Cyclic peptides for capturing interleukin-1β

Abstract

There is provided a compound of formula (I) or a pharma- ceutically acceptable salt thereof, wherein X 1 , X 2 , X 3 , A 1 , A 2 , R 1 ~R 7 , R 8a , R 8b , R 9a , R 9b , R 10 and R 11 is as described herein. (I) is capable of sequestering IL-1β and is expected to have utility as a therapeutic agent, for example, for the treatment of cardiovascular disease and inflammatory disorders. The present disclosure also provides pharmaceutical compositions comprising the compounds disclosed herein or pharma- ceutically acceptable salts thereof. The present disclosure also relates to methods of using the compounds or pharma- ceutically acceptable salts thereof in the therapy and prevention of cardiovascular disease and inflammatory disorders, as well as methods of making medicaments for that purpose. [Formula 1] TIFF2025505490000069.tif90131

Description

[Technical field]

[0001] The present disclosure relates to certain cyclic peptides that trap interleukin-1β (IL-1β), pharmaceutical compositions containing such peptides, and methods of using such compounds to treat, inhibit, or ameliorate one or more cardiovascular disease conditions that may benefit from trapping IL-1β, such as atherosclerosis.

[0002] The XML file, created on May 25, 2023, is named 25545-WO-PCT_SL.XML and is 916,839 bytes in size. [Background technology]

[0003] Atherosclerosis is an arterial disease characterized by the buildup of cholesterol plaque on the inner walls of arteries. As atherosclerosis progresses, arteries become harder and narrower, increasing the risk of plaque rupture. When plaque ruptures, cholesterol globules and other materials are released into the bloodstream, which can result in restricted blood flow to the brain, heart, and other organs. Medically, these are known as major adverse cardiac events (MACE).

[0004] Risk factors for the development and progression of atherosclerotic cardiovascular disease (ASCVD) include high cholesterol, high blood pressure, a diet high in saturated fat, smoking, obesity, diabetes, physical inactivity, and elevated levels of C-reactive protein (CRP), a marker of inflammation.

[0005] The first-line treatment to prevent the progression of ASCVD is a healthy diet and exercise, but adherence is generally low. Pharmacological treatment of ASCVD has focused primarily on cholesterol-lowering drugs such as statins, cholesterol absorption inhibitors, and low-density lipoprotein (LDL) receptor inhibitors. These drugs are highly effective in reducing the buildup of fatty acid deposits and improving arterial health. Other drugs prescribed for ASCVD that do not modify the condition include anticoagulants such as aspirin, which prevent platelet aggregation in narrowed arteries, and blood pressure medications, which reduce the risk and severity of heart attacks. Surgical options for more aggressive interventions when ASCVD progresses include angioplasty, stent placement, endarterectomy (surgical removal of plaque), and bypass surgery.

[0006] Although cholesterol-lowering drugs have served as an important standard of care to slow the progression of atherosclerosis, clinical data support an even more important role for inflammation in the progression of ASCVD that remains untreated. Inflammatory biomarkers such as CRP are associated with increased risk of cardiovascular events, independent of cholesterol levels. The Canakinumab Anti-Inflammatory Thrombosis Outcomes Trial (CANTOS) was the first clinical trial to show that reducing vascular inflammation without concomitant lipid lowering reduces the incidence of cardiovascular events. N Engl J Med 2017;377:1119-1131. Canakinumab is an anti-interleukin-1β (IL-1β) human monoclonal antibody approved for clinical use in rheumatic disorders. IL-1β is a proinflammatory cytokine that induces IL-6, thereby elevating the downstream inflammatory biomarker high-sensitivity CRP (hsCRP). CANTOS therefore provides proof of concept that IL-1β-targeted therapies may reduce the incidence of MACE in select patients, complementing, and potentially adding to, standard LDL-lowering therapies. [Prior art documents] [Non-patent literature]

[0007] [Non-Patent Document 1] N Engl J Med 2017;377:1119-1131 Summary of the Invention [Problem to be solved by the invention]

[0008] Additional non-surgical therapies beyond standard-of-care cholesterol-lowering drugs are needed to slow atherosclerosis progression and reduce the risk of MACE. Furthermore, patients suffering from inflammatory disorders would benefit from an orally administered drug that inhibits the same cytokine IL-1β as canakinumab. [Means for solving the problem]

[0009] The present disclosure provides certain cyclic peptides that bind to IL-1β cytokines, thereby reducing inflammation and preventing their binding to IL-1 receptors, thereby inhibiting downstream proinflammatory signaling. These cyclic peptides can be pharma- ceutical active compounds useful for the treatment of cardiovascular disease and inflammatory disorders. In one aspect, the present disclosure provides compounds of formula (I): [ka] and pharma- ceutically acceptable salts thereof.

[0010] These compounds can capture IL-1β, thereby affecting downstream proinflammatory signaling pathways that may be associated with cardiovascular disorders. Thus, in another aspect, the present invention provides a method for treating cardiovascular disease (e.g., atherosclerosis, vascular inflammation), comprising administering to a subject in need of treatment a therapeutically effective amount of a compound of the present disclosure. In some embodiments, the administration comprises oral administration of the compound.

[0011] The present disclosure further provides methods for making the compounds of the present disclosure, as well as pharmaceutical compositions comprising a compound of the present disclosure and a pharma- ceutically acceptable carrier. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0012] Compounds of the Disclosure In one embodiment, the present disclosure provides a method for producing a pharmaceutical composition comprising: R 1 is CH3C(O)NH-CH2CH2-O- or C 1 and; C 1 but, (i) a 5-6 membered monocyclic aryl or heteroaryl, the heteroaryl containing 1-2 heteroatoms selected from the group consisting of N, O, and S; or (ii) a 5-6 membered monocyclic or bicyclic, saturated cycloalkyl or heterocyclic alkyl containing 1-2 heteroatoms selected from the group consisting of N, O, and S; or (iii) 5-6 membered monocyclic or bicyclic cycloalkyl and; Here, C 1 is unsubstituted or is substituted with 1 to 3 R independently selected from the group consisting of halo, C1-C3 alkyl, C1-C3 fluoroalkyl, carboxy, C1-C3 alkoxy, and C2-C3 acyl. C1 is substituted by a substituent; R 2 is H, C1-C3 alkyl, benzyl, or phenyl-CH2CH2-; R 3 is a 9-10 membered bicyclic aryl or heteroaryl, said heteroaryl containing 1-2 heteroatoms selected from the group consisting of N, O, and S; Here, R 3 is unsubstituted or is substituted with 1 to 3 R independently selected from the group consisting of halo, C1-C3 alkyl, C1-C3 fluoroalkyl, hydroxy, and C1-C3 alkoxy; 3a is substituted by a substituent; R 4 C1-C3 alkyl, HO2C-(CH2) m -, H2NC(O)-(CH2) m -, (CH3)2NC(O)-(CH2) m - or tetrazolyl-(CH2) m - and; R5 But amino, H2N(CH2) n -, H2NC(O)-(CH2) n -, CH3C(O)NH-, CH3C(O)NH(CH2) n -, C 5 , or C 5 -CH2-, C 5 but (i) 5-6 membered monocyclic aryl or heteroaryl, wherein the heteroaryl contains 1-2 heteroatoms selected from the group consisting of N, O, and S; (ii) a 9-10 membered bicyclic aryl or heteroaryl, wherein the bicyclic heteroaryl contains 1-3 heteroatoms selected from the group consisting of N, O, and S; (iii) 5- to 6-membered monocyclic or 9- to 10-membered heterocyclic alkyl [the heterocyclic alkyl is saturated or partially unsaturated and contains 1 to 2 heteroatoms selected from the group consisting of N, O, and S]; (iv) 5-6 membered monocyclic cycloalkyl; (v) 2,3-dihydroindolyl and; Here, C 5 is unsubstituted or independently selected from halo, amino, hydroxy, C1-C3 alkyl, C1-C3 fluoroalkyl, C1-C3 alkoxy, H2N-(CH2) k -, H2NC(O)-(CH2) k R is selected from the group consisting of -, H2C=CH-CH2O-, and phenyl; C5 is substituted by a substituent; R 6 H, C1-C5 alkyl, H2N(CH2) p -, HOCH2-, (CH3)2NCH2-, H3CO-(CH2) q - or C 6 -CH2-; C 6 is a 5- or 6-membered monocyclic saturated heterocycloalkyl containing 1 to 2 heteroatoms selected from the group consisting of N, O, and S; 6is unsubstituted or is substituted with 1 to 3 R independently selected from the group consisting of halo, C1-C3 alkyl, C1-C3 fluoroalkyl, hydroxy, and C1-C3 alkoxy; C6 is substituted by a substituent; R 7 is H or C1-C3 alkyl; R 8a H, C1-C5 alkyl, HOCH2-, H2N(CH2) r -, (CH3)3N + (CH2) r - or CH3C(O)NH(CH2) r - and; R 8b is H or C1-C3 alkyl; R 9a is H or C1-C3 alkyl; R 9b H, C1-C5 alkyl, C 9 -CH2- or C 9 -CH2CH2-; C 9 but, (i) a 5-6 membered monocyclic aryl or heteroaryl, the heteroaryl containing 1-2 heteroatoms selected from the group consisting of N, O, and S; or (ii) 5-6 membered monocyclic, saturated cycloalkyl or heterocyclic alkyl, wherein the heterocyclic alkyl contains 1-2 heteroatoms selected from the group consisting of N, O, and S. and; Here, C 9 is unsubstituted or independently selected from halo, amino, hydroxy, cyano, C1-C3 alkyl, C1-C3 fluoroalkyl, C1-C3 alkoxy, H2N-(CH2) k -, H2NC(O)-(CH2) k R is selected from the group consisting of -, H2NCH2CH2O-, CH3C(O)NH-CH2CH2O-, and morpholinyl-CH2CH2O- C9 is substituted by a substituent; R 10 is H, halo, or C1-C3 alkyl; R 11 is H, halo, or C1-C3 alkyl; each occurrence of subscript k is independently 1 or 2; subscript m is 1 or 2; The subscript n is 1, 2, 3, or 4; subscript p is 1, 2, 3, or 4; The subscript q is 1 or 2; The subscript r is 1, 2, 3, or 4; X 1 , X 2 , and X 3 is independently C(H) or N; and A 1 and A 2 are independently selected from the group consisting of HO2C-, H2NC(O)-, CH3C(O)N(H)-, H2NS(O)2-, CH3S(O)2N(H)-, tetrazolyl, and 5-oxooxadiazolyl, or a pharma- ceutically acceptable salt thereof.

[0013] In another embodiment, the present disclosure provides: R 5 But amino, H2N(CH2) n -, H2NC(O)-(CH2) n -, C 5 , or C 5 -CH2-, C 5 but (i) 5-6 membered monocyclic aryl or heteroaryl [the heteroaryl contains 1-2 heteroatoms selected from the group consisting of N, O, and S]: (ii) a 9-10 membered bicyclic aryl or heteroaryl, the bicyclic heteroaryl containing 1-3 heteroatoms selected from the group consisting of N, O, and S; (iii) a 5- to 6-membered monocyclic or 9- to 10-membered heterocyclic alkyl group, the heterocyclic alkyl group being saturated or partially unsaturated and containing 1 to 2 heteroatoms selected from the group consisting of N, O, and S; or (iv) 2,3-dihydroindolyl and Here, C 5 are unsubstituted or independently selected from halo, amino, hydroxy, C1-C3 alkyl, C1-C3 fluoroalkyl, C1-C3 alkoxy, H2N-(CH2) k -, H2NC(O)-(CH2) k R is selected from the group consisting of -, H2C=CH-CH2O-, and phenyl; C5 is substituted by a substituent; R 6 H, C2-C5 alkyl, H2N(CH2) p -, HOCH2-, (CH3)2NCH2-, H3CO-(CH2) q - or C 6 -CH2-; R 8a is H, C1-C3 alkyl, HOCH2-, or H2N(CH2) r - and; R 9a but H; R 9b H, C1-C3 alkyl, C 9 -CH2- or C 9 -CH2CH2-; R 10 but H; R 11 is H, and the subscript p is 1, 2, or 3; and Compounds of formula (I) are provided, wherein the subscript r is 2, 3, or 4.

[0014] In one embodiment, the compound of formula (I) has the following structural formula (IA): [ka]

[0015] In another embodiment, the present disclosure provides: C 1 is phenyl, pyrimidyl, or piperazinyl, where C 1 is not substituted or 1-2 RC 1is substituted by a substituent; R 3 is naphthyl or indolyl, where R 3 is unsubstituted or has 1 to 2 R 3a is substituted by a substituent; C 5 is phenyl, pyridyl, pyrimidyl, naphthyl, indolyl, 7-azaindolyl, indazolyl, 2,3-dihydroindolyl, piperidinyl, tetrahydropyranyl, or cyclohexyl, where C 5 is unsubstituted or has 1 to 2 R C5 has been replaced by; C 6 is tetrahydropyranyl or morpholinyl; 6 is unsubstituted or has 1 to 2 R C6 has been replaced by; C 9a is H or methyl; C 9b is phenyl, pyridyl, cyclohexyl, morpholinyl, or piperidinyl, where C 9 is unsubstituted or has 1 to 2 R C9 The present invention provides a compound of formula (I) substituted by:

[0016] In another embodiment, the present disclosure provides: X 1 and X 2 is C(H); R 1 is phenyl substituted by carboxy.

[0017] In another embodiment, the present disclosure provides: X 1 and X 2 is C(H); R 1 is CH3C(O)NH-CH2CH2-O-.

[0018] In another embodiment, the present disclosure provides2 is H.

[0019] In another embodiment, the present disclosure provides 3 is indolyl substituted by one halo.

[0020] In another embodiment, the present disclosure provides 3 is naphthyl.

[0021] In another embodiment, the present disclosure provides 4 HO2C-(CH2) m The present invention provides a compound of formula (I),

[0022] In another embodiment, the present disclosure provides a method for the preparation of 3 is C(H).

[0023] In another embodiment, the present disclosure provides 5 But H2N(CH2) n -, indole, 7-azaindole, naphthyl or pyridyl.

[0024] In another embodiment, the present disclosure provides 5 But H2N(CH2) n -, indole, naphthyl or pyridyl.

[0025] In another embodiment, the present disclosure provides 6 is H, HOCH2-, C2-C5 alkyl or H2N(CH2) p The present invention provides a compound of formula (I),

[0026] In another embodiment, the present disclosure provides 6 is C2-C5 alkyl or H2N(CH2) p The present invention provides a compound of formula (I),

[0027] In another embodiment, the present disclosure provides7 is H.

[0028] In another embodiment, the present disclosure provides: R 8a is methyl or H2NCH2CH2-; R 8b is H.

[0029] In another embodiment, the present disclosure provides 9b but [ka] The present invention provides a compound of formula (I):

[0030] In another embodiment, the present disclosure provides 9b but [ka] The present invention provides a compound of formula (I):

[0031] In one embodiment, the present disclosure provides a method for the preparation of a 1 is selected from the group consisting of HO2C-, H2NC(O)-, CH3C(O)N(H)-, H2NS(O)2-, CH3S(O)2N(H)-, tetrazolyl, and 5-oxooxadiazolyl. Those skilled in the art of chemistry will appreciate that such moieties in the amino acid residues can be represented by the following substructures: [ka]

[0032] In one embodiment, the present disclosure provides a method for the preparation of a 2is selected from the group consisting of HO2C-, H2NC(O)-, CH3C(O)N(H)-, H2NS(O)2-, CH3S(O)2N(H)-, tetrazolyl, and 5-oxooxadiazolyl. Those skilled in the art of chemistry will appreciate that such moieties in the amino acid residues can be represented by the following substructures:

[0033] [ka]

[0034] In another embodiment, the present disclosure provides: R 10 but H; R 11 is H, F or Cl.

[0035] In another embodiment, the present disclosure provides a method for producing 1 and A 2 and R are both HO2C- (i.e., carboxy).

