Step S2-1’. Preparation of (l?)-2-((2-chloro-5-nitropyridin-4- yl)amino)propanamide S2-3’: To a stirred solution of 2,4-dichloro-5-nitropyridine S2-1’ (20.0 g, 104 mmol, 1.0 eq.) in acetonitrile (200 mL) was added (A)-2-aminopropanamide hydrochloride S2-2’ (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 concentrate under reduced pressure to afford (A)-2-((2-chloro-5- nitropyridin-4-yl)amino)propanamide S2-3’ (27 g, 97% yield) as a pale yellow solid. LCMS Method S2-L retention time: 0.876 min, 91.45% purity at 220 nm, [M+H]+= 245.0.JH NMR (300 MHz, DMSO-de): 8 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).

[0098] Step S2-2’. Preparation of (l?)-2-((2-chloro-5-nitropyridin-4- yl)amino)propanenitrile S2-4’: To a stirred solution of (A)-2-((2-chloro-5-nitropyridin-4- yl)amino)propanamide S2-3’ (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 the 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-chl oro-5 - nitropyridin-4-yl)amino)propanenitrile S2-4’ (21 g, 89% yield) as a pale yellow solid. LCMS Method S2-L retention time: 1.583 min, 96.3% purity at 220 nm, [M+H]+= 227.0. 'HNMR (400 MHz, DMSO-de): 6 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).

[0099] Step S2-3’. Preparation of (l?)-l-(4-((l-cyanoethyl)amino)-5-nitropyridin-2-yl)- lH-pyrazolo[3,4-b]pyridine-5-carbonitrile S2-6’: To a stirred solution of (R)-2-((2-chl oro-5 - nitropyridin-4-yl)amino)propanenitrile S2-4’ (1.965 g, 8.33 mmol, 1.0 eq.) and 1H- pyrazolo[3,4-Z>]pyridine-5-carbonitrile S2-5’ (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.), 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 N2 atmosphere 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 (7?)-l-(4-((l- cyanoethyl)amino)-5-nitropyridin-2-yl)-U / -pyrazolo[3,4-Z>]pyridine-5-carbonitrile S2-6’ (1.5 g, 53.9% yield) as a yellow solid. 'HNMR (400 MHz, DMSO-d6): 8 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).

[0100] Step S2-4’. Preparation of (7?)-l-(5-amino-4-((l-cyanoethyl)amino)pyridin-2-yl)- l / / -pyrazolo|3.4- / i|pyridine-5-carbonitrile S2-7’: To a stirred solution of (R)- 1 -(4-((l - cyanoethyl)amino)-5-nitropyridin-2-yl)-U / -pyrazolo[3,4-Z>]pyridine-5-carbonitrile S2-6’ (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-Z>]pyridine-5-carbonitrile S2-7’ (1.2 g, 61.5% yield) as brown solid, which was used in the next step without further purification. UPLC Method S2-2: retention time: 0.561 min, [M+H]+= 305.0.

[0101] Step S2-5’. Preparation of (7?)-l-(5-azido-4-((l-cyanoethyl)amino)pyridin-2-yl)- l / / -pyrazolo|3.4- / i|pyridine-5-carbonitrile S2-8’: To a stirred solution of (7?)-l-(5-amino-4- ((l-cyanoethyl)amino)pyridin-2-yl)-U / -pyrazolo[3,4-Z>]pyridine-5-carbonitrile S2-7’ (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 (7?)-l-(5-azido-4-((l- cyanoethyl)amino)pyridin-2-yl)-U7-pyrazolo[3,4-Z>]pyridine-5-carbonitrile S2-8’ (1.3 g, 56.8% yield) as a gummy liquid, which was used in the next step without further purification. LCMS Method S2-1 : retention time: 1.484 min, [M+H]+= 331.0.

[0102] Step S2-6’. Preparation of l-(4-(((R)-l-cyanoethyl)amino)-5-(4-((lr,4R)-4-(2- ((tetrahydro-2H-pyran-2-yl)oxy)ethyl)cyclohexyl)-lH-l,2,3-triazol-l-yl)yridine-2-yl)-lH- pyrazolo[3,4-b]pyridine-5- carbonitrile S2-9’: To a stirred solution of (R)-l-(5-azido-4-((l- cyanoethyl)amino)yridine-2-yl)-lH-pyrazolo[3,4-b]pyridine-5-carbonitrile S2-8’ (430 mg, 1.302mmol) in acetone (6.0 mL) was added 2-(2-((lr,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-((lr,4R)-4-(2-((tetrahydro-2H- pyran-2-yl)oxy)ethyl)cy clohexyl)- 1H- 1 ,2,3 -tri azol- 1 -yl)yridine-2-yl)- lH-pyrazolo[3 ,4- b]pyridine-5- carbonitrile S2-9’ (220 mg, 20.2% yield). LCMS Method S2-L retention time 3.078 min, 67.93% purity at 220 nm, [M+ H]+= 567.0.

