STATEMENT OF U.S. GOVERNMENT SUPPORT

[0001] This invention was made with government support under grant numbers R01 CA197999, R21 CA251151 , GM121316, and CA036727 awarded by the National Institutes of Health. The government has certain rights in the invention.

BACKGROUND

[0002] Cyclin-dependent kinases (CDKs) are members of the CDK family of serine-threonine kinases that are involved in a variety of cellular functions such as the regulation of the cell cycle and transcription. CDKs are activated through association with cyclins or activators that mimic cyclins. A CDK of particular interest in cancer therapy is cyclin-dependent kinase 9 (CDK9), which interacts with Cyclin K and the CDK9/Cyclin K complex to regulate the replication stress response. CDK9 also interacts with cyclin T to form the catalytic subunit of the positive transcription elongation factor b (P-TEFb) that facilitates productive transcriptional elongation. Among others, CDK9/Cyclin T complex regulates transcription of anti-apoptotic protein Mcl-1 and the oncogene Myc. More recently, CDK9 was implicated as a therapeutic target in KRAS- mutant driven pancreatic cancer using the MiaPaCa2 cell line.

[0003] It is possible to develop inhibitors that are selective for CDKs such as CDK9; however these inhibitors are seldom selective for a specific CDK/cyclin combination. Accordingly, there is a need for compounds which can inhibit specific CDK/cyclin combinations to treat diseases and disorders, e.g., cancer.

SUMMARY

[0004] The disclosure provides compounds of Formula 0 and pharmaceutically acceptable salts thereof:

A-L-B (0), wherein

A is a CDK inhibitor/binder moiety;

B is an E3 ligase binding moiety; and

L is selected from

[0005] Also provided are compounds of Formula I and pharmaceutically acceptable salts thereof: wherein

R ¹ is Ci-8 alkyl, C2-8 alkenyl, C2-8 alkynyl, C3-11 cycloalkyl, C4-11 cycloalkenyl, 3-11 membered heterocycloalkyl having 1-3 ring heteroatoms selected from O, S, and N, wherein R ¹ is optionally substituted with from one to six R ⁵ ;

R ² is H, F, -CH ₃ , -CN, or -C(=O)OR ⁷ ;

R ³ is -C(=O)NR ⁹ -, -C(=O)O-, -C(=O)(CR ¹⁰ R ¹¹ ) _n -, or -(CR ¹³ R ¹¹ ) _n -;

R ⁴ is C1.8 alkylene, C2-8 alkenylene, C2-8 alkynylene, C3-11 cycloalkyl, C4-11 cycloalkenyl, 3-11 membered heterocycloalkyl having 1-3 ring heteroatoms selected from O, S, and N, Ce-14 aryl, or 5-14 membered heteroaryl having 1-3 ring heteroatoms selected from O, S, and N, wherein R ⁴ is optionally substituted with from one to three R ⁶ ; each of R ⁵ and R ⁶ is independently selected from halo, NO2, CN, -CF3, -NR ⁷ R ⁸ , - NR ⁷ C(=O)R ⁸ , -NR ⁷ C(=O)OR ⁸ , -NR ⁷ C(=O)NR ⁸ R ⁹ , -NR ⁷ S(=O) ₂ R ⁸ , -NR ⁷ S(=O) ₂ NR ⁸ R ⁹ , - OC(=O)R ⁷ , -OC(=O)OR ⁷ , -C(=O)OR ⁷ , -C(=O)R ⁷ , -C(=O)NR ⁷ R ⁸ , -OC(=O)NR ⁷ R ⁵ , -OC(=O)SR ⁷ , - S(=O)R ⁷ , -S(=O) ₂ R ⁷ , -S(=O) ₂ NR ⁷ R ⁸ , and R ⁷ ; each of R ⁷ , R ⁸ , and R ⁹ is independently selected from H, C1.8 alkyl, C2-8 alkenyl, C2-8 alkynyl, C3-11 cycloalkyl, C4-11 cycloalkenyl, 3-11 membered heterocycloalkyl having 1-3 ring heteroatoms selected from O, S, and N, Ce-14 aryl, or 5-14 membered heteroaryl having 1-3 ring heteroatoms selected from O, S, and N, wherein R ⁷ , R ⁸ , and R ⁹ are each independently optionally substituted with from one to six substituents independently selected from halo, NO2, CN, -CF ₃ , -NR ¹⁰ R ¹¹ , -NR ¹⁰ C(=O)R ¹¹ , -NR ¹⁰ C(=O)OR ¹¹ , -NR ¹⁰ C(=O)NR ¹¹ R ¹² , -NR ¹⁰ S(=O) ₂ R ¹¹ , - NR ¹⁰ S(=O) ₂ NR ¹¹ R ¹² , -OC(=O)R ¹⁰ , -OC(=O)OR ¹⁰ , -C(=O)OR ¹⁰ , -C(=O)R ¹⁰ , -C(=O)NR ¹⁰ R ¹¹ , - OC(=O)NR ¹⁰ R ¹¹ , -OC(=O)SR ¹⁰ , -S(=O)R ¹⁰ , -S(=O) ₂ R ¹⁰ , -S(=O) ₂ NR ¹⁰ R ¹¹ , and R ¹⁰ ; or, when R ⁷ and R ⁸ are as in NR ⁷ R ⁸ , they may instead optionally be connected to form with the nitrogen to which they are attached a 3-7 membered heterocycloalkyl having 1-2 additional ring heteroatoms selected from O, S, and N; each R ¹⁰ , R ¹¹ , and R ¹² is independently selected from H, C1.8 alkyl, C ₂ .s alkenyl, C ₂ .s alkynyl, C3-11 cycloalkyl, C4-11 cycloalkenyl, 3-11 membered heterocycloalkyl having 1-3 ring heteroatoms selected from O, S, and N, Ce-14 aryl, or 5-14 membered heteroaryl having 1-3 ring heteroatoms selected from O, S, and N, wherein R ¹⁰ , R ¹¹ , and R ¹² are each independently optionally substituted with from one to six substituents independently selected from halo, NO ₂ , CN, -CF ₃ , -NR ¹³ R ¹⁴ , -NR ¹³ C(=O)R ¹⁴ , -NR ¹³ C(=O)OR ¹⁴ , -NR ¹³ C(=O)NR ¹⁴ R ¹⁵ , -NR ¹³ S(=O) ₂ R ¹⁴ , - NR ¹³ S(=O) ₂ NR ¹⁴ R ¹⁵ , -OC(=O)R ¹³ , -OC(=O)OR ¹³ , -C(=O)OR ¹³ , -C(=O)R ¹³ , -C(=O)NR ¹³ R ¹⁴ , - OC(=O)NR ¹³ R ¹⁴ , -OC(=O)SR ¹³ , -S(=O)R ¹³ , -S(=O) ₂ R ¹³ , -S(=O) ₂ NR ¹³ R ¹⁴ , and R ¹³ ; each R ¹³ , R ¹⁴ , and R ¹⁵ is independently selected from H, C1.8 alkyl, C ₂ .s alkenyl, C ₂ .s alkynyl, C ₃ -n cycloalkyl, C4-11 cycloalkenyl, 3-11 membered heterocycloalkyl having 1-3 ring heteroatoms selected from O, S, and N, Ce-14 aryl, or 5-14 membered heteroaryl having 1-3 ring heteroatoms selected from O, S, and N, wherein R ¹³ , R ¹⁴ , and R ¹⁵ are each independently optionally substituted with from one to six substituents independently selected from halo, NO ₂ , CN, -CF ₃ , -NR ¹⁶ R ¹⁷ , -NR ¹⁶ C(=O)R ¹⁷ , -NR ¹⁶ C(=O)OR ¹⁷ , -NR ¹⁶ C(=O)NR ¹⁷ R ¹⁸ , -NR ¹⁶ S(=O) ₂ R ¹⁷ , - NR ¹⁶ S(=O) ₂ NR ¹⁷ R ¹⁸ , -OC(=O)R ¹⁶ , -OC(=O)OR ¹⁶ , -C(=O)OR ¹⁶ , -C(=O)R ¹⁶ , -C(=O)NR ¹⁶ R ¹⁷ , - OC(=O)NR ¹⁶ R ¹⁷ , -OC(=O)SR ¹⁶ , -S(=O)R ¹⁷ , -S(=O) ₂ R ¹⁶ , -S(=O) ₂ NR ¹⁶ R ¹⁷ , and R ¹⁶ ; each R ¹⁶ , R ¹⁷ , and R ¹⁸ is independently selected from H, Ci- ₃ alkyl, C ₂ .s alkenyl, C ₂ .s alkynyl, C ₃ -n cycloalkyl, C4-11 cycloalkenyl, 3-11 membered heterocycloalkyl having 1-3 ring heteroatoms selected from O, S, and N, Ce-14 aryl, and 5-14 membered heteroaryl having 1-3 ring heteroatoms selected from O, S, and N; n is 0, 1 , 2, or 3; wherein R ¹⁰ and R ¹¹ in -C(=O)(CR ¹⁰ R ¹¹ ) _n - and -(CR ¹⁰ R ¹¹ ) _n - are for each iteration of n defined independently as recited above;

L is Ci- alkylene interrupted with one or more of (i) non-adjacent heteroatom(s) selected from O, S, and NR ^N , (ii) C(O)NR ^N , and (iii) NR ^N C(O); each R ^N is independently H or CI-B alkyl; and

Het is 3-7 membered heterocycloalkyl having 1-3 ring heteroatoms selected from O, S, and N, and said heterocycloalkyl is optionally substituted with one or more oxo. [0006] In some embodiments, is -C(=O)(CR ¹⁰ R ¹¹ ) _n -. In some embodiments, R ³ is - C(=O)CH2-. In some embodiments, R ⁴ is Ce-14 aryl. In some embodiments, R ⁴ is phenyl.

[0007] Further provided are compounds of Formula IA and pharmaceutically acceptable salts thereof: wherein the variables are defined as above.

[0010] Further provided are methods degrading a cyclin-dependent kinase (CDK) comprising administering to a subject in need thereof a therapeutically effective amount of a composition comprising a CDK inhibitor moiety linked to an E3 ligase binding moiety, e.g., administering to a subject in need thereof a therapeutically effective amount of a composition comprising a compound disclosed herein or a pharmaceutically acceptable salt thereof.

[0011] Further provided herein are methods of treating or preventing a disease or disorder capable of being modulated by CDK degradation, comprising administering to a subject in need thereof a therapeutically effective amount of a composition comprising a CDK inhibitor moiety linked to an E3 ligase binding moiety, e.g., administering to a subject in need thereof a therapeutically effective amount of a composition comprising a compound disclosed herein or a pharmaceutically acceptable salt thereof.

[0012] Other aspects of the disclosure include compositions comprising a CDK inhibitor moiety linked to an E3 ligase binding moiety, e.g., a compound disclosed herein or a pharmaceutically acceptable salt thereof, for use in degrading a cyclin-dependent kinase (CDK), and compositions comprising a CDK inhibitor moiety linked to an E3 ligase binding moiety, e.g., a compound disclosed herein or a pharmaceutically acceptable salt thereof, for use in treating or preventing a disease or disorder capable of being modulated by CDK degradation.

BRIEF DESCRIPTION OF THE DRAWINGS

[0013] Figure 1 shows the structures and physical data for PROTACs disclosed herein (compounds 1-10).

[0014] Figure 2 shows the results of a Western blot analysis of HEK293 cells subjected to increasing concentrations (0.01 - 10 pM) of compounds (top row, left to right) 1 , 3, 5, 7, 9, and (bottom row, left to right) 2, 4, ,6, 8, and 10.

[0015] Figures 3A, 3B, 3C, and 3D show the selectivity profile of compound 2 (also called PROTAC 2). Figures 3A and 3B show a Western blot analysis showing dose-dependent (Figure 3A) and time-dependent (Figure 3B) effects of PROTAC 2 in HEK293 cells. Figures 3C and 3D show in vitro cell free IC50 (Figure 3C) and KD (Figure 3D) profiling of inhibitor 11 and compound 2 (i.e. , PROTAC 2), respectively.

[0016] Figure 4A shows a Western blot analysis showing inhibition of CDK9 degradation upon simultaneous treatment of Pomalidomide and compound 2 (PROTAC 2).

[0017] Figure 4B shows a Western blot analysis showing inhibition of CDK9 degradation upon simultaneous treatment of Flavopiridol and compound 2 (PROTAC 2).

[0018] Figure 4C shows a Western blot analysis showing inhibition of CDK9 degradation upon simultaneous treatment of MG132 and compound 2 (PROTAC 2).

[0019] Figure 5A shows a volcano plot of the kinome in HEK293 cells treated with 1 .M of compound 2 (PROTAC 2) and incubated for 24 hours. [0020] Figure 5B shows a volcano plot of the proteome in MiaPaCa2 cells treated with 1 iM of compound 2 (PROTAC 2) and incubated for 24 hours.

[0021] Figure 6A shows the structures of Bcl2 inhibitors (ABT-263) Navitoclax; Venetoclax (ABT-199); WEHI-539.

[0022] Figure 6B shows the growth inhibitory effects of different inhibitor combinations at 5 pM.

[0023] Figure 6C shows the combination index (Cl) values for the three Bcl2 inhibitor and compound 2 (PROTAC 2) combinations.

[0024] Figure 7 shows a Western blot demonstrating that compound 2 (PROTAC 2) does not induce degradation of Cyclin K.

[0025] Figure 8 shows Western blots demonstrating that compound 2 (PROTAC 2) does not induce degradation of Cyclin K over time.

DETAILED DESCRIPTION

[0026] Analyses of data from large-scale kinome screens suggests that it is possible to develop inhibitors that are selective for CDKs; however these inhibitors are seldom selective for a specific CDK/cyclin combination. This lack of selectivity for various CDKs has been addressed by proteolysis targeting chimeras (PROTACs). Provided herein are compounds which are aminopyrazole-based PROTACs that selectively degrade CDKs. These compounds are useful in the treatment of a variety of diseases and disorders, including but not limited to cancer, inflammatory diseases, and neurological diseases.

[0027] For example, compounds of the disclosure can have selectivity for cyclin-dependent kinase 9 (CDK9), which is involved in transcriptional regulation of several genes, including the oncogene Myc, and is a validated target for pancreatic cancer.