[0036] In some embodiments, the present disclosure provides: R 1 is 4-CH3C(O)-piperazin-1-yl, CH3C(O)NH-CH2CH2-O-, 5-CO2H-pyrimidin-2-yl, or 4-CO2H-phenyl; R 2 is H, ethyl, benzyl, or phenyl-CH2CH2-; R 3 is naphth-1-yl, 4-fluoroindol-3-yl, or 4-chloroindol-3-yl; R 4 But methyl, HO2C-(CH2) m - or H2NC(O)-(CH2) m - and; R 5 But amino, H2N(CH2) n-, naphth-1-yl, indol-3-yl, 7-aza-indol-3-yl, indazol-1-yl, 2,3-dihydroindol-1-yl, pyrid-3-yl, pyrid-4-yl, piperidin-4-yl, 3-aminomethylphenyl, 4-aminomethylphenyl, 3-aminophenyl, 4-aminophenyl, phenyl, pyrid-4-yl-CH-, pyrimidin-5-yl, tetrahydropyran-4-yl-, HNC(O)-(CH)-, 3-biphenyl, 3-CH=CH-CHO-phenyl, CHC(O)NH-, CHC(O)NH(CH)-, or cyclohexyl; R 6 is H, (CH3)2CHCH2-, (CH3)3CCH2-, H2N(CH2) p -, HOCH2-, H3CCH2CH2-, morpholin-4-yl-CH2-, tetrahydropyran-4-yl-CH2-, (CH3)2NCH2-, or H3CO-(CH2) q - and; R 7 is H or methyl; R 8a H, methyl, HOCH2-, H2N(CH2) r -, (CH3)3NCH2CH2-, or CH3C(O)NH(CH2)4-; R 8b is H or methyl; R 9a is H or methyl; R 9b is H, methyl, HCCHCHCHCH-, 4-HO-phenyl-CHCH-, phenyl-CHCH-, cyclohexyl-CHCH-, 5-NC-pyrid-3-yl-CHCH-, 4-FC-phenyl-CH-, 3-FC-phenyl-CH-, 2-FC-phenyl-CH-, morpholin-4-yl-CHCHO-phenyl-CH-, 4-aminophenyl-CH-, 4,4-difluorocyclohexyl-CH-, 4-HNCHCHO-phenyl-CH-, 4-HNCHCHO-pyrid-3-yl-CH-, piperidin-4-yl-CH-, or 4-CHC(O)NH-CHCHO-phenyl-CH-; R 10 is H, fluoro, or methyl; R 11 is H, fluoro, or chloro; A 1 and A 2 but both are HO2C-; and X 1 , X 2 , and X 3 is C(H).

[0037] In certain embodiments, the present disclosure provides a method for the preparation of a pharmaceutical composition comprising: R 1 is 4-CH3C(O)-piperazin-1-yl, CH3C(O)NH-CH2CH2-O-, 5-CO2H-pyrimidin-2-yl, or 4-CO2H-phenyl; R 2 is H, ethyl, benzyl, or phenyl-CH2CH2-; R 3 is naphth-1-yl, 4-fluoroindol-3-yl, or 4-chloroindol-3-yl; R 4 But methyl, HO2C-(CH2) m - or H2N(O)C-(CH2) m - and; R 5 But amino, H2N(CH2) n -, naphth-1-yl, indol-3-yl, 7-aza-indol-3-yl, indazol-1-yl, 2,3-dihydroindol-1-yl, pyrid-3-yl, pyrid-4-yl, piperidin-4-yl, 3-aminomethylphenyl, 4-aminomethylphenyl, 3-aminophenyl, 4-aminophenyl, phenyl, pyrid-4-yl-CH-, pyrimidin-5-yl, tetrahydropyran-4-yl-, HN(O)C-(CH)-, 3-biphenyl, or 3-CH=CH-CHO-phenyl; R 6 is, H, (CH3)2CHCH2-, H2N(CH2) p-, HOCH2-, H3CCH2CH2-, morpholin-4-yl-CH2-, tetrahydropyran-4-yl-CH2-, (CH3)2NCH2-, or H3CO-(CH2) q - and; R 7 is H or methyl; R 8a is H, methyl, HOCH2-, or H2N(CH2) r - and; R 8b is H or methyl; R 9a but H; R 9b is H, methyl, 4-HO-phenyl-CH2CH2-, phenyl-CH2CH2-, 5-NC-pyrid-3-yl-CH2CH2-, 4-F3C-phenyl-CH2-, 3-F3C-phenyl-CH2-, 2-F3C-phenyl-CH2-, morpholin-4-yl-CH2CH2O-phenyl-CH2-, 4-aminophenyl-CH2-, 4,4-difluorocyclohexyl-CH2-, 4-H2NCH2CH2O-phenyl-CH2-, 4-H2NCH2CH2O-pyrid-3-yl-CH2-, piperidin-4-yl-CH2-, or 4-CH3C(O)NH-CH2CH2O-phenyl-CH2-; R 10 and R 11 but both are H; and A 1 and A 2 and

[0038] In some embodiments, the disclosure provides a compound of formula (I), wherein the compound is selected from the group consisting of SEQ ID NOs: 1-215 in Table 1. In certain embodiments, the disclosure provides a compound of formula (I), wherein the compound is selected from the group consisting of SEQ ID NOs: 1-173 in Table 1.

[0039] In a specific embodiment, the disclosure provides a compound of formula (I), wherein the compound is selected from the group consisting of (SEQ ID NOs: 78, 71-72, 67, 65, 70, 69, 68, 66, 64, 77, 79, 209, 193, 215, 212, and 210, respectively, in order of appearance).

[0040] [ka] TIFF0007645434000008.tif203150TIFF0007645434000009.tif137149TIFF0007645434000010.tif216150TIFF0007645434000011.tif75135

[0041] In some embodiments, the disclosure provides a compound of Formula (I), wherein the compound is selected from the group consisting of SEQ ID NOs: 78, 71-72, 67, 65, 70, 69, 68, 66, 64, 77, and 79.

[0042] Without being bound by any particular theory, the applicant believes that the compounds disclosed herein capture interleukin-1β and block signaling through IL-1 receptor, thereby reducing downstream markers IL-6 and CRP.Therefore, the compounds may be useful in treating inflammatory components of cardiovascular disease, such as ASCVD and heart failure with preserved ejection fraction (HFpEF).The compounds may also be useful in treating inflammatory disorders, such as hidradenitis suppurativa (acne inversa), inflammatory bowel disease, and osteoarthritis.

[0043] definition Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs.

[0044] As used throughout this disclosure, "disclosed compounds", "compounds of the disclosure" and "compounds disclosed herein" are used interchangeably and are understood to include the disclosed cyclic peptides and compounds of formula (I). Reference to compounds of formula (I) includes compounds of other general formulas within the scope of formula (I), including but not limited to compounds of formula (IA). Compounds of formula (I) can form salts, which are also within the scope of this disclosure. Reference herein to compounds of the disclosure (or compounds of formula (I)) is understood to include reference to salts thereof, unless otherwise indicated. As used herein, the term "salt" refers to acid salts formed with inorganic and / or organic acids, as well as basic salts formed with inorganic and / or organic bases. Furthermore, when compounds of formula (I) contain both a basic moiety, such as but not limited to an amino group, pyrrolidine, imidazole, and an acidic moiety, such as but not limited to a carboxylic acid, zwitterions ("internal salts") may be formed and are included in the term "salt" as used herein. In one embodiment, the salt is a pharma- ceutically acceptable (i.e., non-toxic, physiologically acceptable) salt. In another embodiment, the salt is other than a pharma- ceutically acceptable salt. Salts of compounds of formula (I) can be formed, for example, by reacting a compound of formula (I) with an amount of acid or base (e.g., an equivalent amount) in a medium such as a medium in which the salt precipitates, or in an aqueous medium, followed by lyophilization.

[0045] "Acyl" means an alkyl-C(O)- group, where alkyl is as defined below. The bond to the parent group is through the carbon atom of the carbonyl group.

[0046] "Alkyl" and other groups having the prefix "alk", such as alkoxy, refer to carbon chains which may be linear or branched or combinations thereof, containing the specified number of carbon atoms. For example, C1-C6 alkyl refers to an alkyl group having from 1 (i.e., methyl) up to 6 (i.e., hexyl) carbon atoms. In certain embodiments, linear alkyl groups have from 1 to 6 carbon atoms, and branched alkyl groups have from 3 to 7 carbon atoms. Examples of alkyl groups include methyl, ethyl, propyl, isopropyl, butyl, sec- and tert-butyl, pentyl, hexyl, heptyl, octyl, nonyl, and the like.

[0047] "Alkoxy" and "alkyl-O-" are used interchangeably and refer to an alkyl group linked to oxygen.

[0048] "Amino" refers to an H2N- group. The bond to the parent group is through the nitrogen atom.

[0049] "Amino acid" refers to naturally occurring α-amino acids and their stereoisomers, as well as non-naturally occurring amino acids (such as β-amino acids and substituted amino acids) and their stereoisomers. In the sequences given for the peptides (compounds) according to this disclosure, the amino acid residues have their conventional meanings. Thus, "G" is glycine, "W" is tryptophan, "A" is alanine, "S" is serine, etc. It is to be understood that "D" isomers are designated by the one-letter code or amino acid name preceded by "d", e.g., dA is the D isomer of L-alanine. Amino acid residues not encompassed above have the definitions provided in the Tables in the Examples section below.

[0050] As used herein, "aryl" refers to a monocyclic 6-membered or bicyclic 10-membered ring system in which at least one ring is aromatic and all ring atoms are carbon.

[0051] "Bicyclic ring system" refers to two linked rings which can be fused, i.e., sharing two adjacent atoms, or "spirocyclic", i.e., sharing only one atom.

[0052] "Carboxy" refers to a HO2C- group. The bond to the parent group is through the carbon atom of the carbonyl moiety.

[0053] "Cycloalkyl" refers to a saturated cyclic hydrocarbon group. In certain embodiments, a cycloalkyl group has 3 to 12 carbon atoms, forming 1 to 3 fused carbon rings. Examples of cycloalkyl include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, adamantyl, and the like.

[0054] "Fluoroalkyl" includes mono- and multiply fluoro-substituted alkyl groups, up to and including perfluoro-substituted alkyl, for example, fluoromethyl, 1,1-difluoroethyl, trifluoromethyl, or 1,1,1,2,2-pentafluorobutyl.

[0055] Unless otherwise specified, "halogen" or "halo" includes fluorine (fluoro), chlorine (chloro), bromine (bromo), and iodine (iodo). In one embodiment, halo is fluoro (-F) or chloro (-Cl).

[0056] "Heterocycloalkyl" means a non-aromatic monocyclic, bicyclic, or tricyclic ring system containing about 3 to about 10 ring atoms, preferably about 5 to about 10 ring atoms, in which one or more atoms in the ring system is an element other than carbon, such as nitrogen, oxygen, or sulfur, either alone or in combination. There are no adjacent oxygen and / or sulfur atoms in the ring system. In some embodiments, a heterocycloalkyl contains about 5 to about 6 ring atoms. The prefix aza, oxa, or thia before the heterocyclic root means that at least a nitrogen, oxygen, or sulfur atom, respectively, is present as a ring atom. In some embodiments, the nitrogen or sulfur atom of a heterocycloalkyl may be optionally oxidized to the corresponding N-oxide, S-oxide, or S,S-dioxide. Non-limiting examples of suitable monocyclic heterocycles include piperidyl, pyrrolidinyl, piperazinyl, morpholinyl, thiomorpholinyl, thiazolidinyl, 1,4-dioxanyl, tetrahydrofuranyl, tetrahydropyranyl, tetrahydrothiophenyl, and the like.

[0057] "Heteroaryl" refers to aromatic monocyclic, bicyclic, and tricyclic ring systems in which one or more atoms in the ring (heteroatoms) is an element other than carbon. Heteroatoms are typically O, S, or N atoms. Examples of heteroaromatic groups include pyridinyl, pyrimidinyl, pyrrolyl, pyridazinyl, isoxazolyl, thiazolyl, oxazolyl, indolyl, benzoxazolyl, benzothiazolyl, and imidazolyl.

[0058] Any variable (e.g., R C1 When R C occurs more than one time in any constituent or in formula (I) or other general formulas herein, its definition at each occurrence is independent of its definition at every other occurrence. Combinations of substituents and / or variables are permissible only if such combinations result in stable compounds. In selecting compounds of the present disclosure, those of skill in the art will appreciate that various substituents (e.g., R C 9It will be appreciated that the substituents ) should be selected in accordance with known principles of connectivity and stability of chemical structures. Unless expressly stated to the contrary, substitution with the designated substituents is permitted at any atom in the ring (e.g., an aryl, heteroaryl ring, or saturated heteroaryl ring) provided that such ring substitution is chemically permissible and results in a stable compound. A "stable" compound is one which can be prepared and isolated and whose structure and properties remain essentially unchanged for a period of time sufficient to use the compound for the purposes described herein (e.g., therapeutic or prophylactic administration to a subject).

[0059] The term "substituted" is deemed to include multiple degrees of substitution with a specified substituent. Where multiple substituent moieties are disclosed or claimed, a substituted compound can be singly or multiply independently substituted with one or more of the disclosed or claimed substituent moieties. Independently substituted means that the (two or more) substituents can be the same or different.

[0060] Unless expressly depicted or otherwise explained, variables depicted in a structural formula with a "floating" bond are allowed on any available carbon atom in the ring to which the variable is attached. When a moiety in Formula (I) or any embodiment thereof is described as being "optionally substituted," this is meant to include compounds that contain the specified substituent(s) on that moiety, as well as compounds that do not contain the specified substituent(s) on that moiety.

[0061] Wavy lines as used herein: [ka] indicates the point of attachment to the remainder of the compound.

[0062] Some of the compounds described herein may exist as tautomers, which differ in the point of attachment of hydrogen with one or more double bond shifts. For example, a ketone and its enol form are keto-enol tautomers. Individual tautomers and mixtures thereof are encompassed by the compounds of the present disclosure.

[0063] In the compounds of the present disclosure, atoms may exhibit natural isotopic abundances, or one or more atoms may be artificially enriched with a particular isotope having the same atomic number but an atomic mass or mass number different from the atomic mass or mass number predominantly found in nature. The present disclosure as described and claimed herein is intended to include all suitable isotopic variations of the compounds of the present disclosure and embodiments thereof. For example, different isotopic forms of hydrogen (H) include protium ( 1 H) and deuterium ( 2 H, also denoted herein as D). Protium is the predominant hydrogen isotope found in nature. Enrichment with deuterium may provide certain therapeutic advantages, such as increased in vivo half-life or reduced dosage requirements, or may result in compounds useful as standards for characterization of biological samples. The isotopically enriched compounds of the present disclosure may be prepared without undue experimentation using the appropriate isotopically enriched reagents and / or intermediates by conventional techniques known to those of skill in the art, or by processes analogous to those described in the schemes and examples herein.

[0064] The term "pharmaceutically acceptable salts" refers to salts prepared from pharmaceutically acceptable non-toxic bases or acids. When a compound of the present disclosure is acidic (or has a functional group that can be anionic), its corresponding salt can be conveniently prepared from pharmaceutically acceptable non-toxic bases, including inorganic bases and organic bases. Examples of suitable inorganic cations include Li + , Na + , and K + Alkali metal ions such as Ca 2+ and Mg 2+ Alkaline earth metal cations such as Al 3+ and Zn +Examples of suitable organic cations include, but are not limited to, ammonium ion (i.e., NH + ) and substituted ammonium ions. Examples of suitable substituted ammonium ions include those derived from methylamine, ethylamine, diethylamine, triethylamine and ethylenediamine. When the compound of the present disclosure is basic, its corresponding salt can be conveniently prepared from pharma- ceutically acceptable non-toxic acids, including inorganic and organic acids. Examples of such acid addition salts include salts formed from hydrohalic acids (e.g., hydrochloric acid, hydrobromic acid, hydroiodic acid), formic acid, acetic acid, capric acid, and citric acid. Salts including acetate, formate, capric acid, chloride, or sodium salts are representative for use with the compounds of the present disclosure. In some embodiments, salts of the compounds of the present disclosure can be formed by anion exchange, such as exchanges well known to those skilled in the art, for example, replacing trifluoroacetate with chloride.

[0065] In addition, the compounds of the present disclosure can exist in amorphous form and / or one or more crystalline forms, and therefore all amorphous and crystalline forms of the compounds of formula (I), including the examples, and mixtures thereof, are intended to be included within the scope of the present disclosure. Furthermore, some of the compounds of the present disclosure may form solvates with water (i.e., hydrates) or common organic solvents, such as, but not limited to, acetic acid or acetonitrile. Such solvates and hydrates of the compounds, particularly pharma- ceutically acceptable solvates and hydrates, are also included within the scope of the present disclosure, along with unsolvated and anhydrous forms.

[0066] Pharmaceutically acceptable prodrug modifications of the compounds of the present disclosure that are converted in vivo to compounds within the scope of the present disclosure are also within the scope of the present disclosure.

