Bifunctional degrading agents of interleukin-1 receptor-related kinases and their therapeutic use

Abstract

To provide a bifunctional degrading agent for interleukin-1 receptor-related kinases and its therapeutic use. [Solution] This disclosure provides a bifunctional compound as an IRAK4 degradation agent via the ubiquitin-proteasome pathway, and a method for treating diseases modulated by IRAK4. The compounds of this disclosure or their pharmaceutical compositions are useful as therapeutic agents for treating inflammatory disorders such as rheumatoid arthritis (RA), inflammatory bowel disease (IBD), Crohn's disease, ulcerative colitis, necrotizing enterocolitis, gout, Lyme disease, arthritis, psoriasis, pelvic inflammatory disease, systemic lupus erythematosus (SLE), Sjögren's syndrome, inflammation associated with gastrointestinal infections including C. difficile, viral myocarditis, acute and chronic tissue injury, non-alcoholic steatohepatitis (NASH), alcoholic hepatitis, and renal diseases including chronic kidney disease and diabetic kidney disease.

Description

[Technical Field]

[0001] Cross-references to related applications This application claims priority to U.S. Provisional Application No. 62 / 978,635, filed February 19, 2020, which is incorporated herein by reference in its entirety.

[0002] background Technical field This invention provides a novel bifunctional compound for degrading interleukin-1 receptor-related kinase 4 (IRAK4) by proteolysis, and a method for treating diseases modulated by IRAK4. [Background technology]

[0003] Explanation of related technologies Interleukin-1 receptor-associated kinase-4 (IRAK4) is a serine / threonine kinase that plays a crucial role in immune cells in mediating Toll-like receptor (TLR) and interleukin-1 receptor (IL1R) signaling to produce pro-inflammatory cytokines. IRAK4 functions as part of the midosome, a large multiprotein complex that assembles in the plasma membrane when its ligand binds to TLR and IL1R receptors. The first step in midsome assembly is the recruitment of the scaffolding protein MyD88, followed by IRAK4 binding to MyD88 via homotypic death-domain (DD) interactions. Next, IRAK4 undergoes self-activation, subsequently phosphorylating the downstream kinases IRAK1 and IRAK2. Because IRAK4 is the most upstream kinase in this complex, it is considered a "major regulator" of midsome signaling. The importance of IRAK4 kinase function was demonstrated in IRAK4 kinase dead mice, which are resistant to TLR-induced septic shock because IRAK4 kinase is unable to produce pro-inflammatory cytokines.

[0004] IRAK4 also reportedly possesses kinase-independent scaffolding function. For example, macrophages derived from IRAK4 kinase-dead mice can still activate NF-κB signaling in response to IL1, TLR2, TLR4, and TLR7 stimulation. Similar scaffolding function has been demonstrated in human fibroblasts, where kinase-dead IRAK4 can restore IL-1-induced NF-κB signaling to levels comparable to that of WT IRAK4.

[0005] Therefore, IRAK4 can be targeted for degradation, thereby offering therapeutic opportunities in the treatment of autoimmune, inflammatory, and oncological diseases. Specific degradation of IRAK4 can be achieved by recruiting IRAK4 to ubiquitin ligases using heterobifunctional small molecules, thus promoting ubiquitination and proteasomal degradation of IRAK4. For example, thalidomide derivatives such as lenalidomide or pomalidomide have been reported to recruit potential protein substrates to cereblon (CRBN), a component of the ubiquitin ligase complex. See, for example, WO2019 / 099926, WO2020 / 023851 and U.S. Publication No. 2019 / 0192668. Further development of therapeutic agents targeting IRAK4 is needed. [Prior art documents] [Patent Documents]

[0006] [Patent Document 1] International Publication No. 2019 / 099926 [Patent Document 2] International Publication No. 2020 / 023851 [Patent Document 3] U.S. Patent Publication Application No. 2019 / 0192668 [Overview of the Initiative] [Means for solving the problem]

[0007] Brief Summary The bifunctional compound represented by formula (I) [Chemical formula] or a pharmaceutically acceptable salt, isotopic form, isolated stereoisomer or mixture of stereoisomers thereof, wherein R 1 is C a alkyl optionally substituted by 1 to 3 R 1~10 ; C a cycloalkyl optionally substituted by 1 to 3 R 3~10 ; or 3- to 12-membered heterocyclyl optionally substituted by 1 to 3 R a ; L is -L1-L2-L3-L4-L5-, and L1, L2, L3, L4 and L5 are each, independently: a) C b cycloalkyl optionally substituted by 1 to 3 R 3~12 ; b) C b aryl optionally substituted by 1 to 3 R 6~12 ; c) 3- to 12-membered heterocyclyl optionally substituted by 1 to 3 R b ; d) 5- to 12-membered heteroaryl optionally substituted by 1 to 3 R b ; e) a direct bond; f) C d alkylene chain optionally substituted by 1 to 3 R 1~12 ; g) C d alkenylene chain optionally substituted by 1 to 3 R 2~12 ; h) C d alkynylene chain optionally substituted by 1 to 3 R 2~12 ; i) one to six ethylene glycol units; j) 1 to 6 propylene glycol units; or k) -C(O)-, -C(O)O-, -O- 、 -N(R c )-, -S-, -C(S)-, -C(S)-O-, -S(O)2-, -S(O)=N-, -S(O)2NH-, -C(O)-N(R c )-, -C=N-, -OC(O)-N(R c )-, - or -OC(O)-O-; and; LHM is the ligature harness portion; R a Each of these independently consists of Halo, -CN, and 1-3 R's. d C is replaced as needed by 1~3 Alkyl, 1 to 3 R d C is replaced as needed by 3~6 Cycloalkyl or -OR c and; R b These are, independently, oxo, imino, sulfoxyimino, halo, nitro, -CN, and C. 1~6 Alkyl, C 2~6 Alkenil, C 2~6 Alkinyl, C 3~15 Cycloalkyl, C 1~8 Haloalkyl, C 6~12 Aryl, 5-12 member heteroaryl, 3-12 member heterocyclyl, -OR c , -C(O)-R c , -C(O)OR c -C(O)-N(R c )(R c ), -N(R c )(R c ), -N(R c )C(O)-R c , -N(R c )C(O)OR c , -N(R c )C(O)N(R c )(R c ), -N(R c )S(O)2(R c ), -NR c S(O)2N(Rc )(R c ), -N(R c )S(O)2O(R c ), -OC(O)R c -OC(O)-N(R c )(R c ), -Si(R c )3, -SR c ,-S(O)R c -S(O)(NH)R c -S(O)2R c or -S(O)2N(R c )(R c ) and C 1~6 Alkyl, C 2~6 Alkenil, C 2~6 Alkinyl, C 3~15 Cycloalkyl, C 1~8 Haloalkyl, C 6~12 Aryls, 5-12 member heteroaryls, and 3-12 member heterocyclines each have 1-3 R d It may be replaced as needed; R c Each of them is independently of hydrogen or C 1~6 It is alkyl; R d Each of these is independently substituted as needed with halo, oxo, -CN, -OH, and 1-3 fluorocarbons. 1~6 Alkyl, or C 3~8 -OC is substituted as needed with cycloalkyl or 1-3 fluorocarbons. 1~6 (It is alkyl.) However, this is provided herein.

[0008] In various further embodiments, LHM targets the E3 ligase VHL, CRBN, or IAP, which are utilized by bifunctional compounds to induce ubiquitination of IRAK4 and subsequent proteasomal degradation.

[0009] In more specific embodiments, LHM is represented by formulas (IIA), (IIB), (IIIA), (IIIB), (IIIC), (IIID), (IIIE), (IVA), (IVB), (IVC), or (IVD), or by their individual substructures.

[0010] In more specific embodiments, the bifunctional compounds are those described in Examples 1 to 192.

[0011] Further embodiments provide pharmaceutical compositions comprising one of the compounds of formula (I) or any substructure thereof, and a pharmaceutically acceptable carrier.

[0012] In one embodiment, the compound of formula (I) or a pharmaceutical composition thereof is useful as a therapeutic agent for treating cancers such as lymphoma, leukemia, acute myeloid leukemia (AML), and myelodysplastic syndrome (MDS).

[0013] In other embodiments, compounds of formula (I) or pharmaceutical compositions thereof are useful as therapeutic agents for treating metabolic disorders such as diabetes mellitus (type 1 and type 2 diabetes mellitus), metabolic syndrome, dyslipidemia, obesity, glucose intolerance, hypertension, elevated serum cholesterol, and elevated triglycerides.

[0014] In other embodiments, compounds of formula (I) or pharmaceutical compositions thereof are useful as therapeutic agents for treating inflammatory disorders such as rheumatoid arthritis (RA), inflammatory bowel disease (IBD), Crohn's disease, ulcerative colitis, necrotizing enterocolitis, gout, Lyme disease, arthritis, psoriasis, pelvic inflammatory disease, systemic lupus erythematosus (SLE), Sjögren's syndrome, inflammation associated with gastrointestinal infections including C. difficile, viral myocarditis, acute and chronic tissue injury, non-alcoholic steatohepatitis (NASH), alcoholic hepatitis, and renal diseases including chronic kidney disease and diabetic kidney disease. [Modes for carrying out the invention]

[0015] Detailed explanation Disclosed are bifunctional compounds capable of mobilizing IRAK4 to an E3 ubiquitin ligase for degradation, as well as methods for their preparation and use. In particular, the bifunctional compounds typically include an IRAK4 binding moiety conjugated by a covalent bond via a linker to a ligase harness moiety for targeting an ubiquitin ligase. Advantageously targeted degradation of IRAK4 enables effective treatment or amelioration of disease states, including IRAK4 function.

[0016] One embodiment is a bifunctional compound of formula (I)

Chemical formula

[0017] The IRAK4 binding portion of the bifunctional compound of formula (I) has the following structure, where the wavy line indicates the bond attached to the remainder of the compound of formula (I). [ka] (In the formula, R 1 This is 1 to 3 R's a C is replaced as needed by 1~10 Alkyl; 1 to 3 R a C is replaced as needed by 3~10 Cycloalkyl; or 1-3 R a (A 3- to 12-member heterocycline that is substituted as needed.) In a more specific embodiment, R 1 teeth a) C which is substituted as necessary with a halo, -OH, or -CN 1~5 alkyl; b) Hello, C 1~5 4- to 8-membered heterocyclines optionally substituted with alkyl, -OH, or -CN; or c) Hello, C 1~5 C is substituted as needed with alkyl, -OH, or -CN. 3~10 Cycloalkyl; That is the case.

[0018] In a more specific embodiment, R 1 These are oxetane, tetrahydrofuran, or tetrahydropyran, each being F, C 1~3 It may be substituted with alkyl, -OH, or -CN as needed.

[0019] In other, more specific embodiments, [ka] The part is one of the following structures (the wavy line indicates the bond to the thiadiazole portion): [ka] It holds. Ligauze harness portion (LHM)

[0020] Von Hipperrindou (VHL) and cereblon (CRBN) proteins are substrate-recognizing subunits of two eccentrically expressed, biologically important Cullin-type RING E3 ubiquitin ligase complexes. Furthermore, inhibitors of apoptosis proteins (IAPs) are a family of proteins involved in the suppression of apoptosis. The human IAP family includes eight members, and numerous other organisms contain IAP homologs. IAPs contain an E3 ligase-specific domain that recognizes substrates and promotes their ubiquitination, as well as a baculovirus IAP repeat (BIR) domain.

[0021] The LHM of the compound of formula (I) targets the VHL, CRBN, or IAP of the E3 ligase, which are utilized by the bifunctional compound to induce ubiquitination of IRAK4 and subsequent proteasomal degradation. A. LHM targeting CRBN

[0022] Thalidomide derivatives such as lenalidomide or pomalidomide can be used to recruit potential substrates to CRBN, a component of the ubiquitin ligase complex.

[0023] One embodiment has the following structure: [ka] (In the formula, W is -C(R g )- or -N-; Y is directly bonded, C 1~4 Alkylene chain, -C(O)-, -C(O)O-, -O-, -N(R g)-, -S--C(S)-, -C(S)-O-, -OC(O)O-, -C(O)-N(R g )-,-OC(O)-N(R g )-and; Ring B is C 6~12 Aryls, 5-12 membered heteroaryls, or 3-12 membered heterocyclines, each consisting of 1-3 R j It is replaced as needed, R j These are, independently, oxo, imino, sulfoxyimino, halo, nitro, -CN, and C. 1~6 Alkyl, C 2~6 Alkenil, C 2~6 Alkinyl, C 3~15 Cycloalkyl, C 1~8 Haloalkyl, C 6~12 Aryl, 5-12 member heteroaryl, 3-12 member heterocyclyl, -OR g , -C(O)-R g , -C(O)OR g -C(O)-N(R g )(R g ), -N(R g )(R g ), -N(R g )C(O)-R g , -N(R g )C(O)OR g , -N(R g )C(O)N(R g )(R g ), -N(R g )S(O)2(R g ), -NR g S(O)2N(R g )(R g ), -N(R g )S(O)2O(R g ), -OC(O)R g -OC(O)-N(R g )(R g ), -Si(R g )3, -SR g ,-S(O)R g -S(O)(NH)R g -S(O)2R g or -S(O)2N(Rg )(R g ) and C 1~6 Alkyl, C 2~6 Alkenil, C 2~6 Alkinyl, C 3~15 Cycloalkyl, C 1~8 Haloalkyl, C 6~12 Aryls, 5-12 member heteroaryls, and 3-12 member heterocyclines each have 1-3 R k It may be replaced as needed; R g is hydrogen or C 1~6 It is alkyl; R k Each of these is independently substituted as needed with halo, oxo, -CN, -OH, and 1-3 fluorocarbons. 1~6 Alkyl, or C 3~8 -OC is substituted as needed with cycloalkyl or 1-3 fluorocarbons. 1~6 (It is alkyl.) The present invention provides a CRBN-targeting LHM having the following characteristics (the dashed line indicates the bond attached to the remainder of the compound of formula (I)).

[0024] In certain specific embodiments, Y is a direct bond, and formula (IIA) has the following structure: [ka] (In the formula, W is -C(R g )- or -N-; Z1 is -C(O)--, -C(S)-, -C(NR g )-,-C(R g )2-, -N=, -N(R g )-,-C(R g )2-C(O)-,-C(O)-N(R g )-,-CR g =CR g -, -C(R g )2-C(S)-, -C(R g )=N- or -C(R g )2-C(R g )2-; Z2 is -C(O)--, -C(S)--, -C(NR g )-,-N(R g )-, -N= or -C(R g )2-; R g is hydrogen or C 1~6 It is alkyl; The E ring is a phenyl, a 5-6 membered heteroaryl, or a 5-6 membered heterocyclyl, each containing 1-3 R rings. j (This is replaced as needed.) It holds.

[0025] In a more specific embodiment, Z2 is -C(O)-, and formula (IIA1) has the following structure: [ka] (In the formula, W is -C(R g )- or -N-; Z1 is -C(O)--, -C(S)-, -C(NR g )-,-C(R g )2-, -C(R g )2-C(O)-,-C(O)-N(R g )-,-CR g =CR g -, -C(R g )=N-, -C(R g )2-C(S)- or -C(R g )2-C(R g )2-; q is 0, 1, or 2; R g is hydrogen or C 1~6 It is alkyl; R 2 C 1~6 alkyl, halo, halo C 1~6 Alkyl, -N(R g )2, CN, nitro, hydroxyl or -OC 1~4 (It is alkyl.) It holds.

[0026] In a more specific embodiment of equation (IIA1'), W is -CH-; Z1 is -C(O)-, -CH2-, -CH2-C(O)-, or -CH=CH-.

[0027] In a particular embodiment, formula (IIA1') is one of the following structures: [ka] It holds. In other embodiments, formula (IIA) has the following structure: [ka] (In the formula, W is -C(R g )- or -N-; Z3 is -C(O)--, -C(S)-, -C(NR g )-,-C(R g )2-, -N=, -N(R g )-,-C(R g )2-C(O)-,-C(O)-N(R g )-,-CR g =CR g -, -C(R g )2-C(S)-, -C(R g )=N-, -C(R g )2-C(R g )2-, -C(R g )2-O-, -C(R g ) 2-S-, -O-, or -S-; Z4 is -C(O)--, -C(S)--, -C(NR g )-,-N(R g )-, -N=, -O-, -S- or -C(R g )2-; R g is hydrogen or C 1~6 It is alkyl; The E ring is a phenyl, a 5-6 membered heteroaryl, or a 5-6 membered heterocyclyl, each containing 1-3 R rings. j (This is replaced as needed.) It holds.

[0028] In a more specific embodiment of formula (IIA2), W is -CtH-; Z3 is -C(R g )2-, -N(R g )-,-C(R g )2-C(O)-,-C(O)-N(R g )-,-CR g =CR g -, -C(R g )2-C(S)-, -C(R g )=N-, -C(R g )2-C(R g )2-, -C(R g )2-O or -C(R g )2-S-; Z4 is -C(O)--, -C(S)--, -C(NR g )- or -C(R g )2-.

[0029] In yet another, more specific embodiment, formula (IIA2) has the following structure: [ka] (wherein q is 0, 1 or 2; R g is hydrogen or C 1~6 Alkyl; R 2 C 1~6 alkyl, halo, halo C 1~6 Alkyl, -N(R g )2, CN, nitro, hydroxyl or -OC 1~4 (It is alkyl.) It holds.

[0030] In a more specific embodiment, formula (IIA2') has the following structure: [ka] It holds.

[0031] In a more specific embodiment of formula (IIA), W is -CH-; Y is directly bonded, C 1~4 Alkylene chain, -C(O)-, -C(O)O-, -O-, -N(R g )-, -S-, -C(S)-, -C(S)-O-, -OC(O)O-, -C(O)-N(R g )-,-OC(O)-N(R g )- and the B ring is a phenyl, a 5-6 membered heteroaryl, or a 5-6 membered heterocyclyl, each having 1-3 R j It is replaced as needed.

[0032] In a particular embodiment, formula (IIA) is one of the following structures: [ka] It holds.

[0033] In another embodiment, the LHM targeting CRBN has the following structure: [ka] (In the formula, W is -C(R g )- or -N-; The D ring is a phenyl, a 5-6 membered heteroaryl, or a 5-6 membered heterocyclyl, each containing 1-3 R rings. j It is replaced as needed, Ring B is C 6~12 Aryls, 5-12 membered heteroaryls, or 3-12 membered heterocyclines, each consisting of 1-3 R j It is replaced as needed, R g is hydrogen or C 1~6 It is alkyl; Rj These are, independently, oxo, imino, sulfoxyimino, halo, nitro, -CN, and C. 1~6 Alkyl, C 2~6 Alkenil, C 2~6 Alkinyl, C 3~15 Cycloalkyl, C 1~8 Haloalkyl, C 6~12 Aryl, 5-12 member heteroaryl, 3-12 member heterocyclyl, -OR g , -C(O)-R g , -C(O)OR g -C(O)-N(R g )(R g ), -N(R g )(R g ), -N(R g )C(O)-R g , -N(R g )C(O)OR g , -N(R g )C(O)N(R g )(R g ), -N(R g )S(O)2(R g ), -NR g S(O)2N(R g )(R g ), -N(R g )S(O)2O(R g ), -OC(O)R g -OC(O)-N(R g )(R g ), -Si(R g )3, -SR g ,-S(O)R g -S(O)(NH)R g -S(O)2R g or -S(O)2N(R g )(R g ) and C 1~6 Alkyl, C 2~6 Alkenil, C 2~6 Alkinyl, C 3~15 Cycloalkyl, C 1~8 Haloalkyl, C 6~12 Aryls, 5-12 member heteroaryls, and 3-12 member heterocyclines each have 1-3 R kIt may be replaced as needed; R k Each of these is independently substituted as needed with halo, oxo, -CN, -OH, and 1-3 fluorocarbons. 1~6 Alkyl, or C 3~8 -OC is substituted as needed with cycloalkyl or 1-3 fluorocarbons. 1~6 (It is alkyl.) It holds.

[0034] In a more specific embodiment, formula (IIB) has the following structure: [ka] (In the formula, Z5 is -C(O)-, -C(S)--, -C(NR g )-,-N(R g )-, -N= or -C(R g )2-; Z6 is -C(O)-, -C(S)-, -C(NR g )-,-C(R g )2-, -N=, -N(R g )-,-C(R g )2-C(O)-,-C(O)-N(R g )-,-CR g =CR g -, -C(R g )2-C(S)-, -C(R g )=N- or -C(R g )2-C(R g )2-; Z7 is -C(O)--, -C(S)--, -C(NR g )-,-N(R g )-, -O-, -S-, -N= or -C(R g )2-; R g is hydrogen or C 1~6 (It is alkyl.) It holds. In a more specific embodiment, formula (IIB1) has the following structure: [ka] It holds.

[0035] More specifically, equation (IB1') has the following structure: [ka] (wherein q is 0, 1 or 2; R 2 C 1~6 alkyl, halo, halo C 1~6 Alkyl, -N(R g )2, CN, nitro, hydroxyl or -OC 1~4 (It is alkyl.) It holds.

[0036] In a more specific embodiment, formula (IB1') has the following structure: [ka] It holds. B. LHM targeting VHL

[0037] In various embodiments, LHMs targeting von Hipperrindou (VHL) ligase have one of the following structures (the wavy lines indicate the bonds attached to the remainder of the compound of formula (I): [ka] (In the formula, V1 is -C(O)-, -C(O)O-, -C(O)O--C(R e )2-,-C(O)-N(R e )-,-C(O)--C(R e )2- or -C(O)-N(R e )-C(R e )2-; V2 is -C(O)-C(Re )2-; The G ring is a phenyl, a 5-6 membered heteroaryl, or a 5-6 membered heterocyclyl, each having 1-3 R rings. j It is replaced as needed; The J ring is a 5-12 member heteroaryl or 5-12 member heterocyclyl, each consisting of 1-3 R rings. j It is replaced as needed, R e Each of them is independently hydrogen and C 1~6 Alkyl or C 3~8 It is a cycloalkyl; R j These are, independently, oxo, imino, sulfoxyimino, halo, nitro, -CN, and C. 1~6 Alkyl, C 2~6 Alkenil, C 2~6 Alkinyl, C 3~15 Cycloalkyl, C 1~8 Haloalkyl, C 6~12 Aryl, 5-12 member heteroaryl, 3-12 member heterocyclyl, -OR g , -C(O)-R g , -C(O)OR g -C(O)-N(R g )(R g ), -N(R g )(R g ), -N(R g )C(O)-R g , -N(R g )C(O)OR g , -N(R g )C(O)N(R g )(R g ), -N(R g )S(O)2(R g ), -NR g S(O)2N(R g )(R g ), -N(R g )S(O)2O(R g ), -OC(O)R g -OC(O)-N(R g )(R g ), -Si(R g )3, -SRg ,-S(O)R g -S(O)(NH)R g -S(O)2R g or -S(O)2N(R g )(R g ) and C 1~6 Alkyl, C 2~6 Alkenil, C 2~6 Alkinyl, C 3~15 Cycloalkyl, C 1~8 Haloalkyl, C 6~12 Aryls, 5-12 member heteroaryls, and 3-12 member heterocyclines each have 1-3 R k It may be replaced as needed; R g Each of them is independently of hydrogen or C 1~6 It is alkyl; R k Each of these is independently substituted as needed with halo, oxo, -CN, -OH, and 1-3 fluorocarbons. 1~6 Alkyl, or C 3~8 -OC is substituted as needed with cycloalkyl or 1-3 fluorocarbons. 1~6 It is alkyl; R 3 is hydrogen or hydroxyl; R 4 is -C(O)R f And R f C 1~6 Alkyl or C 3~8 It has cycloalkyl elements, each of which is optionally substituted with a halo or -CN.

[0038] In a more specific embodiment, formulas (IIIA), (IIIB), (IIIC), (IIID), and (IIIE) have the structure of formulas (IIIA1), (IIIB1), (IIIC1), (IIID1), and (IIIE1), respectively: [ka] (In the formula, p is either 0 or 1; R j This is 1 to 3 R's k It is a 5-6 member heteroaryl that is substituted as needed; R k These are, independently, halo, oxo, -CN, -OH, and C. 1~6 Alkyl, C 3~8 Cycloalkyl or -OC 1~6 It is alkyl; R e Each of them is independently hydrogen and C 1~6 Alkyl or C 3~8 It is a cycloalkyl; R g Each of them is independently of hydrogen or C 1~6 It is alkyl; R 3 is hydrogen or hydroxyl; R 4 is -C(O)R f And R f C 1~6 Alkyl or C 3~8 It has cycloalkyl elements, each of which is optionally substituted with a halo or -CN.

[0039] In some more specific embodiments of any one of the formulas (IIIA1), (IIIB1), (IIIC1), (IIID1), or (IIIE1), p is 1, and R j These are thiazolyl, oxazolyl, isoxazolyl, pyrazolyl, imidazolyl, 1,2,4-oxadiazolyl, and 1,3,4-oxadiazolyl, and these are C 1~6 Alkyl, C 3~8 Substitutions may be made as needed with cycloalkyl, halo, CN, haloalkyl, or hydroxyalkyl elements.

[0040] In a preferred embodiment, R j This is a thiazolyl compound that is optionally substituted with an alkyl group (e.g., methyl group).

[0041] Therefore, a more specific embodiment of formula (IIIA) has the following structure: [ka] It holds.

[0042] A more specific embodiment of formula (IIIB) or (IIIB1) is one of the following structures: [ka] It holds.

[0043] A more specific embodiment of formula (IIIC) or (IIIC1) is one of the following structures: [ka] It holds.

[0044] A more specific embodiment of formula (IIID) or formula (IIID1) is one of the following structures: [ka] It holds.

[0045] A more specific embodiment of formula (IIIE) or (IIIE1) is one of the following structures: [ka] It holds.

[0046] In other embodiments, thiazolyl may be absent (i.e., p is 0). These thiazolyl-free (des-thiazolyl) LHMs are still sufficient to induce degradation. It can be joined to VHL. More specifically, formula (IIIA), (IIIB), (IIIC), or (IIID) is one of the following structures: [ka] It holds. C. LHM targeting IAP

[0047] In various embodiments, LHMs targeting von Hipperrindou (VHL) ligase have one of the following structures (the wavy lines indicate the bonds attached to the remainder of the compound of formula (I): [ka] (In the formula, R 5 Each of them is independently of hydrogen or C 1~6 It is alkyl; R 6 Each of them is independently of hydrogen or C 1~6 It is alkyl; R 7 Each of them is independently hydrogen and C 1~6 Alkyl or C 3~8 It is a cycloalkyl; R 8 Each is independently an aryl, a 5-12 membered cycloalkyl, a 5-12 membered heteroaryl, or a 5-12 membered heterocyclyl, each having 1-3 R j It is replaced as needed, R 9 Each of these is independently hydrogen, halo, or C. 1~6 It is alkyl; R j These are, independently, oxo, imino, sulfoxyimino, halo, nitro, -CN, and C. 1~6 Alkyl, C 2~6 Alkenil, C 2~6 Alkinyl, C 3~15 Cycloalkyl, C 1~8 Haloalkyl, C6~12 Aryl, 5-12 member heteroaryl, 3-12 member heterocyclyl, -OR g , -C(O)-R g , -C(O)OR g -C(O)-N(R g )(R g ), -N(R g )(R g ), -N(R g )C(O)-R g , -N(R g )C(O)OR g , -N(R g )C(O)N(R g )(R g ), -N(R g )S(O)2(R g ), -NR g S(O)2N(R g )(R g ), -N(R g )S(O)2O(R g ), -OC(O)R g -OC(O)-N(R g )(R g ), -Si(R g )3, -SR g ,-S(O)R g -S(O)(NH)R g -S(O)2R g or -S(O)2N(R g )(R g ) and C 1~6 Alkyl, C 2~6 Alkenil, C 2~6 Alkinyl, C 3~15 Cycloalkyl, C 1~8 Haloalkyl, C 6~12 Aryls, 5-12 member heteroaryls, and 3-12 member heterocyclines each have 1-3 R k It may be replaced as needed; R g Each of them is independently of hydrogen or C 1~6 It is alkyl; R k Each of these is independently substituted as needed with halo, oxo, -CN, -OH, and 1-3 fluorocarbons.1~6 Alkyl, or C 3~8 -OC is substituted as needed with cycloalkyl or 1-3 fluorocarbons. 1~6 It is alkyl; U1 is either a direct bond or a -C(O)-; Z is either -CH- or N; The K ring is either phenyl or naphthyl. It holds. More specific embodiments of formulas (IVA), (IVB), (IVC), and (IVD) have the following structures, respectively: [ka] It holds. Linker

[0048] The bifunctional compound of formula (I) includes a linker portion that connects the IRAK4 bond to the LHM. The structure of the linker portion (e.g., length or stiffness) can affect the efficiency or selectivity of the decomposition process. Typically, the linker portion includes multiple segments, which, in addition to providing individual bonding points to the IRAK4 bond and the LHM, contribute to the overall length and stiffness of the linker.

[0049] In a particular embodiment, the linker portion (L) of formula (I) is up to five linker segments (L s The compound of formula (I) has the following structure: (s is 1-5) [ka] (In the formula, L1, L2, L3, L4, and L5 are each independent of the following: a) 1 to 3 R's b C is replaced as needed by 3~10 Cycloalkyl; b) 1 to 3 R's b Arials that are replaced as needed; c) 1 to 3 R's b A 3- to 12-member heterocycline that is substituted as needed; d) 1 to 3 R's b A 5- to 12-member heteroaryl that is substituted as needed; e) direct binding; f) 1 to 3 R's d C is replaced as needed by 1~12 Alkylene chain; g) 1 to 3 R's d C is replaced as needed by 2~12 Alkenylene chain; h) R 1-3 d C is replaced as needed by 2~12 Alkynylene chain; i) 1 to 6 ethylene glycol units; j) 1 to 6 propylene glycol units; and k) -C(O)-, -C(O)O-, -O- 、 -N(R c )-, -S-, -C(S)-, -C(S)-O-, -S(O)2-, -S(O)=N-, -S(O)2NH-, -C(O)-N(R c )-, -C=N-, -OC(O)-N(R c )-, - or -OC(O)-O-; The bivalent part is selected from; R b These are, independently, oxo, imino, sulfoxyimino, halo, nitro, -CN, and C. 1~6 Alkyl, C 2~6 Alkenil, C 2~6 Alkinyl, C 3~15 Cycloalkyl, C 1~8 Haloalkyl, C 6~12 Aryl, 5-12 member heteroaryl, 3-12 member heterocyclyl, -OR c , -C(O)-R c , -C(O)OR c -C(O)-N(R c )(R c ), -N(R c )(R c ), -N(Rc )C(O)-R c , -N(R c )C(O)OR c , -N(R c )C(O)N(R c )(R c ), -N(R c )S(O)2(R c ), -NR c S(O)2N(R c )(R c ), -N(R c )S(O)2O(R c ), -OC(O)R c -OC(O)-N(R c )(R c ), -Si(R c )3, -SR c ,-S(O)R c -S(O)(NH)R c -S(O)2R c or -S(O)2N(R c )(R c ) and C 1~6 Alkyl, C 2~6 Alkenil, C 2~6 Alkinyl, C 3~15 Cycloalkyl, C 1~8 Haloalkyl, C 6~12 Aryls, 5-12 member heteroaryls, and 3-12 member heterocyclines each have 1-3 R d It may be replaced as needed; R c Each of them is independently of hydrogen or C 1~6 It is alkyl; R d Each of these is independently substituted as needed with halo, oxo, -CN, -OH, and 1-3 fluorocarbons. 1~6 C is substituted as needed with alkyl or 1-3 fluorocarbons. 3~8 (It is cycloalkyl) It holds.

[0050] Unless otherwise specified, and provided that the valency is satisfied, the divalent portion described herein (e.g., L or L)s It should be understood that these are not limited to the direction in which they are expressed. For example, in the case of a given linker segment, e.g., -C(O)-NH-, the way in which it is bonded to the rest of the molecule may be in either direction: i.e., -C(O)-NH or -NH-C(O)-, provided that the bond does not violate the rules of valency.

[0051] On the other hand, L is a series of L s When expressed by this, the directionality is in a manner consistent with the structure of formula (I'), and the specified L s The position may be established by the linker segment. For example, it should be understood that linker segment L1 is directly connected to the IRAK4 joint portion, while linker segment L5 is directly connected to the LHM.

[0052] One or more linker segments may be directly coupled. For example, in -L2-L3-L4--, if L3 is directly coupled, then L2 and L4 are directly coupled to each other, so L3 is essentially nonexistent.

[0053] In various specific embodiments, L1 is C 3~15 Cycloalkyl rings are selected from 6-15 membered aryl rings, 3-15 membered heterocyclyl rings, and 5-15 membered heteroaryl rings, each containing up to 3 R rings. d It may be further replaced by (as defined herein). In a more specific embodiment, L1 is C 3~12 Cycloalkyl rings are selected from 6-12 membered aryl rings, 3-12 membered heterocyclyl rings, and 5-12 membered heteroaryl rings, each of which has up to 3 R rings. d It may be further replaced by (as defined herein).

[0054] In various specific embodiments, L1 is one of the following ring portions: [ka] This may also be the case, and each ring may have 1 to 3 R d It may be replaced as needed by R d These are independently: halo, oxo, -CN, -OH, C 1~6 C is substituted as needed with alkyl and 1-3 fluorocarbons. 3~8 -OC is substituted as needed with cycloalkyl or 1-3 fluorocarbons. 1~6 It is alkyl.

[0055] In a more specific embodiment, L1 is one of the following structures: [ka] It holds.

[0056] In a preferred embodiment, L1 is one of the following structures: [ka] It holds.

[0057] In further embodiments, -L2-L3-L4-L5- generally have a linear structure (i.e., not a ring). More specifically, -L2-L3-L4-L5- are -C(O)-, -NH-C(O)-, -C(O)-(CH2) n -, -C(O)-(CH2) n -C(O)-, -C(O)-(CH2) n -O-, -(CH2) n -, -C(O)-(CH2) n -NH-, -C(O)-(CH2CH2O) m -, -C(O)-(CH2CH2O) m -(CH2) n -C(O)-, -C(O)-(CH2CH2O) m -(CH2) n -NH-, -C(O)-(CH2CH2O) m -(CH2) n -,-NH-C(O)-(CH2CH2O)m -(CH2) n -C(O)-, -NH-C(O)-(CH2CH2O) m -(CH2) n -NH-, -NH-C(O)-(CH2) n -C(O)-, -NH-C(O)-(CH2) n -,-NH-C(O)-(CH2CH2O) m -, -NH-C(O)-(CH2) n -O-, -NH-C(O)-(CH2) n -NH- or -NH-C(O)-(CH2CH2O) m -(CH2) n - can be such that m and n are independent integers from 1 to 12, and one or two hydrogen atoms in each of the above linker parts are C 1~3 It may be replaced by an alkyl group (e.g., methyl, ethyl, n-propyl, or isopropyl).

[0058] In a preferred embodiment, m is an integer between 1 and 10; and n is an integer between 1 and 10. In other embodiments, m is 1, 2, 3, 4, 5, or 6, and n is 1, 2, or 3. In various preferred embodiments, m is 1, 2, 3, 4, 5, or 6. In various preferred embodiments, n is 3, 4, 5, 6, 7, 8, 9, 10.

[0059] In a particular embodiment, L1 is [ka] And L has the following structure: [ka] It has the following characteristics. In a preferred embodiment, m is 1, 2, 3, 4, 5 or 6, and n is 1, 2, 3, 4, 5 or 6. In a more preferred embodiment, m is 1, 2 or 3, and n is 1 or 2.

[0060] In other embodiments, L1 is [ka] And L has the following structure: [ka] It has the following characteristics. In a preferred embodiment, m is 1, 2, 3, 4, 5, or 6, and n is 2, 4, or 6. In a more preferred embodiment, m is 1, 2, or 3, and n is 2.

[0061] In other embodiments, L1 is [ka] And L has the following structure: [ka] It has the following characteristics. In a preferred embodiment, m is 1, 2, 3, 4, 5, or 6, and n is 2, 4, or 6. In a more preferred embodiment, m is 1, 2, or 3, and n is 2.

[0062] In other embodiments, L1 is [ka] And L has the following structure: [ka] It has. In a preferred embodiment, m is 1, 2, 3, 4, 5, or 6. In a more preferred embodiment, m is 1, 2, or 3.

[0063] In other embodiments, L1 is [ka] And L has the following structure: [ka] It has the following characteristics. In a preferred embodiment, n is 1, 2, 3, 4, 5, 6, 7, or 8. In a more preferred embodiment, n is 2, 3, 4, or 5.

[0064] In other embodiments, L1 is [ka] And L has the following structure: [ka] It has the following characteristics. In a preferred embodiment, n is 1, 2, 3, 4, 5, or 6. In a more preferred embodiment, n is 1, 3, or 5.

[0065] In other embodiments, L1 is [ka] And L has the following structure: [ka] It has the following characteristics. In a preferred embodiment, n is 4, 5, 6, 7, or 8. In a more preferred embodiment, n is 5 or 7.

[0066] In other embodiments, L1 is [ka] And L is one of the following structures: [ka] (In the formula, R c is hydrogen or C 1~3 It has an alkyl group. In a preferred embodiment, n is 1, 2, 3, or 4. In a more preferred embodiment, n is 1 or 2.

[0067] In other embodiments, L1 is [ka] And L has the following structure: [ka] It has the following characteristics. In a preferred embodiment, n is 1, 2, 3, 4, 5, 6, 7, or 8. In a more preferred embodiment, n is 1, 5, or 7.

[0068] In other embodiments, L1 is [ka] And L has the following structure: [ka] It has the following characteristics. In a preferred embodiment, m is 1, 2, 3, 4, 5, or 6, and n is 2, 4, or 6. In a more preferred embodiment, m is 1, 2, or 3, and n is 2.

[0069] In other embodiments, L1 is [ka] And L has the following structure: [ka] It has the following characteristics. In a preferred embodiment, n is 1, 2, 3, 4, 5, or 6. In a more preferred embodiment, n is 3 or 4.

[0070] In other embodiments, L1 is [ka] And L has the following structure: [ka] It has the following characteristics. In a preferred embodiment, n is 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10. In a more preferred embodiment, n is 2, 3, 4, 5, 7, 7, 9 or 10.

[0071] In other embodiments, L1 is [ka] And L has the following structure: [ka] It has the following characteristics. In a preferred embodiment, m is 1, 2, 3, 4, 5, or 6, and n is 2, 4, or 6. In a more preferred embodiment, m is 1, 3, or 5, and n is 2.

[0072] In other embodiments, L1 is [ka] And L has the following structure: [ka] It has the following characteristics. In a preferred embodiment, n is 1, 2, 3, 4, 5, 6, 7, 8, or 9. In a more preferred embodiment, n is 1, 3, 5, 7, or 9.

[0073] In other embodiments, L1 is [ka] And L has the following structure: [ka] It has the following characteristics. In a preferred embodiment, m is 1, 2, 3, 4, 5, 6, 7, or 8. In a more preferred embodiment, m is 2, 4, or 6.

[0074] In other embodiments, L1 is [ka] And L is one of the following structures: [ka] It has the following characteristics. In a preferred embodiment, n is 1, 2, 3, 4, 5, 6, 7, or 8. In a more preferred embodiment, n is 2, 3, 4, or 5.

[0075] In an additional embodiment, L1 is [ka] And L is one of the following structures: [ka] The formula has (wherein n is 1, 2, or 3). In a preferred embodiment, n is 1.

[0076] In an additional embodiment, L1 is [ka] And L is one of the following structures: [ka] The formula has (wherein n is 1, 2, 3, 4, 5, 6, 7, 8, or 9). In a preferred embodiment, n is 1, 2, or 3. In a more preferred embodiment, n is 1.

[0077] In an additional embodiment, L1 is [ka] And L is one of the following structures: [ka] The formula has (wherein n is 1, 2, 3, 4, 5, 6, 7, 8, or 9). In a preferred embodiment, n is 1, 2, or 3. In a more preferred embodiment, n is 1.

[0078] In an additional embodiment, L1 is [ka] And L has the following structure: [ka] The formula has (wherein n is 1, 2, or 3). In a more preferred embodiment, n is 1.

[0079] In other embodiments, L1 is [ka] And L has the following structure: [ka] It has the following characteristics. In a preferred embodiment, n is 1, 2, or 3. In a more preferred embodiment, n is 1.

[0080] In further embodiments, L1 is a ring, and L2-L3-L4-L5- include at least one ring. The additional rings typically provide further rigidity to the linker portion. In certain embodiments, L1 is as follows: [ka] It is one of the following structures, and -L2-L3-L4-L5- is one of the following structures: [ka] [ka] It holds.

[0081] In a more specific embodiment, L1 is [ka] The linker (L) is one of the following structures: [ka] It holds.

[0082] In a more specific embodiment, L1 is [ka] The linker (L) is one of the following structures: [ka] (In the formula, R c is H or C 1~3 It has (being alkyl).

[0083] In a more specific embodiment, L1 is [ka] The linker (L) is one of the following structures: [ka] It holds.

[0084] In a more specific embodiment, L1 is [ka] The linker (L) is one of the following structures: [ka] It holds.

[0085] In a more specific embodiment, L1 is [ka] The linker (L) is one of the following structures: [ka] It holds.

[0086] In a more specific embodiment, L1 is [ka] The linker (L) is one of the following structures: [ka] It holds.

[0087] In a more specific embodiment, L1 is [ka] The linker (L) has the following structure: [ka] It holds.

[0088] In other embodiments, L1 is not a ring.

[0089] In other embodiments, the linker (L) or a part of the linker (-L1-L s -) is one of the following structures: [ka] [ka] [ka] [ka] It holds. Construction of the compound of formula (I)

[0090] The synthesis or construction of the compound of formula (I) can typically be carried out in several steps, including separately preparing the building blocks of the IRAK4 bond and the LHM moiety, and then linking the individual building blocks by covalent bond formation. Generally speaking, one or both of the building blocks may be linked with one or more linker precursors (L x The linker precursor may be prepared together with one or more linker segments (L s ) contains and has reactive terminal groups for further coupling. The two building blocks are coupled last (L s By forming segments, a compound of formula (I) can be obtained.

[0091] The following scheme demonstrates a general method for preparing building blocks. Examples 1-192 are specific examples of synthesized formula (I), characterized by their individual physicochemical properties. A. General scheme for preparing building blocks for IRAK4 joints [ka]

[0092] The compound of formula 1.5 can be obtained by following the method outlined in Scheme 1. 1-aminopyrrole 1.1 can be condensed with a suitable coupling partner using a suitable catalyst (e.g., HCl) and a suitable solvent (e.g., EtOH) to produce substituted pyrrolo[1,2-b]pyridazine 1.2. Intermediate 1.3 can be formed by halogenation at the indicated position using a known halogenating reagent (e.g., NBS), which can be further substituted by either CH activation or electrophilic aromatic substitution with a suitable reagent (e.g., selectfluor) to produce intermediate 1.4. Intermediate 1.5 can then be obtained by performing a halogen metal exchange of -X to -M using a suitable reagent (e.g., n-BuLi) or a transition metal coupling using a palladium catalyst and metal source (e.g., B2Pin2, Me6Sn2). [ka]

[0093] The compound of formula 2.3 can be obtained by following the method outlined in Scheme 2. Acid 2.1 can be converted to the corresponding acylhydrazine using a coupling reagent (e.g., HATU) in the presence of a base (e.g., DIPEA). Cyclization of compound 2.2 is achieved by heating in the presence of a thiolation reagent (e.g., Lawson's reagent) to obtain compound 2.3. [ka]

[0094] The compound of formula 3.6 can be obtained by following the method outlined in Scheme 3. Dihalopyridine 3.1 can be converted to compound 3.2 by the substitution of one halogen group (e.g., nucleophilic aromatic substitution). Compound 3.3 can be obtained by further functionalization of compound 3.2 using a metal-containing heterocyclic chemical species (e.g., compound 1.5) with a suitable catalyst such as a palladium catalyst. Intermediate 3.4 can be formed by halogenation at the indicated position using a known halogenating agent (e.g., NBS), and this intermediate can be further substituted by a cross-coupling reaction using a suitable catalyst such as a palladium catalyst to obtain compound 3.5. [ka]

[0095] Compound 4.2 can be constructed according to scheme A4. By replacing the halogen group of halothiadianazole 4.1 with a nucleophile (e.g., an amine) (e.g., nucleophilic aromatic substitution), compound 2.3 can be obtained. Intermediate 4.2 can be formed by halogenation at the indicated position using a known halogenating agent (e.g., NBS). [ka]

[0096] The compound of formula 3.5 can also be constructed according to scheme A5. Next, a halogen metal exchange of -X to -M can be achieved using a suitable reagent (e.g., n-BuLi) or a transition metal coupling reaction using a palladium catalyst and a metal source (e.g., B2Pin2, Me6Sn2) to obtain intermediate 5.1. Compound 5.1 can be functionalized using a cross-coupling reaction with compound 4.2 using a suitable catalyst such as a palladium catalyst to obtain compound 3.5.

[0097] Under Scheme A5, L xThis may be a ring having a reactive portion, which can then be linked to another linker segment. For example, BOC protection L x teeth, [ka] Compound 4.2 can be said to be, [ka] The resulting compound 3.5 is an IRAK4 bond building block, i.e., a piperazine ring, having an L1 precursor, which can be further linked to another linker segment by the reactive secondary amine piperazine. [ka]

[0098] An alternative method for obtaining the target compound 3.5 is shown in scheme A6. The corresponding acylhydrazine can be prepared starting from nicotinic acid 6.1 using a coupling reagent (e.g., HATU) in the presence of a base (e.g., DIPEA). Cyclization of compound 6.3 is achieved by heating in the presence of a thiolation reagent (e.g., Lawson's reagent) to obtain compound 6.4. Compound 3.5 can be obtained by further functionalization of compound 6.4 using a metal-containing heterocyclic chemical species (e.g., compound 1.5) with a suitable catalyst such as a palladium catalyst.

[0099] Under scheme A6, L x This may be a ring having a reactive part, which can then be linked to another linker segment. For example, L x teeth, [ka] (If necessary, it may be in a BOC-protected form during synthesis) and the resulting compound 3.5 is a building block of another IRAK4 bond having an L1 precursor, namely a bicyclo[2.2.2]octane ring, which can be further linked to another linker segment by a reactive primary amine.

[0100] A specific example of preparing the building blocks for the IRAK4 connection is described in more detail below.