[0103] Step S2-7’. Preparation of l-(4-(((R)-l-cyanoethyl)amino)-5-(4-((lr,4R)-4-(2- hydroxyethyl)cyclohexyl)-lH-l,2,3-triazol-l-yl)pyridine-2-yl)-lH-pyrazolo[3,4-b]pyridine- 5-carbonitrile S2-10’: To a stirred solution of l-(4-(((R)-l-cyanoethyl)amino)-5-(4-((lr,4R)-4- (2-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)cyclohexyl)-lH- 1 ,2,3-triazol- 1 -yl)pyridine-2-yl)- 1H- pyrazolo[3,4-b]pyridine-5-carbonitrile S2-9’ (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-((lr,4r)-4-(2- hydroxyethyl)cy clohexyl)- 1H-1, 2, 3-tri azol-l-yl)pyri dine-2-yl)- lH-pyrazolo[3,4-b]pyridine-5- carbonitrile S2-10’ (130 mg) as a brown solid, which was used in the next step without further pufication. LCMS Method S2-L retention time 1.994 min, [M+ H]+= 483.2.

[0104] Step S2-8’. Preparation of l-(4-(((l?)-l-cyanoethyl) amino)-5-(4-((lr,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: IBX (151 mg, 0.539 mmol, 2.0 eq.) was added to a solution of l-(4-(((R)-l- cyanoethyl) amino)-5-(4-((lr,4r)-4-(2 -hydroxyethyl) cyclohexyl)-lH-l,2,3-triazol-l-yl) pyridin- 2-yl)-lH-pyrazolo[3,4-b] pyridine-5-carbonitrile S2-10’ (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 l-(4-(((R)-l -cyanoethyl) amino)-5-(4-((lr,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. LCMS Method S2-1 : retention time: 2.297 min, [M+H]+= 481.0.Example S3: (R or 5)-3-methyl-3-(6-((5)-2-methylpiperazin-l-yl)pyridin-3-yl)piperidine-2,6-dione (Cl)

[0105] LCMS Method S3-1. Kinetex XB - Cl 8, 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.

[0106] LCMS Method S3-2. 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 mmAmmonium 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.

[0107] UPLC Method S3 -3. Aquity BEH - C18, 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. MSD positive.

[0108] Step 1’. Preparation of 2-(6-bromopyridin-3-yl)propanenitrile 2’: A solution of 2- (5-bromopyridin-2-yl)acetonitrile 1’ (3.2 g, 16.24 mmol, 1.0 eq.) in THF (3.0 mL) was cooled to -5 °C . To the reaction mixture, 2.0 M sodium tert-butoxide in THF (8.12 mL, 16.24 mmol, 1.0 eq.) was added dropwise at the same temperature. Methyl iodide (0.81 mL, 12.99 mmol, 0.8 eq.) was then added dropwise to the reaction mixture at -5 °C. The resulting reaction mixture was then allowed to warm at RT and stirred for 1.0 h at RT. The reaction mixture was added slowly in ice- cold saturated aq. ammonium chloride solution (20 mL) and extracted with ethyl acetate (3 x 20 mL). Combined organic layers were dried over anhydrous sodium sulfate and filtered. The filtrate was concentrated under reduced pressure to obtain the crude compound. The crude compound was purified by column chromatography using silica gel (230-400 mesh) with 30% ethyl acetate / pet ether to obtain 2-(6-bromopyridin-3-yl)propanenitrile 2’ (0.95 g, 4.44 mmol, 27.4% yield) as a pale yellow liquid. ' H NMR (400 MHz, DMSO-d6): 8 ppm 1.57 (d, J= 7.6 Hz, 3H), 4.39 - 4.45 (m, 1H), 7.72 (d, J= 8.4 Hz, 1H), 7.85 (dd, J= 3.2 Hz and 8.4 Hz, 1H), 8.46 (d, J= 2.8 Hz, 1H).