[0067] The present invention also relates to methods for preparing compounds of formula (I), which are described in the Examples below and which may provide compounds of the present disclosure.

[0068] "Treatment" and "treating" refer to any process that can slow, interrupt, prevent, control, or halt the progression of a disease or disorder as described herein. These terms do not necessarily imply that all symptoms of the disease or disorder are completely eliminated.

[0069] As used herein, "prevent" or "prevention" refers to reducing the likelihood of contracting a disease or disorder described herein or reducing the severity of a disease or disorder described herein.

[0070] The term "therapeutically effective (or potent) amount" and similar descriptions such as "therapeutically effective amount" and "effective dose" refer to an amount of a compound of the present disclosure that elicits the biological or medical response in a tissue, system, animal, or human that is desired by a researcher, veterinarian, physician, or other clinical practitioner. In a preferred embodiment, the term "therapeutically effective amount" refers to an amount of a compound of the present disclosure that reduces at least one clinical symptom in a human patient. The term "prophylactically effective (or potent) amount" and similar descriptions such as "prophylactically effective amount" refer to an amount of a compound of the present disclosure that prevents or reduces the risk of occurrence of a biological or medical event that a researcher, veterinarian, physician, or other clinical practitioner is seeking to prevent in a tissue, system, animal, or human.

[0071] Dosage of Compounds of the Disclosure The dosing regimen using the compounds of the present disclosure is selected according to various factors, including the type, species, age, weight, sex, and condition of the patient; the severity of the condition to be treated; the potency of the compound selected for administration; the route of administration; and the renal and hepatic function of the patient. It is within the competence of a clinical practitioner of ordinary skill to consider these factors in order to determine the therapeutically or prophylactically effective amount required to prevent, inhibit or stop the progression of a condition. It is understood that a particular daily dosage can be both a therapeutically effective amount, for example for treating an oncological condition, and a prophylactically effective amount, for example for preventing an oncological condition.

[0072] While individual needs vary, determining the optimal range of effective amounts of the compounds of the present disclosure is within the skill of the art. For example, a representative dosage of the compounds of the present disclosure for administration to humans in the treatment or prophylactic treatment of the conditions and disorders identified herein can be about 0.05 mg / kg / day to about 50 mg / kg / day. In some embodiments, the patient is administered about 5 mg / day to about 120 mg / day, such as 10 mg / day, 20 mg / day, 30 mg / day, 40 mg / day, 50 mg / day, 60 mg / day, 70 mg / day, 80 mg / day, mg / day, 90 mg / day, or 100 mg / day of the compounds of the present disclosure. In certain embodiments, the patient is administered about 0.2 mg / kg to about 5 mg / kg, such as 0.5 mg / kg, 0.75 mg / kg, 1.0 mg / kg, 1.25 mg / kg, or 1.5 mg / kg of the compounds of the present disclosure. Such doses may be administered in a single dose or may be divided into multiple doses.

[0073] Pharmaceutical Compositions The compounds of the present disclosure and their pharmaceutically acceptable salts can be administered to animals, preferably mammals, particularly humans, as pharmaceuticals, either alone, in a mixture with each other, or in the form of a pharmaceutical composition. The term "subject" or "patient" includes animals, preferably mammals, particularly humans, who use the active agents of the present invention for the prevention or treatment of a medical condition. The administration of the agent to a subject includes both self-administration and administration to the patient by another person. The subject may be a subject who needs or desires treatment of an existing disease or medical condition, or a subject who needs or desires prophylactic treatment to prevent the development of a disease or medical condition or to reduce the risk of development of the disease or medical condition. As used herein, a subject "needs" treatment or prophylactic treatment of an existing condition includes both a medical professional's determination of the need and the patient's desire for such treatment.

[0074] Thus, the present disclosure also provides compounds of the present disclosure and pharma- ceutically acceptable salts thereof for use as pharmaceuticals, their use for modulating the activity of the cytokine IL-1β, in particular in the treatment and prevention of diseases or disorders described below, and their use for preparing medicaments for such purposes. In certain embodiments, the compounds of the present disclosure and pharma-ceutically acceptable salts thereof capture IL-1β.

[0075] The present disclosure further provides pharmaceutical compositions comprising an effective dose of at least one compound of the present disclosure and / or a pharma- ceutically acceptable salt thereof as an active ingredient and a conventional pharma- ceutically acceptable carrier, i.e., one or more pharma- ceutically acceptable carrier substances and / or excipients.

[0076] Thus, the present disclosure provides, for example, a compound of the present disclosure and / or a pharma- ceutically acceptable salt thereof for use as a pharmaceutical composition comprising an effective dose of said compound and / or a pharma- ceutically acceptable salt thereof as an active ingredient and a conventional pharma- ceutically acceptable carrier, as well as the use of said compound and / or a pharma- ceutically acceptable salt thereof in the treatment or prevention of diseases or disorders as described below, such as atherosclerosis, and its use for preparing a medicament for such purposes.

[0077] The pharmaceutical compositions according to the present disclosure may be administered orally, for example in the form of pills, tablets, lacquered tablets, dragees, granules, hard and soft gelatin capsules, aqueous, alcoholic or oily solutions, syrups, emulsions or suspensions, or rectally, for example in the form of suppositories. Administration may also be parenterally, for example subcutaneously, intramuscularly or intravenously in the form of solutions for injection or infusion.

[0078] Other suitable administration forms are, for example, ointments, tinctures, sprays or transdermal therapeutic systems or transdermal or topical administration, for example in the form of microcapsules, implants or rods. The preferred administration form depends, for example, on the disease to be treated and its severity.

[0079] The present disclosure also provides pharmaceutical compositions comprising the compound of formula (I). The compound of formula (I) can be used in combination with any suitable pharmaceutical carrier or excipient. Such pharmaceutical compositions comprise a therapeutically effective amount of one or more compounds of formula (I) and a pharmaceutically acceptable excipient and / or carrier. A particular pharmaceutical composition is adapted to the mode of administration. In certain embodiments, the pharmaceutically acceptable carrier can be water or a buffer.

[0080] The excipients contained in pharmaceutical compositions have different purposes, for example, depending on the nature of the drug and the administration form. Examples of commonly used excipients include saline, buffered saline, dextrose, water for infection, glycerol, ethanol, and combinations thereof, stabilizers, solubilizers and surfactants, buffers and preservatives, isotonicity agents, bulking agents, lubricants (such as talc or silica, and fats such as vegetable stearin, magnesium stearate, or stearic acid), emulsifiers, suspending or viscous agents, inert diluents, fillers (such as cellulose, dibasic calcium phosphate, vegetable fats and oils, lactose, sucrose, glucose, mannitol, sorbitol, calcium carbonate, and magnesium stearate), disintegrants (such as cross-linked polyvinylpyrrolidone, sodium starch glycolate, cross-linked sodium carboxymethylcellulose), binders (starch, gelatin, cellulose, methylcellulose, or modified cellulose). These include, but are not limited to, celluloses, e.g., microcrystalline cellulose, hydroxypropyl cellulose, sugars such as sucrose or lactose, or sugar alcohols such as xylitol, sorbitol or maltitol, polyvinylpyrrolidone, and polyethylene glycols), wetting agents, antimicrobial agents, chelating agents, coating agents (such as cellulose film coating agents, synthetic polymers, shellac, corn protein zein or other polysaccharides, gelatin, etc.), preservatives (including vitamin A, vitamin E, vitamin C, retinyl palmitate, and selenium, cysteine, methionine, citric acid and sodium citrate, and synthetic preservatives such as methylparaben, propylparaben, etc.), sweeteners, fragrances, flavorings, coloring agents, absorption enhancers, administration aids, and combinations thereof.

[0081] Carriers, in the context of pharmaceutical compositions, are compounds and substances that improve and / or extend the delivery of active ingredients to a subject. Carriers may serve to extend the in vivo activity of a drug using sustained release techniques or to delay the release of a drug in a subject. Carriers may also reduce drug metabolism and / or reduce drug toxicity in a subject. Carriers can also be used to deliver drugs to specific cells or tissues in a subject. Common carriers (both hydrophilic and hydrophobic carriers) include lipid emulsions, lipids, PEGylated phospholipids, PEGylated liposomes, PEGylated liposomes coated with cyclic RGD peptides via a PEG spacer, liposomes and lipospheres, microspheres (such as those made of biodegradable polymers or albumin), polymer matrices, biocompatible polymers, protein-DNA complexes, protein complexes, red blood cells, vesicles, nanoparticles, and side chains for carbohydrate staples. The aforementioned carriers can also be used to increase the cell membrane permeability of the compounds of formula (I). In addition to their use in the pharmaceutical compositions of the present disclosure, the carriers may also be used in compositions for other uses, such as in vitro (e.g., for delivery to cultured cells) and / or in vivo research applications.

[0082] Pharmaceutical compositions adapted for oral administration can be provided as discrete units such as capsules or tablets; as powders or granules; as liquids, syrups or suspensions (in aqueous or non-aqueous liquids, or as edible foams or whips, or as emulsions). Suitable excipients for tablets or hard gelatin capsules include lactose, corn starch or derivatives thereof, stearic acid or salts thereof, etc. Suitable excipients for use in soft gelatin capsules include, for example, vegetable oils, waxes, fats, semi-solid or liquid polyols, etc. For the preparation of liquids and syrups, excipients that can be used include water, polyols, and sugars, etc. For the preparation of suspensions, oils, such as vegetable oils, can be used to provide oil-in-water or water-in-oil suspensions. Excipients that promote absorption from the digestive tract can be included, such as penetration enhancers, such as sodium caprate. In certain circumstances, delayed release formulations can be advantageous, and compositions can also be prepared that can deliver the compounds of the present disclosure in a delayed or sustained release manner. Because prolonged residence in the stomach can pose problems of degradation by enzymes present in the stomach, capsules can also be prepared which are enteric coated by standard techniques in the art, so that the active substances are released in the lower part of the digestive tract.

[0083] Pharmaceutical compositions adapted for transdermal administration may be presented as discrete patches intended to remain in intimate contact with the epidermis of the recipient for a prolonged period of time. For example, the active ingredient can be delivered from the patch by iontophoresis, as described in Pharmaceutical Research, 3(6):318 (1986).

[0084] Pharmaceutical compositions adapted for topical administration can be formulated as ointments, creams, suspensions, lotions, powders, solutions, pastes, gels, sprays, aerosols, or oils. When formulated in an ointment, the active ingredient can be used with a paraffinic or water-miscible ointment base. Alternatively, the active ingredient can be formulated in a cream with an oil-in-water cream base or a water-in-oil base. Pharmaceutical compositions adapted for topical administration to the eye include eye drops, in which the active ingredient is dissolved or suspended in a suitable carrier, in particular an aqueous solvent. Pharmaceutical compositions adapted for topical administration in the mouth include lozenges, troches, and mouthwashes.

[0085] Pharmaceutical compositions adapted for rectal administration may be presented as suppositories or enemas.

[0086] Pharmaceutical compositions adapted for nasal administration wherein the carrier is a solid include a coarse powder having a particle size of, for example, 20 to 500 microns, which is administered in the manner of taking a snuff, i.e. by rapid inhalation through the nasal passage from a container of the powder held close to the nose. Suitable compositions wherein the carrier is a liquid, for administration as a nasal spray or drops, include aqueous or oily solutions of the active ingredient.

[0087] Pharmaceutical compositions adapted for administration by inhalation include fine particle dusts or mists, which may be generated by means of various types of metered dose pressurized aerosols, nebulizers or insufflators.

[0088] Pharmaceutical compositions adapted for vaginal administration may be presented as pessaries, tampons, creams, gels, pastes, foams, or spray formulations.

[0089] Pharmaceutical compositions adapted for parenteral administration include aqueous and non-aqueous sterile injection solutions which may contain antioxidants, buffers, bacteriostats and solutes (which render the formulation substantially isotonic with the blood of the intended recipient); and aqueous and non-aqueous sterile suspensions which may contain suspending agents and thickening agents. Excipients which may be used for injection solutions include, for example, water for injection, alcohols, polyols, glycerin and vegetable oils. The compositions may be presented in single-dose or multi-dose containers, for example sealed ampoules and vials, and may be stored in a freeze-dried (lyophilized) condition requiring only the addition of the sterile liquid carrier (e.g., water for injection or saline) immediately prior to use. Extemporaneous injection solutions and suspensions may be prepared from sterile powders, granules and tablets. The pharmaceutical composition may contain preservatives, solubilizers, stabilizers, wetting agents, emulsifiers, sweeteners, colorants, flavoring agents, salts (the substances of the present disclosure may themselves be provided in the form of pharma- ceutically acceptable salts), buffers, coating agents, or antioxidants. It may also contain a therapeutically active agent in addition to the compounds of the present disclosure.

[0090] Methods of Use of the Disclosed Compounds The present application provides a method for inhibiting IL-1 mediated cell signaling comprising contacting a cell with a compound of the present disclosure or a pharma- ceutically acceptable salt thereof. Inhibition of IL-1 mediated cell signaling can be assessed by detecting a decrease in the levels of downstream biomarkers IL-6 and CRP (e.g., hsCRP).

[0091] The present application also provides methods of using the disclosed compounds (or pharma- ceutically acceptable salts thereof) or pharmaceutical compositions comprising such compounds to treat disease conditions, including but not limited to those conditions in which IL-1β is involved.

[0092] In some embodiments, the present disclosure provides a method for treating cardiovascular disease, comprising administering a therapeutically effective amount of a compound of the present disclosure (or a pharma- ceutically acceptable salt thereof) or any of the aforementioned pharmaceutical compositions comprising such a compound to a subject in need of such treatment. In some embodiments, the cardiovascular disease is vascular inflammation. In some embodiments, the cardiovascular disease is atherosclerosis. In some embodiments, the cardiovascular disease is heart failure with preserved ejection fraction (HFpEF). In other embodiments, the cardiovascular disease is heart failure with reduced ejection fraction (HFrEF).

[0093] In some embodiments, the present disclosure provides a method of treating chronic kidney disease, comprising administering to a subject in need of such treatment a therapeutically effective amount of a compound of the present disclosure (or a pharma- ceutically acceptable salt thereof) or any of the aforementioned pharmaceutical compositions comprising such a compound.

[0094] In some embodiments, the present disclosure provides a method of treating an inflammatory disorder, comprising administering a therapeutically effective amount of a compound of the present disclosure (or a pharma- ceutically acceptable salt thereof) or any of the aforementioned pharmaceutical compositions comprising such a compound to a subject in need of such treatment. In certain embodiments, the inflammatory disorder is selected from the group consisting of hidradenitis suppurativa (acne inverse), inflammatory bowel disease, arthritis, and nonalcoholic steatohepatitis (NASH).

[0095] In some embodiments, the inflammatory disorder is hidradenitis suppurativa (acne inversa).

[0096] In certain embodiments, the inflammatory disorder is inflammatory bowel disease, including Crohn's disease and ulcerative bowel disease.

[0097] In some embodiments, the inflammatory disorder is arthritis, for example, osteoarthritis, rheumatoid arthritis, psoriatic arthritis, or gouty arthritis.

[0098] In other embodiments, the inflammatory disorder is non-alcoholic steatohepatitis (NASH).

[0099] Combination therapy One or more additional pharmacologically active agents may be administered in combination with the compound of the present disclosure. By additional active agent(s) is meant an intracellularly active pharmacologic agent(s) that is different from the compound of the present disclosure, such as a prodrug that is converted into a pharmacologic active form after administration, including the free acid, free base, and pharmacologic acceptable salt of said additional active agent. In general, any suitable additional active agent(s) such as, but not limited to, antihypertensive agents, antiatherosclerotic agents such as lipid-modifying compounds, antidiabetic and / or antiobesity agents, anti-inflammatory agents, etc., may be used in any combination with the compound of the present disclosure in a single formulation (fixed dose drug combination), or may be administered to a subject in one or more separate formulations that allow simultaneous or sequential administration of the active agents (co-administration of separate agents).