[0101] BB1:7-(5-(5-(4-aminobicyclo[2.2.2]octan-1-yl)-1,3,4-thiadiazole-2-yl)-4-((tetrahydro-2H-pyran-4-yl)amino)pyridine-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonilicate hydrochloride. [ka]

[0102] Step 1: Methyl 6-chloro-4-((tetrahydro-2H-pyran-4-yl)amino)nicotinate. Methyl 4,6-dichloropyridine-3-carboxylate (4.00 g, 19.4 mmol) and tetrahydropyran-4-amine hydrochloride (4.01 g, 29.1 mmol) were dissolved in THF (20.0 mL), to which DIPEA (10.1 mL, 58.2 mmol) was added. This solution was stirred at 120°C for 12 hours and then concentrated. The crude product was purified by SiO2 chromatography (eluent: 20-100% siRNA / hexane) to obtain methyl 6-chloro-4-((tetrahydro-2H-pyran-4-yl)amino)nicotinate. ES / MS: 271.238(M+H + ).

[0103] Step 2: 6-Chloro-4-((tetrahydro-2H-pyran-4-yl)amino)nicotinohydrazide. A solution of methyl 6-chloro-4-((tetrahydro-2H-pyran-4-yl)amino)nicotinate (3.03 g, 11.2 mmol) and hydrazine hydrate (4.55 g, 90.9 mmol) in ethanol (18.0 mL) was stirred at 80°C for 3 hours and concentrated. This crude substance was carried over to the next step without further purification to obtain 6-chloro-4-((tetrahydro-2H-pyran-4-yl)amino)nicotinohydrazide. ES / MS: 271.201(M+H + ).

[0104] Step 3: tert-butyl(4-(2-(6-chloro-4-((tetrahydro-2H-pyran-4-yl)amino)nicotinoyl)hydrazine-1-carbonyl)bicyclo[2.2.2]octan-1-yl)carbamate. 6-chloro-4-((tetrahydro-2H-pyran-4-yl)amino)nicotinohydrazide (2.70g, 9.97mmol), 4-(tert-butoxycarbonylamino)bicycle[2.2.2] To a solution of octane-1-carboxylic acid (2.82 g, 10.5 mmol) and HATU (4.55 g, 12.0 mmol) in DMF (49.9 mL), DIPEA (5.70 mL, 31.9 mmol) was added. This solution was stirred at room temperature for 30 minutes and concentrated to dryness. Purification of the crude substance by SiO2 chromatography (eluent: 5-15% MeOH / CH2Cl2) yielded tert-butyl(4-(2-(6-chloro-4-((tetrahydro-2H-pyran-4-yl)amino)nicotinoyl)hydrazine-1-carbonyl)bicyclo[2.2.2]octane-1-yl)carbamate. ES / MS: 522.894(M+H + ).

[0105] Step 4: tert-butyl(4-(5-(6-chloro-4-((tetrahydro-2H-pyran-4-yl)amino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl)bicyclo[2.2.2]octan-1-yl)carbamate. A solution of tert-butyl(4-(2-(6-chloro-4-((tetrahydro-2H-pyran-4-yl)amino)nicotinoyl)hydrazine-1-carbonyl)bicyclo[2.2.2]octan-1-yl)carbamate (5.00 g, 9.58 mmol) in 2-MeTHF (47.9 mL) was heated to 65°C (ambient temperature). Lawson's reagent (4.26 g, 10.5 mmol) was then added, and the reaction mixture was stirred at 65°C for 12 hours. This solution was concentrated to dryness and purified by SiO2 chromatography (eluent: 50-100% siRNA / Hex). The product fractions were combined and stirred on 10% carbon-supported palladium (5g) for 1 hour. This slurry was filtered through Celite, washed with CH2Cl2, and the filtrate was concentrated to dryness. Purification of the residue by SiO2 chromatography (eluent: 1-5% MeOH / DCM) yielded tert-butyl(4-(5-(6-chloro-4-((tetrahydro-2H-pyran-4-yl)amino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl)bicyclo[2.2.2]octan-1-yl)carbamate. ES / MS: 520.288(M+H + ).

[0106] Step 5: tert-butyl(4-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-((tetrahydro-2H-pyran-4-yl)amino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl)bicyclo[2.2.2]octan-1-yl)carbamate. Sodium carbonate (2.00 M, 0.205 mL, 0.410 mmol) was added to a solution of tert-butyl(4-(5-(6-chloro-4-((tetrahydro-2H-pyran-4-yl)amino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl)bicyclo[2.2.2]octan-1-yl)carbamate (65.0 mg, 0.103 mmol), 7-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile (41.1 mg, 0.154 mmol), and [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium(II) (15.2 mg, 0.0205 mmol) in DME (2 mL). This solution was degassed with argon for 2 minutes and heated to 120°C (microwave) for 30 minutes. The resulting solution was diluted with THF, filtered, and concentrated to dryness. The crude solution was purified by preparative HPLC (Gemini C18, eluent: 10-65% acetonitrile / H2O / 0.1% TFA) and freeze-dried to obtain tert-butyl(4-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-((tetrahydro-2H-pyran-4-yl)amino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl)bicyclo[2.2.2]octan-1-yl)carbamate. ES / MS: 627.547(M+H + ).

[0107] Step 6: 7-(5-(5-(4-aminobicyclo[2.2.2]octan-1-yl)-1,3,4-thiadiazole-2-yl)-4-((tetrahydro-2H-pyran-4-yl)amino)pyridine-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonile bishydrochloride. tert-butyl(4-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazine-7-yl)-4-((tetrahydro-2H-pyran-4-yl)amino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl)bicyclo[2.2.2]octan-1-yl)carbamate (28 mg, 0.0378 mmol) in a solution of 1,2-dichloroethane (0.189 mL) in 4M dioxane. HCl (4.00 M, 0.09 mL, 0.0378 mmol) was added. This solution was stirred at room temperature for 1 hour and concentrated to dryness to obtain 7-(5-(5-(4-aminobicyclo[2.2.2]octan-1-yl)-1,3,4-thiadiazole-2-yl)-4-((tetrahydro-2H-pyran-4-yl)amino)pyridine-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitride bishydrochloride. ES / MS: 527.366(M+H + ). BB2:7-(5-(5-((trans)-4-aminocyclohexyl)-1,3,4-thiadiazole-2-yl)-4-(isopropylamino)pyridine-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonilicate hydrochloride [ka]

[0108] Step 1: tert-butyl((trans)-4-(2-(6-chloro-4-(isopropylamino)nicotinoyl)hydrazine-1-carbonyl)cyclohexyl)carbamate. 6-chloro-4-(isopropylamino)pyridine-3-carbohydrazide (500 mg, 2.19 mmol), 4-(tert-butoxycarbonylamino)cyclohexanecarboxylic acid (612 mg, 2.52 mmol), and HATU (915 mg, 2.41 mmol) were dissolved in DMF (9 mL) and DIPEA (0.750 mL, 4.31 mmol). This solution was stirred at room temperature for 2 hours and diluted with ELISA. Next, this solution was washed with a 1:1 mix of H2O:saturated aqueous solution NH4Cl, saturated aqueous solution NH4Cl, and brine. The organic layer was dehydrated with MgSO4 and concentrated to dryness. Purification of the crude substance by SiO2 chromatography (eluent: 2-5% MeOH / CH2Cl2) yielded tert-butyl((trans)-4-(2-(6-chloro-4-(isopropylamino)nicotinoyl)hydrazine-1-carbonyl)cyclohexyl)carbamate. ES / MS: 454.944(M+H + ).

[0109] Step 2: tert-butyl((trans)-4-(5-(6-chloro-4-(isopropylamino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl)cyclohexyl)carbamate. A solution of tert-butyl((trans)-4-(2-(6-chloro-4-(isopropylamino)nicotinoyl)hydrazine-1-carbonyl)cyclohexyl)carbamate (739 mg, 1.63 mmol) in THF (15 mL) was heated to 65°C (ambient temperature). Lawson's reagent (978 mg, 2.42 mmol) was then added, and the reaction mixture was stirred at 65°C for 1 hour. The solution was concentrated to dryness and purified by SiO2 chromatography (eluent: 5-35% ethyl acetate (5% MeOH) / Hex). The product fractions were combined and stirred on 10% carbon-supported palladium (1 g) for 1 hour. This slurry was filtered through Celite, washed with CH2Cl2, and the filtrate was concentrated to dryness. Purification of the residue by SiO2 chromatography (eluent: 10-40% acetone / Hex) yielded tert-butyl((trans)-4-(5-(6-chloro-4-(isopropylamino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl)cyclohexyl)carbamate. ES / MS: 452.725(M+H + ).

[0110] Step 3: tert-butyl((trans)-4-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(isopropylamino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl)cyclohexyl)carbamate. To a solution consisting of tert-butyl((trans)-4-(5-(6-chloro-4-(isopropylamino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl)cyclohexyl)carbamate (200 mg, 0.442 mmol), 7-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile (190 mg, 0.706 mmol), and XPhos Pd G3 (28.0 mg, 0.0331 mmol), tripotassium phosphate (2.00 M, 0.450 mL, 0.900 mmol) was added. This solution was degassed with argon for 2 minutes and heated to 120°C (microwave) for 20 minutes. Additional 7-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile (92.0 mg, 0.342 mmol) and XPhos Pd G3 (11.0 mg, 0.0130 mmol) were added, and the solution was heated to 120°C (microwave) for 20 minutes. The resulting solution was diluted with MeOH and concentrated to dryness. The residue was purified by SiO2 chromatography (eluent: 2-5% MeOH / CH2Cl2) to obtain tert-butyl((trans)-4-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazine-7-yl)-4-(isopropylamino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl)cyclohexyl)carbamate. ES / MS: 559.658 (M+H + ).

[0111] Step 4: 7-(5-(5-((trans)-4-aminocyclohexyl)-1,3,4-thiadiazole-2-yl)-4-(isopropylamino)pyridine-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonile bishydrochloride. To a solution of tert-butyl((trans)-4-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazine-7-yl)-4-(isopropylamino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl)cyclohexyl)carbamate (228 mg, 0.408 mmol) in a mixture of CH2Cl2 (4 mL) and MeOH (4 mL), 4M HCl (4.00 M, 2.00 mL, 8.00 mmol) in dioxane was added. This solution was stirred at 45°C for 18 hours and concentrated to dryness to obtain 7-(5-(5-((trans)-4-aminocyclohexyl)-1,3,4-thiadiazole-2-yl)-4-(isopropylamino)pyridine-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitride bishydrochloride. ES / MS: 459.629(M+H + ). BB3:7-(5-(5-((1r,4r)-4-aminocyclohexyl)-1,3,4-thiadiazole-2-yl)-4-(methylamino)pyridine-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile [ka]

[0112] Step 1: Methyl 6-chloro-4-(methylamino)nicotinate. To a solution of methyl 4,6-dichloronicotinate (95.0 g, 461 mmol, 1.00 equivalent) in acetonitrile (1000 mL), methaneamine (288 g, 2.32 mol, 25% purity, 5.03 equivalents) was slowly added at 0°C. The mixture was stirred at 0°C for 0.5 hours, then at 25°C for 2 hours. TLC (petroleum ether:ethyl acetate = 5:1) showed that 4,6-dichloronicotinate (Rf = 0.40) was consumed and a new spot (Rf = 0.30) was formed. The reaction mixture was concentrated under reduced pressure and extracted with ethyl acetate (3 × 500 mL). The combined organic layer was washed with brine (2 × 500 mL), dehydrated with Na₂SO₄, filtered, and the filtrate was concentrated under reduced pressure to obtain the crude product. The crude product was purified by column chromatography (SiO2, petroleum ether:ethyl acetate = 20:1~10:1, Rf = 30). Methyl 6-chloro-4-(methylamino)nicotinate (39.0 g, 184 mmol, 40.0% yield, 95.0% purity) was obtained as a white solid. LCMS: C8H9ClN2O2 Theoretical value (requires): 200.04, Measured value m / z = 201.1 (M+H) + . 1 H NMR: (400 MHz CDCl3) δ 8.65 (s, 1H), 8.08 (s, 1H), 6.54 (s, 1H), 3.88 (s, 3H), 2.92 (d, J = 5.2 Hz, 3H).BB3 is subsequently referred to as 6-chloro-4-(methylamino)pyridine-3-ka Starting with rubohydrazide, the synthesis was carried out using the same reaction sequence as BB2. LCMS:C 22 H 22 N8S Theoretical value: 430.17, Measured value: m / z = 431.39 [M+H] + . BB4:7-(4-(isopropylamino)-5-(5-(piperazin-1-yl)-1,3,4-thiadiazole-2-yl)pyridine-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile [ka]

[0113] Step 1: tert-butyl 4-(5-bromo-1,3,4-thiadiazole-2-yl)piperazine-1-carboxylate. Tert-butyl piperazine-1-carboxylate (1.6 g, 8.6 mmol, 1.05 equivalents) and dibromo-1,3,4-thiadiazole (2.0 g, 8.2 mmol) were combined in dioxane (0.15 M), and then N,N-diisopropylethylamine (2.5 mL, 14.4 mmol) was added. The vial was then stoppered and heated to 110°C for 90 minutes. The reaction mixture was then cooled to room temperature, concentrated on silica gel, and purified by column chromatography (0-5% methanol in DCM) to obtain tert-butyl 4-(5-bromo-1,3,4-thiadiazole-2-yl)piperazine-1-carboxylate (2.0 g, 70%). LCMS:C 11 H 17 BrN4O2S Theoretical value: 348.0, Measured value: m / z = 351.1 [M + H] + .

[0114] Step 2: tert-butyl 4-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(isopropylamino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl)piperazine-1-carboxylate. In a microwave vial, 6-{3-cyanopyrrolo[1,2-b]pyridazin-7-yl}-4-(isopropylamino)pyridine-3-ylboronic acid (1.5 g, 4.7 mmol), cesium carbonate (3.5 g, 10.7 mmol), xanthophos (0.54 g, 0.93 mmol), palladium acetate (105 mg, 0.47 mmol), and tert-butyl 4-(5-bromo-1,3,4-thiadiazole-2-yl)piperazine-1-carboxylate (1.6 g, 4.7 mmol) were combined in dioxane (0.15 M). Nitrogen was passed through the reaction mixture for 1 minute, and then the vial was stoppered. Microwave irradiation was carried out at 145°C for 35 minutes, and then the mixture was cooled to room temperature and filtered through Celite. The Celite pad was washed with ethyl acetate, and the combined organic matter was concentrated on silica gel. Chromatography (0-10% methanol in DCM) yielded tert-butyl 4-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(isopropylamino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl)piperazine-1-carboxylate, which was used directly in the next step.

[0115] Step 3: 7-[4-(isopropylamino)-5-[5-(piperazin-1-yl)-1,3,4-thiadiazole-2-yl]pyridine-2-yl]pyrrolo[1,2-b]pyridazin-3-carbonitride. The tert-butyl 4-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazole-2-yl)piperazin-1-carboxylate was stirred in the smallest amount of dioxane, then 4N dioxane (5 mL) was added and stirred for 5 hours. Next, the reaction product was concentrated on silica using a rotary evaporator and subjected to chromatography (0-20% methanol in DCM) to obtain 7-[4-(isopropylamino)-5-[5-(piperazin-1-yl)-1,3,4-thiadiazole-2-yl]pyridine-2-yl]pyrrolo[1,2-b]pyridazine-3-carbonitrile (0.5 g, total yield 24% in two steps). LCMS:C 22 H 23 N9S Theoretical value: 445.6, Measured value: m / z = 446.4 [M+H] + . BB5:7-[4-(isopropylamino)-5-{5-[4-(piperidine-4-carbonyl)piperazine-1-yl]-1,3,4-thiadiazole-2-yl}pyridine-2-yl]pyrrolo[1,2-b]pyridazine-3-carbonitrile [ka]

[0116] Step 1: 1-(5-bromo-1,3,4-thiadiazole-2-yl)-piperazine. Tert-butyl 4-(5-bromo-1,3,4-thiadiazole-2-yl)piperazine-1-carboxylate (1 g, 2.9 mmol) was dissolved in DCM (0.15 M), then trifluoroacetic acid (0.05 M volume) was added, and the mixture was stirred at room temperature for 3 hours. Next, the reaction product was concentrated, dissolved again in ether, concentrated, and dried under vacuum (0.6 g, 84%). Crude 1-(5-bromo-1,3,4-thiadiazole-2-yl)-piperazine was used directly in the next reaction. LCMS: C6H9BrN4S Theoretical value: 248.0, Measured value: m / z = 249.1 [M + H] + .

[0117] Step 2: tert-butyl 4-[4-(5-bromo-1,3,4-thiadiazole-2-yl)piperazine-1-carbonyl]piperidine-1-carboxylate. 1-(5-bromo-1,3,4-thiadiazole-2-yl)piperazine (300 mg, 1.2 mmol) was added to a solution of 1-(tert-butoxycarbonyl)piperidine-4-carboxylic acid (276 mg, 1.2 mmol) and HATU (570 mg, 1.5 mmol) in DMF (5 mL) and triethylamine (0.6 mL, 4.2 mmol). The reaction mixture was stirred at room temperature for 18 hours. Next, the reaction mixture was partitioned between ethyl acetate and water. The aqueous layer was re-extracted with ethyl acetate. The combined organic matter was washed with brine, then dehydrated with magnesium sulfate, and concentrated on silica gel. tert-butyl 4-[4-(5-bromo-1,3,4-thiadiazole-2-yl)piperazine-1-carbonyl]piperidine-1-carboxylate (0.2 g, 36%) was obtained by silica gel chromatography (0-10% methanol in DCM). LCMS:C 17 H 26 BrN5O3S Theoretical value: 460.4, Measured value: m / z = 484.3 [M + Na] + .

[0118] Step 3: 7-[4-(isopropylamino)-5-{5-[4-(piperidine-4-carbonyl)piperazine-1-yl]-1,3,4-thiadiazole-2-yl}pyridine-2-yl]pyrrolo[1,2-b]pyridazine-3-carbonitride. In a microwave vial, 6-{3-cyanopyrrolo[1,2-b]pyridazin-7-yl}-4-(isopropylamino)pyridine-3-ylboronic acid (150 mg, 0.47 mmol), cesium carbonate (0.42 g, 1.3 mmol), xanthophos (0.11 g, 0.19 mmol), palladium acetate (21 mg, 0.09 mmol), and tert-butyl 4-[4-(5-bromo-1,3,4-thiadiazole-2-yl)piperazine-1-carbonyl]piperidine-1-carboxylate were combined, then dioxane (0.1 M) was added, and N2 was passed through. Microwave irradiation was carried out in a microwave reactor at 145°C for 30 minutes, then cooled and filtered through Celite. The solution was concentrated on silica gel and then subjected to chromatography (0-10% methanol in DCM). Next, 4N HCl (0.15M) in dioxane was added to this substance, and then the mixture was stirred for 2 hours. The reaction product was then concentrated to obtain 7-[4-(isopropylamino)-5-{5-[4-(piperidine-4-carbonyl)piperazin-1-yl]-1,3,4-thiadiazole-2-yl}pyridine-2-yl]pyrrolo[1,2-b]pyridazine-3-carbonitrile (0.1g, 39% in total over both steps). LCMS:C 28 H 32 N 10 OS theoretical value: 556.7, measured value: m / z = 557.4 [M+H] + . BB6:7-[4-(isopropylamino)-5-{5-[4-(piperidine-4-yl)piperazine-1-yl]-1,3,4-thiadiazole-2-yl}pyridine-2-yl]pyrrolo[1,2-b]pyridazine-3-carbonitrile [ka]

[0119] Step 1: tert-butyl 4-[4-(5-bromo-1,3,4-thiadiazole-2-yl)piperazine-1-yl]piperidine-1-carboxylate. 1-(5-bromo-1,3,4-thiadiazole-2-yl)piperazine (300 mg, 1.2 mmol) and tert-butyl 4-oxopiperidine-1-carboxylate were combined in DCE (0.2 M) and TEA (0.5 mL, 3.6 mmol). After stirring for 5 minutes, sodium triacetoxyborohydride (0.45 g, 2.1 mmol) was added in one addition. The reaction mixture was stirred at room temperature for 3 hours, then filtered through Celite and concentrated on silica gel. Chromatography (0-10% methanol in DCM) yielded the desired tert-butyl 4-[4-(5-bromo-1,3,4-thiadiazole-2-yl)piperazine-1-yl]piperidine-1-carboxylate (0.2 g, 39%). LCMS:C 16 H 26 N5O2SBr Theoretical value: 432.4, Measured value: m / z = 456.3 [M + Na] + .

[0120] Step 2: tert-butyl 4-{4-[5-(6-{3-cyanopyrrolo[1,2-b]pyridazin-7-yl}-4-(isopropylamino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl]piperazine-1-yl}piperidine-1-carboxylate. In a microwave vial, 6-{3-cyanopyrrolo[1,2-b]pyridazin-7-yl}-4-(isopropylamino)pyridine-3-ylboronic acid (150 mg, 0.47 mmol), xanthophos (110 mg, 0.19 mmol), cesium carbonate (0.42 g, 1.28 mmol), palladium acetate (21 mg, 0.09 mmol), and tert-butyl 4-[4-(5-bromo-1,3,4-thiadiazole-2-yl)piperazine-1-yl]piperidine-1-carboxylate (200 mg, 0.47 mmol) were combined, and then dioxane (0.15 M) was added. After aerating the reaction mixture with nitrogen for 30 seconds, the vial was stoppered and microwaved in a microwave reactor at 145°C for 30 minutes. Next, the reaction product was cooled, filtered through Celite, and concentrated on silica gel. Chromatography (0-10% methanol in DCM) yielded tert-butyl 4-{4-[5-(6-{3-cyanopyrrolo[1,2-b]pyridazin-7-yl}-4-(isopropylamino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl]piperazine-1-yl}piperidine-1-carboxylate (100 mg, 34%). LCMS:C 32 H 40 N 10 O2S theoretical value: 626.8, measured value: m / z = 629.7 [M+H] + .

[0121] Step 3: 7-[4-(isopropylamino)-5-{5-[4-(piperidine-4-yl)piperazine-1-yl]-1,3,4-thiadiazole-2-yl}pyridine-2-yl]pyrrolo[1,2-b]pyridazine-3-carbonitride. tert-butyl4-{4-[5-(6-{3-cyanopyrrolo[1,2-b]pyridazine-7-yl}-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazole-2-yl]piperazine-1-yl}piperidine-1-carboxylate (100 mg) was dissolved in dioxane (1 mL), and then 4N HCl (1 mL) was added to the dioxane. The reaction mixture was stirred for 2 hours and then concentrated to obtain 7-[4-(isopropylamino)-5-{5-[4-(piperidine-4-yl)piperazine-1-yl]-1,3,4-thiadiazole-2-yl}pyridine-2-yl]pyrrolo[1,2-b]pyridazine-3-carbonitrile hydrochloride (0.08 g, 95%), which was used without further purification. LCMS:C 27 H 32 N 10 S Theoretical value: 528.7, Measured value: m / z = 529.7 [M + H] + . BB7:7-(5-(5-(4-aminopiperidine-1-yl)-1,3,4-thiadiazole-2-yl)-4-(isopropylamino)pyridine-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonilicate hydrochloride [ka]

[0122] Step 1: tert-butyl N-[1-(5-bromo-1,3,4-thiadiazole-2-yl)piperidine-4-yl]carbamate. Dibromo-1,3,4-thiadiazole (1.0 g, 4.1 mmol) and tert-butyl N-(piperidine-4-yl)carbamate (840 mg, 4.2 mmol) were dissolved in dioxane (0.15 M), and then N,N-diisopropylethylamine (1.25 mL, 7.2 mmol) was added. The reaction mixture was heated to 110°C in a sealed vial and stirred for 90 minutes. Next, the reaction mixture was cooled and concentrated on silica gel. Column chromatography (0-5% methanol in DCM) yielded tert-butyl N-[1-(5-bromo-1,3,4-thiadiazole-2-yl)piperidine-4-yl]carbamate (1.0 g, 67%). LCMS:C 12 H 19 BrN4O2S Theoretical value: 363.3, Measured value: m / z = 365.3 [M + H] + .

[0123] Step 2: tert-butyl N-{1-[5-(6-{3-cyanopyrrolo[1,2-b]pyridazin-7-yl}-4-(isopropylamino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl]piperidine-4-yl}carbamate. In a microwave vial, 6-{3-cyanopyrrolo[1,2-b]pyridazin-7-yl}-4-(isopropylamino)pyridine-3-ylboronic acid (90 mg, 0.28 mmol), cesium carbonate (0.25 g, 0.77 mmol), xanthophos (0.06 g, 0.11 mmol), palladium acetate (13 mg, 0.06 mmol), and tert-butyl N-[1-(5-bromo-1,3,4-thiadiazole-2-yl)piperidine-4-yl]carbamate (102 mg, 0.28 mmol) were combined, then dioxane (0.15 M) was added, and the mixture was aerated with nitrogen. The reaction mixture was stirred at room temperature for 3 minutes, and then microwaved at 145°C for 30 minutes. The reaction mixture was cooled, filtered through Celite, and concentrated on silica gel. Column chromatography (0-5% methanol in DCM) yielded tert-butyl N-{1-[5-(6-{3-cyanopyrrolo[1,2-b]pyridazin-7-yl}-4-(isopropylamino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl]piperidine-4-yl}carbamate (0.1 g, 64%).

[0124] Step 3: 7-(5-(5-(4-aminopiperidine-1-yl)-1,3,4-thiadiazole-2-yl)-4-(isopropylamino)pyridine-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonilicate hydrochloride. The title compound was obtained by adding excess 4N HCl to a solution of tert-butyl N-{1-[5-(6-{3-cyanopyrrolo[1,2-b]pyridazine-7-yl}-4-(isopropylamino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl]piperidine-4-yl}carbamate. LCMS:C 23 H 25 N9S Theoretical value: 459.2, Measured value: m / z = 460.5 [M+H] + . BB8:7-(5-(5-(3,9-diazaspiro[5.5]undecane-3-yl)-1,3,4-thiadiazole-2-yl)-4-(isopropylamino)pyridine-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile [ka]

[0125] BB8 was synthesized following the same route as BB4, except that tert-butyl 3,9-diazaspiro[5.5]undecane-3-carboxylate was used as the amine in step 1. LCMS:C 27 H 31 N9S Theoretical value: 513.2, Measured value: m / z = 514.6 [M+H] + . BB9:7-(4-(methylamino)-5-(5-(4-(piperidine-4-carbonyl)piperazine-1-yl)-1,3,4-thiadiazole-2-yl)pyridine-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile [ka]

[0126] BB9 was synthesized following the same route as BB5, except that (6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(methylamino)pyridine-2-yl)boronic acid was used in step 3. LCMS:C 26 H 28 N 10 OS theoretical value: 528.2, measured value: m / z = 529.4 [M+H] + . BB10:7-[4-(methylamino)-5-[5-(piperazin-1-yl)-1,3,4-thiadiazole-2-yl]pyridine-2-yl]pyrolo[1,2-b]pyridazine-3-carbonitrile [ka]

[0127] Step 1: 2-Bromo-N-methylpyridine-4-amine. To a mixture of 2-bromo-4-fluoropyridine (25.0 g, 0.142 mol, 1.0 equivalent), methylamine in methanol (9.8 M) (142 ml, 1.42 mol, 10 equivalents) was added, and the resulting mixture was heated overnight at 80°C. After completion, the reaction mixture was cooled, all volatiles were evaporated under vacuum, dissolved in ethyl acetate, and washed with water. The organic layer was dehydrated with sodium sulfate, filtered, and concentrated to obtain the desired product. (25 g, 89% yield): ESI(+)[M+H] + =188.94; 1 H NMR (300 MHz, DMSO-d6), δ: 7.77 (d, J = 5.8 Hz, 1H), 6.98 - 6.78 (m, 1H), 6.59 (m, 1H), 6.48 (m, 1H), 2.69 (d, J = 4.9 Hz, 3H).

[0128] Step 2: 7-[4-(methylamino)pyridine-2-yl]pyrrolo[1,2-b]pyridazine-3-carbonitrile. 2-bromo-N-methylpyridine-4-amine (6.0 g, 32.08 mmol, 1.0 equivalent), 7-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile (12.09 g, 44.91 mmol, 1.4 equivalent), and Xphos G3 (2.17 g, 2.57 mmol, 0.08 equivalent) were dissolved in anhydrous dimethoxyethane (80 ml, 0.4 M), to which an aqueous solution of 2 M K3PO4 (32.1 ml, 64.16 mmol, 2.0 equivalent) was added. This solution was degassed with argon for 15 minutes, and then heated overnight at 120°C with vigorous stirring. The reaction mixture was filtered through Celite, and the solvent was evaporated under reduced pressure to dryness. The resulting crude residue was purified by chromatography using methanol (0-10%) in dichloromethane to obtain the desired product as a yellow solid (6.1 g, 76% yield). 1 H NMR (300 MHz, DMSO-d6), δ: 8.79 (d, J = 2.2 Hz, 1H,), 8.64 (d, J = 2.2 Hz, 1H,), 8.19 (d, J = 5.6, 1H,), 7.87 (d, J = 2.3, 1H,), 7.76 (d, J = 4.7 Hz, 1H,), 7.08 (d, ESI(+)[M+H] + = 250.36.

[0129] Step 3: 7-[5-bromo-4-(methylamino)pyridine-2-yl]pyrrolo[1,2-b]pyridazine-3-carbonitrile and 7-[4-(methylamino)-2-pyridyl]pyrrolo[1,2-b]pyridazine-3-carbonitrile (5.3 g, 20.05 mmol, 1.0 equivalent) were dissolved in acetonitrile (65 ml, 0.3 M) and dichloromethane (20 ml, 0.7 M). At room temperature, N-bromosuccinimide (3.57 g, 20.05 mmol, 1.0 equivalent) was added in a single addition. The reaction mixture was stirred under ambient conditions for 30 minutes. After completion, the solvent was evaporated from the mixture under reduced pressure, and the resulting residue was purified by chromatography using 0-5% ethyl acetate in dichloromethane, yielding a yellow solid product (5.95 g, 88% yield). 1 1H NMR (300 MHz, DMSO-d6), δ: 8.80 (d, J = 2.2 Hz, 1H), 8.67 (d, J = 2.2 Hz, 1H), 8.36 (s, ESI(+)[M+H] + = 330.16.

[0130] Step 4: 4-(1,3,4-thiadiazole-2-yl)piperazine-1-carboxylate tert-butyl. N,N-diisopropylethylamine (29.57 ml, 169.68 mmol, 4.0 equivalents) was added to a solution of 2-bromo-1,3,4-thiadiazole (7.292 g, 42.424 mmol, 1.0 equivalent) and piperazine-1-carboxylate t-butyl hydrochloride (19.75 g, 106.05 mmol, 2.5 equivalents) in n-butanol (83.18 ml, 0.51 M). This reaction mixture was heated at 120°C for 1 hour. After completion, the resulting mixture was cooled and concentrated under vacuum to obtain the crude product. After chromatographic purification (0-70% ethyl acetate in hexane), the desired compound was obtained as a pink crystalline solid (9.93 g, 86% yield); 1 1H NMR (300 MHz, DMSO-d6), δ: 8.84 (s, 1H), 3.46 (s, 8H), 1.42 (s, 9H); ESI(+)[M+H] + = 272.16.

[0131] Step 5: tert-butyl 4-[5-(6-{3-cyanopyrrolo[1,2-b]pyridazin-7-yl}-4-(methylamino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl]piperazine-1-carboxylate. 7-[5-bromo-4-(methylamino)pyridine-2-yl]pyrrolo[1,2-b]pyridazin-3-carbonitrile (0.5g, 1.524 mmol, 1.0 equivalent), palladium(II) acetate (0.051g, 0.227 mmol, 0.15 equivalent), 4,5-bis(diphenylphosphin)-9,9-dimethylxanthene (0.26g, 0.45 mmol, 0.3 equivalent), cesium carbonate (0.1 g, 3.05 mmol, 2.0 equivalents) and cuprous iodide (0.087 g, 0.457 mmol, 0.3 equivalents) were dissolved in an oven-dried screw-cap vial. To this, 4-(1,3,4-thiadiazole-2-yl)piperazine-1-carboxylate tert-butyl (0.434 g, 1.53 mmol, 1 equivalent) and dioxane (25.39 ml, 0.06 M) were added. The reaction tube was evacuated and back-packed using argon for 20 minutes, sealed, and then heated overnight at 105°C. After completion of the reaction (confirmed by ULC), all volatiles were evaporated under vacuum, and the resulting residue was purified by chromatography (0-31% ethyl acetate in dichloromethane) to obtain the desired product as a yellow crystalline solid (0.57 g, 61% yield). 1 1H NMR (300 MHz, DMSO-d6), δ: 8.83 (s, 1H), 8.73 (s, 1H), 8.48 (m, 2H), 8.14 (s, 1H), 7.86 (s, 1H), 7.12 (d, J = 4.8 Hz, 1H), 3.54 (s, 8H), 3.06 (d, J = 4.9 Hz, 3H), 1.44 (s, 9H); ESI(+)[M+H] + = 518.64.

[0132] Step 6: A solution of tert-butyl 4-[5-(6-{3-cyanopyrrolo[1,2-b]pyridazin-7-yl}-4-(methylamino)-pyridizazole-2-yl]pyridine-2-yl]pyrrolo[1,2-b]pyridazin-3-carbonitride (0.25 g, 0.48 mmol, 1 equivalent) in 1,1,1,3,3,3-hexafluoro-2-propanol (0.769 ml, 7.25 mmol, 15 equivalents) was heated at 140°C for 3 hours using a MW. After evaporating all volatile components under reduced pressure, the remaining residue was purified by chromatography (0-7% methanol in dichloromethane), yielding the target product as a yellow solid (0.15 g, 73% yield): LC-MS:ESI(+)[M+H] + =418.06; 1 H NMR (300 MHz, DMSO-d6), δ: 8.83 (d, J = 2.2 Hz, 1H), 8.72 (s, 1H), 8.54 - 8.42 (m, 2H), 8.13 (s, 1H), 7.85 (d, J = 4.8 Hz, 1H), 7.11 (d, J = 4.8 Hz, 1H), 3.46 (s, 4H), 3.06 (d, J = 4.9 Hz, 3H), 2.84 (s, 4H), 2.61 (br m, 1H). BB11:7-(5-(5-(4-aminopiperidine-1-yl)-1,3,4-thiadiazole-2-yl)-4-(methylamino)pyridine-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile [ka]

[0133] BB11 was synthesized following the same route as BB7, except that (6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(methylamino)pyridine-2-yl)boronic acid was used in step 2. LCMS:C21 H 21 N9S Theoretical value: 431.2, Measured value: m / z = 432.4 [M+H] + . BB12:7-(4-(methylamino)-5-(5-(4-(piperidine-4-yl)piperazine-1-yl)-1,3,4-thiadiazole-2-yl)pyridine-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile [ka]

[0134] BB12 was synthesized following the same route as BB6, except that (6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(methylamino)pyridine-2-yl)boronic acid was used in step 2. LCMS:C 25 H 28 N 10 S Theoretical value: 500.2, Measured value: m / z = 501.5 [M + H] + . BB13:7-(5-(5-(4-aminobicyclo[2.2.2]octan-1-yl)-1,3,4-thiadiazole-2-yl)-4-(methylamino)pyridine-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile [ka]

[0135] BB13 was synthesized following the same route as BB3, except that 4-((tert-butoxycarbonyl)amino)bicyclo[2.2.2]octane-1-carboxylic acid was used in step 1. LCMS:C 24 H 24 N8S Theoretical value: 456.2, Measured value: m / z = 457.1 [M+H] + . BB14:7-(5-(5-((1s,4s)-4-aminocyclohexyl)-1,3,4-thiadiazole-2-yl)-4-(methylamino)pyridine-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile [ka]

[0136] BB14 was synthesized following the same route as BB3, except that cis-4-(tert-butoxycarbonylamino)cyclohexanecarboxylic acid was used in step 1. LCMS:C 22 H 22 N8S Theoretical value: 430.2, Measured value: m / z = 431.3 [M+H] + . BB15:7-(5-(5-(2,6-diazaspiro[3.5]nonane-6-yl)-1,3,4-thiadiazole-2-yl)-4-(methylamino)pyridine-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile [ka]

[0137] BB15 was synthesized following the same route as BB10, except that tert-butyl 2,6-diazaspiro[3.5]nonane-2-carboxylate was used as the amine in step 1. LCMS:C 23 H 23 N9S Theoretical value: 457.2, Measured value: m / z = 458.3 [M+H] + . BB16:7-(4-(methylamino)-5-(5-(4-(piperazine-1-carbonyl)piperazine-1-yl)-1,3,4-thiadiazole-2-yl)pyridine-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile [ka]

[0138] BB10 (55 mg, 0.22 mmol) was stirred with 4-(carboxy)piperazine-1-carboxylate tert-butyl (1 equivalent) in DIEA (2.2 equivalents) and DMF (0.2 M) at room temperature for 5 hours. Next, the reaction mixture was partitioned between ethyl acetate and water. The organic layer was separated, dehydrated with magnesium sulfate, and concentrated. This crude material was directly dissolved in DCM:TFA (4:1 ratio, 0.1 M) and stirred for 18 hours. Next, the reaction mixture was concentrated to dryness and powdered with diethyl ether to obtain the desired product (30 mg, 26% yield). LCMS:C 25 H 27 N 11 OS theoretical value: 529.6, measured value: m / z = 530.5 [M+H] + . BB17:7-(4-(methylamino)-5-(5-(4-(piperazine-1-carbonyl)piperidine-1-yl)-1,3,4-thiadiazole-2-yl)pyridine-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile [ka]

[0139] Step 1: Ethyl 1-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(methylamino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl)piperidine-4-carboxylate. In a microwave vial, 6-{3-cyanopyrrolo[1,2-b]pyridazin-7-yl}-4-(methylamino)pyridine-3-ylboronic acid (200 mg, 0.68 mmol) is mixed with cesium carbonate (2.75 equivalents), xanthophos (0.4 equivalents), and (acetyloxy) palagioacetate (palladio acetate). (0.2 equivalents) and ethyl 1-(5-bromo-1,3,4-thiadiazole-2-yl)piperidine-4-carboxylate (1 equivalent, see Step 1 of BB4) were combined, and then dioxane (8 mL) was added. Next, the reaction mixture was purged with N2 for 1 minute in a microwave reactor, stirred for 3 minutes, and then microwaved at 145°C for 30 minutes. Next, the reaction mixture was filtered through Celite and concentrated on silica gel. The desired product was obtained by chromatography (0-10% methanol in DCM).

[0140] Step 2: 1-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(methylamino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl)piperidine-4-carboxylic acid. Hydrolysis of the ester was carried out using THF / ethanol (10:1) and 2 mL of 2 M LiOH (aqueous solution). The reaction mixture was stirred for 3 hours, then dried on silica gel and subjected to chromatography (C18 column, 0-100% acetonitrile in water) to obtain the desired acid (100 mg, 32% in total over both steps).

[0141] Step 3: 7-(4-(methylamino)-5-(5-(4-(piperazine-1-carbonyl)piperidine-1-yl)-1,3,4-thiadiazole-2-yl)pyridine-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitride. The above carboxylic acid (100 mg) was combined with HATU (1.25 equivalents) and DIEA (5 equivalents) in DMF (0.1 M) and stirred for 10 minutes, then piperazine-1-carboxylic acid tert-butyl (1.2 equivalents) was added. The reaction mixture was stirred for 24 hours and then partitioned between ethyl acetate and water. The organic layer was separated and then redissolved in DCM / TFA (4:1, 0.1 M) and stirred overnight. After concentration, the crude product was obtained and used as is (50 mg, 44% yield): LCMS:C 26 H 28 N 10 OS theoretical value: 528.64, measured value: m / z = 529.6 [M+H] + . BB18:7-[4-(methylamino)-5-{5-[(1r,4r)-4-(methylamino)cyclohexyl]-1,3,4-thiadiazole-2-yl}pyridine-2-yl]pyrrolo[1,2-b]pyridazine-3-carbonitrile [ka]

[0142] Step 1: Methyl(1r,4r)-4-{[(tert-butoxy)carbonyl](methyl)amino}cyclohexane-1-carboxylate. Methyl trans-4-(tert-butoxycarbonylamino)cyclohexanecarboxylate (3.0 g, 11.658 mmol, 1.0 equivalent) was dissolved in DMF (20 ml, 0.6 M) and cooled to 0°C. Next, NaH (0.536 g, 13.99 mmol, 1.2 equivalents) was added, and the reaction mixture was stirred at 0°C for 30 minutes. Then, methyl iodide (1.09 ml, 17.49 mmol, 1.5 equivalents) was added, the cooling bath was removed, and the reaction mixture was stirred at room temperature for 18 hours. This mixture was poured into saturated aqueous ammonium chloride and extracted with ethyl acetate. Purification of the crude product with hexane:SiO yielded 1.4 g (44% yield) of the desired product; 1 H NMR (300 MHz, DMSO-d6) δ 3.58 (s, 3H), 2.64 (s, 3H), 2.25 (tt, J = 11.7, 3.6 Hz, 1H), 1.94 (dt, J = 12.3, 2.6 Hz, 2H), 1.60 - 1.41 (m, 4H), 1.38 (d, J = 1.6 Hz, 12H).

[0143] Step 2: (1r,4r)-4-{[(tert-butoxy)carbonyl]amino}cyclohexane-1-carboxylic acid. Methyl(1r,4r)-4-{[(tert-butoxy)carbonyl](methyl)amino}cyclohexane-1-carboxylate (1.3 g, 4.79 mmol, 1.0 equivalent) was dissolved in THF (18 ml, 0.27 M), then a solution of LiOH (4.8 ml, 4.79 mmol, 2.0 equivalent) was added, and the mixture was stirred at room temperature for 5 hours. TLC showed the residue of the starting material, and an additional volume of LiOH (2.4 ml, 2.39 mmol, 1.0 equivalent) was added, and the reaction mixture was stirred overnight. TLC showed complete conversion, and the mixture was quenched with a saturated solution of KHSO4 until pH < 5, and extracted by DCM to obtain 1.18 g (96% yield) of the desired product as free acid; 1 H NMR (300 MHz, DMSO-d6) δ 3.90- 3.52 (m, 1H) 2.65 (s, 3H), 2.13 (tt, J = 11.7, 3.6 Hz, 1H), 2.02 - 1.91 (m, 2H), 1.66 - 1.45 (m, 4H), 1.39 (d, J = 1.2 Hz, 11H).

[0144] Step 3: tert-butyl N-methyl-N-[(1r,4r)-4-{N'-[6-chloro-4-(methylamino)pyridine-3-carbonyl]hydrazinecarbonyl}cyclohexyl]carbamate. 6-chloro-4-(methylamino)pyridine-3-carbhydrazide (0.84 g, 0.7975 mmol, 1.0 equivalent) and (1r,4r)-4-{[(tert-butoxy)carbonyl]amino}cyclohexane-1-carboxylic acid (1.19 g, 4.61 mmol, 1.1 equivalent) were dissolved in DMF (10 mL), to which DIPEA (2.2 ml, 12.56 mmol, 3.0 equivalent) and HATU (1.91 g, 5.024 mmol, 1.2 equivalents) were added. This mixture was stirred at 25°C for 1 hour. UPLC showed the mass of the desired product. The reaction mixture was quenched with water and extracted with ethyl acetate. The combined organic layer was washed with water and brine, dehydrated with Na2SO4, and concentrated to obtain the crude product. Purification of the crude product by chromatography after elution with DCM:MeOH (0-10%) yielded 1.18 g (64% yield) of the desired product: ESI(+)[M+H] + =440.6; 1 H NMR (300 MHz, DMSO-d6) δ 10.30 (s, 1H), 9.78 (s, 1H),8.33 (s, 1H), 8.11 (s, 1H), 6.66 (s, 1H), 2.82 (d, 3H), 2.65 (s, 3H), 2.13 (tt, J = 11.7, 3.6 Hz, 1H), 2.02 - 1.91 (m, 2H), 1.66 - 1.45 (m, 4H), 1.39 (d, J = 1.2 Hz, 12H).

[0145] Step 4: tert-butyl N-methyl-N-[(1r,4r)-4-{5-[6-chloro-4-(methylamino)pyridine-3-yl]-1,3,4-thiadiazole-2-yl}cyclohexyl]carbamate. Lawson's reagent (1.20 g, 2.95 mmol, 1.1 equivalent) was added to a suspension of tert-butyl N-methyl-N-[(1r,4r)-4-{N'-[6-chloro-4-(methylamino)pyridine-3-carbonyl]hydrazinecarbonyl}cyclohexyl]carbamate (1.18 g, 2.68 mmol, 1.0 equivalent) in dry toluene (50 mL, 0.05 M). The reaction mixture was then stirred under reflux for 1.5 hours. The reaction mixture was then quenched with water, washed with a saturated solution of NaHCO3, extracted by DCM, and concentrated under reduced pressure. Purification of the crude product by flash column chromatography (DCM / MeOH) yielded 0.7 g (60% yield) of the desired product as a white solid: ESI(+)[M+H] + =438.6; 1 H NMR (300 MHz, DMSO-d6) δ 8.66 (d, J = 5.0 Hz, 1H), 8.40 (s, 1H), 6.83 (s, 1H), 3.25 - 3.09 (m, 1H), 2.98 (d, J = 4.9 Hz, 3H), 2.71 (s, 3H), 2.21 (d, J = 10.3 Hz, 2H), 1.69 (d, J = 7.4 Hz, 6H), 1.41 (s, 9H).

[0146] Step 5: tert-butyl N-methyl-N-[(1r,4r)-4-[5-(6-{3-cyanopyrrolo[1,2-b]pyridazin-7-yl}-4-(methylamino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl]cyclohexyl]carbamate. tert-butyl N-methyl-N-[(1r,4r)-4-{5-[6-chloro-4-(methylamino)pyridine-3-yl]-1,3,4-thiadiazole-2-yl}cyclohexyl]carbamate (0.7g, 1.6 mmol, 1.0 equivalent), 7-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile (0.6g, 2.23 mmol, 1.4 equivalents) and Pd(dppf)Cl 2· To a solution of CH2Cl2 (0.328 g, 0.4 mmol, 0.25 equivalents) in dioxane (30 ml), 2 M K2CO3 (1.6 ml, 3.2 mmol, 2.0 equivalents) was added. This solution was degassed with argon for 2-3 minutes, then heated to 120°C and stirred overnight. Complete conversion of the starting materials was demonstrated by UPLC. The resulting solution was diluted with MeOH, filtered through Celite, and concentrated to dryness. Purification of the crude product by chromatography eluted with DCM:MeOH (0-10%) yielded 0.7 g (80% yield) of the desired product: ESI(+)[M+H] + =546.1; 1 H NMR (300 MHz, DMSO-d6) δ 8.85 (d, J = 2.2 Hz, 1H), 8.74 (d, J = 2.2 Hz, 1H), 8.71 - 8.58 (m, 2H), 8.19 (s, 1H), 7.88 (d, J = 4.8 Hz, 1H), 7.13 (d, J = 4.8 Hz, 1H), 3.24 - 3.15 (m, 1H), 3.09 (d, J = 4.9 Hz, 3H), 2.71 (s, 3H), 2.30 - 2.12 (m, 2H), 1.76 - 1.53 (m, 6H), 1.42 (s, 9H).