[0109] Step 2’. Preparation of tert-butyl 4-(6-bromopyridin-3-yl)-4-cyanopentanoate 4’: To a stirred solution of 2-(5-bromopyridin-2-yl)propanenitrile 2’ (1.15 g, 5.45 mmol, 1.0 eq.) in toluene (20.0 mL) was added K2CO3 (1.506 g, 10.90 mmol, 2.0 eq.) followed by benzyltriethylammonium chloride (0.248 g, 1.09 mmol, 0.2 eq.). Then tert-butyl acrylate 3’ (1.397 g, 10.90 mmol, 2.0 eq.) was added dropwise to the resulting reaction mixture at RT. The reaction mixture was heated to 90 °C and maintained for 16 h. The reaction mixture was cooled to RT and filtered through celite bed. Washed the insoluble salts (celite bed) with ethyl acetate (2 x 10 mL). Collected filtrate was then concentrated to get the crude product, and the crude product was purified by silica gel column chromatography using 10% ethyl acetate / pet ether to get tert-butyl 4-(6-bromopyridin-3-yl)-4-cyanopentanoate 4’ (1.5 g, 3.96 mmol, 72.7% yield) as an off-white solid. LCMS Method S3-2: Retention time: 2.67 min, [M+H]+= 339.0 and [M+2+H]+= 341.0.XH NMR (400 MHz, DMSO-d6): 6 ppm 1.35 (s, 9H), 1.75 (s, 3H), 2.08 - 2.14 (m, 1H), 2.22 - 2.34 (m, 3H), 7.74 (dd, J= 0.4 Hz and 8.4 Hz, 1H), 7.88 (dd, J= 2.8 Hz and 8.4 Hz, 1H), 8.53 (dd, J = 0.4 Hz and 2.8 Hz, 1H).

[0110] Step 3’. Preparation of tert-butyl (3S)-4-(5-(5-(tert-butoxy)-2-cyano-5- oxopentan-2-yl)pyridin-2-yl)-3-methylpiperazine-l-carboxylate 6’: To a stirred solution of tert-butyl 4-(6-bromopyridin-3-yl)-4-cyanopentanoate 4’ (1.2 g, 3.54 mmol) in 1,4-dioxane (20.0 mL) was added tert-butyl (S)-3-methylpiperazine-l-carboxylate 5’ (0.850 g, 4.24 mmol) followed by cesium carbonate (2880 mg, 8.84 mmol). The resulting reaction mixture was degassed for 10 min with nitrogen gas, and then, RuPhos Pd G4 (0.180 g, 0.212 mmol) was added. The reaction mixture was heated to 90 °C and stirred for 16 h. The reaction mixture was cooled to room temperature, diluted with water (10 mL) and and extracted with ethyl acetate (2 x 15 mL). Combined organic layers were washed with sat.brine solution (10 mL), dried over anhydrous sodium sulfate and filtered. The filtrate was concentrated under reduced pressure to obtain the crude compound. The crude compound was purified by column chromatography using silica gel (230 - 400 mesh) with 0 - 25% ethyl acetate / pet ether to obtain tert-butyl (3S)-4-(5-(5-(tert- butoxy)-2-cyano-5-oxopentan-2-yl)pyridin-2-yl)-3-methylpiperazine-l-carboxylate 6’ (1.31 g, 69.6% yield) as an off-white solid. LCMS Method S3-2: retention time: 2.59 min, [M+H]+= 459.2.

[0111] Step 4’. Preparation of 3- methyl 3-(6-((S)- 2- methylpi perazin 1- yl)pyridin -3- yl)piperid ine-2,6- dione 7’: To a stirred solution of tert-butyl (3S)-4-(5-(5-(tertbutoxy)-2-cyano- 5-oxopentan-2-yl)pyridin-2-yl)-3-methylpiperazine-l-carboxylate 6’ (0.7 g, 1.875 mmol) in AcOH (5.0 mL) was added sulfuric acid (0.2 mL, 3.75 mmol). The resulting reaction mixture was heated to 120 °C and stirred for 3 h. The reaction mixture was concentrated under reduced pressure to obtain 3- methyl 3-(6-((S)- 2- methylpi perazin 1- yl)pyridin -3- yl)piperid ine-2,6- dione 7’ (1.1 g) as a pale brown thick oil. It was used for the next step without further purification.