Compounds of the Disclosure

[0028] PROTACs are heterobifunctional molecules, wherein ligands that bind to two different proteins are conjugated via a linker. Without wishing to be bound by theory, it is believed that one end of the PROTAC binds to the protein of interest (POI) while the other binds to an E3- ligase to facilitate the formation of a ternary complex (POI:PROTAC:E3-ligase). Without wishing to be bound by theory, it is believed that the formation of a stable ternary complex allows the E3-ligase to ubiquitinate a proximal lysine on the POI to enable proteasomal degradation of POI. Since the distribution of surface exposed lysine residues are different among the various CDKs, a PROTAC generated using a non-selective CDK inhibitor with an appropriate linker could lead to a selective PROTAC. Non-limiting examples of non-selective CDK inhibitors include aminopyrazole, aminothiazole, and 4/7-chromen-4-one moieties, which have been used to generate CDK9 selective PROTACs. Recently, a CDK9 inhibitor (BAY-1143572) has been used in the development of a CDK9 PROTAC, which is similar to a CDK4/6 inhibitor Palbociclib-based CDK6 selective PROTAC.

[0029] Without wishing to be bound by theory, it is believed that linker length and linker composition contribute the selectivity and potency of the compounds disclosed herein. The aminopyrazole-based PROTACs provided herein have an improved iM (DC50) as compared to CDK9 degrading PROTACs known in the art.

[0030] The PROTAC compounds disclosed herein comprise a CDK inhibitor/binder moiety and proteolysis targeting group. The CDK inhibitor/binder moiety may be selected without limitation from a small molecule moiety, a peptide moiety, and an antibody moiety, or a fragment thereof. In some cases, the proteolysis targeting moiety is an E3 ligase binding moiety. The E3 ligase binding moiety can be selected from a small molecule moiety, a peptide moiety, and an antibody moiety or fragment thereof. In some cases, the E3 ligase binding moiety includes, but is not limited to, an iKBa-derived motif (such as a phosphopeptide motif), a HIF-1a derived motif (such as a HIF-1a pentapeptide or octopeptide motif), nutlin, bestatin, m ethyl- bestatin, thalidomide, pomalidomide, lenalidomide, thalidomide analogs, VHL binding molecules (including those described in Galdeano et al, J. Med Chem 2014, 57 (20): 8657- 8663) which is herein incorporated by reference). The E3 ligase binding moieties are capable of binding to and/or recruiting an E3 ligase including but not limited to VHL, cereblon, MDM2, clAP1 , and APC/C ^CDH ' ¹ . In some cases, the E3 ligase binding molecule binds to and/or recruits the E3 ligase cereblon.

[0031] The disclosure provides compounds of Formula 0 and pharmaceutically acceptable salts thereof:

A-L-B (0), wherein A is a CDK inhibitor/binder moiety; B is an E3 ligase binding moiety; and L is selected from

, L s, L is ,

[0032] In some cases, the CDK inhibitor/binder moiety is selected from a small molecule moiety, a peptide moiety, and an antibody moiety, or a fragment thereof. In some cases, the CDK inhibitor/binder moiety is a small molecule moiety. In some cases, the CDK inhibitor/binder moiety is a peptide moiety. In some cases, the CDK inhibitor/binder moiety is an antibody moiety, or a fragment thereof. In some cases, the CDK inhibitor/binder moiety is an antibody moiety. In some cases, the CDK inhibitor/binder moiety is a fragment of an antibody moiety.

[0033] In some cases, the CDK inhibitor/binder moiety is a small molecule moiety adapted to be coupled to L through an amide moiety. In some cases, the CDK inhibitor/binder moiety is derived from a compound comprising a carboxylic acid, which is adapted to be coupled through L through an amide moiety. In some cases, the CDK inhibitor/binder moiety is derived from a compound comprising an amine, which is adapted to be coupled through L through an amide moiety. In some cases, the small molecule moiety is derived from flavopiridol, R-roscovitine, AT7519, dinaciclib, R547, palbociclib, abemaciclib, ribociclib, milciclib, or PHA-793887. In some cases, the small molecule moiety is derived from flavopiridol. In some cases, the small molecule moiety is derived from R-roscovitine. In some cases, the small molecule moiety is derived from AT7519. In some cases, the small molecule moiety is derived from dinaciclib. In some cases, the small molecule moiety is derived from R547. In some cases, the small molecule moiety is derived from palbociclib. In some cases, the small molecule moiety is derived from abemaciclib. In some cases, the small molecule moiety is derived from ribociclib. In some cases, the small molecule moiety is derived from milciclib. In some cases, the small molecule moiety is derived from PHA-793887. [0034] In some cases, the CDK inhibitor/binder moiety is selected from bond to L. In some cases, the CDK inhibitor/binder moiety is In some cases, the CDK inhibitor/binder moiety some cases, the CDK inhibitor/binder moiety is In some cases, the CDK inhibitor/binder moiety is

some cases, the CDK inhibitor/binder moiety is . In some cases, the CDK inhibitor/binder moiety is In some cases, the CDK inhibitor/binder moiety is In some cases, the CDK inhibitor/binder moiety is In some cases, the CDK [0035] In some cases, the E3 ligase binding moiety is selected from a small molecule moiety, a peptide moiety, and an antibody moiety or fragment thereof. In some cases, the E3 ligase binding moiety is a small molecule moiety. In some cases, the E3 ligase binding moiety is a peptide moiety. In some cases, the E3 ligase binding moiety is an antibody moiety or fragment thereof. In some cases, the E3 ligase binding moiety is an antibody moiety. In some cases, the E3 ligase binding moiety is a fragment of an antibody moiety. In some cases, the E3 ligase binding moiety is an iKBa-derived moiety, a HIF-1a derived moiety, nutlin, bestatin, methyl- bestatin, thalidomide, pomalidomide, lenalidomide, a thalidomide analog, or a VHL binding molecule. In some cases, the E3 ligase binding moiety is an IKBa-derived moiety. In some cases, the E3 ligase binding moiety is a HIF-1a derived moiety. In some cases, the E3 ligase binding moiety is nutlin. In some cases, the E3 ligase binding moiety is bestatin. In some cases, the E3 ligase binding moiety is thalidomide. In some cases, the E3 ligase binding moiety is pomalidomide. In some cases, the E3 ligase binding moiety is lenalidomide. In some cases, the E3 ligase binding moiety is a thalidomide analog. In some cases, the E3 ligase binding moiety is a VHL binding molecule.

[0036] In some cases, the E3 ligase binding moiety is capable of binding to and/or recruiting an E3 ligase. In some cases, the E3 ligase is VHL, cereblon, MDM2, clAP1 , or APC/C ^CDH ' ¹ . In some cases, the E3 ligase is VHL. In some cases, the E3 ligase is cereblon. In some cases, the E3 ligase is MDM2. In some cases, the E3 ligase is clAP1. In some cases, the E3 ligase is APC/C ^CDH - ¹ .

[0037] The disclosure provides compounds of Formula I and pharmaceutically acceptable salts thereof: wherein

R ¹ is Ci-8 alkyl, C2-8 alkenyl, C2-8 alkynyl, C3-11 cycloalkyl, C4-11 cycloalkenyl, 3-11 membered heterocycloalkyl having 1-3 ring heteroatoms selected from O, S, and N, wherein R ¹ is optionally substituted with from one to six R ⁵ ;

R ² is H, F, -CH ₃ , -CN, or -C(=O)OR ⁷ ;

R ³ is -C(=O)NR ⁹ -, -C(=O)O-, -C(=O)(CR ¹⁰ R ¹¹ ) _n -, or -(CR ¹³ R ¹¹ ) _n -;

R ⁴ is C1.8 alkylene, C2-8 alkenylene, C2-8 alkynylene, C3-11 cycloalkyl, C4-11 cycloalkenyl, 3-11 membered heterocycloalkyl having 1-3 ring heteroatoms selected from O, S, and N, Ce-14 aryl, or 5-14 membered heteroaryl having 1-3 ring heteroatoms selected from O, S, and N, wherein R ⁴ is optionally substituted with from one to three R ⁶ ; each of R ⁵ and R ⁶ is independently selected from halo, NO2, CN, -CF3, -NR ⁷ R ⁸ , - NR ⁷ C(=O)R ⁸ , -NR ⁷ C(=O)OR ⁸ , -NR ⁷ C(=O)NR ⁸ R ⁹ , -NR ⁷ S(=O) ₂ R ⁸ , -NR ⁷ S(=O) ₂ NR ⁸ R ⁹ , - OC(=O)R ⁷ , -OC(=O)OR ⁷ , -C(=O)OR ⁷ , -C(=O)R ⁷ , -C(=O)NR ⁷ R ⁸ , -OC(=O)NR ⁷ R ⁵ , -OC(=O)SR ⁷ , - S(=O)R ⁷ , -S(=O) ₂ R ⁷ , -S(=O) ₂ NR ⁷ R ⁸ , and R ⁷ ; each of R ⁷ , R ⁸ , and R ⁹ is independently selected from H, Ci-s alkyl, C ₂ .8 alkenyl, C ₂ .8 alkynyl, C3-11 cycloalkyl, C4-11 cycloalkenyl, 3-11 membered heterocycloalkyl having 1-3 ring heteroatoms selected from O, S, and N, Ce-14 aryl, or 5-14 membered heteroaryl having 1-3 ring heteroatoms selected from O, S, and N, wherein R ⁷ , R ⁸ , and R ⁹ are each independently optionally substituted with from one to six substituents independently selected from halo, NO ₂ , CN, -CF ₃ , -NR ¹⁰ R ¹¹ , -NR ¹⁰ C(=O)R ¹¹ , -NR ¹⁰ C(=O)OR ¹¹ , -NR ¹⁰ C(=O)NR ¹¹ R ¹² , -NR ¹⁰ S(=O) ₂ R ¹¹ , - NR ¹⁰ S(=O) ₂ NR ¹¹ R ¹² , -OC(=O)R ¹⁰ , -OC(=O)OR ¹⁰ , -C(=O)OR ¹⁰ , -C(=O)R ¹⁰ , -C(=O)NR ¹⁰ R ¹¹ , - OC(=O)NR ¹⁰ R ¹¹ , -OC(=O)SR ¹⁰ , -S(=O)R ¹⁰ , -S(=O) ₂ R ¹⁰ , -S(=O) ₂ NR ¹⁰ R ¹¹ , and R ¹⁰ ; or, when R ⁷ and R ⁸ are as in NR ⁷ R ⁸ , they may instead optionally be connected to form with the nitrogen to which they are attached a 3-7 membered heterocycloalkyl having 1-2 additional ring heteroatoms selected from O, S, and N; each R ¹⁰ , R ¹¹ , and R ¹² is independently selected from H, C1.8 alkyl, C ₂ .8 alkenyl, C ₂ .8 alkynyl, C3-11 cycloalkyl, C4-11 cycloalkenyl, 3-11 membered heterocycloalkyl having 1-3 ring heteroatoms selected from O, S, and N, Ce-14 aryl, or 5-14 membered heteroaryl having 1-3 ring heteroatoms selected from O, S, and N, wherein R ¹⁰ , R ¹¹ , and R ¹² are each independently optionally substituted with from one to six substituents independently selected from halo, NO ₂ , CN, -CF ₃ , -NR ¹³ R ¹⁴ , -NR ¹³ C(=O)R ¹⁴ , -NR ¹³ C(=O)OR ¹⁴ , -NR ¹³ C(=O)NR ¹⁴ R ¹⁵ , -NR ¹³ S(=O) ₂ R ¹⁴ , - NR ¹³ S(=O) ₂ NR ¹⁴ R ¹⁵ , -OC(=O)R ¹³ , -OC(=O)OR ¹³ , -C(=O)OR ¹³ , -C(=O)R ¹³ , -C(=O)NR ¹³ R ¹⁴ , - OC(=O)NR ¹³ R ¹⁴ , -OC(=O)SR ¹³ , -S(=O)R ¹³ , -S(=O) ₂ R ¹³ , -S(=O) ₂ NR ¹³ R ¹⁴ , and R ¹³ ; each R ¹³ , R ¹⁴ , and R ¹⁵ is independently selected from H, C1.8 alkyl, C ₂ .8 alkenyl, C ₂ .8 alkynyl, C ₃ -n cycloalkyl, C4-11 cycloalkenyl, 3-11 membered heterocycloalkyl having 1-3 ring heteroatoms selected from O, S, and N, Ce-14 aryl, or 5-14 membered heteroaryl having 1-3 ring heteroatoms selected from O, S, and N, wherein R ¹³ , R ¹⁴ , and R ¹⁵ are each independently optionally substituted with from one to six substituents independently selected from halo, NO ₂ , CN, -CF ₃ , -NR ¹⁶ R ¹⁷ , -NR ¹⁶ C(=O)R ¹⁷ , -NR ¹⁶ C(=O)OR ¹⁷ , -NR ¹⁶ C(=O)NR ¹⁷ R ¹⁸ , -NR ¹⁶ S(=O) ₂ R ¹⁷ , - NR ¹⁶ S(=O) ₂ NR ¹⁷ R ¹⁸ , -OC(=O)R ¹⁶ , -OC(=O)OR ¹⁶ , -C(=O)OR ¹⁶ , -C(=O)R ¹⁶ , -C(=O)NR ¹⁶ R ¹⁷ , - OC(=O)NR ¹⁶ R ¹⁷ , -OC(=O)SR ¹⁶ , -S(=O)R ¹⁷ , -S(=O) ₂ R ¹⁶ , -S(=O) ₂ NR ¹⁶ R ¹⁷ , and R ¹⁶ ; each R ¹⁶ , R ¹⁷ , and R ¹⁸ is independently selected from H, C1.8 alkyl, C ₂ .8 alkenyl, C ₂ .8 alkynyl, C ₃ -n cycloalkyl, C4-11 cycloalkenyl, 3-11 membered heterocycloalkyl having 1-3 ring heteroatoms selected from O, S, and N, Ce-14 aryl, and 5-14 membered heteroaryl having 1-3 ring heteroatoms selected from O, S, and N; n is 0, 1 , 2, or 3; wherein R ¹⁰ and R ¹¹ in -C(=O)(CR ¹⁰ R ¹¹ ) _n - and -(CR ¹⁰ R ¹¹ ) _n - are for each iteration of n defined independently as recited above;

L is C1-18 alkylene interrupted with one or more of (i) non-adjacent heteroatom(s) selected from O, S, and NR ^N , (ii) C(O)NR ^N , and (iii) NR ^N C(O); each R ^N is independently H or Ci-e alkyl; and

Het is 3-7 membered heterocycloalkyl having 1-3 ring heteroatoms selected from O, S, and N, and said heterocycloalkyl is optionally substituted with one or more oxo.

[0038] In some cases, R ³ is -C(=O)(CR ¹⁰ R ¹¹ ) _n -. In some cases, R ³ is -C(=0)CH2-.

[0039] In some cases, R ⁴ is Ce-14 aryl. In some cases, In some cases, R ⁴ is phenyl. In some cases, R ⁴ is unsubstituted. In some cases, R ⁴ is substituted with from one to three R ⁶ . In some cases, R ⁴ is substituted with one R ⁶ . In some cases, R ⁴ is substituted with two R ⁶ . In some cases, R ⁴ is substituted with three R ⁶ .