[0147] Step 6: 7-[4-(methylamino)-5-{5-[(1r,4r)-4-(methylamino)cyclohexyl]-1,3,4-thiadiazole-2-yl}pyridine-2-yl]pyrrolo[1,2-b]pyridazine-3-carbonitride. In a sealed vial, tert-butyl N-methyl-N-[(1r,4r)-4-[5-(6-{3-cyanopyrrolo[1,2-b]pyridazin-7-yl}-4-(methylamino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl]cyclohexyl]carbamate (0.67 g, 1.23 mmol, 1.0 equivalent) was dissolved in hexafluoro-2-propanol (4.0 ml, 30.0 equivalents) and irradiated with microwaves at 150°C for 2 hours. UPLC demonstrated complete deprotection of the starting material. Upon evaporation of the solvent to dryness, 0.54 g of the desired product (99% yield) was obtained as a yellow solid:LCMS:ESI(+)[M+H] + =444.97; 1 H NMR (300 MHz, DMSO-d6) δ 8.84 (d, J = 2.3 Hz, 1H), 8.73 (d, J = 2.2 Hz, 1H), 8.68 - 8.47 (m, 2H), 8.18 (s, 1H), 7.88 (d, J = 4.8 Hz, 1H), 7.13 (d, J = 4.8 Hz, 1H), 3.20 - 3.10 (m, 1H), 3.09 (d, J = 4.8 Hz, 3H), 2.36 - 2.26 (m, 4H), 2.20 - 2.08 (m, 2H), 2.07 - 1.96 (m, 2H), 1.70 - 1.49 (m, 3H), 1.33 - 1.11 (m, 2H). BB19:7-[5-(5-{3,8-diazabicyclo[3.2.1]octan-3-yl}-1,3,4-thiadiazole-2-yl)-4-(methylamino)pyridine-2-yl]pyrrolo[1,2-b]pyridazine-3-carbonitrile [ka]

[0148] Step 1: tert-butyl 3-(5-bromo-1,3,4-thiadiazole-2-yl)-3,8-diazabicyclo[3.2.1]octane-8-carboxylate. A stirred suspension of 2,5-dibromo-1,3,4-thiadiazole (1.05 g, 4.305 mmol, 1.0 equivalent), 8-Boc-3,8-diazabicyclo[3.2.1]octane (1.005 g, 4.734 mmol, 1.1 equivalent), and N,N-diisopropylethylamine (1.125 ml, 6.459 mmol, 1.5 equivalent) in dioxane (21.53 ml, 0.2 M) was heated at 120°C for 1 hour. The reaction mixture was diluted with water and extracted by DCM, and the organic phase was concentrated on silica gel. Purification of the crude material by flash chromatography using a toluene / hexane gradient yielded the title compound as a yellow oily substance (0.819 g, 2.182 mmol, 71%): ESI(+)[M+H] + =337.3; 1 H NMR (300 MHz, Chloroform-d) δ 4.37 (s, 2H), 3.72 - 3.26 (m, 4H), 2.03 (m, 2H), 1.82 (m, 2H), 1.50 (d, J = 0.8 Hz, 9H).

[0149] Step 2: 7-[4-(methylamino)-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyrrolo[1,2-b]pyridazin-3-carbonitrile. In a sealed reactor, 7-[5-bromo-4-(methylamino)pyrridazin-2-yl]pyrrolo[1,2-b]pyridazin-3-carbonitrile (1.5 g, 4.57 mmol, 1.0 equivalent) was dissolved in dioxane (25 ml, 0.18 M), and then bis(pinacorato)diborone (1.39 g, 5.49 mmol, 1.2 equivalents) and KOAc (1.39 g, 14.17 mmol, 3.1 equivalents) were added. Argon was passed through the solution for several minutes, and Pd(dppf)Cl2 *DCM (0.373 g, 0.46 mmol, 0.1 equivalent) was added, and the mixture was aerated repeatedly. Next, the reaction mixture was transferred to a preheated oil bath and stirred at 90°C overnight. The reaction mixture was filtered through Celite and evaporated to dryness. The crude product was used in the next step without further purification.

[0150] Step 3: tert-butyl3-[5-(6-{3-cyanopyrrolo[1,2-b]pyridazin-7-yl}-4-(methylamino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl]-3,8-diazabicyclo[3.2.1]octane-8-carboxylate. To a solution of 7-[4-(methylamino)-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridine-2-yl]pyrrolo[1,2-b]pyridazine-3-carbonitrile (0.58 g, 1.55 mmol, 1.0 equivalent) and tert-butyl 3-(5-bromo-1,3,4-thiadiazole-2-yl)-3,8-diazabicyclo[3.2.1]octane-8-carboxylate (0.638 g, 1.70 mmol, 1.1 equivalent) in dioxane (40 ml, 0.04 M), Cs2CO3 (1.26 g, 3.86 mmol, 2.5 equivalents) and Pd(OAc)2 (0.069 g, 0.309 mmol, 0.2 equivalents) were added. The reaction was aerated with argon for several minutes, and then xanthophos (0.358 g, 0.618 mmol, 0.4 equivalents) was added. This solution was degassed with argon for 2-3 minutes, then heated to 120°C and stirred overnight. The reaction mixture was filtered through Celite and evaporated to dryness. The crude product was purified three times by chromatography using DCM:MeOH (0-10%) elution. The major fraction was re-purified with pTLC DCM:MeOH (0-10%), powdered with Et2O, and 0.190 g (23% yield) of the desired product: ESI(+)[M+H] + =544.77; 1 H NMR (300 MHz, DMSO-d6) δ: 8.83 (1 H, d, J 2.2), 8.72 (1 H, d, J 2.3), 8.51 - 8.43 (2 H, m), 8.14 (1 H, s), 7.85 (1 H, d, J 4.8), 7.12 (1 H, d, J 4.8), 4.28 (2 H, s), 3.67 (2 H, d, J 11.8), 3.37 (2 H, d), 3.06 (3 H, d, J 4.9), 1.91 (2 H, d, J 6.3), 1.75 (2 H, d, J 7.4), 1.44 (9 H, s).

[0151] Step 4: 7-[5-(5-{3,8-diazabicyclo[3.2.1]octane-3-yl}-1,3,4-thiadiazole-2-yl)-4-(methylamino)pyridine-2-yl]pyrrolo[1,2-b]pyridazine-3-carbonitride. In a pressure vessel, tert-butyl 3-[5-(6-{3-cyanopyrrolo[1,2-b]pyridazine-7-yl}-4-(methylamino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl]-3,8-diazabicyclo[3.2.1]octane-8-carboxylate (0.120 g, 0.22 mmol, 1.0 equivalent) was mixed with hexafluoro-2-propanol (0.7 ml) and irradiated in microwaves at 150°C for 2.5 hours. The solvent was evaporated to dryness, and the mixture was powdered using Et2O, yielding 0.080 g (82% yield) of the desired product: ESI(+)[M+H] + =444.05; 1 H NMR (300 MHz, DMSO-d6) δ: 8.83 (1 H, d, J 2.3), 8.72 (1 H, d, J 2.2), 8.52 - 8.44 (2 H, m), 8.13 (1 H, s), 7.85 (1 H, d, J 4.8), 7.12 (1 H, d, J 4.8), 3.61 - 3.46 (4 H, m), 3.29 (3 H, s), 3.06 (3 H, m,), 1.70 (4 H, (dd, J 9.8, 6.7). BB20:7-(4-(methylamino)-5-(5-(8-(piperidine-4-yl)-3,8-diazabicyclo[3.2.1]octan-3-yl)-1,3,4-thiadiazole-2-yl)pyridine-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile [ka]

[0152] Step 1: tert-butyl 4-(3-(5-bromo-1,3,4-thiadiazole-2-yl)-3,8-diazabicyclo[3.2.1]octane-8-yl)piperidine-1-carboxylate. 600 mg, 1.6 mmol of tert-butyl 3-(5-bromo-1,3,4-thiadiazole-2-yl)-3,8-diazabicyclo[3.2.1]octane-8-carboxylate (see Step 1 of BB19) was treated with 1:4 TFA / DC (0.1 M) for 2 hours, and then concentrated. The crude product was dissolved in DCE and TEA, and then 1 equivalent of tert-butyl 4-oxopiperidine-1-carboxylate was added. After 10 minutes, STAB (2.2 equivalents) was added, and the reaction mixture was stirred overnight. The reaction mixture was partitioned between DCM and water. The organic layer was separated, dehydrated with magnesium sulfate (mag sulfate), and then concentrated. Chromatography (0-10% methanol in DCM) yielded the desired product (500 mg, 68%). The completion of this synthetic route was carried out as previously described in steps 2 and 3 of BB6. LCMS:C 27 H 30 N 10 S Theoretical value: 526.2, Measured value: m / z = 527.6 [M + H] + . BB21:7-[5-(5-{2,7-diazaspiro[3.5]nonan-2-yl}-1,3,4-thiadiazole-2-yl)-4-(methylamino)pyridine-2-yl]pyrrolo[1,2-b]pyridazine-3-carbonitrile [ka]

[0153] Step 1: tert-butyl 2-(5-bromo-1,3,4-thiadiazole-2-yl)-2,7-diazaspiro[3.5]nonane-7-carboxylate. A stirred suspension of 2,5-dibromo-1,3,4-thiadiazole (970 mg, 3.98 mmol, 1.0 equivalent), tert-butyl 2,7-diazaspiro[3.5]nonane-7-carboxylate (990 mg, 4.37 mmol, 1.1 equivalent), and DIPEA (1.038 ml, 4.61 mmol, 1.5 equivalent) in dioxane (15 mL, 0.21 M) was heated at 120°C for 1 hour. The reaction mixture was diluted with water (10 mL) and extracted with DCM (20 mL). The crude substance was purified by flash chromatography eluted with hexane:SiO, yielding 1.54 g of a yellow oily substance (96% yield):ESI(+)[M+H] + =391.31 1 H NMR (300 MHz, DMSO-d6) δ 3.83 (s, 4H), 3.31 - 3.21 (m, 4H), 1.75 - 1.62 (m, 4H), 1.39 (s, 9H).

[0154] Step 2: 7-[4-(methylamino)-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile. In a pressure vessel, 7-[5-bromo-4-(methylamino)pyrridino-2-yl]pyrrolo[1,2-b]pyridazine-3-carbonitrile (1.5 g, 4.57 mmol, 1.0 equivalent) was dissolved in dioxane (25 ml), and then bis(pinacorato)diborone (1.39 g, 5.49 mmol, 1.2 equivalents) and KOAc (0.89 g, 9.14 mmol, 2.0 equivalents) were added. The solution was passed through with argon for 7 minutes, and Pd(dppf)Cl2 *DCM (0.375 g, 0.457 mmol, 0.1 equivalent) was added, and the mixture was aerated repeatedly. Next, the reaction mixture was transferred to a preheated oil bath and stirred at 90°C overnight. UPLC showed product formation. The reaction mixture was filtered through a Celite cake and evaporated to dryness. This crude product was used in the next step without further purification. ESI(+)[M+H] + =294.2

[0155] Step 3: tert-butyl2-[5-(6-{3-cyanopyrrolo[1,2-b]pyridazin-7-yl}-4-(methylamino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl]-2,7-diazaspiro[3.5]nonane-7-carboxylate. In a pressure vessel, tert-butyl 2-(5-bromo-1,3,4-thiadiazole-2-yl)-2,7-diazaspiro[3.5]nonane-7-carboxylate (1.377 g, 2.57 mmol, 1.0 equivalent) and 7-[4-(methylamino)-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridine-2-yl]pyrrolo[1,2-b]pyridazine-3-carbonitrile (1.0 g, 2.0 mmol, 1.0 equivalent) were dissolved in dioxane (13 ml, 0.2 M), to which cesium carbonate (2.09 g, 6.42 mmol, 2.5 equivalents) and palladium acetate (0.115 g, 0.51 mmol, 0.2 equivalents) were added. The reaction was aerated with argon for 7 minutes, and then xanthophos (0.59 g, 1.03 mmol, 0.4 equivalents) was added. This solution was degassed with argon for 2-3 minutes, then heated to 120°C and stirred overnight. UPLC showed product formation. The reaction mixture was filtered through a Celite cake and evaporated to dryness. Purification of the crude product by chromatography after elution with DCM:MeOH (0-10%) yielded 0.415 g (29% yield) of the desired product. ESI(+)[M+H] + =558.8; 1H NMR (300 MHz, DMSO-d6) δ 8.83 (d, J = 2.2 Hz, 1H), 8.73 (d, J = 2.2 Hz, 1H), 8.48 (s, 2H), 8.14 (s, 1H), 7.85 (d, J = 4.8 Hz, 1H), 7.12 (d, J = 4.8 Hz, 1H), 3.91 (s, 4H), 3.06 (d, J = 4.8 Hz, 3H), 1.75 (t, J = 5.6 Hz, 4H), 1.40 (s, 9H).

[0156] Step 4: 7-[5-(5-{2,7-diazaspiro[3.5]nonane-2-yl}-1,3,4-thiadiazole-2-yl)-4-(methylamino)pyridine-2-yl]pyrrolo[1,2-b]pyridazine-3-carbonitride. In a sealed reactor, tert-butyl 2-[5-(6-{3-cyanopyrrolo[1,2-b]pyridazine-7-yl}-4-(methylamino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl]-2,7-diazaspiro[3.5]nonane-7-carboxylate (0.2 g, 0.359 mmol, 1.0 equivalent) was dissolved in hexafluoro-2-propanol (1.13 mL, 30.0 equivalents) and heated in a microwave at 150°C for 2 hours. UPLC demonstrated complete deprotection of the starting material. The solvent was evaporated to dryness, and the solid was powdered with Et2O to obtain 146 mg (87% yield) of the desired product as a yellow solid. LCMS:ESI(+)[M+H] + =458.08; 1 H NMR (300 MHz, DMSO-d6) δ 8.83 (d, J = 2.2 Hz, 1H), 8.72 (d, J = 2.2 Hz, 1H), 8.47 (s, 2H), 8.13 (s, 1H), 7.85 (d, J = 4.8 Hz, 1H), 7.12 (d, J = 4.8 Hz, 1H), 3.86 (s, 4H), 3.06 (d, J = 4.8 Hz, 3H), 2.63 (t, J = 5.4 Hz, 4H), 1.87 - 1.60 (m, 4H). BB22:7-(5-(5-([4,4'-bipiperidine]-1-yl)-1,3,4-thiadiazole-2-yl)-4-(methylamino)pyridine-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile [ka]

[0157] BB22 was synthesized following the same route as BB10, except that tert-butyl[4,4'-bipiperidine]-1-carboxylate was used as the amine in step 1. LCMS:C 26 H 29 N9S Theoretical value: 499.2, Measured value: m / z = 500.4 [M+H] + . BB23:7-(4-(methylamino)-5-(5-(4-(piperazine-1-yl)piperidine-1-yl)-1,3,4-thiadiazole-2-yl)pyridine-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile [ka]

[0158] BB23 was synthesized following the same route as BB10, except that 4-(piperidine-4-yl)piperazine-1-carboxylate tert-butyl was used as the amine in step 1. LCMS:C 25 H 28 N 10 S Theoretical value: 500.2, Measured value: m / z = 501.4 [M + H] + . BB24:7-[4-(methylamino)-5-{5-[(1r,3r)-3-aminocyclobutyl]-1,3,4-thiadiazole-2-yl}pyridine-2-yl]pyrrolo[1,2-b]pyridazine-3-carbonitrile [ka]

[0159] Step 1: tert-butyl N-[3-({[6-chloro-4-(methylamino)pyridine-3-yl]formohydrazide}carbonyl)cyclobutyl]carbamate. A solution of 6-chloro-4-(methylamino)pyridine-3-carbohydrazide (3.0 g, 19.93 mmol, 1.0 equivalent) and trans-3-((tert-butoxycarbonyl)amino)cyclobutanecarboxylic acid (3.54 g, 16.48 mmol, 1.1 equivalents) in DMF (38 mL) was mixed with DIPEA (7.81 ml, 44.86 mmol, 3.0 equivalents) and HATU (6.82 g, 17.94 mmol, 1.2 equivalents). The mixture was stirred at 25°C for 1 hour. TLC (dichloromethane:methanol = 10:1) showed that the starting materials had been consumed and new spots had formed. The reaction mixture was quenched with water and extracted with ethyl acetate. The combined organic layer was washed with water and brine, dehydrated with Na2SO4, and concentrated to obtain the crude product. Purification of the crude product by chromatography using DCM:MeOH (0-10%) elution yielded 4.73 g (80% yield) of the desired product. ESI(+)[M+H] + =400.5; 1 H NMR (300 MHz, DMSO-d6) δ 10.32 (s, 1H), 9.78 (s, 1H), 8.34 (s, 1H), 8.10 (d, J = 5.1 Hz, 1H), 7.20 (d, J = 8.0 Hz, 1H), 6.67 (s, 1H), 4.13 (q, J = 7.9 Hz, 1H), 2.96 - 2.88 (m, 1H), 2.83 (d, J = 4.9 Hz, 3H), 2.40 - 2.28 (m, 2H), 2.20 - 2.08 (m, 2H), 1.37 (s, 9H).

[0160] Step 2: tert-butyl N-[(1r,3r)-3-{5-[6-chloro-4-(methylamino)pyridine-3-yl]-1,3,4-thiadiazole-2-yl}cyclobutyl]carbamate. To a solution of tert-butyl N-[3-({[6-chloro-4-(methylamino)pyridine-3-yl]formohydrazide}carbonyl)cyclobutyl]carbamate (4.73 g, 1 mmol, 1.0 equivalent) in toluene (94.0 mL, 0.13 M), Lawson's reagent (5.28 g, 13.08 mmol, 1.1 equivalent) was added, and the mixture was stirred at 90°C for 2 hours. The reaction mixture was washed with NaHCO3, extracted by DCM, concentrated, and purified by chromatography after elution with DCM:MeOH (0-10%) to obtain 3.9 g (50% yield) of the desired product with 60% purity. ESI(+)[M+H] + =298.5; 1 H NMR (300 MHz, DMSO-d6) δ 8.68 - 8.60 (m, 1H), 8.39 (s, 1H), 7.39 (d, J = 8.0 Hz, 1H), 6.83 (s, 1H), 4.36 - 4.16 (m, 1H), 2.99 (d, 3H), 2.59 - 2.51 (m, 4H), 1.38 (s, 9H).

[0161] Step 3: tert-butyl N-[(1r,3r)-3-[5-(6-{3-cyanopyrrolo[1,2-b]pyridazin-7-yl}-4-(methylamino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl]cyclobutyl]carbamate. In a pressure vessel, tert-butyl N-[(1r,3r)-3-{5-[6-chloro-4-(methylamino)pyridine-3-yl]-1,3,4-thiadiazole-2-yl}cyclobutyl]carbamate (1.0 g, 1.51 mmol, 1.0 equivalent), {3-cyanopyrrolo[1,2-b]pyridazine-7-yl}boronic acid (0.496 g, 2.12 mmol, 1.4 equivalent), and Pd(dppf)Cl2·CH2Cl2 (0.310 g, 0.379 mmol, 0.25 equivalent) were dissolved in anhydrous dioxane (19 ml, 0.08 M) and 2 M K2CO3 (1.51 ml, 3.03 mmol, 2.0 equivalent). This solution was degassed with argon for 2-3 minutes, then placed in an oil bath, heated to 120°C, and stirred overnight. LC-MS demonstrated complete conversion of the starting materials. The resulting solution was diluted with MeOH, washed through a Celite cake, and concentrated to dryness. Purification of the crude product by chromatography eluted with DCM:MeOH (0-10%), followed by pPTLC DCM:MeOH 4%, yielded 190 mg (25% yield) of the desired product. ESI(+)[M+H] + =503.8; 1 H NMR (300 MHz, DMSO-d6) δ 8.85 (d, J = 2.2 Hz, 1H), 8.75 (d, J = 2.3 Hz, 1H), 8.73 - 8.52 (m, 1H), 8.21 (s, 1H), 7.89 (d, J = 4.8 Hz, 1H), 7.40 (d, J = 8.0 Hz, 1H), 7.13 (d, J = 4.8 Hz, 1H), 4.35 - 4.23 (m, 1H), 4.00 - 3.87 (m, 1H), 3.10 (d, J = 4.9 Hz, 3H), 2.63 - 2.52 (m, 4H), 1.39 (s, 9H).

[0162] Step 4: 7-[4-(methylamino)-5-{5-[(1r,3r)-3-aminocyclobutyl]-1,3,4-thiadiazole-2-yl}pyridine-2-yl]pyrrolo[1,2-b]pyridazin-3-carbonitrile. In a sealed reactor, tert-butyl N-[(1r,3r)-3-[5-(6-{3-cyanopyrrolo[1,2-b]pyridazin-7-yl}-4-(methylamino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl]cyclobutyl]carbamate (0.17 g, 0.338 mmol, 1.0 equivalent) was dissolved in hexafluoro-2-propanol (1.1 ml, 30.0 equivalents) and microwaved at 150°C for 2 hours. LC-MS showed complete deprotection of the starting material. The solvent was evaporated to dryness, and the solid was powdered using Et2O, yielding 101 mg of the desired product (70% yield). LC-MS: ESI(+)[M+H] + =403.03;1H NMR (300 MHz, DMSO-d6) δ 8.83 (d, J = 2.3 Hz, 1H), 8.73 (d, J = 2.2 Hz, 1H), 8.67 - 8.56 (m, 2H), 8.18 (s, 1H), 7.87 (d, J = 4.8 Hz, 1H), 7.12 (d, J = 4.8 Hz, 1H), 3.89 (dq, J = 8.8, 4.3, 3.9 Hz, 1H), 3.64 (q, J = 7.4 Hz, 1H), 3.09 (d, J = 4.8 Hz, 3H), 2.59 - 2.52 (m, 2H), 2.34 - 2.18 (m, 2H). BB25:7-[4-(isopropylamino)-5-{5-[4-(piperidine-4-ylmethyl)piperazine-1-yl]-1,3,4-thiadiazole-2-yl}pyridine-2-yl]pyrrolo[1,2-b]pyridazine-3-carbonitrile [ka]

[0163] BB4 (105 mg, 0.23 mmol) and 4-formylpiperidine-1-carboxylate tert-butyl (50 mg, 0.23 mmol) were combined in DCE (0.1 M), and then TEA (5 equivalents) was added. After 5 minutes, STAB (124 mg, 2.5 equivalents) was added in one dose. After stirring overnight, the reaction mixture was partitioned between DCM and water. The organic layer was separated, dehydrated with magnesium sulfate, and concentrated. The crude material was treated with 4 M dioxane for 3 hours, and then concentrated by rotary evaporator. A yellow solid (50 mg, 39%) was obtained by reverse-phase ISCO (C18 column, 0-100% acetonitrile in water). LCMS:C 28 H 34 N 10 S Theoretical value: 542.3, Measured value: m / z = 543.5 [M + H] + . BB26:7-(5-(5-((1r,4r)-4-(ethylamino)cyclohexyl)-1,3,4-thiadiazole-2-yl)-4-(methylamino)pyridine-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile [ka] BB26 was synthesized in the same manner as BB18, by substituting methyl iodide with ethyl iodide in step 1, to obtain the title compound. LCMS:C 24 H 26 N8S Theoretical value: 458.2, Measured value: m / z = 459.0 [M+H] + B. General scheme for preparing LHM building blocks LHMs targeting CRBNs can generally be prepared in accordance with Scheme B1. [ka]

[0164] In Scheme B1, a functionalized thalidomide (e.g., at position 4 or 5 of the phthalimide ring) is first linked to a linker precursor. The linker precursor (aminoester) comprises "linker A" (representing one or more linker segments including L5) and two reactive end groups, an amine and a protected carboxylic acid in ester form. Step 1 describes the initial coupling step in more detail using an exemplary aminoester linker precursor.

[0165] Step 1: A mixture consisting of 2-(2,6-dioxopiperidine-3-yl)-4-fluoro-2,3-dihydro-1H-isoindole-1,3-dione (0.26 mmol), amino ester (0.26 mmol), ethylbis(propan-2-yl)amine (0.52 mmol), and DMF (1 mL) was stirred overnight at 90°C. The mixture was cooled and purified by HPLC (5-95% MeCN in H2O containing 0.1% TFA) to obtain a tert-butyl ester intermediate.

[0166] This tert-butyl ester intermediate is subsequently hydrolyzed (see step 2) to obtain a building block for CRBN-targeted LHM, having a carboxylic acid-terminated "linker A," which can be further linked to another part by reaction.

[0167] Step 2: A mixture consisting of tert-butyl 4-{[2-(2,6-dioxopiperidine-3-yl)-1,3-dioxo-2,3-dihydro-1H-isoindole-4-yl]aminobutanoate (0.10 mmol), CH2Cl2 (1 mL), and TFA (1 mL) was stirred at room temperature for 2 hours. When this mixture was concentrated, a carboxylic acid product was obtained.

[0168] The following are examples of additional building blocks for CRBN-targeting LHMs that can be prepared in accordance with Scheme B1.

[0169] HCB1:3-(2-((2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-4-yl)amino)ethoxy)propanoic acid. [ka]

[0170] Product of step 1: tert-butyl 3-[2-[[2-(2,6-dioxo-3-piperidyl)-1,3-dioxo-isoindorin-4-yl]amino]ethoxy]propanoate (1.8g, 51.9%). LCMS;C 22 H 27 N3O7 theoretical value: 445, measured value: m / z = 468 [M + Na] + .

[0171] Product of step 2: 3-[2-[[2-(2,6-dioxo-3-piperidyl)-1,3-dioxo-isoindoline-4-yl]amino]ethoxy]propanoic acid (526 mg, 32%). LCMS;C 18 H 19 Theoretical value for N3O7: 389, Measured value: m / z = 390 [M + H] + .

[0172] HCB2:3-(2-(2-(2-((2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-4-yl)amino)ethoxy)ethoxy)ethoxy)propanoic acid. [ka]

[0173] Product of step 1: tert-butyl 3-[2-[2-[2-[[2-(2,6-dioxo-3-piperidyl)-1,3-dioxoisoindorin-4-yl]amino]ethoxy]ethoxy]ethoxy]propanoate (1.6g, 41%). LCMS;C 26 H 35 N3O9 theoretical value: 533, measured value: m / z = 534 [M + H] + .

[0174] Product of step 2: 3-[2-[2-[2-[[2-(2,6-dioxo-3-piperidyl)-1,3-dioxo-isoindorin-4-yl]amino]ethoxy]ethoxy]ethoxy]propanoic acid (1.2g, 73.62%). LCMS;C 22 H 27 N3O9 theoretical value: 477, measured value: m / z = 478 [M + H] + . HCB3:6-((2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-5-yl)amino)hexanoic acid [ka] Step 1: tert-butyl6-{[2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindole-5-yl]amino}hexanoate

[0175] A mixture of 2-(2,6-dioxopiperidine-3-yl)-5-fluoroisoindole-1,3-dione (250 mg, 0.91 mmol) and tert-butyl 6-aminohexanoate hydrochloride (203 mg, 0.91 mmol) in NMP (3 ml) was mixed with N,N-diisopropylethylamine (0.6 mL) and heated overnight to 85°C. The crude reaction mixture was purified by silica gel chromatography using Â100% ethylhexanoate (0-100%) to obtain tert-butyl 6-{[2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindole-5-yl]amino}hexanoate (111 mg, 28%). LCMS:C 23 H 29 N3O6, theoretical value: 443.5, measured value: m / z = 444.4 [M+H] + . Step 2: 6-((2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-5-yl)amino)hexanoic acid

[0176] To a DCM solution of tert-butyl 6-{[2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindole-5-yl]amino}hexanoate (111 mg, 0.25 mmol), TFA (0.5 mL) was added. The reaction mixture was stirred at room temperature for 30 minutes, and then concentrated to obtain 6-((2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindolin-5-yl)amino)hexanoic acid (78 mg, 78%). 1 H NMR (500 MHz, DMSO-d6) δ 12.00 (s, 1H), 11.06 (s, 1H), 7.57 (d, J = 8.3 Hz, 1H), 7.11 (s, 1H), 6.95 (d, J = 2.1 Hz, 1H), 6.85 (dd, J = 8.4, 2.1 Hz, 1H), 5.04 (dd, J = 12.7, 5.4 Hz, 1H), 3.16 (q, J = 6.4 Hz, 2H), 2.23 (t, J = 7.4 Hz, 2H), 2.03 - 1.97 (m, 1H), 1.56 (dq, J = 14.8, 7.2 Hz, 4H), 1.39 (q, J = 7.9 Hz, 2H).LCMS:C 19 H 21 N3O6, theoretical value: 387.4, Measured value: m / z = 388.4 [M + H] + . HCB4:(1s,3s)-3-(2-((2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-4-yl)amino)ethoxy)cyclobutan-1-carboxylic acid [ka] [ka]

[0177] Step 1: Synthesis of cis-tert-butyl-3-(propa-2-ene-1-yloxy)cyclobutane-1-carboxylate: To a solution of cis-tert-butyl-3-hydroxycyclobutane-1-carboxylate (10.0 g, 58.06 mmol) in tetrahydrofuran (100 mL), t-BuOK (64 mL, 1 M in THF) was added dropwise at 0°C under nitrogen, and the mixture was stirred for 10 minutes. To the above solution, 3-bromopropa-1-ene (7.02 g, 58.03 mmol) was added dropwise at 0°C. The resulting mixture was stirred at room temperature for 16 hours. After the reaction was complete, the resulting solution was quenched by adding saturated aqueous NH4Cl. The aqueous phase was extracted with ethyl acetate. The combined organic layers were washed with brine, dehydrated with anhydrous sodium sulfate, filtered, and concentrated under vacuum. When the residue was purified by flash column chromatography using 0-20% ethyl acetate in petroleum ether, cis-tert-butyl-3-(propa-2-en-1-yloxy)cyclobutane-1-carboxylate (11.3 g, 92%) was obtained as a colorless oil. 1 H NMR (300 MHz, Chloroform-d) δ 6.10 - 5.85 (m, 1H), 5.33 - 5.10 (m, 2H), 3.95 - 3.75 (m, 3H), 2.60 - 2.36 (m, 3H), 2.29 - 2.07 (m, 2H), 1.44 (s, 9H).

[0178] Step 2: Synthesis of cis-tert-butyl-3-(2-oxoethoxy)cyclobutane-1-carboxylate: To a solution of cis-tert-butyl-3-(propa-2-en-1-yloxy)cyclobutane-1-carboxylate (1.0 g, 4.71 mmol) in dioxane (30 mL) and H2O (15 mL), K2OsO4,2H2O (86.28 mg, 0.24 mmol), 2,6-dimethylpyridine (1.01 g, 9.43 mmol), and NaIO4 (2.02 g, 9.42 mmol) were added. The resulting mixture was stirred at room temperature for 2 hours. After the reaction was complete, the mixture was diluted with water and extracted with ethyl acetate. The combined organic layers were washed with brine, dehydrated with anhydrous sodium sulfate, filtered, and concentrated under vacuum. When the residue was purified by flash column chromatography using 0-50% ethyl acetate in petroleum ether, cis-tert-butyl-3-(2-oxoethoxy)cyclobutane-1-carboxylate (505 mg, 50%) was obtained as a colorless oil. 1 H NMR (300 MHz, Chloroform-d) δ 9.72 (s, 1H), 6.97 (d, J = 7.8 Hz, 1H), 5.31 (s, 1H), 4.05 - 3.94 (m, 2H), 2.64 - 2.41 (m, 2H), 2.36 - 2.12 (m, 2H), 1.47 (s, 9H).

[0179] Step 3: Synthesis of cis-tert-butyl-3-[2-(benzylamino)ethoxy]cyclobutane-1-carboxylate: To a solution of cis-tert-butyl-3-(2-oxoethoxy)cyclobutane-1-carboxylate (2.0 g, 9.33 mmol) in methanol (20 mL), 1-phenylmethaneamine (3.0 g, 28.00 mmol) and NaBH3CN (1.76 g, 28.00 mmol) were added. The resulting solution was stirred at room temperature for 16 hours and then concentrated under vacuum. The residue was purified by flash column chromatography using 0-100% ethyl acetate in petroleum ether, yielding cis-tert-butyl-3-[2-(benzylamino)ethoxy]cyclobutane-1-carboxylate (1.1 g, 39%) as a colorless oil. MS (ESI) calculated value (C 18 H 27 NO3) [M+H] + , 306.2; measured value 306.1.

[0180] Step 4: Synthesis of cis-tert-butyl-3-(2-aminoethoxy)cyclobutane-1-carboxylate: To a solution of cis-tert-butyl-3-[2-(benzylamino)ethoxy]cyclobutane-1-carboxylate (2.0 g, 6.55 mmol) in methanol (20 mL), Pd / C (10%, 0.5 g) was added. The resulting solution was stirred under hydrogen (40 atm) at room temperature for 72 hours. After the reaction was complete, the solid was filtered off, and the filtrate was concentrated under vacuum to obtain cis-tert-butyl-3-(2-aminoethoxy)cyclobutane-1-carboxylate (1.0 g, crude) as a colorless oil, which was used in the next step without further purification. MS (ESI) calculated value (C 11 H 21 NO3) [M+H] + , 216.2; measured value 216.1.

[0181] Step 5: Synthesis of cis-tert-butyl-3-(2-[[2-(2,6-dioxopiperidine-3-yl)-1,3-dioxo-2,3-dihydro-1H-isoindole-4-yl]amino]ethoxy)cyclobutan-1-carboxylate: To a solution of cis-tert-butyl-3-(2-aminoethoxy)cyclobutan-1-carboxylate (1.0 g, 4.64 mmol) in N,N-dimethylformamide (10 mL), DIEA (6.0 g, 46.43 mmol) and 2-(2,6-dioxopiperidine-3-yl)-4-fluoro-2,3-dihydro-1H-isoindole-1,3-dione (6.72 g, 24.33 mmol) were added. The resulting solution was stirred at 90°C for 4 hours. After the reaction was complete, the resulting solution was diluted with water and extracted with ethyl acetate. The combined organic layers were washed with brine, dehydrated with anhydrous sodium sulfate, and concentrated under vacuum. Purification of the residue by reverse-phase flash column chromatography using 0-100% acetonitrile in water yielded cis-tert-butyl-3-(2-[[2-(2,6-dioxopiperidine-3-yl)-1,3-dioxo-2,3-dihydro-1H-isoindole-4-yl]amino]ethoxy)cyclobutane-1-carboxylate (250 mg, 11%) as a red solid. MS(ESI) calculated value (C 24 H 29 N3O7)[M+H] + , 472.2; measured value 472.1.

[0182] Step 6: Synthesis of cis-3-(2-[[2-(2,6-dioxopiperidine-3-yl)-1,3-dioxo-2,3-dihydro-1H-isoindole-4-yl]amino]ethoxy)cyclobutan-1-carboxylic acid: 850 mg, 1.8 mmol of cis-tert-butyl-3-(2-[[2-(2,6-dioxopiperidine-3-yl)-1,3-dioxo-2,3-dihydro-1H-isoindole-4-yl]amino]ethoxy)cyclobutan-1-carboxylate (850 mg, 1.8 mmol) was dissolved in 10 mL of dichloromethane, to which 10 mL of trifluoroacetic acid was added. The resulting solution was stirred at room temperature for 3 hours and then concentrated under vacuum. The residue was purified by reverse-phase flash column chromatography using 0-100% acetonitrile in water to obtain the crude product, which was then subjected to the following conditions [Column: Ultimate XB-NH2, 21.2 * Further purification by non-chiral preparative SFC using [250 mm; 5 μm; mobile phase A: CO2: 50, mobile phase B: MeOH -- preparative: 50; flow rate: 40 mL / min; 220 nm] yielded cis-3-(2-[[2-(2,6-dioxopiperidine-3-yl)-1,3-dioxo-2,3-dihydro-1H-isoindole-4-yl]amino]ethoxy)cyclobutan-1-carboxylic acid (359.1 mg, 37) as a yellow solid. MS (ESI) calculated value (C 20 H 21 N3O7)[M+H] + , 416.4; measured value 416.2. 1 1H NMR (400 MHz, DMSO-d6) δ 12.16 (s, 1H), 11.11 (s, 1H), 7.63 - 7.55 (m, 1H), 7.15 (d, J = 8.0 Hz, 1H), 7.05 (d, J = 8.0 Hz, 1H), 6.59 (t, J = 5.6 Hz, 1H), 5.12 - 5.03 (m, 1H), 3.96 - 3.84 (m, 1H), 3.59 - 3.40 (m, 4H), 2.98 - 2.82 (m, 1H), 2.64 - 2.52 (m, 3H), 2.48 - 2.37 (m, 2H), 2.08 - 1.91 (m, 3H).

[0183] Scheme B2 presents an alternative method for preparing building blocks of LHM targeting CRBN. [ka]

[0184] Step 1: 2-(2,6-dioxopiperidine-3-yl)-5-fluoroisoindorin-1,3-dione. A mixture of 5-fluoro-1,3-dihydro-2-benzofuran-1,3-dione (5.0 g, 30.10 mmol), 3-aminopiperidine-2,6-dione hydrochloride (6.9 g, 42.14 mmol), and NaOAc (4.2 g, 51.17 mmol) in HOAc (50 mL) was stirred at 120°C for 5 hours and then concentrated under vacuum. The residue was washed with water, and the solid was collected by filtration. The crude product was washed twice with water and twice with ethyl acetate, and dried in an oven to obtain 2-(2,6-dioxopiperidine-3-yl)-5-fluoroisoindorin-1,3-dione (7.7 g, 92%) as a light brown solid. 1 1H NMR (300 MHz, DMSO-d6) δ 11.16 (s, 1H), 8.03 - 8.00 (m, 1H), 7.87 - 7.85 (m, 1H), 7.75 - 7.70 (m, 1H), 5.19 - 5.15 (m, 1H), 2.94 - 2.86 (m, 1H), 2.63 - 2.48 (m, 2H), 2.12 - 2.06 (m, 1H).F NMR (300 MHz, DMSO-d6) δ -102.078.

[0185] Step 2: Amine substitution of aryl fluoride. To a solution of 2-(2,6-dioxopiperidine-3-yl)-5-fluoro-2,3-dihydro-1H-isoindole-1,3-dione (1.0 g, 3.62 mmol) in N-methylpyrrolidone (10 mL), amine (3.60 mmol) and DIEA (1.4 g, 10.83 mmol) were added. The resulting solution was stirred at 80°C for 16 hours. The reaction mixture was cooled to room temperature and purified by reverse-phase flash chromatography to obtain the corresponding final product.

[0186] Step 3: Oxidation of alcohol to aldehyde. Dess-Martin periodinane (2.12 mmol) was added to a mixture of alcohols (1.06 mmol) in CH2Cl2 (10 mL). This mixture was stirred at room temperature for 1 hour. The mixture was purified by column chromatography to obtain the desired aldehyde.

[0187] The following are examples of additional building blocks for CRBN-targeting LHMs that can be prepared in accordance with Scheme B2. HCB5:(3S)-1-(2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-5-yl)pyrrolidine-3-carbaldehyde [ka]

[0188] Step 1: 2-(2,6-dioxopiperidine-3-yl)-5-fluoroisoindoline-1,3-dione. This is the same as Step 1 of Scheme B2.

[0189] Step 2: 2-(2,6-dioxopiperidine-3-yl)-5-((S)-3-(hydroxymethyl)pyrrolidine-1-yl)isoindorin-1,3-dione. 2-(2,6-dioxopiperidine-3-yl)-5-fluoroisoindorin-1,3-dione was reacted with (S)-pyrrolidine-3-yl methanol to obtain 2-(2,6-dioxopiperidine-3-yl)-5-((S)-3-(hydroxymethyl)pyrrolidine-1-yl)isoindorin-1,3-dione (643.1 mg, 33%) as a yellow solid. 1 H NMR (300 MHz, DMSO-d6) δ 11.08 (s, 1H), 7.64 (d, J = 8.4 Hz, 1H), 6.89 (d, J = 2.1 Hz, 1H), 6.80 (dd, J = 8.4, 2.1 Hz, 1H), 5.06 (dd, J = 12.9, 5.4 Hz, 1H), 4.78 (t, J = 5.4 Hz, 1H), 3.59 - 3.41 (m, 5H), 3.22 - 3.17 (m, 1H), 2.95 - 2.83 (m, 1H), 2.67 - 2.44 (m, 3H), 2.12 - 1.88 (m, 2H), 1.87 - 1.76 (m, 1H). MS (ESI) calculated value (C 18 H 19 N3O5)[M+H] + , 358.1; measured value 358.1.

[0190] Step 3: (3S)-1-(2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindolin-5-yl)pyrrolidine-3-carboaldehyde. A mixture of 2-(2,6-dioxopiperidine-3-yl)-5-[(3S)-3-(hydroxymethyl)pyrrolidine-1-yl]isoindole-1,3-dione (258 mg, 0.72 mmol) in DCM (5 mL) is mixed with 1,1-bis(acetyloxy)-3-oxo-1λ 52-Benziodaoxol-1-yl acetate (0.61 g, 1.44 mmol) was added. After 90 minutes, silica gel was added, and the mixture was concentrated to dryness. The resulting powder was transferred to a loading cartridge, and the mixture was purified by flash chromatography on a 24 g column eluted with 0-100% ethyl acetate / hexane to obtain (3S)-1-[2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindole-5-yl]pyrrolidine-3-carboaldehyde (198 mg, 77%). LCMS C 18 H 17 N3O5 theoretical value: 355, measured value: m / z = 356 [M + H] + . HCB6:3-{4-[2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindole-5-yl]piperazine-1-yl}propanoic acid [ka]

[0191] Step 1: tert-butyl 3-(4-(2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindoline-5-yl)piperazine-1-yl)propanoate. 400.00 mg, 1.87 mmol of tert-butyl 3-(piperazine-1-yl)propanoate and 515.56 mg, 1.87 mmol of 2-(2,6-dioxopiperidine-3-yl)-5-fluoroisoindole-1,3-dione (515.56 mg, 1.87 mmol) were added to 10 ml of NMP, and N,N-diisopropylethylamine (0.65 mL, 0.48 g, 3.73 mmol) was heated at 85-90°C for 16 hours. The mixture was then partitioned between ethyl acetate and water (x2), and the organic layer was washed with brine, dried, and concentrated. 823 mg of the desired product was obtained by silica gel column purification using 10-100% HCl / hexane. LC-MS: C24H30N4O6, Theoretical value: 470.5, Measured value: m / z = 471.8 [M+H] + .

[0192] Step 2: 3-{4-[2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindole-5-yl]piperazine-1-yl}propanoic acid. Tert-butyl 3-{4-[2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindole-5-yl]piperazine-1-yl}propanoate (820.00 mg, 1.74 mmol) was dissolved in trifluoroacetic acid (9.94 g, 87.14 mmol), and the TFA was evaporated after 1 hour. The product was freeze-dried to dryness to obtain 3-{4-[2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindole-5-yl]piperazine-1-yl}propanoic acid (722 mg, 100% yield). LCMS: C20H22N4O6, Theoretical value: 414.4, Measured value: m / z = 415.4 [M+H] + . HCB7:2-(2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-5-yl)-2,7-diazaspiro[3.5]nonane-7-yl)acetic acid [ka]

[0193] Step 1: Benzyl 2-{2-[2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindole-5-yl]-2,7-diazaspiro[3.5]nonan-7-yl}acetate. A mixture of 2-(2,6-dioxopiperidine-3-yl)-5-fluoroisoindole-1,3-dione (70.00 mg, 0.25 mmol) and benzyl 2-{2,7-diazaspiro[3.5]nonanan-7-yl}acetate (69.53 mg, 0.25 mmol) in NMP (2 ml) was mixed with N,N-diisopropylethylamine (0.13 mL) and heated overnight at 85 °C. Purification of the crude mixture by column chromatography eluted with Â10-100% hexane yielded benzyl 2-{2-[2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindole-5-yl]-2,7-diazaspiro[3.5]nonane-7-yl}acetate (68 mg, 51%). LCMS C29 H 30 N4O6 theoretical value: 530, measured value: m / z = 532 [M + H] + .

[0194] Step 2: 2-(2-(2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindoline-5-yl)-2,7-diazaspiro[3.5]nonane-7-yl)acetic acid. To a solution of benzyl 2-{2-[2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindole-5-yl]-2,7-diazaspiro[3.5]nonane-7-yl}acetate (68.00 mg, 0.13 mmol) in EtOH (5 mL) and DCM (2 mL), carbon-supported palladium (6 mg, 0.06 mmol) was added. Hydrogen was blown into this reaction mixture and maintained under 1 atmosphere of hydrogen using a balloon, and stirred at room temperature for 48 hours. The reaction mixture was filtered through a Celite pad and concentrated to obtain benzyl 2-{2-[2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindole-5-yl]-2,7-diazaspiro[3.5]nonane-7-yl}acetate (56 mg, 99%). LCMS C 22 H 24 N4O6 theoretical value: 440, measured value: m / z = 441 [M + H] + . HCB8:2-(1-(2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-5-yl)piperidine-4-yl)acetaldehyde [ka]

[0195] Step 1: 2-(2,6-dioxopiperidine-3-yl)-5-fluoroisoindoline-1,3-dione. This is the same as Step 1 of Scheme B2.

[0196] Step 2: 2-(2,6-dioxopiperidine-3-yl)-5-(4-(2-hydroxyethyl)piperidine-1-yl)isoindorin-1,3-dione. 2-(2,6-dioxopiperidine-3-yl)-5-fluoroisoindorin-1,3-dione was reacted with 2-(piperidine-4-yl)ethane-1-ol to obtain 2-(2,6-dioxopiperidine-3-yl)-5-(4-(2-hydroxyethyl)piperidine-1-yl)isoindorin-1,3-dione (823 mg, 59%) as a yellow solid. 1 H NMR (300 MHz, DMSO-d6) δ11.09 (s, 1H), 7.65 (d, J = 8.4 Hz, 1H), 7.30 (d, J = 2.4 Hz, 1H), 7.23 (dd, J = 8.4, 2.4 Hz, 1H), 5.07 (dd, J = 12.6, 5.4 Hz, 1H), 4.40 (t, J = 5.1 Hz, 1H), 4.04 (d, J = 13.2 Hz, 2H), 3.64 - 3.40 (m, 2H), 3.09 - 2.79 (m, 3H), 2.70 - 2.51 (m, 2H), 2.07 - 1.94 (m, 1H), 1.77 - 1.66 (m, 3H), 1.41 - 1.34 (m, 2H), 1.24 - 1.12 (m, 2H).MS(ESI) calculation value (C 20 H 23 N3O5)[M+H] + 386.2; measured value 386.1.

