Small molecule cyclin-dependent kinase 4 / 6 (CDK4 / 6) and IKZF2 (HELIOS) degraders and methods of use thereof

Abstract

The present application provides bifunctional compounds that act as proteolysis-inducing moieties of cyclin-dependent kinase 4 (CDK4), cyclin-dependent kinase 6 (CDK6) and Ikaros family zinc finger 2 (IKZF2), also known as Helios. Methods for the treatment of disorders regulated by CDK4, CDK6, and IKZF2 are also disclosed.

Description

[Technical field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims the benefit of priority under 35 U.S.C. §119(e) to U.S. Provisional Patent Application No. 63 / 222,646, filed July 16, 2021, which is incorporated by reference in its entirety.

[0002] Government support This disclosure was made with Government support under Grant No. R01 CA218278 awarded by the National Institutes of Health. The Government has certain rights in this disclosure. [Background technology]

[0003] Helios (IKZF2), a member of the IKZF zinc finger transcription factor family (IKZF) family, is a key regulator of T cell activity and function. Genetic deletion of Helios resulted in enhanced antitumor immune responses (Kim et al., Science 350:334-339(2015)). In particular, Helios is highly expressed in regulatory T cells (Elkord et al. Biol. Ther. 12:1423-1425(2012)), a subpopulation of T cells that limits the activity of effector T cells. Selective deletion of Helios in regulatory T cells (Tregs) resulted in both loss of suppressive activity and gain of effector T cell function (Najagawa et al., Proc. Natl. Acad. Sci. USA 113:6248-6253 (2016); Yates et al., Proc. Natl. Acad. Sci. USA 115:2162-2167 (2018)). Thus, Helios is a key factor limiting T cell effector function in Tregs.

[0004] Helios expression has also been shown to be associated with chronic viral infections (Crawford et al., Immunity 40:289-302 (2014), Doering et al., Immunity 371130-1144 (2012); Scott-Browne et al., Immunity 45:1327-1340 (2016)) and tumors (Martinez et al., Immunity 42:265-278 (2015); Mognol et al., Proc. Natl. Acad. Sci. USA 114:E2776-E2785 (2017); Pereira et al., J. Leukoc. Biol. 102:601-615 (2017); Singer et al., Cell 166:1500-1511 (2016); Schietinger et al., Immunity 45:389-401 (2016)), as well as in dysfunctional chimeric antigen receptor (CAR) T cells (Long et al., Nat. Med. 21:581-590 (2015)). Overexpression or aberrant expression of Helios and various splice isoforms has been reported in several hematological malignancies, including T cell leukemias and lymphomas (Nakase et al., Exp. Hematol. 30:313-317 (2002); Tabayashi et al., Cancer Sci. 98:182-188 (2007); Asanuma et al., Cancer Sci. 104:1097-1106 (2013)). Furthermore, knockdown of Helios in a model of mixed lineage leukemia (MLL)-driven myeloid leukemia potently suppressed proliferation and increased cell death (Park et al., J. Clin. Invest. 125:1286-1298 (2015); Park et al., Cell Stem Cell 24:153-165 (2019)).

[0005] Cyclin-dependent kinases (CDKs) integrate multiple signaling pathways to control either the cell cycle or gene transcription. CDK1, 2, 4 and 6 are key enzymes that drive cell cycle transitions. For example, CDK1 is a key determinant of mitotic progression, CDK2 regulates DNA replication in S phase, and CDK4 / 6 drive the cell cycle from G0 or G1 to S phase by phosphorylating Rb protein to activate the expression of genes involved in cell cycle control. CDK4 / 6 inhibitors have been shown to have potent immunostimulatory effects on T cells both in ex vivo human T cells and in multiple in vivo mouse tumor models. CDK7, 9 and 12 regulate transcription instead of directly promoting the cell cycle. CDK7 is an enzymatic component of the TFIIH complex responsible for regulating transcription initiation, while CDK9 and CDK12 regulate transcription elongation and processing.

[0006] Deregulation of CDKs has been shown to have significant effects on cellular status and is often identified as oncogenic. Although numerous selective or pan-CDK small molecule inhibitors have been identified, most of the known inhibitors have failed in clinical trials due to the lack of high systemic drug concentrations. More recently, three CDK4 / 6 inhibitors, abemaciclib, palbociclib, and ribociclib, have been clinically applied in combination with hormone therapy to treat hormone receptor (HR)-positive human epidermal growth factor receptor 2 (HER-2)-negative metastatic breast cancer. Summary of the Invention [Means for solving the problem]

[0007] A first aspect of the present disclosure is a compound represented by formula (I): [ka] or a pharma- ceutically acceptable salt or stereoisomer thereof, wherein the targeting ligand binds to CDK4 and / or CDK6 and the linker comprises an alkylene chain or a polyethylene glycol chain.

[0008] Another aspect of the present disclosure is directed to a pharmaceutical composition comprising a therapeutically effective amount of a bifunctional compound of formula (I) or a pharma- ceutically acceptable salt or stereoisomer thereof and a pharma- ceutically acceptable carrier. In some embodiments, the pharmaceutical composition is in solid form. In some embodiments, the pharmaceutical composition is in tablet or capsule form. In some embodiments, the pharmaceutical composition is in liquid form.

[0009] Another aspect of the present disclosure is directed to methods of making the bifunctional compounds of formula (I) and their pharma- ceutically acceptable salts and stereoisomers.

[0010] A further aspect of the present disclosure is directed to a method of treating a disease or disorder characterized by abnormal activity of CDK4 and / or CDK6 and Helios, comprising administering to a subject in need thereof a therapeutically effective amount of a bifunctional compound of formula (I) or a pharma- ceutically acceptable salt or stereoisomer thereof. In some embodiments, the disease or disorder is cancer. In some embodiments, the cancer is characterized by a solid tumor. In some embodiments, the cancer is selected from breast cancer, brain cancer, endometrial cancer, head and neck cancer, gastrointestinal cancer, lung cancer, ovarian cancer, prostate cancer, uterine cancer, hepatocellular carcinoma, liposarcoma, and melanoma. In some embodiments, the cancer is a hematological cancer. In some embodiments, the hematological cancer is selected from leukemia, lymphoma, and myeloma.

[0011] Yet a further aspect of the present disclosure is directed to a method of reducing levels of CDK4 and / or CDK6, and Helios in a cell, either in vitro or in vivo, comprising contacting the cell with an effective amount of a bifunctional compound of formula (I) or a pharmaceutically acceptable salt or stereoisomer thereof.

[0012] CDK4 / 6 inhibitors have been shown to have potent immune stimulatory effects on T cells in both ex vivo human T cells and multiple in vivo mouse tumor models. Conventional CDK4 / 6 inhibitors have not been shown to induce the degradation of Helios. Conventional small molecule degraders of Helios have been shown to destabilize the anergic and suppressive phenotypes of regulatory T cells. The bifunctional compounds of the present disclosure can reprogram the CRL4 (CRBN) E3 ligase complex to target Helios, CDK4, and / or CDK6 for ubiquitination and subsequent proteasomal degradation, thus promoting antitumor immunity. They can also be used in combination with existing immunotherapies such as immune checkpoint inhibitors (e.g., anti-PD-1, anti-PD-L1) or cell therapies (e.g., CAR-T cells).

[0013] Without intending to be bound by any particular theory of action, it is believed that by inducing the degradation of Helios, the bifunctional compounds of the present disclosure may enhance anti-tumor immune responses by converting regulatory T cells into effector T cells and by rescuing effector T cell function in exhausted T cells or CAR-T cells. Furthermore, it is believed that the compounds of the present disclosure exert their therapeutic (e.g., anti-cancer) effects or benefits by a combination of anti-proliferative and immunomodulatory effects. Mechanistically, it is believed that the disclosed compounds induce the degradation of Helios by forming a "molecular glue" with cereblon (CRBN).

[0014] Molecular glue compounds induce protein-protein interactions that lead to protein degradation in the context of ubiquitin ligases (Stanton et al., Science 359:eaao5902 (2018)). Unlike proteolysis-targeting chimeric molecules (PROTACs), molecular glue compounds are small molecules (also known as small molecule degraders) that induce interactions between the substrate receptor of an E3 ubiquitin ligase and a target protein, leading to targeted protein degradation. Examples of molecular adhesives that induce targeted protein degradation include IMiDs (immunomodulatory drugs; e.g., thalidomide), which generate a novel interaction between a substrate (e.g., IKZF1 / 3) and cereblon, a substrate receptor (also known as DCAF) for Cullin-RING ubiquitin ligase 4 (CRL4) (Besten and Lipford, Nat. Chem. Biol. 16(11):1157-1158 (2020)). Unlike traditional enzyme inhibitors, these molecular adhesive degraders act stoichiometrically to catalyze the rapid depletion of previously inaccessible targets (Chopra et al., Drug Discov. Today. Technol. 31:5-13 (2019)). Although highly desirable, molecular adhesive degraders have only been discovered serendipitously. There are limited strategies available to identify or design these compounds (Slabicki et al., Nature DOI: 10.1038 / s41586-020-2374-x (2020)).

[0015] In the context of the present disclosure, the compounds of formula (I) act as molecular glue in the sense that they recruit a ubiquitin ligase (in this case CRBN) to a target protein (in this case Helios) and function as a catalyst for the degradation of the target protein. In doing so, the compounds are believed to alter the substrate binding site of CRBN such that the target protein becomes a neosubstrate (Burslem et al., Chem. Rev. 117:11269-11301(2017)). Dissociation of the molecular glue after the ubiquitination step allows the target protein to subsequently function on a different molecule (Che et al.Med. Chem. Lett. 28:2585-2592(2018)).

[0016] As shown in the examples herein, the bifunctional compounds of formula (I) (also referred to herein as decomposition agents) promote the co-degradation of CDK4, CDK6, and Helios, while substantially sparing other CDK and IKZF isoforms.Therefore, the bifunctional compounds of the present disclosure serve as a series of new chemical tools for CDK4, CDK6, and Helios knockdown, and exemplify a broadly applicable approach to arrive at decomposition agents that are more selective than non-selective binding ligands, and can provide effective treatment of CDK4, CDK6, and Helios-mediated diseases and disorders, such as cancer, neurodegenerative diseases, and autoimmune diseases. [Brief description of the drawings]

[0017] [Figure 1A] FIG. 1A is a set of immunoblots of Jurkat cells treated with 1 μM of target compounds (DKY709 and Ia-1 to Ia-8) for 4 h, showing the levels of Ikaros, Helios, CDK4, CDK6, and tubulin for each compound.

[0018] [Figure 1B]FIG. 1B is a set of immunoblots of Jurkat cells treated for 4 hours with the Helios degrader DKY709, the triple degrader Ia-8, and the paired negative control compound 17, showing the levels of Ikaros, Helios, CDK4, CDK6, and tubulin for each compound at 0.1 μM and 1 μM.

[0019] [Diagram 2] FIG. 2 is a set of immunoblots of Jurkat cells treated with 0.1 μM and 1 μM of target compounds (palbociclib, DKY709, Ia-8, and 17) for 16 hours.

[0020] [Figure 3A] Figure 3A is a set of histograms and quantification of DNA content using propidium iodide (PI) staining after treating Jurkat cells with 100 nM of target compounds (palbociclib, DKY709, 18, Ia-6, 16, Ia-8, and 17) for 24 hours.

[0021] [Figure 3B] FIG. 3B is a graph of normalized luminescence versus concentration (log[inhibitor], M) of target compounds (palbociclib, Ia-8, and 17) assessed by CellTiterGlo after 3 days of treatment of Jurkat cells.

[0022] [Figure 4] FIG. 4 is a graph of [IL2] for target compounds in Jurkat cells upon TCR stimulation with α-CD3 / CD28. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0023] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the art to which the subject matter of this specification belongs. As used in this specification and the appended claims, unless expressly stated to the contrary, the following terms have the meanings set forth to facilitate the understanding of this disclosure.

[0024] As used in this specification and the appended claims, the singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise. Thus, for example, reference to "a composition" includes mixtures of two or more such compositions, reference to "an inhibitor" includes mixtures of two or more such inhibitors, etc.

[0025] Unless otherwise specified, the term "about" means within 10% (eg, within 5%, 2%, or 1%) of the particular value that is modified by the term "about."

[0026] The transitional phrase "comprising," which is synonymous with "including," "containing," or "characterized by," is inclusive or open-ended and does not exclude additional unrecited elements or method steps. In contrast, the transitional phrase "consisting of" excludes any element, step, or ingredient not specified in the claim. The transitional phrase "consisting essentially of" limits the claim to "particular materials or steps" of the claimed disclosure and those that do not materially affect the basic and novel characteristics.

[0027] With respect to the compounds of the present disclosure, to the extent the following terms are used herein to further describe them, the following definitions apply.

[0028] As used herein, the term "alkyl" refers to a saturated linear or branched monovalent hydrocarbon radical. Unless otherwise defined for any particular group in the compounds of formula (I), in one embodiment, an alkyl group is a C1-C 18 In other embodiments, the alkyl group is a C0-C6, C0-C5, C0-C3, C1-C 12 , C1-C8, C1-C6, C1-C5, C1-C4 or C1-C3 groups (wherein CO alkyl means a bond). Examples of alkyl groups include methyl, ethyl, 1-propyl, 2-propyl, i-propyl, 1-butyl, 2-methyl-1-propyl, 2-butyl, 2-methyl-2-propyl, 1-pentyl, n-pentyl, 2-pentyl, 3-pentyl, 2-methyl-2-butyl, 3-methyl-2-butyl, 3-methyl-1-butyl, 2-methyl-1-butyl, 1-hexyl, 2-hexyl, 3-hexyl, 2-methyl-2-pentyl, 3-methyl-2-pentyl, 4-methyl-2-pentyl, 3-methyl-3-pentyl, 2-methyl-3-pentyl, 2,3-dimethyl-2-butyl, 3,3-dimethyl-2-butyl, heptyl, octyl, nonyl, decyl, undecyl and dodecyl. In some embodiments, the alkyl group is a C1 to C3 alkyl group.

[0029] As used herein, the term "alkylene" refers to a straight or branched divalent hydrocarbon chain, consisting solely of carbon and hydrogen, containing no unsaturation, having 1 to 12 carbon atoms, linking the remainder of the molecule to a radical group, e.g., methylene, ethylene, propylene, n-butylene, etc. In some embodiments, the alkylene chain or divalent alkylene chain may be interrupted by at least one other group and / or terminated (at either or both ends) with at least one other group. In some embodiments, an alkylene chain or a divalent alkylene chain is -O-, -S-, -N(R')-, -C≡C-, -C(O)-, -C(O)O-, -OC(O)-, -OC(O)O-, -C(NOR')-, -C(O)N(R')-, -C(O)N(R')C(O)-, -C(O)N(R')C(O)N(R')-, -N(R')C(O)-, -N(R')C(O)N(R')-, -N(R')C(O)O-, -OC(O)N(R')-, - C(NR')-, -N(R')C(NR')-, -C(NR')N(R')-, -N(R')C(NR')N(R')-, -OB(Me)O-, -S(O)2-, -OS(O)-, -S(O)O-, -S(O)-, -OS(O )2-, -S(O)2O-, -N(R')S(O)2-, -S(O)2N(R')-, -N(R')S(O)-, -S(O)N(R')-, -N(R')S(O)2N(R')-, -N(R')S(O)N(R')-, C3~C 12It may be interrupted and / or terminated (at either or both ends) with at least one of carbocyclene, 3-12 membered heterocyclene, 5-12 membered heteroarylene or any combination thereof, where R' is H or C1-C6 alkyl, and the interrupting group and one or both terminal groups may be the same or different. Unless otherwise defined for any particular group in the compounds of formula (I), the alkylene chain may be attached to the rest of the molecule through a single bond and to the radical group through a single bond. In some embodiments, the alkylene group contains 1-8 carbon atoms (C1-C8 alkylene). In other embodiments, the alkylene group contains 1-5 carbon atoms (C1-C5 alkylene). In other embodiments, the alkylene group contains 1-4 carbon atoms (C1-C4 alkylene). In other embodiments, the alkylene group contains 1-3 carbon atoms (C1-C3 alkylene). In other embodiments, the alkylene group contains 1 to 2 carbon atoms (C1-C2 alkylene). In other embodiments, the alkylene group contains 1 carbon atom (C1 alkylene).

[0030] As used herein, the term "alkenyl" refers to a straight or branched chain monovalent hydrocarbon radical having at least one carbon-carbon double bond. Unless otherwise defined for any particular group in the compounds of formula (I), alkenyl includes radicals having "cis" and "trans" orientations, alternatively "E" and "Z" orientations. In one example, an alkenyl radical is a C2-C 18 In another embodiment, the alkenyl group is a C-C 12 , C2~C 10 , C2-C8, C2-C6 or C2-C3 groups. Examples include ethenyl or vinyl, prop-1-enyl, prop-2-enyl, 2-methylprop-1-enyl, but-1-enyl, but-2-enyl, but-3-enyl, buta-1,3-dienyl, 2-methylbuta-1,3-dienyl, hex-1-enyl, hex-2-enyl, hex-3-enyl, hex-4-enyl and hex-1,3-dienyl.