[0040] In some cases, the compound of Formula I has the structure of Formula IA:

[0041] In some cases, R ¹ is C3-8 cycloalkyl. In some cases, R ¹ is C3-5 cycloalkyl. In some cases, R ¹ is C4 cycloalkyl. In some cases, R ¹ is unsubstituted. In some cases, R ¹ is substituted with from one to six R ⁵ . In some cases, R ¹ is substituted with one R ⁵ . In some cases, R ¹ is substituted with two R ⁵ . In some cases, R ¹ is substituted with three R ⁵ . In some cases, R ¹ is substituted with four R ⁵ . In some cases, R ¹ is substituted with five R ⁵ . In some cases, R ¹ is substituted with six R ⁵ .

[0042] In some cases, R ² is H or C1.6 alkyl. In some cases, R ² is H. In some cases, R ² is Ci- 6 alkyl.

[0043] In some cases, Het is 3-7 membered heterocycloalkyl having 1-3 ring heteroatoms selected from O, S, and N. In some cases, Het is 3 membered heterocycloalkyl having 1-3 ring heteroatoms selected from O, S, and N. In some cases, Het is 4 membered heterocycloalkyl having 1-3 ring heteroatoms selected from O, S, and N. In some cases, Het is 5 membered heterocycloalkyl having 1-3 ring heteroatoms selected from O, S, and N. In some cases, Het is 6 membered heterocycloalkyl having 1-3 ring heteroatoms selected from O, S, and N. In some cases, Het is 7 membered heterocycloalkyl having 1-3 ring heteroatoms selected from O, S, and N.

[0044] In some cases, Het is unsubstituted. In some cases, Het is substituted with one or more oxo. In some cases, Het is substituted with one oxo. In some cases, Het is substituted with two oxo. In some cases, Het is 6 membered heterocyloalkyl substituted with two oxo. In some cases, Het i

[0045] The “L” comprises C1-18 alkylene interrupted with one or more of (i) non-adjacent heteroatom(s) selected from O, S, and NR ^N , (ii) C(O)NR ^N , and (iii) NR ^N C(O). In some cases, L is interrupted with 1-3 of any combination of groups (i)-(iii). In some cases, L is interrupted with 1 group of (i)-(iii). In some cases, L is uninterrupted Ci- alkylene. In some cases, L is Ci- alkylene interrupted with one or more of (i) non-adjacent heteroatom(s) selected from O and NR ^N , (ii) C(O)NR ^N , and (iii) NR ^N C(O).

[0046] In some cases, L comprises no O. In some cases, L comprises one O. In some cases, L comprises no NR ^N . In some cases, L comprises one NR ^N . In some cases, L comprises no NH. In some cases, L comprises one NH. In some cases, L comprises at least one C(O)NR ^N . In some cases, L comprises one C(O)NR ^N . In some cases, L comprises at least one C(O)NH. In some cases, L comprises one C(O)NH.

[0047] In some cases, L is Ci-w alkylene interrupted with one each of (i) O, (ii) NR ^N , and (iii) C(O)NR ^N or NR ^N C(O). In some cases, L is Ci- alkylene interrupted with one each of (i) O, (ii) NH, and (iii) C(O)NH or NHC(O).

[0048] In some cases, each R ^N is H. In some cases, each R ^N is Ci-e alkyl. ,

[0050] Specific compounds contemplated include those listed in Table 1, and pharmaceutically acceptable salts thereof:

Table 1

[0051] In some cases, the compound is compound 2: or a pharmaceutically acceptable salt thereof.

[0052] The compounds disclosed herein can be in the form of a pharmaceutically acceptable salt. As used herein, the term “pharmaceutically acceptable salt” refers to those salts which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of humans and lower animals without undue toxicity, irritation, allergic response and the like, and are commensurate with a reasonable benefit/risk ratio. Pharmaceutically acceptable salts are well known in the art. For example, S. M. Berge et al. describe pharmaceutically acceptable salts in detail in J. Pharmaceutical Sciences, 1977, 66, 1-19, which is incorporated herein by reference. Pharmaceutically acceptable salts of the compounds of this invention include those derived from suitable inorganic and organic acids and bases. Examples of pharmaceutically acceptable, nontoxic acid addition salts are salts of an amino group formed with inorganic acids such as hydrochloric acid, hydrobromic acid, phosphoric acid, sulfuric acid and perchloric acid or with organic acids such as acetic acid, trifluoroacetic acid, oxalic acid, maleic acid, tartaric acid, citric acid, succinic acid or malonic acid or by using other methods used in the art such as ion exchange. Other pharmaceutically acceptable salts include adipate, alginate, ascorbate, aspartate, benzenesulfonate, benzoate, bisulfate, borate, butyrate, camphorate, camphorsulfonate, citrate, cyclopentanepropionate, digluconate, dodecylsulfate, ethanesulfonate, formate, fumarate, glucoheptonate, glycerophosphate, gluconate, glutamate, hemisulfate, heptanoate, hexanoate, hydroiodide, 2-hydroxy-ethanesulfonate, lactobionate, lactate, laurate, lauryl sulfate, malate, maleate, malonate, methanesulfonate, 2- naphthalenesulfonate, nicotinate, nitrate, oleate, oxalate, palmitate, pamoate, pectinate, persulfate, 3-phenylpropionate, phosphate, picrate, pivalate, propionate, stearate, succinate, sulfate, tartrate, thiocyanate, p-toluenesulfonate, undecanoate, valerate salts, and the like. Salts of compounds containing a carboxylic acid or other acidic functional group can be prepared by reacting with a suitable base. Such salts include, but are not limited to, alkali metal, alkaline earth metal, aluminum salts, ammonium, N ⁺ (Ci-4alkyl)4 salts, and salts of organic bases such as trimethylamine, triethylamine, morpholine, pyridine, piperidine, picoline, dicyclohexylamine, N.NEdibenzylethylenediamine, 2-hydroxyethylamine, bis-(2- hydroxyethyl)amine, tri-(2-hydroxyethyl)amine, procaine, dibenzylpiperidine, dehydroabietylamine, N.NEbisdehydroabietylamine, glucamine, N-methylglucamine, collidine, quinine, quinoline, and basic amino acids such as lysine and arginine. This invention also envisions the quaternization of any basic nitrogen-containing groups of the compounds disclosed herein. Water or oil-soluble or dispersible products may be obtained by such quaternization. Representative alkali or alkaline earth metal salts include sodium, lithium, potassium, calcium, magnesium, and the like. Further pharmaceutically acceptable salts include, when appropriate, nontoxic ammonium, quaternary ammonium, and amine cations formed using counterions such as halide, hydroxide, carboxylate, sulfate, phosphate, nitrate, lower alkyl sulfonate and aryl sulfonate.

Definitions

[0053] As used herein, the term “alkyl” refers to straight chained and branched saturated hydrocarbon groups containing one to thirty carbon atoms, for example, one to twenty carbon atoms, or one to ten carbon atoms. The term C _n means the alkyl group has “n” carbon atoms. For example, C4 alkyl refers to an alkyl group that has 4 carbon atoms. Ci-Cs alkyl refers to an alkyl group having a number of carbon atoms encompassing the entire range (e.g., 1 to 8 carbon atoms), as well as all subgroups (e.g., 1-8, 2-8, 3-8, 4-8, 5-8, 6-8, 7-8, 1 , 2, 3, 4, 5, 6, 7, and 8 carbon atoms). Nonlimiting examples of alkyl groups include, methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl (2-methylpropyl), t-butyl (1 ,1 -dimethylethyl), 3,3-dimethylpentyl, and

2-ethylhexyl. Unless otherwise indicated, an alkyl group can be an unsubstituted alkyl group or a substituted alkyl group.

[0054] The term “alkylene” used herein refers to an alkyl group having a substituent. For example, the term “alkylenehalo” refers to an alkyl group substituted with a halo group. For example, an alkylene group can be -CH2CH2- or -CH2-. The term C _n means the alkylene group has “n” carbon atoms. For example, C1.18 alkylene refers to an alkylene group having a number of carbon atoms encompassing the entire range, as well as all subgroups, as previously described for “alkyl” groups. Unless otherwise indicated, an alkylene group can be an unsubstituted alkylene group or a substituted alkylene group.

[0055] The term “alkenyl” used herein refers to an unsaturated aliphatic group analogous in length and possible substitution to an alkyl group described above, but that contains at least one double bond. For example, the term “alkenyl” includes straight chain alkenyl groups (e.g., ethenyl, propenyl, butenyl, pentenyl, hexenyl, heptenyl, octenyl, nonenyl, decenyl), and branched alkenyl groups. For example, a straight chain or branched alkenyl group can have six or fewer carbon atoms in its backbone (e.g., C2-C6 for straight chain, C3-C6 for branched chain). The term “C2-C6” includes chains having a number of carbon atoms encompassing the entire range (e.g., 2 to 6 carbon atoms), as well as all subgroups (e.g., 2-6, 2-5, 2-4, 3-6, 2, 3, 4, 5, and 6 carbon atoms). The term “Cs-Ce” includes chains having a number of carbon atoms encompassing the entire range (e.g., 3 to 6 carbon atoms), as well as all subgroups (e.g., 3-6,

3-5, 3-4, 3, 4, 5, and 6 carbon atoms). Unless otherwise indicated, an alkenyl group can be an unsubstituted alkenyl group or a substituted alkenyl group.

[0056] The term “alkenylene” used herein refers to an alkenyl group having a substituent. For example, the term “alkenylenehalo” refers to an alkyl group substituted with a halo group. For example, an alkylene group can be -CH=CH-. The term C _n means the alkenylene group has “n” carbon atoms. For example, C2-6 alkenylene refers to an alkenylene group having a number of carbon atoms encompassing the entire range, as well as all subgroups, as previously described for “alkenyl” groups. Unless otherwise indicated, an alkenylene group can be an unsubstituted alkenylene group or a substituted alkenylene group.

[0057] As used herein, an alkylene which is “interrupted” is understood to be an alkylene group in which at one or more (e.g., 1-5, 1-4, 1-3, 1-2, 1 , 2, 3, 4, or 5) positions on the alkylene chain is inserted a group selected from one or more of (i) non-adjacent heteroatom(s) selected from O, S, and NR ^N , (ii) C(O)NR ^N , and (iii) NR ^N C(O). The interruptions can be consecutive for various combinations of these interrupting groups (e.g., a heteroatom next to a C(O)NR ^N moiety), except that two heteroatoms cannot be adjacent or consecutive to each other.

[0058] As used herein an alkylene which is interrupted with “one or more” groups is understood to be interrupted with from 1 to n-1 groups, wherein n is the number of carbon atoms in the alkylene chain. For example, a Ce-alkylene which is optionally interrupted with one or more groups can be interrupted with one, two, three, four, or five groups.

[0059] The term “alkynyl” used herein refers to an unsaturated aliphatic group analogous in length and possible substitution to an alkyl group described above, but that contains at least one triple bond. For example, the term “alkynyl” includes straight chain alkynyl groups (e.g., ethynyl, propynyl, butynyl, pentynyl, hexynyl, heptynyl, octynyl), and branched alkynyl groups. For example, a straight chain or branched alkynyl group can have eight or fewer carbon atoms in its backbone (e.g., C2-C8 for straight chain, C4-C8 for branched chain). The term “C2-C8” includes chains having a number of carbon atoms encompassing the entire range (e.g., 2 to 8 carbon atoms), as well as all subgroups (e.g., 2-6, 2-5, 2-4, 3-6, 2, 3, 4, 5, and 6 carbon atoms). The term “C4-C8” includes chains having a number of carbon atoms encompassing the entire range (e.g., 4 to 8 carbon atoms), as well as all subgroups (e.g., 4-6, 4-5, 4, 5, and 6 carbon atoms). Unless otherwise indicated, an alkynyl group can be an unsubstituted alkynyl group or a substituted alkynyl group.

[0060] The term “alkynylene” used herein refers to an alkynyl group having a substituent. For I — c=c — I example, an alkylene group can be » « . The term C _n means the alkynylene group has

“n” carbon atoms. For example, C2-8 alkynylene refers to an alkynylene group having a number of carbon atoms encompassing the entire range, as well as all subgroups, as previously described for “alkynyl” groups. Unless otherwise indicated, an alkynylene group can be an unsubstituted alkynylene group or a substituted alkynylene group.

[0061] As used herein, the term “cycloalkyl” refers to an aliphatic cyclic hydrocarbon group containing three to eleven carbon atoms (e.g., 3, 4, 5, 6, 7, 8, 9, 10, or 11 carbon atoms). The term C _n means the cycloalkyl group has “n” carbon atoms. For example, C5 cycloalkyl refers to a cycloalkyl group that has 5 carbon atoms in the ring. Ce-Cn cycloalkyl refers to cycloalkyl groups having a number of carbon atoms encompassing the entire range (e.g., 6 to 11 carbon atoms), as well as all subgroups (e.g., 6-7, 6-8, 7-8, 6-9, 6, 7, 8, 9, 10, and 11 carbon atoms). Nonlimiting examples of cycloalkyl groups include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and cyclooctyl. Unless otherwise indicated, a cycloalkyl group can be an unsubstituted cycloalkyl group or a substituted cycloalkyl group. The cycloalkyl groups described herein can be isolated or fused to another cycloalkyl group. When a cycloalkyl group is fused to another cycloalkyl group, then each of the cycloalkyl groups can contain three to twelve carbon atoms unless specified otherwise. Unless otherwise indicated, a cycloalkyl group can be unsubstituted or substituted.

[0062] The term “cycloalkenyl” is defined similarly as “cycloalkyl” except that the ring comprises at least one double bond, without being aromatic. The cycloalkenyl groups described herein can be isolated or fused to another cycloalkenyl group. Unless otherwise indicated, a cycloalkenyl group can be unsubstituted or substituted.

[0063] As used herein, the term “heterocycloalkyl” is defined similarly as cycloalkyl, except the ring contains one to three heteroatoms independently selected from oxygen, nitrogen, and sulfur. In particular, the term “heterocycloalkyl” refers to a ring containing a total of three to eleven atoms (e.g., three to seven, or five to eleven), of which 1 , 2, 3 or three of those atoms are heteroatoms independently selected from the group consisting of oxygen, nitrogen, and sulfur, and the remaining atoms in the ring are carbon atoms. Nonlimiting examples of heterocycloalkyl groups include piperdine, pyrazolidine, tetrahydrofuran, tetrahydropyran, dihydrofuran, morpholine, and the like. The heterocycloalkyl groups described herein can be isolated or fused to another heterocycloalkyl group. Heterocycloalkyl groups can be saturated or partially unsaturated ring systems. Unless otherwise indicated, a heterocycloalkyl group can be unsubstituted or substituted.