[0112] Step 5’. Preparation of tert-butyl (3S)-3-methyl-4- (5-(3-methyl-2,6- dioxopiperidin-3-yl)pyridin-2-yl)piperazine-l-carboxylate 8’: To a stirred solution of 3- methyl-3-(6-((S)-2-methylpiperazin-l-yl)pyri din-3 -yl)piperidine-2, 6-dione 7’ (1.0 g, 3.31 mmol) in acetonitrile (5 mL) at 0 °C was added DIPEA (5.78 mL, 33.1 mmol) and stirred for 5 min. Then di- / c / 7-butyl decarbonate (1.15 mL, 4.96 mmol) was added to the reaction mixture, and the resulting reaction mixture was stirred at RT for 16 h. Then, the reaction mixture was diluted with ethyl acetate (25 mL) and washed with water (3 x 25 mL). Combined organic layers were washed with sat. brine solution (25 mL), dried over anhydrous sodium sulfate and filtered. The filtrate was concentrated under reduced pressure to obtain the crude compound. The crude compound was trituration with pet ether (2 x 5 mL) to obtain the crude tert-butyl (3S)-3-methyl-4-(5-(3-methyl- 2, 6-dioxopiperi din-3 -yl)pyridin-2-yl)piperazine-l -carboxylate 8’ (0.651 g, 43.0% yield) as an off- white solid. UPLC Method S3-3: retention time: 0.828 min, [M+H]+= 403.2

[0113] Step 6’. The preparation of tert-butyl (R or S)-2-methyl-4-(5-((S)-3-methyl-2,6- dioxopiperidin-3-yl)pyridin-2-yl)piperazine-l-carboxylate 9a’(peak-l) and tert-butyl (R or S)-2-methyl-4-(5-((S)-3-methyl-2,6-dioxopiperidin-3-yl)pyridin-2-yl)piperazine-l- carboxylate 9b’(peak-2): 720 mg of the material 8’ was purified by chiral SFC to obtain tertbutyl (R or S)-2-methyl-4-(5-((S)-3-methyl-2,6-dioxopiperidin-3-yl)pyridin-2-yl)piperazine-l- carboxylate 9a’ (302 mg, peak-1) and tert-butyl (R or S)-2-methyl-4-(5-((S)-3-methyl-2,6- dioxopiperidin-3-yl)pyridin-2-yl)piperazine-l -carboxylate 9b’ (320 mg, peak-2).

[0114] SFC Method: Chiralpak-IH, 250 x 4.6 mm, 5.0 pm, Flow: 3.0 mL / min. Co-Solvent: 50.0% IP A

[0115] tert-butyl (R or S)-2-methyl-4-(5-((S)-3-methyl-2,6-dioxopiperidin-3-yl)pyridin-2- yl)piperazine-l -carboxylate 9a’ (peak-1): RT 2.176 min; ee 100.0%. LCMS Method S3-1 : retention time: 2.377 min, [M+H]+= 403.2. 'H NMR (400 MHz, DMSO-d6): 8 ppm 0.99 - 1.04 (d, J= 6.8 Hz, 3H), 1.40 - 1.45 (m, 12H), 2.02 - 2.18 (m, 2H), 2.31 - 2.36 (m, 1H), 2.44 - 2.49 (m, 1H), 2.85 - 3.20 (m, 3H), 3.76 - 3.80 (m, 1H), 3.95 - 3.98 (m, 2H), 4.44 - 4.46 (m, 1H), 6.77 (d, J= 9.2 Hz, 1H), 7.46 - 7.48 (m, 1H), 8.01 (d, J= 2.8 Hz, 1H), 10.87 (s, 1H).

[0116] tert-butyl (R or S)-2-methyl-4-(5-((S)-3-methyl-2,6-dioxopiperidin-3-yl)pyridin-2- yl)piperazine-l -carboxylate 9b’ (peak-2): RT 3.147 min; ee 99.94%. LCMS Method S3-2: retention time: 2.06 min, [M+H]+= 403.2. 'H NMR (400 MHz, DMSO-d6): 6 ppm 1.01 (d, J = 6.4 Hz, 3H), 1.40 - 1.42 (m, 12H), 2.08 - 2.18 (m, 2H), 2.31 - 2.36 (m, 1H), 2.44 - 2.45 (m, 1H), 2.85 - 3.20 (m, 3H), 3.78 - 3.81 (m, 1H), 3.96 - 3.99 (m, 2H), 4.46 - 4.47 (m, 1H), 6.77 (d, J= 8.8 Hz, 1H), 7.46 - 7.49 (m, 1H), 8.01 (d, J= 2.4 Hz, 1H), 10.88 (s, 1H).