[0197] Step 3: 2-(1-(2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-5-yl)piperidine-4-yl)acetaldehyde. Following Scheme B2, 2-(2,6-dioxopiperidine-3-yl)-5-(4-(2-hydroxyethyl)piperidine-1-yl)isoindorin-1,3-dione was oxidized to obtain the title compound. LCMS C 20 H 21 Theoretical value for N3O5: 383, Measured value: m / z = 384 [M + H] + . HCB9:1-(2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-5-yl)piperidine-4-carbaldehyde [ka]

[0198] Step 1: 2-(2,6-dioxopiperidine-3-yl)-5-fluoroisoindoline-1,3-dione. This is the same as Step 1 of Scheme B2.

[0199] Step 2: 2-(2,6-dioxopiperidine-3-yl)-5-(4-(hydroxymethyl)piperidine-1-yl)isoindorin-1,3-dione. 2-(2,6-dioxopiperidine-3-yl)-5-fluoroisoindorin-1,3-dione was reacted with piperidine-4-yl methanol to obtain 2-(2,6-dioxopiperidine-3-yl)-5-(4-(hydroxymethyl)piperidine-1-yl)isoindorin-1,3-dione (939 mg, 70%) as a yellow solid. 1 1H NMR (300 MHz, DMSO-d6) δ 11.09 (s, 1H), 7.65 (d, J = 8.4 Hz, 1H), 7.30 (d, J = 2.4 Hz, 1H), 7.23 (dd, J = 8.4, 2.4 Hz, 1H), 5.07 (dd, J = 12.6, 5.4 Hz, 1H), 4.51 (t, J = 5.1 Hz, 1H), 4.07 (d, J = 13.2 Hz, 2H), 3.27 (t, J = 5.7 Hz, 2H), 2.99 - 2.80 (m, 3H), 2.62 - 2.55 (m, 2H), 2.17 - 1.95 (m, 1H), 1.76 - 1.67 (m, 3H), 1.24 - 1.12 (m, 2H).MS(ESI) calculated value (C 19 H 21 N3O5)[M+H] + ,372. 1; Measured value: 372.2.

[0200] Step 3: 1-(2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-5-yl)piperidine-4-carboaldehyde. In accordance with Scheme B2, 2-(2,6-dioxopiperidine-3-yl)-5-(4-(hydroxymethyl)piperidine-1-yl)isoindorin-1,3-dione was oxidized to obtain 1-(2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-5-yl)piperidine-4-carboaldehyde. LCMS C 19 H 19 Theoretical value for N3O5: 369, Measured value: m / z = 370 [M + H] + . HCB10:1-[2-(2,6-dioxopiperidine-3-yl)-1-oxoisoquinoline-6-yl]piperidine-4-carbaldehyde [ka]

[0201] Step 1: Synthesis of 2-bromopentanedioic acid. To a solution of L-glutamic acid (100.0 g, 0.7 mol) and NaBr (244.7 g, 2.4 mol) in HBr (1 L, 40% in water), a solution of NaNO2 (84.4 g, 1.2 mol, in 200 mL of water) was added dropwise at 0°C under a nitrogen atmosphere. The resulting solution was stirred at 0-5°C for 2 hours. The reaction was quenched at 0°C by adding 30 mL of concentrated H2SO4 and stirred for 10 minutes. The resulting mixture was extracted with ethyl acetate. The combined organic layers were washed with brine, dehydrated with anhydrous Na2SO4, and filtered. The filtrate was concentrated under vacuum to obtain 2-bromopentanedioic acid (51 g, crude) as a light brown oily substance, which was used in the next step without further purification.

[0202] Step 2: Synthesis of dimethyl 2-bromopentanedioate. A solution of 2-bromopentanedioic acid (51.0 g, 241.69 mmol) in MeOH (500 mL) was mixed with concentrated H₂SO₄ (10 mL, 187.60 mmol). This mixture was stirred at 80°C for 3 hours and then concentrated under vacuum. The residue was diluted with water and extracted with ethyl acetate. The combined organic layers were washed with brine, dehydrated with anhydrous Na₂SO₄, and filtered. The filtrate was concentrated under vacuum. The residue was purified by flash column chromatography using 0-30% ethyl acetate in petroleum ether, yielding dimethyl 2-bromopentanedioate (36 g, 62%) as a pale yellow oil. 1H NMR (300 MHz, Chloroform-d) δ 4.38 (dd, J = 8.4, 5.7 Hz, 1H), 3.79 (s, 3H), 3.69 (s, 3H), 2.55 - 2.51 (m, 2H), 2.47 - 2.20 (m, 2H).

[0203] Step 3: Synthesis of 4-(hydroxymethyl)piperidine-1-carboxylate tert-butyl. A mixture of piperidine-4-ylmethanol (5.0 g, 43.41 mmol) and Et3N (5.3 g, 52.37 mmol) in THF (50 mL) was added dropwise at -5°C in a solution of Boc2O (10.4 g, 47.65 mmol in 10 mL of THF). The resulting mixture was warmed to room temperature and stirred for 16 hours. The solvent was removed under vacuum, and the residue was partitioned between ethyl acetate and water. The collected organic layer was washed with 5% aqueous HCl solution, water, and brine, dehydrated with anhydrous Na2SO4, and filtered. The filtrate was concentrated under vacuum. The crude product was powdered with hexane to obtain 4-(hydroxymethyl)piperidine-1-carboxylate tert-butyl (7.7 g, 82%) as a white solid. MS(ESI) calculated value (C11H21NO3)[M+H] + , 216.2; measured value 216.0.

[0204] Step 4: Synthesis of 4-(benzyloxymethyl)piperidine-1-carboxylate tert-butyl. To a mixture of NaH (42.0 g, 1021.86 mmol, 60%) in THF (500 mL), a solution of 4-(hydroxymethyl)piperidine-1-carboxylate tert-butyl (100.0 g, 464.483 mmol, in 500 mL of THF) was added dropwise at 0°C and the mixture was stirred at room temperature for 30 minutes under a nitrogen atmosphere. Next, benzyl bromide (174.8 g, 1021.88 mmol) was added dropwise to the above mixture at room temperature. The resulting solution was stirred at 80°C for 2 hours under a nitrogen atmosphere. The reaction mixture was quenched by carefully adding saturated aqueous NH4Cl. The aqueous layer was extracted with ethyl acetate. The combined organic layers were washed with brine, dehydrated with anhydrous Na2SO4, and filtered. The filtrate was concentrated under vacuum. Purification of the residue by flash column chromatography using 0-10% ethyl acetate in petroleum ether yielded 4-(benzyloxymethyl)piperidine-1-carboxylate tert-butyl (115.6 g, 81%) as a pale yellow oil. MS(ESI) calculation value (C18H27NO3)[M+H] + , 306.2; measured value 306.0.

[0205] Step 5: Synthesis of 4-((benzyloxy)methyl)piperidine. A solution of 4-(benzyloxymethyl)piperidine-1-carboxylate tert-butyl (94.0 g, 307.2 mmol) in HCl (4M in dioxane) (1000 mL) was stirred at room temperature for 2 hours. The solvent was removed under vacuum. The residue was partitioned between ethyl acetate and a 10% potassium carbonate aqueous solution. The collected organic layer was dehydrated with anhydrous Na₂SO₄ and filtered. The filtrate was concentrated under vacuum to obtain 4-[(benzyloxy)methyl]piperidine (54.0 g, 85%) as a yellow oily substance. MS(ESI) calculation value: (C13H19NO)[M+H] + , 206.2; measured value 206.2.

[0206] Step 6: Synthesis of 6-(4-(benzyloxymethyl)piperidine-1-yl)isoquinoline-1(2H)-one. To a degassed solution of 6-bromo-2H-isoquinoline-1-one (4.0 g, 17.85 mmol) in t-amyl alcohol (50 mL), 4-[(benzyloxy)methyl]piperidine (4.4 g, 21.42 mmol), t-BuONa (5.2 g, 53.91 mmol), and RuPhos-PdCl-2nd G (1.39 g, 1.78 mmol) were added. This mixture was stirred under a nitrogen atmosphere at 100°C for 3 hours. The reaction mixture was quenched by adding saturated citric acid aqueous solution and extracted with ethyl acetate. The combined organic layers were washed with brine, dehydrated with anhydrous Na2SO4, and filtered. The filtrate was concentrated under vacuum. Purification of the residue by flash column chromatography using 0-10% methanol in dichloromethane yielded 6-(4-(benzyloxymethyl)piperidine-1-yl)isoquinoline-1(2H)-one (5.5g, 88%) as a brown solid. MS(ESI) calculation value: (C22H24N2O2)[M+H] + , 349.2; measured value 349.2.

[0207] Step 7: Synthesis of dimethyl 2-(6-(4-(benzyloxymethyl)piperidine-1-yl)-1-oxoisoquinoline-2(1H)-yl)pentanedioate. To a solution of 6-(4-(benzyloxymethyl)piperidine-1-yl)isoquinoline-1(2H)-one (6.2 g, 17.79 mmol) in DMF (60 mL), dimethyl 2-bromopentanedioate (5.0 g, 20.91 mmol) and Cs2CO3 (17.4 g, 53.40 mmol) were added. The resulting mixture was stirred under a nitrogen atmosphere at 100°C for 16 hours. This mixture was diluted with saturated citric acid aqueous solution (aquous solution) and extracted with ethyl acetate. The combined organic layers were washed with brine, dehydrated with anhydrous Na2SO4, and filtered. When the filtrate was concentrated under vacuum, dimethyl 2-(6-(4-(benzyloxymethyl)piperidine-1-yl)-1-oxoisoquinoline-2(1H)-yl)pentanedioate (6g, crude) was obtained as a brown oily substance, which was used in the next step without further purification. MS(ESI) calculation value (C29H34N2O6)[M+H] + , 507.2; measured value 507.2.

[0208] Step 8: Synthesis of 2-(6-(4-(benzyloxymethyl)piperidine-1-yl)-1-oxoisoquinoline-2(1H)-yl)pentanedioic acid: To a solution of 1,5-dimethyl 2-(6-[4-[(benzyloxymethyl]piperidine-1-yl]-1-oxoisoquinoline-2-yl)pentanedioate (20.0 g, 39.48 mmol) in MeOH (80 mL), THF (80 mL), and H2O (80 mL), LiOH (5.67 g, 236.87 mmol) was added. This mixture was stirred at room temperature for 16 hours. The organic solvent was removed under vacuum, and the residue was diluted with water and extracted with ethyl acetate. The collected aqueous layer was acidified to pH 5-6 with citric acid and extracted with ethyl acetate. The combined organic layers were washed with brine, dehydrated with anhydrous Na2SO4, and filtered. When the filtrate was concentrated under vacuum, 2-(6-(4-(benzyloxymethyl)piperidine-1-yl)-1-oxoisoquinoline-2(1H)-yl)pentanedioic acid (15g, crude) was obtained as a brown oily substance, which was used in the next step without further purification. MS(ESI) calculation value (C27H30N2O6)[M+H] + , 479.2; measured value 479.0.

[0209] Step 9: Synthesis of 3-(6-(4-(benzyloxymethyl)piperidine-1-yl)-1-oxoisoquinoline-2(1H)-yl)piperidine-2,6-dione. Urea (2.0 g, 33.30 mmol) was added to a solution of 2-(6-(4-(benzyloxymethyl)piperidine-1-yl)-1-oxoisoquinoline-2(1H)-yl)pentanedioic acid (1.60 g, 3.34 mmol) in NMP (15 mL). The mixture was stirred at 180 °C for 4 hours under a nitrogen atmosphere. The resulting mixture was cooled to room temperature and diluted with water. This mixture was extracted with ethyl acetate. The combined organic layers were washed with brine, dehydrated with anhydrous Na₂SO₄, and filtered. The filtrate was concentrated under vacuum. Reverse-phase flash column chromatography using 5-65% acetonitrile in water purified the residue, yielding 3-(6-(4-(benzyloxymethyl)piperidine-1-yl)-1-oxoisoquinoline-2(1H)-yl)piperidine-2,6-dione (350 mg, 22%) as an off-white solid. MS(ESI) calculation value: (C27H29N3O4)[M+H] + , 460.2; measured value 460.2.

[0210] Step 10: Synthesis of 3-(6-(4-(hydroxymethyl)piperidine-1-yl)-1-oxoisoquinoline-2(1H)-yl)piperidine-2,6-dione. To a solution of 3-(6-(4-(benzyloxymethyl)piperidine-1-yl)-1-oxoisoquinoline-2(1H)-yl)piperidine-2,6-dione (1.8 g, 3.91 mmol) in THF (60 mL), Pd(OH)2 / C (10%, 1.8 g) and cyclohexene (3.2 g, 38.96 mmol) were added. The mixture was stirred under a nitrogen atmosphere at 80°C for 24 hours. Once the reaction was complete, the solid was filtered off and the filtrate was concentrated under vacuum. Purification of the residue by high-pressure flash column chromatography using the following conditions: [column, C18 silica gel; mobile phase, MeCN in water (0.1% NH4HCO3), gradient from 15% to 40% over 30 minutes; detector, UV 254 nm] yielded 3-(6-(4-(hydroxymethyl)piperidine-1-yl)-1-oxoisoquinoline-2(1H)-yl)piperidine-2,6-dione (800 mg, 55%) as an off-white solid. 1 H NMR (300 MHz, DMSO-d6) δ 10.72 (s, 1H), 7.94 (d, J = 9.0 Hz, 1H), 7.23 (d, J = 7.5 Hz, 1H), 7.14 (dd, J = 9.0, 2.4 Hz, 1H), 6.92 (d, J = 2.4 Hz, 1H), 6.44 (d, J = 7.5 Hz, 1H), 5.40 (s, 1H), 4.47 (t, J = 5.1 Hz, 1H), 3.97 - 3.94 (m, 2H), 3.27 - 3.24 (m, 2H), 2.85 - 2.78 (m, 3H), 2.58 - 2.52 (m, 2H), 1.98 - 1.94 (m, 1H), 1.81 - 1.67 (m, 2H), 1.65 - 1.59 (m, 1H), 1.26 - 1.12 (m, 2H).MS(ESI) calculated value (C20H23N3O4)[M+H] + 370.2; measured value, 370.3.

[0211] Step 11: 1-[2-(2,6-dioxopiperidine-3-yl)-1-oxoisoquinoline-6-yl]piperidine-4-carboaldehyde. Dissolve 3-{6-[4-(hydroxymethyl)piperidine-1-yl]-1-oxoisoquinoline-2-yl}piperidine-2,6-dione (150.00 mg, 0.41 mmol) in CH2Cl2 (2 mL) and prepare 1,1-bis(acetyloxy)-3-oxo-1λ 5 172 mg, 0.41 mmol of 2-benzoiodaoxol-1-yl acetate was added in a single addition at room temperature. After 5 hours, the reaction mixture was diluted with NaHCO3 (2 mL saturated aqueous solution), and Na2S2O3 (saturated aqueous solution) was added. The mixture was stirred for 30 minutes. The organic phase was removed. The aqueous layer was extracted (2 × 20 ml CH2Cl2), the combined organic phase was dried (Na2SO4), filtered, and concentrated. Purification by silica gel column chromatography (2-6% MeOH / CH2Cl2) yielded 1-[2-(2,6-dioxopiperidine-3-yl)-1-oxoisoquinoline-6-yl]piperidine-4-carboaldehyde (120 mg, 80%). LCMS C 20 H 21 Theoretical value for N3O4: 367.2, Measured value: m / z = 368.4 [M + H] + . HCB11:rac-(R)-1-(4-(1-(2,6-dioxopiperidine-3-yl)-4-methyl-5-oxo-4,5-dihydro-1H-1,2,4-triazole-3-yl)phenyl)piperidine-4-carboaldehyde [ka]

[0212] Step 1: Synthesis of 4-(4-(hydroxymethyl)piperidine-1-yl)methyl benzoate. To a solution of methyl 4-fluorobenzoate (25.0 g, 162.190 mmol) in DMF (250 mL), piperidine-4-yl methanol (18.6 g, 162.18 mmol) and K2CO3 (44.8 g, 324.38 mmol) were added. The resulting solution was stirred at 120°C for 16 hours. Once the reaction was complete, the reaction mixture was cooled to room temperature and quenched by adding water. The resulting mixture was extracted with ethyl acetate. The combined organic layers were washed with brine, dehydrated with anhydrous Na2SO4, and filtered. The filtrate was concentrated under vacuum. The residue was purified by silica gel column chromatography using 0-50% ethyl acetate in petroleum ether, yielding 4-(4-(hydroxymethyl)piperidine-1-yl)methyl benzoate (14 g, 34%) as a white solid. MS (ESI) calculated value (C 14 H 19 NO3) [M+H] + , 250.1; measured value 250.0.

[0213] Step 2: Synthesis of 4-(4-((tert-butyldimethylsilyloxy)methyl)piperidine-1-yl)methyl benzoate. To a solution of 4-(4-(hydroxymethyl)piperidine-1-yl)methyl benzoate (40.0 g, 160.44 mmol) in DMF (400 mL), imidazole (21.8 g, 320.88 mmol), DMAP (1.9 g, 16.04 mmol), and t-butyldimethylchlorosilane (29.0 g, 192.53 mmol) were added. The resulting mixture was stirred at room temperature for 3 hours under a nitrogen atmosphere. The reaction mixture was quenched by adding water and extracted with ethyl acetate. The combined organic layers were washed with water and brine, dehydrated with anhydrous Na2SO4, and filtered. The filtrate was concentrated under vacuum. Purification of the residue by silica gel column chromatography using 0-20% ethyl acetate in petroleum ether yielded methyl 4-(4-((tert-butyldimethylsilyloxy)methyl)piperidine-1-yl)benzoate (35.0 g, 60%) as a white solid. MS (ESI) calculated value (C 20 H33 NO3Si)[M+H] + , 364.2; measured value 364.2.

[0214] Step 3: Synthesis of 4-(4-((tert-butyldimethylsilyloxy)methyl)piperidine-1-yl)benzohydrazide. A solution of methyl 4-(4-((tert-butyldimethylsilyloxy)methyl)piperidine-1-yl)benzoate (35.0 g, 96.26 mmol) in EtOH (150 mL) was mixed with hydrazine (150 mL, 80%). This mixture was stirred at 90°C for 6 hours and then concentrated under vacuum. The crude residue was diluted with water and extracted with ethyl acetate. The combined organic layers were washed with brine, dehydrated with anhydrous Na2SO4, and filtered. The filtrate was concentrated under vacuum to obtain 4-(4-((tert-butyldimethylsilyloxy)methyl)piperidine-1-yl)benzohydrazide (29.0 g, crude) as a white solid, which was used in the next step without further purification. MS(ESI) calculated value (C 19 H 33 N3O2Si)[M+H] + , 364.2; measured value 364.0.

[0215] Step 4: Synthesis of 2-(4-(4-((tert-butyldimethylsilyloxy)methyl)piperidine-1-yl)benzoyl)-N-methylhydrazinecarboxamide. 2,5-Dioxopyrrolidine-1-yl N-methylcarbamate (20.6 g, 119.64 mmol) and DIEA (30.9 g, 239.28 mmol) were added to a solution of 4-(4-((tert-butyldimethylsilyloxy)methyl)piperidine-1-yl)benzohydrazide (29.0 g, 79.76 mmol) in MeCN (300 mL). The mixture was stirred at room temperature under a nitrogen atmosphere for 16 hours. The reaction mixture was quenched by adding water and extracted with ethyl acetate. The combined organic layers were washed with brine, dehydrated with anhydrous Na2SO4, and filtered. When the filtrate was concentrated under vacuum, 2-(4-(4-((tert-butyldimethylsilyloxy)methyl)piperidine-1-yl)benzoyl)-N-methylhydrazine carboxamide (38.0 g, crude) was obtained as an off-white solid, which was used in the next step without further purification. MS(ESI) calculated value (C 21 H 36 N4O3Si)[M+H] + , 421.3; measured value 421.0.

[0216] Step 5: Synthesis of 5-(4-(4-((tert-butyldimethylsilyloxy)methyl)piperidine-1-yl)phenyl)-4-methyl-2H-1,2,4-triazole-3(4H)-one. 2-(4-(4-((tert-butyldimethylsilyloxy)methyl)piperidine-1-yl)benzoyl)-N-methylhydrazinecarboxamide (38.0 g, 90.34 mmol) was added to a solution of NaOH (7.2 g, 180.68 mmol) in water (300 mL). This mixture was stirred at 100°C for 3 hours under a nitrogen atmosphere. After cooling to room temperature, the resulting mixture was extracted with ethyl acetate. The combined organic layers were washed with brine, dehydrated with anhydrous Na2SO4, and concentrated under vacuum. Purification of the residue by silica gel column chromatography using 0-10% methanol in dichloromethane yielded 5-(4-(4-((tert-butyldimethylsilyloxy)methyl)piperidine-1-yl)phenyl)-4-methyl-2H-1,2,4-triazole-3(4H)-one (12g, 33%) as an off-white solid. MS(ESI) calculation value (C 21 H 34 N4O2Si)[M+H] + , 403.2; measured value 403.2.

[0217] Step 6: Synthesis of 3-(3-(4-(4-((tert-butyldimethylsilyloxy)methyl)piperidine-1-yl)phenyl)-4-methyl-5-oxo-4,5-dihydro-1,2,4-triazole-1-yl)piperidine-2,6-dione. 5-(4-(4-((tert-butyldimethylsilyloxy)methyl)piperidine-1-yl)phenyl)-4-methyl-2H-1,2,4-triazole-3(4H)-one (9.6 g, 23.84 mmol) was dissolved in DMF (100 mL) and NaH (60%, 2.6 g, 65.0 mmol) was added in fractions at 0°C. This mixture was stirred at room temperature under a nitrogen atmosphere for 30 minutes. 3-bromopiperidine-2,6-dione (7.7 g, 40.10 mmol) was added to the above mixture at 0°C. The resulting mixture was stirred under a nitrogen atmosphere at 40°C for 4 hours. The reaction mixture was carefully poured into a saturated aqueous solution of NH4Cl and then extracted with ethyl acetate. The combined organic layers were washed with brine, dehydrated with anhydrous sodium sulfate, and filtered. The filtrate was concentrated under vacuum. Purification of the residue by flash column chromatography using 0-8% methanol in dichloromethane yielded 3-(3-(4-(4-((tert-butyldimethylsilyloxy)methyl)piperidine-1-yl)phenyl)-4-methyl-5-oxo-4,5-dihydro-1,2,4-triazole-1-yl)piperidine-2,6-dione (8.2 g, 66%) as a brown oily substance. MS(ESI) calculation value (C 26 H 39 N5O4Si)[M+H] + 514.3; measured value 514.3.

[0218] Step 7: Synthesis of 3-(3-(4-(4-(hydroxymethyl)piperidine-1-yl)phenyl)-4-methyl-5-oxo-4,5-dihydro-1,2,4-triazole-1-yl)piperidine-2,6-dione. The mixture of 3-(3-(4-(4-((tert-butyldimethylsilyloxy)methyl)piperidine-1-yl)phenyl)-4-methyl-5-oxo-4,5-dihydro-1,2,4-triazole-1-yl)piperidine-2,6-dione (350 mg, 0.68 mmol) in HCl (4 M in 1,4-dioxane, 5 mL) was stirred at room temperature for 2 hours. The solvent was removed under vacuum. The crude residue was diluted with DMF and made basic to pH 8-9 using triethylamine. The mixture obtained by reverse-phase flash chromatography using the following conditions: [column, C18 silica gel; mobile phase, ACN (0.05% NH4HCO3) in water, gradient from 10% to 35% over 30 minutes; detector, UV 254 nm] was purified to obtain 3-(3-(4-(4-(hydroxymethyl)piperidine-1-yl)phenyl)-4-methyl-5-oxo-4,5-dihydro-1,2,4-triazole-1-yl)piperidine-2,6-dione (90 mg, 22%) as a white solid. 1 1H NMR (300 MHz, DMSO-d6 + D2O) δ 7.55 - 7.41 (m, 2H), 7.12 - 6.88 (m, 2H), 5.15 - 5.05 (m, 1H), 3.85 - 3.76 (m, 2H), 3.34 - 3.20 (m, 5H), 2.91 - 2.58 (m, 4H), 2.48 - 2.34 (m, 1H), 2.21 - 2.05 (m, 1H), 1.78 - 1.66 (m, 2H), 1.63 - 1.48 (m, 1H), 1.24 - 1.08 (m, 2H).MS(ESI) calculated value (C 20 H 25 N5O4)[M+H] + , 400.2; measured value 400.1.

[0219] Step 8: rac-(R)-1-(4-(1-(2,6-Dioxopiperidin-3-yl)-4-methyl-5-oxo-4,5-dihydro-1H-1,2,4-triazol-3-yl)phenyl)piperidine-4-carbaldehyde. To a solution of 3-(3-(4-(4-(Hydroxymethyl)piperidin-1-yl)phenyl)-4-methyl-5-oxo-4,5-dihydro-1,2,4-triazol-1-yl)piperidine-2,6-dione (0.30 g, 0.75 mmol) in DCM (5 mL) was added Dess-Martin periodinane (0.38 g, 0.90 mmol). The reaction mixture was stirred at room temperature for 2 h, filtered through a pad of celite, concentrated on silica gel, and then purified by column chromatography (0 - 100% EtOAc / hexane) to give the title compound. LCMS C 20 H 23 N5O4 Calcd: 397.2, Found: m / z = 398.4 [M+H] + Minor, 416.4 [M+H2O] + Major. HCB12: 3-(2-((2-(2,6-Dioxopiperidin-3-yl)-1,3-dioxoisoindolin-5-yl)amino)ethoxy)propanoic acid [Chemical formula]

[0220] 2-(2,6-Dioxopiperidin-3-yl)-4-fluoroisoindoline-1,3-dione was replaced with 2-(2,6-dioxopiperidin-3-yl)-5-fluoroisoindoline-1,3-dione and prepared in a similar manner to HCB1 to give the title compound. LCMS C 18 H 19 N3O7 Calcd: 389.1, Found: m / z = 387.8 [M-H] - HCB13: 1-(1-(2-(2,6-Dioxopiperidin-3-yl)-1,3-dioxoisoindolin-5-yl)piperidin-4-yl)azetidine-3-carboxylic acid [Chemical formula]

[0221] Step 1: Benzyl 4-{3-[(tert-butoxy)carbonyl]azetidin-1-yl}piperidine-1-carboxylate. Acetic acid (2.46 ml, 42.94 mmol, 1.5 eq) was added to a solution of tert-butyl azetidine-3-carboxylate (4.5 g, 28.62 mmol, 1.0 eq) and 1-(benzyloxycarbonyl)-4-piperidinone (7.35 g, 31.49 mmol, 1.10 eq) in DCE (136 mL, 0.2 M), and the reaction was stirred at room temperature for 1 h. Then, NaBH(OAc)3 (9.71 g, 45.8 mmol, 1.6 eq) was added and the reaction was stirred at room temperature overnight. The reaction mixture was quenched with aqueous NaHCO3, extracted with DCM (3×), washed with brine and dried over Na2SO4, and concentrated to dryness. Purification by flash column chromatography eluting with DCM:MeOH (0-10%) gave 9.39 g (88% yield) of the desired product as a white solid. ESI[M+H] + =375.6

[0222] Step 2: tert-Butyl 1-(piperidin-4-yl)azetidine-3-carboxylate. A solution of benzyl 4-{3-[(tert-butoxy)carbonyl]azetidin-1-yl}piperidine-1-carboxylate (9.39 g, 25.07 mmol, 1.0 eq) in MeOH (250 ml, 0.1 M) was degassed three times and filled with argon. Next, Pd(OH)2 (0.7 g, 5.0 mmol, 0.2 eq) was added and the mixture was degassed three times again and filled with argon. Then, the reaction mixture (RM) was degassed and a H2 balloon was introduced, and the reaction was stirred at room temperature overnight. Cleavage of the Cbz was confirmed by UPLC. The reaction mixture was filtered through a Celite pad and the filtrate was concentrated to give 5.81 g (96% yield) of the desired product.

[0223] Step 3: tert-butyl 1-{1-[2-(2,6-dioxopiperidine-3-yl)-1,3-dioxo-2,3-dihydro-1H-isoindole-5-yl]piperidine-4-yl}azetidine-3-carboxylate. To a solution of 2-(2,6-dioxopiperidine-3-yl)-5-fluoro-2,3-dihydro-1H-isoindole-1,3-dione (6.05 g, 21.9 mmol, 1.0 equivalent) in DMSO (43.8 mL, 0.5 M), tert-butyl 1-(piperidine-4-yl)azetidine-3-carboxylate (5.79 g, 24.09 mmol, 1.1 equivalent) and DIPEA (7.63 mL, 43.8 mmol, 2.0 equivalent) were added. Next, the reaction mixture was transferred to a bath preheated to 90°C and stirred overnight under an Ar atmosphere. UPLC demonstrated the formation of the desired product. The reaction mixture (RM) was quenched with water and extracted with DCM (×3), and the organic phase was washed with ice-cold water. The crude product was purified by FC elution with DCM:acetone (0-10%) to obtain 6.95 g (64% yield) of the product as a yellow solid. ESI[M+H] + = 497.4.

[0224] Step 4: 1-{1-[2-(2,6-dioxopiperidine-3-yl)-1,3-dioxo-2,3-dihydro-1H-isoindole-5-yl]piperidine-4-yl}azetidine-3-carboxylate hydrochloride. To a solution of tert-butyl 1-{1-[2-(2,6-dioxopiperidine-3-yl)-1,3-dioxo-2,3-dihydro-1H-isoindole-5-yl]piperidine-4-yl}azetidine-3-carboxylate (4.95 g, 9.97 mmol, 1.0 equivalent) in anhydrous DCM (100 mL, 0.1 M), 2 M HCl (50 ml, 99.69 mmol, 10.0 equivalent) in Et2O was added. The reaction mixture was then stirred at room temperature for 2 hours, and the presence of the starting material (SM) was shown by UPLC. Further amounts of HCl in Et2O (50 ml, 99.69 mmol, 10.0 equivalents) were added, and the reaction mixture was stirred for a further 3 hours. UPLC showed 10% of the starting material. The precipitate was filtered, redissolved in DCM, and then 2M HCl in Et2O (50 ml, 99.69 mmol, 10.0 equivalents) was added. The reaction mixture was sonicated for 45 minutes. The precipitated solid was filtered off, washed with Et2O, and dried under vacuum to obtain 4.83 g (quantitative yield) of the desired product as an HCl salt. LCMS (254 nm): RT=2.83 min, 94.5%, ESI[M+H] + =441.07; 1 H NMR (300 MHz, D2O) δ 7.68 (d, J = 8.5 Hz, 1H), 7.37 (s, 1H), 7.22 (dd, J = 8.6, 2.3 Hz, 1H), 5.14 (dd, J = 12.8, 5.6 Hz, 1H), 4.49 - 4.28 (m, 4H), 4.08 (d, J = 13.6 Hz, 2H), 3.80 (t, J = 9.0 Hz, 1H), 3.69 - 3.56 (m, 1H), 3.03 (t, J = 12.8 Hz, 2H), 2.92 - 2.73 (m, 2H), 2.61 (qd, J = 12.8, 5.6 Hz, 1H), 2.24 - 2.09 (m, 2H), 1.59 - 1.42 (m, 2H). HCB14:2-(4-(2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-5-yl)piperazine-1-yl)acetic acid [ka]

[0225] Step 1: tert-butyl 2-{4-[2-(2,6-dioxopiperidine-3-yl)-1,3-dioxo-2,3-dihydro-1H-isoindole-5-yl]piperazine-1-yl}acetate. To a solution of tert-butylpiperazine-1-yl acetate dihydrochloride (4.46 g, 0.0163 mmol, 1.1 equivalents) in DMSO (29.7 mL, 0.5 M), DIPEA (3.93 mL, 0.0297 mmol, 2 equivalents) and 2-(2,6-dioxopiperidine-3-yl)-5-fluoroisoindorin-1,3-dione (4.1 g, 0.0148 mmol, 1 equivalent) were added. The reaction mixture was heated under argon at 90°C for 40 hours. The reaction mixture was cooled to room temperature and 5 mL of water was added dropwise. A bright yellow precipitate was formed, which was filtered off and washed twice with water on a filter. The filtrate was extracted twice by DCM. The combined DCM layers were concentrated under vacuum and combined with the precipitate. Purification of the crude product by flash column chromatography yielded tert-butyl 2-{4-[2-(2,6-dioxopiperidine-3-yl)-1,3-dioxo-2,3-dihydro-1H-isoindole-5-yl]piperazine-1-yl}acetate (5.49 g, 81% yield) as a yellow solid. ESI[M+H] + =457.7; 1H NMR (300 MHz, Chloroform-d) δ 8.07 (s, 1H), 7.72 (d, J = 8.5 Hz, 1H), 7.31 (d, J = 2.3 Hz, 1H), 7.08 (dd, J = 8.6, 2.4 Hz, 1H), 4.96 (dd, J = 12.2, 5.2 Hz, 1H), 3.55 - 3.45 (m, 4H), 3.21 (s, 2H), 2.98 - 2.81 (m, 2H), 2.81 - 2.72 (m, 5H), 2.24 - 2.09 (m, 1H), 1.50 (s, 9H).

[0226] Step 2: Trifluoroacetate of 2-{4-[2-(2,6-dioxopiperidine-3-yl)-1,3-dioxo-2,3-dihydro-1H-isoindole-5-yl]piperazine-1-yl}acetic acid. To a solution of tert-butyl 2-{4-[2-(2,6-dioxopiperidine-3-yl)-1,3-dioxo-2,3-dihydro-1H-isoindole-5-yl]piperazine-1-yl}acetate (5.49 g, 12.03 mmol, 1 equivalent) in DCM (100 mL, 0.12 M), TFA (50.6 mL, 661 mmol, 55 equivalents) was added. The reaction mixture was stirred at room temperature for 16 hours, and then concentrated under reduced pressure. The resulting bright yellow viscous solid was sonicated with 200 mL of anhydrous diethyl ether and stirred for 1 hour. The resulting precipitate was filtered, washed twice with anhydrous Et2O, and dried under reduced pressure to obtain a bright yellow solid (6.55 g, quantitative). LCMS (254 nm): RT=2.69 min, 98.59%, ESI[M+H] + =401.14; 1 H NMR (400 MHz, DMSO-d6) δ 11.11 (s, 1H), 10.87 (br. s, 1H), 7.77 (d, J = 8.4 Hz, 1H), 7.48 (d, J = 2.3 Hz, 1H), 7.34 (dd, J = 8.6, 2.3 Hz, 1H), 6.27 (br. s, 2H), 5.10 (dd, J = 12.9, 5.4 Hz, 1H), 4.22 (s, 2H), 4.11 (br. s, 2H), 3.45 (br. s, 6H), 3.38 (dd, J = 139.7, 7.0 Hz, 8H), 2.90 (ddd, J = 17.4, 14.1, 5.5 Hz, 1H), 2.65 - 2.52 (m, 2H), 2.10 - 1.97 (m, 1H). HCB15:6-((2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-4-yl)amino)hexanoic acid [ka]

[0227] The title compound was obtained by substituting 2-(2,6-dioxopiperidine-3-yl)-5-fluoroisoindoline-1,3-dione with 2-(2,6-dioxopiperidine-3-yl)-4-fluoroisoindoline-1,3-dione and preparing it in a manner similar to that of HCB3. LCMS C 19 H 21 N3O6 theoretical value: 387.1, measured value: m / z = 385.9 [MH] - HCB16:8-((2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-5-yl)amino)octanoic acid [ka]

[0228] The title compound was obtained by substituting tert-butyl 8-aminooctanoate with tert-butyl 6-aminohexanoate hydrochloride and preparing it in a manner similar to that of HCB3. LCMS C 21 H 25 N3O6 theoretical value: 415.2, measured value: m / z = 414.2 [MH] - HCB17:2-(4-(2-(1-methyl-2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-5-yl)piperazine-1-yl)acetic acid [ka]

[0229] The title compound was synthesized by substituting 2-(2,6-dioxopiperidine-3-yl)-5-fluoroisoindorin-1,3-dione with 5-fluoro-2-(1-methyl-2,6-dioxopiperidine-3-yl)isoindorin-1,3-dione using a method similar to that for HCB14. LCMS C 20 H 22 N4O6 theoretical value: 414.2, measured value: m / z = 415.4 [M + H] + HCB18: N-(2,6-dioxopiperidine-3-yl)-5-(4-formylpiperidine-1-yl)pyridine-2-carboxamide [ka]

[0230] Step 1: Methyl 5-{4-[(benzyloxy)methyl]piperidine-1-yl}pyridine-2-carboxylate. Methyl 5-bromopyridine-2-carboxylate (1.0 g, 4.63 mmol, 1.0 equivalent), 4-[(benzyloxy)methyl]piperidine (950 mg, 4.63 mmol, 1.0 equivalent), rac-BINAP (288 mg, 463 μmol, 0.1 equivalent), Pd2(dba)3 (432 mg, 463 μmol, 0.1 equivalent), and Cs2CO3 (4.52 g, 13.9 mmol, 3.0 equivalent) were suspended in toluene (30 mL), and the mixture was heated to 100°C for 12 hours. The reaction mixture was cooled to room temperature, diluted with 100 mL of butyl, and then purified by filtration (elution with SiO2, 10 → 100% butyl / hexane, 70%) to obtain methyl 5-{4-[(benzyloxy)methyl]piperidine-1-yl}pyridine-2-carboxylate (1.1 g, 67%). LCMS:C 20 H 24 Theoretical value of N2O3: 340, Measured value: m / z = 341 [M + H] + .

[0231] Step 2: 5-{4-[(benzyloxy)methyl]piperidine-1-yl}pyridine-2-carboxylic acid. Methyl 5-{4-[(benzyloxy)methyl]piperidine-1-yl}pyridine-2-carboxylate (510 mg, 1.50 mmol, 1.0 equivalent) was suspended in MeOH / H2O (1:4), and NaOH (90 mg, 2.25 mmol, 1.5 equivalents) was added in one step at room temperature. After 16 hours, HCl (1 M, aqueous solution) was added to make a pH=5 solution. The solid was collected by filtration, yielding 5-{4-[(benzyloxy)methyl]piperidine-1-yl}pyridine-2-carboxylic acid (800 mg, 83%). LCMS:C 19 H 22 Theoretical value of N2O3: 326, Measured value: m / z = 327 [M + H] + .

[0232] Step 3: 5-{4-[(benzyloxy)methyl]piperidine-1-yl}-N-(2,6-dioxopiperidine-3-yl)pyridine-2-carboxamide. 5-{4-[(benzyloxy)methyl]piperidine-1-yl}pyridine-2-carboxylic acid (510 mg, 1.56 mmol, 1.0 equivalent) was dissolved in DMF (1 mL), and HATU (594 mg, 1.56 mmol, 1.0 equivalent) was added at room temperature. After 5 minutes, 3-aminopiperidine-2,6-dione hydrochloride (257 mg, 1.56 mmol, 1.0 equivalent) and DIPEA (1.09 mL, 6.25 mmol, 4.0 equivalent) were added. The mixture was stirred for 16 hours and then partitioned between butyl / H2O (20 mL each). The organic phase was washed (2 × 5 mL H₂O, 1 × 5 mL brine), dried (Na₂SO₄), filtered, and concentrated. Purification (SiO₂, 0 → 4% MeOH / CH₂Cl₂) yielded 5-{4-[(benzyloxy)methyl]piperidine-1-yl}-N-(2,6-dioxopiperidine-3-yl)pyridine-2-carboxamide (625 mg, 92%) as a white solid. LCMS:C 24 H 28 N4O4 theoretical value: 436, measured value: m / z = 437 [M + H] + .

[0233] Step 4: N-(2,6-dioxopiperidine-3-yl)-5-[4-(hydroxymethyl)piperidine-1-yl]pyridine-2-carboxamide. Under an H2 atmosphere (balloon), 5-{4-[(benzyloxy)methyl]piperidine-1-yl}-N-(2,6-dioxopiperidine-3-yl)pyridine-2-carboxamide (500 mg, 1.15 mmol, 1.0 equivalent), acetate (196 μL, 3.44 mmol, 3.0 equivalent), Pd(OH)2 (50 mg) and Pd / C (50 mg) were suspended in EtOH (100 mL). The mixture was heated to 40°C for 18 hours, then cooled, filtered, and concentrated. Purification (SiO2, 0 → 8% MeOH / CH2Cl2) yielded N-(2,6-dioxopiperidine-3-yl)-5-[4-(hydroxymethyl)piperidine-1-yl]pyridine-2-carboxamide (233 mg, 58%) as a white solid. 1H NMR (500 MHz, DMSO-d6) δ 10.84 (s, 1H), 8.69 (d, J = 8.3 Hz, 1H), 8.30 (d, J = 3.0 Hz, 1H), 7.83 (d, J = 8.8 Hz, 1H), 7.40 (dd, J = 9.0, 3.0 Hz, 1H), 4.74 (dq, J = 13.3, 6.4, 5.8 Hz, 1H), 3.95 (d, J = 12.7 Hz, 2H), 3.28 (d, J = 6.1 Hz, 2H), 2.88 - 2.73 (m, 3H), 2.53 (s, 2H), 2.23 - 2.11 (m, 1H), 2.01 (d, J = 13.1 Hz, 1H), 1.78 - 1.71 (m, 2H), 1.61 (s, 1H), 1.21 (h, J = 11.0, 10.6 Hz, 2H).LCMS:C 17 H 22 N4O4 theoretical value: 346, measured value: m / z = 347 [ M+H] + .

[0234] Step 5: N-(2,6-dioxopiperidine-3-yl)-5-(4-formylpiperidine-1-yl)pyridine-2-carboxamide. A mixture consisting of N-(2,6-dioxopiperidine-3-yl)-5-[4-(hydroxymethyl)piperidine-1-yl]pyridine-2-carboxamide (200 mg, 580 μmol, 1.0 equivalent) and Et3N (321 μL, 2.31 mmol, 4.0 equivalents) was dissolved in DMSO (500 μL) and CH2Cl2 (500 μL). This reaction mixture was cooled to 0°C and SO 3·Pyridine (184 mg, 1.15 equivalents, 2.0 equivalents, solution in 300 μL of DMSO) was added dropwise. The reaction mixture was warmed to room temperature and stirred for 30 minutes, after which NaHCO3 (5 mL, saturated aqueous solution) was added. After 1 minute, this suspension was diluted with CH2Cl2 (10 mL), and the aqueous phase was extracted (3 × 10 mL CH2Cl2). The combined organic matter was washed (2 × 5 mL H2O, 1 × 5 mL brine), dried (Na2SO4), filtered, and concentrated. Purification (SiO2, 0 → elution with 10% MeOH / CH2Cl2, 5%) yielded N-(2,6-dioxopiperidine-3-yl)-5-(4-formylpiperidine-1-yl)pyridine-2-carboxamide as a white foam (190 mg, 95%). LCMS:C 17 H 20 N4O4 theoretical value: 344, measured value: m / z = 345 [M+H] + . HCB19:N-(2,6-dioxopiperidine-3-yl)-4-formylbenzamide [ka]

[0235] 4-Formylbenzoic acid (500 mg, 3.33 mmol) and HATU were combined in DMF, then DIPEA (4 equivalents, 13.3 mmol) was added and the mixture was stirred for 5 minutes. Next, 3-aminopiperidine-2,6-dione hydrochloride was added, and the reaction mixture was stirred for 18 hours. The mixture was then directly purified by reverse-phase chromatography (C18 column, 0-100% acetonitrile in water) to obtain the desired product (0.6 g, 69% yield).

[0236] LCMS:C 13 H 12 Theoretical value for N2O4: 260.1, Measured value: m / z = 261.2 [M + H] + . HCB20: (3R)-1-(1-(2,6-dioxopiperidine-3-yl)-3-methyl-2-oxo-2,3-dihydro-1H-benzo[d]imidazole-5-yl)pyrrolidine-3-carbaldehyde [ka]

[0237] Step 1: tert-butyl(3R)-3-[[tert-butyl(diphenyl)silyl]oxymethyl]pyrrolidine-1-carboxylate. A mixture of tert-butyl (3R)-3-(hydroxymethyl)pyrrolidine-1-carboxylate (25.0 g, 124 mmol) and imidazole (10.1 g, 149 mmol) in DCM (500 mL) was mixed with TBDPSCl (32.3 mL, 124 mmol) under nitrogen at 0°C. The mixture was stirred at 23°C for 16 hours and diluted with water (300 mL). The organic phase was washed with water (100 mL) and brine (3 × 100 mL), dried (Na₂SO₄), filtered, and concentrated to obtain the title compound as an oil (54.0 g, 99%). m / z:ES + [M-C6H5-tBu+H] + =306.2, LCMS(A05);t R =2.47 minutes. ; 1 H NMR (500 MHz, CDCl3) δ 7.67 - 7.60 (m, 4H), 7.46 - 7.34 (m, 6H), 3.64 - 3.55 (m, 2H), 3.45 - 3.37 (m, 1H), 3.37 - 3.22 (m, 1H), 3.17 - 3.07 (m, 1H), 2.42 (m, 1H), 1.97 - 1.86 (m, 1H), 1.74 - 1.62 (m, 1H), 1.60 (s, 1H), 1.46 (s, 9H), 1.08 - 1.02 (m, 9H);

[0238] Step 2: tert-butyl-diphenyl-[[(3R)-pyrrolidine-3-yl]methoxy]silane 2,2,2-trifluoroacetic acid. A mixture of tert-butyl(3R)-3-[[tert-butyl(diphenyl)silyl]oxymethyl]pyrrolidine-1-carboxylate (54.0 g, 123 mmol) in DCM (200 mL) was mixed with TFA (50 mL) under nitrogen at 23°C. The mixture was stirred and concentrated at 23°C for 1.5 hours. The residue was diluted with PhMe (150 mL) and concentrated (this process was repeated twice) to obtain the title compound as an oily substance (55.7 g, quantitative). m / z:ES + [M+H-TFA] + =340.3, LCMS(A05);t R =2.32 minutes. 1 H NMR (500 MHz, CDCl3) δ 8.82 (s, 2H), 7.65 - 7.57 (m, 4H), 7.52 - 7.40 (m, 6H), 3.65 (d, J = 6.4 Hz, 2H), 3.35 - 3.27 (m, 1H), 3.24 - 3.10 (m, 2H), 3.01 - 2.91 (m, 1H), 2.58 - 2.52 (m, 1H), 2.04 - 1.93 (m, 1H), 1.74 - 1.63 (m, 1H), 1.01 (s, 9H);

[0239] Step 3: Dimethyl 2-bromopentanediate. A mixture of (2S)-2-aminopentanediate (30 g, 204 mmol), NaBr (73.2 g, 711 mol), and HBr (50 mL, 48% in water) in water (100 mL) was added to a solution of NaNO2 (25.5 g, 370 mmol) in water (50 mL) under nitrogen at 0°C (internal temperature maintained below 10°C). This mixture was stirred at 23°C for 6 hours, and H2SO4 (25.0 mL) was added at 23°C. This mixture was extracted with Et2O (4 × 70.0 mL), and the combined organic phase was washed with brine (2 × 50.0 mL), dried (Na2SO4), filtered, and concentrated. H2SO4 (10.0 mL) was added to the mixture of residues in MeOH (80.0 mL) under nitrogen at 23°C. This mixture was refluxed for 16 hours and cooled to 23°C to concentrate. The residue was diluted with Et2O (100 mL) and water (100 mL). This aqueous phase was extracted with Et2O (4 × 50.0 mL). The combined organic layers were washed with water (60.0 mL), NaHCO3 (2 × 60.0 mL), and brine (2 × 50.0 mL), dried (Na2SO4), filtered, and concentrated to obtain the title compound as an oily substance (19 g, 39%). 1 H NMR (400 MHz, CDCl3) δ 4.34 (dd, J = 8.5, 5.8 Hz, 1H), 3.75 (s, 3H), 3.65 (s, 3H), 2.52 - 2.45 (m, 2H), 2.40 - 2.30 (m, 1H), 2.26 (m, 1H).