[0100] Examples of further active agents that can be used in the treatment of cardiovascular disorders include angiotensin-converting enzyme inhibitors (e.g., alacepril, benazepril, captopril, ceronapril, cilazapril, delapril, enalapril, enalaprilat, fosinopril, imidapril, lisinopril, moveltipril, perindopril, quinapril, ramipril, cipirapril, temocapril, or trandolapril), angiotensin II receptor antagonists (e.g., losartan, i.e., C OZAAR®, valsartan (including in combination with sacubitril), candesartan, olmesartan, telmesartan, and any of these agents in combination with hydrochlorothiazide, such as HYZAAR®; sGC activators (e.g., riociguat and vericiguat), PCSK9 inhibitors (e.g., evolocumab, alirocumab, MK-0616, and those disclosed in WO2019 / 246349), neutral endopeptidase inhibitors (e.g., thiorphan and phosphoramidon), aldosterone antagonists, aldosterone synthase inhibitors, renin inhibitors, endothelin receptor antagonists, phosphodiesterase-5 inhibitors (e.g., sildenafil, tadalafil and vardenafil), vasodilators, calcium channel blockers (e.g., amlodipine, nifedipine, verapamil, diltiazem, gallopamil, niludipine, nimodipine, nicardipine), potassium channel activators (e.g., nicorandil, pina sidil, cromakalim, minoxidil, aprikalim, loprazolam), diuretics (e.g., hydrochlorothiazide), sympatholytics, beta-adrenergic blocking agents (e.g., propranolol, atenolol, bisoprolol, carvedilol, metoprolol, or metoprolol tartrate), alpha-adrenergic blocking agents (e.g., doxazosin, prazosin, or alpha-methyldopa), central alpha-adrenergic agonists, peripheral vasodilators (e.g., hydralazine);Lipid-lowering agents, for example HMG-CoA reductase inhibitors such as simvastatin and lovastatin, marketed as ZOCOR® and MEVACOR®, which are in lactone prodrug form and function as inhibitors after administration, and pharma- ceutically acceptable salts of dihydroxy open-acid HMG-CoA reductase inhibitors such as atorvastatin (particularly the calcium salt sold as LIPITOR®), rosuvastatin (particularly the calcium salt sold as CRESTOR®), pravastatin (particularly the sodium salt sold as PRAVACHOL®), fluvastatin (particularly the sodium salt sold as LESCOL®), crivastatin, and pitavastatin; cholesterol absorption inhibitors such as ezetimibe (ZETIA®), and any other lipid-lowering agents such as the HMG-CoA reductase inhibitors mentioned above, in particular simvastatin, rosuvastatin, pravastatin (particularly the sodium salt sold as PRAVACHOL®), fluvastatin (particularly the sodium salt sold as LESCOL®), crivastatin, and pitavastatin; Ezetimibe in combination with statins (VYTORIN®) or atorvastatin calcium; niacin in immediate or sustained release form, and / or with HMG-CoA reductase inhibitors; niacin receptor agonists such as acipimox and acifran, and niacin receptor partial agonists; insulin and insulin mimetics (e.g., insulin degludec, insulin glargine, insulin lispro), dipeptidyl peptidase-IV (DPP-IV), insulin agonists ... 4) inhibitors (e.g., sitagliptin, alogliptin, omarigliptin, linagliptin, vildagliptin); insulin sensitizers, such as (i) PPARy agonists, such as the glitazones (e.g., pioglitazone, mitoglitazone, lobeglitazone, rosiglitazone and balaglitazone), and other PPAR ligands, such as (1) PPARα / γ dual agonists (e.g., tiglitazar, muraglitazar, aleglitazar, soderglitazar and naveglitazar);(2) PPARα agonists, such as fenofibric acid derivatives (e.g., gemfibrozil, clofibrate, ciprofibrate, fenofibrate, bezafibrate), (3) selective PPARγ modulators (SPPARγMs), (e.g., those disclosed in WO02 / 060388, WO02 / 08188, WO2004 / 019869, WO2004 / 020409, WO2004 / 020408, and WO2004 / 066963); and (4) PPARγ partial agonists; (ii) metformin and pharma- ceutically acceptable salts thereof, in particular metformin. biguanides, such as sucralose hydrochloride, and sustained release formulations thereof, such as Grumezza®, Fortamet®, and Glucophage XR®; and (iii) protein tyrosine phosphatase-1B (PTP-1B) inhibitors; insulin or insulin analogs (e.g., insulin detemir, insulin glulisine, insulin degludec, insulin glargine, insulin lispro, and their respective inhaled formulations); leptin and leptin derivatives and agonists; amylin and amylin analogs (e.g., pramlintide); sulfonylureas and non-sulfonylurea derivatives. Urea insulin secretagogues (e.g., tolbutamide, glyburide, glipizide, glimepiride, mitiglinide, meglitinides, nateglinide, and repaglinide); alpha-glucosidase inhibitors (e.g., acarbose, voglibose, and miglitol); glucagon receptor antagonists; incretin mimetics such as GLP-1, GLP-1 analogs, derivatives, and mimetics; and GLP-1 receptor agonists (e.g., dulaglutide, semaglutide, albiglutide, exenatide, liraglutide, lixisenatide, including nasal, transdermal, and once-weekly formulations). , taspoglutide); bile acid sequestrants (e.g. colestilan, colestimide, colesevalam hydrochloride, colestipol, cholestyramine and dialkylaminoalkyl derivatives of cross-linked dextran), acyl-CoA:cholesterol acyltransferase inhibitors (e.g. avasimibe); anti-obesity compounds; drugs intended for use in inflammatory conditions such as aspirin, nonsteroidal anti-inflammatory drugs or NSAIDs, glucocorticoids and selective cyclooxygenase-2 or COX-2 inhibitors; glucokinase activators (GKAs);11β-hydroxysteroid dehydrogenase type 1 inhibitors (e.g., those disclosed in U.S. Patent No. 6,730,690); fructose 1,6-bisphosphatase inhibitors (e.g., those disclosed in U.S. Patent Nos. 6,054,587; 6,110,903; 6,284,748; 6,399,782; and 6,489,476); acetyl-CoA carboxylase-1 or 2 inhibitors (ACC1 or ACC2); AMP-activated protein kinase (AMPK) activators; other G protein-coupled Receptor agonists: (i) GPR-109, (ii) GPR-119, and (iii) GPR-40; SSTR3 antagonists (e.g., those disclosed in WO2009 / 001836); Neuromedin U receptor agonists (e.g., those disclosed in WO2009 / 042053, including but not limited to Neuromedin S (MS)); SCD modulators; GPR-105 antagonists (e.g., those disclosed in WO2009 / 000087); SGLT inhibitors (e.g., empagliflozin, dapagliflozin, canagliflozin, acyl-coenzyme A: diacylglycerol acyltransferase 1 and 2 (DGAT-1 and DGAT-2) inhibitors; fatty acid synthase inhibitors; acyl-coenzyme A: monoacylglycerol acyltransferase 1 and 2 (MGAT-1 and MGAT-2) inhibitors; TGR5 receptor (also known as GPBAR1, BG37, GPCR19, GPR131, and M-BAR) agonists; ileal bile acid transporter inhibitors agents; PACAP, PACAP mimetics, and PACAP receptor 3 agonists; PPAR agonists; protein tyrosine phosphatase-1B (PTP-1B) inhibitors; IL-1β antibodies (e.g., gevokizumab and canakinumab); and metabolic modifiers including bromocriptine mesylate and rapid release formulations thereof, or, where chemically feasible, in combination with other drugs useful in the treatment of said conditions or disorders, including free acid, free base and pharmacologic acceptable salt forms of said active agents;

[0101] Examples of additional active agents that can be used to treat inflammatory disorders include, but are not limited to, steroidal and nonsteroidal anti-inflammatory agents, glucocorticoids, and therapeutic hormones. In certain embodiments, in the treatment of hidradenitis suppurativa (acne inversa), the additional active agent can be an antibiotic, an injectable steroid, a therapeutic hormone, a TNF inhibitor (e.g., infliximab, adalimumab, etanercept, golimumab, certolizumab), an analgesic (e.g., codeine, hydrocodone, morphine, pregabalin, gabapentin, intralesional triamcinolone, a corticosteroid, naproxen, ketoprofen, diclofenac, ibuprofen, acetaminophen). In other embodiments, in the treatment of inflammatory bowel disease, the additional active agent can be methotrexate, a TNF inhibitor, an oral sphingosine 1-phosphate receptor modulator (e.g., fingolimod, siponimod, ozanimod, ponesimod), or a selective JAK inhibitor (e.g., tofacitinib, baricitinib, upadacitinib). In some embodiments, in the treatment of osteoarthritis, the additional active agent can be an analgesic (examples above). In other embodiments, in the treatment of gouty arthritis, the additional active agent can be colcichine, a nonsteroidal anti-inflammatory agent, or a glucocorticoid.

[0102] Methods for making the disclosed compounds The compounds described herein can be prepared according to the procedures in the following schemes and examples using appropriate materials, and are further illustrated by the following specific examples. The examples also include methods for testing such compounds in cellular assays. However, the compounds shown in the examples should not be construed as forming the only genus that is considered to be disclosed.

[0103] The examples further illustrate the details of the preparation of the compounds of the present disclosure. Those skilled in the art will readily recognize that known variations of the conditions and processes of the following preparation procedures can be used to prepare these compounds. For example, in some cases the order of carrying out the steps of the reaction schemes can be altered to facilitate the reaction or to avoid undesired reaction products. Raw materials and intermediates for the final compounds are purchased, prepared by known procedures, or as otherwise described. The examples are provided for the purpose of further illustration only and are not intended to be limitations on the present disclosure.

[0104] NMR data were obtained on 300 MHz or 400 MHz instruments in CDCl3, DMSO-d6 or methanol-d4 and chemical shifts are reported relative to a tetramethylsilane standard. Resonance signals are reported with the following abbreviations: s = singlet, d = doublet, t = triplet, q = quartet, dd = doublet of doublets, m = multiplet or overlap of unequal resonances. Coupling constants (J) are reported in Hertz (Hz).

[0105] Throughout the synthetic schemes and examples, unless otherwise stated, abbreviations and acronyms may be used with the following meanings:

[0106] Abbreviation [Table 1] TIFF0007645434000014.tif255148TIFF0007645434000015.tif255149TIFF0007645434000016.tif255150TIFF00076454340 00017.tif255148TIFF0007645434000018.tif255148TIFF0007645434000019.tif255149TIFF0007645434000020.tif238153

[0107] Intermediate synthesis: Synthesis scheme 1 [ka] (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(1H-indazol-1-yl)propanoic acid Step 1: A 2-liter four-necked round-bottom flask, purged with and maintained under an inert atmosphere of nitrogen, was charged with DMSO (150 mL), 1H-indazole (15 g, 127 mmol), (S)-2-amino-3-hydroxypropanoic acid (14.01 g, 133 mmol), and pH 8 0.2 M potassium phosphate buffer (600 mL, 127 mmol) at 65° C. [(4-formyl-5-hydroxy-6-methylpyridin-3-yl)methoxy]phosphonic acid (PLP) (96 mg, 0.00284 equiv.) in HO (2 mL) and PfTrpB-0B2 (1.05 g, 10 wt %) in 0.2 M potassium phosphate buffer (2 mL) were added to the solution. The resulting solution was stirred at 65° C. for 16 h. The progress of the reaction was monitored by HPLC-MS.

[0108] Step 2: To the resulting solution was added Na2CO3 (21.55 g, 2.0 equiv.) and Fmoc-OSu (37.68 g, 1.1 equiv.) in THF (150 mL) at 0° C. The solution was stirred at 25° C. for 2 h. The progress of the reaction was monitored by HPLC-MS. The solution was adjusted to pH 3 with hydrogen chloride (6 M). The solution was extracted with EtOAc (3×500 mL). The organic layers were combined and washed with H2O (2×1 L) and NaCl (1×1 L). The organic layer was dried over anhydrous sodium sulfate and concentrated under reduced pressure. The crude product was slurried with DCM (15 vol.) and the solid was collected by filtration. HPLC-MS: (ESI, m / z): 428 [M+H] + . 1H NMR: (400 MHz, DMSO-d6, ppm): δ 13.08 (s, 1H), 8.10 (d, J = 1.0 Hz, 1H), 7.87 (dt, J = 7.7, 1.0 Hz, 2H), 7.81 - 7.71 (m, 2H), 7.71 - 7.65 (m, 1H), 7.58 (d, J = 7.5 Hz, 1H), 7.54 (d, J = 7.5 Hz, 1H), 7.41 (dtd, J = 6.0, 4.4, 4.3, 3.2 Hz, 2H), 7.40 - 7.33 (m, 1H), 7.36 - 7.24 (m, 1H), 7.29 - 7.20 (m, 1H), 7.16 - 7.08 (m, 1H), 4.83 - 4.65 (m, 2H), 4.61 - 4.51 (m, 1H), 4.22 - 4.06 (m, 3H).

[0109] Synthesis scheme 2 [ka] TIFF0007645434000023.tif29133 (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(indolin-1-yl)propanoic acid Step 1: In a 10 L three-necked round bottom flask was added indoline (125 g, 1.05 mol, 1.00 equiv), DMSO (625 mL), pH 8 0.2 M potassium phosphate buffer (5625 mL) and PLP (778.32 mg, 2.94 mmol), L-serine (115.75 g, 1.10 mol, 1.05 equiv) and PfTrpB-4D11 (12.50 g) at 25° C. The resulting solution was stirred at 65° C. under nitrogen for 4 h. The progress of the reaction was monitored by HPLC-MS.

[0110] Step 2: To the above solution was added Na2CO3 (129 g, 2.00 equiv), Fmoc-OSu (306 g, 1.50 equiv) and THF (2.5 L) at 0°C. The reaction mixture was stirred under nitrogen at 25°C for 3 h. The solution was acidified with 6M HCl to pH 3. The mother liquor was collected by filtration and extracted with EtOAc (3 x 1 L). The organic layers were combined, washed with H2O (2 x 800 mL) and NaCl (1 x 800 mL), dried over Na2SO4 and concentrated under reduced pressure. The residue was purified by silica gel chromatography eluting with PE / EtOAc (3:1) to give the crude product. The crude product was then purified by preparative HPLC reverse phase (C18) eluting with acetonitrile / water + 0.05% TFA (50% to 70% gradient in 30 min). The mixture was concentrated under reduced pressure at 20° C. to remove MeCN, extracted with EtOAc (3×1 L), dried over Na2SO4, and concentrated under reduced pressure. The pure product was slurried in hexane (1 L) for 30 min. The solid was collected by filtration. This gave (2S)-3-(2,3-dihydroindol-1-yl)-2-[[(9H-fluoren-9-ylmethoxy)carbonyl]amino]propanoic acid. HPLC-MS: (ESI, m / z): 429 [M+H] + . 1 H NMR: (400 MHz, DMSO-d6, ppm) δ 7.89 (d, J = 7.5 Hz, 2H), 7.74 - 7.67 (m, 2H), 7.41 (t, J = 7.5 Hz, 2H), 7.35 - 7.25 (m, 2H), 7.00 (ddd, J = 17.4, 7.5, 1.3 Hz, 2H), 6.61 - 6.48 (m, 2H), 4.35 - 4.18 (m, 4H), 3.47 - 3.28 (m, 4H), 2.87 (t, J = 8.2 Hz, 2H).

[0111] Synthesis scheme 3 [ka] (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(6-(4-acetylpiperazin-1-yl)pyridin-3-yl)propanoic acid Step 1: To a mixture of 1-(piperazin-1-yl)ethan-1-one (21.85 g, 170 mmol) in DMF (150 mL) was added 5-bromo-2-fluoropyridine (15 g, 85 mmol) at room temperature under argon. The reaction was stirred at 100° C. for 2 h, diluted with 300 mL of EtOAc, washed with H2O (3×80 mL), saturated aqueous NaCl (80 mL), dried over Na2SO4, and filtered. The filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography eluting with a gradient of 0% to 100% EtOAc / PE. Fractions containing the desired product were combined and concentrated under reduced pressure to give 1-(4-(5-bromopyridin-2-yl)piperazin-1-yl)ethan-1-one. MS ESI:C 11 H 15 BrNO[M+H] + Calculated values: 284.03 and 286.03, measured values: 283.90 and 285.90.

[0112] Step 2: A mixture of nickel(II) chloride ethylene glycol dimethyl ether complex (0.696 g, 3.17 mmol) and 1,10-phenanthroline (0.571 g, 3.17 mmol) in DMA (2 mL) was heated at 50° C. for 0.5 h. A mixture of 1-(4-(5-bromopyridin-2-yl)piperazin-1-yl)ethan-1-one (4.5 g, 15.84 mmol), tert-butyl (R)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-bromopropanoate (7.78 g, 17.42 mmol) and tetrabutylammonium iodide (5.85 g, 15.84 mmol) in DMA (2 mL) was added at 25° C. Then zinc powder (2.071 g, 31.7 mmol) was added. The resulting mixture was stirred at 50° C. for 1 h. The resulting mixture was poured into water (300 mL) and extracted with EtOAc (3×300 mL). The organic layer was washed with water (100 mL) and brine (2×80 mL), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The residue was purified by column chromatography on silica gel eluting with DCM-MeOH (10:1) to give tert-butyl (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(6-(4-acetylpiperazin-1-yl)pyridin-3-yl)propanoate. MS ESI:C 33 H 39 N4O5[M+H] + Calculated value: 571.28, actual value: 571.40.