[0031] The term "alkoxyl" or "alkoxy" as used herein refers to an alkyl group, as defined above, having an oxygen radical attached to it, which is the point of attachment. Representative alkoxyl groups include methoxy, ethoxy, propyloxy, tert-butoxy, and the like. An "ether" is two hydrocarbons covalently linked by an oxygen. Thus, the substituent of an alkyl that makes it an ether is or resembles an alkoxyl, as can be represented by one of -O-alkyl, -O-alkenyl, and -O-alkynyl.

[0032] As used herein, the term "alkoxylenes" refers to compounds of the general formula (-OC n H 2n -) where n represents an integer (e.g., 1, 2, 3, 4, 5, 6, or 7) and includes both straight and branched chain groups. Unless otherwise defined for any particular group in the compounds of formula (I), the alkoxylene chain may be attached to the rest of the molecule through a single bond and to the radical group through a single bond. In some embodiments, the alkoxylene group contains 1 to 3 carbon atoms (-O-C1-C3 alkoxyylene). In other embodiments, the alkoxyylene group contains 1 to 5 carbon atoms (-O-C1-C5 alkoxyylene).

[0033] As used herein, the term "halogen" (or "halo" or "halide") refers to fluorine, chlorine, bromine, or iodine.

[0034] As used herein, the term "cyclic group" is used alone or as part of a larger moiety and broadly refers to any group that contains a saturated, partially saturated or aromatic ring system, such as carbocyclic (cycloalkyl, cycloalkenyl), heterocyclic (heterocycloalkyl, heterocycloalkenyl), aryl and heteroaryl groups. Unless otherwise defined for any particular group in the compounds of formula (I), a cyclic group may have one or more (e.g., fused) ring systems. Thus, for example, a cyclic group may contain one or more carbocyclic, heterocyclic, aryl or heteroaryl groups.

[0035] As used herein, the term "carbocycle" (also referred to as "carbocyclyl"), used alone or as part of a larger moiety, refers to a group that contains a saturated, partially unsaturated, or aromatic ring system having from 3 to 20 carbon atoms, either alone or as part of a larger moiety (e.g., an alkcarbocyclic group). Unless otherwise defined for any particular group in the compounds of Formula (I), the term carbocyclyl includes monocyclic, bicyclic, tricyclic, fused, bridged, and spirocyclic rings, and combinations thereof. In one embodiment, a carbocyclyl is a ring system having from 3 to 15 carbon atoms (C3-C4). 15 In one embodiment, carbocyclyl is an alkyl group having 3 to 12 carbon atoms (C 12 In another embodiment, carbocyclyl is C3-C8, C3-C 10 Or C5~C 10 In another embodiment, the carbocyclyl, as a monocycle, includes C3-C8, C3-C6, or C5-C6. In some embodiments, the carbocyclyl, as a bicycle, includes C7-C 12 In another embodiment, the carbocyclyl, as a spiro system, is a C5-C 12Representative examples of monocyclic carbocyclyls include cyclopropyl, cyclobutyl, cyclopentyl, 1-cyclopent-1-enyl, 1-cyclopent-2-enyl, 1-cyclopent-3-enyl, cyclohexyl, perdeuterocyclohexyl, 1-cyclohex-1-enyl, 1-cyclohex-2-enyl, 1-cyclohex-3-enyl, cyclohexadienyl, cycloheptyl, cyclooctyl, cyclononyl, cyclodecyl, cycloundecyl, phenyl, and cyclododecyl; bicyclic carbocyclyls having 7 to 12 ring atoms include [4,3], [4,4], [4,5], [5,5], [5,6], or [6,6] ring systems, such as bicyclo[2.2.1]heptane, bicyclo[2.2.2]octane, naphthalene, and bicyclo[3.2.2]nonane. Representative examples of spirocarbocyclyl include spiro[2.2]pentane, spiro[2.3]hexane, spiro[2.4]heptane, spiro[2.5]octane and spiro[4.5]decane. The term carbocyclyl includes aryl ring systems as defined herein. The term carbocyclyl also includes cycloalkyl rings (e.g., saturated or partially unsaturated monocyclic, bicyclic, or spirocyclic carbocycles). The term carbocyclic group also includes carbocyclic rings fused to one or more (e.g., 1, 2, or 3) different cyclic groups (e.g., aryl or heterocyclic rings), where a radical or point of attachment is on the carbocyclic ring.

[0036] Thus, the term carbocyclic, as used herein, also refers to a ring having the formula -R c -carbocyclyl (wherein R c is an alkylene chain). The term carbocyclic, as used herein, also includes carbocyclylalkyl groups of the formula -OR c -carbocyclyl (wherein R c is an alkylene chain).

[0037] As used herein, the term "heterocyclyl" used alone or as part of a larger moiety refers to a "carbocyclyl" containing a saturated, partially unsaturated, or aromatic ring system in which one or more (e.g., 1, 2, 3, or 4) carbon atoms are replaced with a heteroatom (e.g., O, N, N(O), S, S(O), or S(O)2). The term heterocyclyl includes monocyclic, bicyclic, tricyclic, fused, bridged, and spiro ring systems, and combinations thereof. Unless otherwise defined for any particular group in the compounds of formula (I), in some embodiments, heterocyclyl refers to a 3-15 membered heterocyclyl ring system. In some embodiments, heterocyclyl refers to a 3-12 membered heterocyclyl ring system. In some embodiments, heterocyclyl refers to a saturated ring system, such as a 3-12 membered saturated heterocyclyl ring system. In some embodiments, heterocyclyl refers to a heteroaryl ring system, such as a 5-14 membered heteroaryl ring system. The term heterocyclyl also includes C3-C8 heterocycloalkyl, which are saturated or partially unsaturated monocyclic, bicyclic or spirocyclic ring systems containing 3 to 8 carbons and one or more (1, 2, 3 or 4) heteroatoms.

[0038] In some embodiments, the heterocyclyl group contains 3 to 12 ring atoms, including monocyclic, bicyclic, tricyclic, and spirocyclic ring systems, where the ring atoms are carbon and 1 to 5 ring atoms are heteroatoms (e.g., nitrogen, sulfur, or oxygen). In some embodiments, the heterocyclyl contains a 3- to 7-membered monocyclic ring having one or more heteroatoms selected from nitrogen, sulfur, or oxygen. In some embodiments, the heterocyclyl contains a 4- to 6-membered monocyclic ring having one or more heteroatoms selected from nitrogen, sulfur, or oxygen. In some embodiments, the heterocyclyl contains a 3-membered monocyclic ring. In some embodiments, the heterocyclyl contains a 4-membered monocyclic ring. In some embodiments, the heterocyclyl contains a 5- to 6-membered monocyclic ring. In some embodiments, the heterocyclyl group contains 0 to 3 double bonds. In any of the foregoing embodiments, the heterocyclyl contains 1, 2, 3, or 4 heteroatoms. Any nitrogen or sulfur heteroatom may be optionally oxidized (e.g., NO, SO, SO2), and any nitrogen heteroatom may be optionally quaternized (e.g., [NR4] + Cl - , [NR4] + OH -Representative examples of heterocyclyl include oxiranyl, aziridinyl, thiiranyl, azetidinyl, oxetanyl, thietanyl, 1,2-dithietanyl, 1,3-dithietanyl, pyrrolidinyl, dihydro-1H-pyrrolyl, dihydrofuranyl, tetrahydropyranyl, dihydrothienyl, tetrahydrothienyl, imidazolidinyl, piperidinyl, piperazinyl, morpholinyl, thiomorpholinyl, 1,1-dioxo-thiomorpholinyl, dihydropyranyl, tetrahydropyranyl, hexahydrothiopyranyl, hexahydropyrimidinyl, oxazinanyl. , thiazinanyl, thioxanyl, homopiperazinyl, homopiperidinyl, azepanyl, oxepanyl, thiepanyl, oxazepinyl, oxazepanyl, diazepanyl, 1,4-diazepanyl, diazepinyl, thiazepinyl, thiazepanyl, tetrahydrothiopyranyl, oxazolidinyl, thiazolidinyl, isothiazolidinyl, 1,1-dioxoisothiazolidinonyl, oxazolidinonyl, imidazolidinonyl, 4,5,6,7-tetrahydro[2H]indazolyl, tetrahydrobenzimidazolyl, 4,5,6,7-tetrahydrobenzo[d ]imidazolyl, 1,6-dihydroimidazole[4,5-d]pyrrolo[2,3-b]pyridinyl, thiazinyl, thiophenyl, oxazinyl, thiadiazinyl, oxadiazinyl, dithiazinyl, dioxazinyl, oxathiazinyl, thiatriazinyl, oxatriazinyl, dithiadiazinyl, imidazolinyl, dihydropyrimidyl, tetrahydropyrimidyl, 1-pyrrolinyl, 2-pyrrolinyl, 3-pyrrolinyl, indolinyl, thiapyranyl, 2H-pyranyl, 4H-pyranyl, dioxanyl, 1,3-dioxolanyl, pyrazolinyl, pyrazolinyl, Zolidinyl, dithianyl, dithiolanyl, pyrimidinonyl, pyrimidindionyl, pyrimidine-2,4-dionyl, piperazinonyl, piperazinedionyl, pyrazolidinylimidazolinyl, 3-azabicyclo[3.1.0]hexanyl, 3,6-diazabicyclo[3.1.1]heptanyl, 6-azabicyclo[3.1.1]heptanyl, 3-azabicyclo[3.1.1]heptanyl, 3-azabicyclo[4.1.0]heptanyl, azabicyclo[2.2.2]hexanyl, 2-azabicyclo[3.2.1]octanyl, 8-azabicyclo[3.2.1]octanyl, 2-azabicyclo[2.2.2]octanyl, 8-azabicyclo[2.2.2]octanyl, 7-oxabicyclo[2.2.1]heptane, azaspiro[3.5]nonanyl, azaspiro[2.5]octanyl, azaspiro[4.5]decanyl, 1-azaspiro[4.5]decan-2-one, azaspiro[5.5]undecanyl, tetrahydroindolyl, octahydroindolyl, tetrahydroisoindolyl, tetrahydroindazolyl, 1,1-dioxohexahydrothiopyranyl. Examples of 5-membered heterocyclyls containing a sulfur or oxygen atom and 1 to 3 nitrogen atoms are thiazolyl, including thiazol-2-yl and thiazol-2-yl N-oxide, thiadiazolyl, including 1,3,4-thiadiazol-5-yl and 1,2,4-thiadiazol-5-yl, oxazolyl, such as oxazol-2-yl, and oxadiazolyl, such as 1,3,4-oxadiazol-5-yl and 1,2,4-oxadiazol-5-yl. Examples of 5-membered heterocyclyls containing 2 to 4 nitrogen atoms include imidazolyls such as imidazol-2-yl, triazolyls such as 1,3,4-triazol-5-yl, 1,2,3-triazol-5-yl, 1,2,4-triazol-5-yl, and tetrazolyls such as 1H-tetrazol-5-yl. Representative examples of benzo-fused 5-membered heterocyclyls are benzoxazol-2-yl, benzothiazol-2-yl and benzimidazol-2-yl. Examples of 6-membered heterocyclyls contain 1 to 3 nitrogen atoms and optionally sulfur or oxygen atoms, such as pyridyl, e.g. pyrid-2-yl, pyrid-3-yl and pyrid-4-yl; pyrimidyl, e.g. pyrimidyl and pyrimid-4-yl; triazinyl, e.g. 1,3,4-triazin-2-yl and 1,3,5-triazin-4-yl; pyridazinyl, especially pyridazin-3-yl and pyrazinyl. Pyridine N-oxide and pyridazine N-oxide, as well as pyridyl, pyrimid-2-yl, pyrimid-4-yl, pyridazinyl and 1,3,4-triazin-2-yl groups, are further examples of heterocyclyl groups. In some embodiments, a heterocyclic group comprises a heterocycle fused to one or more (e.g., 1, 2, or 3) different cyclic groups (e.g., carbocycles or heterocycles), where the radical or point of attachment is on the heterocycle, and in some embodiments, the point of attachment is a heteroatom contained in the heterocycle.

[0039] Thus, the term "heterocyclic" includes N-heterocyclyl groups, which as used herein refer to heterocyclyl groups containing at least one nitrogen, where the point of attachment of the heterocyclyl group to the remainder of the molecule is through a nitrogen atom in the heterocyclyl group. Representative examples of N-heterocyclyl groups include 1-morpholinyl, 1-piperidinyl, 1-piperazinyl, 1-pyrrolidinyl, pyrazolidinyl, imidazolinyl and imidazolidinyl. The term "heterocyclic" also includes C-heterocyclyl groups, which as used herein refer to heterocyclyl groups containing at least one heteroatom, where the point of attachment of the heterocyclyl group to the remainder of the molecule is through a carbon atom in the heterocyclyl group. Representative examples of C-heterocyclyl groups include 2-morpholinyl, 2- or 3- or 4-piperidinyl, 2-piperazinyl, and 2- or 3-pyrrolidinyl. The term "heterocycle" also refers to the ring of formula -R, as disclosed above. c Heterocyclyl (wherein R c is an alkylene chain). The term "heterocycle" as used herein also includes heterocyclylalkyl groups, which refer to groups of the formula -OR c -heterocyclyl (wherein R c is an alkylene chain), including heterocyclylalkoxy groups.

[0040] As used herein, the term "aryl" used alone or as part of a larger moiety (e.g., "aralkyl" where the atom of the terminal carbon on the alkyl is the point of attachment, e.g., a benzyl group), "aralkoxy" where the oxygen is the point of attachment, or "aralkoxyalkyl" where the point of attachment is on the aryl) refers to a group that includes a monocyclic, bicyclic, or tricyclic carbocyclic ring system, including fused rings, where at least one ring in the system is aromatic. Unless otherwise defined for any particular group in the compounds of formula (I), in some embodiments, the aralkoxy group is a benzoxy group. The term "aryl" may be used interchangeably with the term "aryl ring." In one embodiment, aryl includes groups having 6 to 18 carbon atoms. In another embodiment, aryl includes groups having 6 to 10 carbon atoms. Examples of aryl groups include phenyl, naphthyl, anthracyl, biphenyl, phenanthrenyl, naphthacenyl, 1,2,3,4-tetrahydronaphthalenyl, 1H-indenyl, 2,3-dihydro-1H-indenyl, naphthyridinyl, and the like, which may be substituted or independently substituted by one or more substituents described herein. A specific aryl is phenyl. In some embodiments, an aryl group comprises an aryl ring fused to one or more (e.g., one, two or three) different cyclic groups (e.g., carbocyclic or heterocyclic rings), where the radical or point of attachment is on the aryl ring. The structure of any aryl group that may have differently positioned double bonds is considered to include any and all such resonance structures.

[0041] Thus, the term aryl refers to a group of the formula -R c Aryl (wherein R c is an alkylene chain such as methylene or ethylene), in some embodiments, the aralkyl group is an optionally substituted benzyl group. The term aryl, as used herein, also includes aryl groups of the formula -OR c -aryl (wherein R cincludes aralkoxy groups, which refer to groups attached through an oxygen atom of an alkylene chain such as methylene or ethylene.

[0042] As used herein, the term "heteroaryl," used alone or as part of a larger moiety (e.g., "heteroarylalkyl" (also "heteroaralkyl"), or "heteroarylalkoxy" (also "heteroaralkoxy"), refers to a monocyclic, bicyclic, or tricyclic ring system having 5 to 14 ring atoms, in which at least one ring is aromatic and contains at least one heteroatom. In one embodiment, heteroaryl includes a 5-6 membered monocyclic aromatic group in which one or more ring atoms are independently optionally substituted nitrogen, sulfur, or oxygen. In another embodiment, heteroaryl includes a 5-6 membered monocyclic aromatic group in which one or more ring atoms are nitrogen, sulfur, or oxygen. Representative examples of heteroaryl groups include thienyl, furyl, imidazolyl, pyrazolyl, thiazolyl, isothiazolyl, oxazolyl, isoxazolyl, triazolyl, thiadiazolyl, oxadiazolyl, tetra ... Zolyl, thiatriazolyl, oxatriazolyl, pyridyl, pyrimidyl, imidazopyridyl, pyrazinyl, pyridazinyl, triazinyl, tetrazinyl, tetrazolo[1,5-b]pyridazinyl, purinyl, deazapurinyl, benzoxazolyl, benzofuryl, benzothiazolyl, benzothiadiazolyl, benzotriazolyl, benzimidazolyl, indolyl, 1,3-thiazol-2-yl, 1,3,4-triazol-5-yl, 1,3-oxazol-2-yl The term "heteroaryl" also includes groups in which a heteroaryl is fused to one or more cyclic (e.g., carbocyclyl, or heterocyclyl) rings where the radical or point of attachment is on the heteroaryl ring.Non-limiting examples include indolyl, indolizinyl, isoindolyl, benzothienyl, benzothiophenyl, methylenedioxyphenyl, benzofuranyl, dibenzofuranyl, indazolyl, benzimidazolyl, benzodioxazolyl, benzothiazolyl, quinolyl, isoquinolyl, cinnolinyl, phthalazinyl, quinazolinyl, quinoxalinyl, 4H-quinolizinyl, carbazolyl, acridinyl, phenazinyl, phenothiazinyl, phenoxazinyl, tetrahydroquinolinyl, tetrahydroisoquinolinyl, and pyrido[2,3-b]-1,4-oxazin-3(4H)-one. Heteroaryl groups may be monocyclic, bicyclic, or tricyclic. In some embodiments, a heteroaryl group comprises a heteroaryl ring fused to one or more (e.g., 1, 2 or 3) different cyclic groups (e.g., carbocyclic or heterocyclic rings), where the radical or point of attachment is on the heteroaryl ring, and in some embodiments, the point of attachment is a heteroatom contained in the heterocyclic ring. The structure of any heteroaryl group that can have differently positioned double bonds is considered to encompass any and all such resonance structures.