[0117] Step 7’. Preparation of (R) or (S) 3- methyl 3-(6-((S)-2- methylpiperazin-1- yl)pyridin-3-yl)piperidine-2,6- dione (Cl): To a stirred solution of tert-butyl (R or S)-2-methyl- 4-(5-((S)-3-methyl-2,6-dioxopiperidin-3-yl)pyridin-2-yl)piperazine-l-carboxylate 9a’ (0.3 g, 0.745 mmol) in DCM (5.0 mL, 0.103 M) at RT was added 4.0 N HC1 in 1,4-dioxane (0.93 mL, 3.73 mmol). The resulting reaction mixture was stirred at RT for 4 h. The reaction mixture was concentrated under reduced pressure to obtain crude (Cl) (0.290 g, 94% yield) as an off-white solid. It was used for the next step without further purification. LCMS Method S3-2: retention time: 0.356 min, [M+H]+= 303.2.Example S4: Preparation of l-(4-(((l?)-l-cyanoethyl)amino)-5-(4-((15',41?)-4-(2-((5')-3- methyl-4-(5-((l? or 5)-3-methyl-2,6-dioxopiperidin-3-yl)pyridin-2-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)

[0118] LCMS Method S4-1. Kinetex XB - C18, 50 x 4.6 mm, 5.0 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 2.5 min. MSD positive.

[0119] LCMS Method S4-2. 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.

[0120] UPLC Method S4-3. Aquity 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. MSD positive.

[0121] l-(4-((R)-l-cyanoethyl)amino)-5-(4-((lS,4R)-4-(2-((3S)-3-methyl-4-(5-(3-methyl-2,6-Dioxopiperidin-3-yl)pyridine-2-yl)piperazin-l-yl)ethyl)cyclohexyl)-lH-l,2,3-triazol-l- yl)pyridine-2-yl)-lH-pyrazolo[3,4-b]pyridine-5-carbonitrile (P-1): To a stirred solution of 1- (4-((R)-l-cyanoethyl)amino)-5-(4-((lr,4r)-4-(2-oxoethyl)cyclohexyl)-lH-l,2,3-triazol-l-yl) pyridine-2-yl)-lH-pyrazolo[3,4-b] pyridine-5-carbonitrile Al (100 mg, 0.162 mmol) and (R) or (S) 3- methyl 3-(6-((5)-2-methylpiperazin-l-yl)pyridine-3-yl)piperidine-2, 6-dione, 2 HC1 (61.5 mg, 0.162 mmol) in DMSO (1.5 mL) at RT was added DIPEA (0.085 mL, 0.487 mmol) followed by sodium triacetoxyborohydride (STAB, 51.6 mg, 0.243 mmol). The resulting reaction mixture was stirred at RT for 1 h. The reaction mixture was quenched with four drops of water, purified by Prep-HPLC purification to afford l-(4-((R)-l-cyanoethyl)amino)-5-(4-((lS,4R)-4-(2-((3S)-3- methyl-4-(5-(3-methyl-2,6-Dioxopiperidin-3-yl)pyridine-2-yl)piperazin-l-yl)ethyl)cyclohexyl)- lH-l,2,3-triazol-l-yl)pyridine-2-yl)-lH-pyrazolo[3,4-b]pyridine-5-carbonitrile (P-1), 2 TFA (31.71 mg, 19.43% yield) as a beige solid.

[0122] Prep-HPLC Method: Kinetex Biphenyl (250 x 19)mm, 5 p; Mobile phase A: 0.2% TFA in Water; Mobile phase B: 0.2% TFA in ACN; Retention time: 14.330 min.

[0123] LCMS Method S4-2: retention time: 1.717 min, [M+H]+= 767.2.