[0240] Step 4: 5-Bromo-N-methyl-2-nitroaniline. To a mixture of 4-bromo-2-fluoro-1-nitrobenzene (50.0 g, 227 mmol) in EtOH (455 mL), methylamine (56.6 mL, 455 mmol, 33 wt% in EtOH) was added under nitrogen at 23°C. This mixture was stirred at 23°C for 30 minutes, filtered, and washed with cold EtOH (200 mL) to obtain the title compound as a solid (48.2 g, 92%). m / z(ES) + )[M+H] + =231.0, LCMS(A05);t R =2.51 minutes;.1 H NMR (400 MHz, DMSO-d6) δ 8.23 ​​(d, J = 4.3 Hz, 1H), 7.98 (d, J = 9.1 Hz, 1H), 7.17 (d, J = 2.0 Hz, 1H), 6.82 (dd, J = 9.1, 2.1 Hz, 1H), 2.95 (d, J = 5.0 Hz, 3H)

[0241] Step 5: [(3R)-1-[3-(methylamino)-4-nitrophenyl]pyrrolidine-3-yl]methanol. RuPhos-Pd-G3 (2.71 g, 3.25 mmol) was added to a mixture of 5-bromo-N-methyl-2-nitro-aniline (25 g, 108 mmol), tert-butyl-diphenyl-[[(3R)-pyrrolidine-3-yl]methoxy]silane 2,2,2-trifluoroacetic acid (60.0 g, 119 mmol, 90% purity) and Cs2CO3 (106 g, 325 mmol) in PhMe (600 mL) under nitrogen at 23°C. The mixture was degassed by passing nitrogen through it at 23°C for 15 minutes, stirred at 100°C for 19 hours, cooled to 23°C, filtered, and concentrated. The product was purified by silica gel chromatography using hexane and butyl (0-50%) (2 x 330 g cartridges in series), yielding the title compound as a solid (41.0 g, 77%). m / z:ES + [M+H] + =490.4; 1H NMR (400 MHz, DMSO-d6) δ 8.36 (d, J = 4.9 Hz, 1H), 7.91 (d, J = 9.6 Hz, 1H), 7.64 - 7.57 (m, 4H), 7.50 - 7.37 (m, 6H), 6.07 (dd, J = 9.6, 2.5 Hz, 1H), 5.50 (d, J = 2.4 Hz, 1H), 3.68 (d, J = 6.6 Hz, 2H), 3.54 - 3.46 (m, 1H), 3.46 - 3.37 (m, 2H), 3.27 - 3.21 (m, 1H), 2.90 (d, J = 5.0 Hz, 3H), 2.64 - 2.55 (m, 1H), 2.16 - 2.04 (m, 1H), 1.90 - 1.79 (m, 1H), 1.01 (s, 9H)

[0242] Step 6: 4-[(3R)-3-ethylpyrrolidine-1-yl]-N2-methylbenzene-1,2-diamine. A solution of [(3R)-1-[3-(methylamino)-4-nitrophenyl]pyrrolidine-3-yl]methanol (20.0 g, 40.8 mmol) in THF (100 mL) and EtOH (100 mL) was added to 10% Pd / C (4.4 g, 4.1 mmol, 50% wet) under nitrogen at 23°C. The mixture was refluxed and hydrazine hydrate (16 mL, 163 mmol) was added over 30 minutes. The mixture was refluxed for 2 hours and cooled to 23°C, filtered (Celite), washed with ELISA (200 mL) and EtOH (200 mL), and concentrated to obtain the title compound as an oil (18.0 g, 96%). m / z ESI + [M-Ph-tBu+H] + =328.16; 1H NMR (400 MHz, DMSO-d6) δ 7.65 - 7.59 (m, 4H), 7.48 - 7.38 (m, 6H), 6.45 - 6.40 (m, 1H), 5.71 (d, J = 2.5 Hz, 1H), 5.66 (dd, J = 8.1, 2.5 Hz, 1H), 4.50 (d, J = 4.9 Hz, 1H), 3.65 (d, J = 6.8 Hz, 2H), 3.33 (br. s, 2H), 3.21 (dd, J = 9.1, 7.6 Hz, 1H), 3.14 - 3.07 (m, 2H), 2.97 (dd, J = 9.2, 5.9 Hz, 1H), 2.68 (d, J = 4.2 Hz, 3H), 2.56 - 2.51 (m, 1H), 2.05 - 1.95 (m, 1H), 1.76 - 1.67 (m, 1H), 1.01 (s, 9H);

[0243] Step 7: 5-[(3R)-3-ethylpyrrolidine-1-yl]-3-methyl-1H-benzimidazole-2-one. A mixture of ISN-4-[(3R)-3-ethylpyrrolidine-1-yl]-N2-methylbenzene-1,2-diamine (38.0 g, 82.7 mmol) and DIPEA (115 mL, 661 mmol) in DCM (300 mL) was added to a mixture of triphosgene (8.09 g, 27.3 mmol) in DCM (30 mL) under nitrogen at 0°C. The mixture was stirred at 0°C for 30 minutes and diluted with water (300 mL). The aqueous phase was extracted with DCM (2 × 100 mL), and the combined organic phase was washed with brine (50.0 mL), dried (MgSO4), filtered, and concentrated. The product was purified by silica gel chromatography (2 × 330 g cartridge) using DCM and MeOH (0-10%), yielding the title compound as a solid (21 g, 52%). m / z:ES + [M+H] + = 486.4.

[0244] Step 8: Dimethyl 2-[5-[(3R)-3-ethylpyrrolidine-1-yl]-3-methyl-2-oxo-benzimidazole-1-yl]pentanediate. To a mixture of 5-[(3R)-3-ethylpyrrolidine-1-yl]-3-methyl-1H-benzimidazole-2-one (10.0 g, 20.6 mmol) and Cs2CO3 (20.3 g, 62.3 mmol) in DMF (100 mL), dimethyl 2-bromopentanedioate (10.9 g, 30.9 mmol, 68% purity) was added under nitrogen at 23°C. The mixture was stirred at 100°C for 18 hours and cooled to 23°C, then diluted with ELISA (200 mL) and water (100 mL). The aqueous phase was extracted with ELISA (2 × 100 mL), and the combined organic phase was washed with brine (2 × 50 mL), dried (MgSO4), filtered, and concentrated. The product was purified by silica gel chromatography (220 g cartridge) using hexane and ELISA (0-50%), yielding the title compound as a solid (9.00 g, 68%). m / z:ES + [M+H] + =644.4, LCMS(A05);t R =2.33 minutes.

[0245] Step 9: 2-[5-[(3R)-3-ethylpyrrolidine-1-yl]-3-methyl-2-oxo-benzimidazole-1-yl]pentanedioic acid. To a mixture of dimethyl 2-[5-[(3R)-3-ethylpyrrolidine-1-yl]-3-methyl-2-oxo-benzimidazole-1-yl]pentanedioate (9.00 g, 14.0 mmol) in a mixture of THF and water (200 mL, 1:1 v / v), aqueous NaOH (5 M, 14.0 mL, 70.0 mmol) was added at 23°C under nitrogen. The mixture was stirred at 23°C for 1 hour and diluted with ELISA (100 mL) and aqueous HCl (1 M, 80.0 mL). The aqueous phase was extracted with butyl (3 × 50.0 mL), and the combined organic phase was washed with brine (2 × 50.0 mL), dried (Na₂SO₄), filtered, and concentrated to obtain the title compound as a solid (8.6 g, quantitative). 1H NMR (500 MHz, DMSO-d6) δ 7.67 - 7.58 (m, 4H), 7.51 - 7.36 (m, 6H), 6.93 - 6.81 (m, 1H), 6.41 - 6.30 (m, 1H), 6.27 - 6.17 (m, 1H), 4.95 (dd, J = 10.8, 5.0 Hz, 1H), 3.69 (d, J = 6.6 Hz, 2H), 3.38 - 3.31 (m, 1H), 3.29 (s, 3H), 3.27 - 3.19 (m, 2H), 3.12 - 3.03 (m, 1H), 2.65 - 2.55 (m, 1H), 2.41 - 2.21 (m, 2H), 2.21 - 2.02 (m, 3H), 1.86 - 1.79 (m, 1H), 1.02 (s, 9H).

[0246] Step 10: 3-[5-[(3R)-3-ethylpyrrolidine-1-yl]-3-methyl-2-oxo-benzimidazole-1-yl]piperidine-2,6-dione. A mixture of 2-[5-[(3R)-3-ethylpyrrolidine-1-yl]-3-methyl-2-oxo-benzimidazole-1-yl]pentanedioic acid (5.0 g, 8.12 mmol), trifluoroacetamide (1.01 g, 8.93 mmol), and DIPEA (5.66 mL, 32.5 mmol) in DMF (50.0 mL) was added with HATU (6.792 g, 17.9 mmol) under nitrogen at 23°C. The mixture was concentrated by stirring at 23°C for 18 hours. The product was purified by silica gel chromatography (120 g cartridge) using DCM and MeOH (0-5%), yielding the title compound as a solid (3.30 g, 68%). 1 H NMR (500 MHz, DMSO-d6) δ 11.04 (s, 1H), 7.65 - 7.60 (m, 4H), 7.50 - 7.39 (m, 6H), 6.93 - 6.87 (m, 1H), 6.35 (d, J = 2.1 Hz, 1H), 6.20 (dd, J = 8.6, 2.2 Hz, 1H), 5.26 (dd, J = 12.8, 5.4 Hz, 1H), 3.69 (d, J = 6.6 Hz, 2H), 3.29 (s, 3H), 3.26 - 3.20 (m, 2H), 3.08 - 3.02 (m, 1H), 2.93 - 2.85 (m, 1H), 2.70 (s, 2H), 2.67 - 2.55 (m, 2H), 2.13 - 2.04 (m, 1H), 1.98 - 1.95 (m, 1H), 1.86 - 1.76 (m, 1H), 1.02 (s, 9H).

[0247] Step 11: 3-[5-[(3R)-3-(Hydroxymethyl)pyrrolidin-1-yl]-3-methyl-2-oxo-benzimidazol-1-yl]piperidine-2,6-dione. To a mixture of 3-[5-[(3R)-3-ethylpyrrolidin-1-yl]-3-methyl-2-oxo-benzimidazol-1-yl]piperidine-2,6-dione (3.20 g, 5.36 mmol) in THF (20 mL) was added TBAF (8.00 mL, 8.00 mmol, 1 M in THF) at 23 °C under nitrogen. The mixture was stirred at 23 °C for 3 h and concentrated. The product was purified by silica gel chromatography (220 g cartridge) using DCM and MeOH (0 - 12%) to give the title compound as a solid (1.30 g, 67%). m / z: ES + [M] + = 358.2; 1H NMR (400 MHz, DMSO-d6) δ 11.03 (s, 1H), 6.89 (d, J = 8.5 Hz, 1H), 6.37 (d, J = 2.2 Hz, 1H), 6.21 (dd, J = 8.6, 2.2 Hz, 1H), 5.25 (dd, J = 12.9, 5.4 Hz, 1H), 4.69 (t, J = 5.2 Hz, 1H), 3.48 - 3.36 (m, 2H), 3.37 - 3.32 (m, 1H), 3.29 (s, 3H), 3.27 - 3.15 (m, 2H), 3.05 - 2.97 (m, 1H), 2.95 - 2.83 (m, 1H), 2.73 - 2.55 (m, 2H), 2.48 - 2.37 (m, 1H), 2.08 - 1.92 (m, 2H), 1.79 - 1.68 (m, 1H);

[0248] Step 12: (3R)-1-(1-(2,6-dioxopiperidine-3-yl)-3-methyl-2-oxo-2,3-dihydro-1H-benzo[d]imidazole-5-yl)pyrrolidine-3-carboaldehyde. A mixture of (3RS)-3-{5-[(3R)-3-(hydroxymethyl)pyrrolidine-1-yl]-3-methyl-2-oxo-1,3-benzodiazole-1-yl}piperidine-2,6-dione (33.50 mg, 0.09 mmol) in DMSO (1.00 mL) was mixed with triethylamine (0.26 mL, 0.19 g, 1.87 mmol), followed by sulfur trioxide pyridine complex (148.77 mg, 0.93 mmol). After 25 minutes, water was added, and the mixture was extracted twice by DCM. When the combined organic layers were concentrated, the title compound was obtained without further purification. m / z:ES + [M] +=357.2. Building blocks of LHM targeting VHL can generally be prepared in accordance with scheme B3, in which the LHM is first linked to a linker precursor containing "linker A" (representing one or more linker segments) and two reactive end groups. One of the reactive groups is a carboxylic acid or a reactive equivalent thereof; the other reactive group X can be, for example, a carboxylic acid, hydroxyl, or aldehyde group. The resulting building block of LHM has a reactive moiety (X), which can be further linked to another moiety. HCB21: N-(2,6-dioxopiperidine-3-yl)-4-(4-formylpiperidine-1-yl)-N-methylbenzamide [ka]

[0249] Step 1: 3-[benzyl(methyl)amino]piperidine-2,6-dione. A mixture of 3-bromopiperidine-2,6-dione (6.00 g, 31.2 mmol) and N-methyl-1-phenyl-methaneamine (10.0 g, 82.5 mmol) in DMF (30.0 mL) was stirred at 23°C for 16 hours. The mixture was concentrated. The residue was diluted with toluene (100 mL) and DCM (20.0 mL). The solid was filtered off. The filtrate was further diluted with water (200 mL), ether (100 mL), and ELISA (200 mL). The organic phase was separated, and the aqueous phase was extracted with ELISA (3 × 100 mL). The combined organic phase was washed with brine (3 × 50.0 mL), dried (Na₂SO₄), filtered, and concentrated. The residue was suspended in hexane / ether (10:1 ratio by volume; 200 mL) and stirred for 10 minutes. The solid was collected by filtration and dried under high vacuum to obtain the title compound as an off-white solid (5.65 g, 78%). 1 1H NMR (500 MHz, DMSO-d6) δ 10.63 (s, 1H), 7.36 - 7.29 (m, 4H), 7.27 - 7.21 (m, 1H), 3.78 (s, 2H), 3.61 (dd, J = 12.1, 4.7 Hz, 1H), 2.66 - 2.57 (m, 1H), 2.51 (s, 1H), 2.25 (s, 3H), 2.15 - 2.05 (m, 1H), 1.98 - 1.90 (m, 1H).

[0250] Step 2: 3-(methylamino)piperidine-2,6-dione. In a Parr shaker container, 10% Pd / C (1.58 g, 1.49 mmol) was mixed with a solution of 3-[benzyl(methyl)amino]piperidine-2,6-dione (5.65 g, 24.3 mmol) in siRNA (60.0 mL) under nitrogen. The mixture was purged with hydrogen and stirred at 50 psi at 23°C for 18 hours. The mixture was filtered through Celite and washed with siRNA (100 mL) and DCM (100 mL). The filtrate was concentrated to obtain the title compound as a bright blue solid (3.10 g, 90%). 1 H NMR (400 MHz, DMSO-d6) δ 10.63 (s, 1H), 3.16 (dd, J = 10.3, 4.8 Hz, 1H), 2.52 - 2.41 (m, 2H), 2.27 (s, 3H), 2.26 - 2.22 (m, 1H), 2.02 - 1.93 (m, 1H), 1.71 - 1.57 (m, 1H).m / z(ES + ), [M+H] + 143.1

[0251] Step 3: Benzyl 4-(1,3-dioxolan-2-yl)piperidine-1-carboxylate. A solution of benzyl 4-formylpiperidine-1-carboxylate (25.0 g, 101 mmol), PTSA (515 mg, 2.71 mmol), and ethylene glycol (35.0 mL, 142 mmol) in toluene (120 mL) was refluxed for 7 hours using a Dean-Stark apparatus. After cooling to 23°C, the mixture was concentrated. The residue was diluted with saturated NaHCO3 (100 mL) and ELISA (200 mL). The organic phase was separated, washed with saturated NaHCO3 (2 × 50.0 mL) and brine (2 × 100 mL), dried (Na2SO4), filtered, and concentrated to obtain the title compound as a pale yellow oil (29.5 g, 100%). 1 H NMR (400 MHz, CDCl3) δ 7.36 - 7.24 (m, 5H), 5.09 (s, 2H), 4.61 (d, J = 4.6 Hz, 1H), 4.30 - 4.08 (m, 2H), 3.94 - 3.86 (m, 2H), 3.86 - 3.78 (m, 2H), 2.83 - 2.63 (m, 2H), 1.77 - 1.64 (m, 3H), 1.40 - 1.21 (m, 2H).

[0252] Step 4: 4-(1,3-dioxolan-2-yl)piperidine. To 10% Pd / C (7.20 g, 6.76 mmol), a solution of benzyl 4-(1,3-dioxolan-2-yl)piperidine-1-carboxylate (29.5 g, 422 mmol) in EtOH (120 mL) was added under nitrogen. This suspension was purged with hydrogen and stirred at 23°C for 4 hours. The mixture was filtered through Celite and washed with DCM (200 mL). When the filtrate was concentrated, the title compound was obtained as a colorless oil (15.1 g, 95%). 1 H NMR (500 MHz, CDCl3) δ 4.60 (dd, J = 5.1, 2.3 Hz, 1H), 3.96 - 3.89 (m, 2H), 3.84 (dd, J = 4.2, 2.2 Hz, 2H), 3.08 (d, J = 2.0 Hz, 2H), 2.58 (tt, J = 12.3, 2.3 Hz, 2H), 1.70 (s, 2H), 1.68 - 1.61 (m, 1H), 1.34 - 1.24 (m, 2H), 1.21 (d, J = 3.3 Hz, 1H).

[0253] Step 5: Methyl 4-[4-(1,3-dioxolan-2-yl)-1-piperidyl]benzoate. A mixture of 4-(1,3-dioxolan-2-yl)piperidine (8.00 g, 50.9 mmol), methyl 5-fluoropyridine-2-carboxylate (7.85 g, 50.9 mmol), and K2CO3 (7.04 g, 50.9 mmol) in anhydrous DMSO (20.0 mL) was heated at 80°C for 4 hours. After cooling to 23°C, water (200 mL) was added. The solid was collected by filtration and dried under high vacuum to obtain the title compound as an off-white solid (13.8 g, 93%). m / z(ES) + ), [M+H] + 292.2

[0254] Step 6: 4-[4-(1,3-dioxolan-2-yl)-1-piperidyl]benzoic acid. Methyl 4-[4-(1,3-dioxolan-2-yl)-1-piperidyl]benzoate (10.0 g, 34.3 mmol) in a water / THF mixture (1:1 v / v, 200 mL) was mixed with aqueous NaOH (5 M, 35.0 mL, 175 mmol). This solution was stirred at 23°C for 2 hours. The reaction mixture was diluted with HCl (100 mL) and the pH was adjusted to 4 by adding aqueous HCl (1 M). The organic phase was separated, and the aqueous phase was extracted with HCl (4 × 100 mL). The combined organic phase was washed with brine (100 mL), dried (Na₂SO₄), filtered, and concentrated to obtain the title compound as a light brown solid (7.10 g, 75%). m / z(ES) + ), [M+H] + 278.2

[0255] Step 7: 4-[4-(1,3-dioxolan-2-yl)-1-piperidyl]-N-(2,6-dioxo-3-piperidyl)-N-methyl-benzamide. A mixture of 4-[4-(1,3-dioxolan-2-yl)-1-piperidyl]benzoic acid (2.50 g, 9.01 mmol), HATU (6.86 g, 18.0 mmol), and 3-(methylamino)piperidine-2,6-dione (1.54 g, 10.8 mmol) in anhydrous DMF (20.0 mL) was mixed with DIPEA (3.58 mL, 20.6 mmol) at 23 °C. The mixture was stirred at 23 °C for 2 hours and diluted with water (200 mL). The aqueous phase was extracted with a mixture of iPrOH / CHCl3 (1 / 9 v / v, 4 × 100 mL). The combined organic phases were washed with brine (100 mL), dried (Na2SO4), filtered, and concentrated. The residue was suspended in Et2O (200 mL) and sonicated for 5 minutes. The solid was collected by filtration and dried under high vacuum to obtain the title compound as a colorless solid (1.70 g, 45%). m / z(ES) + ), [M+H] + 402.3

[0256] Step 8: N-(2,6-dioxo-3-piperidyl)-4-(4-formyl-1-piperidyl)-N-methyl-benzamide. A mixture of 4-[4-(1,3-dioxolan-2-yl)-1-piperidyl]-N-(2,6-dioxo-3-piperidyl)-N-methyl-benzamide (1.60 g, 3.99 mmol) in THF (16.0 mL) and water (60.0 mL) was mixed with aqueous HCl (3 M, 10.0 mL, 30 mmol) at 23 °C. This solution was heated at 55 °C for 5 hours. After cooling to 0 °C, a solution of NaHCO3 (1.20 g, 13.9 mmol) in aqueous solution (100 mL) was slowly added. The aqueous phase was extracted with a mixture of iPrOH / CHCl3 (1:9 v / v, 8 × 100 mL). The combined organic phases were washed with brine (100 mL), dried (Na2SO4), filtered, and concentrated. The residue was suspended in Et2O (200 mL) and sonicated for 5 minutes. The solid was collected by filtration and dried under high vacuum to obtain the title compound as a colorless solid (1.10 g, 77%). 1 H NMR (500 MHz, DMSO-d6) δ 10.86 (s, 1H), 9.64 (s, 1H), 7.39 - 7.21 (m, 2H), 7.00 - 6.92 (m, 2H), 5.14 - 4.52 (m, 1H), 3.72 (d, J = 12.8 Hz, 2H), 3.04 - 2.86 (m, 4H), 2.87 - 2.63 (m, 2H), 2.61 - 2.51 (m, 2H), 2.44 - 2.30 (m, 1H), 2.03 - 1.84 (m, 3H), 1.66 - 1.46 (m, 2H).m / z(ES + ), [M+H] + 358.2. [ka]

[0257] The following are examples of additional building blocks for LHM targeting VHL that can be prepared in accordance with Scheme B3. HVB1:5-(((S)-1-((2S,4R)-4-hydroxy-2-((4-(4-methylthiazole-5-yl)benzyl)carbamoyl)pyrrolidine-1-yl)-3,3-dimethyl-1-oxobutan-2-yl)amino)-5-oxopentanoic acid [ka]

[0258] To a solution of glutaric acid (135 mg, 1.0 mmol) in THF (10 mL) and methanol (5 mL), HATU (0.39 g, 1.0 mmol) and N,N-diisopropylethylamine (0.33 mL, 1.9 mmol) were added. This reaction mixture was stirred for 5 minutes, and then (2S,4R)-1-[(2S)-2-amino-3,3-dimethylbutanoyl]-4-hydroxy-N-{[4-(4-methyl-1,3-thiazole-5-yl)phenyl]methyl}pyrrolidine-2-carboxamide (0.40 g, 0.93 mmol) was added. This reaction mixture was stirred for 16 hours, then quenched with 4N dioxane (0.25 mL), and the crude mixture was concentrated on silica gel and purified by reverse-phase chromatography. LCMS C 27 H 36 N4O6S Theoretical value: 544, Measured value: m / z = 567.5 [M + Na] + .

[0259] 1 H NMR (500 MHz, DMSO-d6) δ 12.01 (s, 1H), 9.00 (s, 1H), 8.58 (d, J = 6.4 Hz, 1H), 7.91 (d, J = 9.3 Hz, 1H), 7.43 (p, J = 7.8, 6.7 Hz, 4H), 5.14 (d, J = 3.7 Hz, 1H), 4.55 (d, J = 9.2 Hz, 1H), 4.53 - 4.43 (m, 2H), 4.37 (s, 1H), 4.23 (dd, J = 16.0, 5.2 Hz, 1H), 3.78 - 3.52 (m, 2H), 2.46 (s, 3H), 2.28 (dt, J = 15.7, 7.7 Hz, 1H), 2.25 - 2.15 (m, 3H), 2.05 (t, J = 10.6 Hz, 1H), 1.98 - 1.83 (m, 1H), 1.72 (h, J = 6.4 Hz, 2H), 0.95 (s, 9H). HVB2:(1r,4r)-4-{[(2S)-1-[(2S,4R)-4-hydroxy-2-({[4-(4-methyl-1,3-thiazole-5-yl)phenyl]methyl}carbamoyl)pyrrolidine-1-yl]-3,3-dimethyl-1-oxobutan-2-yl]carbamoyl}cyclohexane-1-carboxylic acid [ka]

[0259] To a solution of hexafluoro-lambda 5-phosphanoid (0.34 g, 0.89 mmol) and (1r,4r)-cyclohexane-1,4-dicarboxylic acid (154 mg, 0.89 mmol) stirred in [(dimethylamino)({[1,2,3]triazolo[4,5-b]pyridine-3-yloxy})methylidene]dimethylazanium; THF:DCM (in a 1:2 ratio) and N,N-diisopropylethylamine (0.26 g, 2.0 mmol), (2S,4R)-1-[(2S)-2-amino-3,3-dimethylbutanoyl]-4-hydroxy-N-{[4-(4-methyl-1,3-thiazole-5-yl)phenyl]methyl}pyrrolidine-2-carboxamide (350 mg, 0.81 mmol) was added and stirred for 16 hours. Next, the reaction product was quenched with 4N HCl in excess dioxane and then concentrated on silica gel. Reverse-phase column chromatography (0-100% acetonitrile in water) yielded (1r,4r)-4-{[(2S)-1-[(2S,4R)-4-hydroxy-2-({[4-(4-methyl-1,3-thiazole-5-yl)phenyl]methyl}carbamoyl)pyrrolidine-1-yl]-3,3-dimethyl-1-oxobutan-2-yl]carbamoyl}cyclohexane-1-carboxylic acid (0.17 g, 36%). LCMS:C 30 H 40 N4O6S Theoretical value: 584.73, Measured value: m / z = 607.6 [M + Na] + . HVB3:(2S,4R)-1-[(2S)-3,3-dimethyl-2-{[(1r,4r)-4-formylcyclohexyl]formamide}butanoyl]-4-hydroxy-N-{[4-(4-methyl-1,3-thiazole-5-yl)phenyl]methyl}pyrrolidine-2-carboxamide [ka]

[0260] Step 1: (2S,4R)-1-[(2S)-3,3-dimethyl-2-{[(1r,4r)-4-(hydroxymethyl)cyclohexyl]formamide}butanoyl]-4-hydroxy-N-{[4-(4-methyl-1,3-thiazole-5-yl)phenyl]methyl}pyrrolidine-2-carboxamide. [(dimethylamino)({[1,2,3]triazolo[4,5-b]pyridine-3-yloxy})methylidene]dimethylazanium; hexafluoro-lambda 5-phosphanoid (0.34 g, 0.89 mmol) and (1r,4r)-4-(hydroxymethyl)cyclohexane-1-carboxylic acid (141 mg, 0.89 mmol) were stirred in THF:DCM (1:2 ratio) and N,N-diisopropylethylamine (0.26 g, 2.0 mmol), to which (2S,4R)-1-[(2S)-2-amino-3,3-dimethylbutanoyl]-4-hydroxy-N-{[4-(4-methyl-1,3-thiazole-5-yl)phenyl]methyl}pyrrolidine-2-carboxamide (350 mg, 0.81 mmol) was added and stirred for 16 hours. Next, this reaction was quenched with 4N HCl in 2 drops of dioxane, and then concentrated on silica gel. Reverse-phase chromatography (0-100% acetonitrile in water) yielded (2S,4R)-1-[(2S)-3,3-dimethyl-2-{[(1r,4r)-4-(hydroxymethyl)cyclohexyl]formamide}butanoyl]-4-hydroxy-N-{[4-(4-methyl-1,3-thiazole-5-yl)phenyl]methyl}pyrrolidine-2-carboxamide (0.33 g, 71%). LCMS:C 30 H 42 N4O5S Theoretical value: 570.8, Measured value: m / z = 571.6 [M+H] + .

[0261] Step 2: (2S,4R)-1-[(2S)-3,3-dimethyl-2-{[(1r,4r)-4-formylcyclohexyl]formamide}butanoyl]-4-hydroxy-N-{[4-(4-methyl-1,3-thiazole-5-yl)phenyl]methyl}pyrrolidine-2-carboxamide. (2S,4R)-1-[(2S)-3,3-dimethyl-2-{[(1r,4r)-4-(hydroxymethyl)cyclohexyl]formamide}butanoyl]-4-hydroxy-N-{[4-(4-methyl-1,3-thiazole-5-yl)phenyl]methyl}pyrrolidine-2-carboxamide (330 mg, 0.58 mmol) was dissolved in DCM (0.1 M), and then 1,1-bis(acetyloxy)-3-oxo-1-lambda5,2-benzoiodaoxol-1-yl acetate (0.3 g, 0.7 mmol) was added. The reaction mixture was stirred for 2 hours, then filtered through Celite and concentrated on silica gel. Chromatography (0-10% methanol in DCM) yielded (2S,4R)-1-[(2S)-3,3-dimethyl-2-{[(1r,4r)-4-formylcyclohexyl]formamide}butanoyl]-4-hydroxy-N-{[4-(4-methyl-1,3-thiazole-5-yl)phenyl]methyl}pyrrolidine-2-carboxamide (0.2 g, 61%) as a white solid. LCMS:C 30 H 40 N4O5S Theoretical value: 568.7, Measured value: m / z = 569.6 [M+H] + . Scheme B4 demonstrates another method for generating building blocks of LHMs targeting VHLs via different binding points to LHMs: [ka]

[0262] Scheme B4 begins with coupling the linker precursor to a VHL-targeted LHM, namely (2S,4R)-1-[(2S)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxy-N-{[2-hydroxy-4-(4-methyl-1,3-thiazole-5-yl)phenyl]methyl}pyrrolidine-2-carboxamide. The VHL-targeted LHM is prepared according to the following steps. [ka]

[0263] Step 1: 2-Hydroxy-4-(4-methyl-1,3-thiazole-5-yl)benzonitrile. A solution of 4-bromo-2-hydroxybenzonitrile (25 g, 126.25 mmol), 4-methylthiazole (25.035 g, 252.5 mmol, 2.0 equivalents), and anhydrous KOAc (24.78 g, 252.5 mmol) in DMF (210.42 mL, 0.6 M) was barbotated with argon in an ultrasonic bath for 10 minutes. Next Pd(OAc)2 (0.567 g, 2.52 mmol) was added. The mixture was stirred under argon at 110°C for 5 hours, with additional amounts of Pd(OAc)2 (0.283 g, 1.26 mmol) added three times at 1-hour intervals (total amount of Pd(OAc)2 (1.417 g, 6.31 mmol)). The reaction mixture was cooled to room temperature, filtered through Celite, diluted with water, and extracted with ethyl acetate. The combined organic layers were washed with brine, dehydrated with anhydrous sodium sulfate, and concentrated under reduced pressure. Purification of the residue by silica gel column chromatography (DCM / MeOH) yielded 2-hydroxy-4-(4-methyl-1,3-thiazole-5-yl)benzonitrile (17.64 g, 64.6%) as a yellow solid. LCMS:C 11 H8N2OS Theoretical value: 216.3, Measured value: m / z = 217.49 [M+H] + ; 1 1H NMR (300 MHz, DMSO-d6) δ 11.36 (s, 1H), 9.08 (s, 1H), 7.71 (d, J = 8.1 Hz, 1H), 7.14 (d, J = 1.6 Hz, 1H), 7.08 (dd, J = 8.0, 1.7 Hz, 1H), 2.50 (s, 3H).

[0264] Step 2: 2-(aminomethyl)-5-(4-methyl-1,3-thiazole-5-yl)phenol. To a solution of LAH (1M in THF) (203.9 mL, 203.92 mmol), a solution of 2-hydroxy-4-(4-methyl-1,3-thiazole-5-yl)benzonitrile (17.64 g, 81.57 mmol) in THF (203.92 mL, 0.4 M) was slowly added under argon at -10°C. After the addition was complete, the reaction mixture was slowly brought to room temperature for 5 hours. Na2SO 4· The reaction was quenched by adding 10H2O and concentrated under reduced pressure. Purification of the residue by silica gel column chromatography (DCM / MeOH) yielded 2-(aminomethyl)-5-(4-methyl-1,3-thiazole-5-yl)phenol (9.18 g, 52%) as an amber-colored oil. LCMS:C 11 H 12 N2OS Theoretical value: 220.3, Measured value: m / z = 221.5 [M+H] + ; 1 H NMR (300 MHz, DMSO-d6) δ 8.96 (s, 1H), 7.23 - 7.15 (m, 1H), 6.87 - 6.81 (m, 2H), 3.88 (s, 2H), 2.45 (s, 3H).

[0265] Step 3: Methyl(2S,4R)-1-[(2S)-2-{[(tert-butoxy)carbonyl]amino}-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-carboxylate. When HATU (70.8g, 0.186mol) was added in small quantities as a solid at 10°C to a solution of methyl(2S)-2-{[(tert-butoxy)carbonyl]amino}-3,3-dimethylbutanoic acid (41.0g, 0.177mol) and DIPEA (46.3mL, 0.266mol) in anhydrous THF (1770mL, 0.1M), an activated ester was formed within 30 minutes. In a separate reactor, a solution of (2S,4R)-4-hydroxypyrrolidine-2-carboxylate hydrochloride (48.0 g, 1.266 mol) and DIPEA (46.3 mL, 0.266 mol, 1.5 equivalents) was prepared and cooled to -45°C under an inert atmosphere. The activated ester solution was added dropwise over 0.5 hours at -45 to 40°C, and the reaction mixture was slowly warmed to room temperature overnight. Water (approximately 500 mL) was added in one batch to quench the reaction, and the volatiles were concentrated under vacuum. The oily residue was extracted with HCl (3 × 400 mL), washed with saturated aqueous NaHCO3 (250 mL), 10% aqueous KHSO4 (250 mL), and brine (300 mL), dehydrated with MgSO4, filtered, and the solvent evaporated to obtain the crude product, which was purified by FC. Concentration of the corresponding fraction yielded methyl(2S,4R)-1-[(2S)-2-{[(tert-butoxy)carbonyl]amino}-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-carboxylate as a pale yellow oil (64 g, 99%). LCMS:C 17 H 30 Theoretical value of N2O6: 358.44, Measured value: m / z = 359.3 [M + H] + ; 1 H NMR (300 MHz, DMSO-d6) δ 6.54 (d, J = 9.3 Hz, 1H), 5.23 (d, J = 3.8 Hz, 1H), 4.42 - 4.29 (m, 2H), 4.16 (d, J = 9.4 Hz, 1H), 3.71 - 3.61 (m, 2H), 2.11 (dd, J = 12.2, 9.2 Hz, 1H), 1.95 - 1.85 (m, 1H), 1.38 (s, 10H), 0.94 (s, 9H).

[0266] Step 4: (2S,4R)-1-[(2S)-2-{[(tert-butoxy)carbonyl]amino}-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-carboxylic acid. To a solution of methyl (2S,4R)-1-[(2S)-2-{[(tert-butoxy)carbonyl]amino}-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-carboxylate (63.54 g, 0.177 mol) in THF (220 mL, 0.8 M), LiOH·H2O (14.88 g, 0.355 mol) was immediately added as an aqueous solution (86 mL, 0.2 M) at room temperature. The reaction mixture was stirred at room temperature for 3 hours and monitored by TLC / UPLC. Once the reaction was complete, 10% aqueous KHSO4 was added until the pH was approximately 3. THF was concentrated using a rotary evaporator, and the residue was extracted with ELISA (3 × 400 mL). The combined organic fraction was washed with 10% aqueous KHSO4 (200 mL) and brine (300 mL), dehydrated with MgSO4, filtered, and evaporated to dryness. The highly viscous, pale yellow oily residue was sonicated with anhydrous THF (300 mL) to obtain an off-white precipitate. This precipitate was filtered and dried under vacuum at 50°C to obtain 69.6 g of (2S,4R)-1-[(2S)-2-{[(tert-butoxy)carbonyl]amino}-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-carboxylic acid (69.6 g, containing approximately 15 wt% THF). LCMS:C 16 H 28 Theoretical value of N2O6: 344.4, Measured value: m / z = 345.2 [M + H] + ; 1 H NMR (300 MHz, DMSO-d6) δ 12.43 (s, 1H), 6.49 (d, J = 9.4 Hz, 1H), 5.18 (d, J = 3.7 Hz, 1H), 4.33 (bs, 1H), 4.26 (t, J = 8.4 Hz, 1H), 4.16 (d, J = 9.4 Hz, 1H), 3.69-3.52 (m, 2H),2.18 - 2.02 (m, 1H), 1.89 (ddd, J = 13.2, 9.1, 4.6 Hz, 1H), 1.38 (s, 9H), 0.94 (s, 9H).

[0267] Step 5: tert-butyl N-[(2S)-1-[(2S,4R)-4-hydroxy-2-({[2-hydroxy-4-(4-methyl-1,3-thiazole-5-yl)phenyl]methyl}carbamoyl)pyrrolidine-1-yl]-3,3-dimethyl-1-oxobutan-2-yl]carbamate. To a solution of (2S,4R)-1-[(2S)-2-{[(tert-butoxy)carbonyl]amino}-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-carboxylic acid (14.352 g, 41.67 mmol) in DMF (138.9 mL, 0.3 M), cooled in an ice bath, DIPEA (10.89 mL, 62.51 mmol) and HATU (16.644 g, 43.76 mmol) were added under argon. The resulting mixture was allowed to rise to room temperature for 0.5 hours, and then slowly added dropwise to a solution of 2-(aminomethyl)-5-(4-methyl-1,3-thiazole-5-yl)phenol (9.180 g, 41.67 mmol) and DIPEA (7.26 mL, 42.67 mmol) in DMF (83.34 mL, 0.5 M) under argon at -40°C. After addition, the reaction mixture was placed in a cooling bath and slowly brought to room temperature over 5 hours. The reaction was quenched by adding 5 mL of water and concentrated under reduced pressure. Purification of the residue by silica gel flash chromatography (DCM / MeOH) yielded (2S,4R)-1-[(2S)-2-{[(tert-butoxy)carbonyl]amino}-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-carboxylic acid (13.36 g, 58.64%) as a yellow solid. LCMS:C 27 H 38 N4O6S Theoretical value: 546.7, Measured value: m / z = 547.9 [M+H] +After purification by flash chromatography, a double acylation byproduct, 2-({[(2S,4R)-1-[(2S)-2-{[(tert-butoxy)carbonyl]amino}-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}methyl)-5-(4-methyl-1,3-thiazole-5-yl)phenyl(2S)-1-(2-{[(tert-butoxy)carbonyl]amino}-3,3-dimethylbutanoyl)pyrrolidine-2-carboxylate), was also obtained. The acyl group can be cleaved according to step 5a. 1 H NMR (300 MHz, Chloroform-d) δ 9.28 (br s, 1H), 8.70 (s, 1H), 8.11 (t, J = 6.6 Hz, 1H), 7.13 (d, J = 7.8 Hz, 1H), 6.98 (d, J = 1.8 Hz, 1H), 6.88 (dd, J = 7.7, 1.8 Hz, 1H), 5.19 (d, J = 8.9 Hz, 1H), 4.77 (t, J = 7.9 Hz, 1H), 4.51 (dd, J = 15.0, 6.9 Hz, 2H), 4.12 (td, J = 20.4, 8.4 Hz, 3H), 3.57 (dd, J = 11.4, 3.6 Hz, 1H), 2.85 (br s, 2H), 2.53 (m, 4H), 2.11 (dd, J = 13.5, 8.1 Hz, 1H), 1.56 - 1.43 (m, 2H), 1.41 (s, 9H), 0.84 (s, 9H).

[0268] Step 5a: tert-butyl N-[(2S)-1-[(2S,4R)-4-hydroxy-2-({[2-hydroxy-4-(4-methyl-1,3-thiazole-5-yl)phenyl]methyl}carbamoyl)pyrrolidine-1-yl]-3,3-dimethyl-1-oxobutan-2-yl]carbamate. 2-({[(2S,4R)-1-[(2S)-2-{[(tert-butoxy)carbonyl]amino}-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}methyl)-5-(4-methyl-1,3-thiazole-5-yl)phenyl(2S)-1-(2-{[(tert-butoxy)carbonyl]amino}-3,3-dimethylbutanoyl)pyrrolidine-2-carboxylate (3 g, 3.5 mmol) was dissolved in MeOH (70 mL, 0.05 M) and K2CO3 (0.484 g, 3.5 mmol) was added. The reaction mixture was stirred at room temperature for 12 hours. The reaction mixture was concentrated, the residue was diluted with water and neutralized with KHSO4, extracted by DCM (3 times), and the resulting organic layer was dried under Na2SO4 and concentrated under reduced pressure. The resulting residue was purified by silica gel flash chromatography (5% DCM / MeOH) to obtain tert-butyl N-[(2S)-1-[(2S,4R)-4-hydroxy-2-({[2-hydroxy-4-(4-methyl-1,3-thiazole-5-yl)phenyl]methyl}carbamoyl)pyrrolidine-1-yl]-3,3-dimethyl-1-oxobutan-2-yl]carbamate (2.14 g, 99%) as a yellow solid. LCMS:C 27 H 38 N4O6S Theoretical value: 546.7, Measured value: m / z = 547.2 [M+H] + ; 1 H NMR (300 MHz, Chloroform-d) δ 9.29 (s, 1H), 8.80 (s, 1H), 8.19 (s, 1H), 7.14 (d, J = 7.8 Hz, 1H), 6.98 (d, J = 1.8 Hz, 1H), 6.87 (dd, J = 7.7, 1.8 Hz, 1H), 5.14 (d, J = 8.9 Hz, 1H), 4.81 (t, J = 7.9 Hz, 1H), 4.56 (q, J = 7.8 Hz, 2H), 4.12 (td, J = 13.6, 12.6, 4.7 Hz, 3H), 3.56 (dd, J = 11.4, 3.5 Hz, 1H), 2.56 (s, 4H), 2.19 - 2.05 (m, 1H), 0.83 (s, 10H).

[0269] Step 6: (2S,4R)-1-[(2S)-2-amino-3,3-dimethylbutanoyl]-4-hydroxy-N-{[2-hydroxy-4-(4-methyl-1,3-thiazole-5-yl)phenyl]methyl}pyrrolidine-2-carboxamide. To a solution of tert-butyl N-[(2S)-1-[(2S,4R)-4-hydroxy-2-({[2-hydroxy-4-(4-methyl-1,3-thiazole-5-yl)phenyl]methyl}carbamoyl)pyrrolidine-1-yl]-3,3-dimethyl-1-oxobutan-2-yl]carbamate (2.14 g) (5.27 g, 9.64 mmol) in DCM (48.2 mL, 0.2 M), which had been cooled in an ice bath, 2 M HCl (38.56 mL, 77.12 mmol) in Et2O was added. The reaction mixture was stirred at room temperature for 2 hours. The solid was powdered in an ultrasonic bath and filtered off, washed on a filter using DCM, and dried under vacuum to obtain (2S,4R)-1-[(2S)-2-amino-3,3-dimethylbutanoyl]-4-hydroxy-N-{[2-hydroxy-4-(4-methyl-1,3-thiazole-5-yl)phenyl]methyl}pyrrolidine-2-carboxamide (5.05 g, 99%) as a white solid. LCMS:C 22 H 30 N4O4S Theoretical value: 446.6, Measured value: m / z = 447.7 [M+H] + ; 1H NMR (300 MHz, D2O) δ 9.50 (d, J = 1.0 Hz, 1H), 7.30 (d, J = 7.8 Hz, 1H), 7.04 - 6.89 (m, 2H), 4.58 (dd, J = 9.9, 7.6 Hz, 1H), 4.52 (s, 1H), 4.44 - 4.23 (m, 2H), 4.08 (s, 1H), 3.80 (d, J = 11.9 Hz, 1H), 3.68 (dd, J = 11.9, 3.4 Hz, 1H), 3.46 (q, J = 7.1 Hz, 1H), 2.45 (s, 3H), 2.28 (dd, J = 13.9, 7.7 Hz, 1H), 2.01 (ddd, J = 14.0, 9.9, 4.2 Hz, 1H), 1.08 (t, J = 7.1 Hz, 2H), 0.98 (s, 9H).