[0043] The term heteroaryl, as used herein, also refers to the heteroaryl group defined above and includes N-heteroaryl groups containing at least one nitrogen atom, the point of attachment of the N-heteroaryl group to the remainder of the molecule being through a nitrogen atom in the heteroaryl group. The term heteroaryl, as used herein, further includes C-heteroaryl groups, which refer to the heteroaryl group defined above and the point of attachment of the heteroaryl group to the remainder of the molecule being through a carbon atom in the heteroaryl group. The term heteroaryl, as used herein, further includes heteroarylalkyl groups, which are represented by the formula -R, as disclosed above. c -heteroaryl, where R c is an alkylene chain as defined above. The term heteroaryl as used herein refers to a group of the formula -OR c -heteroaryl (wherein R cFurther encompassed are heteroaralkoxy (or heteroarylalkoxy) groups which refer to groups attached through an oxygen atom of an alkylene group, as defined above.

[0044] Unless otherwise stated, and unless further defined for any particular group(s), any of the groups described herein may be substituted or unsubstituted. As used herein, the term "substituted" refers broadly to all permissible substituents, with the implicit proviso that such substitution results in a stable compound, i.e., a compound that does not spontaneously undergo transformation by rearrangement, cyclization, elimination, and the like, according to the permissible valences of the substituted atom and the substituent. Representative substituents include halogens, hydroxyl groups, and any other organic group containing any number of carbon atoms, e.g., 1-14 carbon atoms, which may contain one or more (e.g., 1, 2, 3, or 4) heteroatoms, such as oxygen, sulfur, and nitrogen, categorized in a linear, branched, or cyclic structural format.

[0045] Thus, unless further defined for any particular group, representative examples of substituents include alkyl, substituted alkyl (e.g., C1-C6, C 1~5 , C 1~4 , C 1~3 , C 1~2 , C1), alkoxy (e.g., C1 to C6, C 1~5 , C 1~4 , C 1~3 , C 1~2 , C1), substituted alkoxy (e.g., C1-C6, C 1~5 , C 1~4 , C 1~3 , C 1~2 , C1), haloalkyl (e.g., CF3), alkenyl (e.g., C2-C6, C 2~5 , C 2~4 , C 2~3 , C2), substituted alkenyl (e.g., C2-C6, C 2~5 , C 2~4 , C 2~3 , C2), alkynyl (e.g., C2-C6, C 2~5 , C 2~4 , C2~3 , C2), substituted alkynyl (e.g., C2-C6, C 2~5 , C 2~4 , C 2~3 , C2), cyclic (e.g., C3-C 12 , C5-C6), substituted cyclic (e.g., C3-C 12 , C5-C6), carbocyclic (e.g., C3-C 12 , C5-C6), substituted carbocyclic rings (e.g., C3-C 12 , C5-C6), heterocycles (e.g., C3-C 12 , C5-C6), substituted heterocycles (e.g., C3-C 12 , C5-C6), aryl (e.g., benzyl and phenyl), substituted aryl (e.g., substituted benzyl or phenyl), heteroaryl (e.g., pyridyl or pyrimidyl), substituted heteroaryl (e.g., substituted pyridyl or pyrimidyl), aralkyl (e.g., benzyl), substituted aralkyl (e.g., substituted benzyl), halo, hydroxyl, aryloxy (e.g., C6-C 12 , C6), substituted aryloxy (e.g., C6-C 12 , C6), alkylthio (e.g., C1-C6), substituted alkylthio (e.g., C1-C6), arylthio (e.g., C6-C 12 , C6), substituted arylthio (e.g., C6-C 12 , C6), cyano, carbonyl, substituted carbonyl, carboxyl, substituted carboxyl, amino, substituted amino, amido, substituted amido, thio, substituted thio, sulfinyl, substituted sulfinyl, sulfonyl, substituted sulfonyl, sulfinamide, substituted sulfinamide, sulfonamide, substituted sulfonamide, urea, substituted urea, carbamate, substituted carbamate, amino acid, and peptide groups.

[0046] The zinc finger transcription factor Helios is important for maintaining the identity, anergic phenotype, and suppressive activity of regulatory T cells [PMID:26472910, 27185917, 29440380], and acute pharmacological degradation of Helios can destabilize regulatory T cells ex vivo [PMID:34035522]. Separately, considering the fundamental role that cyclin-dependent kinase 4 / 6 (CDK4 / 6) has in regulating cell cycle progression, inhibitors of CDK4 / 6 have been found to have potent antiproliferative effects [PMID:27030077, 29935901, 26658964]. Recently, these CDK4 / 6 inhibitors have also been found to have immunomodulatory effects, including directly promoting T cell activity [PMID:29101163], enhancing the formation of memory T cells [PMID:33941591], modulating regulatory T cell proliferation [PMID:28813415], and modulating tumor cell immunogenicity [PMID:28813415, 29160310, 30388455]. Thus, targeting both Helios and CDK4 / 6 may have profound effects on both reducing tumor cell proliferation and promoting antitumor immune responses.

[0047] A first aspect of the present disclosure is a compound represented by formula (I): [ka] or a pharma- ceutically acceptable salt or stereoisomer thereof, wherein the targeting ligand binds to CDK4 and / or CDK6 and the linker comprises an alkylene chain or a polyethylene glycol chain.

[0048] The targeting ligand represents a moiety that binds to CDK4 and / or CDK6. In some embodiments, the targeting ligand is derived from palbociclib, ribociclib, or abemaciclib. Palbociclib derivatives are known in the art and are described, for example, in U.S. Patent Nos. 6,936,612, 7,456,168, 10,723,730, and RE47739. Ribociclib derivatives are known in the art and are described, for example, in U.S. Patent Nos. 8,324,225, 8,415,355, 8,685,980, 8,962,630, 9,193,732, 9,416,136, 9,868,739, and 10,799,506. Derivatives of abemaciclib are known in the art and are described, for example, in U.S. Pat. No. 7,855,221.

[0049] In some embodiments, the targeting ligand is represented by any one of the following structures: [ka]

[0050] Thus, in some embodiments, the bifunctional compound has a structure represented by Formula (Ia), Formula (Ib), or Formula (Ic): [ka] or a pharma- ceutically acceptable salt or stereoisomer thereof.

[0051] In some embodiments, the linker contains an alkylene chain (e.g., having 1 to 20 alkylene units). In some embodiments, the linker includes 1 to 6 alkylene units. In some embodiments, the alkylene chain or divalent alkylene chain is -O-, -S-, -N(R')-, -C≡C-, -C(O)-, -C(O)O-, -OC(O)-, -OC(O)O-, -C(NOR')-, -C(O)N(R')-, -C(O)N(R')C(O)-, -C(O)N(R')C(O)N(R')-, -N(R')C(O)-, -N(R')C(O)N(R')-, -N(R')C(O)O-, -OC(O)N(R')-, - C(NR')-, -N(R')C(NR')-, -C(NR')N(R')-, -N(R')C(NR')N(R')-, -OB(Me)O-, -S(O)2-, -OS(O)-, -S(O)O-, -S(O)-, -OS(O )2-, -S(O)2O-, -N(R')S(O)2-, -S(O)2N(R')-, -N(R')S(O)-, -S(O)N(R')-, -N(R')S(O)2N(R')-, -N(R')S(O)N(R')-, C3~C 12 It may be interrupted and / or terminated (at either or both termini) by at least one of a carbocyclene, a 3- to 12-membered heterocyclene, a 5- to 12-membered heteroarylene, or any combination thereof, where R' is H or a C1-C6 alkyl, and the interrupting group and either or both terminus groups may be the same or different.

[0052] In some embodiments, the alkylene chain is interrupted and / or terminated at either or both ends by at least one of -O- or -NH-.

[0053] In other embodiments, the linker comprises a polyethylene glycol chain, the polyethylene glycol chain being selected from the group consisting of -S-, -N(R')-, -C≡C-, -C(O)-, -C(O)O-, -OC(O)-, -OC(O)O-, -C(NOR')-, -C(O)N(R')-, -C(O)N(R')C(O)-, -C(O)N(R')C(O)N(R')-, -N(R')C(O)-, -N(R')C(O)N(R')-, -N(R')C(O)O-, -OC(O)N(R')-, -C(NR')-, -N (R')C(NR')-, -C(NR')N(R')-, -N(R')C(NR')N(R')-, -OB(Me)O-, -S(O)2-, -OS(O)-, -S(O)O-, -S(O)-, -OS(O)2-, -S(O)2O-, -N(R')S(O)2-, -S(O)2N(R')-, -N(R')S(O)-, -S(O)N(R')-, -N(R')S(O)2N(R')-, -N(R')S(O)N(R')-, C 3~12 It may be terminated (at either or both termini) with at least one of a carbocyclene, a 3- to 12-membered heterocyclene, a 5- to 12-membered heteroarylene, or any combination thereof, where R' is H or a C1-C6 alkyl, and one or both termini may be the same or different.

[0054] In some embodiments, the polyethylene glycol chain is terminated at either or both ends with at least one of -O- or -NH-. In some embodiments, the linker comprises 1 to 3 ethylene glycol units. In some embodiments, the linker has the structure: [ka] In some embodiments, the linker is attached to the phenyl group at the 3-position (meta). In some embodiments, the linker is attached to the phenyl group at the 4-position (para).

[0055] The term "binding", when referring to the interaction between a targeting ligand and two targeting proteins, which in this disclosure are CDK4 and / or CDK6, typically refers to a molecular interaction that is preferential (also referred to herein as "selective") in that the binding of the targeting ligand to other proteins present in the cell, including other CDK isoforms, is substantially less and functionally insignificant, at least from the perspective of degradation. The terms "selective" and "selectivity" refer to the ability of a bifunctional compound to discriminate between molecular targets. The selective dual CDK4 / 6 degraders described herein are capable of selectively binding to one or more of CDK1, CDK2, CDK7, CDK8, CDK9, CDK11, CDK12, CDK13, CDK14, etc., and other kinases. 50 DCs with at least about 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10-fold lower CDK4 / 6 activity than 50 (half-maximal degradation concentration). Thus, although various bifunctional compounds of the present disclosure bind to other CDK proteins, they exhibit selective degradation of CDK4 / 6, even if with similar or much lower affinity.

[0056] The term "binding" with respect to the interaction between the group attached to the linker on the opposite side of the targeting ligand of formula (I) and an E3 ubiquitin ligase typically refers to an intermolecular interaction that may or may not exhibit an affinity level that is equal to or greater than the affinity between the targeting ligand and the target protein, but which affinity is nevertheless sufficient to achieve targeted degradation and recruitment of the ligase to the selective degradation of the targeted protein.

[0057] The term "linked" when referring to the interaction between the groups attached to the linkers on opposite sides of the targeting ligand of formula (I) and Helios typically refers to the compound acting as a molecular glue in the sense of recruiting a ubiquitin ligase (in this case CRBN) to a target protein (in this case Helios) to function as a catalyst for targeted protein degradation.

[0058] Representative bifunctional compounds of the present disclosure are as follows: [ka] [ka] [ka] or a pharma- ceutically acceptable salt or stereoisomer thereof.

[0059] The bifunctional compounds of formula (I) may be in the form of free acid or free base, or pharma- ceutically acceptable salts. As used herein, the term "pharma-ceutically acceptable" in the context of salts refers to salts of compounds that do not abrogate the biological activity or properties of the compounds and are relatively non-toxic, i.e., the salt form of the compound can be administered to a subject without causing undesirable biological effects (such as dizziness or stomach upset) or interacting in a detrimental manner with any of the other components of the composition in which it is contained. The term "pharma-ceutically acceptable salts" refers to the products obtained by reacting the compounds of the present disclosure with a suitable acid or base. Examples of pharma-ceutically acceptable salts of the compounds of the present disclosure include those derived from suitable inorganic bases, such as Li, Na, K, Ca, Mg, Fe, Cu, Al, Zn and Mn salts. Examples of pharma- ceutically acceptable non-toxic acid addition salts are salts of amino groups formed with inorganic acids, such as hydrochloride, hydrobromide, hydroiodide, nitrate, sulfate, bisulfate, phosphate, isonicotinate, acetate, lactate, salicylate, citrate, tartrate, pantothenate, bitartrate, ascorbate, succinate, maleate, gentisinate, fumarate, gluconate, glucuronate, saccharate, formate, benzoate, glutamate, methanesulfonate, ethanesulfonate, benzenesulfonate, 4-methylbenzenesulfonate or p-toluenesulfonate, etc. Certain compounds of the present disclosure can form pharma- ceutically acceptable salts with various organic bases, such as lysine, arginine, guanidine, diethanolamine or metformin.

[0060] The bifunctional compounds of formula (I) may have at least one chiral center and therefore may be in the form of stereoisomers, which as used herein includes all isomers of the individual compounds that differ only in the orientation of their atoms in space. The term stereoisomer includes mirror isomers (enantiomers containing the (R-) or (S-) configuration of the compound), mixtures of mirror isomers of the compounds (physical mixtures of enantiomers, and racemates or racemic mixtures), geometric (cis / trans or E / Z, R / S) isomers of the compounds, and isomers of compounds with two or more chiral centers that are not mirror images of each other (diastereoisomers). The chiral centers of the compounds may undergo epimerization in vivo; therefore, for these compounds, administration of the compound in its (R-) form is considered equivalent to administration of the compound in its (S-) form. Thus, the compounds of the disclosure can be prepared and used in the form of individual isomers, substantially free of other isomers, or in the form of mixtures of various isomers, e.g., racemic mixtures of stereoisomers.

[0061] In some embodiments, the bifunctional compounds of formula (I) are isotopically derivatives in that they have at least one desired isotopic substitution of an atom that is greater than the natural abundance of the isotope, i.e., enriched. In one embodiment, the compound contains deuterium or multiple deuterium atoms. Deuterium, i.e. 2 Substitution with heavier isotopes, such as H, can confer certain therapeutic advantages resulting from greater metabolic stability, for example increased in vivo half-life or reduced dosage requirements, and therefore may be advantageous in some circumstances.

[0062] Additionally, the bifunctional compounds of formula (I) encompass the use of N-oxides of the compounds, crystalline forms (also known as polymorphs), active metabolites of the compounds having the same type of activity, tautomers, and unsolvated and solvated forms with pharma- ceutically acceptable solvents such as water, ethanol, etc. Solvated forms of the conjugates presented herein are also considered to be disclosed herein.

[0063] Synthesis method In some embodiments, the present disclosure is directed to a method for making a bifunctional compound of formula (I) or its pharma- ceutically acceptable salt or stereoisomer.Generally, the compound or its pharma- ceutically acceptable salt or stereoisomer can be prepared by any process known to be applicable to the preparation of chemically related compounds.The compounds of the present disclosure will be better understood in conjunction with the synthetic schemes described in various examples, which illustrate the non-limiting methods by which the compounds of the present disclosure can be prepared.

[0064] Pharmaceutical Compositions Another aspect of the present disclosure is directed to a pharmaceutical composition comprising a therapeutically effective amount of a bifunctional compound of formula (I) or a pharma- ceutically acceptable salt or stereoisomer thereof and a pharma- ceutically acceptable carrier. The term "pharma- ceutically acceptable carrier" as known in the art refers to a pharma- ceutically acceptable material, composition or vehicle suitable for administering the compounds of the present disclosure to a mammal. Suitable carriers include, for example, liquids (both aqueous and non-aqueous, and combinations thereof), solids, encapsulating materials, gases, and combinations thereof (e.g., semi-solids), and gases, which function to carry or transport the compound from one organ or part of the body to another organ or part of the body. A carrier is "acceptable" in the sense of being physiologically inert and compatible with the other ingredients of the formulation and not harmful to the subject or patient. Depending on the type of formulation, the composition may include one or more pharma- ceutically acceptable excipients.