[0064] As used herein, the term “aryl” refers to a monocyclic aromatic hydrocarbon group, such as phenyl or a bicyclic aromatic hydrocarbon group, such as naphthyl. Unless otherwise indicated, an aryl group can be unsubstituted or substituted with one or more groups. Aryl groups can be isolated (e.g., phenyl) or fused to another aryl group (e.g., naphthyl, anthracenyl), a cycloalkyl group (e.g. tetraydronaphthyl), a heterocycloalkyl group, and/or a heteroaryl group. Exemplary aryl groups include, but are not limited to, phenyl, chlorophenyl, methylphenyl, methoxyphenyl, trifluoromethylphenyl, nitrophenyl, 2,4-methoxychlorophenyl, and the like. Throughout, the abbreviation “Ph” refers to phenyl and “Bn” refers to benzyl (i.e. , CH2phenyl).

[0065] As used herein, the term “heteroaryl” refers to a monocyclic or bicyclic aromatic ring having 5 to 14 total ring atoms, and containing one to three heteroatoms selected from nitrogen, oxygen, and sulfur atom in the aromatic ring. Unless otherwise indicated, a heteroaryl group can be unsubstituted or substituted. Examples of heteroaryl groups include, but are not limited to, thienyl, furyl, pyridyl, pyrrolyl, oxazolyl, triazinyl, triazolyl, isothiazolyl, isoxazolyl, imidazolyl, pyrazinyl, pyrimidinyl, thiazolyl, and thiadiazolyl.

[0066] As used herein, the term “oxo” refers to a =0 group.

[0067] As used herein, the term “halo” refers to a F (fluoro), Cl (chloro), Br (bromo), or I

(iodo) group. [0068] As used herein, the term “substituted,” when used to modify a chemical functional group, refers to the replacement of at least one hydrogen radical on the functional group with a substituent. Substituents can include, but are not limited to, alkyl, cycloalkyl, alkenyl, cycloalkenyl, alkynyl, heterocycloalkyl, aryl, heteroaryl, hydroxyl, oxy, alkoxy, heteroalkoxy, ester, thioester, carboxy, cyano, nitro, amino, amido, acetamide, and halo (e.g., fluoro, chloro, bromo, or iodo). When a chemical functional group includes more than one substituent, the substituents can be bound to the same carbon atom or to two or more different carbon atoms.

[0069] As used herein, the phrase “optionally substituted” means unsubstituted (e.g., substituted with a H) or substituted. As used herein, the term “substituted” means that a hydrogen atom is removed and replaced by a substituent. It is understood that substitution at a given atom is limited by valency. The use of a substituent (radical) prefix name such as alkyl without the modifier “optionally substituted” or “substituted” is understood to mean that the particular substituent is unsubstituted.

[0070] As used herein, the term “therapeutically effective amount” means an amount of a compound or combination of therapeutically active compounds (e.g., a CDK degrader or combination of CDK degrader) that ameliorates, attenuates or eliminates one or more symptoms of a particular disease or condition (e.g., cancer), or prevents or delays the onset of one of more symptoms of a particular disease or condition.

[0071] As used herein, the terms “patient” and “subject” may be used interchangeably and mean animals, such as dogs, cats, cows, horses, and sheep (e.g., non-human animals) and humans. Particular patients or subjects are mammals (e.g., humans). The terms patient and subject include males and females.

[0072] As used herein, the term “pharmaceutically acceptable” means that the referenced substance, such as a compound of the present disclosure, or a formulation containing the compound, or a particular excipient, are safe and suitable for administration to a patient or subject. The term “pharmaceutically acceptable excipient” refers to a medium that does not interfere with the effectiveness of the biological activity of the active ingredient(s) and is not toxic to the host to which it is administered.

[0073] As used herein the terms “treating”, “treat” or “treatment” and the like include preventative (e.g., prophylactic) and palliative treatment.

[0074] As used herein, the term “excipient” means any pharmaceutically acceptable additive, carrier, diluent, adjuvant, or other ingredient, other than the active pharmaceutical ingredient (API). Pharmaceutical formulations, dosing, and routes of administration

[0075] Further provided are pharmaceutical formulations comprising a compound as described herein (e.g., compounds of Formula 0, Formula I, Formula IA, the compounds of Table 1, or pharmaceutically acceptable salts of the compounds) and a pharmaceutically acceptable excipient.

[0076] The compounds described herein can be administered to a subject in a therapeutically effective amount (e.g., in an amount sufficient to prevent or relieve the symptoms of a disorder capable of being modulated by CDK degradation). The compounds can be administered alone or as part of a pharmaceutically acceptable composition or formulation. In addition, the compounds can be administered all at once, multiple times, or delivered substantially uniformly over a period of time. It is also noted that the dose of the compound can be varied over time.

[0077] A particular administration regimen for a particular subject will depend, in part, upon the compound, the amount of compound administered, the route of administration, and the cause and extent of any side effects. The amount of compound administered to a subject (e.g., a mammal, such as a human) in accordance with the disclosure should be sufficient to effect the desired response over a reasonable time frame. Dosage typically depends upon the route, timing, and frequency of administration. Accordingly, the clinician titers the dosage and modifies the route of administration to obtain the optimal therapeutic effect, and conventional rangefinding techniques are known to those of ordinary skill in the art.

[0078] Purely by way of illustration, the method comprises administering, e.g., from about 0.1 mg/kg up to about 100 mg/kg of compound or more, depending on the factors mentioned above. In other embodiments, the dosage ranges from 1 mg/kg up to about 100 mg/kg; or 5 mg/kg up to about 100 mg/kg; or 10 mg/kg up to about 100 mg/kg. Some conditions require prolonged treatment, which may or may not entail administering lower doses of compound over multiple administrations. If desired, a dose of the compound is administered as two, three, four, five, six or more sub-doses administered separately at appropriate intervals throughout the day, optionally, in unit dosage forms. The treatment period will depend on the particular condition and type of pain, and may last one day to several months.

[0079] Suitable methods of administering a physiologically-acceptable composition, such as a pharmaceutical composition comprising the compounds disclosed herein (e.g., compounds of compounds of Formula 0, Formula I, Formula IA, the compounds of Table 1, or pharmaceutically acceptable salts of the compounds), are well known in the art. Although more than one route can be used to administer a compound, a particular route can provide a more immediate and more effective reaction than another route. Depending on the circumstances, a pharmaceutical composition comprising the compound is applied or instilled into body cavities, absorbed through the skin or mucous membranes, ingested, inhaled, and/or introduced into circulation. For example, in certain circumstances, it will be desirable to deliver a pharmaceutical composition comprising the agent orally, through injection by intravenous, intraperitoneal, intracerebral (intra-parenchymal), intracerebroventricular, intramuscular, intraocular, intraarterial, intraportal, intralesional, intramedullary, intrathecal, intraventricular, transdermal, subcutaneous, intraperitoneal, intranasal, enteral, topical, sublingual, urethral, vaginal, or rectal means, by sustained release systems, or by implantation devices. If desired, the compound is administered regionally via intrathecal administration, intracerebral (intra- parenchymal) administration, intracerebroventricular administration, or intraarterial or intravenous administration feeding the region of interest. Alternatively, the composition is administered locally via implantation of a membrane, sponge, or another appropriate material onto which the desired compound has been absorbed or encapsulated. Where an implantation device is used, the device is, in one aspect, implanted into any suitable tissue or organ, and delivery of the desired compound is, for example, via diffusion, timed-release bolus, or continuous administration.

[0080] To facilitate administration, the compound is, in various aspects, formulated into a physiologically-acceptable composition comprising a carrier (e.g., vehicle, adjuvant, or diluent). The particular carrier employed is limited only by chemico-physical considerations, such as solubility and lack of reactivity with the compound, and by the route of administration. Physiologically- acceptable carriers are well known in the art. Illustrative pharmaceutical forms suitable for injectable use include sterile aqueous solutions or dispersions and sterile powders for the extemporaneous preparation of sterile injectable solutions or dispersions (for example, see U.S. Patent No. 5,466,468). Injectable formulations are further described in, e.g., Pharmaceutics and Pharmacy Practice, J. B. Lippincott Co., Philadelphia. Pa., Banker and Chalmers, eds., pages 238-250 (1982), and ASHP Handbook on Injectable Drugs, Toissel, 4th ed., pages 622-630 (1986)). A pharmaceutical composition comprising the compound is, in one aspect, placed within containers, along with packaging material that provides instructions regarding the use of such pharmaceutical compositions. Generally, such instructions include a tangible expression describing the reagent concentration, as well as, in certain embodiments, relative amounts of excipient ingredients or diluents (e.g., water, saline or PBS) that may be necessary to reconstitute the pharmaceutical composition.

[0081] Compositions suitable for parenteral injection may comprise physiologically acceptable sterile aqueous or nonaqueous solutions, dispersions, suspensions, or emulsions, and sterile powders for reconstitution into sterile injectable solutions or dispersions. Examples of suitable aqueous and nonaqueous carriers, diluents, solvents, or vehicles include water, ethanol, polyols (propylene glycol, polyethylene glycol, glycerol, and the like), suitable mixtures thereof, vegetable oils (such as olive oil) and injectable organic esters such as ethyl oleate. Proper fluidity can be maintained, for example, by the use of a coating such as lecithin, by the maintenance of the required particle size in the case of dispersions, and by the use of surfactants.

[0082] These compositions may also contain adjuvants such as preserving, wetting, emulsifying, and dispersing agents. Microorganism contamination can be prevented by adding various antibacterial and antifungal agents, for example, parabens, chlorobutanol, phenol, sorbic acid, and the like. It may also be desirable to include isotonic agents, for example, sugars, sodium chloride, and the like. Prolonged absorption of injectable pharmaceutical compositions can be brought about by the use of agents delaying absorption, for example, aluminum monostearate and gelatin.

[0083] Solid dosage forms for oral administration include capsules, tablets, powders, and granules. In such solid dosage forms, the active compound is admixed with at least one inert customary excipient (or carrier) such as sodium citrate or dicalcium phosphate or (a) fillers or extenders, as for example, starches, lactose, sucrose, mannitol, and silicic acid; (b) binders, as for example, carboxymethylcellulose, alginates, gelatin, polyvinylpyrrolidone, sucrose, and acacia; (c) humectants, as for example, glycerol; (d) disintegrating agents, as for example, agar-agar, calcium carbonate, potato or tapioca starch, alginic acid, certain complex silicates, and sodium carbonate; (a) solution retarders, as for example, paraffin; (f) absorption accelerators, as for example, quaternary ammonium compounds; (g) wetting agents, as for example, cetyl alcohol and glycerol monostearate; (h) adsorbents, as for example, kaolin and bentonite; and (i) lubricants, as for example, talc, calcium stearate, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate, or mixtures thereof. In the case of capsules, and tablets, the dosage forms may also comprise buffering agents. Solid compositions of a similar type may also be used as fillers in soft and hard filled gelatin capsules using such excipients as lactose or milk sugar, as well as high molecular weight polyethylene glycols, and the like.

[0084] Solid dosage forms such as tablets, dragees, capsules, pills, and granules can be prepared with coatings and shells, such as enteric coatings and others well known in the art. The solid dosage forms may also contain opacifying agents. Further, the solid dosage forms may be embedding compositions, such that they release the active compound or compounds in a certain part of the intestinal tract in a delayed manner. Examples of embedding compositions that can be used are polymeric substances and waxes. The active compound can also be in micro-encapsulated form, optionally with one or more excipients.

[0085] Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions, solutions, suspensions, syrups, and elixirs. In addition to the active compounds, the liquid dosage form may contain inert diluents commonly used in the art, such as water or other solvents, solubilizing agents and emulsifiers, as for example, ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butylene glycol, dimethylformamide, oils, in particular, cottonseed oil, groundnut oil, corn germ oil, olive oil, castor oil, and sesame seed oil, glycerol, tetrahydrofurfuryl alcohol, polyethylene glycols and fatty acid esters of sorbitan, or mixtures of these substances, and the like.

[0086] Besides such inert diluents, the composition can also include adjuvants, such as wetting agents, emulsifying and suspending agents, sweetening, flavoring, and perfuming agents. Suspensions, in addition to the active compound, may contain suspending agents, as for example, ethoxylated isostearyl alcohols, polyoxyethylene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum metahydroxide, bentonite, agar-agar, and tragacanth, or mixtures of these substances, and the like.

[0087] Compositions for rectal administration are preferably suppositories, which can be prepared by mixing the compounds of the disclosure with suitable non-irritating excipients or carriers such as cocoa butter, polyethylene glycol or a suppository wax, which are solid at ordinary room temperature, but liquid at body temperature, and therefore, melt in the rectum or vaginal cavity and release the active component.

[0088] The compositions used in the methods of the invention may be formulated in micelles or liposomes. Such formulations include sterically stabilized micelles or liposomes and sterically stabilized mixed micelles or liposomes. Such formulations can facilitate intracellular delivery, since lipid bilayers of liposomes and micelles are known to fuse with the plasma membrane of cells and deliver entrapped contents into the intracellular compartment.

[0089] Upon formulation, solutions will be administered in a manner compatible with the dosage formulation and in such amount as is therapeutically effective. The formulations are easily administered in a variety of dosage forms such as injectable solutions, drug release capsules and the like. For parenteral administration in an aqueous solution, for example, the solution should be suitably buffered if necessary and the liquid diluent first rendered isotonic with sufficient saline or glucose. These particular aqueous solutions are especially suitable for intravenous, intramuscular, subcutaneous and intraperitoneal administration.

[0090] The frequency of dosing will depend on the pharmacokinetic parameters of the agents and the routes of administration. The optimal pharmaceutical formulation will be determined by one of skill in the art depending on the route of administration and the desired dosage. See, for example, Remington’s Pharmaceutical Sciences, 18th Ed. (1990) Mack Publishing Co., Easton, PA, pages 1435-1712, incorporated herein by reference. Such formulations may influence the physical state, stability, rate of in vivo release and rate of in vivo clearance of the administered agents. Depending on the route of administration, a suitable dose may be calculated according to body weight, body surface areas or organ size. Further refinement of the calculations necessary to determine the appropriate treatment dose is routinely made by those of ordinary skill in the art without undue experimentation, especially in light of the dosage information and assays disclosed herein, as well as the pharmacokinetic data observed in animals or human clinical trials.

[0091] The precise dosage to be employed depends upon several factors including the host, whether in veterinary medicine or human medicine, the nature and severity of the condition, e.g., disease or disorder, being treated, the mode of administration and the particular active substance employed. The compounds may be administered by any conventional route, in particular enterally, and, in one aspect, orally in the form of tablets or capsules. Administered compounds can be in the free form or pharmaceutically acceptable salt form as appropriate, for use as a pharmaceutical, particularly for use in the prophylactic or curative treatment of a disease of interest. These measures will slow the rate of progress of the disease state and assist the body in reversing the process direction in a natural manner.