[0113] Step 3: To a mixture of tert-butyl (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(6-(4-acetylpiperazin-1-yl)pyridin-3-yl)propanoate (7.3 g, 12.79 mmol) in DCM (70 mL) at room temperature under argon was added TFA (70 mL, 909 mmol). The reaction was stirred at room temperature for 1 h and concentrated under reduced pressure. The residue was dissolved in THF (20 mL) and the resulting mixture was purified by reversed-phase Combi-Flash with the following conditions: column C18 silica gel column (330 g), 20-35 μm; mobile phase A: 5 mM TFA in water; mobile phase B: MeCN; (gradient: 0% B hold 10 min, to 42.3% B in 35 min, to 42.3% B hold 3.2 min; to 95% B in 2 min, to 95% B hold 10 min); flow rate: 60 mL / min; detector: UV254 nm; RT: 35.32 min. Product-containing fractions were collected and concentrated under reduced pressure to give (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(6-(4-acetylpiperazin-1-yl)pyridin-3-yl)propanoic acid. MS ESI: C 29 H 31 N4O5[M+H] + Calculated value: 515.22, actual value: 515.15. 1 H NMR (300 MHz, methanol-d4) δ 7.87 - 7.79 (m, 4H), 7.62 - 7.55 (m, 2H), 7.42 - 7.27 (m, 4H), 7.11 - 7.09 (m, 1H), 4.51 - 4.46 (m, 1H), 4.32 - 4.09 (m, 3H), 3.65 - 3.54 (m, 8H), 3.29 - 3.20 (m, 1H), 2.94 - 2.89 (m, 1H), 2.12 (s, 3H).

[0114] Synthesis scheme 4 [ka] TIFF0007645434000026.tif29133 (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(6-(2-acetamidoethoxy)pyridin-3-yl)propanoic acid Step 1: To a mixture of N-(2-hydroxyethyl)acetamide (17.58 g, 170 mmol) in t-BuOH (150 mL) at room temperature under argon was added 5-bromo-2-fluoropyridine (15 g, 85 mmol) followed by potassium tert-butoxide (19.13 g, 170 mmol). The reaction was stirred at room temperature for 1 h and concentrated under reduced pressure. The residue was diluted with 500 mL of EtOAc, washed with saturated aqueous NaHCO3 (3 x 250 mL), saturated aqueous NaCl (250 mL), dried over Na2SO4, and filtered. The filtrate was concentrated under reduced pressure to give crude N-(2-((5-bromopyridin-2-yl)oxy)ethyl)acetamide. MS ESI: CH 12 BrN2O2[M+H] + Calculated values: 259.00 and 261.00, actual values: 258.90 and 261.90.

[0115] Step 2: A mixture of nickel(II) chloride ethylene glycol dimethyl ether complex (1.187 g, 5.40 mmol) and 1,10-phenanthroline (0.974 g, 5.40 mmol) in DMA (70 mL) was heated at 50° C. for 0.5 h. A mixture of N-(2-((5-bromopyridin-2-yl)oxy)ethyl)acetamide (7 g, 27.0 mmol), tert-butyl (R)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-bromopropanoate (13.26 g, 29.7 mmol) and tetrabutylammonium iodide (9.98 g, 27.0 mmol) in DMA (70 mL) was added at 25° C. Then, zinc powder (3.53 g, 54.0 mmol) was added and the resulting mixture was stirred at 50° C. for 1 h. The reaction mixture was diluted with 300 mL of EtOAc, washed with saturated aqueous NaHCO3 (3 x 80 mL), brine (80 mL), dried over Na2SO4, and filtered. The filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography eluting with a gradient of 0% to 20% DCM / MeOH. Fractions containing the desired product were combined and concentrated under reduced pressure to give tert-butyl (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(6-(2-acetamidoethoxy)pyridin-3-yl)propanoate. MS ESI:C 31 H 36 N3O6[M+H] + Calculated value: 546.25, actual value: 546.40.

[0116] Step 3: To a mixture of tert-butyl (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(6-(2-acetamidoethoxy)pyridin-3-yl)propanoate (8 g, 14.66 mmol) in DCM (80 mL) at room temperature under argon was added TFA (80 mL, 1038 mmol). The reaction was stirred at room temperature for 1 h and concentrated under reduced pressure. The residue was purified by RP flash column chromatography under the following conditions: Column: C18 silica gel column (330 g), 20-35 μm; Mobile phase A: 5 mM NH4CO3 in water; Mobile phase B: MeCN; (Gradient: 0% B hold 5 min, to 45% B in 20 min, hold 45% B for 10 min; to 95% B in 15 min, hold 95% B for 10 min); Flow rate: 60 mL / min; Detector: UV254×210 nm; RT: 35.32 min. Product-containing fractions were collected and concentrated under reduced pressure to give (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(6-(2-acetamidoethoxy)pyridin-3-yl)propanoic acid. MS ESI: C 27 H 28 N3O6[M+H] + Calculated value: 490.19, actual value: 490.10. 1 H NMR (400 MHz, methanol-d4) δ 8.02 - 8.00 (m, 1H), 7.79 - 7.77 (m, 2H), 7.67 - 7.64 (m, 1H), 7.61 - 7.57 (m, 2H), 7.40 - 7.36 (m, 2H), 7.31 - 7.27 (m, 2H), 6.82 - 6.80 (m, 1H), 4.42 - 4.39 (m, 1H), 4.33 - 4.22 (m, 4H), 4.16 - 4.13 (m, 1H), 3.53 - 3.51 (m, 2H), 3.20 - 3.15 (m, 1H), 2.93 - 2.87 (m, 1H), 1.92 (s, 3H).

[0117] Synthesis scheme 5 [ka] (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(6-(4-(tert-butoxycarbonyl)phenyl)pyridin-3-yl)propanoic acid Step 1: To a stirred solution of NiCl2-glyme (0.918 g, 4.18 mmol) in DMA (100 mL) was added 1,10-phenanthroline (0.905 g, 4.18 mmol) at room temperature under nitrogen atmosphere. The resulting solution was stirred at 50° C. for 1 h. 2-Chloro-5-iodopyridine (5 g, 20.88 mmol), tert-butyl (R)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-iodopropanoate (12.36 g, 25.06 mmol), TBAI (8.01 g, 20.88 mmol) and Zn (2.73 g, 41.8 mmol) were added to the above mixture at room temperature and the resulting mixture was stirred at 25° C. for 2 h. The reaction was quenched with H2O (200 mL) and extracted with EtOAc (2×500 mL). The combined organic layers were washed with brine (3 x 200 mL), dried over anhydrous Na2SO4, and filtered. The filtrate was concentrated under reduced pressure, and the residue was purified by silica gel chromatography eluting with 0-30% EtOAc / PE to give tert-butyl (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(6-chloropyridin-3-yl)propanoate. MS ESI:C 27 H 28 ClN2O4[M+H] + Calculated value: 479.17, actual value: 479.20.

[0118] Step 2: To a stirred solution of tert-butyl (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(6-chloropyridin-3-yl)propanoate (5 g, 10.48 mmol) in DCM (5 mL) was added TFA (10 mL) at room temperature. The solution was stirred at 25° C. for 1 h. The solvent was concentrated under reduced pressure and the residue was purified by RP-flash under the following conditions: column: flash C18 (330 g); mobile phase A: water (0.1% TFA), mobile phase B: MeCN; (gradient: hold 5% B for 5 min, up to 30% B in 15 min, hold 30% B for 5 min; up to 95% B in 20 min, hold 95% B for 10 min); flow rate: 90 mL / min; detector: UV 210 nm; RT=40 min. The product-containing fractions were collected and evaporated under reduced pressure to give (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(6-chloropyridin-3-yl)propanoic acid. MS ESI:C 23 H 20 ClN2O4[M+H] + Calculated value: 423.10, actual value: 423.10.

[0119] Step 3: To a stirred solution of (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(6-chloropyridin-3-yl)propanoic acid (3 g, 7.09 mmol) in THF (25 mL) and water (5 mL) at 25 °C under nitrogen was added (4-(tert-butoxycarbonyl)phenyl)boronic acid (1.890 g, 8.51 mmol) and K3PO4 (7.53 g, 35.5 mmol). The resulting solution was stirred at 25 °C for 10 min. Pd(dtbpf)Cl2 (0.694 g, 1.064 mmol) was added to the solution and the mixture was stirred at 60 °C for 16 h. The reaction was cooled to room temperature, quenched with H2O (200 mL) and extracted with EtOAc (2 x 500 mL). The combined organic layers were washed with brine (3 x 200 mL), dried over anhydrous Na2SO4, and filtered. The filtrate was concentrated under reduced pressure and the residue was purified by RP-flash under the following conditions: Column: Flash C18 (330 g); Mobile phase A: Water (0.1% TFA), Mobile phase B: MeCN; (Gradient: 5% B hold 5 min, to 60% B in 15 min, to 60% B hold 15 min; to 95% B in 10 min, to 95% B hold 10 min); Flow rate: 90 mL / min; Detector: UV 210 nm; RT = 55 min. Product-containing fractions were collected and evaporated under reduced pressure to give (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(6-(4-(tert-butoxycarbonyl)phenyl)pyridin-3-yl)propanoic acid. MS ESI: C 34 H 33 N2O6[M+H] + Calculated value: 565.23, Observed value: 565.15; 1H NMR (400 MHz, methanol-d4) δ 8.66 (d, J = 1.9 Hz, 1H), 8.12 - 8.07 (m, 3H), 7.99 - 7.92 (m, 3H), 7.77 (d, J = 7.5 Hz, 2H), 7.59 - 7.56 (m, 2H), 7.37 - 7.33 (m, 2H), 7.30 - 7.22 (m, 2H), 4.60 - 4.56 (m, 1H), 4.29 - 4.27 (m, 2H), 4.14 - 4.10 (m, 1H), 3.45 - 3.41 (m, 1H), 3.14 - 3.10 (m, 1H), 1.62 (s, 9H).

[0120] Synthesis scheme 6 [ka] (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-4-morpholinobutanoic acid Step 1: To a stirred mixture of ((benzyloxy)carbonyl)-L-homoserine (2.5 g, 9.87 mmol) and NaHCO3 (2.488 g, 29.6 mmol) in DMF (50 mL) was added benzyl bromide (2.026 g, 11.85 mmol) under nitrogen atmosphere at 0 °C. The mixture was stirred at 25 °C for 16 h, quenched with water (200 mL) and extracted with EtOAc (3 x 200 mL). The combined organic layers were washed with brine (3 x 100 mL) and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure and the residue was purified by silica gel column chromatography eluting with 0-45% EtOAc / PE to give benzyl ((benzyloxy)carbonyl)-L-homoserinate. MS ESI:C 19 H 21 NO5Na[M+Na] + Calculated value: 366.14, actual value: 366.15.

[0121] Step 2: In a two-necked 500 mL flask under N2 atmosphere, I2 (22.17 g, 87 mmol) was added to a solution of PPh3 (17.19 g, 65.5 mmol) and 1H-imidazole (4.46 g, 65.5 mmol) in dry CHCl2 (100 mL) at 0 °C. The reaction was warmed to 25 °C and stirred for 15 min. A solution of benzyl ((benzyloxy)carbonyl)-L-homoserinate (15 g, 43.7 mmol) in CHCl2 (100 mL) was added over 10 min and stirred at room temperature for 2 h. The mixture was washed with saturated Na2S2O3 solution (3 x 200 mL) and then with brine (3 x 50 mL). The organic layer was dried over Na2SO4, the filtrate was concentrated under reduced pressure, and the residue was purified by silica gel chromatography eluting with 0-40% EtOAc / PE to give benzyl (S)-2-(((benzyloxy)carbonyl)amino)-4-iodobutanoate. MS ESI:C 19 H 21 INO4[M+H] + Calculated value: 454.04, actual value: 454.05. 1 H NMR (400 MHz, CDCl3) δ 7.45 - 7.31 (m, 10H), 5.37 - 5.35 (m, 1H), 5.19 - 5.15 (m, 2H), 5.11 (s, 2H), 4.48 - 4.41 (m, 1H), 3.15 - 3.10 (m, 2H), 2.46 - 2.44 (m, 1H), 2.25 - 2.20 (m, 1H).

[0122] Step 3: To a stirred mixture of benzyl (S)-2-(((benzyloxy)carbonyl)amino)-4-iodobutanoate (13.6 g, 30.0 mmol) in THF (140 mL) was added morpholine (20.91 g, 240 mmol) at room temperature. The resulting mixture was stirred at 50° C. for 16 h. The reaction mixture was concentrated under reduced pressure and the residue was purified by silica gel chromatography eluting with 0-80% EtOAc / PE, the product-containing fractions were collected and concentrated under reduced pressure to give benzyl (S)-2-(((benzyloxy)carbonyl)amino)-4-morpholinobutanoate. MS ESI:C23 H 29 N2O5[M+H] + Calculated value: 413.21, actual value: 413.35. 1 H NMR (400 MHz, CDCl3) δ 7.36 - 7.34 (m, 10H), 6.67 (s, 1H), 5.24 - 5.02 (m, 4H), 4.52 - 4.47 (m, 1H), 3.67 - 3.52 (m, 4H), 2.38 - 2.51 (m, 6H), 2.15 - 1.81 (m, 2H).

[0123] Step 4: Benzyl (S)-2-(((benzyloxy)carbonyl)amino)-4-morpholinobutanoate (6.5 g, 15.76 mmol) was dissolved in EtOH (65 mL) and the resulting mixture was evacuated and covered with nitrogen at ambient temperature. Pd-C (2.6 g, dry, 20 wt%) was then added under nitrogen. The suspension was degassed under reduced pressure and purged with H2 several times and the reaction solution was stirred at 25° C. under 1 atm H2 for 5 h. The suspension was filtered and the filtrate was concentrated under reduced pressure to give crude (S)-2-amino-4-morpholinobutanoic acid. MS ESI: CH 17 N2O3[M+H] + Calculated value: 189.12, actual value: 189.25.

[0124] Step 5: To a solution of (S)-2-amino-4-morpholinobutanoic acid (2.97 g, 15.76 mmol) in THF (25 mL) and water (25 mL) at 25 °C, NaHCO3 (6.63 g, 79 mmol) and Fmoc-OSu (4.85 g, 14.20 mmol) were added. The reaction mixture was stirred at 25 °C for 16 h. Upon completion, the pH value of the solution was adjusted to 1M. The pH was adjusted to 3 with HCl. The mixture was purified by reversed-phase flash chromatography under the following conditions: C18 column (120 g); mobile phase A: water (0.05% TFA), mobile phase B: MeCN; (gradient: hold 5% B for 5 min, to 38% B in 30 min, hold 38% B for 2.6 min; to 95% B in 2 min, hold 95% B for 5 min); flow rate: 60 mL / min; detector: UV 210 nm; RT = 31 min. Product-containing fractions were collected and concentrated under reduced pressure to give (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-4-morpholinobutanoic acid. MS ESI: C 23 H 27 N2O5[MH] - Calculated value: 411.19, actual value: 411.30. 1 H NMR (300 MHz, methanol-d4) δ 7.81 - 7.79 (m, 2H), 7.68 - 7.65 (m, 2H), 7.42 - 7.37 (m, 2H), 7.33 - 7.29 (m, 2H), 4.48 - 4.45 (m, 2H), 4.42 - 4.23 (m, 2H), 4.01 - 3.77 (m, 4H), 3.33 - 3.30 (m, 2H), 3.29 - 3.14 (m, 4H), 2.45 - 2.05 (m, 2H).