[0065] In general, the bifunctional compounds of formula (I) and their pharma- ceutically acceptable salts and stereoisomers can be formulated into a given type of composition according to conventional pharmaceutical practices such as conventional mixing, dissolving, granulating, dragee-making, levigating, emulsifying, encapsulating, entrapping and compression processes (see, for example, Remington: The Science and Practice of Pharmacy (20th ed.), ed. A R Gennaro, Lippincott Williams & Wilkins, 2000 and Encyclopedia of Pharmaceutical Technology, eds. J. Swarbrick and J. C. Boylan, 1988-1999, Marcel Dekker, New York). The type of formulation depends on the mode of administration, which may include enteral (e.g., oral, buccal, sublingual and rectal), parenteral (e.g., subcutaneous (sc), intravenous (iv), intramuscular (im) and intrasternal injection or infusion techniques, intraocular, intraarterial, intramedullary, intrathecal, intraventricular, transdermal, intradermal, intravaginal, intraperitoneal, mucosal, intranasal, intratracheal instillation, bronchial instillation and inhalation) and topical (e.g., transdermal). In general, the most appropriate route of administration will depend on a variety of factors, including, for example, the nature of the agent (e.g., its stability in the environment of the gastrointestinal tract) and / or the condition of the subject (e.g., whether the subject can tolerate oral administration). For example, parenteral (e.g., intravenous) administration may also be advantageous in that the compound may be administered relatively quickly, such as in the case of single-dose treatments and / or acute conditions.

[0066] In some embodiments, the bifunctional compounds are formulated for oral or intravenous administration (eg, systemic intravenous injection).

[0067] Thus, the bifunctional compounds of the present disclosure can be formulated into solid compositions (e.g., powders, tablets, dispersible granules, capsules, cachets, and suppositories), liquid compositions (e.g., solutions in which the compound is dissolved, suspensions in which solid particles of the compound are dispersed, emulsions, and solutions containing liposomes, micelles, or nanoparticles, syrups and elixirs); semi-solid compositions (e.g., gels, suspensions, and creams); and gases (e.g., propellants for aerosol compositions). The compounds can also be formulated for immediate, intermediate, or sustained release.

[0068] Oral solid dosage forms include capsules, tablets, pills, powders, and granules.In these solid dosage forms, the active compound is mixed with a carrier such as sodium citrate or dicalcium phosphate, and a) a filler or extender such as starch, lactose, sucrose, glucose, mannitol, and silicic acid, b) a binder such as methylcellulose, microcrystalline cellulose, hydroxypropylmethylcellulose, carboxymethylcellulose, sodium carboxymethylcellulose, alginate, gelatin, polyvinylpyrrolidinone, sucrose, and acacia, c) a wetting agent such as glycerol, d) a crosslinked polymer (e.g., crosslinked polyvinylpyrrolidone (crospovidone), crosslinked sodium carboxymethylcellulose, (croscarmellose sodium), sodium starch glycolate, agar-agar, calcium carbonate, potato or tapioca starch, alginic acid, certain silicates, and sodium carbonate), e) disintegrating agents such as paraffin, f) absorption accelerators such as quaternary ammonium compounds, g) wetting agents such as cetyl alcohol and glycerol monostearate, h) absorbents such as kaolin and bentonite clay, and i) lubricants such as talc, calcium stearate, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate, and mixtures thereof. In the case of capsules, tablets, and pills, the dosage form may also include buffering agents. Similar types of solid compositions may also be used as fillers in soft and hard-filled gelatin capsules, using excipients such as lactose or milk sugar and high molecular weight polyethylene glycols. Solid dosage forms of tablets, dragees, capsules, pills, and granules can be prepared with coatings and shells, such as enteric coatings and other coatings. They may further contain an opacifying agent.

[0069] In some embodiments, the bifunctional compound of the present disclosure can be formulated in hard or soft gelatin capsules.Representative excipients that can be used include pregelatinized starch, magnesium stearate, mannitol, sodium stearyl fumarate, lactose anhydrous, microcrystalline cellulose and croscarmellose sodium.Gelatin shells can include gelatin, titanium dioxide, iron oxide and coloring agents.

[0070] Liquid dosage forms for oral administration include solutions, suspensions, emulsions, microemulsions, syrups and elixirs.In addition to the compound, liquid dosage forms may contain aqueous or non-aqueous carriers (depending on the solubility of the compound) commonly used in the art, such as water or other solvents, solubilizers and emulsifiers, such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butylene glycol, dimethylformamide, oils (especially cottonseed oil, peanut oil, corn oil, germ oil, olive oil, castor oil and sesame oil), glycerol, tetrahydrofurfuryl alcohol, polyethylene glycol and fatty acid esters of sorbitan, and mixtures thereof.Oral compositions may also contain excipients such as wetting agents, suspending agents, coloring agents, sweeteners, flavoring agents and perfumes.

[0071] Injectable preparations may include sterile aqueous or oily suspensions. They may be formulated according to standard techniques using suitable dispersing or wetting agents and suspending agents. Sterile injectable preparations may also be sterile injectable solutions, suspensions or emulsions in non-toxic parenterally acceptable diluents or solvents, for example, solutions in 1,3-butanediol. Among the acceptable vehicles and solvents that may be used are water, Ringer's solution, USP and isotonic sodium chloride solution. In addition, sterile fixed oils are conventionally used as a solvent or suspending medium. For this purpose, any non-irritating fixed oil may be used, including synthetic mono- or diglycerides. In addition, fatty acids such as oleic acid are used in injectable preparations. Injectable preparations may be sterilized, for example, by filtration through a bacteria-retaining filter, or by incorporating sterilizing agents in the form of sterile solid compositions that can be dissolved or dispersed in sterile water or other sterile injectable medium before use. The effect of a compound can be prolonged by slowing its absorption, which can be accomplished by the use of a liquid suspension of poor water solubility or of crystalline or amorphous material. Prolonged absorption of a compound from a parenterally administered formulation can also be accomplished by suspending the compound in an oil vehicle.

[0072] In certain embodiments, the bifunctional compounds of formula (I) can be administered locally rather than systemically, for example, via injection of the conjugate directly into an organ, often in a depot or sustained release formulation. In certain embodiments, long-acting formulations are administered by implantation (e.g., subcutaneously or intramuscularly) or by intramuscular injection. Injectable depot forms are made by forming microencapsule matrices of the compound in biodegradable polymers, such as polylactide-polyglycolide, poly(orthoesters) and poly(anhydrides). The release rate of the compound can be controlled by varying the ratio of compound to polymer and the nature of the particular polymer used. Depot injectable formulations are also prepared by entrapping the compound in liposomes or microemulsions that are compatible with body tissues. Furthermore, in other embodiments, the compound is delivered in a targeted drug delivery system, for example, liposomes coated with organ-specific antibodies. In such embodiments, the liposomes are targeted to and taken up selectively by the organ.

[0073] The bifunctional compounds can be formulated for buccal or sublingual administration and examples include tablets, lozenges, and gels.

[0074] The bifunctional compound can be formulated for administration by inhalation. Various forms suitable for administration by inhalation include aerosol, mist or powder. The pharmaceutical composition can be delivered in the form of aerosol spray presentation from a pressurized pack or nebulizer using a suitable propellant (e.g., dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide or other suitable gas). In some embodiments, the dosage unit of the pressurized aerosol can be determined by providing a valve to deliver a metered amount. In some embodiments, capsules and cartridges containing, for example, gelatin for use in an inhaler or insufflator can be formulated to contain a powder mixture of the compound and a suitable powder base such as lactose or starch.

[0075] The bifunctional compound of formula (I) can be formulated for topical administration, and topical administration as used herein refers to intradermal administration by applying the formulation to the epidermis.These types of compositions are typically in the form of ointments, pastes, creams, lotions, gels, solutions and sprays.

[0076] Representative examples of carriers useful for formulating compositions for topical application include solvents (e.g., alcohol, polyalcohol, water), creams, lotions, ointments, oils, plasters, liposomes, powders, emulsions, microemulsions, and buffer solutions (e.g., hypotonic or buffered saline). Creams can be formulated with saturated or unsaturated fatty acids, such as, for example, stearic acid, palmitic acid, oleic acid, palmito-oleic acid, cetyl, or oleyl alcohol. Creams can also contain nonionic surfactants, such as polyoxy-40-stearate.

[0077] In some embodiments, topical formulations may also include excipients, examples of which are penetration enhancers. These agents can preferably transport pharmacologically active compounds through the stratum corneum to the epidermis or dermis with little or no systemic absorption. A wide variety of compounds have been evaluated for their effectiveness in enhancing the penetration rate of drugs through the skin. For example, see Percutaneous Penetration Enhancers, Maibach HI and Smith HE (eds.), CRC Press, Inc., Boca Raton, Fla. (1995), which reviews the use and testing of various skin penetration enhancers, and Buyuktimkin et al., Chemical Means of Transdermal Drug Permeation Enhancement in Transdermal and Topical Drug Delivery Systems, Gosh TK, Pfister WR, Yum SI (Eds.), Interpharm Press Inc., Buffalo Grove, Ill. (1997). Representative examples of penetration enhancers include triglycerides (e.g., soybean oil), aloe compositions (e.g., aloe vera gel), ethyl alcohol, isopropyl alcohol, octylphenyl polyethylene glycol, oleic acid, polyethylene glycol 400, propylene glycol, N-decylmethyl sulfoxide, fatty acid esters (e.g., isopropyl myristate, methyl laurate, glycerol monooleate, and propylene glycol monooleate), and N-methylpyrrolidone.

[0078] Representative examples of further excipients that may be included in topical formulations and other types of formulations (to the extent that they are compatible) include preservatives, antioxidants, moisturizers, emollients, buffers, solubilizers, skin protectants, and surfactants. Suitable preservatives include alcohols, quaternary amines, organic acids, parabens, and phenols. Suitable antioxidants include ascorbic acid and its esters, sodium bisulfite, butylated hydroxytoluene, butylated hydroxyanisole, tocopherol, and chelating agents such as EDTA and citric acid. Suitable moisturizers include glycerin, sorbitol, polyethylene glycol, urea, and propylene glycol. Suitable buffers include citric acid, hydrochloric acid, and lactic acid buffers. Suitable solubilizers include quaternary ammonium chlorides, cyclodextrins, benzyl benzoate, lecithin, and polysorbates. Suitable skin protectants include vitamin E oil, allatoin, dimethicone, glycerin, petrolatum, and zinc oxide.

[0079] Transdermal formulations typically use transdermal delivery devices and transdermal delivery patches in which the compound is formulated in a lipophilic emulsion or buffered aqueous solution dissolved and / or dispersed in a polymer or adhesive. Patches can be constructed for continuous, pulsatile, or on-demand delivery of pharmaceuticals. Transdermal delivery of compounds can be achieved by iontophoretic patches. Transdermal patches can provide controlled delivery of compounds, where the absorption rate is slowed by using a rate-controlling membrane or by trapping the compound within a polymer matrix or gel. Absorption enhancers can be used to increase absorption, examples of which include absorbable pharma- ceutically acceptable solvents that aid in passage through the skin.

[0080] Ophthalmic preparations include eye drops.

[0081] Formulations for rectal administration include enemas, rectal gels, rectal foams, rectal aerosols, and retention enemas, which may contain conventional suppository bases such as cocoa butter or other glycerides, as well as synthetic polymers such as polyvinylpyrrolidone, PEG, etc. Compositions for rectal or vaginal administration can also be formulated as suppositories, which can be prepared by mixing the compound with suitable non-irritating carriers and excipients, such as cocoa butter, mixtures of fatty acid glycerides, polyethylene glycol, suppository waxes, and combinations thereof, all of which are solid at ambient temperature but liquid at body temperature, and thus melt in the rectum or vaginal cavity and release the compound.

[0082] Dosage As used herein, the term "therapeutically effective amount" refers to an amount of a bifunctional compound of formula (I) or a pharma- ceutically acceptable salt or stereoisomer thereof; or a composition comprising a bifunctional compound of formula (I) or a pharma- ceutically acceptable salt or stereoisomer thereof, effective to produce a desired therapeutic response in a particular patient suffering from a disease or disorder characterized or mediated by the abnormal activity of any one or more of CDK4 / 6 and Helios. Thus, the term "therapeutically effective amount" includes an amount of a bifunctional compound of the present disclosure or a pharma- ceutically acceptable salt or stereoisomer thereof, when administered, that is sufficient to induce a positive alteration in the disease or disorder being treated, or to prevent the onset or progression of the disease or disorder, or to alleviate to some extent one or more of the symptoms of the disease or disorder being treated in a subject, or to simply kill or inhibit the growth of diseased (e.g., neuroblastoma) cells, or to reduce the amount of CDK4 / 6 and Helios in diseased cells.

[0083] The total daily dosage of bifunctional compound and its method of use can be determined according to standard medical practice, for example, by attending physician using sound medical judgment.The specific therapeutically effective dose for any specific subject can depend on various factors, including the disease or disorder to be treated and its severity (e.g., its current condition); the subject's age, weight, general health, sex and diet; administration time, administration route and excretion rate of the specific compound used; treatment period; the drug used in combination or simultaneously with the bifunctional compound; and similar factors well known in the medical field (for example, see Goodman and Gilman's, The Pharmacological Basis of Therapeutics, 10th Edition, A. Gilman, J. Hardman and L. Limbird, eds., McGraw-Hill Press, 155-173, 2001).

[0084] The bifunctional compounds of formula (I) and their pharma- ceutically acceptable salts and stereoisomers may be effective over a wide dosage range. In some embodiments, the total daily dosage (e.g., for an adult human) may range from about 0.001 to about 1600 mg, 0.01 to about 1600 mg, 0.01 to about 500 mg, about 0.01 to about 100 mg, about 0.5 to about 100 mg, 1 to about 100 to about 400 mg per day, about 1 to about 50 mg per day, and about 5 to about 40 mg per day, and in still other embodiments, may range from about 10 to about 30 mg per day. Individual dosages may be formulated to contain the desired dosage, depending on the number of times the compound is administered per day. By way of example, capsules may be formulated with about 1 to about 200 mg of the bifunctional compound (e.g., 1, 2, 2.5, 3, 4, 5, 10, 15, 20, 25, 50, 100, 150, and 200 mg). In some embodiments, individual doses may be formulated to contain the desired dosage depending on the number of times the compound is to be administered per day.

[0085] How to use In some aspects, the present disclosure is directed to a method of treating a disease or disorder associated with aberrant (e.g., dysfunctional or dysregulated) CDK4 / 6 and / or Helios activity, comprising administering to a subject in need thereof a therapeutically effective amount of a bifunctional compound of formula (I) or a pharma- ceutically acceptable salt or stereoisomer thereof. In some embodiments, the present disclosure is directed to a method of reducing levels of CDK4 and / or CDK6, and Helios in a cell, either in vitro or in vivo, comprising contacting the cell with a bifunctional compound of formula (I).

[0086] The disease or disorder is characterized by or mediated by abnormal activity of CDK4 / 6 and / or Helios (e.g., elevated levels of CDK4 / 6 and / or Helios, or otherwise functionally abnormal CDK4 / 6 and / or Helios, compared to non-pathological conditions). A "disease" is generally considered to be a state of health of a subject in which the subject is unable to maintain homeostasis and the subject's health continues to deteriorate if the disease is not ameliorated. In contrast, a "disorder" in a subject is a state of health in which the subject is able to maintain homeostasis, but the subject's health is less favorable than in the absence of the disorder. If left untreated, the disorder does not necessarily cause further deterioration of the animal's health. In some embodiments, the compounds of formula (I) may be useful in the treatment of cell proliferation diseases and disorders (e.g., cancer or benign neoplasms). As used herein, the term "cell proliferation disease or disorder" refers to a condition characterized by deregulated or abnormal cell proliferation, or both, including neoplasms, precancerous conditions, benign tumors, and noncancerous conditions such as cancer.

[0087] The term "subject" (or "patient") as used herein includes all members of the animal kingdom susceptible to or afflicted with the indicated disease or disorder. In some embodiments, the subject is a mammal, e.g., a human or non-human mammal. The method is also applicable to companion animals such as dogs and cats, as well as livestock such as cows, horses, sheep, goats, pigs, and other domesticated and wild animals. A subject "in need" of treatment according to the present disclosure may "suffer or be suspected of suffering from" a particular disease or disorder, have been positively diagnosed, or otherwise exhibit a sufficient number of risk factors or a sufficient number or combination of signs or symptoms to allow a medical professional to diagnose or suspect that the subject suffers from the disease or disorder. Thus, subjects suffering from and suspected of suffering from a particular disease or disorder are not necessarily two distinct groups.