[0092] It will be appreciated that the pharmaceutical compositions and treatment methods of the invention are useful in fields of human medicine and veterinary medicine. Thus the subject to be treated is in one aspect a mammal. In another aspect, the mammal is a human.

[0093] In jurisdictions that forbid the patenting of methods that are practiced on the human body, the meaning of “administering” of a composition to a human subject shall be restricted to prescribing a controlled substance that a human subject will self-administer by any technique (e.g., orally, inhalation, topical application, injection, insertion, etc.). The broadest reasonable interpretation that is consistent with laws or regulations defining patentable subject matter is intended. In jurisdictions that do not forbid the patenting of methods that are practiced on the human body, the “administering” of compositions includes both methods practiced on the human body and also the foregoing activities.

Methods of use

[0094] Provided herein are methods of degrading a cyclin-dependent kinase (CDK) comprising administering to a subject in need thereof a therapeutically effective amount of a composition comprising a CDK inhibitor moiety linked to an E3 ligase binding moiety. Also provided are methods of treating or preventing a disease or disorder capable of being modulated by CDK degradation, comprising administering to a subject in need thereof a therapeutically effective amount of a composition comprising a CDK inhibitor moiety linked to an E3 ligase binding moiety.

[0095] Also provided are compositions comprising a CDK inhibitor moiety linked to an E3 ligase binding moiety for use in a method of degrading a cyclin-dependent kinase (CDK). Also provided are compositions comprising a CDK inhibitor moiety linked to an E3 ligase binding moiety for use in a method of treating or preventing a disease or disorder capable of being modulated by CDK degradation. [0096] In some cases, the CDK inhibitor moiety binds to CDK5. In some cases, the CDK inhibitor moiety binds to CDK9. Without being bound by theory, the compounds disclosed herein (e.g., compounds of Formula 0, Formula 1, Formula 1A, Table 1, and pharmaceutically acceptable salts thereof) bind to CDKs and cause degradation of the bound CDK protein. In some cases, the compounds and salts disclosed herein target specific CDKs. In some cases, the compounds disclosed herein (e.g., compounds of Formula 0, Formula 1 , Formula 1A, Table 1 , and pharmaceutically acceptable salts thereof) bind to and selectively degrade CDK1 , CDK2, CDK3, CDK4, CDK5, CDK6, CDK7, CDK8, CDK9, CDK10, CDK11, CDK12, or CDK13. In some cases, the compounds disclosed herein (e.g., compounds of Formula 0, Formula 1 , Formula 1A, Table 1, and pharmaceutically acceptable salts thereof) bind to and selectively degrade CDK5. In some cases, the compounds disclosed herein (e.g., compounds of Formula 0, Formula 1, Formula 1A, Table 1, and pharmaceutically acceptable salts thereof) bind to and selectively degrade CDK9.

[0097] In some cases, the composition comprising a CDK inhibitor moiety linked to an E3 ligase binding moiety is a compound or salt disclosed herein, e.g., a compound of Formula 0, Formula 1, Formula 1A, Table 1, and pharmaceutically acceptable salts thereof.

[0098] In some cases, the disease or disorder capable of being modulated by CDK degradation is selected from the group consisting of cancer, inflammatory diseases, and neurological diseases.

[0099] In some cases, the disease or disorder is cancer. In some cases, the cancer is leukemia (e.g., acute lymphoblastic leukemia, acute myeloid leukemia, chronic lymphocytic leukemia, chronic myelogenous leukemia), lymphoma (ex. Hodgkin lymphoma, Non-Hodgkin lymphoma), multiple myeloma, breast cancer, prostate cancer, pancreatic cancer, colon cancer, thyroid cancer, bladder cancer, liver cancer, neuroblastoma, brain cancers (gliomas, meningiomas, pituitary adenomas etc.), lung cancer, ovarian cancer, stomach cancer, skin cancer (melanoma), cervical cancer, testicular cancer, kidney cancer, carcinoid tumors, or bone cancer. In some cases, the cancer is pancreatic cancer. In some cases, the cancer is resistant to treatment by Bcl-xL, Bcl2, or Bcl-w inhibition.

[00100] In some cases, the disease or disorder is an inflammatory disease. In some cases, the inflammatory disease is arthritis, atherosclerosis, inflammatory bowel disease, rheumatoid arthritis, colitis, pancreatitis, hepatitis, thyroiditis, Crohn’s disease, asthma, or pelvic inflammatory disease.

[00101] In some cases, the disease or disorder is a neurological disease. In some cases, the neurological disease is Alzheimer’s disease, Parkinson’s disease, traumatic brain injury, stroke, Amyotrophic Lateral Sclerosis, Huntington’s disease, ischemia, attention deficit disorders, or epilepsy. [00102] The compounds described herein (e.g., the compounds of compounds of Formula 0, Formula I, Formula I A, the compounds of Table 1 , or pharmaceutically acceptable salts of the compounds) can degrade a CDK. In some embodiments, the compounds bind to any of CDK1, CDK2, CDK3, CDK4, CDK5, CDK6, CDK7, CDK8, CDK9, CDK10, CDK11 , CDK12, or CDK13 leading to their degradation, e.g., the compounds trigger or inhibit CDK-mediated biological activity, such as gene expression. In various cases, the compounds are CDK modulators, e.g., the compounds change, inhibit, or prevent one or more of a CDK’s biological activities.

[00103] The compounds disclosed herein are particularly advantageous for the treatment of diseases or disorders caused by aberrant expression or activity of a CDK. The incidence and/or intensity of diseases or disorders associated with aberrant expression or activity of a CDK is reduced.

[00104] Selective CDK degraders can be used for cancer prevention and treatment. The relationship between CDK activity and cancer have been in various studies, and CDK has been implicated in the regulation of the transcription of the anti-apoptotic protein Mcl-1 and the oncogene Myc. Compounds of compounds of Formula 0, Formula I, Formula I A, the compounds of Table 1 , and pharmaceutically acceptable salts of the compounds display high selectivity for growth inhibition and/or induction of apoptosis in cancer cells, e.g., in pancreatic cancer cells.

[00105] The disclosed methods include methods for treating disease or disorder capable of being modulated by degradation of CDK, e.g., cancer, comprising administering to a subject a compound that degrades a CDK or a component of a CDK ternary complex with a protein of interest (POI) and an E3-ligase.

[00106] Provided herein are methods of degrading CDK in a cell, comprising contacting the cell with a compound or a composition as disclosed herein (e.g., the compounds of compounds of Formula 0, Formula I, Formula IA, the compounds of Table 1 , or pharmaceutically acceptable salts of the compounds) in an amount sufficient to degrade the CDK. The contacting of the cell can occur in vitro or in vivo. In some cases, contacting of the cell occurs in vitro. In other cases, contacting of the cell occurs in vivo. Therefore, the disclosure includes administering one or more of a compound described herein to a subject, such as a human, in need thereof. In some embodiments, the subject suffers from a disease or disorder associated with aberrant activity of a CDK. Disorders associated with aberrant activity of a CDK include, but are not limited to, cancer (e.g., pancreatic cancer), inflammatory diseases, and neurological diseases. Specifically contemplated cancers include ovarian cancer, breast cancer, prostate cancer, colon cancer, liver cancer, brain cancer, kidney cancer, lung cancer, leukemia, lymphoma, multiple myeloma, thyroid cancer, bone cancer, esophageal cancer, and pancreatic cancer. [00107] In some cases, the methods comprise sensitizing the cancer to a Bcl2 inhibitor. In some cases, the methods disclosed herein further comprise administration of a therapeutic agent. In some cases, the therapeutic agent is ABT-263 (Navitoclax), ABT-199 (Venetoclax), or WEHI-539. In some cases, the therapeutic agent is ABT-199 (Venetoclax).

[00108] The disclosed methods utilize compounds that degrade CDK, for treating, e.g., cancer. Methods for assessing the usefulness of a compound for treating cancer are known to those of skill in the art. For example, compounds may be assessed using models of cancer, including cells (such as pancreatic cancer cells), animal models (such as mouse xenograph or other cancer models), or in human subjects having, e.g., pancreatic cancer.

[00109] The compounds described herein can be used to decrease or prevent cancer in human subjects with e.g., pancreatic cancer. In a particular example, a compound or mixture is administered orally, such as by mixing with distilled water. In another example, a test compound or mixture is administered intravenously, such as in saline or distilled water. In some examples, treatment with test compound may be a single dose or repeated doses. The test compound may be administered about every 6 hours, about every 12 hours, about every 24 hours (daily), about every 48 hours, about every 72 hours, or about weekly. Treatment with repeated doses may continue for a period of time, for example for about 1 week to 12 months, such as about 1 week to about 6 months, or about 2 weeks to about 3 months, or about 1 to 2 months. Administration of a compound may also continue indefinitely. Doses of test compound are from about 0.1 mg/kg to about 400 mg/kg, such as about 1 mg/kg to about 300 mg/kg, about 2 mg/kg to 200 mg/kg, about 10 mg/kg to about 100 mg/kg, about 20 mg/kg to about 75 mg/kg, or about 25 mg/kg to about 50 mg/kg.

[00110] It will be understood that the methods and compositions described herein for treating cancer, comprising administering a compound that degrades CDK, are applicable to methods of treating other diseases related to CDK activity, such as those described above. The methods for assessing the effectiveness of test compounds for treating such diseases in cells, appropriate animal models, or affected subjects are known to one of skill in the art.

[00111] Uses of the compounds disclosed herein in the preparation of a medicament for treating diseases or disorders related to CDK activity also are provided herein.

[00112] The disclosure herein will be understood more readily by reference to the following examples, below.

EXAMPLES

[00113] The following examples are provided for illustration and are not intended to limit the scope of the disclosure. Synthetic procedures for compounds of Formula I

General Experimental Procedures.

[00114] Cell lines and Materials: HEK293 and MiaPaCa2 cells were grown in Dulbeccos Modified Eagle Medium (DMEM) with High Glucose (HyClone #SH30022.FS) supplemented with 10% Fetal Bovine Serum (Gibco by Life Technologies #26140-079) and 1x-pencillin- streptomycin (HyClone #SV30010). Cells were maintained at 37°C and 5% CO2.

[00115] Western Blotting: Following treatment, cells were washed three times with cold 1X phosphate-buffered saline (PBS, HyClone #SH30028.02) and scraped before being lysed in a buffer comprised of radioimmunoprecipitation assay (RIPA) buffer (Thermo Scientific #89900), sodium orthovandate (NasVO^, sodium fluoride (NaF), p-glycerophosphate and 1mM phenylmethylsulfonyl fluoride (PMSF). Samples were then incubated on ice for 30 minutes, vortexed in 10-minute intervals, and pelleted by centrifugation at 14,000 g for 10 minutes at 4°C. The supernatant was collected, and protein quantification was determined using BCA Protein Assay (Pierce #23225). 40 pg of total protein were loaded per well and run on a 4-15% gradient gel (BioRad) in 1X TRIS-Glycine-SDS Buffer (Research Products International Corporation #T32080) at 120V for ~80 minutes and separated by SDS-PAGE (sodium dodecyl sulfate-polyacrylamide gel) electrophoresis. The proteins were transferred onto a polyvinylidene fluoride (PVDF) methanol activated membrane using a semi-dry transfer method

(ThermoScientific #35035) run at 18V for 35 minutes. Membranes were blocked by shaking in a solution of 5% (w/v) non-fat dry milk in 1X-Tris Buffered Saline with 0.1% Tween (IxTBST) for 1 hour at room temperature. Membranes were incubated in primary antibodies (listed below) in 5% milk in IxTBST at 4°C overnight with gentle rocking. Complimentary HRP-conjugated secondary antibodies diluted at 1:10,000 or 1:5,000 incubated in 5% milk in IxTBST, rocked at room temperature for 1 hour. The membranes were then incubated with ECL Prime (Cytiva #RPN2236) to detect protein expression. Imaged was used for the quantification of the Western blots.

[00116] Antibody Information: a-tubulin - Cell Signaling Technologies # 3873 (1:5,000 dilution) CDK9 - Cell Signaling Technologies #2316 (1:1 ,000 dilution) CDK12 - Cell Signaling Technologies #11973 (1:1,000 dilution) CDK8 - Cell Signaling Technologies #17395 (1:1,000 dilution) CDK7 - Cell Signaling Technologies #2916 (1:2,000 dilution) CDK6 - Cell Signaling Technologies #13331 (1:1,000 dilution) CDK5 - Cell Signaling Technologies #2506 (1:1 ,000 dilution) CDK4 - Cell Signaling Technologies #12790 (1:1,000 dilution) CDK2 - Cell Signaling Technologies #2546 (1:2,000 dilution) CDK1- Cell Signaling T echnologies #77055 (1 : 1 ,000 dilution). [00117] Sample Preparation for Label-free and TMT mass spectrometry experiments:

Samples were prepared in a buffer comprised of radioimmunoprecipitation assay (RIPA) buffer (Thermo Scientific #89900), sodium orthovandate (NasVC ), sodium fluoride (NaF), p- glycerophosphate and 1 mM phenylmethylsulfonyl fluoride (PMSF). Extracted protein samples for both label-free quantitation (LFQ) and TMT experiments were acetone precipitated and washed to remove detergent before redissolving in 8 M urea, 100 mM tris/HCI, pH 7.8 (for LFQ experiments) and 7 M urea, 2 M thiourea, 0.5 M triethylammonium bicarbonate (TEAB) buffer, pH 8.5 (for TMT experiments). Samples were assayed for protein and 100pg (LFQ)/200|jg (TMT) was reduced and alkylated, then digested using Lys-C and trypsin. Before labeling of digests with TMT 10plex reagents (Thermofisher Scientific) according to the manufacturer’s instructions, samples were desalted using Sep-Pak® C18 SPE columns (Waters Corp, Milford, MA). Labels were randomized within each 10plex set. Labeling efficiency was calculated as >99.3% labeled for both 10-plex sets. 200 pg of the combined 10-plex mix was subfractionated offline into 96 fractions using high pH reverse phase C18 chromatography (ACQUITY UPLC® BEH C18, 1.7 pm, 2.1 x 150 mm, Waters Corp) in ammonium formate, pH 10. The 96 fractions were recombined to give a total of 12 fractions according to the concatenated strategy of Yang et al (2012). For each LFQ analysis 3pg of peptides was analyzed by mass spectrometry and for each TMT fraction 2.5 pg of peptides was analyzed.