[0270] Step 7: (2S,4R)-1-[(2S)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxy-N-{[2-hydroxy-4-(4-methyl-1,3-thiazole-5-yl)phenyl]methyl}pyrrolidine-2-carboxamide. To a solution of 1-fluorocyclopropane-1-carboxylic acid (1.337 g, 12.85 mmol) in DMF (128 mL, 0.1 M), cooled in an ice bath, HATU (5.129 g, 13.49 mmol) and DIPEA (3.36 mL, 19.27 mmol) were added. The resulting mixture was allowed to rise to room temperature for 0.5 hours, and then added dropwise at -40°C to a solution of (2S,4R)-1-[(2S)-2-amino-3,3-dimethylbutanoyl]-4-hydroxy-N-{[2-hydroxy-4-(4-methyl-1,3-thiazole-5-yl)phenyl]methyl}pyrrolidine-2-carboxamide (5.05 g, quantitative yield) (6.674 g, 12.85 mmol) and DIPEA (7.83 mL, 44.97 mmol) in DMF (42 mL, 0.3 M). After the addition, the reaction mixture was placed in a cooling bath and slowly brought to room temperature over 16 hours. Next, the reaction product was diluted with water and extracted with ethyl acetate. The combined organic layers were washed with brine, dehydrated with anhydrous sodium sulfate, and concentrated under reduced pressure. Purification of the residue by silica gel column chromatography (DCM / MeOH) yielded (2S,4R)-1-[(2S)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxy-N-{[2-hydroxy-4-(4-methyl-1,3-thiazole-5-yl)phenyl]methyl}pyrrolidine-2-carboxamide (5.05 g, 74%) as a yellow solid. LCMS:C 26 H 33 N4O5SF Theoretical value: 532.6, Measured value: m / z = 533.8 [M+H] + ; 1 H NMR (300 MHz, Chloroform-d) δ 9.29 (s, 1H), 8.70 (s, 1H), 8.09 (dd, J = 7.5, 5.5 Hz, 1H), 7.13 (d, J = 7.8 Hz, 1H), 7.01 (dd, J = 8.5, 3.7 Hz, 1H), 6.98 (d, J = 1.8 Hz, 1H), 6.88 (dd, J = 7.7, 1.8 Hz, 1H), 4.73 (t, J = 7.9 Hz, 1H), 4.53 (br s, 1H), 4.51 - 4.40 (m, 2H), 4.18 (dd, J = 14.6, 5.4 Hz, 1H), 3.99 (d, J = 11.3 Hz, 1H), 3.63 (dd, J = 11.2, 3.7 Hz, 1H), 2.53 (s, 3H), 2.47 (ddd, J = 12.9, 7.9, 4.6 Hz, 1H), 2.15 - 2.01 (m, 1H), 1.36 - 1.22 (m, 4H), 0.91 (s, 9H).

[0271] The following are examples of additional building blocks for LHM targeting VHL that can be prepared in accordance with Scheme B4. HVB4:6-(2-(((2S,4R)-1-((S)-2-(1-fluorocyclopropane-1-carboxamide)-3,3-dimethylbutanoyl)-4-hydroxypyrrolidine-2-carboxamide)methyl)-5-(4-methylthiazole-5-yl)phenoxy)hexanoic acid [ka]

[0272] Step 1: tert-butyl 6-[2-({[(2S,4R)-1-[(2S)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}methyl)-5-(4-methyl-1,3-thiazole-5-yl)phenoxy]hexanoate. (2S,4R)-1-[(2S)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxy-N-{[2-hydroxy-4-(4-methyl-1,3-thiazole-5-yl)phenyl]methyl}pyrrolidine-2-carboxamide (1.29 g, 2.42 mmol, 1.0 equivalent) in anhydrous DMF (16 mL, 0.15 M) was mixed with Cs2CO3 (1.184 g, 3.63 mmol, 1.5 equivalents) and tert-butyl 6-bromohexanoate (CAS 65868-63-5, 0.85 g, 3.4 mmol, 1.4 equivalents). The reaction mixture was purged with argon, sealed, and stirred at 25°C for 16 hours. The solid was filtered, washed with ELISA (5 mL), and discarded. The resulting filtrate was diluted with water (60 mL) and extracted with ethyl acetate (3 × 20 mL). The combined organic layers were washed with brine (20 mL), dehydrated with Na₂SO₄, filtered, and concentrated under reduced pressure to obtain the crude product. This was purified by flash chromatography (hexane / ethyl acetate) to obtain 1.38 g of the desired product as a white solid (81.1% yield). ESI(+)[M+H] + =703.8

[0273] Step 2: 6-[2-({[(2S,4R)-1-[(2S)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}methyl)-5-(4-methyl-1,3-thiazole-5-yl)phenoxy]hexanoic acid. To a solution of tert-butyl 6-[2-({[(2S,4R)-1-[(2S)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}methyl)-5-(4-methyl-1,3-thiazole-5-yl)phenoxy]hexanoate (1.38 g, 1.96 mmol, 1.0 equivalent) in anhydrous DCM (147.3 mL, 0.4 M), HCl (2 M, 30 mL) in diethyl ether was added. The reaction mixture was stirred overnight at room temperature. The solvent was evaporated under reduced pressure to obtain a residue, which was dissolved in THF (10 mL), powdered with aqueous ammonia (3 M, 5 mL) for 10 minutes, and then concentrated again. The obtained crude product was purified by reverse-phase flash chromatography to yield 6-(2-(((2S,4R)-1-((S)-2-(1-fluorocyclopropane-1-carboxamide)-3,3-dimethylbutanoyl)-4-hydroxypyrrolidine-2-carboxamide)methyl)-5-(4-methylthiazole-5-yl)phenoxy)hexanoic acid (614 mg, 48%) as an off-white amorphous solid. LCMS 254 nm, RT=2.59 min, 95.62% purity, ESI(+)=647.13[M+H] + ; 1 H NMR (300 MHz, Methanol-d4) δ 8.86 (s, 1H), 7.50 (dd, J = 19.7, 9.1 Hz, 2H), 7.07 - 6.92 (m, 2H), 4.80 - 4.66 (m, 1H), 4.63 (t, J = 8.3 Hz, 1H), 4.50 (d, J = 3.2 Hz, 1H), 4.42 (d, J = 9.6 Hz, 1H), 4.07 (t, J = 6.2 Hz, 2H), 3.91 - 3.62 (m, 2H), 2.48 (s, 3H), 2.34 (t, J = 7.2 Hz, 2H), 2.27 - 2.02 (m, 2H), 1.87 (p, J = 6.6 Hz, 2H), 1.65 (dp, J = 33.1, 8.5, 7.8 Hz, 4H), 1.47 - 1.18 (m, 5H), 1.03 (s, 10H). HVB5:8-(2-(((2S,4R)-1-((S)-2-(1-fluorocyclopropane-1-carboxamide)-3,3-dimethylbutanoyl)-4-hydroxypyrrolidine-2-carboxamide)methyl)-5-(4-methylthiazole-5-yl)phenoxy)octanoic acid

[0274] HVB5 was prepared in the same manner as HVB4, except that hexanoic acid was replaced with octanoic acid, and the title compound was obtained. LCMS:C 34 H 47 FN4O7S Theoretical value: 674.3, Measured value: m / z = 672.7 [MH] - HVB6:10-(2-(((2S,4R)-1-((S)-2-(1-fluorocyclopropane-1-carboxamide)-3,3-dimethylbutanoyl)-4-hydroxypyrrolidine-2-carboxamide)methyl)-5-(4-methylthiazole-5-yl)phenoxy)decanoic acid [ka]

[0275] Step 1: tert-butyl 10-bromodecanoate. To a solution of 10-bromodecanoate (CAS: 50530-12-6, 10.0 g, 39.8 mmol, 1.0 equivalent) in anhydrous dichloromethane (0.25 M), tert-butyl alcohol (18.9 mL, 199 mmol, 5.0 equivalents) was added under nitrogen at 0°C, followed by DMAP (0.96 g, 4.0 mmol, 0.1 equivalents). After 5 minutes, dicyclohexylcarbodiimide (9.04 g, 44 mmol, 1.1 equivalents) was added to this solution at 0°C. The reaction mixture was warmed to room temperature and stirred for 20 hours. The volatiles were concentrated, and the crude product was then loaded directly onto silica (5-10% ethyl acetate in hexane). The desired product was isolated (9.0 g), which contained DCC as an impurity. 1 (By 1H NMR analysis). Further purification was performed by FC (eluent: 10-50% DCM in hexane), yielding 5.8 g of tert-butyl 10-bromodecanate (47% yield) as a colorless oil. 1 H NMR (300 MHz, Chloroform-d) δ 3.42 (t, J = 6.9 Hz, 2H), 2.22 (t, J = 7.5 Hz, 2H), 1.87 (p, J = 6.9 Hz, 2H), 1.68 - 1.51 (m, 2H), 1.46 (s, 9H), 1.45 - 1.37 (m, 2H), 1.31 (s, 8H).

[0276] Step 2: tert-butyl10-[2-({[(2S,4R)-1-[(2S)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}methyl)-5-(4-methyl-1,3-thiazole-5-yl)phenoxy]decanoate. (2S,4R)-1-[(2S)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxy-N-{[2-hydroxy-4-(4-methyl-1,3-thiazole-5-yl)phenyl]methyl}pyrrolidine-2-carboxamide (0.8 g, 1.5 mmol, 1.0 equivalent) in anhydrous DMF (15 mL, 0.1 M) was mixed with Cs2CO3 (0.734 g, 2.25 mmol, 1.5 equivalent) and tert-butyl 10-bromodecanate (0.646 g, 2.10 mmol, 1.4 equivalent). The reaction mixture was purged with argon, sealed, and stirred at 25°C for 16 hours. The solid was filtered, washed with ELISA (5 mL), and discarded. The resulting filtrate was diluted with water (60 mL) and extracted with ethyl acetate (3 × 20 mL). The combined organic layers were washed with brine (20 mL), dehydrated with Na₂SO₄, filtered, and concentrated under reduced pressure to obtain the crude product. This was purified by flash chromatography (hexane / ethyl acetate) to obtain 0.99 g of the desired product as a white solid (87.1% yield). ESI(+)[M+H] + =782.4; 1H NMR (300 MHz, Chloroform-d) δ 8.70 (s, 1H), 7.34 (d, J = 7.7 Hz, 1H), 7.25 (t, J = 5.9 Hz, 1H), 7.05 (dd, J = 8.7, 3.6 Hz, 1H), 6.96 (dd, J = 7.6, 1.6 Hz, 1H), 6.89 (d, J = 1.6 Hz, 1H), 4.76 (t, J = 7.7 Hz, 1H), 4.61 - 4.49 (m, 3H), 4.44 (dd, J = 14.8, 5.4 Hz, 1H), 4.09 - 3.97 (m, 3H), 3.64 (dd, J = 11.3, 3.9 Hz, 1H), 2.65 - 2.56 (m, 1H), 2.55 (s, 3H), 2.22 (t, J = 7.5 Hz, 2H), 2.15 (d, J = 2.6 Hz, 1H), 1.87 (p, J = 6.6 Hz, 2H), 1.59 (t, J = 7.1 Hz, 2H), 1.52 (m, 2H), 1.46 (s, 9H), 1.43 - 1.32 (m, 10H), 0.96 (s, 9H).

[0277] Step 3: 10-[2-({[(2S,4R)-1-[(2S)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}methyl)-5-(4-methyl-1,3-thiazole-5-yl)phenoxy]decanoic acid. To a solution of tert-butyl 10-[2-({[(2S,4R)-1-[(2S)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}methyl)-5-(4-methyl-1,3-thiazole-5-yl)phenoxy]decanoate (0.993 g, 1.31 mmol, 1.0 equivalent) in anhydrous DCM (6.5 mL, 0.2 M), TFA (2.00 mL, 26.17 mmol, 20 equivalents) was added. The reaction mixture was stirred at 25°C for 3 hours. The solvent was evaporated under vacuum, and the resulting oily substance was treated with aqueous ammonia (20%, 5 mL). After stirring for 1 hour, an oily substance was formed. The supernatant was decanted. The oily substance was dried under vacuum and purified using reverse-phase flash chromatography (20% to 60% acetonitrile / 0.1% formic acid aqueous solution) to obtain 0.703 g of the title compound (76.5% yield) as a white solid. LC-MS (254 nm): RT = 3.037 min, 100.00% purity, ESI [M+H] + =703.18; 1 H NMR (300 MHz, DMSO-d6) δ 12.00 (s, 1H), 8.99 (s, 1H), 8.51 (t, J = 5.9 Hz, 1H), 7.41 (d, J = 7.8 Hz, 1H), 7.31 (dd, J = 9.3, 2.9 Hz, 1H), 7.01 (d, J = 1.7 Hz, 1H), 6.96 (dd, J = 7.7, 1.6 Hz, 1H), 5.19 (s, 1H), 4.66 - 4.57 (m, 1H), 4.53 (t, J = 8.2 Hz, 1H), 4.36 (s, 1H), 4.25 (qd, J = 16.7, 5.9 Hz, 2H), 4.05 (t, J = 6.3 Hz, 2H), 3.73 - 3.56 (m, 2H), 2.47 (s, 3H), 2.19 (t, J = 7.3 Hz, 2H), 2.15 - 2.09 (m, 1H), 1.93 (ddd, J = 13.0, 8.9, 4.5 Hz, 1H), 1.76 (p, J = 6.4 Hz, 2H), 1.57 - 1.38 (m, 6H), 1.38 - 1.15 (m, 12H), 0.97 (s, 9H). HVB7:3-{2-[2-({[(2S,4R)-1-[(2S)-2-[(1-fluorocyclopropyl)formamide]-3-methylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}methyl)-5-(4-methyl-1,3-thiazole-5-yl)phenoxy]ethoxy}propanoic acid [ka]

[0278] Step 1: tert-butyl 3-(2-bromoethoxy)propanoate. A solution of tert-butyl 3-(2-hydroxyethoxy)propanoate (3.0 g, 15.7 mmol, 1 equivalent) and carbon tetrabromide (3.9 g, 11.87 mmol, 1.5 equivalents) in dichloromethane (15 mL, 0.5 mL) was prepared in a 50 mL flask and cooled to 0°C. Triphenylphosphine (3.1 g, 11.87 mmol, 1.5 equivalents) was added in small portions from a powder funnel over 30 minutes while vigorously stirring. Upon addition of phosphine, the colorless solution turned light brown and was stirred for a further 2 hours at room temperature. This mixture was concentrated and quickly added to stirred hexane (50 mL). The white precipitate was filtered, the residual solution was concentrated, and the resulting residue was purified by FC (elution with DCM / MeOH -9 / 1) to obtain 4.1 g of the desired product (DP) as a white solid (yield 62.8%).

[0279] Step 2: tert-butyl 3-{2-[2-({[(2S,4R)-1-[(2S)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}methyl)-5-(4-methyl-1,3-thiazole-5-yl)phenoxy]ethoxy}propanoate. To a solution of (2S,4R)-1-[(2S)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxy-N-{[2-hydroxy-4-(4-methyl-1,3-thiazole-5-yl)phenyl]methyl}pyrrolidine-2-carboxamide (1.5 g, 2.82 mmol, 1.0 equivalent) in DMF (18.77 mL, 0.15 M), Cs2CO3 (1.376 g, 4.22 mmol, 1.5 equivalent) and tert-butyl 3-(2-bromoethoxy)propanoate (2.18 g, 3.94 mmol, 1.4 equivalent) were added. The resulting mixture was stirred overnight at room temperature. The reaction mixture was diluted with water and extracted with ELISA (three times). The organic layer was dried under Na2SO4 and concentrated. The residue was purified by elution with DCM / MeOH-9 / 1 and FC, yielding 1.8 g of the desired product as a pale yellow oily substance (quantitative yield). UPLC (12 min, 254 nm): RT=6.25 min, 100% purity, ESI[M+H] + ] + =705.55

[0280] Step 3: 3-{2-[2-({[(2S,4R)-1-[(2S)-2-[(1-fluorocyclopropyl)formamide]-3-methylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}methyl)-5-(4-methyl-1,3-thiazole-5-yl)phenoxy]ethoxy}propanoic acid. To a solution of tert-butyl 3-{2-[2-({[(2S,4R)-1-[(2S)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}methyl)-5-(4-methyl-1,3-thiazole-5-yl)phenoxy]ethoxy}propanoate (1.8 g, 2.64 mmol, 1 equivalent) in DCM (17.6 mL, 0.15 M), TFA (13.2 mL, 0.2 M) was added dropwise at 0°C. The reaction mixture was stirred at room temperature for 1 hour. The reaction mixture was concentrated, and the residue was diluted with 50 mL of aqueous NH4OH (to pH=11), and left in an ultrasonic bath for 0.5 hours, then simply stirred for 1 hour. The obtained slurry was concentrated and purified twice by RF: first, eluted with ACN / H2O to obtain 0.3 g of the desired product; second, eluted with ACN / H2O (0.1% formic acid) to obtain 1 g of the desired product. After neutralization with NH4OH, the product was obtained in the form of an ammonium salt, which was dissociated with formic acid during the second purification. Combining all the amounts, 1.3 g of the desired product (76% yield) was obtained. LCMS (254 nm): RT=2.29 min: 99% purity, ESI[M+H] + =649.10; 1 H NMR (300 MHz, Chloroform-d) δ 8.70 (s, 1H), 7.37 (d, J = 7.8 Hz, 2H), 7.09 - 7.03 (m, 1H), 6.99 (dd, J = 7.7, 1.6 Hz, 1H), 6.91 (d, J = 1.6 Hz, 1H), 4.76 (t, J = 8.1 Hz, 1H), 4.64 - 4.51 (m, 3H), 4.41 (dd, J = 14.3, 5.2 Hz, 1H), 4.20 (t, J = 4.2 Hz, 2H), 4.03 (d, J = 11.3 Hz, 1H), 3.89 (td, J = 8.6, 7.8, 4.4 Hz, 4H), 3.77 (dd, J = 11.3, 3.7 Hz, 1H), 2.66 (ddd, J = 19.7, 14.9, 5.1 Hz, 2H), 2.54 (s, 3H), 2.33 - 2.14 (m, 2H), 1.41 - 1.23 (m, 4H), 1.03 (s, 9H).

[0281] HVB8:1-(2-(((2S,4R)-1-((S)-2-(1-fluorocyclopropane-1-carboxamide)-3,3-dimethylbutanoyl)-4-hydroxypyrrolidine-2-carboxamide)methyl)-5-(4-methylthiazole-5-yl)phenoxy)-3,6,9,12,15-pentaoxaoctadecane-18-acid

[0282] HVB8 was prepared in a similar manner to HVB7 by replacing tert-butyl 3-{2-[2-(2-bromoethoxy)ethoxy]ethoxy}propanoate with tert-butyl 3-(2-hydroxyethoxy)propanoate in step 1, and the title compound was obtained as a white solid. LCMS (254 nm): RT=2.27 min, 96.35% purity, ESI[M+H] + = 736.88.

[0284] 1 H NMR (300 MHz, DMSO-d6) δ 8.98 (s, 1H), 8.51 (t, J = 6.0 Hz, 1H), 7.41 (d, J = 7.8 Hz, 1H), 7.31 (dd, J = 9.2, 2.9 Hz, 1H), 7.04 (d, J = 1.7 Hz, 1H), 6.97 (dd, J = 7.7, 1.6 Hz, 1H), 5.19 (s, 1H), 4.60 (d, J = 9.1 Hz, 1H), 4.51 (t, J = 8.2 Hz, 1H), 4.35 (s, 1H), 4.28 (d, J = 6.1 Hz, 1H), 4.25 - 4.14 (m, 3H), 3.79 (dd, J = 5.8, 3.4 Hz, 2H), 3.66 - 3.46 (m, 12H), 2.46 (s, 3H), 2.42 (t, J = 6.3 Hz, 2H), 2.10 (dd, J = 13.0, 8.0 Hz, 1H), 1.92 (ddd, J = 13.1, 9.0, 4.4 Hz, 1H), 1.49 - 1.28 (m, 2H), 1.21 (tq, J = 8.4, 4.6, 3.8 Hz, 2H), 0.96 (s, 9H).

[0283] HVB9:1-(2-(((2S,4R)-1-((S)-2-(1-fluorocyclopropane-1-carboxamide)-3,3-dimethylbutanoyl)-4-hydroxypyrrolidine-2-carboxamide)methyl)-5-(4-methylthiazole-5-yl)phenoxy)-3,6,9,12,15-pentaoxaoctadecane-18-ate tert-butyl

[0284] HVB9 was prepared in a similar manner to HVB7 by replacing 1-bromo-3,6,9,12,15-pentaoxaoctadecane-18-ate tert-butyl with 3-(2-hydroxyethoxy)propanoate tert-butyl in step 1, and the title compound was obtained as a white solid. LCMS (254 nm): RT=2.27 min, 99.8% purity, ESI(+)[M+H] + =825.21

[0287] 1H NMR (300 MHz, DMSO-d6) δ 8.98 (s, 1H), 8.50 (t, J = 6.0 Hz, 1H), 7.41 (d, J = 7.8 Hz, 1H), 7.29 (dd, J = 9.4, 2.9 Hz, 1H), 7.04 (d, J = 1.7 Hz, 1H), 6.96 (dd, J = 7.8, 1.6 Hz, 1H), 4.60 (d, J = 9.2 Hz, 1H), 4.51 (t, J = 8.2 Hz, 1H), 4.35 (s, 1H), 4.28 (d, J = 6.0 Hz, 1H), 4.24 - 4.11 (m, 3H), 3.79 (dd, J = 5.5, 3.7 Hz, 2H), 3.67 - 3.42 (m, 22H), 2.46 (s, 3H), 2.42 (t, J = 6.4 Hz, 2H), 2.13 - 2.03 (m, 1H), 1.96 - 1.86 (m, 1H),1.46 - 1.27 (m, 2H), 1.23 (dq, J = 8.6, 4.1 Hz, 2H), 0.96 (s, 9H). HVB10:3-{[(2S)-1-[(2S,4R)-4-hydroxy-2-{[(1S)-1-[4-(4-methyl-1,3-thiazole-5-yl)phenyl]ethyl]carbamoyl}pyrrolidine-1-yl]-3,3-dimethyl-1-oxobutan-2-yl]carbamoyl}propanoic acid [ka]

[0285] To a solution of [(dimethylamino)({[1,2,3]triazolo[4,5-b]pyridine-3-yloxy})methylidene]dimethylazanium;hexafluoro-lambda 5-phosphanoid (1.41 g, 3.71 mmol) and succinic acid (398 mg, 3.37 mmol) stirred in THF:DCM (1:2 ratio), (2S,4R)-1-[(2S)-2-amino-3,3-dimethylbutanoyl]-4-hydroxy-N-{[4-(4-methyl-1,3-thiazole-5-yl)phenyl]methyl}pyrrolidine-2-carboxamide (1.5 g, 3.37 mmol) was added. N,N-diisopropylethylamine (0.72 mL, 8.43 mmol) was added, and the reaction mixture was stirred for 16 hours. Next, the reaction was quenched with 4N HCl in excess dioxane and then concentrated on silica gel. The title compound was obtained by reverse-phase column chromatography (0-100% acetonitrile in water). LC-MS:C 27 H 36 N4O6S Theoretical value: 544.24, Measured value: m / z = 545.6 [M+H] + . HVB11:3-[3-[[(1S)-1-[(2S,4R)-4-hydroxy-2-[[(1S)-1-[4-(4-methylthiazole-5-yl)phenyl]ethyl]carbamoyl]pyrrolidine-1-carbonyl]-2,2-dimethylpropyl]amino]-3-oxopropoxy]propanoic acid [ka]

[0286] To a 30 mL solution of 3-(2-carboxyethoxy)propanoic acid (1.5 g, 9.4 mmol) and HATU (2.6 g, 6.9 mmol) in DCM (30 mL), DIPEA (5.3 mL, 31 mmol) was slowly added, and the solution was stirred at room temperature for 5 minutes. To this mixture, (2S,4R)-1-[(2S)-2-amino-3,3-dimethyl-butanoyl]-4-hydroxy-N-[(1S)-1-[4-(4-methylthiazole-5-yl)phenyl]ethyl]pyrrolidine-2-carboxamide hydrochloride 2 (3.0 g, 6.2 mmol) was added, and the reaction mixture was stirred for 30 minutes. This mixture was diluted with 1 M NaOH (5.0 mL) and stirred for 5 minutes. Next, the mixture was acidified to pH 5 using 5% citric acid. The layers were separated, and the aqueous layer was extracted with RINKAN (7 × 50 mL) and DCM (3 × 50 mL). The combined organic layers were dried (sodium sulfate), filtered, and concentrated under reduced pressure. The substance was purified by C18 reverse-phase chromatography using a 10–30% gradient of MeCN and water (containing 0.1% ammonium formate / formic acid), and the title compound was obtained as a solid (1.28 g, 35%). MS(ESI)[M+H] + = 589.3. 1 H NMR (500 MHz, DMSO) δ 8.99 (s, 1H), 8.39 (d, J = 7.8 Hz, 1H), 7.87 (d, J = 9.3 Hz, 1H), 7.47 - 7.41 (m, 2H), 7.39 (s, 2H), 4.92 (p, J = 7.0 Hz, 1H), 4.53 (d, J = 9.4 Hz, 1H), 4.44 (t, J = 8.0 Hz, 1H), 4.28 (s, 1H), 3.65 - 3.49 (m, 6H), 2.46 (s, 3H), 2.37 (t, J = 6.7 Hz, 2H), 2.39 - 2.31 (m, 1H), 2.05 - 1.99 (m, 1H), 1.80 (ddd, J = 12.9, 8.4, 4.7 Hz, 1H), 1.37 (t, J = 8.2 Hz, 3H), 0.94 (s, 9H). HVB12:4-[2-({[(2S,4R)-1-[(2S)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}methyl)-5-(4-methyl-1,3-thiazole-5-yl)phenoxy]butanoic acid [ka]

[0287] Step 1: Ethyl 4-[2-({[(2S,4R)-1-[(2S)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}methyl)-5-(4-methyl-1,3-thiazole-5-yl)phenoxy]butanoate. (2S,4R)-1-[(2S)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxy-N-{[2-hydroxy-4-(4-methyl-1,3-thiazole-5-yl)phenyl]methyl}pyrrolidine-2-carboxamide (50.00 mg, 0.09 mmol) and potassium carbonate (20.73 mg, 0.15 mmol) were combined and suspended in N,N-dimethylformamide (2.00 mL). Ethyl 4-bromobutanoate (0.02 mL, 21.97 mg, 0.11 mmol) was added and the mixture was stirred at room temperature for 3 days. The mixture was quenched with water and extracted with ethyl acetate. The mixture was washed twice more with water, and then once with brine. The mixture was dehydrated with sodium sulfate, filtered, and concentrated. The reaction product was carried over to the next step as a crude product. 1 H NMR (500 MHz, Chloroform-d) δ 7.35 (d, J = 7.7 Hz, 1H), 6.99 (dd, J = 7.7, 1.6 Hz, 2H), 4.78 (t, J = 7.7 Hz, 2H), 4.63 - 4.47 (m, 3H), 4.47 - 4.38 (m, 1H), 4.10 (d, J = 5.7 Hz, 7H), 4.07 (s, 4H), 3.69 - 3.58 (m, 1H), 3.50 (t, J = 6.5 Hz, 5H), 2.20 (p, J = 6.8 Hz, 7H), 2.12 (s, 2H), 1.59 (s, 4H), 1.29 (t, J = 7.2 Hz, 13H), 0.96 (s, 8H).

[0288] Step 2: 4-[2-({[(2S,4R)-1-[(2S)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}methyl)-5-(4-methyl-1,3-thiazole-5-yl)phenoxy]butanoic acid. Ethyl 4-[2-({[(2S,4R)-1-[(2S)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}methyl)-5-(4-methyl-1,3-thiazole-5-yl)phenoxy]butanoate (50.00 mg, 0.08 mmol) was dissolved in tetrahydrofuran (2.00 mL) and water (0.50 mL), and lithium hydroxide hydrate (32.44 mg, 0.77 mmol) was added. The mixture was stirred at room temperature for 2 days. It was quenched with saturated ammonium chloride and extracted with ethyl acetate. It was washed with brine and then dehydrated with sodium sulfate. The solution was filtered and concentrated until it became a white solid, yielding 4-[2-({[(2S,4R)-1-[(2S)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}methyl)-5-(4-methyl-1,3-thiazole-5-yl)phenoxy]butanoic acid (0.0400 g, 83.6%). ESI theoretical value 618.25; measured value 641.7 (M+Na + ) HVB13:6-{[(2S)-1-[(2S,4R)-4-hydroxy-2-{[(1S)-1-[4-(4-methyl-1,3-thiazole-5-yl)phenyl]ethyl]carbamoyl}pyrrolidine-1-yl]-3,3-dimethyl-1-oxobutan-2-yl]carbamoyl}hexanoic acid [ka]

[0289] (2S,4R)-1-[(2S)-2-amino-3,3-dimethylbutanoyl]-4-hydroxy-N-[(1S)-1-[4-(4-methylthiazole-5-yl)phenyl]ethyl]pyrrolidine-2-carboxamide hydrochloride 2 (1.75 g, 3.64 mmol), heptanediol (874 mg, 5.46 mmol), and HATU (1.94 g, 5.09 mmol) were mixed in DCM (70.0 mL) and DIPEA (3.11 mL, 18.2 mmol) at 0°C. The reaction mixture was stirred for 2 hours. The mixture was diluted with 1 M NaOH (50 mL) and stirred for 1 hour. The layers were separated, and the organic layer was extracted with 1 M NaOH (2 × 30 mL). The combined aqueous layer was acidified to pH 5-6 and extracted with ELISA (5 × 50 mL). The combined organic layers were dried (sodium sulfate), filtered, and concentrated under reduced pressure. Further purification of the substance was performed by C18 reverse-phase chromatography using a 10–60% gradient of MeCN and water (containing 0.1% ammonium formate / formic acid), yielding the title compound as a solid (0.924 g, 43%). LCMS:C 30 H 42 N4O6S Theoretical value: 586.75, Measured value: m / z = 587.3 [M+H] + .

[0294] 1 H NMR (500 MHz, DMSO) δ 8.99 (s, 1H), 8.37 (d, J = 7.8 Hz, 1H), 7.79 (d, J = 9.3 Hz, 1H), 7.46 - 7.41 (m, 2H), 7.40 - 7.36 (m, 2H), 4.92 (p, J = 7.0 Hz, 1H), 4.52 (d, J = 9.4 Hz, 1H), 4.43 (t, J = 8.1 Hz, 1H), 4.30 - 4.26 (m, 1H), 3.65 - 3.57 (m, 2H), 3.46 - 3.33 (m, 1H), 2.46 (s, 3H), 2.28 - 2.20 (m, 1H), 2.18 (t, J = 7.4 Hz, 2H), 2.15 - 2.06 (m, 1H), 2.04 - 1.97 (m, 1H), 1.80 (ddd, J = 12.9, 8.5, 4.7Hz, 1H), 1.54 - 1.42 (m, 4H), 1.38 (d, J = 7.0 Hz, 3H), 1.28 - 1.20 (m, 2H), 0.94 (s, 9H). HVB14:(3S)-3-{[(2S,4R)-1-[(2S)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}-3-[4-(4-methyl-1,3-thiazole-5-yl)phenyl]propanoic acid [ka]

[0290] Step 1: (3S)-3-amino-3-(4-bromophenyl)propanoate methyl hydrochloride. To a solution of (3S)-3-(4-bromophenyl)-3-{[(tert-butoxy)carbonyl]amino}propanoic acid (8 g, 1.453 mmol, 1.0 equivalent) in MeOH (140 mL, 0.3 M), 3 M HCl (200 mL, 0.15 M) in MeOH cooled to 0°C was slowly added. This mixture was stirred at room temperature for 16 hours. The MeOH was then removed by evaporation under vacuum, and the resulting residue was powdered with Et2O to obtain the desired product, (3S)-3-amino-3-(4-bromophenyl)propanoate methyl hydrochloride (90% yield), as a foamy white solid: ESI(+)[M+H] +=257.9 and 259.9 (Br pattern); 1H NMR (300 MHz, DMSO-d6): 8.91 (d, J = 5.5 Hz, 3H), 7.62 (d, J = 8.5 Hz, 2H), 7.54 (d, J = 8.6 Hz, 2H), 4.57 (q, J = 5.5, 4.9 Hz, 1H), 3.54 (s, 3H), 3.25 (dd, J = 16.4, 5.6 Hz, 1H), 3.03 (dd, J = 16.3Hz, 9.0Hz, 1H). Step 2: Methyl(3S)-3-(4-bromophenyl)-3-{[(2S,4R)-1-[(2R)-2-{[(tert-butoxy)carbonyl]amino}-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}propanoate

[0291] 1. A mixture of (2S,4R)-1-[(2R)-2-{[(tert-butoxy)carbonyl]amino}-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-carboxylic acid (11.925 g, 34.628 mmol, 1.20 equivalents) and DIPEA (7.5 mL, in 1.5 equivalents of DMF (0.17 M)) was slowly added at 0°C with HATU (11.52 g, 30,298 mmol, 1.05 equivalents) dissolved in DMF (0.15 M). The reaction mixture was stirred at room temperature for 30 minutes.

[0292] 2. To a solution of (3S)-3-amino-3-(4-bromophenyl)propanoate methyl hydrochloride (8.5 g, 28.85 mmol, 1.0 equivalent) in DMF (55 mL, 0.6 M), DIPEA (20.11 mL, 4 equivalents) was added at -40°C and stirred for 5 minutes at -40°C.

[0293] 3. Reactant 1 was slowly added to Reactant 2 at -40°C. The mixture was stirred at room temperature for 16 hours.

[0294] The reaction mixture was diluted with water, then extracted with DCM, the organic layer was washed with brine, dried under Na2SO4, and the resulting crude product was purified by flash chromatography after elution with DCM / MeOH-9 / 1 to obtain the product methyl(3S)-3-(4-bromophenyl)-3-{[(2S,4R)-1-[(2R)-2-{[(tert-butoxy)carbonyl]amino}-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}propanoate (yield 71%) as a pale yellow solid. ESI(+)[M+H] + = 585.85.

[0300] 1 H NMR (300 MHz, DMSO-d6) δ 8.50 (d, J = 8.1 Hz, 1H), 7.52 - 7.46 (m, 2H), 7.30 - 7.21 (m, 2H), 6.45 (d, J = 9.2 Hz, 1H), 5.17 - 5.07 (m, 2H), 4.36 (t, J = 8.0 Hz, 1H), 4.25 (s, 1H), 4.12 (d, J = 9.3 Hz, 1H), 3.62-3.49 (m, 5H), 2.83 - 2.76 (m, 1H), 1.99-1.92 (m, 1H), 1.73-1.64 (m, 1H), 1.38 (s, 9H), 0.92 (s, 9H).

[0295] Step 3: Methyl(3S)-3-{[(2S,4R)-1-[(2R)-2-{[(tert-butoxy)carbonyl]amino}-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}-3-[4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenyl]propanoate. Bis(pinacorato)diborone (8.689g, 34.21mmol, 2 equivalents) dissolved in 285mL of 1,4-dioxane (0.06M), potassium acetate (5.037g, 51.32mmol, 3 equivalents), methyl(3S)-3-(4-bromophenyl)-3-{[(2S,4R)-1-[(2R)-2-{[(tert-butoxy)carbonyl]amino}-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2- [Iyl]formamide propanoate (10 g, 14.542 mmol, 17.10 mmol, 1 equivalent) was stirred with argon for a while, then [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium(II) complex with dichloromethane (1.39 g, 1.71 mmol, 0.1 equivalent) was added, and the mixture was further stirred with argon. The mixture was then placed in a preheated 95°C oil bath and stirred for 16 hours. The reaction mixture was concentrated, then redissolved in DCM, and purified by two flash chromatography steps using DCM / MeOH98 / 2 to obtain the product. Further flash purification after elution with hexane / siRNA-0 (>80%) yielded 8 g of the desired product, methyl(3S)-3-{[(2S,4R)-1-[(2R)-2-{[(tert-butoxy)carbonyl]amino}-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}-3-[4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenyl]propanoate (yield 71%), as a light brown foamy substance. ESI(+)[M+H] + =632.0; 1H NMR (300 MHz, DMSO-d6) δ 8.49 (d, J = 8.1 Hz, 1H), 7.65 - 7.57 (m, 2H), 7.30 (d, J = 8.0 Hz, 2H), 6.44 (d, J = 9.2 Hz, 1H), 5.22 - 5.05 (m, 2H), 4.43 - 4.34 (m, 1H), 4.25 (s, 1H), 4.13 (d, J = 9.4 Hz, 1H), 3.55 (s, 5H), 2.88 - 2.70 (m, 1H), 1.98 - 1.90 (m, 1H), 1.68 (ddd, J = 12.8, 8.2, 4.7 Hz, 1H), 1.38 (s, 9H), 1.28 (s, 12H), 0.93 (s, 9H).

[0296] Step 4: Methyl(3S)-3-{[(2S,4R)-1-[(2R)-2-{[(tert-butoxy)carbonyl]amino}-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}-3-[4-(4-methyl-1,3-thiazole-5-yl)phenyl]propanoate. 5-bromo-4-methylthiazole (3.383 g, 19.0 mmol, 1.5 equivalents), [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium(II) (complex with dichloromethane) (1.034 g, 1.266 mmol, 0.1 equivalents), potassium carbonate (5.02 g, 36.35 mmol, 2.87 equivalents), methyl(3S)-3-{[(2S,4R) The reaction mixture consisting of -1-[(2R)-2-{[(tert-butoxy)carbonyl]amino}-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}-3-[4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenyl]propanoate (8.0 g, 12.667 mmol, 1.0 equivalent) was stirred under argon for a while, and then placed in a preheated 100°C oil bath and stirred for 16 hours. Next, the reaction mixture was filtered through a Celite pad, the filtrate was concentrated, and purified by flash chromatography after elution with DCM / MeOH (10-30% MeOH). 4.2 g of the desired product, methyl(3S)-3-{[(2S,4R)-1-[(2R)-2-{[(tert-butoxy)carbonyl]amino}-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}-3-[4-(4-methyl-1,3-thiazole-5-yl)phenyl]propanoate (yield 51%), was obtained as a light brown solid. ESI(+)[M+H] + =589.3; 1H NMR (300 MHz, DMSO-d6) δ 8.99 (s, 1H), 8.55 (d, J = 7.9 Hz, 1H), 7.49 - 7.34 (m, 4H), 6.45 (d, J = 9.2 Hz, 1H), 5.15-5.08(m, 2H), 4.40 (t, J = 8.1 Hz, 1H), 4.27 (s, 1H), 4.14 (d, J = 9.3 Hz, 1H), 3.62 - 3.54 (m, 2H), 2.75 - 2.58 (m, 2H), 2.45 (s, 3H), 2.00 (d, J = 3.7Hz, 1H), 1.75 (s, 1H), 1.39 (s, 9H), 0.93 (s, 9H).

[0297] Step 5: Methyl(3S)-3-{[(2S,4R)-1-[(2R)-2-amino-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}-3-[4-(4-methyl-1,3-thiazole-5-yl)phenyl]propanoate hydrochloride. To a solution of methyl(3S)-3-{[(2S,4R)-1-[(2R)-2-{[(tert-butoxy)carbonyl]amino}-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}-3-[4-(4-methyl-1,3-thiazole-5-yl)phenyl]propanoate (4.9 g, 8.1 mmol, 1.0 equivalent) in MeOH (68 mL, 0.3 M), 3 M HCl (43.5 mL, 0.15 M) cooled at 0°C in MeOH was slowly added. The mixture was stirred at room temperature for 16 hours. Subsequently, MeOH was removed by evaporation under vacuum, and the resulting residue was powdered with Et2O to obtain the desired product, methyl(3S)-3-{[(2S,4R)-1-[(2R)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}-3-[4-(4-methyl-1,3-thiazole-5-yl)phenyl]propanoate hydrochloride (yield 70%), as a foamy white solid. ESI(+)[M+H] + =503.3; 1H NMR (300 MHz, DMSO-d6) δ 9.06 (s, 1H), 8.74 (d, J = 8.1 Hz, 1H), 8.10 (s, 4H), 7.43 (q, J = 8.4 Hz, 4H), 5.19 (d, J = 7.6 Hz, 1H), 4.51 (d, J = 8.4 Hz, 1H), 4.30 (s, 1H), 3.89 (d, J = 5.3 Hz, 1H), 3.73 (d, J = 11.0 Hz, 1H), 3.61 - 3.47 (m, 5H), 2.87-2.82 (m, 2H), 2.46 (s, 3H), 2.13 - 2.00 (m, 1H), 1.77-1.66 (m Hz, 1H), 1.02 (s, 9H). Step 6: Methyl(3S)-3-{[(2S,4R)-1-[(2R)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}-3-[4-(4-methyl-1,3-thiazole-5-yl)phenyl]propanoate

[0298] 1. To a mixture of 1-fluorocyclopropanecarboxylic acid (0.983 g, 9.45 mmol, 1.04 equivalents) and DIPEA (2.4 mL, 1.5 equivalents) in DMF (0.17 M), HATU (3.6 g, 9.5 mmol, 1.04 equivalents) dissolved in DMF (0.15 M) was slowly added at 0°C. The reaction mixture was stirred at room temperature for 30 minutes.

[0299] 2. To a solution of methyl(3S)-3-{[(2S,4R)-1-[(2R)-2-amino-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}-3-[4-(4-methyl-1,3-thiazole-5-yl)phenyl]propanoate (4.9 g, 9.2 mmol, 1.0 equivalent) in DMF (55 mL, 0.16 M), DIPEA (8.4 mL, 5 equivalents) was added at -40°C and the mixture was stirred at -40°C for 5 minutes.

[0300] 3. Reactant 1 was slowly added to Reactant 2 at -40°C. The mixture was stirred at room temperature for 16 hours.

[0301] The reaction mixture was diluted with water, then extracted with DCM, the organic layer was washed with brine, and dried under Na2SO4. The resulting crude product was purified by flash chromatography after elution with DCM / MeOH-9 / 1, yielding the product (3S)-3-{[(2S,4R)-1-[(2R)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}-3-[4-(4-methyl-1,3-thiazole-5-yl)phenyl]propanoic acid (yield 47%) as a pale yellow solid. ESI(+)[M+H] + = 589.35

[0308] 1 H NMR (300 MHz, DMSO-d6) δ 8.99 (s, 1H), 8.59 (d, J = 8.1 Hz, 1H), 7.44 (t, J = 7.5 Hz, 4H), 7.28 (dd, J = 9.2, 2.9 Hz, 1H), 5.25 - 5.11 (m, 2H), 4.57 (d, J = 9.2 Hz, 1H), 4.43 (t, J = 8.3 Hz, 1H), 4.27 (s, 1H), 3.63-3.52 (m, 5H), 2.86-2.81 (m, 1H), 2.46 (s, 3H), 2.08-2.00 (m, 1H), 1.79-1.68 (m, 1H), 1.42 - 1.29 (m, 2H), 1.24 - 1.18 (m, 2H), 0.96 (d, J = 6.4 Hz, 9H).

[0302] Step 7: (3S)-3-{[(2S,4R)-1-[(2R)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}-3-[4-(4-methyl-1,3-thiazole-5-yl)phenyl]propanoic acid. (3S)-3-{[(2S,4R)-1-[(2R)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}-3-[4-(4-methyl-1,3-thiazole-5-yl)phenyl]propanoic acid (2.8 g, 4.756 mmol, 1.0 equivalent) and lithium hydroxide monohydrate (0.409 g, 9.518 mmol, 2 equivalents) were dissolved in a mixture of tetrahydrofuran (2.8 ml, 1.7 M) and water (10.12 ml, 0.47 M) and stirred at room temperature for 2 hours. Subsequently, THF was removed under vacuum, the resulting aqueous residue was neutralized to pH 4 with KHSO4, and the formed solid was filtered to obtain the product (3S)-3-{[(2S,4R)-1-[(2S)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}-3-[4-(4-methyl-1,3-thiazole-5-yl)phenyl]propanoic acid (yield 86%), which was an off-white solid. Obtained by: LCMS (254nm): RT=2.787 min, 93.13% purity. ESI(+)[M+H] + = 575.24. 1 H NMR (300 MHz, DMSO-d6) δ 8.99 (s, 1H), 8.55 (d, J = 7.6 Hz, 1H), 7.47 - 7.37 (m, 4H), 7.28 (dd, J = 9.2, 3.0 Hz, 1H), 5.17-5.09 (m, 2H), 4.57 (d, J = 9.2 Hz, 1H), 4.43 (t, J = 8.3 Hz, 1H), 4.27 (s, 1H), 3.59 (dd, J = 12.0, 8.3 Hz, 2H), 2.86 - 2.62 (m, 2H), 2.46 (s, 3H), 2.04 (t, J = 10.6 Hz, 1H), 1.73 (ddd, J = 13.1, 8.9, 4.5 Hz, 1H), 1.36 (ddd, J = 18.2, 5.7, 3.1Hz, 2H), 1.25 - 1.16 (m, 2H), 0.96 (s, 9H). HVB15:2-[2-({[(2S,4R)-1-[(2S)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}methyl)-5-(4-methyl-1,3-thiazole-5-yl)phenoxy]acetic acid [ka]

[0303] The preparation was the same as for HVB4, except that tert-butyl bromoacetate was used instead of tert-butyl 6-bromohexanoate. LCMS:C 28 H 35 FN4O7S Theoretical value: 590.22, Measured value: m / z = 591.3 [M+H] + . HVB16:(S)-3-((2S,4R)-1-((R)-2-(1-fluorocyclopropane-1-carboxamide)-3,3-dimethylbutanoyl)-4-hydroxypyrrolidine-2-carboxamide)-3-phenylpropanoic acid [ka]

[0304] Step 1: Methyl(3S)-3-{[(2S,4R)-1-[(2R)-2-amino-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}-3-(4-bromophenyl)propanoate hydrochloride. Methyl(3S)-3-(4-bromophenyl)-3-{[(2S,4R)-1-[(2R)-2-{[(tert-butoxy)carbonyl]amino}-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}propanoate (1.5 g, 2.566 mmol, 1.0 equivalent) was slowly added to a solution of MeOH (21 mL, 0.3 M) with a methanol solution of 3 M HCl cooled to 0°C (160 mL, 0.15 M). The mixture was stirred at room temperature for 64 hours (overnight). Subsequently, MeOH was removed, and the resulting residue was powdered with Et2O to obtain the product methyl(3S)-3-{[(2S,4R)-1-[(2S)-2-amino-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}-3-(4-bromophenyl)propanoate hydrochloride (1.32 g, 2.534 mmol, 94%) as a pale yellow solid. ESI(-)[MH] - =482; 1 H NMR (300 MHz, DMSO-d6) 8.72 (d, J = 8.0 Hz, 1H), 8.10 (s, 3H), 7.53 - 7.48 (m, 2H), 7.30 - 7.23 (m, 2H), 5.11 (d, J = 7.7 Hz, 1H), 4.47 (t, J = 8.4 Hz, 1H), 4.28 (s, 1H), 3.88 (d, J = 5.1 Hz, 2H), 3.71 (d, J = 11.0 Hz, 1H), 3.48 (dd, J = 11.0, 3.8 Hz, 1H), 3.17 (s, 3H), 2.88 - 2.71 (m, 2H), 2.10 - 1.99 (m, 1H), 1.66 (s, 1H), 1.02 (s, 9H). Step 2: Methyl(3S)-3-(4-bromophenyl)-3-{[(2S,4R)-1-[(2R)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}propanoate

[0305] 1. To 1.5 mL of DMF (0.5 M), 1-fluorocyclopropanecarboxylic acid (0.274 g, 2.63 mmol, 1.04 equivalents) was added with DIPEA (0.663 mL, 1.5 equivalents) at 0°C. Next, HATU (1 g, 2.635 mmol, 1.04 equivalents) was dissolved in 5 mL of DMF and slowly added to the above mixture at 0°C. The reaction mixture was stirred at room temperature for 30 minutes.