[0088] Exemplary types of non-cancerous (e.g., cell proliferative) diseases or disorders that may be amenable to treatment with the compounds of the present disclosure include inflammatory diseases and conditions, autoimmune diseases, neurodegenerative diseases, cardiac diseases, viral diseases, chronic and acute kidney diseases or injuries, metabolic diseases, and allergic and genetic diseases.

[0089] Representative examples of specific non-cancerous diseases and disorders include rheumatoid arthritis, alopecia areata, lymphoproliferative conditions, autoimmune blood disorders (e.g., hemolytic anemia, aplastic anemia, anhidrotic ectodermal dysplasia, pure red cell anemia, and idiopathic thrombocytopenia), cholecystitis, acromegaly, rheumatoid spondylitis, osteoarthritis, gout, scleroderma, sepsis, septic shock, dacryocystitis, cryopyrin-associated periodic syndromes (CAPS), endotoxic shock, endometritis, gram-negative sepsis, keratoconjunctivitis sicca, and toxic shock syndrome. , asthma, adult respiratory distress syndrome, chronic obstructive pulmonary disease, chronic pneumonia, chronic transplant rejection, hidradenitis suppurativa, inflammatory bowel disease, Crohn's disease, Behçet's disease, systemic lupus erythematosus, glomerulonephritis, multiple sclerosis, early-onset diabetes mellitus, autoimmune uveoretinitis, autoimmune vasculitis, thyroiditis, Addison's disease, lichen planus, appendicitis, bullous pemphigus, pemphigus vulgaris, pemphigus foliaceus, paraneoplastic pemphigus, myasthenia gravis, immunoglobulin A nephropathy, Hashimoto's disease, Sjogren's syndrome, vitiligo, Wegener's granulomatosis, granulomatous orchitis, autoimmune nephropathy ... Autoimmune oophoritis, sarcoidosis, rheumatic carditis, ankylosing spondylitis, Graves' disease, autoimmune thrombocytopenic purpura, psoriasis, psoriatic arthritis, eczema, dermatitis herpetiformis, ulcerative colitis, pancreatic fibrosis, hepatitis, hepatic fibrosis, CD14-mediated sepsis, non-CD14-mediated sepsis, acute and chronic kidney disease, irritable bowel syndrome, empyema, restenosis, cervicitis, stroke and ischemic injury, neurotrauma, acute and chronic pain, allergic rhinitis, allergic conjunctivitis, chronic heart failure, congestive heart failure, acute coronary syndrome, cachexia, myelopathy, rheumatoid arthritis ... Laria, leprosy, leishmaniasis, Lyme disease, Reiter's syndrome, acute synovitis, muscle degeneration, bursitis, tendinitis, tenosynovitis, herniated disc, ruptured disc-prolapse syndrome, osteopetrosis, rhinosinusitis, thrombosis, silicosis, pulmonary sarcosis, bone resorption diseases such as osteoporosis, fibromyalgia, AIDS and other viral diseases such as shingles, herpes simplex I or II, influenza virus, cytomegalovirus, diabetes mellitus type I and II, obesity, insulin resistance and diabetic retinopathy, 22q11.2 deletion syndrome, Angelman syndrome, Canavan disease, celiac disease, Charcot-Marie-Tooth disease, color blindness, cri de chat, Down syndrome, cystic fibrosis, Duchenne muscular dystrophy, hemophilia, Klinefelter syndrome, neurofibromatosis, phenylketonuria, Prader-Willi syndrome, sickle cell disease, Tay-Sachs disease, Turner syndrome, urea cycle disorders, thalassemia, otitis, pancreatitis, parotitis, pericarditis, peritonitis, pharyngitis, pleuritis, phlebitis, pneumonia, uveitis, polymyositis, proctitis, interstitial pulmonary fibrosis, dermatomyositis, atherosclerosis, arteriosclerosis, amyotrophic lateral sclerosis, asocial disorders, varicose veins, vaginitis, depression, and sudden infant death syndrome.

[0090] In some embodiments, the bifunctional compounds may be useful in the treatment of neurodegenerative diseases and disorders. As used herein, the term "neurodegenerative diseases and disorders" refers to conditions characterized by progressive degeneration or death, or both, of nerve cells, including problems with movement (ataxia) or mental function (dementia). Representative examples of such diseases and disorders include Alzheimer's disease (AD) and AD-related dementia, Parkinson's disease (PD) and PD-related dementia, prion diseases, motor neuron diseases (MND), Huntington's disease (HD), Pick's syndrome, spinocerebellar ataxia (SCA), spinal muscular atrophy (SMA), primary progressive aphasia (PPA), amyotrophic lateral sclerosis (ALS), traumatic brain injury (TBI), multiple sclerosis (MS), dementia (e.g., vascular dementia (VaD), Lewy body dementia (LBD), semantic dementia, and frontotemporal dementia (FTD).

[0091] In some embodiments, bifunctional compounds can be useful in the treatment of autoimmune diseases and disorders.As used herein, the term "autoimmune disease" refers to the condition in which immune system produces antibodies that attack normal body tissue.Representative examples of such diseases include Sjogren's syndrome, Hashimoto's thyroiditis, chronic rheumatoid arthritis, juvenile (type 1) diabetes, polymyositis, scleroderma, Addison's disease, lupus including systemic lupus erythematosus, vitiligo, pernicious anemia, glomerulonephritis, pulmonary fibrosis, celiac disease, polymyalgia rheumatica, multiple sclerosis, ankylosing spondylitis, alopecia areata, vasculitis, and temporal arteritis.

[0092] In other embodiments, the method is directed to treating subjects with cancer.In general, the bifunctional compounds of the present disclosure can be effective in treating carcinomas (solid tumors, including both primary and metastatic tumors), sarcomas, melanomas, and hematological cancers (cancers that affect the blood, including lymphocytes, bone marrow, and / or lymph nodes), such as leukemia, lymphoma, and multiple myeloma.Includes adult tumors / cancers and pediatric tumors / cancers.Cancers can be vascularized or not yet substantially vascularized or non-vascularized tumors.

[0093] Representative examples of cancer include adrenal cortical carcinoma, AIDS-related cancer (e.g., Kaposi's sarcoma and AIDS-related lymphoma), appendix cancer, childhood cancer (e.g., childhood cerebellar astrocytoma, childhood cerebral astrocytoma), basal cell carcinoma, skin (non-melanoma) cancer, biliary tract cancer, extrahepatic bile duct cancer, intrahepatic bile duct cancer, bladder cancer, urinary bladder cancer, brain tumors (e.g., gliomas and glioblastomas (e.g., brain stem glioma, gestational trophoblastic tumor glioma, cerebellar astrocytoma, cerebral astrocytoma / malignant glioma, ependymoma, medulloblastoma, supratentorial primitive neuroectodermal tumor, visual pathway and hypothalamic glioma), breast cancer, bronchial adenoma / carcinoid, carcinoma, id tumors, cancers of the nervous system (e.g., central nervous system cancer, central nervous system lymphoma), cervical cancer, chronic myeloproliferative disorders, colorectal cancer (e.g., colon cancer, rectal cancer), lymphatic tumors, mycosis fungoides, Sezary syndrome, endometrial cancer, esophageal cancer, extracranial germ cell tumors, extragonadal germ cell tumors, extrahepatic bile duct cancer, eye cancer, intraocular melanoma, retinoblastoma, gallbladder cancer, gastrointestinal cancer (e.g., gastric cancer, small intestine cancer, gastrointestinal carcinoid tumors, gastrointestinal stromal tumors (GIST)), cholangiocarcinoma, germ cell tumors, ovarian germ cell tumors, head and neck cancer, neuroendocrine tumors, Hodgkin's cancer, Ann Arbor stage III and IV childhood non-small cell lung cancer Hodgkin's lymphoma, ROS1-positive refractory non-Hodgkin's lymphoma, leukemia, lymphoma, multiple myeloma, hypopharyngeal carcinoma, intraocular melanoma, eye cancer, pancreatic islet cell tumors (pancreatic endocrine tumors), kidney cancer (e.g., Wilms' tumor, renal cell carcinoma), liver cancer, lung cancer (e.g., non-small cell lung cancer and small cell lung cancer), ALK-positive anaplastic large cell carcinoma, ALK-positive advanced malignant solid tumors, Waldenstrom's macroglobulinoma, melanoma, intraocular (eye) melanoma, Merkel cell carcinoma, mesothelioma, primary occult metastatic squamous cell cervical cancer, multiple endocrine neoplasia (MEN), myelodysplastic syndromes, myelodysplasia / myeloma Proliferative diseases, nasopharyngeal carcinoma, neuroblastoma, oral cancer (e.g., lip cancer, oral cavity cancer, tongue cancer, oropharynx cancer, pharyngeal cancer, laryngeal cancer), ovarian cancer (e.g., ovarian epithelial cancer, ovarian germ cell tumor, ovarian low malignant potential tumor), pancreatic cancer, islet cell pancreatic cancer, paranasal sinus and nasal cancer, parathyroid cancer, penile cancer, pharyngeal cancer, pheochromocytoma, pineoblastoma, metastatic anaplastic thyroid cancer, anaplastic thyroid cancer, papillary thyroid cancer, pituitary tumor, plasma cell neoplasm / multiple myeloma, pleuropulmonary blastoma, prostate cancer, retinoblastoma, rhabdomyosarcoma, salivary gland cancer, uterine cancer, (e.g., endometrial uterine carcinoma, uterine sarcoma, endometrial carcinoma), squamous cell carcinoma, testicular cancer, thymoma,These include thymic carcinoma, thyroid carcinoma, juvenile xanthogranuloma, transitional cell carcinoma of the renal pelvis, ureter and other urinary tracts, urethral carcinoma, gestational trophoblastic neoplasm, vaginal cancer, vulvar cancer, hepatoblastoma, rhabdominal tumor, and Wilms' tumor.

[0094] Sarcomas that may be treatable with the bifunctional compounds of the present disclosure include both soft tissue and bone cancers, representative examples of which include osteosarcoma or osteogenic sarcoma (bone) (e.g., Ewing's sarcoma), chondrosarcoma (cartilage), leiomyosarcoma (smooth muscle), rhabdomyosarcoma (skeletal muscle), mesothelioma or mesothelioma (membranous lining of body cavities), fibrosarcoma (fibrous tissue), angiosarcoma or hemangioendothelioma (blood vessels), liposarcoma (fatty tissue), glioma or astrocytoma (neurogenic connective tissue found in the brain), myxosarcoma (primitive embryonic connective tissue), mesenchymal or mixed mesodermal tumor (mixed connective tissue type), and histiocytic sarcoma (immune cancer).

[0095] In some embodiments, the methods of the disclosure involve treatment of a subject having a cell proliferative disease or disorder of the blood system, liver, brain, lung, colon, pancreas, prostate, ovaries, breast, skin, and endometrium.

[0096] As used herein, "cell proliferative diseases or disorders of the blood system" includes lymphoma, leukemia, myeloid neoplasm, mast cell neoplasm, myelodysplasia, benign monoclonal gammopathy, lymphomatoid papulosis, polycythemia vera, chronic myeloid leukemia, myeloid metaplasia of unknown etiology, and essential thrombocythemia. Representative examples of hematological cancers are thus multiple myeloma, lymphoma (including T-cell lymphoma, Hodgkin's lymphoma, non-Hodgkin's lymphoma (diffuse large B-cell lymphoma (DLBCL), follicular lymphoma (FL), mantle cell lymphoma (MCL) and ALK+ anaplastic large cell lymphoma (e.g., B-cell non-Hodgkin's lymphoma selected from diffuse large B-cell lymphoma (e.g., germinal center B-cell-like diffuse large B-cell lymphoma or activated B-cell-like diffuse large B-cell lymphoma)), Burkitt's lymphoma / leukemia, mantle cell lymphoma, mediastinal (thymic) large B-cell lymphoma, follicular lymphoma, ALK+ anaplastic large cell lymphoma (e.g., B-cell non-Hodgkin's lymphoma selected from diffuse large B-cell lymphoma (e.g., germinal center B-cell-like diffuse large B-cell lymphoma or activated B-cell-like diffuse large B-cell lymphoma)), ALK+ anaplastic large cell lymphoma (e.g., B-cell non-Hodgkin's lymphoma ... lymphoma, marginal zone lymphoma, lymphoplasmacytic lymphoma / Waldenstrom macroglobulinemia, metastatic pancreatic adenocarcinoma, refractory B-cell non-Hodgkin's lymphoma, and relapsed B-cell non-Hodgkin's lymphoma, childhood lymphomas, as well as lymphomas of lymphocytic and cutaneous origin (e.g., small lymphocytic lymphoma), leukemias (including childhood leukemia, hairy cell leukemia, acute lymphocytic leukemia, acute myeloid leukemia, acute myeloid leukemia (e.g., acute monocytic leukemia), chronic lymphocytic leukemia, small lymphocytic leukemia, chronic myelogenous leukemia, chronic myelogenous leukemia, and mast cell leukemia), myeloid neoplasms, and mast cell neoplasms.

[0097] As used herein, "hepatic cell proliferation disease or disorder" includes all forms of cell proliferation disorder that affect the liver.Hepatic cell proliferation disorder can include liver cancer (e.g., hepatocellular carcinoma, intrahepatic cholangiocarcinoma and hepatoblastoma), precancerous or precancerous conditions of the liver, benign growth or lesion of the liver, and malignant growth or lesion of the liver, as well as metastatic lesions in the tissues and organs of the body other than the liver.Hepatic cell proliferation disorder can include hepatic hyperplasia, metaplasia, and dysplasia.

[0098] As used herein, "brain cell proliferation disease or disorder" includes all forms of cell proliferation disorder that affect the brain.Brain cell proliferation disorder can include brain cancer (e.g., glioma, glioblastoma, meningioma, pituitary adenoma, vestibular schwannoma, and primitive neuroectodermal tumor (medulloblastoma)), brain precancer or precancerous condition, brain benign growth or lesion, and brain malignant growth or lesion, as well as metastatic lesion in body tissue and organ other than brain.Brain cell proliferation disorder can include brain hyperplasia, metaplasia, and dysplasia.

[0099] As used herein, "pulmonary cell proliferative disease or disorder" includes all forms of cell proliferative disorders affecting lung cells. Pulmonary cell proliferative disorders include lung cancer, precancerous and precancerous conditions of the lung, benign growths or lesions of the lung, hyperplasia, metaplasia, and dysplasia of the lung, and metastatic lesions in tissues and organs of the body other than the lung. Lung cancer includes all forms of lung cancer, such as malignant lung neoplasms, carcinoma in situ, typical carcinoid tumors, and atypical carcinoid tumors. Lung cancer includes small cell lung cancer ("SLCL"), non-small cell lung cancer ("NSCLC"), adenocarcinoma, small cell carcinoma, large cell carcinoma, squamous cell carcinoma, and mesothelioma. Lung cancer may include "scar carcinoma", bronchioloalveolar carcinoma, giant cell carcinoma, spindle cell carcinoma, and large cell neuroendocrine carcinoma. Lung cancer also includes lung neoplasms with histological and ultrastructural heterogeneity (e.g., mixed cell types). In some embodiments, compounds of the present disclosure may be used to treat non-metastatic or metastatic lung cancer (e.g., NSCLC, ALK-positive NSCLC, NSCLC with ROS1 rearrangements, lung adenocarcinoma, and lung squamous cell carcinoma).

[0100] As used herein, "colon cell proliferative disease or disorder" includes all forms of cell proliferative disorders that affect colon cells, including colon cancer, colon precancer or precancerous conditions, colon adenomatous polyps, and colon metachronous lesions.Colon cancer includes sporadic and hereditary colon cancer, malignant colon neoplasms, intraepithelial carcinoma, typical carcinoid tumors, and atypical carcinoid tumors, adenocarcinoma, squamous cell carcinoma, and squamous cell carcinoma.Colon cancer may be associated with hereditary syndromes, such as hereditary nonpolyposis colorectal cancer, familial adenomatous polyposis, MYH-associated polyposis, Gardner syndrome, Peutz-Jeghers syndrome, Turcot syndrome, and juvenile polyposis.Colon cell proliferative disorders may also be characterized by colon hyperplasia, metaplasia, or dysplasia.

[0101] As used herein, "pancreatic cell proliferation disease or disorder" includes all forms of cell proliferation disorder that affect pancreatic cells.Pancreatic cell proliferation disorder can include pancreatic cancer, pancreatic precancer or precancerous condition, pancreatic hyperplasia, pancreatic dysplasia, pancreatic benign growth or lesion, and pancreatic malignant growth or lesion, as well as metastatic lesions in tissues and organs of the body other than pancreas.Pancreatic cancer includes all forms of pancreatic cancer, including ductal adenocarcinoma, adenosquamous carcinoma, pleomorphic giant cell carcinoma, mucinous adenocarcinoma, osteoclast-like giant cell carcinoma, mucinous cystadenocarcinoma, acinar carcinoma, unclassified large cell carcinoma, small cell carcinoma, pancreatoblastoma, papillary tumor, mucinous cystadenoma, papillary cystic tumor, and serous cystadenoma, as well as pancreatic tumor with histochemical and ultrastructural heterogeneity (e.g., mixed cells).