[00118] LC-MS/MS analysis of label-free and TMT-labeled samples: Peptide samples were analyzed by LC-MS/MS on an RSLCnano system (ThermoFisher Scientific) coupled to a Q-Exactive HF mass spectrometer (ThermoFisher Scientific). The samples were first injected onto a trap column (Acclaim PepMap™ 100, 75 pm x 2 cm, ThermoFisher Scientific) before switching in-line with the main column (Acquity UPLC® M-class, Peptide CSH™ 130A, 1.7 pm 75 pm x 250 mm, Waters Corp). Mass spectra were acquired on a Q Exactive HF mass spectrometer in data-dependent mode using a mass range of m/z 375-1500 and MS1 resolution of 120,000. Data-dependent MS2 spectra were acquired by HCD. For the label-free samples the MS2 settings were: 15,000 resolution, AGC target 1e ⁵ ions, maximum ion time 250msec, top20 with a dynamic exclusion time of 60sec. For the TMT, MS2 settings were: 45,000 resolution, AGC target 5e ⁵ ions, maximum ion time 86 msec, top10 with a dynamic exclusion time of 30 sec and the isolation window set to 0.7 m/z to reduce co-isolation.

[00119] Data analysis: Data were analyzed in Proteome Discoverer 2.4 software (ThermoFisher Scientific). Mascot 2.6.2 was used to search the databases; the common contaminants database cRAP (116 entries, www.theGPM.org) and the Human Reference Proteome UniProtKB (74,034 entries from 10/24/2019). For the LFQ approach used for the MiaPaCa2 cells experiment, methionine oxidation, asparagine and glutamine deamidation, protein N-terminal acetylation, cysteine carbamidomethylation, and serine, threonine, tyrosine phosphorylation was set as variable modifications. For the TMT-labeled experiment used for the HEK293 cells, the same variable modifications were used with the exception of the phosphorylation, whilst TMTIOplex (K) and TMTIOplex (N-term) were specified as fixed modifications. The search included a maximum of two trypsin missed cleavages with the precursor mass tolerance set to 10 ppm and the fragment mass tolerance to 0.02 Da, respectively. Peptide validations were done by Percolator with a 0.01 posterior error probability (PEP) threshold. The data were searched using a decoy database to set the false discovery rate to 1% (high confidence). The peptides for MiaPaCa2 cells were quantified using the precursor abundance based on intensity. The peak abundance was normalized for differences in sample loading using total peptide amount where the peptide group abundances are summed for each sample and the maximum sum across all runs is determined. The significance of differential expression reported as Iog2 fold change was tested using an ANOVA test, which provides adjusted p-values using the Benjamini-Hochberg method for all the calculated ratios. The peptide quantification for HEK293 cells was processed using the peak intensity of the TMT reporter ion in the MS2 spectrum, with the co-isolation threshold set to 50% and the average S/N to 10.

[00120] Cell Viability: MiaPaCa2 cells were plated at a density of 4,000 cells/well in a 96- well plate (Thermo Scientific #249946) and allowed to adhere overnight at 37°C, 5% CO2. The following day, cells were treated with indicated concentration of inhibitors. PrestoBlue cell viability (10 pL) reagent (Invitrogen #A13262) was added to cells after 72-hour drug incubation and incubated for 15 min at 37°C to assess the growth inhibition. Fluorescence (560 _e x/590 _e m) was measured using the SpectraMax M5e instrument to assess the growth inhibition. Percentage growth inhibition was calculated using 100 - [100 x (samples - To)/(T o-To)], whereTo is the vehicle control reading immediately following drug addition and Two is the vehicle control reading at the end of the 72-hr incubation.

[00121] Calcusyn: To determine fraction affected as a decimal of 1 , percent growth inhibition data was divided by 100. If a value exceeded 100%, 0.999 was assumed. If a negative value was observed a value of 0.001 was assumed. Using Calcusyn software, combination index (Cl) values were calculated as a mean of Cl values calculated for each clinically relevant effect dose. Clinically relevant effect doses and their corresponding Cl values were determined from the following ED values: ED75, ED90, and ED90, where ED75 is the dose at which 75% of the cells are affected.

[00122] General methods: All reagents were purchased from commercial sources and were used without further purification. Flash chromatography was carried out on silica gel (200-400 mesh). Thin layer chromatography (TLC) were run on pre-coated EMD silica gel 60 F254 plates and observed under UV light at 254 nm and with basic potassium permanganate dip. Column chromatography was performed with silica gel (230-400 mesh, grade 60, Fisher scientific, USA). 1 H NMR (500 MHz) and 13C NMR (125 MHz) spectra were recorded in chloroform-d or DMSO-d6 on a Varian-500 and Varian-600 spectrometer (DMSO-d6 was 2.50 ppm for 1 H and 39.55 ppm for 13C and CDCI3 was 7.27 ppm for 1H and 77.23 ppm for 13C. Proton and carbon chemical shifts were reported in ppm relative to the signal from residual solvent proton and carbon. Final compounds purity was determined by analytical HPLC and was found to be >95% pure. Analysis of sample purity was performed on a Waters Alliance 2695 HPLC system equipped with a Waters 2996 photodiode array detector and an auto-sampler with Phenomenex Luna RP-C18 column (5 pm, 4.6 mm x 250 mm, 100 A). HPLC conditions: solvent A, H2O containing 0.1% formic acid (FA); solvent B, CH3CN containing 0.1% FA; gradient, 10% B to 100% B over 15 min followed by 100% B over 2 min; flow rate, 1 mL/min. Purity was determined at =254 nm. Retention times for each final compound are providedbelow. HRMS for the compounds was generated on an Agilent 6230 LC/TOF system with UV detector (254 nm).

[00123] Example 1 : Synthesis of intermediates

[00124] Synthesis of intermediate 3: To a solution of 2 (700 mg, 2.89 mmol) in DMF was added K2CO3 (798 mg, 5.78 mmol). The solution was heated at 60 °C for 10 minutes followed by addition of tert-butyl 2-bromoacetate (677 mg, 3.47 mmol). The reaction mixture was stirred for 4h. The crude mixture was then poured into brine, extracted with ethyl acetate, and the organic phase was collected and extracted consecutively with water, and dried with MgSO4. Evaporation of the solvent gave a residue which was purified by column chromatography to afford intermediate 3. ¹ H NMR (500 MHz, CDCI3) 6 (ppm) 7.35 - 7.30 (m, 5H), 7.21 (d, 2H, J = 8.5 Hz), 6.85 (d, 2H, J = 8.5 Hz), 5.12 (s, 2H), 4.50 (s, 2H), 3.61 (s, 2H), 1.49 (s, 9H). ¹³ C NMR (125 MHz, CDCI3) 6 (ppm) 171.47, 167.91 , 157.03, 135.79, 130.29, 128.45, 128.11 , 128.02, 126.81 , 114.65, 82.23, 66.47, 65.70, 40.35, 27.96.

[00125] Synthesis of intermediate 4: Intermediate 3 (1.16 g) was combined with 20 mL of dry ethyl acetate via syringe. To the reaction mixture was added Pd/C (116 mg, 5 % by weight on activated carbon) and argon gas was bubbled though the reaction mixture for about 10 minutes. Reaction mixture was gently vacuumed and was kept under hydrogen atmosphere for 16 hours. After completion of reaction, mixture was passed through a bed of celite and column chromatographed using hexane and ethyl acetate gradient to afford intermediate 4. ¹ H NMR (500 MHz, CDCI3) 6 (ppm) 7.18 (d, 2H, J = 8.5 Hz), 6.84 (d, 2H, J = 8.5 Hz), 4.49 (s, 2H), 3.56 (s, 2H), 1.48 (s, 9H). ¹³ C NMR (125 MHz, CDCI3) 6 (ppm) 177.03, 167.98, 157.24, 130.44, 126.23, 114.80, 82.39, 65.77, 39.98, 28.03.

[00126] Synthesis of intermediate 5: A solution of intermediate 2 (500 mg, 2.00 mmol) in DMF was added K2CO3 (553 mg, 4.00 mmol), the solution was heated at 60 °C for 15 minutes followed by addition of tert-butyl 4-bromobutanoate (553 mg, 2.47 mmol). The reaction mixture was stirred for 6h. The crude mixture was then poured into brine, extracted with ethyl acetate, and the organic phase was collected and extracted consecutively with water, and dried with MgSC>4. Evaporation of the solvent gave a residue which was purified by column chromatography to afford 5. ¹ H NMR (500 MHz, CDCh) 6 (ppm) 7.35 - 7.31 (m, 5H), 7.19 (d, 2H, J = 8.5 Hz), 6.85 (d, 2H, J = 8.5 Hz), 5.13 (s, 2H), 3.98 (t, 2H, J = 6.0 Hz), 3.60 (s, 2H), 2.42 (t, 2H, J = 7.5 Hz), 2.09 - 2.03 (m, 2H), 1.46 (s, 9H). ¹³ C NMR (125 MHz, CDCh) 6 (ppm) 172.50, 171.69, 158.04, 135.88, 130.32, 130.27, 128.55, 128.50, 128.16, 128.09, 125.95, 114.94, 114.59, 80.32, 66.87, 66.51, 40.41 , 32.00, 28.09, 24.74.

[00127] Synthesis of intermediate 6: Intermediate 5 (500 mg) was taken in a round bottom flask and added 10 mL of dry ethyl acetate via syringe. To the reaction mixture was added Pd/C (70 mg, 5 % by weight on activated carbon) and argon gas was bubbled though the reaction mixture for about 10 minutes. Reaction mixture was gently vacuumed and was kept under hydrogen atmosphere for 16 hours. After completion of reaction, mixture was passed through a bed of celite and column chromatographed using hexane and ethyl acetate gradient to obtain intermediate 6. ¹ H NMR (500 MHz, CDCh) 6 (ppm) 7.17 (d, 2H, J = 8.5 Hz), 6.84 (d, 2H, J = 8.5 Hz), 3.96 (t, 2H, J = 6.0 Hz), 3.57 (s, 2H), 2.41 (t, 2H, J = 7.5 Hz), 2.07 - 2.02 (m, 2H), 1.44 (s, 9H). ¹³ C NMR (125 MHz, CDCh) 6 (ppm) 177.70, 172.60, 158.18, 130.36, 125.34, 114.64, 80.40, 66.86, 60.41, 40.08, 32.00, 28.08, 24.73, 21.00, 14.16.

[00128] Synthesis of intermediate 8: Intermediate 8 was synthesized as described in Zhang, X. et al, Journal of the American Chemical Society 2013, 135 (25), 9248-51.

[00129] Synthesis of intermediate 9: To a solution of intermediate 8 (473 mg, 1.76 mmol) in acetone (10 ml) was added Nal (754 mg 5.29 mmol). The reaction mixture was stirred at reflux temperature for 24 h, then the solvent was removed under vacuum and crude product was dissolved in ethyl acetate and washed with aqueous 10% NaHSCh, brine, dried (MgSCU) and evaporated under vacuum to yield intermediate 9. ¹ H NMR (500 MHz, CDCh) 6 (ppm) 3.90 (s, 2H), 3.64 - 3.54 (m, 10H), 3.14 (s, 2H), 1.35 (s, 9H). ¹³ C NMR (125 MHz, CDCh) 6 (ppm) 169.16, 80.96, 71.51, 70.28, 70.17, 69.77, 68.61, 27.74.

[00130] Synthesis of intermediate 10: To a solution of intermediate 2 (323 mg, 1.34 mmol) in DMF was added K2CO3 (370 mg, 2.68 mmol), the solution was heated at 60 °C for 10 minutes followed by addition of tert-butyl 2-(2-(2-(2-iodoethoxy)ethoxy)ethoxy)acetate (600 mg, 1.60 mmol). The reaction mixture was stirred for additional 16h at 60 °C. The crude mixture was then poured into brine, extracted with ethyl acetate, and the organic phase was collected and extracted consecutively with water, and dried with MgSC>4. Evaporation of the solvent gave a residue which was purified by column chromatography to afford intermediate 10. ¹ H NMR (500 MHz, CDCh) 6 (ppm) 7.35 - 7.30 (m, 5H), 7.18 (d, 2H, J = 8.5 Hz), 6.86 (d, 2H, J = 8.5 Hz), 4.11 (t, J = 5.0 Hz), 4.02 (s, 2H), 3.84 (t, 2H, J = 5.0 Hz), 3.74 - 3.68 (m, 8H), 3.59 (s, 2H), 1.47 (s, 9H). 2.42 (t, 2H, J = 7.5 Hz), 2.09 - 2.03 (m, 2H), 1.46 (s, 9H). ¹³ C NMR (125 MHz, CDCh) 6 171.59, 169.57, 157.88, 135.82, 130.20, 128.44, 128.10, 128.02, 126.07, 114.66, 81.41, 70.72, 70.65, 70.57, 70.56, 69.64, 68.96, 67.37, 66.44, 40.35, 28.03.

[00131] Synthesis of intermediate 11 : Intermediate 10 (510 mg) was taken in a round bottom flask and added 10 mL of dry ethyl acetate via syringe. To the reaction mixture was added Pd/C (70 mg, 5 % by weight on activated carbon) and argon gas was bubbled though the reaction mixture for about 10 minutes. Reaction mixture was gently vacuumed and was kept under hydrogen atmosphere for 16 hours. After completion of reaction, mixture was passed through a bed of celite and column chromatographed using hexane and ethyl acetate gradient to yield intermediate 11. ¹ H NMR (500 MHz, CDCh) 6 (ppm) 7.15 (d, 2H, J = 8.5 Hz), 6.84 (d, 2H, J = 8.5 Hz), 4.11 - 4.07 (m, 2H), 4.00 (s, 2H), 3.82 (t, 2H, J = 5.0 Hz), 3.72 - 3.66 (m, 8H), 3.55 (s, 2H), 1.45 (s, 9H). 2.42 (t, 2H, J = 7.5 Hz), 2.09 - 2.03 (m, 2H), 1.46 (s, 9H). ¹³ C NMR (125 MHz, CDCh) 6 177.05, 169.66, 157.93, 130.27, 125.63, 114.69, 81.52, 70.67, 70.62, 70.53, 70.50, 69.62, 68.93, 67.32, 60.36, 40.04, 28.01 , 20.94, 14.09.

[00132] Synthesis of intermediate 12: Intermediate 12 was synthesized as described in Robb, C. M. et al., Chemical communications 2017, 53 (54), 7577-7580.

[00133] Synthesis of intermediate 13: To a stirred solution of intermediate 4 (25 mg, 0.10 mmol), tert-butyl 3-amino-5-cyclobutyl-1 H-pyrazole-1-carboxylate 12 (15 mg, 0.06 mmol) in dichloromethane (1 mL) was added and triethyl amine (32 mg, 0.32 mmol). The reaction mixture was stirred for 10 minutes followed by addition of 50% T3P in ethyl acetate (30 mg, 0.09 mmol). The reaction was stirred overnight and the progress of the reaction was monitored by thin layer chromatography. The crude mixture was dissolved in dichloromethane, washed brine. The organic phase was collected, dried with MgSC>4 and the solvent was evaporated and purified by flash column chromatography with hexane/ethyl acetate to give intermediate 13. ¹ H NMR (500 MHz, CDCI3) 5 (ppm) 10.23 (s, 1 H), 7.28 (d, 2H, J = 3.0 Hz), 6.92 (d, 2H, J = 3.0 Hz), 6.87 (s, 1 H), 4.52 (s, 2H), 3.71 (s, 2H), 3.56 - 3.51 (m, 1 H), 2.34 - 1.88 (m, 6H), 1.63 (s, 9H), 1.51 (s, 9H). ¹³ C NMR (125 MHz, CDCI3) 167.88, 161.06, 157.48, 150.93, 141.35, 130.52, 126.62, 115.18, 95.49, 86.26, 82.35, 65.82, 43.62, 34.33, 28.38, 28.03, 27.92, 18.68.