[0306] 2. To a solution of methyl(3S)-3-{[(2S,4R)-1-[(2S)-2-amino-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}-3-(4-bromophenyl)propanoate hydrochloride (1.320 g, 2.534 mmol, 1.0 equivalent) in DMF (5 mL, 0.5 M), DIPEA (2.2 mL, 5 equivalents) was added at -40°C and the mixture was stirred at -40°C for 5 minutes.

[0307] 3. Reactant 1 was slowly added to reactant 2 at -40°C. The mixture was stirred at room temperature for 1 hour.

[0308] Subsequently, the resulting reaction mixture was diluted with water, then extracted with DCM, the organic layer was washed with brine, and dried under Na2SO4 to obtain the crude product. This crude product was purified by elution with DCM / MeOH-9 / 1 and FC to obtain the desired product, methyl(3S)-3-(4-bromophenyl)-3-{[(2S,4R)-1-[(2S)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}propanoate (1.117 g, 1.958 mmol, 73%). ESI(+)[M+H] + =570.2

[0316] 1 H NMR (300 MHz, DMSO-d6) δ 8.57 (d, J = 8.0 Hz, 1H), 7.55 - 7.46 (m, 2H), 7.31 - 7.23 (m, 3H), 5.17 - 5.09 (m, 2H), 4.57 (d, J = 9.1 Hz, 1H), 4.40 (t, J = 8.3 Hz, 1H), 4.25 (s, 1H), 3.56 (s, 4H), 3.20 - 3.03 (m, 1H), 2.84 - 2.70 (m, 2H), 2.05 - 1.92 (m, 1H), 1.69 (td, J = 8.6, 4.4 Hz, 1H), 1.36 (dd, J = 18.5, 3.5 Hz, 1H), 0.95 (d, J = 7.0 Hz, 9H).

[0309] Step 3: Methyl(3S)-3-{[(2S,4R)-1-[(2R)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}-3-phenylpropanoate. Methyl(3S)-3-{[(2S,4R)-1-[(2R)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}-3-phenylpropanoate) (0.834 g, 1.46 mmol, 1 equivalent) dissolved in i-PrOH (0.5 M) was degassed, Pd(OAc)2 (0.4 equivalents) was added, and the mixture was stirred overnight under H2 (1 atm, balloon). The conversion was monitored by LCMS, NMR and TLC. After complete consumption of the starting materials, the reaction mixture was filtered through a Celite pad and concentrated under reduced pressure to obtain the desired product, methyl(3S)-3-{[(2S,4R)-1-[(2R)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}-3-phenylpropanoate (0.6 g, 1.22 mmol, 87% yield). ESI(-)[MH] - =490.30;ESI(+)[M+H]+ =492.25; 1 H NMR (300 MHz, DMSO-d6) δ 8.53 (d, J = 8.2 Hz, 1H), 7.35 - 7.20 (m, 6H), 5.23 - 5.08 (m, 2H), 4.61 - 4.53 (d, J = 9.35 Hz, 1H), 4.42 (t, J = 8.2 Hz, 1H), 4.26 (s, 1H), 3.60 (m, 3.62-3.53 Hz, 5H), 2.88 - 2.70 (m, 2H), 2.05 - 1.96 (m, 1H), 1.74-1.65 (m, 1H), 1.42 - 1.31 (m, 2H), 1.25 - 1.17 (m, 3H), 0.96 (s, 9H).

[0310] Step 4: (S)-3-((2S,4R)-1-(R)-2-(1-fluorocyclopropane-1-carboxamide)-3,3-dimethylbutanoyl)-4-hydroxypyrrolidine-2-carboxamide)-3-phenylpropanoic acid. Methyl (3S)-3-{[(2S,4R)-1-[(2R)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}-3-phenylpropanoate (0.6 g, 1.22 mmol, 1 equivalent) was dissolved in a mixture of THF / H2O-5 / 1 (0.5 M), to which lithium hydroxide monohydrate (0.042 g, 2.44 mmol, 2 equivalents) was added, and the mixture was stirred at room temperature for 16 hours. The reaction product was monitored by TLC and LCMS. The THF was evaporated, the aqueous residue was neutralized with NaHSO4 (2 equivalents), and the mixture was concentrated. The resulting dried residue was powdered by DCM to obtain the desired product, methyl(3S)-3-{[(2S,4R)-1-[(2R)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxypyrrolidine-2-yl]formamide}-3-phenylpropanoate (0.350 g, 0.732 mmol, 81% yield):LCMS:254nm, RT=2.09 min, 87.76%, ESI(-)[MH] - =475.98; 1 H NMR (300 MHz, DMSO-d6) δ 12.24 (s, 1H), 8.46 (d, J = 8.1 Hz, 1H), 7.33 - 7.20 (m, 6H), 5.16 - 5.03 (m, 2H), 4.57 (d, J = 9.1 Hz, 1H), 4.42 (t, J = 8.3 Hz, 1H), 4.25 (s, 1H), 3.64 - 3.49 (m, 2H), 2.77 (dd, J = 15.7, 6.6 Hz, 1H), 2.64 (dd, J = 15.6, 8.2 Hz, 1H), 2.06 - 1.95 (m, 1H), 1.70 (ddd, J = 12.8, 8.7, 4.4 Hz, 1H), 1.42 - 1.31 (m, 2H), 1.24 - 1.17 (m, 3H), 0.96 (s, 9H). HVB17(2S,4S)-1-((S)-2-amino-3,3-dimethylbutanoyl)-4-hydroxy-N-(4-(4-methylthiazole-5-yl)benzyl)pyrrolidine-2-carboxamide [ka]

[0311] Step 1: Synthesis of N-[(4-bromophenyl)methyl]carbamate tert-butyl. To a solution of (4-bromophenyl)methaneamine (22.8 g, 122.55 mmol, 15.51 mL, 1 equivalent) and TEA (18.60 g, 183.82 mmol, 25.59 mL, 1.5 equivalents) in DCM (150 mL), tert-butoxycarbonyl tert-butyl carbonate (29.42 g, 134.80 mmol, 30.97 mL, 1.1 equivalents) was added. This mixture was stirred at 25°C for 2 hours. TLC (petroleum ether / ethyl acetate = 10:1) showed that the substance (4-bromophenyl)methaneamine had been consumed and a major novel spot was detected. The mixture was poured into water (150 mL) to separate the organic layer, which was washed with aqueous 1N HCl (150 mL) and brine (100 mL). The organic layer was then dehydrated with anhydrous Na2SO4 and filtered, and the filtrate was concentrated. The residue was purified by powdering in petroleum ether (120 mL), collected by filtration, and the filter cake was dried under vacuum to obtain N-[(4-bromophenyl)methyl]carbamate tert-butyl (26.3 g, 91.91 mmol, 75% yield) as a white solid.

[0312] Step 2: Synthesis of tert-butyl N-[[4-(4-methylthiazole-5-yl)phenyl]methyl]carbamate. To a stirred solution of tert-butyl N-[(4-bromophenyl)methyl]carbamate (26.3 g, 91.91 mmol, 1 equivalent) in DMA (150 mL), 4-methylthiazole (18.23 g, 183.81 mmol, 16.72 mL, 2 equivalents), KOAc (18.04 g, 183.81 mmol, 2 equivalents), and Pd(OAc)2 (1.03 g, 4.60 mmol, 0.05 equivalents) were added under an N2 atmosphere. The resulting mixture was stirred at 120 °C for 16 hours. LC-MS showed the detection of a major peak with the desired MS. TLC (petroleum ether / ethyl acetate = 5:1) showed that the substance N-[(4-bromophenyl)methyl]carbamate tert-butyl was consumed, and a major new spot was detected. This mixture was poured into water (200 mL) and the aqueous mixture was converted to ethyl acetate (150 mL). * The mixture was extracted using method 2), washed with brine (100 mL), dehydrated with anhydrous Na2SO4, and concentrated. The residue was powdered using petroleum ether:ethyl acetate = 10:1 (80 mL) to obtain tert-butyl N-[[4-(4-methylthiazole-5-yl)phenyl]methyl]carbamate (16.8 g, 55.19 mmol, 60.05% yield) as a yellow solid. MS[M+H] + =305.0. 1 1H NMR (400MHz, CDCl3) δ 8.70 (s, 1H), 7.46 - 7.41 (m, 2H), 7.40 - 7.34 (m, 2H), 4.98 - 4.87 (m, 1H), 4.39 (d, J = 5.9 Hz, 2H), 2.55 (s, 3H), 1.50 (s, 9H).

[0313] Step 3: Synthesis of [4-(4-methylthiazole-5-yl)phenyl]methaneamine. A mixture of tert-butyl N-[[4-(4-methylthiazole-5-yl)phenyl]methyl]carbamate (16.8 g, 55.19 mmol, 1 equivalent) in HCl / dioxane (4 M, 50 mL, 3.62 equivalents) was stirred at 25°C for 1 hour. LC-MS showed the detection of a major peak with the desired mass. Upon evaporation of the solvent, [4-(4-methylthiazole-5-yl)phenyl]methaneamine (13.3 g, crude, HCl) was obtained as a yellow solid, which was used directly in the next step without any purification. MS[M+H] + = 205.1.

[0314] Step 4: Synthesis of tert-butyl(2S,4S)-4-hydroxy-2-[[4-(4-methylthiazole-5-yl)phenyl]methylcarbamoyl]pyrrolidine-1-carboxylate. To a mixture of (2S,4S)-1-tert-butoxycarbonyl-4-hydroxypyrrolidine-2-carboxylic acid (12.77 g, 55.24 mmol, 1.0 equivalent) and DIPEA (14.28 g, 110.49 mmol, 19.24 mL, 2 equivalents) in DMF (120 mL), HATU (23.11 g, 60.77 mmol, 1.1 equivalents) was added. This mixture was stirred at 25°C for 30 minutes, then [4-(4-methylthiazole-5-yl)phenyl]methaneamine (13.3 g, 55.24 mmol, 1 equivalent, HCl) was added, and the resulting mixture was stirred at 25°C for 1.5 hours. LC-MS showed that the substance [4-(4-methylthiazole-5-yl)phenyl]methaneamine had been consumed and the desired mass was detected. This mixture was poured into water (100 mL), and the resulting aqueous solution was dissolved in ethyl acetate (100 mL). *2) The combined organic layer was extracted and concentrated by dehydration with anhydrous Na2SO4. Purification of the residue by chromatography (elution by silica gel, DCM:MeOH = 100:1, 50:1) yielded tert-butyl(2S,4S)-4-hydroxy-2-[[4-(4-methylthiazole-5-yl)phenyl]methylcarbamoyl]pyrrolidine-1-carboxylate (17 g, 38.19 mmol, 69.13% yield, 93.8% purity) as a pale yellow oil. MS[M+H] + = 418.3.

[0315] Step 5: Synthesis of (2S,4S)-4-hydroxy-N-[[4-(4-methylthiazole-5-yl)phenyl]methyl]pyrrolidine-2-carboxamide. A mixture of tert-butyl(2S,4S)-4-hydroxy-2-[[4-(4-methylthiazole-5-yl)phenyl]methylcarbamoyl]pyrrolidine-1-carboxylate (5 g, 11.23 mmol, 1 equivalent) in HCl / dioxane (4 M, 50 mL, 17.80 equivalents) was stirred at 25°C for 1 hour. LCMS showed that the substance tert-butyl(2S,4S)-4-hydroxy-2-[[4-(4-methylthiazole-5-yl)phenyl]methylcarbamoyl]pyrrolidine-1-carboxylate was consumed and a major peak with the desired mass was detected. Upon evaporation of the solvent, (2S,4S)-4-hydroxy-N-[[4-(4-methylthiazole-5-yl)phenyl]methyl]pyrrolidine-2-carboxamide (4g, crude, HCl) was obtained as a pale yellow solid, which was used directly in the next step without any purification. MS(M+H) + = 318.1.

[0316] Step 6: Synthesis of tert-butyl N-[(1S)-1-[(2S,4S)-4-hydroxy-2-[[4-(4-methylthiazole-5-yl)phenyl]methylcarbamoyl]pyrrolidine-1-carbonyl]-2,2-dimethylpropyl]carbamate. To a 30 mL solution of (2S,4S)-4-hydroxy-N-[[4-(4-methylthiazole-5-yl)phenyl]methyl]pyrrolidine-2-carboxamide (4 g, 11.30 mmol, 1 equivalent, HCl) and (2S)-2-(tert-butoxycarbonylamino)-3,3-dimethylbutanoic acid (2.61 g, 11.30 mmol, 1 equivalent) in DMF (30 mL), HATU (4.73 g, 12.43 mmol, 1.1 equivalents) and DIPEA (2.92 g, 22.61 mmol, 3.94 mL, 2 equivalents) were added at 0 °C, and the mixture was then stirred at 25 °C for 2 hours. LC-MS showed that the substance (2S,4S)-4-hydroxy-N-[[4-(4-methylthiazole-5-yl)phenyl]methyl]pyrrolidine-2-carboxamide was consumed, and a major peak with the desired mass was detected. This mixture was poured into water (100 mL), and the resulting water was dissolved in ethyl acetate (100 mL). * Extraction was performed according to (2). The combined organic layers were dehydrated with anhydrous Na2SO4 and concentrated. The residue was purified by reverse flash MPLC (FA), and tert -Butyl-N-[(1S)-1-[(2S,4S)-4-hydroxy-2-[[4-(4-methylthiazole-5-yl)phenyl]methylcarbamoyl]pyrrolidine-1-carbonyl]-2,2-dimethylpropyl]carbamate (1.65 g, 3.01 mmol, 26.60% yield, 96.7% purity) was obtained as a pale yellow gum-like substance. MS[M+H] + = 531.2.

[0317] Step 7: Synthesis of (2S,4S)-1-[(2S)-2-amino-3,3-dimethyl-butanoyl]-4-hydroxy-N-[[4-(4-methylthiazole-5-yl)phenyl]methyl]pyrrolidine-2-carboxamide. To a solution of tert-butyl N-[(1S)-1-[(2S,4S)-4-hydroxy-2-[[4-(4-methylthiazole-5-yl)phenyl]methylcarbamoyl]pyrrolidine-1-carbonyl]-2,2-dimethyl-propyl]carbamate (3.35 g, 6.31 mmol, 1 equivalent) in dioxane (10 mL), HCl / dioxane (4 M, 20 mL, 12.67 equivalents) was added. The mixture was stirred at 25 °C for 1 hour. LC-MS showed that a major peak with the desired mass was detected. The solvent was evaporated. The residue was powdered in petroleum ether / ethyl acetate (10:1, 80 mL) and collected by filtration, yielding (2S,4S)-1-[(2S)-2-amino-3,3-dimethyl-butanoyl]-4-hydroxy-N-[[4-(4-methylthiazole-5-yl)phenyl]methyl]pyrrolidine-2-carboxamide (2.77 g, 5.63 mmol, 89.20% yield, 94.8% purity, HCl) as a white powder. MS[M+H] + = 431.3. 1 H NMR (400MHz, CD3OD) δ 9.78 (s, 1H), 7.63 - 7.48 (m, 4H), 4.65 - 4.59 (m, 1H), 4.57 - 4.54 (m, 1H), 4.06 (s, 1H), 4.00 - 3.94 (m, 1H), 3.67 - 3.63 (m, 1H), 3.62 (s, 2H), 2.60 (s, 3H), 2.57 - 2.49 (m, 1H), 1.99 (d, J = 13.4 Hz, 1H), 1.15 (s, 9H). HVB18:(2S,4R)-1-[(2S)-2-amino-3,3-dimethylbutanoyl]-N-[(1S)-1-(4-bromophenyl)ethyl]-4-hydroxypyrrolidine-2-carboxamide [ka]

[0318] The same protocol as for the synthesis of (2S,4R)-1-[(2S)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxy-N-{[2-hydroxy-4-(4-methyl-1,3-thiazole-5-yl)phenyl]methyl}pyrrolidine-2-carboxamide (Scheme B4) was followed, except that (S)-1-(4-bromophenyl)ethane-1-amine was used instead of 2-(aminomethyl)-5-(4-methyl-1,3-thiazole-5-yl)phenol. LCMS:C 19 H 28 Theoretical value for BrN3O3: 425.13, Measured value: m / z = 426.67 [M+H] + . HVB19: (2S,4R)-1-[(2S)-2-amino-3,3-dimethylbutanoyl]-4-hydroxy-N-[(1S)-1-phenylethyl]pyrrolidine-2-carboxamide [ka]

[0319] The title compound was obtained by following the same protocol as in the synthesis of (2S,4R)-1-[(2S)-2-[(1-fluorocyclopropyl)formamide]-3,3-dimethylbutanoyl]-4-hydroxy-N-{[2-hydroxy-4-(4-methyl-1,3-thiazole-5-yl)phenyl]methyl}pyrrolidine-2-carboxamide (Scheme B4), except that (S)-1-phenylethane-1-amine was used instead of 2-(aminomethyl)-5-(4-methyl-1,3-thiazole-5-yl)phenol. LCMS:C 19 H 29 Theoretical value of N3O3: 347.22, Measured value: m / z = 348.13 [M + H] + . HVB20: (3R)-3-((2R,4S)-4-hydroxy-1-(3-methyl-2-(3-methylisoxazole-5-yl)butanoyl)pyrrolidine-2-carboxamide)-3-(4-(4-methylthiazole-5-yl)phenyl)propanoic acid [ka]

[0320] Step 1: Methyl (3S)-3-amino-3-(4-bromophenyl)propanoate. To a solution of (3S)-3-(4-bromophenyl)-3-{[(tert-butoxy)carbonyl]amino}propanoic acid (8.0 g, 0.023 mmol, 1.0 equivalent) in methanol (100 ml, 0.01 M), a chilled solution of HCl (3 M in MeOH, 160 mL, 0.01 M) was slowly added at 0°C. The mixture was stirred at room temperature for 16 hours. The crude reaction product was concentrated under vacuum at 30°C, then 40 ml of a 3 M solution of HCl in Et2O was added, and the mixture was concentrated under vacuum to obtain methyl (3S)-3-amino-3-(4-bromophenyl)propanoate as a foamy white solid. The product was isolated as an HCl salt and used in the next step without further purification (5.71 g, 95% yield). ESI(+)[M+H] + =258.00; 1 H NMR (300 MHz, Methanol-d4), δ: 7.63 (d, J = 8.2 Hz, 2H), 7.40 (d, J = 8.2 Hz, 2H), 4.73 (t, J = 7.1 Hz, 1H), 3.70 (s, 3H), 3.19 - 2.97 (m, 2H).

[0321] Step 2: tert-butyl(2S,4R)-2-{[(1S)-1-(4-bromophenyl)-3-methoxy-3-oxopropyl]carbamo-yl}-4-hydroxypyrrolidine-1-carboxylate. (2S,4R)-1-[(tert-butoxy)carbonyl]-4-hydroxypyrrolidine-2-carboxylic acid (5.71 g, 24.7 mmol, 1.15 equivalents) was dissolved in DMF (45 mL, 0.5 M) and DIPEA (6 ml, 1.5 equivalents) was added at 0°C. Next, HATU (8.53 g, 22.5 mmol, 1.04 equivalents) was slowly added to the previous solution in DMF (45 mL, 0.5 M) at 0°C. The reaction mixture was stirred at room temperature for 0.5 hours, and then slowly added at -30°C to a chilled DMF (35 ml, 0.6 M) solution of methyl (3S)-3-amino-3-(4-bromophenyl)propanoate (6.7 g, 21.5 mmol, 1 equivalent) pretreated with DIPEA (20 mL, 5 equivalents). The mixture was stirred at -30°C and slowly warmed to room temperature over 2 hours (controlled by TLC, ULC, and NMR). The crude reaction mixture was then poured onto crushed ice and extracted by DCM (6 × 500 mL). The organic layer was dehydrated with Na₂SO₄, concentrated under vacuum, and purified by flash column chromatography (eluent DCM / MeOH₇:1) to obtain the desired compound as a foamy white solid (10.54 g, quantitative yield). ESI(+)[M+H] + =471.10; 1 ¹H NMR (300 MHz, DMSO-d6) Methanol-d4) δ 7.47 (t, J = 7.9 Hz, 2H), 7.36 - 7.20 (m, 2H), 5.32 (t, J = 7.4 Hz, 1H), 4.29 (dd, J = 15.4, 6.9 Hz, 2H), 3.62 (s, 3H), 3.60 - 3.40 (m, 1H), 3.03 - 2.73 (m, 2H), 2.34 - 2.08 (m, 1H), 2.03 - 1.76 (m, 1H), 1.47 (s, 3H), 1.40 - 1.29 (s, 6H). No Boc protons were found.

[0322] Step 3: Methyl-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1,3-thiazole. A mixture of 5-bromo-4-methyl-1,3-thiazole (7.5 g, 42.1 mmol, 1 equivalent), KOAc (12.4 g, 126.4 mmol, 3.4 equivalents), and 4,4,5,5-tetramethyl-2-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1,3,2-dioxaborolan (21.4 g, 1.85 mmol, 2 equivalents) and Pd(PPh3)4 (10 g, 20 mol%) was dissolved in dioxane (375 ml, 0.1 M), purged with argon for 10 minutes, and stirred at 95°C for 16 hours. Next, the mixture was cooled to room temperature, filtered through a Celite pad, concentrated under vacuum, and purified by short manual column chromatography (eluent hexane / siRNA 1:1) to obtain the title product as an off-white solid (10.25 g, 52% yield, contaminated with 50 wt% pinacol derivative). 1 H NMR (300 MHz, Chloroform-d), δ: 8.92 (s, 1H), 2.70 (s, 3H), 1.34 (s, 12H).

[0323] Step 4: (3S)-3-{[(2S,4R)-1-[(tert-butoxy)carbonyl]-4-hydroxypyrrolidine-2-yl]formami-do}-3-[4-(4-methyl-1,3-thiazole-5-yl)phenyl]propanoate methyl ester. The mixture of tert-butyl(2S,4R)-2-{[(1S)-1-(4-bromophenyl)-3-methoxy-3-oxopropyl]carbamoyl}-4-hydroxypyrrolidine-1-carboxylate (9g, 19.09 mmol, 1 equivalent), 4-methyl-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1,3-thiazole (9.91g, 21 mmol, 1.2 equivalents), K2CO3 (13.2g, 95.5 mmol, 5 equivalents), and Pd(dppf)Cl2.DCM (1.6g, 10 mol%) in dioxane / H2O (5:1, 380 mL, 0.05 M) was purged with argon for 20 minutes and stirred at 110°C for 2 hours (completion of Suzuki coupling). Next, the mixture was cooled to room temperature and filtered through a Celite pad. The filtrate was concentrated under vacuum and purified by flash column chromatography (eluent DCM / MeOH / AcOH 8:2:0.2% to 6:4:0.2%). The desired product was concentrated under vacuum, dissolved in DCM / MeOH / AcOH 9:1:0.1%, filtered, and the final silica gel was removed. The filtrate was concentrated under vacuum and then precipitated in diethyl ether to obtain the desired product as a gray solid (6.6 g, 76% yield). ESI(+)[M+H] + =476.07; 1 H NMR (300 MHz, Methanol-d4), δ: 8.88 (s, 1H), 7.47 (m, 4H), 5.54 - 5.28 (m, 1H), 4.33 (d, J = 9.9 Hz, 2H), 3.68 - 3.40 (m, 2H), 3.60 (s, 3H), 2.88 (m, 2H), 2.48 (s, 3H), 2.31 - 2.14 (m, 1H), 1.99 (s, 1H), 1.48 (s, 3H), 1.33 (s, 6H).

[0324] Step 5: Methyl (3S)-3-{[(2S,4R)-4-hydroxypyrrolidine-2-yl]formamide}-3-[4-(4-methyl-1,3-thiazole-5-yl)phenyl]propanoate. A mixture consisting of (3S)-3-{[(2S,4R)-1-[(tert-butoxy)carbonyl]-4-hydroxypyrrolidine-2-yl]formamide (forma-mido)}-3-[4-(4-methyl-1,3-thiazole-5-yl)phenyl]methyl propanoate (0.3 g, 0.61 mmol, 1 equivalent) and 2N HCl (10 equivalents) in methanol was stirred under ambient conditions for 2 hours (reaction controlled by ULC and NMR). The solvent was removed under vacuum, and the resulting solid was powdered with dry diethyl ether to obtain the desired salt product, which was obtained as a highly viscous brown oily substance (0.22 g, 83% yield). ESI(+)[M+H] + =390.45; 1 H NMR (300 MHz, DMSO-d6) δ: 9.89 (s, 1H), 9.32 (d, J = 7.9 Hz, 1H), 9.03 (s, 1H), 8.65 (s, 1H), 7.51 - 7.39 (m, 4H), 4.33 (s, 2H), 3.61 (s, 3H), 3.51 (s, 1H), 3.41 (s, 2H), 3.07 (d, J = 4.7 Hz, 1H), 2.88 (d, J = 7.5 Hz, 2H), 2.33 (s, 1H), 1.78 (m, 1H).

[0325] Step 6: Methyl 2-(3-methyl-1,2-oxazol-5-yl)acetate. To a solution of 3-methyl-5-isoxazoleacetic acid (0.8 g, 5.67 mmol, 1 equivalent) in MeOH (10 ml, 0.55 M), thionyl chloride (1.5 equivalents) was added dropwise at 0°C, and the resulting mixture was stirred at 50°C for 4 hours. UPLC monitoring was used. Next, this reaction mixture was poured into saturated ammonium chloride and converted to ethyl acetate. Further extraction, washing with saturated NaHCO3, drying, and concentration under vacuum yielded the desired product as a brown oily substance (0.78 g, 89% yield). 1 H NMR (300 MHz, Chloroform-d), δ: 6.11 (s, 1H), 3.80 (s, 2H), 2.76 (s, 3H), 2.30 (s, 3H).

[0326] Step 7: Methyl 3-methyl-2-(3-methyl-1,2-oxazole-5-yl)butanoate. A mixture of methyl 2-(3-methyl-1,2-oxazole-5-yl)acetate (0.14 g, 0.9 mmol, 1 equivalent), cesium carbonate (0.32 g, 0.99 mmol, 1.1 equivalents), and 2-iodopropane (0.16 g, 0.94 mmol, 1.05 equivalents) in DMSO (2.3 ml, 0.4 M) was stirred at 65-70°C for 5-8 hours (LCMS control was applied). After the reaction was complete, the reaction mixture was poured into a dilute aqueous HCl solution, extracted twice with ethyl acetate, dried, and the solvent evaporated under vacuum. The crude product was purified by flash chromatography using ELSD (the product is not UV-reactive) to obtain the desired product (0.12 g, 64% yield). 1 H NMR (300 MHz, DMSO-d6), δ: 6.30 (s, 1H), 3.76 (d, J = 8.6 Hz, 1H), 3.66 (s, 3H), 2.35 - 2.26 (m, 1H), 2.21 (s, 3H), 0.93 (d, J = 6.7 Hz, 3H), 0.83 (d, J = 6.7 Hz, 3H).

[0327] Step 8: 3-Methyl-2-(3-methyl-1,2-oxazol-5-yl)butanoic acid. To the initial solution of methyl 3-methyl-2-(3-methyl-1,2-oxazol-5-yl)butanoate (0.59 g, 2.99 mmol, 1 equivalent) in THF-water (3:1; 0.14 M), sodium hydroxide (0.18 g, 4.5 mmol, 1.5 equivalents) was added, and the resulting mixture was stirred at room temperature until the reaction was complete (controlled by TLC). Next, the THF was evaporated under low pressure, and the aqueous residue was acidified to pH = 4-3 with 1N aqueous HCl. The resulting solution was extracted twice with ELISA and dried, and after all volatiles had evaporated, the desired compound was obtained as a white solid (0.5 g, 90% yield). 1 H NMR (300 MHz, DMSO-d6), δ: 12.84 (s, 1H), 6.27 (s, 1H), 3.58 (d, J = 8.7 Hz, 1H), 2.35 - 2.23 (m, 1H), 2.21 (s, 3H), 0.96 (d, J = 6.7 Hz, 3H), 0.82 (d, J = 6.7 Hz, 3H).

[0328] Step 9: (3S)-3-{[(2S,4R)-4-hydroxy-1-[3-methyl-2-(3-methyl-1,2-oxazol-5-yl)butanoyl]pyrrolidine-2-yl]formamide}-3-[4-(4-methyl-1,3-thiazole-5-yl)phenyl]propanoic acid. To a solution of methyl(3S)-3-{[(2S,4R)-4-hydroxypyrrolidine-2-yl]formamide}-3-[4-(4-methyl-1,3-thiazole-5-yl)phenyl]propanoate (0.23 g, 0.56 mmol, 1 equivalent) and 3-methyl-2-(3-methyl-1,2-oxazole-5-yl)butanoic acid (0.11 g, 0.62 mmol, 1.1 equivalents) in DCM (6 ml, 0.1 M), DIPEA (0.22 ml, 1.7 mmol, 3.00 equivalents) and HATU (0.32 g, 0.84 mmol, 1.5 equivalents) were added. This mixture was stirred overnight at 25°C. UPLC control was used. The reaction mixture was quenched with water and extracted with ethyl acetate. The combined organic layers were washed with acidic water and brine, dehydrated with Na2SO4, and concentrated to obtain the crude product, which was purified by flash chromatography as a viscous oily substance (0.28 g, 78% yield). ESI(+)[M+H] + =556.04; 1 H NMR (300 MHz, DMSO-d6), δ: 8.72 (s, 1H), 7.47-7.35 (br m, 4H), 6.10 (m, 1H), 5.35 (m, 1H), 4.60 (m, 2H), 3.72 (m, 4H), 3.60 (s, 3H), 2.77 (m, 2H), 2.50 (m, 4H), 2.25 (m, 4H), 2.06 (m, 1H), 1.07 (m, 3H), 0.89 (m, 3H).

[0329] Step 10: (3S)-3-{[(2S,4R)-4-hydroxy-1-[3-methyl-2-(3-methyl-1,2-oxazol-5-yl)butanoyl]pyrrolidine-2-yl]formamide}-3-[4-(4-methyl-1,3-thiazole-5-yl)phenyl]propanoic acid. To a methanol-water (3:1; 0.14M) solution of the starting compound (3S)-3-{[(2S,4R)-4-hydroxy-1-[3-methyl-2-(3-methyl-1,2-oxazole-5-yl)butanoyl]pyrrolidine-2-yl]formamide}-3-[4-(4-methyl-1,3-thiazole-5-yl)phenyl]propanoic acid (0.28 g, 0.54 mmol, 1 equivalent) in methanol-water (3:1; 0.14 M), sodium hydroxide (0.03 g, 0.75 mmol, 1.5 equivalents) was added, and the resulting mixture was stirred under ambient conditions until complete (controlled by ULC). Next, the organic solvent was evaporated under low pressure, and the water residue was acidified to pH=4-3 with 1N HCl. The resulting solution was purified by reverse-phase flash chromatography (5-29% acetonitrile in water). After solvent evaporation, the title compound was obtained as a white solid (0.1 g, 37% yield). LCMS: 254nm, RT=2.443min, 98.32% purity, ESI(+)[M+H] + =542.66; 1 H NMR (300 MHz, DMSO-d6), δ: 8.90 (s, 1H), 7.47 (m, 4H), 6.25 (d, J = 5.9 Hz, 1H), 5.38 (m, 1H), 4.62 - 4.37 (m, 2H), 3.94 - 3.41 (m, 4H), 3.10 - 2.78 (m, 2H), 2.50 (m, 4H), 2.25 (m, 4H), 1.98 (m, 1H), 1.07 (d, J = 7.6 Hz, 3H), 0.94 - 0.82 (m, 3H). C. General scheme for coupling IRAK4 joints with LHM building blocks

[0330] The L portion typically has up to five linker segments (-L1-L2-L3-L4-L5-), one of which is formed by coupling the building blocks and LHM blocks of IRAK4 described herein by bond formation (e.g., amide). The following general methods A to D illustrate bond formations in which building blocks can be linked to yield a compound of formula (I). General method A (amide coupling): [ka] General method B (reduced amination): [ka] General method C (replacement): [ka] General method D (amide coupling, in-situ BOC deprotection)

[0331] General method D is similar to general method A, except that the terminal portion of the amine (e.g., the building block of IRAK4) can be initially protected with BOC. Amide coupling can be carried out by in-situ BOC deprotection to form an amide bond having the terminal portion of the carboxylic acid (e.g., the building block of LHM). See, for example, the synthesis of Example 50. definition

[0332] The following descriptions illustrate exemplary methods, parameters, etc. However, it should be recognized that such descriptions are not intended to limit the scope of this disclosure, but rather are presented as descriptions of exemplary embodiments.

[0333] A dash ("-") that does not exist between two letters or symbols is used to indicate bonding points for substituents. For example, -C(O)NH2 is bonded via a carbon atom. Dashes at the beginning or end of a chemical group are for convenience only, and chemical groups may be illustrated with or without one or more dashes without losing their original meaning. A wavy line illustrated across a line in a structure indicates a bonding point of a group. Unless chemically or structurally necessary, direction is not indicated, nor is it implied by the order in which chemical groups are listed or named.

[0334] Prefix “C” u~v " indicates that the following group has u to v carbon atoms. For example, "C 1~6 The term "alkyl" indicates that this alkyl group has 1 to 6 carbon atoms.

[0335] References to values ​​or parameters "about" in this specification include (and are described) embodiments that apply to the value or parameter itself. In certain embodiments, the term "about" includes ±10% of the stated amount. In other embodiments, the term "about" includes ±5% of the stated amount. In certain other embodiments, the term "about" includes ±1% of the stated amount. Similarly, the term "with respect to X" includes the description of "X". Furthermore, the singular forms "a" and "the" include plural references unless the context specifically indicates otherwise. Thus, for example, "compound" includes multiple such compounds, and "assay" includes one or more assays and their equivalents known to those skilled in the art as references.

[0336] "Alkyl" refers to a branched saturated hydrocarbon chain that does not contain unsaturated carbon atoms. When used herein, alkyl refers to a chain with 1 to 20 carbon atoms (i.e., C 1~20 Alkyl), 1 to 12 carbon atoms (i.e., C 1~12 Alkyl), 1 to 8 carbon atoms (i.e., C 1~8 Alkyl), 1 to 6 carbon atoms (i.e., C 1~6 Alkyl) or 1 to 4 carbon atoms (i.e., C1~4 Alkyl compounds include methyl, ethyl, propyl, isopropyl, n-butyl, sec-butyl, isobutyl, tert-butyl, pentyl, 2-pentyl, isopentyl, neopentyl, hexyl, 2-hexyl, 3-hexyl, and 3-methylpentyl. When an alkyl residue having a specific number of carbon atoms is named by its chemical name or identified by its molecular formula, all positional isomers having that number of carbon atoms can be considered; for example, "butyl" includes n-butyl (i.e., -(CH2)3CH3), sec-butyl (i.e., -CH(CH3)CH2CH3), isobutyl (i.e., -CH2CH(CH3)2), and tert-butyl (i.e., -C(CH3)3); and "propyl" includes n-propyl (i.e., -(CH2)2CH3) and isopropyl (i.e., -CH(CH3)2).

[0337] "Alkylene" or "alkylene chain" refers to a divalent, unbranched or branched hydrocarbon chain that is unsaturated and has 1 to 20 carbon atoms, or more typically 1 to 12 carbon atoms, or 1 to 8 carbon atoms, linking the rest of the molecule to a radical group, such as methylene, ethylene, propylene, n-butylene, etc. The alkylene chain can be bonded to the rest of the molecule and the radical group via one carbon in the chain or via any two carbons in the chain.

[0338] "Alkenyl" is defined as a molecule containing at least one carbon-carbon double bond and 2 to 20 carbon atoms (i.e., C 2~20 Alkenyls), or more typically, 2 to 12 carbon atoms (i.e., C 2~12 Alkenyl), 2 to 8 carbon atoms (i.e., C 2~8 Alkenyl), 2 to 6 carbon atoms (i.e., C 2~6 Alkenyl) or 2-4 carbon atoms (i.e., C 2~4This refers to an alkyl group having an alkenyl group. Examples of alkenyl groups include ethenyl, propenyl, and butadienyl (including 1,2-butadienyl and 1,3-butadienyl).

[0339] "Alkenylene" and "alkenylene chain" refer to divalent unbranched or branched hydrocarbon chains that contain at least one double bond and have 2 to 20 carbon atoms, or more typically 2 to 12 carbon atoms, or 2 to 8 carbon atoms, linking the rest of the molecule to a radical group, such as etenylene, propenylene, n-butenylene, etc. Alkenylene chains are bonded to the rest of the molecule via single bonds and to the radical group via double or single bonds. The bonding points of an alkenylene chain to the rest of the molecule and to the radical group can be via one carbon or any two carbons within the chain.

[0340] "Alkynyl" refers to a compound that contains at least one carbon-carbon triple bond and 2 to 20 carbon atoms (i.e., C 2~20 Alkynnyl) or more typically, 2 to 12 carbon atoms (i.e., C 2~12 Alkynnyl) or more typically, 2 to 8 carbon atoms (i.e., C 2~8 Alkynyl), 2-6 carbon atoms (i.e., C 2~6 Alkynyl) or 2-4 carbon atoms (i.e., C 2~4 This refers to an alkyl group having an alkynyl bond. The term "alkynyl" also includes such groups having one triple bond and one double bond.

[0341] "Alkynylene" and "alkynylene chain" refer to divalent, unbranched or branched hydrocarbon chains that contain at least one triple bond and have 2 to 20 carbon atoms, or more typically 2 to 12 carbon atoms, or 2 to 8 carbon atoms, linking the rest of the molecule to a radical group. Alkynylene chains are bonded to the rest of the molecule via single bonds and to the radical group via double or single bonds. The bonding points of an alkynylene chain to the rest of the molecule and to the radical group can be via one carbon or any two carbons within the chain.

[0342] "Alkoxy" refers to the group "alkyl-O-". Examples of alkoxy groups include methoxy, ethoxy, n-propoxy, iso-propoxy, n-butoxy, tert-butoxy, sec-butoxy, n-pentoxy, n-hexoxy, and 1,2-dimethylbutoxy.

[0343] A "haloalkoxy" refers to an alkoxy group as defined above, in which one or more hydrogen atoms are replaced by halogens.

[0344] "Alkylthio" refers to the group "alkyl-S-".

[0345] "Amino" means -NR y R y It refers to the base, R y Each of these is independently selected from the group consisting of hydrogen, alkyl, alkenyl, alkynyl, aryl, heterocyclyl, cycloalkyl, or heteroaryl, and each of these is substituted as necessary as defined herein.

[0346] "Aryl" refers to an aromatic carbocyclic group having a monocyclic (e.g., monocyclic) or polycyclic (e.g., bicyclic or tricyclic) structure, including condensed systems. As used herein, aryl has 6 to 20 ring carbon atoms (i.e., C 6~20 aryl), 6 to 15 carbocyclic atoms (i.e., C 6~15aryl) or 6-10 carbon ring atoms (i.e., C 6~10 It contains an aryl group. Examples of aryl groups include phenyl, naphthyl, fluorenyl, and anthryl. However, aryl does not include, nor overlap with, heteroaryls as defined below. When one or more aryl groups are fused with a heteroaryl, the resulting ring system is a heteroaryl. When one or more aryl groups are fused with a heterocyclyl, the resulting ring system is a heterocyclyl.

[0347] "Cyano" refers to the -CN group.

[0348] "Keto" or "oxo" refers to the oxygen group (=O).

[0349] "Carbamoyl" is -OC(O)NR y R z The group referred to is the "O-carbamoyl" group, and -NR y C(O)OR z The term "N-carbamoyl" refers to both the group and the R group. y and R z These are independently selected from the group consisting of hydrogen, alkyl, aryl, haloalkyl, or heteroaryl, and each may be substituted as needed.

[0350] "Carboxyl" or "carboxylic acid" refers to -C(O)OH.

[0351] "Ester" refers to both -OC(O)R and -C(O)OR, where R is a substituent, which may be substituted as necessary, as defined herein.

[0352] "Cycloalkyl" refers to saturated or partially unsaturated cyclic alkyl groups having monocyclic or polycyclic structures, including fused ring systems, bridging ring systems, and spiro-ring systems. The term "cycloalkyl" includes a cycloalkenyl group (i.e., a cyclic group having at least one double bond). As used herein, cycloalkyl refers to a ring of 3 to 15 carbon atoms (i.e., C 3~20 Cycloalkyl), 3 to 12 ring carbon atoms (i.e., C 3~12 Cycloalkyl), 3 to 10 ring carbon atoms (i.e., C 3~10 Cycloalkyl), 3 to 8 ring carbon atoms (i.e., C 3~8 Cycloalkyl) or 3-6 ring carbon atoms (i.e., C 3~6 It has a cycloalkyl group. Examples of cycloalkyl groups include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, and bicyclo[2.2.2]octan-1-yl. The cycloalkyl group may be bonded to the remainder of the molecule by a monocyclic atom (e.g., as a substituent) or by two ring atoms (e.g., as a linker).

[0353] An "ethylene glycol unit" refers to a divalent monomer having the structure -CH2CH2O-, which may be repeated or extended into a longer chain. A linker segment may have up to 12 ethylene glycol units, or more typically, up to 6 ethylene glycol units.

[0354] A "propylene glycol unit" refers to a divalent monomer having the structure -CH(CH3)-CH2O-, which may be repeated or extended into a longer chain. The linker segment may have up to 12 propylene glycol units, or more typically, up to 6 propylene glycol units.

[0355] "Halogen" or "halo" includes fluoro, chloro, bromo, and iodine.

[0356] A "haloalkyl" refers to an alkyl group that is unbranched or branched as defined above, in which one or more hydrogen atoms are replaced by halogens. For example, if a residue is substituted with more than one halogen, it may be referred to using a prefix corresponding to the number of halogenated groups. Dihaloalkyls and trihaloalkyls refer to alkyl groups substituted with two ("di") or three ("tri") halo groups, where the halo groups may be the same halogen, but are not necessarily the same. Examples of haloalkyls include difluoromethyl (-CHF2) and trifluoromethyl (-CF3).

[0357] A "heteroalkyl" refers to an alkyl group in which one or more carbon atoms (and any bonded hydrogen atoms) are independently replaced by the same or different heteroatoms (such as N, O, or S). The term "heteroalkyl" includes unbranched or branched saturated chains having carbon and heteroatoms. For example, one, two, or three carbon atoms may be independently replaced by the same or different heteroatoms. Heteroatomic groups include, but are not limited to, -N(R)-, -O-, -S-, -S(O)-, -S(O)2-, etc., where R is H, alkyl, aryl, cycloalkyl, heteroalkyl, heteroaryl, or heterocyclyl, each of which may be substituted as needed. Examples of heteroalkyl groups include -OCH3, -CH2OCH3, -SCH3, -CH2SCH3, -NRCH3, and -CH2NRCH3, where R is hydrogen, alkyl, aryl, arylalkyl, heteroalkyl, or heteroaryl, each of which may be substituted as needed. As used herein, a heteroalkyl group comprises 1 to 10 carbon atoms, 1 to 8 carbon atoms, or 1 to 4 carbon atoms; and 1 to 3 heteroatoms, 1 to 2 heteroatoms, or 1 heteroatom.

[0358] "Heteroaryl" refers to an aromatic group having a monocyclic, polycyclic, or fused polycyclic structure with 5 to 15 members, or more typically 5 to 12 members, and having 1 to 3 ring heteroatoms independently selected from nitrogen, oxygen, and sulfur. As used herein, heteroaryl refers to an aromatic group having 3 to 12 ring carbon atoms (i.e., C 3~12 Heteroaryl) or 3 to 8 carbon ring atoms (i.e., C 3~8 Heteroaryl groups include, but are not limited to, benzo[d]thiazolyl, quinolinyl, isoquinolinyl, benzo[b]thiophenyl, indazolyl, benzo[d]imidazolyl, pyrazolo[1,5-a]pyridinyl, pyrazolo[1,5-a]pyridinyl, pyrazolo[1,5-a]pyridinyl, and pyrazolo[1,5-a]pyridinyl, and heteroaryl groups may be bonded via any of the rings in the condensation system. Any aromatic ring having a monocyclic or polycyclic group containing at least one heteroatom is considered a heteroaryl group regardless of its bond to the rest of the molecule (i.e., via any one of the condensation rings). A heteroaryl does not encompass, nor overlap with, the aryls defined above. A heteroaryl may be bonded to the remainder of the molecule by a monocyclic atom (e.g., as a substituent) or by two ring atoms (e.g., as a linker).