[0102] As used herein, "prostate cell proliferation disease or disorder" includes all forms of cell proliferation disorder that affect the prostate.Prostate cell proliferation disorder can include prostate cancer, prostate pre-cancer or pre-cancerous condition, prostate benign growth or lesion, and prostate malignant growth or lesion, as well as metastatic lesion in body tissues and organs other than prostate.Prostate cell proliferation disorder can include prostate hyperplasia, metaplasia, and dysplasia.

[0103] As used herein, "cell proliferative disease or disorder of the ovary" includes all forms of cell proliferative disorders affecting cells of the ovary. Cell proliferative disorders of the ovary may include precancer or precancerous conditions of the ovary, benign growths or lesions of the ovary, ovarian cancer, and metastatic lesions in tissues and organs of the body other than the ovary. Cell proliferative disorders of the ovary may include ovarian hyperplasia, metaplasia, and dysplasia.

[0104] As used herein, "cell proliferative disease or disorder of the breast" includes all forms of cell proliferative disorders that affect breast cells. Cell proliferative disorders of the breast may include breast cancer, pre-cancer or pre-cancerous conditions of the breast, benign growths or lesions of the breast, and metastatic lesions in body tissues and organs other than the breast. Cell proliferative disorders of the breast may include hyperplasia, metaplasia, and dysplasia of the breast.

[0105] As used herein, "cell proliferative diseases or disorders of the skin" include all forms of cell proliferative disorders affecting skin cells. Cell proliferative disorders of the skin may include precancerous or precancerous conditions of the skin, benign growths or lesions of the skin, melanoma, malignant melanoma or other malignant growths or lesions of the skin, and metastatic lesions in tissues and organs of the body other than the skin. Cell proliferative disorders of the skin may include hyperplasia, metaplasia, and dysplasia of the skin.

[0106] As used herein, "endometrial cell proliferative disease or disorder" includes all forms of cell proliferative disorders that affect cells of the endometrium. Endometrial cell proliferative disorders may include pre-cancer or pre-cancerous conditions of the endometrium, benign endometrial proliferations or lesions, endometrial cancer, and metastatic lesions in tissues and organs of the body other than the endometrium. Endometrial cell proliferative disorders may include endometrial hyperplasia, metaplasia, and dysplasia.

[0107] In some embodiments, the disease or disorder is cancer. In some embodiments, the cancer is characterized by a solid tumor. In some embodiments, the cancer is selected from breast cancer, brain cancer, endometrial cancer, head and neck cancer, gastrointestinal cancer, lung cancer, ovarian cancer, prostate cancer, uterine cancer, hepatocellular carcinoma, liposarcoma, and melanoma. In some embodiments, the cancer is a blood cancer. In some embodiments, the blood cancer is selected from leukemia, lymphoma, and myeloma.

[0108] The bifunctional compounds of formula (I) may be administered to patients, e.g., cancer patients, as monotherapy or in combination therapy. The treatment may be a "front / first line" treatment, i.e., as an initial treatment in patients who have not received a previous anticancer treatment regimen, alone or in combination with other treatments; or a "second line" treatment, i.e., as a treatment in patients who have received a previous anticancer treatment regimen, alone or in combination with other treatments; or a "third line," "fourth line," etc. treatment, alone or in combination with other treatments. The treatment may also be given to patients who have previously received unsuccessful or partially successful treatments but have become intolerant to the treatment. The treatment may also be given as an adjuvant treatment, i.e., to prevent recurrence of cancer in patients who currently have no detectable disease, or after surgical removal of a tumor. Thus, in some embodiments, the bifunctional compounds may be administered to patients who have received another therapy, such as chemotherapy, radioimmunotherapy, surgical therapy, immunotherapy, radiation therapy, targeted therapy, or any combination thereof.

[0109] The disclosed method may involve administering a bifunctional compound of formula (I) or a pharmaceutical composition thereof to a patient in a single dose or multiple doses (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 10, 15, 20 or more doses). For example, the dosing frequency may range from once a day to about once every 8 weeks. In some embodiments, the dosing frequency ranges from about once a day to 1, 2, 3, 4, 5, or 6 weeks, with a 28-day cycle including daily dosing for 3 weeks (21 days) followed by a 7-day "off" period. In other embodiments, the bifunctional compound may be administered twice a day (BID) for 2 1 / 2 days (a total of 5 doses), or once a day (QD) for 2 days (a total of 2 doses). In other embodiments, the bifunctional compound may be administered once a day (QD) for 5 days.

[0110] Combination therapy The bifunctional compounds of formula (I) can be used in combination or simultaneously with at least one other active agent, such as an anti-cancer agent or regimen, in the treatment of diseases and disorders. In some embodiments, the bifunctional compounds of formula (I) can be used in combination with existing immunotherapy, such as immune checkpoint inhibitors (e.g., anti-PD-1 or anti-PD-L1) or cell therapy (e.g., CAR-T cells). The terms "in combination" and "simultaneously" in this context mean that the agents are administered simultaneously, including substantially contemporaneous administration, by the same or separate dosage forms, and by the same or different modes of administration, or sequentially, for example, as part of the same treatment regimen, or by sequential treatment regimens. Thus, when given sequentially at the start of administration of the second compound, the first of the two compounds can still be detected at effective concentrations at the treatment site, in some cases. The order and time intervals can be determined so that they can act together (e.g., synergistically) to provide a greater benefit than if they were administered otherwise. For example, therapeutic agents can be administered at the same time or sequentially in any order at different times, but if not administered at the same time, they can be administered sufficiently close in time to provide the desired therapeutic effect, which may be in a synergistic manner. Thus, the terms are not limited to administration of active agents at exactly the same time.

[0111] In some embodiments, the treatment regimen may include administration of the bifunctional compound of formula (I) in combination with one or more additional therapeutic agents known for use in treating a disease or condition (e.g., cancer). The dosage of the additional anti-cancer therapeutic agent may be the same as or lower than the known or recommended dose. See Hardman et al., eds., Goodman & Gilman's The Pharmacological Basis of Therapeutics, 10th ed., McGraw-Hill, New York, 2001; Physician's Desk Reference 60th ed., 2006. For example, anti-cancer agents that may be suitable for use in combination with the bifunctional compound are known in the art. See, for example, U.S. Pat. No. 9,101,622 (section 5.2 thereof) and U.S. Pat. No. 9,345,705 B2 (columns 12-18 thereof). Representative examples of additional active agents and treatment regimens include radiation therapy, chemotherapeutic agents (e.g., antimitotic agents, angiogenesis inhibitors, antihormones, autophagy inhibitors, alkylating agents, intercalating antibiotics, growth factor inhibitors, antiandrogens, signal transduction pathway inhibitors, antimicrotubule agents, platinum coordination complexes, HDAC inhibitors, proteasome inhibitors, and topoisomerase inhibitors), immunomodulatory agents, therapeutic antibodies (e.g., monospecific and bifunctional antibodies), and CAR-T therapy.

[0112] In some embodiments, the bifunctional compound of Formula (I) and the additional (e.g., anticancer) therapeutic agent may be administered less than 5 minutes apart, less than 30 minutes apart, less than 1 hour apart, about 1 hour apart, about 1 to about 2 hours apart, about 2 to about 3 hours apart, about 3 to about 4 hours apart, about 4 to about 5 hours apart, about 5 to about 6 hours apart, about 6 to about 7 hours apart, about 7 to about 8 hours apart, about 8 to about 9 hours apart, about 9 to about 10 hours apart, about 10 to about 11 hours apart, about 11 to about 12 hours apart, about 12 to 18 hours apart, 18 to 24 hours apart, 24 to 36 hours apart, 36 to 48 hours apart, 48 to 52 hours apart, 52 to 60 hours apart, 60 to 72 hours apart, 72 to 84 hours apart, 84 to 96 hours apart, or 96 to 120 hours apart. Two or more (eg, anti-cancer) therapeutic agents may be administered within the same patient visit.

[0113] In some embodiments involving cancer therapy, the bifunctional compound of formula (I) and the additional anti-cancer or therapeutic agent are administered periodically. Cycling therapy involves the administration of one anti-cancer therapeutic agent for a period of time, followed by the administration of a second anti-cancer therapeutic agent for a period of time, and repeating this sequential administration (i.e., cycle), thereby reducing the occurrence of resistance to one or both of the anti-cancer therapeutic agents, avoiding or reducing the side effects of one or both of the anti-cancer therapeutic agents, and / or improving the efficacy of the treatment. In one example, cycling therapy involves the administration of a first anti-cancer therapeutic agent for a period of time, followed by the administration of a second anti-cancer therapeutic agent for a period of time, optionally followed by the administration of a third anti-cancer therapeutic agent for a period of time, etc., and repeating this sequential administration, i.e., cycle, to reduce the occurrence of resistance to one of the anti-cancer therapeutic agents, avoiding or reducing the side effects of one of the anti-cancer therapeutic agents, and / or improving the efficacy of the anti-cancer therapeutic agents.

[0114] In some embodiments, depending on the particular cancer being treated, the compounds of the disclosure may be administered with or without the use of any of the following: paclitaxel (e.g., ovarian cancer, breast cancer, lung cancer, Kaposi's sarcoma, cervical cancer, and pancreatic cancer), topotecan (e.g., ovarian cancer and lung cancer), irinotecan (e.g., colon cancer and small cell lung cancer), etoposide (e.g., testicular cancer, lung cancer, lymphoma, and non-lymphocytic leukemia), vincristine (e.g., leukemia), leucovorin (e.g., colon cancer), altretamine (e.g., ovarian cancer), daunorubicin (e.g., acute myeloid leukemia (AML), acute lymphocytic leukemia (ALL), or combination chemotherapy (e.g., chemotherapy with other chemotherapy agents). , chronic myeloid leukemia (CML), and Kaposi's sarcoma), trastuzumab (e.g., breast cancer, gastric cancer, esophageal cancer), rituximab (e.g., non-Hodgkin's lymphoma), cetuximab (e.g., colorectal cancer, metastatic non-small cell lung cancer, head and neck cancer), pertuzumab (e.g., metastatic HER2-positive breast cancer), alemtuzumab (e.g., chronic lymphocytic leukemia (CLL), cutaneous T-cell lymphoma (CTCL), and T-cell lymphoma), panitumumab (e.g., colon and rectal cancer), tamoxifen (e.g., breast cancer), fulvestrant (e.g., breast cancer), letrozole (e.g., breast cancer) , exemestane (e.g., breast cancer), azacitidine (e.g., myelodysplastic syndromes), mitomycin C (e.g., gastrointestinal cancer, anal cancer, and breast cancer), dactinomycin (e.g., Wilms' tumor, rhabdomyosarcoma, Ewing's sarcoma, trophoblastic tumor, testicular cancer, and ovarian cancer), erlotinib (e.g., non-small cell lung cancer and pancreatic cancer), sorafenib (e.g., renal cancer and liver cancer), temsirolimus (e.g., renal cancer), bortezomib (e.g., multiple myeloma and mantle cell lymphoma), pegaspagases (e.g., acute lymphoblastic leukemia, etc.), cabozantinib (e.g., bronchial ascites ... (e.g., hepatocellular carcinoma, medullary thyroid carcinoma, renal cell carcinoma, etc.), pembrolizumab (e.g., cervical cancer, gastric cancer, hepatocellular carcinoma, Hodgkin's lymphoma, melanoma, Merkel cell carcinoma, non-small cell lung cancer, urothelial carcinoma, and head and neck squamous cell carcinoma), nivolumab (e.g., colorectal cancer, hepatocellular carcinoma, melanoma, non-small cell lung cancer, renal cell carcinoma, small cell lung cancer, and urothelial carcinoma), regorafenib (e.g., colorectal cancer, gastrointestinal stromal tumor, hepatocellular carcinoma), Cemiplimab (e.g., cutaneous squamous cell carcinoma (CSCC)), avelumab (e.g., Merkel cell carcinoma, urothelial carcinoma, and renal cell carcinoma),It may be used in combination with at least one other anti-cancer agent, such as durvalumab (e.g., bladder and lung cancer), atezolizumab (e.g., urothelial carcinoma, non-small cell lung cancer (NSCLC), triple-negative breast cancer (TNBC), small cell lung cancer (SCLC), and hepatocellular carcinoma (HCC)), and ipilimumab (e.g., melanoma, non-small cell lung cancer (NSCLC), small cell lung cancer (SCLC), bladder cancer, and prostate cancer).

[0115] Medicine Kit The bifunctional compounds of the present invention and / or compositions containing them can be assembled into kits or pharmaceutical systems.The kits or pharmaceutical systems according to this aspect of the present disclosure include a carrier or package such as a box, carton, tube, etc., having one or more containers such as vials, tubes, ampoules, or bottles, which contain the bifunctional compounds of formula (I) or pharmaceutical compositions thereof, in close confinement therein.The kits or pharmaceutical systems of the present disclosure may also include printed instructions for using the bifunctional compounds and compositions. EXAMPLES

[0116] These and other aspects of the present disclosure will be further understood in light of the following examples, which are intended to illustrate certain embodiments of the present disclosure but are not intended to limit its scope as defined by the claims.

[0117] General method

[0118] Unless otherwise stated, reagents and solvents were used as received from commercial suppliers. All reactions were monitored using a Waters® Acquity ultra-performance liquid chromatography / mass spectrometry (UPLC / MS) system using an Acquity UPLC® BEH C18 column (2.1×50 mm, 1.7 μm particle size). UPLC Method A: Solvent gradient = 90% A at 0 min, 5% A at 1.8 min; Method B: Solvent gradient = 85% A at 0 min, 1% A at 1.8 min; Solvent A = 0.1% formic acid in HO; Solvent B = 0.1% formic acid in acetonitrile; Flow rate: 0.6 mL / min. Purification of reaction products was performed by flash chromatography using a CombiFlash® Rf equipped with a Teledyne ISCO RediSep® normal phase silica flash column; or by a Waters® high performance liquid chromatography (HPLC) system using a SunFire® C18 column (19×100 mm, 5 μm particle size): solvent gradient 0%-99% acetonitrile in H2O (0.035% trifluoroacetic acid (TFA) as additive); flow rate: 20 mL / min; or by a SunFire® C18 column (30×250 mm, 5 μm particle size): solvent gradient 0%-99% acetonitrile in H2O (0.035% TFA as additive); flow rate: 40 mL / min. All compounds were >95% pure and analyzed on a Waters® UPLC system. 1 H NMR and 13 C NMR spectra were obtained using a Bruker Avance III spectrometer ( 1 H is 500MHz, 13 The chemical shifts were obtained using a 125 MHz C laser. 1 H NMR is reported relative to deuterated methanol (δ = 3.31) or dimethylsulfoxide (DMSO) (δ = 2.50). Spectra are given in ppm (δ), br = broad, s = singlet, d = doublet, t = triplet, q = quartet, m = multiplet, and coupling constants (J) are reported in Hertz.

[0119] Example 1: General synthetic route. [ka]

[0120] Compounds Ia-1 to Ia-8 can be prepared according to the general synthetic route described above using the appropriate hydroxybenzaldehyde, such as 4-hydroxybenzaldehyde 8, or 3-hydroxybenzaldehyde, the appropriate dibromo or dichloro compound, such as 9a to 9e, and intermediate 7 described below in Example 2. An exemplary synthesis of Ia-8 is described in Example 3.

[0121] Example 2: Synthesis of intermediate 7. [ka]

[0122] 3-(5-Bromo-1-oxoisoindolin-2-yl)piperidine-2,6-dione (3). [ka]

[0123] Potassium carbonate (2.29 g, 16.57 mmol) was added to a solution of methyl 4-bromo-2-(bromomethyl)benzoate 1 (1.70 g, 5.52 mmol) and 3-aminopiperidine-2,6-dione hydrochloride 2 (1 g, 6.08 mmol) in N,N-dimethylformamide (DMF, 18.4 mL, 0.30 M) and stirred at 70° C. for 17 h. The reaction mixture was concentrated under vacuum and water was added to the residue to give a precipitate that was collected by gravity filtration and dried to give the title compound as a solid (1.5 g, 84% yield). LC-MS (ES+): 322.9 m / z [M+H]+.

[0124] tert-Butyl 4-(2-(2,6-dioxopiperidin-3-yl)-1-oxoisoindolin-5-yl)-3,6-dihydropyridine-1(2H)-carboxylate (5). [ka]

[0125] 3-(5-Bromo-1-oxoisoindolin-2-yl)piperidine-2,6-dione 3 (1.5 g, 4.64 mmol) and tert-butyl 4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-3,6-dihydropyridine-1(2H)-carboxylate 4 (1.87 g, 6.03 mmol) were dissolved in DMF (30 mL, 0.155 M) under nitrogen atmosphere. Pd(dppf)Cl2-CH2Cl2 (189 mg, 0.232 mmol) and potassium phosphate (K3PO4, 1.182 g, 5.57 mmol) were added and the reaction mixture was stirred at 100 °C for 15 h. Upon cooling to room temperature, the reaction mixture was diluted with EtOAc. The organic layer was washed with water 2x, brine, dried over Na2SO4, filtered and concentrated in vacuo. Purification (SiO2: 0-100% EtOAc in hexanes) afforded the title compound as a light brown solid (1.5 g, 76%). LC-MS (ES+): 426.2 m / z [M+H]+.