[0125] Synthesis scheme 7 [ka] (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(4'-(tert-butoxycarbonyl)-[1,1'-biphenyl]-4-yl)propanoic acid A mixture of (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(4-bromophenyl)propanoic acid (6 g, 12.87 mmol), (4-(tert-butoxycarbonyl)phenyl)boronic acid (4.29 g, 19.30 mmol) and K3PO4 (8.19 g, 38.6 mmol) in THF (40 mL) was sparged with argon gas for 10 min and [1,1'-bis(di-tert-butylphosphino)ferrocene]dichloropalladium(II) (0.839 g, 1.287 mmol) was added. The resulting mixture was stirred at 50 °C for 16 h, after which it was diluted with EtOAc (300 mL), washed with saturated aqueous NaHCO3 (3 x 80 mL), brine (2 x 40 mL), dried over Na2SO4 and filtered. The filtrate was concentrated under reduced pressure. The residue was purified by silica gel chromatography eluting with a gradient of 0% to 50% EtOAc / PE. The product-containing fractions were collected and rotary evaporated under reduced pressure. The residue was purified again by combi-flash under the following conditions: Column: C18 gel column (330 g), 20-35 μm; Mobile phase A: 0.5% TFA in water; Mobile phase B: MeCN; (Gradient: 0% B hold 10 min, to 62.3% B in 25 min, hold 62.3% B for 6.2 min; to 95% B in 2 min, hold 95% B for 10 min); Flow rate: 90 mL / min; Detector: UV254×; RT: 32.32 min. The product-containing fractions were collected and concentrated under reduced pressure to give (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(4'-(tert-butoxycarbonyl)-[1,1'-biphenyl]-4-yl)propanoic acid. MS ESI:C 35 H 34 NO6[M+1] + Calculated value: 564.23, actual value: 564.15. 1H NMR (300 MHz, methanol-d4) δ 7.97 - 7.95 (m, 2H), 7.78 - 7.76 (m, 2H), 7.61 - 7.53 (m, 6H), 7.38 - 7.21 (m, 2H), 4.51 - 4.11 (m, 4H), 3.32 - 3.25 (m, 1H), 3.03 - 2.95 (m, 1H), 1.61 (s, 9H).

[0126] Synthesis scheme 8 [ka] (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-4-(5-cyanopyridin-3-yl)butanoic acid Step 1: To a stirred solution of (((9H-fluoren-9-yl)methoxy)carbonyl)-L-homoserine (15 g, 43.9 mmol) in DCM (200 mL) at room temperature was added tert-butyl (Z)-N,N'-diisopropylcarbamimidate (35.2 g, 176 mmol). The solution was stirred at 30°C for 4 h. The solids were filtered off and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography eluting with 0-32% EtOAc / PE to give tert-butyl (((9H-fluoren-9-yl)methoxy)carbonyl)-L-homoserinate. MS ESI:C 23 H 28 NO5[M+H] + Calculated value: 398.19, actual value: 398.15. 1 H NMR (300 MHz, CDCl3) δ 7.81 - 7.74 (m, 2H), 7.64 - 7.56 (m, 2H), 7.45 - 7.37 (m, 2H), 7.37 - 7.28 (m, 2H), 5.61 (d, J = 7.7 Hz, 1H), 4.54 - 4.32 (m, 3H), 4.22 (t, J = 6.9 Hz, 1H), 3.76 - 3.52 (m, 2H), 2.25 - 2.07 (m, 1H), 1.71 - 1.55 (m, 1H), 1.48 (s, 9H).

[0127] Step 2: To a stirred solution of PPh3 (12.67 g, 48.3 mmol) and 1H-imidazole (4.38 g, 64.4 mmol) in DCM (200 mL) was added I2 (12.26 g, 48.3 mmol) at room temperature under nitrogen atmosphere. The mixture was stirred at room temperature for 10 min. tert-Butyl (((9H-fluoren-9-yl)methoxy)carbonyl)-L-homoserinate (12.8 g, 32.2 mmol) was added to the mixture and stirred at 25 °C for 2 h. The solvent was concentrated under reduced pressure and the residue was purified by silica gel chromatography eluting with 0-18% EtOAc / PE to give tert-butyl (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-4-iodobutanoate. MS ESI: C 23 H 26 INO4Na[M+Na] + Calculated value: 530.09, actual value: 530.20. 1 H NMR (300 MHz, CDCl3) δ 7.83 - 7.71 (m, 2H), 7.65 - 7.56 (m, 2H), 7.47 - 7.36 (m, 2H), 7.36 - 7.28 (m, 2H), 5.35 (d, J = 8.3 Hz, 1H), 4.52 - 4.35 (m, 2H), 4.34 - 4.16 (m, 2H), 3.21 - 3.04 (m, 2H), 2.52 - 2.32 (m, 1H), 2.28 - 2.07 (m, 1H), 1.48 (s, 9H).

[0128] Step 3: To a stirred solution of 1,10-phenanthroline (1.083 g, 6.01 mmol) in DMA (300 mL) at room temperature under argon atmosphere was added NiCl2-glyme (1.321 g, 6.01 mmol). The mixture was stirred at 50° C. for 1 h and then cooled to room temperature. Then, tert-butyl (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-4-iodobutanoate (15.25 g, 30.1 mmol), 5-bromonicotinonitrile (5.5 g, 30.1 mmol), TBAI (15.25 g, 30.1 mmol) and zinc (3.93 g, 60.1 mmol) were added to the mixture at room temperature and stirred at 35° C. for 2.5 h. The mixture was cooled to room temperature and diluted with water (500 mL) and EtOAc (800 mL). The solids were filtered off and the organic layer was separated. The organic layer was washed with brine (3 x 150 mL), dried over anhydrous Na2SO4, and filtered. The filtrate was concentrated under reduced pressure and the residue was purified by silica gel column chromatography eluting with 0-40% EtOAc / PE to give tert-butyl (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-4-(5-cyanopyridin-3-yl)butanoate. MS ESI:C 29 H 30 N3O4[M+H] + Calculated value: 484.22, actual value: 484.20. 1 H NMR (300 MHz, CDCl3) δ 8.75 (s, 1H), 8.66 (s, 1H), 7.87 - 7.71 (m, 3H), 7.61 (d, J = 7.4 Hz, 2H), 7.47 - 7.28 (m, 4H), 5.40 (d, J = 7.8 Hz, 1H), 4.55 - 4.37 (m, 2H), 4.36 - 4.18 (m, 2H), 2.85 - 2.59 (m, 2H), 2.26 - 2.08 (m, 1H), 2.02 - 1.82 (m, 1H), 1.49 (s, 9H).

[0129] Step 4: To a stirred solution of tert-butyl (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-4-(5-cyanopyridin-3-yl)butanoate (7.5 g, 15.51 mmol) in DCM (50 mL) was added TFA (100 mL) at room temperature. The solution was stirred at 25° C. for 3 h and then concentrated under reduced pressure. The residue was purified by RP-flash with the following conditions: C18 column, 330 g, 5%-5% 10 min, 5% to 55% 40 min, 55%-55% 10 min, MeCN / water (0.05% TFA) to give (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-4-(5-cyanopyridin-3-yl)butanoic acid. MS ESI: C 25 H 22 N3O4[M+H] + Calculated value: 428.15, actual value: 428.05. 1 H NMR (400 MHz, DMSO-d6) δ 12.78 (br, 1H), 8.87 (s, 1H), 8.75 (s, 1H), 8.18 (s, 1H), 7.92 - 7.90 (m, 2H), 7.76 - 7.73 (m, 3H), 7.44 - 7.33 (m, 4H), 4.39 - 4.20 (m, 3H), 3.94 - 3.89 (m, 1H), 2.79 - 2.67 (m, 2H), 2.09 - 1.89 (m, 2H).

[0130] Synthesis scheme 9 [ka] (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(4-(4-acetylpiperazin-1-yl)phenyl)propanoic acid Step 1: To a stirred solution of (S)-3-(4-bromophenyl)-2-((tert-butoxycarbonyl)amino)propanoic acid (6 g, 17.43 mmol) in toluene (180 mL) at 25° C. under nitrogen was added XPhos Pd G2 (2.057 g, 2.61 mmol). The resulting solution was stirred at 100° C. for 10 min. 1-(piperazin-1-yl)ethan-1-one (2.234 g, 17.43 mmol) and Cs2CO3 (5.04 g, 26.1 mmol) were added and the resulting solution was stirred at 110° C. for 2 h. The reaction was cooled to room temperature, quenched with H2O (500 mL), and extracted with EtOAc (2×500 mL). The combined organic layers were washed with brine (3×200 mL), dried over anhydrous Na2SO4, and filtered. The filtrate was concentrated under reduced pressure and the residue was purified by silica gel chromatography eluting with 0-60% EtOAc / PE to give (S)-3-(4-(4-acetylpiperazin-1-yl)phenyl)-2-((tert-butoxycarbonyl)amino)propanoic acid. MS ESI:C 20 H 30 N3O5[M+H] + Calculated value: 392.21, actual value: 392.25. 1 H NMR (400 MHz, methanol-d4) δ 7.12 (d, J = 8 Hz, 2H), 6.89 (d, J = 8 Hz, 2H), 4.16 - 4.13 (m, 1H), 3.72 - 3.65 (m, 4H), 3.14 - 3.04 (m, 4H), 2.93 - 2.81 (m, 2H), 2.13 (s, 3H), 1.38 - 1.29 (m, 9H).

[0131] Step 2: To a stirred solution of (S)-3-(4-(4-acetylpiperazin-1-yl)phenyl)-2-((tert-butoxycarbonyl)amino)propanoic acid (10 g, 25.5 mmol) in DCM (30 mL) was added TFA (30 mL) at room temperature. The solution was stirred at 25° C. for 1 h and concentrated under reduced pressure. The crude (S)-3-(4-(4-acetylpiperazin-1-yl)phenyl)-2-aminopropanoic acid was used directly in the next step without further purification. MS ESI:C15 H 22 N3O3[M+H] + Calculated value: 292.16, actual value: 292.20.

[0132] Step 3: To a stirred solution of (S)-3-(4-(4-acetylpiperazin-1-yl)phenyl)-2-(carboxyamino)propanoic acid (7 g, 20.87 mmol) in THF (25 mL) and water (25 mL) at 25° C. under nitrogen was added Fmoc-OSu (6.34 g, 18.79 mmol) followed by NaHCO3 (8.77 g, 104 mmol). The resulting mixture was stirred at 25° C. for 2 h. The pH was adjusted to 5 with 1N HCl and extracted with EtOAc (2×200 mL). The combined organic layers were washed with brine (3×100 mL), dried over anhydrous Na2SO4 and filtered. The filtrate was concentrated under reduced pressure and the residue was purified by reverse-phase flash chromatography under the following conditions: Column: C18 silica gel (330 g); Mobile phase A: water (0.1% TFA), Mobile phase B: MeCN; (Gradient: 5% B hold 5 min, to 55% B in 15 min, to 55% B hold 5 min; to 95% B in 20 min, to 95% B hold 5 min); Flow rate: 90 mL / min; Detector: UV 210 nm; RT = 45 min. Product-containing fractions were collected and concentrated under reduced pressure to give (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(4-(4-acetylpiperazin-1-yl)phenyl)propanoic acid. MS ESI: C 30 H 32 N3O5[M+H] + Calculated value: 514.23, actual value: 514.30. 1H NMR (400 MHz, methanol-d4) δ 7.78 - 7.76 (m, 2H), 7.61 - 7.52 (m, 2H), 7.40 - 7.37 (m, 2H), 7.32 - 7.22 (m, 4H), 7.08 - 6.98 (m, 2H), 4.47 - 4.43 (m, 1H), 4.33 - 4.31 (m, 1H), 4.14 - 4.02 (m, 2H), 3.68 - 3.63 (m, 4H), 3.23 - 3.08 (m, 5H), 2.91 - 2.85 (m, 1H), 2.11 (s, 3H).

[0133] Synthesis scheme 10 [ka] (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(4-(5-(tert-butoxycarbonyl)pyrimidin-2-yl)phenyl)propanoic acid Step 1: To a mixture of 2-chloropyrimidine-5-carboxylic acid (10 g, 63.1 mmol) in t-BuOH (100 mL) was added DMAP (0.771 g, 6.31 mmol) and Boc2O (16.52 g, 76 mmol) at room temperature under argon. The resulting mixture was stirred at 50° C. for 16 h, then it was cooled to room temperature and concentrated under reduced pressure. The residue was purified by silica gel chromatography eluting with 0-60% EtOAc / PE to give tert-butyl 2-chloropyrimidine-5-carboxylate. MS ESI: CH 12 ClN2O2[M+H] + Calculated value: 215.05, actual value: 214.95.

[0134] Step 2: To a mixture of tert-butyl 2-chloropyrimidine-5-carboxylate (4 g, 18.64 mmol) in 1,4-dioxane (40 mL) and water (8 mL) was added (S)-3-(4-boronophenyl)-2-((tert-butoxycarbonyl)amino)propanoic acid (8.64 g, 28.0 mmol), PdCl2(dppf) (1.364 g, 1.864 mmol) and K2CO3 (7.73 g, 55.9 mmol) at room temperature under argon. The resulting mixture was stirred at 80° C. for 3 h, then the pH was adjusted to 4 with 1N HCl and extracted with EtOAc (2×300 mL). The combined organic layers were washed with brine (3×50 mL), dried over anhydrous Na2SO4 and filtered. The filtrate was concentrated under reduced pressure. The residue was purified by silica gel chromatography eluting with 0-100% EtOAc / PE to give (S)-2-((tert-butoxycarbonyl)amino)-3-(4-(5-(tert-butoxycarbonyl)pyrimidin-2-yl)phenyl)propanoic acid. MS ESI:C 23 H 30 N3O6[M+H] + Calculated value: 444.21, actual value: 444.35. 1 H NMR (400 MHz, CDCl3) δ 9.27 (s, 2H), 8.42 (d, J = 7.8 Hz, 2H), 7.36 (d, J = 7.9 Hz, 2H), 4.69 - 4.68 (m, 1H), 3.29 - 3.24 (m, 2H), 1.63 (s, 9H), 1.44 (s, 9H).

[0135] Step 3: To a mixture of (S)-2-((tert-butoxycarbonyl)amino)-3-(4-(5-(tert-butoxycarbonyl)pyrimidin-2-yl)phenyl)propanoic acid (7 g, 15.78 mmol) in DCM (70 mL) at room temperature under argon was added TFA (14 mL, 182 mmol). After stirring at room temperature for 1 h, the reaction was concentrated to give (S)-2-amino-3-(4-(5-(tert-butoxycarbonyl)pyrimidin-2-yl)phenyl)propanoic acid. MS ESI:C 18 H 22 N3O4[M+H]+ Calculated value: 344.15, actual value: 344.20.

[0136] Step 4: To a mixture of (S)-2-amino-3-(4-(5-(tert-butoxycarbonyl)pyrimidin-2-yl)phenyl)propanoic acid (4 g, 11.65 mmol) in THF (40 mL) and H2O (40 mL) was added NaHCO3 (4.89 g, 58.2 mmol) and Fmoc-OSu (3.54 g, 10.48 mmol) at room temperature under argon. After stirring at room temperature for 1 h, the pH was adjusted to 4 with 1N HCl and the solution was extracted with EtOAc (2 x 300 mL). The combined organic layers were washed with brine (150 mL), dried over anhydrous Na2SO4 and filtered. The filtrate was concentrated under reduced pressure and the residue was purified by RP-flash under the following conditions: Column: C18 column (330 g); Mobile phase A: Water (0.05% TFA); Mobile phase B: MeCN; (Gradient: 0% B hold 5 min, to 73% B in 30 min, to 73% B hold 10 min; to 95% B in 4 min, to 95% B hold 10 min); Flow rate: 60 mL / min; Detector: UV254&210 nm; RT: 35.32 min to give (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(4-(5-(tert-butoxycarbonyl)pyrimidin-2-yl)phenyl)propanoic acid. MS ESI: C 33 H 32 N3O6[M+H] + Calculated value: 566.22, actual value: 566.35. 1 H NMR (300 MHz, DMSO-d6) δ 12.85 (s, 1H), 9.24 (s, 2H), 8.37 (d, J = 8.0 Hz, 2H), 7.88 - 7.79 (m, 3H), 7.66 - 7.14 (m, 8H), 4.30 - 4.14 (m, 4H), 3.22 - 3.16 (m, 1H), 3.05 - 2.82 (m, 1H), 1.59 (s, 9H).