[00134] Synthesis of intermediate 14: To a stirred solution of intermediate 13 (220 mg) in CH2CI2 (2 mL) at 0 °C was added 1 mL of trifluoroacetic acid dropwise and reaction mixture was stirred for 2h. After completion, the reaction mixture was concentrated in vacuo to give intermediate 14, which was dissolved in 3 mL of dry DMF and stored at -20 °C before use in the next step.

[00135] Synthesis of intermediate 15: To a stirred solution of amine 12 (31 mg, 0.13 mmol), 2-(4-(4-(tert-butoxy)-4-oxobutoxy)phenyl)acetic acid 6 (50 mg, 0.17 mmol) in dichloromethane (2 mL) was added and triethyl amine (66 mg, 0.65 mmol). The reaction mixture was stirred for 10 minutes followed by dropwise addition of 50% T3P in ethyl acetate (120 jiL, 0.19 mmol). The reaction was stirred for 4h and the progress of the reaction was monitored by thin layer chromatography. The crude mixture was dissolved in dichloromethane, washed brine. The organic phase was collected, dried with MgSC>4 and the solvent was evaporated and purified by flash column chromatography with hexane/ethyl acetate to give intermediate 15. ¹ H NMR (500 MHz, CDCI3) 5 (ppm) 10.19 (s, 1 H), 7.22 (d, 2H, J = 8.5 Hz), 6.89 (d, 2H, J = 3.0 Hz), 6.84 (s, 1 H), 3.99 (t, 2H, J = 6.0Hz), 3.68 (s, 2H), 3.56 - 3.49 (m, 1 H), 2.42 (t, 2H, 7.5 Hz), 2.32 - 1.86 (m, 6H), 1.60 (s, 9H), 1.45 (s, 9H). ¹³ C NMR (125 MHz, CDCI3) 172.48, 168.12, 161.09, 158.47, 150.92, 141.41 , 130.50, 125.70, 115.09, 95.44, 86.23, 80.36, 67.00, 43.65, 34.34, 32.02, 28.40, 28.11 , 27.92, 24.74, 18.69.

[00136] Synthesis of intermediate 16: To a stirred solution of intermediate 15 (59 mg) in CH2CI2 (2 mL) at 0 °C was added 1 mL of trifluoroacetic acid dropwise and reaction mixture was stirred for 2h. After completion, the reaction mixture was concentrated in vacuo to give intermediate 16 which was dissolved in 2 mL of dry DMF and stored at -20 °C for use in the next step.

[00137] Synthesis of intermediate 17: To a stirred solution of amine 12 (71 mg, 0.30 mmol), 2-(4-((13,13-dimethyl-11-oxo-3, 6,9, 12-tetraoxatetradecyl)oxy)phenyl)acetic acid 11 (150 mg, 0.38 mmol) in dichloromethane (5 mL) was added and triethyl amine (46 mg, 0.45 mmol). The reaction mixture was stirred for 10 minutes followed by dropwise addition of 50% T3P in ethyl acetate (1 mL, 1.5 mmol). The reaction was stirred for 4h and the progress of the reaction was monitored by thin layer chromatography. The crude mixture was dissolved in dichloromethane, washed brine. The organic phase was collected, dried with MgSC>4 and the solvent was evaporated and purified by flash column chromatography with hexane/ethyl acetate to give intermediate 17. ¹ H NMR (500 MHz, CDCI3) 5 (ppm) 10.19 (s, 1 H), 7.22 (d, 2H, J = 8.5 Hz), 6.91 (d, 2H, J = 8.5 Hz), 6.84 (s, 1 H), 4.13 - 4.11 (m, 2H), 4.01 (s, 2H), 3.88 - 3.85 (t, 2H), 3.73 - 3.68 (m, 10H), 3.55 - 3.49 (m, 1 H), 2.31 - 2.25 (m, 2H), 2.21 - 2.14 (m, 2H), 2.04 - 1.86 (m, 2H), 1.60 (s, 9H), 1.46 (s, 9H). ¹³ C NMR (125 MHz, CDCI ₃ ) 169.61 , 168.05, 161.04, 158.32, 150.87, 141.38, 130.44, 125.85, 115.19, 95.43, 86.25, 81.48, 70.79, 70.70, 70.62, 70.61 , 69.66, 69.01 , 67.50, 43.63, 34.30, 28.37, 28.08, 27.91 , 18.67.

[00138] Synthesis of intermediate 18: To a stirred solution of intermediate 17 (110 mg) in CH2CI2 (5 mL) at 0 °C was added 2 mL of trifluoroacetic acid dropwise and the reaction mixture was stirred for 3h. After completion, the reaction mixture was concentrated in vacuo to give intermediate 18 which was dissolved in 3 mL of dry DMF and stored at -20 °C for use in the next step.

[00139] Synthesis of intermediates 19, 20, and 24: Intermediates 19, 20, and 24 were synthesized as described in Rana, S. et al. Bioorganic & medicinal chemistry letters 2019, 29 (11), 1375-1379.

[00140] Synthesis of intermediate 22: To a stirred solution of intermediate 21 (100 mg, 0.36 mmol) in DMA (2 mL) was added tert-butyl (2-(2-(2-aminoethoxy)ethoxy)ethyl)carbamate (90 mg, 0.35 mmol) and DIPEA (0.2 mL, 1.08 mmol). The reaction mixture was stirred at 90 °C for 2 h. The mixture was cooled to room temperature, poured into brine, extracted twice with ethyl acetate, and dried with MgSC . After filtration and evaporation, the crude residue was purified by column chromatography to give intermediate 22. ¹ H NMR (500 MHz, CDCI3) 5 (ppm) 8.53 (s, 1 H), 7.48 (t, 1 H, 8.0 Hz), 7.09 (d, 1 H, 7.0 Hz), 6.89 (d, 1 H, 8.0 Hz), 6.50 (s, 1 H), 5.05 - 4.9 (m, 2H), 3.72 - 3.30 (m, 12H), 2.87 - 2.72 (m, 3H), 2.12 - 2.10 (m, 1 H), 1.41 (s, 9H). ¹³ C NMR (125 MHz, CDC ) 171.14, 171.10, 169.29, 168.43, 167.56, 156.03, 146.76, 135.99, 132.51 , 116.67, 111.63, 110.36, 79.22, 70.71 , 70.32, 70.11 , 69.34, 60.34, 48.85, 42.29, 40.36, 31.35, 28.37, 22.80, 20.99, 14.15.

[00141] Synthesis of intermediate 23: To a stirred solution of intermediate 22 (50 mg) in CH ₂ CI ₂ (1 mL) at 0 °C was added 1 mL of trifluoroacetic acid dropwise and reaction mixture was stirred for 3h. After completion, the reaction mixture was concentrated in vacuo to give intermediate 23 which was dissolved in 3 mL of dry DMF and stored at -20 °C for use in the next step.

[00142] Example 2: Synthesis of Compound 1 (N-(5-cyclobutyl-1 H-pyrazol-3-yl)-2-(4-(2- ((4-((2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoiso indolin-4-yl)amino)butyl)amino)-2- oxoethoxy)phenyl)acetamide):

[00143] A solution of intermediate 14 (10 mg, 0.03 mmol) and intermediate 19 (10 mg, 0.023 mmol) in DMF (1 mL) was stirred followed by adding EDC (8.5 mg, 0.04 mmol), HOBT (6.6 mg, 0.04 mmol) and NEts (20 pL, 0.04 mmol) and stirred overnight at the ambient temperature. The mixture was rotavaped and purified by reverse phase HPLC (5-95% CH3CN in H ₂ O) to give Compound 1. ¹ H NMR (500 MHz, CD3OD) 5 (ppm) 7.51 (t, 1 H, J = 6.0 Hz), 7.28 (br, 2H), 7.07 - 6.94 (m, 4H), 5.07 - 5.04 (m, 1 H), 3.97 (t, 2H, J = 5.5 Hz), 4.50 (s, 2H), 3.65 - 3.62 (m, 2H), 3.52 (br, 1 H), 2.88 - 2.66 (m, 3H), 2.36 - 2.34 (m, 2H), 2.19 - 2.16 (m, 2H), 2.10 - 2.02 (m, 2H), 1.94 - 1.93 (m, 2H), 1.66 - 1.54 (m, 4H), 0.96 - 0.88 (m, 3H). HRMS-ESI (+) calcd m/z for C34H38N7C 656.2827 (M+H) ⁺ , found 656.2830. HPLC purity >95%, t= 13.77 min.

[00144] Example 3: Synthesis of Compound 2 (4-(4-(2-((5-cyclobutyl-1H-pyrazol-3- yl)amino)-2-oxoethyl)phenoxy)-N-(4-((2-(2,6-dioxopiperidin-3 -yl)-1,3-dioxoisoindolin-4- yl)amino)butyl)butanamide):

[00145] Compound 2 was synthesized in a similar fashion to compound 1. ¹ H NMR (500 MHz, CD ₃ OD) 5 (ppm) 7.55 - 7.47 (m, 1 H), 7.23 (br, 2H), 7.01 - 7.00 (m, 2H), 6.88 (br, 2H), 5.07 - 5.03 (m, 1 H), 3.97 (t, 2H, J = 5.5 Hz), 3.65 - 3.61 (m, 1 H), 3.28 - 3.20 (m, 4H), 2.95 - 2.66 (m, 3H), 2.39 - 2.36 (m, 4H), 2.19 (br, 2H), 2.09 - 2.04 (m, 4H), 1.94 (br, 1 H), 1.66 - 1.54 (m, 4H), 1.32 - 1.29 (m, 2H). HRMS-ESI (+) calcd m/z for C36H42N7CV 684.3140 (M+H) ⁺ , found 684.3141. HPLC purity >95%, t _R = 13.20 min.

[00146] Example 4: Synthesis of Compound 3 (N-(5-cyclobutyl-1 H-pyrazol-3-yl)-2-(4-(2- ((8-((2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindolin-4-yl)a mino)octyl)amino)-2- oxoethoxy)phenyl)acetamide):

[00147] Compound 3 was synthesized in a similar fashion to compound 1. ¹ H NMR (500 MHz, CD3OD) 5 (ppm) 7.52 (t, 1 H, J = 7.5 Hz), 7.28 - 7.26 (m, 2H), 7.02 - 7.00 (m, 2H), 6.95 -

6.94 (m, 2H), 6.28 (br, 1 H), 5.06 - 5.02 (m, 1 H), 4.48 (s, 1 H), 3.65 - 3.54 (m, 3H), 3.30 - 3.23 (m, 8H), 2.86 - 2.65 (m, 4H), 2.38 - 2.36 (m, 2H), 2.24 - 2.16 (m, 2H), 2.10 - 2.04 (m, 2H),

1 .94 - 1.92 (m, 1 H), 1.66 - 1.60 (m, 2H), 1 .54 - 1.48 (m, 2H), 1.40 - 1.29 (m, 8H). HRMS-ESI (+) calcd m/z for C4oH ₅ oN ₇ Oio ⁺ 712.3453 (M+H) ⁺ , found 712.3457. HPLC purity >95%, t _R = 15.11 min. [00148] Example 5: Synthesis of Compound 4 (N-(5-cyclobutyl-1 H-pyrazol-3-yl)-2-(4-(2-

((2-(2-(2-((2-(2,6-dioxopiperidin -3-yl)-1,3-dioxoisoindolin-4- yl)amino)ethoxy)ethoxy)ethyl)amino)-2-oxoethoxy) phenyl)acetamide)

[00149] Compound 4 was synthesized in a similar fashion to compound 1. ¹ H NMR (500 MHz, CD ₃ OD) 5 (ppm) 7.49 (t, 1 H, J = 7.5 Hz), 7.29 - 7.22 (m, 2H), 7.01 - 6.99 (m, 2H), 6.91 - 6.89 (m, 2H), 6.28 (br, 1 H), 5.04 - 5.00 (m, 1 H), 4.45 (s, 1 H), 3.70 - 3.68 (m, 2H), 3.60 - 3.57 (m, 8H), 3.47 - 3.42 (m, 4H), 2.86 - 2.63 (m, 3H), 2.39 - 2.34 (m, 2H), 2.22 - 2.16 (m, 2H), 2.09 - 2.04 (m, 2H), 1.95 - 1.92 (m, 1 H). HRMS-ESI (+) calcd m/z for C36H42N7CV 716.3039 (M+H) ⁺ , found 716.3039. HPLC purity >95%, t _R = 11.24 min.

[00150] Example 6: Synthesis of Compound 5 (4-(4-(2-((5-cyclobutyl-1H-pyrazol-3- yl)amino)-2-oxoethyl)phenoxy)-N-(2-(2-(2-((2-(2,6-dioxopiper idin-3-yl)-1,3-dioxoisoindolin- 4-yl)amino)ethoxy)ethoxy)ethyl)butanamide):

[00151] Compound 5 was synthesized in a similar fashion to compound 1. ¹ H NMR (500 MHz, CD3OD) 5 (ppm) 7.52 (t, 1 H, J = 7.5 Hz), 7.22 - 7.20 (m, 2H), 7.05 - 7.02 (m, 2H), 6.86 - 6.84 (m, 2H), 6.28 (br, 1 H), 5.05 - 5.02 (m, 1 H), 3.94 (t, 2H, J = 6.0 Hz), 3.67 (t, 2H, J = 5.0 Hz), 3.60 - 3.57 (m, 4H), 3.52 (t, 2H, J = 5.0 Hz), 3.45 (t, 2H, J = 5.0 Hz), 3.35 (t, 2H, J = 5.0 Hz), 2.86 - 2.64 (m, 3H),2.36 - 2.33 (m, 4H), 2.23 - 2.16 (m, 2H), 2.09 - 2.00 (m, 4H). HRMS- ESI (+) calcd m/z for C38H ₄ 6N ₇ O ₉ ⁺ 744.3352 (M+H) ⁺ , found 744.3355. HPLC purity >95%, t _R = 12.90 min.