[0126] tert-Butyl 4-(2-(2,6-dioxopiperidin-3-yl)-1-oxoisoindolin-5-yl)piperidine-1-carboxylate (6). [ka]

[0127] tert-Butyl 4-(2-(2,6-dioxopiperidin-3-yl)-1-oxoisoindolin-5-yl)-3,6-dihydropyridine-1(2H)-carboxylate 5 (1.5 g, 3.53 mmol) was dissolved in DMF (20 mL, 0.18 M) in a 100 mL round bottom flask equipped with a stir bar under a N2 atmosphere. Palladium on carbon (10 wt%, 375 mg, 0.10 equiv) was added and the flask was fitted with a H2 balloon and flushed with H2. After stirring at room temperature under a H2 atmosphere for 20 h, the reaction mixture was filtered through Celite to remove the Pd catalyst and rinsed with DMF. The filtrate was concentrated in vacuo to give the title compound as an off-white solid (1.51 g, 99%) which was used further without purification. LC-MS 372.1m / z[M+H-tBu]+, 328.11m / z[M+H-Boc]+.

[0128] 3-(1-oxo-5-(piperidin-4-yl)isoindolin-2-yl)piperidine-2,6-dione hydrochloride (7). [ka]

[0129] A solution of 4M HCl in dioxane (10 mL) was added to a solution of tert-butyl 4-(2-(2,6-dioxopiperidin-3-yl)-1-oxoisoindolin-5-yl)piperidine-1-carboxylate 6 (1.51 g, 3.53 mmol) in CH2Cl2 (10 mL, 0.35 M) and stirred at room temperature for 12 h. The reaction mixture was concentrated in vacuo to give the title compound as a solid (1.156 g, 90%) which was carried forward without purification. 1H NMR(500MHz,DMSO-d6)δ10.98(s,1H), 8.95(d,J=9.7Hz,1H), 8.88-8.72(m,1H), 7.70(d,J=7.9Hz,1H), 7.46(s,1H), 7.41-7.34(m,1H), 5.11(dd,J=13.3,5.1Hz, 1H), 4.45(d,J=17.3Hz,1H), 4.32(d,J=17.3Hz,1H), 3.44-3.36(m,2H), 3.04-2. 88(m,4H), 2.63-2.57(m,1H), 2.39(ddd,J=13.3,4.5Hz,1H), 2.03-1.85(m,5H). LC-MS(ES+):328.05m / z[M+H]+.

[0130] Example 3: Synthesis of (Ia-8). [ka] 3-(5-(1-(4-(2-(4-(6-((6-acetyl-8-cyclopentyl-5-methyl-7-oxo-7,8-dihydropyrido[2,3-d]pyrimidin-2-yl)amino)pyridin-3-yl)piperazin-1-yl)ethoxy)benzyl)piperidin-4-yl)-1-oxoisoindolin-2-yl)piperidine-2,6-dione (Ia-8)

[0131] 4-(2-Bromoethoxy)benzaldehyde (10a). [ka]

[0132] To a solution of 4-hydroxybenzaldehyde 8 (302 mg, 2.47 mmol) in DMF (10 mL, 0.25 M) was added potassium carbonate (410 mg, 2.96 mmol) and stirred at room temperature for 10 min. 1,2-Dibromoethane 9a was added via syringe and the reaction mixture was stirred at room temperature for 15 h. The reaction mixture was extracted with ethyl acetate (EtOAc) and washed with saturated aqueous NH4Cl, water 2x, and brine. The organic layer was collected, dried over Na2SO4, filtered, and concentrated under vacuum. Purification by silica flash chromatography afforded the title compound as a yellow solid (376 mg, 67% yield). LC-MS (ES+): 228.9 m / z [M+H]+.

[0133] 4-(2-(4-(6-((6-acetyl-8-cyclopentyl-5-methyl-7-oxo-7,8-dihydropyrido[2,3-d]pyrimidin-2-yl)amino)pyridin-3-yl)piperazin-1-yl)ethoxy)benzaldehyde (12a). [ka]

[0134] N,N-Diisopropylethylamine (DIPEA, 440 μL, 2.52 mmol) was added to a solution of 4-(2-bromoethoxy)benzaldehyde 10a (376 mg, 0.839 mmol) and palbociclib 11 (250 mg, 1.09 mmol) in DMF (4 mL, 0.21 M) and stirred at 80 °C for 15 h. Upon cooling to room temperature, the reaction mixture was extracted with EtOAc, washed with water 2x, brine, dried over Na2SO4, filtered, and concentrated under vacuum. Purification by silica flash chromatography (0-80% EtOAc / CH2Cl2, then 0-20% MeOH / CH2Cl2) afforded the title compound as a yellow solid (310 mg, 62%). LC-MS (ES+): 596.23 m / z [M+H]+, 298.68 m / z [M+2H]2+.

[0135] 3-(5-(1-(4-(2-(4-(6-((6-acetyl-8-cyclopentyl-5-methyl-7-oxo-7,8-dihydropyrido[2,3-d]pyrimidin-2-yl)amino)pyridin-3-yl)piperazin-1-yl)ethoxy)benzyl)piperidin-4-yl)-1-oxoisoindolin-2-yl)piperidine-2,6-dione (Ia-8). [ka]

[0136] Sodium triacetoxyborohydride (NaBH(OAc), 64 mg, 0.300 mmol) was added to a solution of 4-(2-(4-(6-((6-acetyl-8-cyclopentyl-5-methyl-7-oxo-7,8-dihydropyrido[2,3-d]pyrimidin-2-yl)amino)pyridin-3-yl)piperazin-1-yl)ethoxy)-benzaldehyde 12a (89.5 mg, 0.150 mmol) and 3-(1-oxo-5-(piperidin-4-yl)isoindolin-2-yl)piperidine-2,6-dione hydrochloride 7 (59 mg, 0.180 mmol) in 4:1 DMF / CHCl (3 mL, 0.05 M). Purification by flash chromatography on silica (0-80% EtOAc / CH2Cl2 then 0-20% MeOH / CH2Cl2) gave the title compound as a yellow solid (5.1 mg). 1H NMR(500MHz,DMSO-d6)δ10.97(s,1H), 10.08(s,1H), 8.94(s,1H), 8.05(d,J=3 .0Hz,1H), 7.84(d,J=9.0Hz,1H), 7.63(d,J=7.9Hz,1H), 7.49-7.45(m,2H), 7.3 9(d,J=8.0Hz,1H), 7.23(t,J=8.4Hz,2H), 6.92(d,J=8.5Hz,2H), 5.81(p,J=8.5 Hz,1H), 5.09(dd,J=13.3,5.1Hz,1H), 4.41(d,J=17.2Hz,1H), 4.28(d,J=17.4H) z,1H), 4.12(t,J=5.7Hz,2H), 3.18-3.16(m,3H), 2.95-2.86(m,4H), 2.77(t,J =5.6Hz,2H), 2.69-2.64(m,4H), 2.62-2.56(m,2H), 2.43-2.36(m,4H), 2.30(s, 2H), 2.26-2.21(m,2H), 2.20-2.16(m,2H), 2.09-2.03(m,2H), 2.01-1.95(m,2H) ), 1.89-1.84(m,2H), 1.80-1.74(m,4H), 1.72-1.67(m,2H), 1.61-1.55(m,2H). LC-MS(ES+):907.48m / z[M+H]+,454.44m / z[M+2H]2+.

[0137] Example 4: Synthesis of (Ia-1). [ka] 3-(5-(1-(3-(2-(2-(4-(6-((6-acetyl-8-cyclopentyl-5-methyl-7-oxo-7,8-dihydropyrido[2,3-d]pyrimidin-2-yl)amino)pyridin-3-yl)piperazin-1-yl)ethoxy)ethoxy)benzyl)piperidin-4-yl)-1-oxoisoindolin-2-yl)piperidine-2,6-dione (Ia-1).

[0138] The title compound was prepared according to Example 1. 1H NMR(500MHz,DMSO-d6)δ10.96(s,1H), 10.08(s,1H), 8.94(s,1H), 8.03(d,J=2. 8Hz,1H), 7.84(d,J=9.0Hz,1H), 7.61(d,J=7.8Hz,1H), 7.46(s,1H), 7.44(dd,J= 9.1,2.8Hz,1H), 7.37(d,J=7.9Hz,1H), 7.23(t,J=8.0Hz,1H), 6.94-6.87(m,2H) , 6.83(dd,1H), 5.81(p,J=8.8Hz,1H), 5.08(dd,J=13.3,5.0Hz,1H), 4.39(d,J=1 7.2Hz,1H), 4.26(d,J=17.2Hz,1H), 4.12-4.07(m,2H), 3.77-3.72(m,2H), 3.63 (t,J=5.6Hz,2H), 3.47(s,2H), 3.15-3.09(m,4H), 2.95-2.85(m,3H), 2.64-2.54 (m,8H), 2.42(s,3H), 2.37(dd,J=13.2,4.5Hz,1H), 2.30(s,3H), 2.27-2.19(m,2 H), 2.09-1.94(m,3H), 1.90-1.83(m,2H), 1.81-1.64(m,6H), 1.62-1.53(m,2H). LC-MS(ES+):951.54m / z[M+H]+,476.48m / z[M+2H]2+

[0139] Example 5: Synthesis of (Ia-2). [ka] 3-(5-(1-(4-(2-(2-(4-(6-((6-acetyl-8-cyclopentyl-5-methyl-7-oxo-7,8-dihydropyrido[2,3-d]pyrimidin-2-yl)amino)pyridin-3-yl)piperazin-1-yl)ethoxy)ethoxy)benzyl)piperidin-4-yl)-1-oxoisoindolin-2-yl)piperidine-2,6-dione (Ia-2).

[0140] The title compound was prepared according to Example 1. 1H NMR(500MHz,DMSO-d6)δ10.97(s,1H), 10.09(s,1H), 8.94(s,1H), 8.04(d,J=2.9Hz,1H), 7.84(d,J=9.0Hz,1H), 7.62(d,J=7.9Hz,1H), 7.48-7.42(m,2H), 7.37(d,J=7.8Hz,1H), 7.33-7.20(m,2H), 6.93(d,J=8.1Hz,2H), 5.81(p,J=8.8Hz,1H), 5.09(dd,J=13.3,5.1Hz,1H ), 4.40(d,J=17.2Hz,1H), 4.27(d,J=17.2Hz,1H), 4.14-4.05(m,2H), 3.76-3.72(m,2H), 3.69-3.42(m,4H), 3.21-2.84(m,7H), 2.74-2 .54(m,8H), 2.42(s,3H), 2.40-2.33(m,1H), 2.30(s,3H), 2.28-2.18(m,3H), 2.02-1.94(m,1H), 1.93-1.68(m,9H), 1.62-1.53(m,2H). LC-MS(ES+):951.56m / z[M+H]+,476.48m / z[M+2H]2+.

[0141] Example 6: Synthesis of (Ia-3). [ka] 3-(5-(1-(3-(2-(2-(2-(4-(6-((6-acetyl-8-cyclopentyl-5-methyl-7-oxo-7,8-dihydropyrido[2,3-d]pyrimidin-2-yl)amino)pyridin-3-yl)piperazin-1-yl)ethoxy)ethoxy)ethoxy)benzyl)piperidin-4-yl)-1-oxoisoindolin-2-yl)piperidine-2,6-dione (Ia-3).

[0142] The title compound was prepared according to Example 1. 1H NMR(500MHz,DMSO-d6)δ10.97(s,1H), 10.08(s,1H), 8.94(s,1H), 8.02(d,J=2.9Hz,1H), 7.84(d,J=9.0Hz,1H), 7.61(d,J=7.8Hz,1H), 7.46(s,1H), 7 .42(dd,J=9.1,3.0Hz,1H), 7.36(d,J=7.9Hz,1H), 7.23(t,J=8.0Hz,1H), 6 .91-6.88(m,2H), 6.83(dd,J=8.1,2.3Hz,1H), 5.83-5.78(m,1H), 5.09(dd, J=13.3,5.1Hz,1H), 4.39(d,J=17.2Hz,1H), 4.27(d,J=17.2Hz,1H), 4.08( dd,J=5.5,3.7Hz,2H), 3.76(dd,J=5.5,3.7Hz,2H), 3.62-3.59(m,3H), 3.58 -3.54(m,6H), 3.13-3.10(m,4H), 2.92-2.89(m,2H), 2.59-2.53(m,8H), 2. 42(s,3H), 2.30(s,3H), 1.90(s,3H), 1.78-1.67(m,8H), 1.59-1.54(m,3H). LC-MS(ES+):995.58m / z[M+H]+,498.47m / z[M+2H]2+.

[0143] Example 7: Synthesis of (Ia-4). [ka] 3-(5-(1-(4-(2-(2-(2-(4-(6-((6-acetyl-8-cyclopentyl-5-methyl-7-oxo-7,8-dihydropyrido[2,3-d]pyrimidin-2-yl)amino)pyridin-3-yl)piperazin-1-yl)ethoxy)ethoxy)ethoxy)benzyl)piperidin-4-yl)-1-oxoisoindolin-2-yl)piperidine-2,6-dione (Ia-4).

[0144] The title compound was prepared according to Example 1. 1H NMR(500MHz,DMSO-d6)δ10.97(s,1H), 10.16(s,1H), 8.95(s,1H), 8.02(s,1H), 7.88(d,J=8.5Hz,1H), 7.65(d,J=7.8Hz,1H), 7.60-7.51(m,2H) ), 7.47(d,J=7.6Hz,1H), 7.41(s,1H), 7.33(d,J=7.6Hz,1H), 7.02(d,J=8.4Hz,2H), 5.81(p,J=8.8Hz,1H), 5.08(dd,J=13.2,5.0Hz,1H), 4.41 (d,J=17.3Hz,1H), 4.28(d,J=17.3Hz,1H), 4.16-4.10(m,2H), 3.80-3.74(m,3H), 3.65-3.57(m,6H), 3.23-3.08(m,5H), 3.04(q,J=7.2Hz,3H) , 3.00-2.85(m,4H), 2.63-2.55(m,2H), 2.41(s,4H), 2.30(s,3H), 2.27 -2.05(m,5H), 1.99-1.84(m,6H), 1.79-1.72(m,2H), 1.59-1.54(m,2H). LC-MS(ES+):995.58m / z[M+H]+,498.48m / z[M+2H]2+.

[0145] Example 8: Synthesis of (Ia-5). [ka] 3-(5-(1-(3-(4-(4-(6-((6-acetyl-8-cyclopentyl-5-methyl-7-oxo-7,8-dihydropyrido[2,3-d]pyrimidin-2-yl)amino)pyridin-3-yl)piperazin-1-yl)butoxy)benzyl)piperidin-4-yl)-1-oxoisoindolin-2-yl)piperidine-2,6-dione (Ia-5).

[0146] The title compound was prepared according to Example 1. 1H NMR(500MHz,DMSO-d6)δ10.97(s,1H), 10.08(s,1H), 8.94(s,1H), 8.04(d,J=2.8Hz,1H), 7.84(d,J=9.0Hz,1H), 7.62(d,J=7.8Hz,1H), 7. 48-7.43(m,2H), 7.38(d,J=7.8Hz,1H), 7.23(t,J=7.8Hz,1H), 6.93-6.87(m,2H), 6.82(d,J=7.1Hz,1H), 5.81(p,J=8.8Hz,1H), 5.09(dd, J=13.3,5.1Hz,1H), 4.40(d,J=17.2Hz,1H), 4.27(d,J=17.2Hz,1H), 4.05-3.96(m,3H), 3.17-3.12(m,4H), 2.99-2.84(m,3H), 2.61-2.52 (m,4H), 2.44-2.35(m,7H), 2.30(s,3H), 2.27-2.17(m,4H), 1.98-1.94(m,1H), 1.90-1.83(m,3H), 1.79-1.71(m,8H), 1.65-1.56(m,5H). LC-MS(ES+):935.53m / z[M+H]+,468.45m / z[M+2H]2+.

[0147] Example 9: Synthesis of (Ia-6). [ka] 3-(5-(1-(4-(4-(4-(6-((6-acetyl-8-cyclopentyl-5-methyl-7-oxo-7,8-dihydropyrido[2,3-d]pyrimidin-2-yl)amino)pyridin-3-yl)piperazin-1-yl)butoxy)benzyl)piperidin-4-yl)-1-oxoisoindolin-2-yl)piperidine-2,6-dione (Ia-6).