[0359] A "heterocyclyl" refers to a 3- to 15-membered, or more typically 5- to 12-membered saturated or unsaturated cyclic alkyl group having 1 to 3 ring heteroatoms independently selected from nitrogen, oxygen, and sulfur. The term "heterocyclyl" includes heterocycloalkenyl groups (i.e., heterocyclyl groups having at least one double bond), bicyclic heterocyclyl groups, bridging heterocyclyl groups, condensed heterocyclyl groups, and spiroheterocyclyl groups. Heterocyclyls may be monocyclic or polycyclic, and polycyclics may be condensed, bridging, or spiro. Any non-aromatic ring containing at least one heteroatom is considered a heterocyclyl, regardless of bonding (i.e., it can be bonded via carbon atoms or heteroatoms). Furthermore, the term heterocyclyl is intended to encompass any non-aromatic ring containing at least one heteroatom, and the above rings may be condensed to an aryl ring or heteroaryl ring, regardless of bonding to the rest of the molecule. As used herein, a heterocyclyl has 3 to 15 ring atoms (e.g., a 3 to 15-membered heterocyclyl, a 3 to 12-membered heterocyclyl, a 4 to 10-membered heterocyclyl, a 4 to 8-membered heterocyclyl, or a 4 to 6-membered heterocyclyl) having 1 to 5 ring heteroatoms, 1 to 4 ring heteroatoms, 1 to 3 ring heteroatoms, 1 to 2 ring heteroatoms, or 1 ring heteroatom, which are independently selected from nitrogen, sulfur, or oxygen. The heterocyclyl may also contain one or more oxo groups and / or thioxo groups. Examples of heterocyclyl groups include pyrrolidinyl, piperidinyl, piperazinyl, oxetanyl, dioxolanil, azetidinyl, azetidinyl, morpholinyl, thiomorpholinyl, 4-7 membered sultams, 4-7 membered cyclic carbamates, 4-7 membered cyclic carbonates, 4-7 membered cyclic sulfides, and morpholinyl.As used herein, heterocyclyl can include a crosslinked structure (i.e., “crosslinked heterocyclyl”), in which case a heterocyclyl having one or more (e.g., one or two) 4- to 10-membered cyclic moieties linked at two non-adjacent atoms of a heterocyclyl, each heteroatom independently selected from nitrogen, oxygen, and sulfur. As used herein, crosslinked heterocyclyls include bicyclic and tricyclic systems. Similarly, as used herein, the term “spiro-heterocyclyl” means a cyclic system in which a 3- to 10-membered heterocyclyl has one or more additional rings, in which case the one or more additional rings are a 3- to 10-membered cycloalkyl or a 3- to 10-membered heterocyclyl, and a single atom of the one or more additional rings is also an atom of the 3- to 10-membered heterocyclyl. Examples of spiro-heterocyclyl rings include bicyclic and tricyclic ring systems such as 2-oxa-7-azaspiro[3.5]nonanyl, 2-oxa-6-azaspiro[3.4]octanyl, and 6-oxa-1-azaspiro[3.3]heptanyl. Examples of condensed heterocyclyl rings include, but are not limited to, 1,2,3,4-tetrahydroisoquinolinyl, 1-oxo-1,2,3,4-tetrahydroisoquinolinyl, 1-oxo-1,2-dihydroisoquinolinyl, 4,5,6,7-tetrahydrothieno[2,3-c]pyridinyl, indolinyl, 2,3-dihydro-1H-isoindolyl, and isoindolinyl, and heterocyclyls can be linked via any of the rings in the condensed system. As used herein, a bicyclic heterocyclyl group is a heterocyclyl group bonded at two points to another cyclic group, the other cyclic group may itself be a heterocyclic or carbocyclic group. The heteroaryl may be bonded to the remainder of the molecule by a monocyclic atom (e.g., as a substituent) or by two cyclic atoms (e.g., as a linker).

[0360] "Condensed" refers to a ring that is connected to an adjacent ring and shares two adjacent ring atoms that form a covalent bond.

[0361] "Bridged" refers to a ring condensation in which non-adjacent atoms on a ring are bonded by divalent substituents such as alkenyl groups, one or two heteroatoms, or alkenyl groups containing a single heteroatom. Quinuclidinyl and adamantanyl are examples of bridged ring systems.

[0362] "Spiro" refers to a ring substituent linked by two bonds at the same carbon atom. Examples of spiro groups include 1,1-diethylcyclopentane, dimethyl-dioxolane, and 4-benzyl-4-methylpiperidine, with cyclopentane and piperidine being spiro substituents, respectively.

[0363] "Hydroxyl" or "hydroxy" refers to the -OH group. "Hydroxyalkyl" refers to an alkyl group that is unbranched or branched as defined above, in which one or more hydrogen atoms are replaced by hydroxyl groups.

[0364] "Nitro" refers to the -NO2 group.

[0365] "Imino" means C=NR y Or =NC(O)R y This refers to groups containing a C=N double bond, such as R. y The imino is selected from the group consisting of hydrogen, alkyl, aryl, cyano, haloalkyl, or heteroaryl, and each of these may be substituted as needed. The imino may be a linker segment, bonded to the remainder at carbon and nitrogen, respectively. "Sulfoximin" or "sulfoximino" is defined by the following general formula: [ka]

[0366] (In the formula, R yis selected from the group consisting of hydrogen, alkyl, amino, aryl, cyano, haloalkyl, heterocyclyl, or heteroaryl; V and W are independently selected from bond, alkyl, amino, aryl, haloalkyl, heterocyclyl, or heteroaryl, and these may each be substituted as needed, R y and V, R y The substituted or unsubstituted portions refer to the (and W and V and W may be linked together with the atoms to which they are bonded to form a ring). The sulfoximine may also be a linker segment, bonded to the remaining portion at sulfur and nitrogen, respectively.

[0367] "Sulfonyl" refers to the -S(O)2R group, where R is a substituent or a defined group.

[0368] "Alkylsulfonyl" refers to the -S(O)2R group, where R is a substituent or a defined group.

[0369] "Alkyl sulfinyl" refers to the -S(O)R group, where R is a substituent or a defined group.

[0370] "Thiocyanate" - SCN.

[0371] "Thiol" refers to the -SR group, where R is a substituent or a defined group.

[0372] "Thioxo" or "thione" refers to a (=S) or (S) group.

[0373] Certain commonly used alternative chemical names may be used. For example, divalent groups such as divalent "alkyl" groups and divalent "aryl" groups may also be referred to as "alkylene" or "alkenyl" groups, or "arylene" or "aryrenyl" groups, respectively. Similarly, unless otherwise explicitly indicated, if a combination of groups is referred to herein as one part, for example, arylalkyl, the last specified group contains the atoms to which this part is bonded in the remainder of the molecule.

[0374] "Optionally" or "optionally" means that the event or situation described thereafter may or may not occur, and this description includes both the cases in which such event or situation occurs and the cases in which it does not occur. Similarly, the term "optionally substituted" means that any one or more hydrogen atoms of the specified atom or group may or may not be replaced by a non-hydrogen part. "Optionally substituted" ranges from zero to the maximum number of possible substitutions and is independent of each occurrence. When the term "substituted" is used, the substitution must be made at the replaceable hydrogen atoms of the specified substituent. Optional substitutions may or may not be the same as (required) substitutions.

[0375] If a part is "substituted as needed," and refers to any of the general terms such as "alkyl," "alkenyl," "alkynyl," "haloalkyl," "cycloalkyl," "aryl," or "heteroaryl," then that general term is (C 1~3 (Alkyl), (C 4~6 Alkyl), -O(C 1~4 (Alkyl), (C 3~10 Cycloalkyl), O-(C 3~10 It can refer to any of the terms specifically listed before it, such as cycloalkyl. For example, "any aryl" includes both "aryl" and "-O(aryl)," as well as examples of aryls such as phenyl or naphthyl. Similarly, the term "any heterocyclyl" includes both the terms "heterocyclyl" and "O-(heterocyclyl)," as well as examples of heterocyclyls (such as oxetanyl, tetrahydropyranil, morpholino, and piperidinyl). Similarly, the term "any heteroaryl" includes both the terms "heteroaryl" and "O-(heteroaryl (heteroryl))." This includes certain heteroaryl compounds (such as pyridine).

[0376] Some compounds of formula (I) may exist as "stereoisomers" or mixtures of stereoisomers. Stereoiomers are compounds that are composed of the same atoms bonded together by the same bonds but have different three-dimensional structures and are not interconvertible. The compounds of this disclosure or their pharmaceutically acceptable salts may contain one or more chiral centers, thus resulting in enantiomers (two stereoisomers whose molecules are mirror images of each other but do not overlap), diastereomers, and other stereoisomers that can be defined as (R)- or (S)- with respect to absolute stereochemistry. This disclosure is intended to include all such possible isomers, as well as their racemic mixtures (i.e., equal amounts of (R) and (S) enantiomers) and optically pure forms. Optically active (+) and (-), (R)- and (S)- isomers can be prepared using chiral synthons or chiral reagents, or they can be separated using conventional techniques such as HPLC using chiral columns.

[0377] This disclosure also includes “deuterated analogs” of the compounds of formula I, in which 1 to n hydrogens (where n is the number of hydrogens in the molecule) bonded to a carbon atom are replaced by deuterium. Such compounds have shown improved resistance to metabolism and are therefore useful in increasing the half-life of any of the compounds of formula I when administered to mammals, particularly humans. See, for example, Foster, “Deuterium Isotope Effects in Studies of Drug Metabolism,” Trends Pharmacol. Sci. 5(12):524-527 (1984). Such compounds can be synthesized by means of art known, for example, by using starting materials in which one or more hydrogen atoms are replaced by deuterium.

[0378] Deuterium-labeled or substituted therapeutic compounds of the present disclosure may have improved DMPK (drug metabolism and pharmacokinetic) properties related to distribution, metabolism, and elimination (ADME). Substitution with heavier isotopes such as deuterium may result in certain therapeutic benefits, such as greater metabolic stability, e.g., increased in vivo half-life, reduced dose requirements, and / or improved therapeutic index. 18 Fluorine-labeled compounds may be useful for PET or SPECT studies. The isotope-labeled compounds of this disclosure can generally be prepared by performing the procedures disclosed in the scheme or examples and the preparations described below, by substituting readily available isotope-labeled reagents for unlabeled reagents. In this context, deuterium is understood to be a substituent in the compounds of formula I.

[0379] The concentration of such heavier isotopes, specifically deuterium, can be defined by the isotopic enrichment factor. In the compounds of this disclosure, any atom not specifically indicated as a particular isotope is intended to represent any stable isotope of that atom. Unless otherwise specified, where a position is specifically indicated as "H" or "hydrogen," that position is understood to have hydrogen in its naturally occurring isotopic composition. Therefore, in the compounds of this disclosure, any atom specifically indicated as deuterium (D) is intended to represent deuterium.

[0380] In many cases, the compounds of this disclosure can form acid salts and / or base salts due to the presence of an amino group and / or a carboxyl group, or a similar group.

[0381] Also provided herein are pharmaceutically acceptable salts, hydrates, or solvates of the compounds described herein. "pharmaceutically acceptable" or "physiologically acceptable" means compounds, salts, compositions, dosage forms, and other substances that are useful for preparing pharmaceutical compositions suitable for veterinary use or human medicinal use.

[0382] The term "pharmaceutically acceptable salt" of a given compound refers to a salt that retains the biological efficacy and properties of the given compound and is not biologically or otherwise undesirable. "pharmaceutically acceptable salt" or "physiologically acceptable salt" includes, for example, salts with inorganic acids and salts with organic acids. Furthermore, if the compounds described herein are obtained as acid addition salts, the free base can be obtained by making a solution of the acid salt basic. Conversely, if the product is a free base, the addition salt, in particular a pharmaceutically acceptable addition salt, can be produced by dissolving the free base in a suitable organic solvent and treating this solution with an acid, in accordance with the conventional procedure for preparing acid addition salts from basic compounds. Those skilled in the art are aware of the various synthetic methods that can be used to prepare non-toxic pharmaceutically acceptable addition salts. pharmaceutically acceptable acid addition salts can be prepared from inorganic and organic acids. Salts derived from inorganic acids include hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, and the like. Salts derived from organic acids include acetic acid, propionic acid, glycolic acid, pyruvic acid, oxalic acid, malic acid, malonic acid, succinic acid, maleic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, and salicylic acid. Similarly, pharmaceutically acceptable base addition salts can be prepared from inorganic and organic bases. Salts derived from inorganic bases include, but are not limited to, salts of sodium, potassium, lithium, ammonium, calcium, and magnesium.Salts derived from organic bases include, but are not limited to, alkylamines (i.e., NH2(alkyl)), dialkylamines (i.e., HN(alkyl)2), trialkylamines (i.e., N(alkyl)3), substituted alkylamines (i.e., NH2(substituted alkyl)), di(substituted alkyl)amines (i.e., HN(substituted alkyl)2), tri(substituted alkyl)amines (i.e., N(substituted alkyl)3), alkenylamines (i.e., NH2(alkenyl)), dialkenylamines (i.e., HN(alkenyl)2), and trialkenylamines (i.e., These include salts of primary, secondary, and tertiary amines such as N(alkenyl)3), substituted alkenylamines (i.e., NH2(substituted alkenyl)), di(substituted alkenyl)amines (i.e., HN(substituted alkenyl)2), tri(substituted alkenyl)amines (i.e., N(substituted alkenyl)3), mono, di, or tricycloalkylamines (i.e., NH2(cycloalkyl), HN(cycloalkyl)2, N(cycloalkyl)3), mono, di, or triarylamines (i.e., NH2(aryl), HN(aryl)2, N(aryl)3), or mixed amines. Specific examples of suitable amines include, but are not limited to, isopropylamine, trimethylamine, diethylamine, tri(isopropyl)amine, tri(n-propyl)amine, ethanolamine, 2-dimethylaminoethanol, piperazine, piperidine, morpholine, and N-ethylpiperidine.

[0383] The term “substituted” means that one or more hydrogen atoms on a specified atom or group are replaced by one or more substituents other than hydrogen, provided that the normal valency of the specified atom is not exceeded. The one or more substituents include, but are not limited to, alkyl, alkenyl, alkynyl, alkoxy, acyl, amino, amide, amidino, aryl, azide, carbamoyl, carboxyl, carboxyl ester, cyano, guanidino, halo, haloalkyl, haloalkoxy, heteroalkyl, heteroaryl, heterocyclyl, hydroxy, hydrazino, imino, oxo, nitro, alkylsulfinyl, sulfonic acid, alkylsulfonyl, thiocyanate, thiol, thion, or combinations thereof. Polymers or similar indeterminate structures achieved by defining substituents having an infinitely increasing number of further substituents (e.g., substituted aryls having a substituted alkyl, where the substituted alkyl is itself substituted by a substituted aryl group, and the substituted aryl group is further substituted by a substituted heteroalkyl group) are not intended to be included herein. Unless otherwise specified, the maximum number of consecutive substitutions in the compounds described herein is three. For example, consecutive substitution of a substituted aryl group by two other substituted aryl groups is limited to ((substituted aryl)substituted aryl)substituted aryl. Similarly, the above definitions are not intended to include unacceptable substitution patterns (e.g., a methyl group substituted with five fluorine atoms, or a heteroaryl group having two adjacent oxygen ring atoms). Such unacceptable substitution patterns are well known to those skilled in the art. The term “substituted” may describe other chemical groups as defined herein when used to modify a chemical group. Unless otherwise specified, if a group is described as being substituted as necessary, then any substituents on that group are themselves unsubstituted. For example, in some embodiments, the term “substituted alkyl” refers to an alkyl group having one or more substituents, including hydroxyl, halo, alkoxy, cycloalkyl, heterocyclyl, aryl, and heteroaryl.In other embodiments, one or more substituents may be further substituted with halo, alkyl, haloalkyl, hydroxyl, alkoxy, cycloalkyl, heterocyclyl, aryl, or heteroaryl compounds, each being substituted. In other embodiments, substituents may be further substituted with halo, alkyl, haloalkyl, alkoxy, hydroxyl, cycloalkyl, heterocyclyl, aryl, or heteroaryl compounds, each being unsubstituted. Those skilled in the art will recognize that the substituents and other parts of the compounds in the general formulas herein should be selected to result in compounds that are stable enough to yield pharmaceutically useful compounds that can be formulated into pharmaceutical compositions that are stable to an acceptable degree. Compounds having such stability are intended to be within the scope of the present invention. It should be understood by those skilled in the art that none of the above definitions and combinations of substituents should result in unmanipulable chemical species or compounds.

[0384] As used herein, “pharmaceutically acceptable carrier” or “pharmaceutically acceptable excipient” includes all kinds of solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic agents and absorption retarders, etc. The use of such media and agents for pharmaceutically active substances is well known in the art. Any conventional media or agent is intended for use in a therapeutic composition unless it is incompatible with the active ingredient. Co-active ingredients may also be incorporated into this composition.

[0385] A "solvate" is formed by the interaction of a solvent and a compound. Solvates of salts of the compounds described herein are also provided. Hydrations of the compounds described herein are also provided. Target IRAK4 decomposition

[0386] The compounds of this disclosure are demonstrated by cell-based profiling that selectively degrades IRAK4.

[0387] The degradation mechanisms and selectivity of two representative compounds (formula (IIA)) possessing LHMs targeting CRBN and two representative compounds (formula (IIB)) possessing LHMs targeting VHL were evaluated and discussed herein. For comparison, three compounds known to degrade IRAK4 were also evaluated. Table 1 shows the structures of the selected compounds. [Table 1-1] [Table 1-2] [Table 1-3]

[0388] More specifically, the cyto-degradation of IRAK4 by the selected compounds was evaluated using three different assay formats: HiBiT assay, HTRF assay, and Western blotting. All compounds showed consistent and reproducible degradation across these three assays. In particular, among the selected compounds, compound 47 was shown to be the most effective degrader in terms of Dmax, achieving 99% degradation as assessed by Western blot analysis. Furthermore, representative compounds showed equivalent or superior degradation (D) compared to known compounds with similar LHM (compounds a1, a2, and b1). max ) was shown.

[0389] Furthermore, to confirm that IRAK4 degradation is mediated by the ubiquitin-proteasome system, the compound of formula (I) was profiled in the presence of proteasome inhibitors, ligase inhibitors, or in excess concentrations of corresponding monofunctional compounds, such as compounds containing only the IRAK4 binding moiety or compounds containing only the LHM. Pretreatment under any of these conditions restored IRAK4 protein levels to those of untreated cells, demonstrating the on mechanism of the bifunctional compounds.

[0390] The specificity of IRAK4 degradation by the compounds disclosed herein was assessed by first estimating the degradation of the CRBN neosubstrates Ikaros, Aiolos, and GSPT1, and secondly estimating the effect of the degrading agents on the highly relevant target IRAK1. Profiling of the neosubstrates demonstrated that comparative compound a2, one of the known compounds, degraded both Ikaros and Aiolos, while none of compounds 13, 24, 47, and 35 showed any degradation of the neosubstrates. Furthermore, none of the assayed compounds affected IRAK1 levels, demonstrating specificity towards IRAK4 rather than IRAK1 degradation. Ultimately, none of the assayed compounds affected cell viability, as estimated by CellTiter-Glo. Table 2 summarizes the degradation results for selected compounds targeting CRBN.

[0391] [Table 2] Table 3 summarizes the degradation results for selected compounds targeting VHL.

[0392] [Table 3]

[0393] Pharmaceutical compositions and uses of the difunctional compound of formula (I)

[0394] The bifunctional compound of formula (I) degrades IRAK4 and is therefore demonstrated to be useful in treating disease indications or disorders involving IRAK4 functions such as signal transduction or scaffold formation.

[0395] Various embodiments provide pharmaceutical compositions comprising a compound of formula (I), or any one of the substructures or compounds in Table 5, and a pharmaceutically acceptable carrier.

[0396] Further embodiments provide a method for treating cancer, inflammatory disorders, autoimmune disorders, or metabolic disorders, comprising the step of administering to a subject in need of such treatment a therapeutically effective amount of a compound of formula (I), or one of its substructures or compounds from Table 5.

[0397] Examples of cancers that can be treated include lymphomas and leukemias, such as acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS).

[0398] Examples of metabolic disorders include, but are not limited to, diabetes mellitus including type 1 and type 2 diabetes, metabolic syndrome, dyslipidemia, obesity, glucose intolerance, hypertension, elevated serum cholesterol, and elevated triglycerides.

[0399] Examples of inflammatory disorders include rheumatoid arthritis (RA), inflammatory bowel disease (IBD), Crohn's disease, ulcerative colitis, necrotizing enterocolitis, gout, Lyme disease, arthritis, psoriasis, pelvic inflammatory disease, systemic lupus erythematosus (SLE), Sjögren's syndrome, inflammation associated with gastrointestinal infections including C. difficile, viral myocarditis, acute and chronic tissue injury, non-alcoholic steatohepatitis (NASH), alcoholic hepatitis, and renal diseases including chronic kidney disease and diabetic kidney disease.

[0400] Further embodiments provide a method for treating an inflammation-related disease or condition, or a metabolic disorder, gastrointestinal disorder, or cancer, the method comprising the step of administering a compound of formula (I) in combination with one or more compounds useful for treating such diseases to a subject in need, particularly a human subject.

[0401] In some embodiments, the compounds of the Disclosure are co-formulated with one or more additional active ingredients. In some embodiments, the other active ingredients are administered in separate dosage forms at approximately the same time. In some embodiments, the other active ingredients may be administered sequentially at different times with respect to the compounds of the Disclosure. [Examples]

[0402] Preparation of the compound of formula (I) (Example 1) [ka] N-(4-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazine-7-yl)-4-((tetrahydro-2H-pyran-4-yl)amino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl)bicyclo[2.2.2]octan-1-yl)-3-(2-(2-((2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-4-yl)amino)ethoxy)ethoxy)propanamide

[0403] A mixture of 7-(5-(5-(4-aminobicyclo[2.2.2]octan-1-yl)-1,3,4-thiadiazole-2-yl)-4-((tetrahydro-2H-pyran-4-yl)amino)pyridine-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitride bishydrochloride (BB1, 16.0 mg, 0.0249 mmol), 3-[2-[2-[[2-(2,6-dioxo-3-piperidyl)-1,3-dioxo-isoindorin-4-yl]amino]ethoxy]ethoxy]propanoic acid (13.0 mg, 0.0299 mmol), and HATU (9.97 mg, 0.0262 mmol) in DMF (0.125 mL) was mixed with DIPEA (0.0143 mL, 0.0799 mmol). The resulting solution was stirred at room temperature for 12 hours. The crude solution was purified by preparative HPLC (Gemini C18, eluent: 10-64% acetonitrile / H2O / 0.1% TFA) and freeze-dried to obtain N-(4-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-((tetrahydro-2H-pyran-4-yl)amino)pyridine-3-yl)-1,3,4-thiadiazole-2-6-yl)bicyclo[2.2.2]octan-1-yl)-3-(2-(2-((2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-4-yl)amino)ethoxy)ethoxy)propanamide as the TFA salt. ES / MS: 942.476(M+H + ); 1 H NMR (400 MHz, Methanol-d4) δ 10.17 (d, J = 7.7 Hz, 1H), 9.03 (s, 1H), 8.68 (s, 1H), 8.66 - 8.58 (m, 2H), 8.11 (d, J = 5.1 Hz, 1H), 7.79 (s, 1H), 7.59 (dd, J = 8.6, 7.1 Hz, 1H), 7.21 (d, J = 5.1 Hz, 1H), 7.08 (t, J = 7.8 Hz, 2H), 6.34 (s, 1H), 4.97 (dd, J = 12.2, 5.3 Hz, 1H), 4.01 (dt, J = 12.1, 3.9 Hz, 2H), 3.74 (t, J = 5.3 Hz, 2H), 3.70 - 3.56 (m, 9H), 3.49 (t, J = 5.3 Hz, 2H), 2.83 - 2.64 (m, 3H), 2.30 (t, J = 6.0 Hz, 2H), 2.19 - 2.11 (m, 2H), 2.11 - 2.01 (m, 9H), 1.85 - 1.73 (m, 3H). (Example 2) [ka]

[0404] N-(4-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazine-7-yl)-4-((tetrahydro-2H-pyran-4-yl)amino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl)bicyclo[2.2.2]octan-1-yl)-3-(2-(2-(2-((2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-4-yl)amino)ethoxy)ethoxy)ethoxy)propanamide

[0405] N-(4-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazine-7-yl)-4-((tetrahydro-2H-pyran-4-yl)amino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl)bicyclo[2.2.2]octan-1-yl)-3-(2-(2-(2-((2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-4-yl)amino)ethoxy)ethoxy)ethoxy)propanamide is 7-(5-(5-(4-aminobicyclo[2.2.2]octan-1-yl)-1,3,4-thiadiazole-2-yl)-4-((tetrahydro-2H-pyran-4-yl)amino)py Starting with lysine-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitride bishydrochloride (BB1, 18.0 mg, 0.0281 mmol), the preparation was carried out according to the procedure of Example 2, with 3-(2-(2-(2-((2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindolin-4-yl)amino)ethoxy)ethoxy)ethoxy)propanoic acid (14.8 mg, 0.0309 mmol) replaced with 3-[2-[2-[[2-(2,6-dioxo-3-piperidyl)-1,3-dioxoisoindolin-4-yl]amino]ethoxy]ethoxy]propanoic acid (13.0 mg, 0.0299 mmol). ES / MS: 986.572 (M+H + ); 1 H NMR (400 MHz, Acetonitrile-d3) δ 10.19 (d, J = 7.6 Hz, 1H), 9.12 (s, 1H), 8.68 - 8.59 (m, 3H), 8.11 (d, J = 5.1 Hz, 1H), 7.74 (s, 1H), 7.56 (dd, J = 8.6, 7.1 Hz, 1H), 7.22 (d, J = 5.1 Hz, 1H), 7.06 (dd, J = 19.2, 7.7 Hz, 2H), 6.34 (s, 1H), 4.96 (dd, J = 12.4, 5.4 Hz, 1H), 4.01 (dt, J = 11.9, 3.8 Hz, 2H), 3.72 (t, J = 5.3 Hz, 2H), 3.65 (tt, J = 5.3, 3.1 Hz, 6H), 3.61 - 3.54 (m, 3H), 3.49 (t, J = 5.3 Hz, 2H), 2.86 - 2.58 (m, 2H), 2.31 (t, J = 6.1 Hz, 2H), 2.19 - 2.03 (m, 12H), 1.84 - 1.73 (m, 2H). (Example 3) [ka]

[0406] Synthesis of N-((1r,4r)-4-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazine-7-yl)-4-(isopropylamino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl)cyclohexyl)-3-(2-(2-((2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-4-yl)amino)ethoxy)ethoxy)propanamide

[0407] A mixture of 7-(5-(5-((trans)-4-aminocyclohexyl)-1,3,4-thiadiazole-2-yl)-4-(isopropylamino)pyridine-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitride bishydrochloride (BB2, 10.0 mg, 0.0188 mmol), 3-(2-(2-((2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-4-yl)amino)ethoxy)ethoxy)propanoic acid (10.0 mg, 0.0231 mmol), and HATU (10.0 mg, 0.0263 mmol) in DMF (0.5 mL) was mixed with DIPEA (0.0170 mL, 0.0976 mmol). The resulting solution was stirred at room temperature for 20 minutes. The crude solution was purified by preparative HPLC (Gemini C18, eluent: 10-45% acetonitrile / H2O / 0.1% TFA) and freeze-dried to obtain N-((trans)-4-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(isopropylamino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl)cyclohexyl)-3-(2-(2-((2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-4-yl)amino)ethoxy)ethoxy)propenamide as the TFA salt. ES / MS: 874.659(M+H + ); 1 H NMR (400 MHz, Methanol-d4) δ 8.79 (d, J = 2.1 Hz, 1H), 8.72 (d, J = 1.8 Hz, 2H), 8.10 (d, J = 5.1 Hz, 1H), 7.97 (s, 1H), 7.58 (dd, J = 8.6, 7.1 Hz, 1H), 7.26 (d, J = 5.1 Hz, 1H), 7.13 (d, J = 8.5 Hz, 1H), 7.05 (d, J = 7.0 Hz, 1H), 5.09 (dd, J = 12.5, 5.5 Hz, 1H), 4.34 (p, J = 6.4 Hz, 1H), 3.77 (td, J = 5.6, 2.6 Hz, 5H), 3.73 - 3.62 (m, 3H), 3.60 - 3.48 (m, 2H), 3.23 - 3.09 (m, 1H), 2.89 (ddd, J = 17.7, 14.3, 5.0 Hz, 1H), 2.83 - 2.66 (m, 2H), 2.45 (t, J = 5.9 Hz, 2H), 2.30 - 1.99 (m, 5H), 1.81 - 1.63 (m, 2H), 1.52 (d, J = 6.4 Hz, 6H), 1.50 - 1.35 (m, 3H). (Example 4) [ka]

[0408] N-((1r,4r)-4-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazine-7-yl)-4-(isopropylamino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl)cyclohexyl)-3-(2-(2-(2-((2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-4-yl)amino)ethoxy)ethoxy)ethoxy)propanamide

[0409] N-((trans)-4-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazine-7-yl)-4-(isopropylamino)pyridine-3-yl)-1,3,4-thiadiazole-2-yl)cyclohexyl)-3-(2-(2-(2-((2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-4-yl)amino)ethoxy)ethoxy)ethoxy)propenamide is 7-(5-(5-((trans)-4-aminocyclohexyl)-1,3,4-thiadiazole-2-yl)-4-(isopropylamino)pyridine-2-yl)pyrrolo[1,2- b) Starting with pyridazine-3-carbonitride bishydrochloride (BB2, 10.0 mg, 0.0188 mmol), the solution was prepared according to the procedure of Example 3, replacing 3-(2-(2-((2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-4-yl)amino)ethoxy)ethoxy)propanoic acid (10.0 mg, 0.0231 mmol) with 3-(2-(2-((2-((2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-4-yl)amino)ethoxy)ethoxy)ethoxy)propanoic acid (10.0 mg, 0.0209 mmol). ES / MS: 918.750 (M+H + ); 1 1H NMR (400 MHz, Methanol-d4) δ 8.79 (d, J = 2.2 Hz, 1H), 8.72 (d, J = 2.2 Hz, 1H), 8.70 (s, 1H), 8.10 (d, J = 5.1 Hz, 1H), 7.97 (s, 1H), 7.55 (dd, J = 8.6, 7.1 Hz, 1H), 7.26 (d, J = 5.1 Hz, 1H), 7.11 (d, J = 8.6 Hz, 1H), 7.01 (d, J = 7.0 Hz, 1H), 5.07 (dd, J = 12.4, 5.5 Hz, 1H), 4.34 (p, J = 6.4 Hz, 1H), 3.80 - 3.72 (m, 5H), 3.70 (s, 4H), 3.69 - 3.59 (m, 4H), 3.53 (t, J = 5.2 Hz, 2H), 3.24 (tt, J = 12.0, 3.6 Hz, 1H), 2.89 (ddd, J = 17.8, 14.2, 5.2 Hz, 1H), 2.83 - 2.65 (m, 2H), 2.44 (t, J = 6.0 Hz, 2H), 2.27 (d, J = 13.1 Hz, 2H), 2.19 - 2.04 (m, 3H), 1.75 (qd, J = 13.0, 3.3 Hz, 2H), 1.52 (d, J = 6.4 Hz, 6H), 1.45 (dd, J = 12.8, 3.4 Hz, 2H). (Example 5) [ka] 7-(5-(5-(4-(8-((2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-5-yl)amino)octanoyl)piperazine-1-yl)-1,3,4-thiadiazole-2-yl)-4-(isopropylamino)pyridine-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile

[0410] HATU (19 mg, 0.05 mmol) and 8-{[2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindole-5-yl]aminooctanoic acid (14 mg, 0.03 mmol) were dissolved in DMF (0.15 M) and triethylamine (7 mg, 0.07 mmol). After stirring the reaction mixture at room temperature for 10 minutes, 7-[4-(isopropylamino)-5-[5-(piperazine-1-yl)-1,3,4-thiadiazole-2-yl]pyridine-2-yl]pyrrolo[1,2-b]pyridazine-3-carbonitrile (15 mg, 0.03 mmol) and BB4 were added. Next, the reaction mixture was stirred for 16 hours, then filtered through a syringe filter, and purified by HPLC to obtain 7-(5-{5-[4-(8-{[2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindole-5-yl]amino}octanoyl)piperazine-1-yl]-1,3,4-thiadiazole-2-yl}-4-(isopropylamino)pyridine-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile (3.6 mg, 13%). LCMS:C 43 H 46 N 12 O5S theoretical value: 843.0, measured value: m / z = 843.9 [M+H] + ; 1 H NMR (500 MHz, DMSO-d6) δ 11.06 (s, 1H), 9.50 (s, 1H), 8.98 (d, J = 2.1 Hz, 1H), 8.85 (d, J = 2.1 Hz, 1H), 8.57 (s, 1H), 8.06 (d, J = 4.9 Hz, 1H), 8.02 (s, 1H), 7.57 (d, J = 8.3 Hz, 1H), 7.24 (d, J = 5.0 Hz, 1H), 7.12 (s, 1H), 6.95 (d, J = 2.1 Hz, 1H), 6.85 (dd, J = 8.2, 2.1 Hz, 1H), 6.56 (s, 1H), 5.03 (dd, J = 12.7, 5.4 Hz, 1H), 4.18 (s, 1H), 3.68 (d, 2.60 (s, 1H), 2.39 (d, J = 7.5 Hz, 1H), 2.00 (d, J = 12.9 Hz, 1H), 1.62 - 1.54 (m, 2H), 1.53 (d, J = 6.8 Hz, 3H), 1.38 (d, J = 6.4 Hz, 8H), 1.36 - 1.32 (m, 6H). (Example 6) [ka] 7-(5-(5-(4-(6-((2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-5-yl)amino)hexanoyl)piperazine-1-yl)-1,3,4-thiadiazole-2-yl)-4-(isopropylamino)pyridine-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile

[0411] The title compound was synthesized from BB4 and 6-((2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-5-yl)amino)hexanoic acid by amide coupling using general method A. LCMS:C 41 H 42 N 12 O5S theoretical value: 814.3, measured value: m / z = 815.9 [M+H] + ; 1 1H NMR (500 MHz, DMSO-d6) δ 11.06 (s, 1H), 9.20 (s, 1H), 8.94 (d, J = 2.3 Hz, 1H), 8.82 (d, J = 2.2 Hz, 1H), 8.56 (s, 1H), 8.09 (s, 1H), 8.00 (d, J = 4.7 Hz, 1H), 7.55 (dd, J = 19.1, 8.3 Hz, 1H), 7.20 (d, J = 4.8 Hz, 1H), 7.13 (s, 1H), 6.96 (d, J = 2.2 Hz, 1H), 6.85 (ddd, J = 10.6, 8.3, 2.1 Hz, 1H), 6.54 (s, 3H), 5.03 (dd, J = 12.7, 5.2 Hz, 1H), 4.11 (s, 1H), 3.67 (d, J = 5.3 Hz, 2H), 3.18 (s, 2H), 2.88 (ddd, J = 16.5, 13.6, 5.4 Hz, 1H), 2.60 (s, 1H), 2.48 (s, 2H), 2.41 (t, J = 7.3 Hz, 1H), 2.00 (d, J = 12.9 Hz, 1H), 1.60 (tt, J = 15.2, 7.6 Hz, 3H), 1.43 (d, J = 7.5 Hz, 1H), 1.38 (d, J = 6.4 Hz, 6H). (Example 7) [ka]

[0412] 7-(5-(5-(4-(3-(2-((2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-4-yl)amino)ethoxy)propanoyl)piperazine-1-yl)-1,3,4-thiadiazole-2-yl)-4-(isopropylamino)pyridine-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile

[0413] The title compound was synthesized from BB4 and 3-(2-((2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-4-yl)amino)ethoxy)propanoic acid by amide coupling using general method A. LCMS:C 40 H 40 N 12O6S Theoretical value: 816.3 Measured value: m / z = 817.7 [M+H] + ; 1 H NMR (500 MHz, DMSO-d6) δ 11.11 (s, 1H), 9.37 (s, 1H), 8.97 (d, J = 2.1 Hz, 1H), 8.84 (d, J = 2.2 Hz, 1H), 8.52 (s, 1H), 8.05 (d, J = 6.1 Hz, 2H), 7.61 - 7.55 (m, 1H), 7.23 (d, J = 4.9 Hz, 1H), 7.14 (d, J = 8.6 Hz, 1H), 7.00 (d, J = 7.0 Hz, 1H), 6.56 (t, J = 5.5 Hz, 1H), 5.06 (dd, J = 12.8, 5.4 Hz, 1H), 4.16 (s, 1H), 3.73 (t, J = 6.3 Hz, 2H), 3.66 (dt, J = 14.4, 5.2 Hz, 6H), 3.61 - 3.52 (m, 4H), 3.48 (q, J = 5.4 Hz, 2H), 2.90 (ddd, J = 17.4, 13.8, 5.5 Hz, 1H), 2.67 (t, J = 6.3 Hz, 2H), 2.63 (s, 1H), 2.57 (d, J = 15.6 Hz, 1H), 2.10 - 2.03 (m, 1H), 1.39 (d, J = 6.3 Hz, 6H). (Example 8) [ka] 7-(5-(5-(4-(8-((2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-4-yl)amino)octanoyl)piperazine-1-yl)-1,3,4-thiadiazole-2-yl)-4-(isopropylamino)pyridine-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile

[0414] The title compound was synthesized from BB4 and 8-((2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-4-yl)amino)octanoic acid by amide coupling using general method A. LCMS:C 43 H 46 N 12 O5S theoretical value: 842.3, measured value: m / z = 843.8 [M+H] + ; 1 1H NMR (500 MHz, DMSO-d6) δ 11.10 (s, 1H), 9.42 (s, 1H), 8.97 (d, J = 2.2 Hz, 1H), 8.84 (d, J = 2.2 Hz, 1H), 8.56 (s, 1H), 8.04 (d, J = 6.6 Hz, 2H), 7.59 (dd, J = 8.6, 7.0 Hz, 1H), 7.23 (d, J = 4.9 Hz, 1H), 7.11 (d, J = 8.6 Hz, 1H), 7.03 (d, J = 7.0 Hz, 1H), 6.54 (t, J = 6.0 Hz, 1H), 5.06 (dd, J = 12.7, 5.5 Hz, 1H), 4.16 (s, 1H), 3.67 (dd, J = 7.1, 3.7 Hz, 4H), 3.57 (d, J = 5.5 Hz, 2H), 3.31 (q, J = 6.4 Hz, 2H), 2.89 (ddd, J = 16.8, 13.7, 5.4 Hz, 1H), 2.63 - 2.56 (m, 1H), 2.49 (s, 1H), 2.38 (t, J = 7.4 Hz, 2H), 2.08 - 2.01 (m, 1H), 1.59 (t, J = 6.9 Hz, 2H), 1.53 (t, J = 7.2 Hz, 2H), 1.40 - 1.33 (m, 12H). (Example 9) [ka] 7-(5-(5-(4-(6-((2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-4-yl)amino)hexanoyl)piperazine-1-yl)-1,3,4-thiadiazole-2-yl)-4-(isopropylamino)pyridine-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile

[0415] The title compound was synthesized from BB4 and 6-((2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-4-yl)amino)hexanoic acid by amide coupling using general method A. LCMS:C 41 H 42 N 12 O5S theoretical value: 814.3, measured value: m / z = 817.7 [M+H] + ; 1 1H NMR (500 MHz, DMSO-d6) δ 11.10 (s, 1H), 9.26 (s, 1H), 8.95 (d, J = 2.3 Hz, 1H), 8.82 (d, J = 2.3 Hz, 1H), 8.56 (s, 1H), 8.08 (s, 1H), 8.01 (d, J = 4.9 Hz, 1H), 7.63 - 7.56 (m, 1H), 7.21 (d, J = 4.9 Hz, 1H), 7.12 (d, J = 8.6 Hz, 1H), 7.02 (d, J = 7.0 Hz, 1H), 6.55 (t, J = 6.2 Hz, 1H), 5.06 (dd, J = 12.8, 5.4 Hz, 1H), 4.12 (s, 1H), 3.67 (dd, J = 6.9, 3.7 Hz, 4H), 3.56 (d, J = 5.4 Hz, 2H), 3.32 (q, J = 6.6 Hz, 2H), 2.90 (ddd, J = 16.8, 13.8, 5.4 Hz, 1H), 2.65 - 2.56 (m, 1H), 2.41 (t, J = 7.4 Hz, 2H), 2.07 - 2.01 (m, 1H), 1.60 (dp, J = 15.1, 7.3 Hz, 4H), 1.38 (d, J = 6.4 Hz, 8H). (Example 10) [ka]

[0416] 7-(5-(5-(4-(3-(2-((2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-5-yl)amino)ethoxy)propanoyl)piperazine-1-yl)-1,3,4-thiadiazole-2-yl)-4-(isopropylamino)pyridine-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile

[0417] The title compound was synthesized from BB4 and 3-(2-((2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-5-yl)amino)ethoxy)propanoic acid by amide coupling using general method A. LCMS:C 40 H 40 N 12 O6S theoretical value: 816.3, measured value: m / z = 817.6 [M+H] + ; 1H NMR (500 MHz, DMSO-d6) δ 11.05 (s, 1H), 9.43 (s, 1H), 8.97 (d, J = 2.2 Hz, 1H), 8.84 (d, J = 2.2 Hz, 1H), 8.54 (s, 1H), 8.07 - 8.02 (m, 2H), 7.54 (d, J = 8.4 Hz, 1H), 7.23 (d, J = 4.9 Hz, 1H), 7.15 (s, 1H), 7.01 (d, J = 2.2 Hz, 1H), 6.90 (dd, J = 8.4, 2.2 Hz, 1H), 5.01 (dd, J = 12.9, 5.4 Hz, 1H), 4.16 (q, J = 6.6 Hz, 1H), 3.76 - 3.58 (m, 8H), 3.57 (s, 2H), 3.36 (t, J = 5.4 Hz, 2H), 2.85 (ddd, J = 17.4, 14.0, 5.5 Hz, 1H), 2.68 (t, J = 6.4 Hz, 2H), 2.58 - 2.53 (m, 1H), 2.50 - 2.43 (m, 0H), 1.97 (dtd, J = 13.0, 6.1, 2.9 Hz, 1H), 1.38 (d, J = 6.3 Hz, 6H). (Example 11) [ka]

[0418] 7-(5-(5-(4-(3-(2-(2-(2-((2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-4-yl)amino)ethoxy)ethoxy)ethoxy)propanoyl)piperazine-1-yl)-1,3,4-thiadiazole-2-yl)-4-(isopropylamino)pyridine-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile

[0419] The title compound was synthesized from BB4 and 3-(2-(2-(2-((2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-4-yl)amino)ethoxy)ethoxy)ethoxy)propanoic acid by amide coupling using general method A. LCMS:C 44 H 48 N 12 O8S Theoretical value: 904.3, Measured value: m / z = 906.1 [M+H] + ; 1 H NMR (500 MHz, DMSO-d6) δ 11.10 (s, 1H), 8.96 (d, J = 2.1 Hz, 1H), 8.83 (d, J = 2.2 Hz, 1H), 8.53 (s, 1H), 8.07 - 8.00 (m, 2H), 7.56 (t, J = 7.8 Hz, 1H), 7.22 (d, J = 4.9 Hz, 1H), 7.13 (d, J = 8.6 Hz, 1H), 7.01 (d, J = 7.0 Hz, 1H), 6.59 (t, J = 5.7 Hz, 1H), 5.06 (dd, J = 12.7, 5.4 Hz, 1H), 4.13 (s, 1H), 3.65 (dp, J = 17.0, 5.3 Hz, 10H), 3.59 - 3.46 (m, 10H), 3.46 (d, J = 5.6 Hz, 2H), 2.89 (ddd, J = 17.5, 13.8, 5.3 Hz, 1H), 2.67 - 2.56 (m, 3H), 2.05 (dd, J = 9.9, 4.4 Hz, 1H), 1.37 (d, J = 6.3 Hz, 6H). (Example 12) [ka]

[0420] 7-(5-(5-(4-(1-(((3R)-1-(2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-5-yl)pyrroridine-3-yl)methyl)piperidine-4-carbonyl)piperazine-1-yl)-1,3,4-thiadiazole-2-yl)-4-(isopropylamino)pyridine-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile

[0421] The title compound was synthesized from BB5 and (3S)-1-(2-(2,6-dioxopiperidine-3-yl)-1,3-dioxoisoindorin-5-yl)pyrrolidine-3-carboaldehyde by reductive amination using general method B. LCMS:C 46 H 49 N 13 O5S theoretical value: 895.4, measured value: m / z = 896.8 [M+H] + ; 1 H NMR (500 MHz, DMSO-d6) δ 11.09 (s, 1H), 9.28 (s, 1H), 9.13 (s, 2H), 8.94 (d, J = 2.3 Hz, 1H), 8.81 (d, J = 2.2 Hz, 1H), 8.58 (s, 1H), 8.11 (s, 1H), 7.99 (d, J = 4.8 Hz, 1H), 7.74 - 7.68 (m, 1H), 7.21 (d, J = 4.9 Hz, 1H), 6.96 (d, J = 2.2 Hz, 1H), 6.85 (dd, J = 8.5, 2.2 Hz, 1H), 5.08 (dd, J = 12.8, 5.4 Hz, 1H), 4.10 (s, 1H), 3.98 (s, 3H), 3.77 (s, 2H), 3.61 - 3.56 (m, 4H), 3.43 (q, J = 8.5 Hz, 1H), 3.27 (dt, J = 27.8, 7.7 Hz, 2H), 3.04 (s, 2H), 3.01 (s, 1H), 2.92 - 2.83 (m, 2H), 2.64 - 2.57 (m, 1H), 2.28 (s, 1H), 2.03 (d, J = 12.1 Hz, 1H), 1.93 (s, 5H), 1...

Claims

【Request Item 1】 【Chemistry 451-1】 【Chemistry 451-2】 【Chemistry 451-3】 【Chemistry 451-4】 【Chemistry 451-5】 【Chemistry 451-6】 【Chemistry 451-7】 【Chemistry 451-8】 【Chemistry 451-9】 【Chemistry 451-10】 【Chemistry 451-11】 【Chemistry 451-12】 【Chemistry 451-13】 【Chemistry 451-14】 【Chemistry 451-15】 【Chemistry 451-16】 【Chemistry 451-17】 【Chemistry 451-18】 【Chemistry 451-19】 【Chemistry 451-20】 【Chemistry 451-21】 A compound having the structure.

2. A pharmaceutical composition comprising the compound described in Claim 1 and a pharmaceutically acceptable carrier.

3. A composition comprising the compound according to claim 1 or a pharmaceutical composition according to claim 2 for use in the treatment of cancer.

4. The composition according to claim 3, wherein the cancer is lymphoma, leukemia, acute myeloid leukemia (AML), and myelodysplastic syndrome (MDS).

5. A composition comprising the compound according to claim 1 or a pharmaceutical composition according to claim 2 for use in the treatment of metabolic disorders.

6. The composition according to claim 5, wherein the metabolic disorder is diabetes (type 1 and type 2 diabetes), metabolic syndrome, dyslipidemia, obesity, glucose intolerance, hypertension, elevated serum cholesterol, and elevated triglycerides.

7. A composition comprising the compound according to claim 1 or a pharmaceutical composition according to claim 2 for use in the treatment of inflammatory disorders.

8. The composition according to claim 7, wherein the inflammatory disorder includes rheumatoid arthritis (RA), inflammatory bowel disease (IBD), Crohn's disease, ulcerative colitis, necrotizing enterocolitis, gout, Lyme disease, arthritis, psoriasis, pelvic inflammatory disease, systemic lupus erythematosus (SLE), Sjögren's syndrome, inflammation associated with gastrointestinal infections including C. difficultile, viral myocarditis, acute and chronic tissue injury, non-alcoholic steatohepatitis (NASH), alcoholic hepatitis, and kidney diseases including chronic kidney disease and diabetic kidney disease.

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