[00152] Example 7: Synthesis of Compound 6 (N-(5-cyclobutyl-1 H-pyrazol-3-yl)-2-(4- ((16-((2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoiso indolin-4-yl)amino)-2-oxo-7,10,13-trioxa-3- azahexadecyl)oxy)phenyl)acetamide):

[00153] Compound 6 was synthesized in a similar fashion to compound 1. ¹ H NMR (500 MHz, CD3OD) 5 (ppm) 7.51 (t, 1 H, J = 7.5 Hz), 7.28 - 7.27 (m, 2H), 7.04 - 6.99 (m, 2H), 6.95 - 6.94 (m, 2H), 6.28 (br, 1 H), 5.05 - 5.01 (m, 1 H), 4.46 (s, 2H), 3.65 - 3.51 (m, 10H), 3.47 (t, 2H, J = 6.0 Hz), 3.40 (t, 2H, J = 6.5 Hz), 3.35 (t, 2H, J = 6.5 Hz), 2.74 - 2.65 (m, 3H), 2.38 - 2.36 (m, 2H), 2.24 - 2.18 (m, 2H), 2.09 - 2.04 (m, 2H), 1.90 - 1.85 (m, 2H), 1.78 - 1.73 (m, 2H). HRMS-ESI (+) calcd m/z for C4OH ₅ ON ₇ OIO ⁺ 788.3614 (M+H) ⁺ , found 788.3619. HPLC purity >95%, t _R = 13.41 min.

[00154] Example 8: Synthesis of Compound 7 (N-(5-cyclobutyl-1 H-pyrazol-3-yl)-2-(4- ((16-((2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindolin-4-yl) amino)-11-oxo-3,6,9-trioxa-12- azahexadecyl)oxy)phenyl)acetamide):

[00155] Compound 7 was synthesized in a similar fashion to compound 1. ¹ H NMR (500 MHz, CD ₃ OD) 5 (ppm) 7.51 (t, 1 H, J = 7.5 Hz), 7.28 - 7.27 (m, 2H), 7.02 - 6.99 (m, 2H), 6.89 (br, 2H), 5.06 - 5.02 (m, 1 H), 4.43 - 4.08 (m, 2H), 3.95 (s, 2H), 3.85 - 3.79 (m, 3H), 3.66 - 3.64 (m, 12H), 2.88 - 2.68 (m, 3H), 2.66 (s, 1 H), 2.37 (br, 2H), 2.20 (br, 2H), 2.10 - 2.08 (m, 2H), 1.62 (br, 4H). HRMS-ESI (+) calcd m/z for C4OH ₅ ON ₇ OIO ⁺ 788.3614 (M+H) ⁺ , found 788.3618. HPLC purity >95%, t _R = 12.66 min.

[00156] Example 9: Synthesis of Compound 8 (4-(4-(2-((5-cyclobutyl-1H-pyrazol-3- yl)amino)-2-oxoethyl)phenoxy)-N-(3-(2-(2-(3-((2-(2,6-dioxopi peridin-3-yl)-1,3- dioxoisoindolin-4-yl)amino)propoxy)ethoxy)ethoxy)propyl) butanamide):

[00157] Compound 8 was synthesized in a similar fashion to compound 1. ¹ H NMR (500 MHz, CD3OD) 5 (ppm) 7.53 (t, 1 H, J = 7.5 Hz), 7.23 (br, 2H), 7.06 - 7.01 (m, 2H), 6.89 (br, 2H), 5.06 - 5.02 (m, 1 H), 3.96 (t, 2H, J = 5.5 Hz), 3.67 - 3.40 (m, 13H), 3.24 (t, 2H, J = 7.0 Hz), 2.86

- 2.66 (m, 3H), 2.37 - 2.34 (m, 2H), 2.00 (br, 1 H), 2.06 - 2.02 (m, 2H), 1.90 - 1.88 (m, 2H), 1.74 - 1.69 (m, 2H). HRMS-ESI (+) calcd m/z for C ₄ 2H ₅ 4N ₇ OIO ⁺ 816.3927 (M+H) ⁺ , found 816.3928. HPLC purity >95%, t _R = 13.44 min.

[00158] Example 10: Synthesis of Compound 9 (N-(5-cyclobutyl-1 H-pyrazol-3-yl)-2-(4- ((1-((2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindolin-4-yl)a mino)-10-oxo-3,6,12,15,18- pentaoxa-9-azaicosan-20-yl)oxy)phenyl)acetamide):

[00159] Compound 9 was synthesized in a similar fashion to compound 1. ¹ H NMR (500 MHz, CD ₃ OD) 5 (ppm) 7.56 - 7.51 (m, 1 H), 7.23 (br, 2H), 7.06 - 7.04 (m, 2H), 6.88 (br, 2H), 5.05 - 5.02 (m, 1 H), 4.09 (m, 2H), 3.94 (br, 2H), 3.81 - 3.80 (m, 2H), 3.65 - 3.61 (m, 14H), 3.55 (t, 2H, J = 5.0 Hz), 3.47 - 3.39 (m, 4H), 2.89 - 2.66 (m, 3H), 2.36 (br, 2H), 2.19 (br, 2H), 2.09 - 2.02 (m, 2H). HRMS-ESI (+) calcd m/z for C42H ₅ 4N ₇ Oi2 ⁺ 848.3825 (M+H) ⁺ , found 848.3822. HPLC purity >95%, t _R = 13.83 min.

[00160] Example 11 : Synthesis of Compound 10 (N-(5-cyclobutyl-1H-pyrazol-3-yl)-2-(4- ((25-((2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoiso indolin-4-yl)amino)-11-oxo-3,6,9,16, 19,22- hexaoxa-12-azapentacosyl) oxy)phenyl) acetamide):

[00161] Compound 10 was synthesized in a similar fashion to compound 1. ¹ H NMR (500 MHz, CD3OD) 5 (ppm) 7.53 (t, 1 H, J = 7.0 Hz), 7.24 (br, 2H), 7.06 - 7.02 (m, 2H), 6.90 (br, 2H), 5.06 - 5.02 (m, 1 H), 4.11(br, 2H), 3.94 (br, 2H), 3.83 - 3.82 (m, 2H), 3.70 - 3.54 (M, 18H), 3.48 (T, 2H, J = 6.0 Hz ), 3.42 (T, 2H, J = 6.0 Hz), 2.89 - 2.66 (m, 3H), 2.36 (br, 2H), 2.19 (br, 2H), 2.10 - 2.01 (m, 2H), 1.94 - 1.88 (m, 2H), 1.75 (t, 2H, J = 6.0 Hz). HRMS-ESI (+) calcd m/z for C ₄ 6H ₆ 2N ₇ Oi3 ⁺ 920.4400 (M+H) ⁺ , found 920.4404. HPLC purity >95%, t _R = 13.58 min.

Biological assay data

[00162] Example 12: CDK9 Degradation Study.

[00163] Compounds 1-10 were screened in a dose-response and time-course study to identify the optimal linker that induces CDK9 degradation. Briefly, HEK293 cells were subjected to increasing concentrations (0.01 - 10 mM) of the PROTACs. CDK9 degradation by the PROTACs were assessed by western blot analysis of the above lysates (Figure 2). [00164] The screen identified Compound 2 as the most potent CDK9 degrader with a DC50 value of 158 ± 6 nM and almost complete degradation was observed at 1 iM (Figure 2). It was also determined that compound 2 does not degrade the CDK9 binding partner Cyclin K (Figure 7).

[00165] Decreasing the linker length by 2-carbon atoms, 1 -carbon atom on either side of the amide, in Compound 1 resulted in complete loss of activity. Increasing the linker length by two additional carbon atoms in compound 3 resulted in partial CDK9 degradation with DC50 ~ 1 piM (Figure 2). Surprisingly, linkers with oxygen atoms exhibited little to no CDK9 degradation (Figure 2). These studies suggest that selection of linker length and linker composition such that they can form a stable ternary complex can allow for efficient CDK9 degradation.

[00166] Example 13: CDK Family Degradation Study.

[00167] The selectivity of compound 2 across other CDK family members was also studied in dose-dependent (Figure 3A) and time-dependent (Figure 3B) studies. HEK293 cells were treated with indicated concentrations of compound 2 for 24 h and the lysates were subjected to Western blot analyses and probed for the levels of indicated CDKs (Figure 3A). In the dosedependent study, compound 2 selectively degraded CDK9 without affecting the levels of other CDK family members. Next, HEK293 cells to were treated with 1 DM of compound 2 and the resulting lysates were probed for various CDKs at indicated time points (Figure 3B). Compound 2 degraded CDK9 as early as the 4 h time point and the degradation was sustained for 24 h. It is important to note that compound 2 did not degrade any other CDKs probed (Figure 3B). Together these studies clearly show that compound 2 selectively degrades CDK9 in dose- and time-dependent studies.

[00168] Example 14: Mechanistic Studies.

[00169] It has been shown that aminopyrazole analogs are CDK 2/5 inhibitors, and the data herein demonstrate that the aminopyrazole based PROTAC compound 2 selectively degrades CDK9. To determine if the selective degradation of CDK9 is a result of loss of binding to CDK2/5 due to the modifications of the aminopyrazole inhibitor to convert it to a PROTAC, compound 2 and CDK inhibitor 11 (structure shown in Figure 3C) were evaluated in in vitro cell- free kinase assays. Inhibitor 11 and compound 2 exhibited nM potency (IC50) in blocking the kinase activities of CDK2 and CDK5. The CDK2, CDK5 and CDK9 kinase activity inhibition profile of Inhibitor 11 and compound 2 was similar (Figure 3C). Likewise, the binding affinity (KD) profile of Inhibitor 11 and compound 2 for CDK2, CDK5 and CDK9 were also similar (Figure 3D). Without wishing to be bound by theory, since the binding affinity profile and the kinase activity inhibitory profile of analog 11 and compound 2 are similar, the selective degradation of CDK9 by compound 2 can be attributed to the differential topographical distributions of lysine residues among CDK2, CDK5 and CDK9. [00170] Next, competition experiments were conducted to evaluate the mechanism of action of compound 2. Without wishing to be bound by theory, it is believed that since degradation of CDK9 by compound 2 follows the formation of a ternary complex between CDK9:compound 2:cerblon (CRBN)-E3 ligase, two sets of competition experiments were conducted with compound 2 and pomalidomide (a CRBN ligand), and compound 2 and flavopiridol (a CDK9 inhibitor). HEK293 cells were treated with either 10 .M of pomalidomide or flavopiridol alone and in combination with 1 iM of compound 2 for 24 h and 8 h, respectively and the lysates were subjected to Western blot analysis. The results show that pomalidomide or flavopiridol by itself did not affect CDK9 levels but was able to block compound 2 mediated degradation of CDK9 when treated in combination (Figure 4A and 4B). No such changes were observed in CDK7 levels another CDK involved in transcriptional regulation, which was used as a control. These competition studies demonstrate the need for simultaneous engagement of CDK9 and a CRBN E3 ligase (ternary complex) by compound 2 to facilitate CDK9 degradation. Since the PROTAC-based strategy involves the ubiquitination of target protein followed by its proteasomal degradation, HEK293 cells were also treated with increasing concentrations of proteasome inhibitor MG132 and compound 2 for 24 h. CDK9 degradation was abrogated in the presence of MG132. The membranes were also probed for CDK1 and CDK12 and no such changes were observed in their levels (Figure 4C). Collectively, these studies confirmed the formation of the ternary complex between CDK9:compound 2:CRBN, followed by CDK9 ubiquitination and subsequent proteasomal degradation as the mechanism of action of compound 2.

[00171] Example 15: Kinome and Proteome Selectivity Studies.

[00172] To assess kinome and proteome selectivity of compound 2, quantitative mass spectrometry was performed with lysates from HEK293 and MiaPaCa2 cell lines treated with Compound 2 for 24 h (Figures 5A and 5B). MiaPaCa2 cells were selected for the proteome wide profiling as they were used to validate CDK9 as a therapeutic target for pancreatic cancers.9229 unique kinases were quantified, including 13 CDKs in the HEK293 lysate and CDK9 was the only kinase identified as a hit with a 1.5-fold change and a significance threshold of P < 0.001 (Figure 5A). 3433 proteins were quantified in the MiaPaCa2 lysate and three kinases CDK9, CDK2 and RPS6KA1 were identified as hits with a 1.5-fold change and a significance threshold of P < 0.001 (Figure 5B).

[00173] Example 16: Synergism Studies.

[00174] Recent studies have shown that concurrent inactivation of Mcl-1 , and Bcl-xL resulted in robust induction of apoptosis. Consequently, targeting Mcl-1, and Bcl-xL is considered a therapeutic strategy for pancreatic cancer therapy. Direct inhibitors of Bcl-xL/Bcl2/Bcl-w have been developed (Figure 6A). However, resistance to Bcl-xL/Bcl2/Bcl-w inhibition has been attributed to compensatory activity by Mcl-1 and Mcl-1 inactivation sensitized cancer cells to Bcl2 inhibitors. Since CDK9 activity regulates the levels of pro-survival protein Mcl-1, the ability of the selective CDK9 degrader compound 2 to sensitize MiaPaCa2 cells to Bcl2 inhibitors was tested.

[00175] To test the above, three Bcl2 inhibitors were selected with varying Bcl2 selectivity profiles: ABT-263 (Navitoclax) is a clinical candidate that underwent a phase II trial and is a Bcl2/Bcl-xL/Bcl-w inhibitor, ABT- 199 (Venetoclax) is a FDA approved Bcl2 inhibitor, and WEHI- 539 is an experimental Bcl-xL inhibitor (Figure 6A). Growth inhibition studies were performed to determine if compound 2 sensitizes MiaPaCa2 cells to the above Bcl2 inhibitors. Briefly, MiaPaCa2 cells were treated individually with ABT-263 (Bcl2/Bcl-xL/Bcl-w), ABT-199 (Bcl2), WEHI-537 (Bcl-xL), compound 2 (CDK9), and the combination of Bcl2 inhibitors and compound 2, respectively. Following a 3-day incubation, cell viability was measured using a PrestoBlue assay. The selective compounds compound 2, ABT199 and WEHI-539 showed minimal growth inhibitory effects individually at equimolar concentration, while the non-selective inhibitor ABT- 263 induced -50% growth inhibition. However, compound 2 sensitized MiPaCa2 cells to ABT- 263 or ABT-199. Interestingly, compound 2 did not sensitize MiaPaCa2 to the Bcl-xL selective inhibitor WEHI-539 (Figure 6B). Consistently, the combination index (Cl) values determined using CalcuSyn at effective doses (ED) of 50, 75, and 90, showed that compound 2 exhibited strong synergism with ABT-263 and ABT-199 but not with WEHI-539 (Figure 6C). Together, these data show that compound 2 potently sensitizes MiaPaCa2 cells to the FDA approved Bcl2 inhibitor Venetoclax.

[00176] In view of the many possible embodiments to which the principles of the disclosure may be applied, it should be recognized that the illustrated embodiments are only examples and should not be taken as limiting the scope of the invention.