[0137] Synthesis scheme 11 [ka] (2S,4R)-1-(((9H-fluoren-9-yl)methoxy)carbonyl)-4-(benzylamino)pyrrolidine-2-carboxylic acid Step 1: To a mixture of 1-(tert-butyl) 2-methyl (2S,4R)-4-aminopyrrolidine-1,2-dicarboxylate (10 g, 40.9 mmol) in DCM (100 mL) was added TEA (14.26 mL, 102 mmol) and NsCl (10.89 g, 49.1 mmol) at room temperature. The reaction was stirred at room temperature for 4 h. The resulting solution was quenched with water (150 mL) and extracted with EtOAc (3 x 300 mL). The organic layers were combined, washed with brine (2 x 150 mL), dried over anhydrous Na2SO4, and filtered. The filtrate was concentrated under reduced pressure. The residue was purified by silica gel chromatography eluting with a gradient of EtOAc / PE 0% to 45% to give 1-(tert-butyl) 2-methyl(2S,4R)-4-((4-nitrophenyl)sulfonamido)pyrrolidine-1,2-dicarboxylate. MS ESI:C 17 H 24 N3O8S[M+H-Boc] + Calculated value: 330.12, actual value: 330.10. 1 H NMR (400 MHz, CDCl3) δ 8.18 - 8.16 (m, 1H), 7.91 - 7.87 (m, 1H), 7.79 - 7.75 (m, 2H), 4.40 - 4.29 (m, 1H), 4.18 - 4.15 (m, 1H), 3.72 (s, 3H), 3.66 - 3.62 (m, 1H), 3.26 - 3.19 (m, 1H), 2.29 - 2.15 (m, 2H), 1.38 (s, 9H).

[0138] Step 2: To a mixture of 1-(tert-butyl) 2-methyl (2S,4R)-4-((4-nitrophenyl)sulfonamido)pyrrolidine-1,2-dicarboxylate (17 g, 39.6 mmol) in DCM (60 mL) at ambient temperature was added TFA (30 mL, 389 mmol). The reaction was stirred at ambient temperature for 2 h and then concentrated under reduced pressure to give methyl (2S,4R)-4-((4-nitrophenyl)sulfonamido)pyrrolidine-2-carboxylate. MS ESI:C 12 H 16N3O6S[M+H] + Calculated value: 330.07, actual value: 330.10.

[0139] Step 3: To a solution of methyl (2S,4R)-4-((4-nitrophenyl)sulfonamido)pyrrolidine-2-carboxylate (18.5 g, 39.3 mmol) in THF (150 mL) and water (150 mL) at ambient temperature was added NaHCO3 (16.52 g, 197 mmol) and Alloc-OSu (5.46 mL, 35.4 mmol). The reaction was stirred at ambient temperature for 4 h, then diluted with water (100 mL) and extracted with EtOAc (3 x 250 mL). The organic layers were combined, washed with brine (2 x 150 mL), dried over anhydrous Na2SO4, and filtered. The filtrate was concentrated under reduced pressure. The residue was purified by silica gel chromatography eluting with a gradient of EtOAc / PE 0% to 55% to give 1-allyl 2-methyl(2S,4R)-4-((4-nitrophenyl)sulfonamido)pyrrolidine-1,2-dicarboxylate. MS ESI:C 16 H 20 N3O8S[M+H] + Calculated value: 414.09, actual value: 413.95. 1 H NMR (300 MHz, CDCl3) δ 8.18 - 8.15 (m, 1H), 7.91 - 7.88 (m, 1H), 7.82 - 7.77 (m, 2H), 5.84 - 5.70 (m, 1H), 5.30 - 5.16 (m, 2H), 4.58 - 4.52 (m, 2H), 4.50 - 4.48 (m, 1H), 4.47 - 4.15 (m, 1H), 3.78 - 3.73 (m, 4H), 3.32 - 3.28 (m, 1H), 2.30 - 2.19 (m, 2H).

[0140] Step 4: To a solution of 1-allyl 2-methyl(2S,4R)-4-((4-nitrophenyl)sulfonamido)pyrrolidine-1,2-dicarboxylate (8.1 g, 19.59 mmol) in DMF (100 mL) at ambient temperature was added (bromomethyl)benzene (4.02 g, 23.51 mmol) and K2CO3 (8.12 g, 58.8 mmol). The reaction was stirred at ambient temperature for 4 h, then quenched with water (100 mL) and extracted with EtOAc (3 x 250 mL). The organic layers were combined, washed with brine (4 x 200 mL), dried over anhydrous Na2SO4, and filtered. The filtrate was concentrated under reduced pressure. The residue was purified by silica gel chromatography eluting with a gradient of 0% to 55% EtOAc / PE to give 1-allyl 2-methyl(2S,4R)-4-((N-benzyl-4-nitrophenyl)sulfonamido)pyrrolidine-1,2-dicarboxylate. MS ESI:C 23 H 26 N3O8S[M+H] + Calculated value: 504.14, actual value: 504.20. 1 H NMR (300 MHz, CDCl3) δ 7.88 - 7.84 (m, 1H), 7.68 - 7.67 (m, 2H), 7.66 - 7.55 (m, 1H), 7.29 - 7.23 (m, 5H), 5.85 - 5.75 (m, 1H), 5.25 - 5.18 (m, 2H), 4.19 - 4.15 (m, 6H), 3.73 - 3.25 (m, 5H), 2.23 - 2.15 (m, 2H).

[0141] Step 5: To a solution of 1-allyl 2-methyl(2S,4R)-4-((N-benzyl-4-nitrophenyl)sulfonamido)pyrrolidine-1,2-dicarboxylate (9.3 g, 18.47 mmol) in DMF (40 mL) at ambient temperature was added 4-methoxybenzenethiol (3.11 g, 22.16 mmol) and K2CO3 (7.66 g, 55.4 mmol). The reaction was stirred at ambient temperature for 1 h, then diluted with water (100 mL) and extracted with EtOAc (3 x 150 mL). The organic layers were combined, washed with brine (2 x 100 mL), dried over anhydrous Na2SO4, and filtered. The residue was purified by RP-Flash under the following conditions: Column: Flash C18 (330 g); Mobile phase A: Water (0.05% TFA), Mobile phase B: MeCN; (Gradient: 5% B hold 5 min, to 32% B in 15 min, hold 32% B for 6 min; to 95% B in 5 min, hold 95% B for 5 min); Flow rate: 90 mL / min; Detector: UV 210 nm; RT=36 min. Product-containing fractions were collected and concentrated under reduced pressure to give 1-allyl 2-methyl(2S,4R)-4-(benzylamino)pyrrolidine-1,2-dicarboxylate. MS ESI: C 17 H 23 N2O4[M+H] + Calculated value: 319.16, actual value: 319.10. 1 H NMR (300 MHz, CDCl3) δ 7.39 - 7.34 (m, 5H), 5.90 - 5.85 (m, 1H), 5.32 - 5.17 (m, 2H), 4.60 - 4.36 (m, 3H), 3.96 - 3.57 (m, 8H), 2.07 - 1.99 (m, 2H).

[0142] Step 6: To a solution of 1-allyl 2-methyl(2S,4R)-4-(benzylamino)pyrrolidine-1,2-dicarboxylate (4.5 g, 12.72 mmol) in DCM (40 mL) at ambient temperature was added Boc2O (4.16 g, 19.08 mmol) and TEA (5.32 mL, 38.2 mmol). The reaction was stirred at ambient temperature for 4 h, then quenched with water (30 mL) and extracted with EtOAc (3 x 200 mL). The organic layers were combined, washed with brine (2 x 100 mL), dried over anhydrous Na2SO4, and filtered. The filtrate was concentrated under reduced pressure. The residue was purified by silica gel chromatography eluting with a gradient of 0% to 40% EtOAc / PE to give 1-allyl 2-methyl(2S,4R)-4-(benzyl(tert-butoxycarbonyl)amino)pyrrolidine-1,2-dicarboxylate. MS ESI:C 22 H 31 N2O6[M+H] + Calculated value: 419.21, actual value: 419.15. 1 H NMR (300 MHz, CDCl3) δ 7.34 - 7.15 (m, 5H), 5.90 - 5.85 (m, 1H), 5.25 - 5.18 (m, 2H), 4.58 - 4.32 (m, 6H), 3.74 - 3.44 (m, 5H), 2.49 - 2.38 (m, 1H), 2.12 - 1.99 (m, 1H), 1.42 (s, 9H).

[0143] Step 7: To a mixture of 1-allyl 2-methyl(2S,4R)-4-(benzyl(tert-butoxycarbonyl)amino)pyrrolidine-1,2-dicarboxylate (3 g, 5.73 mmol) in THF (15 mL) was added LiOH (11.47 mL, 11.47 mmol, 1 M aqueous solution) at 0 °C. The reaction was stirred at ambient temperature for 2 h, then acidified to pH 3-4 with aqueous HCl and extracted with EtOAc (3 x 150 mL). The organic layers were combined, washed with brine (2 x 80 mL), dried over anhydrous Na2SO4, and filtered. The filtrate was concentrated under reduced pressure to give (2S,4R)-1-((allyloxy)carbonyl)-4-(benzyl(tert-butoxycarbonyl)amino)pyrrolidine-2-carboxylic acid. MS ESI: C 21 H 29 N2O6[M+H] + Calculated value: 405.19, actual value: 405.30. 1 H NMR (300 MHz, CDCl3) δ 7.35 - 7.14 (m, 5H), 5.89 - 5.75 (m, 1H), 5.14 - 5.19 (m, 2H), 4.61 - 4.30 (m, 6H), 3.82 - 3.32 (m, 2H), 2.47 - 2.18 (m, 2H), 1.42 (s, 9H).

[0144] Step 8: To a mixture of (2S,4R)-1-((allyloxy)carbonyl)-4-(benzyl(tert-butoxycarbonyl)amino)pyrrolidine-2-carboxylic acid (2.6 g, 5.46 mmol) in DCM (150 mL) was added AcOH (0.751 mL, 13.11 mmol) and Pd(Ph3P)4 (0.126 g, 0.109 mmol) at ambient temperature, and then Bu3SnH (1.749 g, 6.01 mmol) was added to the reaction. After the reaction was stirred at ambient temperature for 4 hours, it was concentrated under reduced pressure to give (2S,4R)-4-(benzyl(tert-butoxycarbonyl)amino)pyrrolidine-2-carboxylic acid. MS ESI:C 17 H 25 N2O4[M+H] + Calculated value: 321.17, actual value: 321.15.

[0145] Step 9: To a mixture of (2S,4R)-4-(benzyl(tert-butoxycarbonyl)amino)pyrrolidine-2-carboxylic acid (3.2 g, 4.99 mmol) in THF (50 mL) and water (50 mL) was added NaHCO3 (2.098 g, 24.97 mmol) to pH 8-9, then Fmoc-OSu (1.516 g, 4.49 mmol) was added to the reaction. The resulting mixture was stirred at ambient temperature for 4 h, then acidified to pH 3-4 with aqueous HCl and extracted with EtOAc (3 x 200 mL). The organic layers were combined, washed with brine (2 x 100 mL), dried over anhydrous Na2SO4, and filtered. The filtrate was concentrated under reduced pressure to give the crude product. The residue was purified by RP-flash under the following conditions: Column: Flash C18 (330 g); Mobile phase A: Water (0.05% TFA), Mobile phase B: MeCN; (Gradient: 5% B hold 5 min, to 70% B in 25 min, hold 70% B for 8 min; to 95% B in 2 min, hold 95% B for 5 min); Flow rate: 90 mL / min; Detector: UV 210 nm; RT=45 min. Product-containing fractions were collected and concentrated under reduced pressure to give (2S,4R)-1-(((9H-fluoren-9-yl)methoxy)carbonyl)-4-(benzyl(tert-butoxycarbonyl)amino)pyrrolidine-2-carboxylic acid. MS ESI: C 32 H 35 N2O6[M+H] + Calculated value: 543.24, actual value: 543.20. 1 H NMR (400 MHz, DMSO-d6) δ 12.93 (s, 2H), 7.88 (d, J = 7.6 Hz, 2H), 7.61 - 7.57 (m, 2H), 7.43 - 7.18 (m, 9H), 4.43 - 4.15 (m, 8H), 3.53 - 3.25 (m, 2H), 2.51 - 2.50 (m, 1H), 2.03 - 1.92 (m, 1H), 1.37 (s, 2H).

[0146] Synthesis scheme 12 [ka] TIFF0007645434000035.tif41136 (2S,4R)-1-(((9H-fluoren-9-yl)methoxy)carbonyl)-4-(phenethylamino)pyrrolidine-2-carboxylic acid Step 1: To a mixture of 1-allyl 2-methyl(2S,4R)-4-((4-nitrophenyl)sulfonamido)pyrrolidine-1,2-dicarboxylate (7.2 g, 17.42 mmol), DIAD (5.08 mL, 26.1 mmol), and PPh3 (6.85 g, 26.1 mmol) in THF (70 mL) at ambient temperature was added 2-phenylethan-1-ol (3.19 g, 26.1 mmol). The reaction was stirred at ambient temperature for 4 h, then quenched with water (100 mL) and extracted with EtOAc (3 x 250 mL). The organic layers were combined, washed with brine (4 x 200 mL), dried over anhydrous Na2SO4, and filtered. The filtrate was concentrated under reduced pressure. The residue was purified by silica gel chromatography eluting with a gradient of EtOAc / PE 0% to 55% to give 1-allyl 2-methyl(2S,4R)-4-((4-nitro-N-phenethylphenyl)sulfonamido)pyrrolidine-1,2-dicarboxylate. MS ESI:C 24 H 28 N3O8S[M+H] + Calculated value: 518.15, actual value: 518.20.

[0147] Step 2: To a mixture of 1-allyl 2-methyl(2S,4R)-4-((4-nitro-N-phenethylphenyl)sulfonamido)pyrrolidine-1,2-dicarboxylate (7.2 g, 13.91 mmol) in DMF (80 mL) at ambient temperature was added 4-methoxybenzenethiol (1.950 g, 13.91 mmol) and K2CO3 (1.923 g, 13.91 mmol). The reaction was stirred at ambient temperature for 1 h, then diluted with water (200 mL) and extracted with EtOAc (3 x 150 mL). The organic layers were combined, washed with brine (2 x 500 mL), dried over anhydrous Na2SO4, and filtered. The filtrate was concentrated under reduced pressure and the residue was purified by RP-Flash under the following conditions: Column: Flash C18 (330 g); Mobile phase A: Water (0.05% TFA), Mobile phase B: MeCN; (Gradient: 5% B hold 5 min, to 32% B in 15 min, to 32% B hold 6 min; to 95% B in 5 min, to 95% B hold 5 min); Flow rate: 90 mL / min; Detector: UV 210 nm; RT=36 min. Product-containing fractions were collected and concentrated under reduced pressure to give 1-allyl 2-methyl(2S,4R)-4-(phenethylamino)pyrrolidine-1,2-dicarboxylate. MS ESI: C 18 H 25 N2O4[M+H] + Calculated value: 333.17, actual value: 333.25.

[0148] Step 3: To a mixture of 1-allyl 2-methyl(2S,4R)-4-(phenethylamino)pyrrolidine-1,2-dicarboxylate (4 g, 12.03 mmol) in DCM (40 mL) at ambient temperature was added Boc2O (3.93 g, 18.04 mmol) and TEA (3.35 mL, 24.07 mmol). The reaction was stirred at ambient temperature for 4 h, then quenched with water (30 mL) and extracted with EtOAc (3 x 200 mL). The organic layers were combined, washed with brine (2 x 100 mL), dried over anhydrous Na2SO4, and filtered. The filtrate was concentrated under reduced pressure. The residue was purified by silica gel chromatography eluting with a gradient of EtOAc / PE 0% to 40% to give 1-allyl 2-methyl(2S,4R)-4-((tert-butoxycarbonyl)(phenethyl)amino)pyrrolidine-1,2-dicarboxylate. MS ESI:C 23 H 33 N2O6[M+H] + Calculated value: 433.23, actual value: 433.25.

[0149] Step 4: To a mixture of 1-allyl 2-methyl(2S,4R)-4-((tert-butoxycarbonyl)(phenethyl)amino)pyrrolidine-1,2-dicarboxylate (3.84 g, 8.88 mmol) in THF (30 mL) was added LiOH (17.76 mL, 17.76 mmol, 1 M aqueous solution) at 0 °C. The reaction was stirred at ambient temperature for 2 h, then acidified to pH 3-4 with aqueous HCl and extracted with EtOAc (3 x 150 mL). The organic layers were combined, washed with brine (2 x 80 mL), dried over anhydrous Na2SO4, and filtered. The filtrate was concentrated under reduced pressure and the crude product was used directly in the next step without further purification. MS ESI:C 22 H 31 N2O6[M+H] + Calculated value: 419.21, actual value: 419.35.