[0124] 1H NMR (400 MHz, DMSO-d6): 5 ppm 1.16 - 1.29 (m, 6H), 1.42 - 1.60 (m, 6H), 1.61(d, .7= 7.2 Hz, 3H), 1.63 - 1.75 (m, 2H), 1.86 - 1.91 (m, 2H), 2.02 - 2.20 (m, 4H), 2.31 - 2.41 (m, 1H), 2.71 - 2.83 (m, 1H), 2.97 - 3.09 (m, 1H), 3.15 - 3.30 (m, 4H), 3.50 - 3.59 (m, 2H), 4.31 - 4.40 (m, 1H), 4.75 - 4.83 (s, 1H), 4.99 - 5.07 (m, 1H), 6.92 (d, J= 8.8 Hz, 1H), 7.19 (d, J= 8.4 Hz, 1H), 7.54 - 7.60 (m, 1H), 7.68 (s, 1H), 8.04 - 8.05 (m, 1H), 8.37 (s, 1H), 8.41 (s, 1H), 8.73 (s, 1H), 9.06 - 9.08 (m, 2H), 10.91 (s, 1H).Biological ExamplesExample Bl. Reagent Preparations

[0125] 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.

[0126] The Cell titer Gio (CTG) buffer and substrate (CellTiter-Glo Luminescent Cell Viability Assay, Promega Ref.# G7573) were stored at -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.

[0127] 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.

[0128] 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.

[0129] 20X Antibody Solution aliquots were stored at -80 °C and the Detection Buffer was stored at 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

[0130] 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 preparedin 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.

[0131] Dosing was conducted using the following method. To dose assay plates and CTG plate 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.

[0132] 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 1 (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 IX antibody mix. Using a multichannel repeater, 4 pL of IX antibody mix was added to each well. The 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 an Envision plate reader.

[0133] 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).

[0134] A summary of the LVL THP1 HTRF data for the tested compound is provided in the following table:Example B3. Pharmacokinetic Studies in Mice.

[0135] Due to the higher molecular weight and lipophilicity, targeted protein degraders generally occupy a physiochemical property space that is beyond Lipinski’s rule of 5 and asconsequence are expected to have poor oral bioavailability and overall pharmacokinetic properties. Compounds of the present disclosure can overcome these liabilities as demonstrated by the highly favorable PK properties shown for the selected compounds. The plasma pharmacokinetics (PK) of P-1 were characterized in female C57B16 mice following intravenous (IV; 0.5 mg / kg) administration. Plasma from the tail vein was withdrawn in composite fashion: one group of mice (n = 3) provided samples at 3 min, 1 h, and 24 h, while another group (n = 3) provided samples at 15 min and 3 h, and a third group of mice (n = 3) provided samples at 30 min and 7 h. The concentration of test article in these samples was measured using reversephase LC-MS-MS and then plotted as a function of time. The resulting concentration-time data was processed using noncompartmental analysis for the calculation of clearance (Cl), volume of distribution at steady state (Vss), half-life, mean residence time (MRT), and area under the time curve (AUC).

[0136] Similarly, the PK were characterized following oral (PO; 3 mg / kg) administration. Again, plasma from the tail vein was withdrawn in composite fashion: one group of mice (n = 3) provided samples at 15 min, 1 h, and 7 h, while another group (n = 3) provided samples at 30 min, 3 h, and 24 h. Oral bioavailability (Fpo) was calculated by comparing the results of the studies described, dividing the dose-normalized AUCLAST.PO by the dose-normalized AUCLAST.IV. The results of the PK study in mice for compound P-1 are shown in the following table, and highlight the highly favorable PK properties and oral bioavailability of P-1 for molecules of this size:IV vehicle: 85% D5W, 5% DMSO, 10% SOLUTOL (solution)PO vehicle: 0.5% MC and 0.25% TWEEN 80 in water (suspension)

[0137] 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 the following structural formula:or a pharmaceutically acceptable salt thereof.

2. The compound of claim 1, of the following structural formula:or a pharmaceutically acceptable salt thereof.

3. A compound of the following structural formula:or a pharmaceutically acceptable salt thereof.

4. A pharmaceutical composition comprising the compound or pharmaceutically acceptable salt thereof of any one of claims 1-3 , and a pharmaceutically acceptable excipient.

5. A method of modulating interleukin- 1 (IL1) receptor-associated kinase 4 (IRAK4) activity comprising contacting IRAK4 with an effective amount of the compound or pharmaceutically acceptable salt thereof of any one of claims 1-3, or the pharmaceutical composition of claim 4.

6. 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 or pharmaceutically acceptable salt thereof of any one of claims 1-3, or the pharmaceutical composition of claim 4, optionally wherein the inflammatory or autoimmune disease is atopicdermatitis, 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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