[0148] The title compound was prepared according to Example 1. 1H NMR(500MHz,DMSO-d6)δ10.97(s,1H), 10.09(s,1H), 8.94(s,1H), 8.05(d,J=2.8Hz,1H), 7.84(d,J=9.0Hz,1H), 7.63(d ,J=7.8Hz,1H), 7.50-7.43(m,2H), 7.38(d,J=7.8Hz,1H), 7.28(s,2H), 6.92(d,J=6.7Hz,2H), 5.81(p,J=8.7Hz,1H),5. 09(dd,J=13.3,5.1Hz,1H), 4.41(d,J=17.3Hz,1H), 4.28(d,J=17.2Hz,1H), 4.05-3.94(m,3H), 3.16(s,4H), 3.04-2.85 (m,3H), 2.62-2.53(m,4H), 2.46-2.35(m,7H), 2.34-2.19(m,7H), 2.01-1.96(m,1H), 1.87(s,3H), 1.82-1.53(m,13H). LC-MS(ES+):935.54m / z[M+H]+,468.46m / z[M+2H]2+.

[0149] Example 10: Synthesis of (Ia-7). [ka] 3-(5-(1-(4-(2-(4-(6-((6-acetyl-8-cyclopentyl-5-methyl-7-oxo-7,8-dihydropyrido[2,3-d]pyrimidin-2-yl)amino)pyridin-3-yl)piperazin-1-yl)ethoxy)benzyl)piperidin-4-yl)-1-oxoisoindolin-2-yl)piperidine-2,6-dione (Ia-7).

[0150] The title compound was prepared according to Example 1. 1H NMR (500MHz, DMSO-d6) δ10.97(s,1H), 10.08(s,1H), 8.95(s,1H), 8.04(d,J =2.6Hz,1H), 7.84(d,J=9.0Hz,1H), 7.62(d,J=7.8Hz,1H), 7.48-7.43(m,2H ), 7.38(d,J=7.9Hz,1H), 7.21(d,J=8.1Hz,2H), 6.87(d,J=8.3Hz,2H), 5.82 (p,J=8.8Hz,1H), 5.09(dd,J=13.3,5.1Hz,1H), 4.40(d,J=17.2Hz,1H), 4.2 7(d,J=17.2Hz,1H), 3.94(t,J=6.3Hz,2H), 3.16-3.12(m,4H), 2.95-2.86(m ,3H), 2.59(d,J=16.9Hz,2H), 2.42(s,3H), 2.38(dd,J=13.4,4.7Hz,1H), 2. 35-2.28(m,6H), 2.28-2.20(m,3H), 2.05-1.95(m,3H), 1.92-1.85(m,3H), 1 .78-1.67(m,8H), 1.60-1.55(m,2H), 1.51-1.41(m,5H), 1.39-1.34(m,2H). LC-MS(ES+): 963.60m / z[M+H]+, 482.50m / z[M+2H]2+.

[0151] Example 11: Synthesis of (16).

change

[0152] tert-ブチル4-(2-(1-メチル-2,6-ジオキソピペリジン-3-イル)-1-オキソイソインドリン-5-イル)ピペリジン-1-カルボン acid (14).

change

[0153] Potassium carbonate was added to a solution of tert-butyl 4-(2-(2,6-dioxopiperidin-3-yl)-1-oxoisoindolin-5-yl)piperidine-1-carboxylate 6 (350 mg, 0.819 mmol) in DMF (12 mL, 0.07 M) and stirred at room temperature for 10 min. Methyl iodide (0.080 mL, 1.23 mmol) was added and stirred at room temperature for 6 h. The reaction mixture was diluted with EtOAc and washed with water 2× and brine. The organic layer was dried over Na2SO4, filtered and concentrated in vacuo. Purification by purification (SiO2: 0-100% EtOAc / Hexanes) afforded the title compound (340 mg, 94%). LC-MS (ES+): 386.14 m / z [M+H-tBu]+, 342.15 m / z [M+H-Boc]+.

[0154] 1-Methyl-3-(1-oxo-5-(piperidin-4-yl)isoindolin-2-yl)piperidine-2,6-dione hydrochloride (15). [ka]

[0155] A solution of 4M HCl in dioxane (4 mL) was added to a solution of tert-butyl 4-(2-(1-methyl-2,6-dioxopiperidin-3-yl)-1-oxoisoindolin-5-yl)piperidine-1-carboxylate 14 (340 mg, 0.77 mmol) in CH2Cl2 (4 mL) and stirred at room temperature for 12 h. The reaction mixture was concentrated in vacuo to give the title compound as a solid (278 mg, 91%) which was carried forward without purification. 1H NMR(500MHz,DMSO-d6)δ9.14-9.01(m,2H), 7.70(d,J=7.5Hz,1H), 7.46(s,1H), 7.37(s,1H), 5.17(dd,J=12.8,4.2Hz,1H), 4.44(d,J=17.1 Hz,1H), 4.31(d,J=17.0Hz,1H), 3.36(d,J=11.2Hz,2H), 3.03-2.94(m,7H), 2.76(d,J=16.7Hz,1H), 2.43-2.34(m,1H), 2.02-1.90(m,5H). LC-MS(ES+):342.1m / z[M+H]+.

[0156] 3-(5-(1-(4-(4-(4-(6-((6-acetyl-8-cyclopentyl-5-methyl-7-oxo-7,8-dihydropyrido[2,3-d]pyrimidin-2-yl)amino)pyridin-3-yl)piperazin-1-yl)butoxy)benzyl)piperidin-4-yl)-1-oxoisoindolin-2-yl)-1-methylpiperidine-2,6-dione (16). [ka]

[0157] Reductive amination of 1-methyl-3-(1-oxo-5-(piperidin-4-yl)isoindolin-2-yl)piperidine-2,6-dione hydrochloride 15 and 4-(4-(4-(6-((6-acetyl-8-cyclopentyl-5-methyl-7-oxo-7,8-dihydropyrido[2,3-d]pyrimidin-2-yl)amino)pyridin-3-yl)piperazin-1-yl)butoxy)benzaldehyde 12b was carried out as described in Example 3 to give the title compounds. 1H NMR(500MHz,DMSO-d6)δ10.97(s,1H), 10.08(s,1H), 8.94(s,1H), 8.05(d,J=3.0Hz,1H), 7.84(d,J=9.0Hz,1H), 7.63(d,J=7.9Hz,1H), 7.49-7 .45(m,2H), 7.39(d,J=8.0Hz,1H), 7.23(t,J=8.4Hz,2H), 6.92(d,J=8.5Hz,2H), 5.81(p,J=8.5Hz,1H), 5.09(dd,J=13.3,5.1Hz,1H), 4.41(d,J =17.2Hz,1H), 4.28(d,J=17.4Hz,1H), 4.12(t,J=5.7Hz,2H), 3.19-3.15(m,4H), 2.97-2.86(m,5H), 2.77(t,J=5.6Hz,2H), 2.69-2.56(m,7H), 2.42-2.36(m,4H), 2.30(s,3H), 2.26-2.21(m,2H), 2.18(s,2H), 2.09- 2.03(m,2H), 2.01-1.95(m,2H), 1.89-1.84(m,2H), 1.81-1.53(m,11H). LC-MS(ES+):949.6m / z[M+H]+,475.5m / z[M+2H]2+.

[0158] Example 12: Synthesis of (17). [ka] 3-(5-(1-(4-(2-(4-(6-((6-acetyl-8-cyclopentyl-5-methyl-7-oxo-7,8-dihydropyrido[2,3-d]pyrimidin-2-yl)amino)pyridin-3-yl)piperazin-1-yl)ethoxy)benzyl)piperidin-4-yl)-1-oxoisoindolin-2-yl)-1-methylpiperidine-2,6-dione (17).

[0159] Reductive amination of 1-methyl-3-(1-oxo-5-(piperidin-4-yl)isoindolin-2-yl)piperidine-2,6-dione hydrochloride 15 and 12a was carried out as described in Example 3 to give the title compounds. 1H NMR(500MHz,DMSO-d6)δ10.08(s,1H), 8.95(s,1H), 8.05(d,J=3.0Hz,1H), 7.84(d,J =9.0Hz,1H), 7.64(d,J=7.8Hz,1H), 7.50-7.45(m,2H), 7.39(d,J=7.9Hz,1H), 7.24(d ,J=8.1Hz,2H), 6.92(d,J=8.1Hz,2H), 5.81(p,J=8.8Hz,1H), 5.16(dd,J=13.4,5.1Hz ,1H), 4.41(d,J=17.2Hz,1H), 4.27(d,J=17.1Hz,1H), 4.12(t,J=5.7Hz,2H), 3.45(s, 2H), 3.17(t,J=5.1Hz,4H), 3.00(s,3H), 2.95(d,J=5.5Hz,2H), 2.77(d,J=5.2Hz,3H ), 2.73(d,J=3.8Hz,1H), 2.66(t,J=5.0Hz,4H), 2.42(s,3H), 2.40-2.35(m,1H), 2.30 (s,3H), 2.24(t,J=9.7Hz,2H), 2.02-1.97(m,2H), 1.88(q,J=8.5,7.4Hz,2H), 1.76(d ,J=9.3Hz,5H), 1.70(d,J=11.8Hz,1H), 1.57(q,J=6.0Hz,2H), 1.23(d,J=2.8Hz,1H). LC-MS(ES+):920.51m / z[M+H]+.

[0160] Example 13: Co-degradation of CDK4, CDK6, and Helios by Ia-1 to Ia-8

[0161] As shown in Figure 1A, immunoblots from Jurkat cells treated for 4 hours with the title compounds (DKY709, and Ia-1 through Ia-8) revealed co-degradation of CDK4, CDK6, and Helios while sparing Ikaros, demonstrating the feasibility of redirecting neo-substrate specificity of the E3 ligase-binding components of bifunctional degradative molecules. The structure of DKY709 (3-(5-(1-benzylpiperidin-4-yl)-1-oxoisoindolin-2-yl)piperidine-2,6-dione) is shown here: [ka]

[0162] Example 14: Co-degradation of CDK4, CDK6, and Helios by Ia-8

[0163] As shown in Figure 1B, immunoblots from Jurkat cells treated with 1 μM of the title compounds (DKY709, triple degrader Ia-8, and paired negative control compound 17) for 4 hours are shown. The parent Helios degrader DKY709 only induced the degradation of Helios and did not affect the abundance of CDK4 or CDK6, whereas the triple degrader Ia-8 induced the degradation of CDK4, CDK6 and Helios, but not Ikaros, in Jurkat cells. The negative control compound 17 had no degrading activity.

[0164] Example 15: Protein abundance of CDK4, CDK6, Helios, and phosphorylated Rb

[0165] As shown in Figure 2 (immunoblots from Jurkat cells treated with 1 μM of the indicated compounds for 16 h), palbociclib (a CDK4 / 6 inhibitor) reduced the levels of phosphorylated Rb without affecting the protein abundance of Helios, CDK4, or CDK6, whereas DKY709 induced Helios degradation without affecting the levels of phosphorylated Rb. Ia-8 induced triple degradation of Helios, CDK4, and CDK6 and reduced the levels of phosphorylated Rb, whereas the negative control compound 17 had little effect.

[0166] Example 16: Induction of G1 arrest

[0167] As shown in FIG. 3A (propidium iodide staining for DNA content of Jurkat cells treated with 100 nM of the indicated compound for 24 hours), palbociclib induced G1 arrest, but DKY709 did not, whereas 18 and Ia-8 induced G1 arrest, whereas the negative control compound 17 did not. The structure of 18 (N-(4-(4-(6-((6-acetyl-8-cyclopentyl-5-methyl-7-oxo-7,8-dihydropyrido[2,3-d]pyrimidin-2-yl)amino)pyridin-3-yl)piperazin-1-yl)butyl)-2-((2-(1-methyl-2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindolin-4-yl)amino)acetamide) is shown here: [ka]

[0168] Example 17: Measurement of antiproliferative activity

[0169] As shown in Figure 3B, Jurkat cells were treated with a dose curve of the indicated compounds for 3 days and cell proliferation was quantified by CellTiter-Glo, revealing that the antiproliferative activity of the negative control compound 17 (244 nM) was less potent than that of Ia-8 (166 nM).

[0170] Example 18: CDK4-CDK6-Helios co-degradation enhances IL-2 secretion

[0171] Quantification of IL-2 levels by ELISA from Jurkat cells pretreated with 1 μM of the indicated compounds for 24 h and then TCR stimulated for 18 h. Results are shown in FIG. 4 as mean ± SD (untreated, n=3; other conditions, n=4) (*p<0.05). Both lenalidomide and DKY709 treatments led to an increase in IL-2 secretion, but did not reach statistical significance. Treatment with the CDK4 / CDK6 selective degrader 19 has similar activity to palbociclib, while the CDK4 / CDK6 / Ikaros / Aiolos degrader 18 induced even more IL-2 secretion. Notably, treatment with the CDK4 / CDK6 / Helios triple degrader, Ia-8, induced an increase in levels of IL2 comparable to that of 18, whereas the inactive chemical control compound, 17, had activity similar to that of palbociclib. The structure of 19 (N-(4-(4-(6-((6-acetyl-8-cyclopentyl-5-methyl-7-oxo-7,8-dihydropyrido[2,3-d]pyrimidin-2-yl)amino)pyridin-3-yl)piperazin-1-yl)butyl)-2-((2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindolin-4-yl)oxy)acetamide) is shown here: [ka]

[0172] All patent publications and non-patent publications are indicative of the level of skill of those skilled in the art to which this disclosure pertains. All these publications, including any specific portions thereof referenced, are herein incorporated by reference to the same extent as if each individual publication was specifically and individually indicated to be incorporated by reference.

[0173] Although the disclosure herein has been described with reference to particular embodiments, it is to be understood that these embodiments are merely illustrative of the principles and applications of the disclosure. It is thus to be understood that numerous modifications can be made to the illustrative embodiments and other configurations can be devised without departing from the spirit and scope of the disclosure as defined by the appended claims.

Claims

1. A compound having the structure represented by formula (I): 【Chemical 1】 or a pharmaceutically acceptable salt or stereoisomer thereof, wherein the targeting ligand is: [Chemical Formula 2] and the linker contains an alkylene chain or a polyethylene glycol chain, the compound, or a pharmaceutically acceptable salt or stereoisomer thereof.

2. Formula (Ia): [Chemical Formula 3] The compound according to claim 1, having the structure represented by, or a pharmaceutically acceptable salt or stereoisomer thereof.

3. Formula (Ib): 【Chemical 4】 The compound according to claim 1, having the structure represented by, or a pharmaceutically acceptable salt or stereoisomer thereof.

4. Formula (Ic): 【Chemical Formula 5】 The compound according to claim 1, having the structure represented by, or a pharmaceutically acceptable salt or stereoisomer thereof.

5. The linker contains an alkylene chain, optionally, the alkylene chain is interrupted by at least one of -O- or -NH-, and / or one or both ends thereof are terminated by at least one of -O- or -NH-, or the linker contains 1 to 6 alkylene units, The compound according to claim 1.

6. The linker contains a polyethylene glycol chain, optionally, the linker is terminated at one or both ends by at least one of -O- or -NH-, or the linker contains 2 to 3 ethylene glycol units, The compound according to claim 1.

7. The linker is a) any one of the structures: [Chemical Formula 6] or b) attached to the phenyl group at the 3-position (meta), or c) attached to the phenyl group at the 4-position (para), The compound according to claim 1.

8. The formula: 【Chemical Formula 7】 【Chemical Formula 8】 The compound according to claim 1, represented by, or a pharmaceutically acceptable salt or stereoisomer thereof.

9. A pharmaceutical composition comprising the compound according to any one of claims 1 to 8 or a pharmaceutically acceptable salt or stereoisomer thereof and a pharmaceutically acceptable carrier, optionally, the pharmaceutical composition is in solid or liquid form, further optionally, the solid pharmaceutical composition is in the form of tablets or capsules, A pharmaceutical composition.

10. A pharmaceutical composition comprising the compound according to any one of claims 1 to 8, or a pharmaceutically acceptable salt or stereoisomer thereof, i) A pharmaceutical composition for use in treating a disease or disorder characterized by abnormal activity of CDK4 and / or CDK6, and Helios, optionally, wherein said disease or disorder is cancer, optionally, wherein said cancer is, a) characterized by solid tumors, b) selected from breast cancer, brain tumor, endometrial cancer, head and neck cancer, gastrointestinal cancer, lung cancer, ovarian cancer, prostate cancer, uterine cancer, hepatocellular carcinoma, liposarcoma, and melanoma, or c) a hematological cancer, optionally, wherein said hematological cancer is selected from leukemia, lymphoma, and myeloma, or, ii) A pharmaceutical composition for use in reducing the levels of CDK4 and / or CDK6, and Helios in cells, either in vitro or in vivo, which is a pharmaceutical composition.

Images