[0150] Step 5: To a mixture of (2S,4R)-1-((allyloxy)carbonyl)-4-((tert-butoxycarbonyl)(phenethyl)amino)pyrrolidine-2-carboxylic acid (7 g, 16.73 mmol) in DCM (150 mL) was added AcOH (2.411 g, 40.1 mmol) and Pd(Ph3P)4 (3.87 g, 3.35 mmol) at ambient temperature, then Bu3SnH (5.36 g, 18.40 mmol) was added to the reaction. The reaction was stirred at ambient temperature for 4 h and then concentrated under reduced pressure. The crude product was used directly in the next step without further purification. MS ESI:C 18 H 27 N2O4[M+H] + Calculated value: 335.19, actual value: 335.25.

[0151] Step 6: To a mixture of (2S,4R)-4-((tert-butoxycarbonyl)(phenethyl)amino)pyrrolidine-2-carboxylic acid (5.4 g, 16.15 mmol) in THF (50 mL) and water (50 mL) at room temperature was added NaHCO3 (8.56 g, 81 mmol) followed by Fmoc-OSu (4.89 g, 14.54 mmol). After the reaction was stirred at ambient temperature for 4 h, it was acidified to pH 3-4 with aqueous HCl and extracted with EtOAc (3 x 400 mL). The organic layers were combined, washed with brine (2 x 200 mL), dried over anhydrous Na2SO4 and filtered. The filtrate was concentrated under reduced pressure. The residue was purified by RP-flash under the following conditions: Column: Flash C18 (330 g); Mobile phase A: Water (0.05% TFA), Mobile phase B: MeCN; (Gradient: 5% B hold 5 min, to 70% B in 25 min, to 70% B hold 8 min; to 95% B in 2 min, to 95% B hold 5 min); Flow rate: 90 mL / min; Detector: UV 210 nm; RT=45 min. Product-containing fractions were collected and concentrated under reduced pressure to give (2S,4R)-1-(((9H-fluoren-9-yl)methoxy)carbonyl)-4-((tert-butoxycarbonyl)(phenethyl)amino)pyrrolidine-2-carboxylic acid. MS ESI: C 33 H 37 N2O6Na[M+Na] + Calculated value: 579.26, actual value: 579.25. 1H NMR (300 MHz, methanol-d4) δ 7.78 - 7.76 (m, 2H), 7.63 - 7.57 (m, 2H), 7.44 - 7.24 (m, 6H), 7.20 - 7.17 (m, 3H), 4.51 - 4.28 (m, 4H), 4.25 - 4.15 (m, 1H), 3.60 - 3.41 (m, 1H), 3.39 - 3.31 (m, 3H), 2.82 - 2.77 (m, 2H), 2.45 - 2.44 (m, 1H), 1.98 - 1.94 (m, 1H), 1.46 (s, 9H).

[0152] Synthesis scheme 13 [ka] (2S,4R)-1-(((9H-fluoren-9-yl)methoxy)carbonyl)-4-(ethylamino)pyrrolidine-2-carboxylic acid Step 1: To a mixture of 1-allyl 2-methyl(2S,4R)-4-((4-nitrophenyl)sulfonamido)pyrrolidine-1,2-dicarboxylate (9.6 g, 23.22 mmol) in DMF (100 mL) was added iodoethane (4.35 g, 27.9 mmol) and K2CO3 (9.63 g, 69.7 mmol) at room temperature. The reaction was stirred at room temperature for 12 h, then quenched with water (80 mL) and extracted with EtOAc (3 x 200 mL). The organic layers were combined, washed with brine (4 x 150 mL), dried over anhydrous Na2SO4, and filtered. The filtrate was concentrated under reduced pressure. The residue was purified by silica gel chromatography eluting with a gradient of 0% to 55% EtOAc / PE to give 1-allyl 2-methyl(2S,4R)-4-((N-ethyl-4-nitrophenyl)sulfonamido)pyrrolidine-1,2-dicarboxylate. MS ESI:C 18 H 24 N3O8S[M+H] + Calculated value: 442.12, actual value: 442.05. 1H NMR (300 MHz, CDCl3) δ 8.02 - 8.01 (m, 1H), 7.73 - 7.62 (m, 3H), 5.95 - 5.78 (m, 1H), 5.29 - 5.18 (m, 2H), 4.58 - 4.43 (m, 4H), 3.76 - 3.73 (m, 4H), 3.38 - 3.29 (m, 3H), 2.29 - 2.26 (m, 2H), 1.26 - 1.20 (m, 3H).

[0153] Step 2: To a stirred mixture of 1-allyl 2-methyl(2S,4R)-4-((N-ethyl-4-nitrophenyl)sulfonamido)pyrrolidine-1,2-dicarboxylate (5.5 g, 12.46 mmol) and K2CO3 (5.17 g, 37.4 mmol) in DMF (80 mL) was added 4-methoxybenzenethiol (3.49 g, 24.92 mmol) at room temperature. After the mixture was stirred at room temperature for 1 h, it was quenched with water (200 mL) and extracted with EtOAc (3 x 200 mL). The combined organic layers were washed with brine (3 x 100 mL) and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure and the residue was purified by silica gel column chromatography eluting with 0% to 3% MeOH / DCM to give 1-allyl 2-methyl(2S,4R)-4-(ethylamino)pyrrolidine-1,2-dicarboxylate. MS ESI:C 12 H 21 N2O4[M+H] + Calculated value: 257.14, measured value: 257.15. 1 H NMR (300 MHz, CDCl3) δ 6.02 - 5.76 (m, 1H), 5.40 - 5.11 (m, 2H), 4.68 - 4.38 (m, 3H), 3.91 - 3.77 (m, 1H), 3.77 - 3.67 (m, 3H), 3.61 - 3.42 (m, 1H), 3.42 - 3.21 (m, 1H), 2.68 (q, J = 7.2 Hz, 2H), 2.28 - 2.00 (m, 2H), 1.14 (t, J = 7.1 Hz, 3H).

[0154] Step 3: To a stirred solution of 1-allyl 2-methyl(2S,4R)-4-(ethylamino)pyrrolidine-1,2-dicarboxylate (2.9 g, 11.31 mmol) and TEA (3.15 mL, 22.63 mmol) in DCM (50 mL) was added BocO (3.94 mL, 16.97 mmol) at room temperature. The resulting solution was stirred at 25° C. for 16 h and then concentrated under reduced pressure. The residue was purified by silica gel column chromatography eluting with 0-30% EtOAc / PE to give 1-allyl 2-methyl(2S,4R)-4-((tert-butoxycarbonyl)(ethyl)amino)pyrrolidine-1,2-dicarboxylate. MS ESI:C 17 H 29 N2O6[M+H] + Calculated value: 357.19, actual value: 357.30. 1 H NMR (400 MHz, CDCl3) δ 6.01 - 5.79 (m, 1H), 5.37 - 5.14 (m, 2H), 4.70 - 4.40 (m, 4H), 3.85 - 3.75 (m, 1H), 3.73 (d, J = 7.8 Hz, 3H), 3.49 - 3.36 (m, 1H), 3.28 - 3.06 (m, 2H), 2.53 - 2.38 (m, 1H), 2.22 - 2.07 (m, 1H), 1.46 (s, 9H), 1.11 (t, J = 7.0 Hz, 3H).

[0155] Step 4: To a stirred solution of 1-allyl 2-methyl(2S,4R)-4-((tert-butoxycarbonyl)(ethyl)amino)pyrrolidine-1,2-dicarboxylate (3.8 g, 10.66 mmol) in THF (50 mL) was added LiOH (32 mL, 32.0 mmol, 1N aqueous solution) at room temperature. After stirring the solution at 25° C. for 2 h, the pH of the solution was adjusted to 3 with 1N HCl and then extracted with EtOAc (3×100 mL). The combined organic layers were washed with brine (100 mL) and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure to give (2S,4R)-1-((allyloxy)carbonyl)-4-((tert-butoxycarbonyl)(ethyl)amino)pyrrolidine-2-carboxylic acid. MS ESI:C 16 H 27 N2O6[M+H] + Calculated value: 343.18, actual value: 343.30.

[0156] Step 5: To a stirred solution of (2S,4R)-1-((allyloxy)carbonyl)-4-((tert-butoxycarbonyl)(ethyl)amino)pyrrolidine-2-carboxylic acid (3.4 g, 9.93 mmol), AcOH (1.431 g, 23.83 mmol) and Pd(Ph3P)4 (2.295 g, 1.986 mmol) in DCM (150 mL) was added Bu3SnH (3.18 g, 10.92 mmol) dropwise at room temperature. The resulting solution was stirred at 25° C. for 1 h and then concentrated under reduced pressure to give (2S,4R)-4-((tert-butoxycarbonyl)(ethyl)amino)pyrrolidine-2-carboxylic acid. MS ESI:C 12 H 23 N2O4[M+H] + Calculated value: 259.16, actual value: 259.30.

[0157] Step 6: To a stirred solution of (2S,4R)-4-((tert-butoxycarbonyl)(ethyl)amino)pyrrolidine-2-carboxylic acid (2.5 g, 9.68 mmol) and Na2CO3 (3.08 g, 29.0 mmol) in THF (60 mL) and water (60 mL) was added Fmoc-OSu (3.26 g, 9.68 mmol) at room temperature. After stirring the resulting mixture at 25° C. for 16 h, the pH was adjusted to 3 with 1N HCl and then extracted with EtOAc (3×100 mL). The combined organic layers were washed with brine (2×50 mL) and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure and the residue was purified by RP-flash with the following conditions: 330 g C18 column, 5%-5% 5 min, 5% to 65% MeCN / water (0.05% TFA) in 40 min to give (2S,4R)-1-(((9H-fluoren-9-yl)methoxy)carbonyl)-4-((tert-butoxycarbonyl)(ethyl)amino)pyrrolidine-2-carboxylic acid. MS ESI:C 27 H 33 N2O6[M+H] + Calculated value: 481.23, actual value: 481.10. 1 H NMR (400 MHz, DMSO-d6) δ 12.98 (br, 1H), 7.90 (d, J = 7.6 Hz, 2H), 7.68 - 7.64 (m, 2H), 7.44 - 7.41 (m, 2H), 7.35 - 7.31 (m, 2H), 4.43 - 4.18 (m, 5H), 3.60 - 3.56 (m, 1H), 3.29 - 3.22 (m, 1H), 3.16 - 3.12 (m, 2H), 2.42 - 2.38 (m, 1H), 2.08 - 2.01 (m, 1H), 1.40 (s, 9H), 1.05 - 1.00 (m, 3H).

[0158] Synthesis scheme 14 [ka] (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(2-(4-acetylpiperazin-1-yl)pyrimidin-5-yl)propanoic acid Step 1: To a mixture of 5-bromo-2-fluoropyrimidine (7 g, 39.6 mmol) in DMF (70 mL) at room temperature was added 1-(piperazin-1-yl)ethan-1-one (10.14 g, 79 mmol) and K2CO3 (10.93 g, 79 mmol). The reaction mixture was stirred at 100° C. for 2 h and then cooled back to room temperature. The reaction mixture was extracted with 500 mL of EtOAc, washed with H2O (3×100 mL) and brine (80 mL), dried over Na2SO4, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography eluting with a gradient of 0% to 80% EtOAc / PE. Fractions containing the desired product were combined and concentrated under reduced pressure to give 4-(5-bromopyrimidin-2-yl)piperazine-1-carbaldehyde. MS ESI:C 10 H 14 BrNO[M+H] + Calculated values: 285.03, 287.03, actual values: 284.95, 286.95. 1 H NMR (300 MHz, CDCl3) δ 8.32 (s, 2H), 3.90 - 3.74 (m, 4H), 3.74 - 3.64 (m, 2H), 3.58 - 3.48 (m, 2H), 2.15 (s, 3H).

[0159] Step 2: A mixture of nickel(II) chloride ethylene glycol dimethyl ether complex (0.693 g, 3.16 mmol) and 1,10-phenanthroline (0.569 g, 3.16 mmol) in DMA (50 mL) was heated at 50° C. for 0.5 h. A solution of 1-(4-(5-bromopyrimidin-2-yl)piperazin-1-yl)ethan-1-one (4.5 g, 15.78 mmol), tert-butyl (R)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-iodopropanoate (11.68 g, 23.67 mmol) and TBAI (5.83 g, 15.78 mmol) in DMA (50 mL) was added at 25° C. Then Zn (2.064 g, 31.6 mmol) was added. After stirring the resulting mixture at 30° C. for 24 hours, the reaction mixture was filtered and washed with DCM. The organic layer was concentrated under reduced pressure. The residue was purified by RP-flash under the following conditions: Column: C18 column (330 g); Mobile phase A: Water (0.05% TFA); Mobile phase B: MeCN; (Gradient: 0% B hold 5 min, to 82% B in 30 min, hold 82% B for 6 min; to 95% B in 2 min, hold 95% B for 3 min); Flow rate: 60 mL / min; Detector: UV254×210 nm; RT: 35 min to give tert-butyl (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(2-(4-acetylpiperazin-1-yl)pyrimidin-5-yl)propanoate. MS ESI: C 32 H 38 N5O5[M+H] + Calculated value: 572.28, actual value: 572.30. 1H NMR (300 MHz, CDCl3) δ 8.19 (s, 2H), 7.77 (d, J = 7.6 Hz, 2H), 7.63 - 7.52 (m, 2H), 7.46 - 7.26 (m, 4H), 4.48 - 4.42 (m,2H), 4.35 - 4.30 (m, 1H), 4.22 - 4.17 (m, 1H), 3.89 - 3.81 (m, 4H), 3.71 - 3.68 (m, 2H), 3.56 - 3.52 (m, 2H), 3.08 - 3.02 (m, 1H), 2.92 - 2.87 (m, 1H), 2.17 (s, 3H), 1.48 (s, 9H).

[0160] Step 3: To a mixture of tert-butyl (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(2-(4-acetylpiperazin-1-yl)pyrimidin-5-yl)propanoate (4.6 g, 8.05 mmol) in DCM (40 mL) at room temperature under argon was added TFA (80 mL, 1038 mmol). The reaction was stirred at room temperature for 3 h and then concentrated under reduced pressure. The residue was purified by RP-flash under the following conditions: Column: C18 gel column (330 g); Mobile phase A: Water (0.05% TFA); Mobile phase B: MeCN; (Gradient: 0% B hold 5 min, to 72% B in 30 min, to 72% B hold 6 min; to 95% B in 2 min, to 95% B hold 10 min); Flow rate: 60 mL / min; Detector: UV254&210 nm; RT: 35 min to give (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(2-(4-acetylpiperazin-1-yl)pyrimidin-5-yl)propanoic acid. MS ESI: C 28 H 30 N5O5[M+H] + Calculated value: 516.22, actual value: 516.35. 1H NMR (300 MHz, DMSO-d6) δ 8.31 (s, 2H), 7.89 (d, J = 7.5 Hz, 2H), 7.81 (d, J = 8.6 Hz, 1H), 7.74 - 7.62 (m, 2H), 7.59 - 7.22 (m, 4H), 4.29 - 4.09 (m, 4H), 3.74 - 3.60 (m, 4H), 3.47 - 3.46 (m, 4H), 2.98 - 2.92 (m, 1H), 2.76 - 2.68 (m, 1H), 2.03 (s, 3H).

[0161] Synthesis scheme 15 [ka] (2S,3S)-2-Amino-3-(4-fluoro-1H-indol-3-yl)butanoic acid In a 1-liter three-necked round-bottom flask purged and maintained with an inert atmosphere of nitrogen, 4-fluoro-1H-indole (10 g, 1.00 equiv.), L-threonine (10.6 g, 1.20 equiv.), DMSO (100 mL), potassium phosphate buffer (0.2 M, 300 mL, pH=7.4) were added. The reaction mixture was heated to 65° C., and then PfTrpB-7E6 (2.5 g, 25 wt %) and 3-hydroxy-2-methyl-5-([phosphonooxy]methyl)-4-pyridinecarboxaldehyde (0.078 g, 0.004 equiv.) were added. The resulting solution was stirred at 65° C. overnight. The mixture was cooled to room temperature and used directly in the next step.

[0162] To the above reaction mixture was added THF (100 mL), sodium carbonate (23.56 g, 3.0 equiv.) and 2,5-dioxopyrrolidin-1-yl 9H-fluoren-9-yl methyl carbonate (29.96 g, 1.20 equiv.) at 0° C. The resulting solution was stirred at room temperature overnight. The pH was adjusted to 4 with 3M HCl and the solid precipitate was filtered off. The resulting solution was extracted with EtOAc (3×500 mL). The organic fractions were combined, washed with brine (1 L), dried over anhydrous sodium sulfate and concentrated under reduced pressure. The mixture was purified on a silica gel column with MeOH:DCM=1:5. HPLC-MS: (ES, m / z): [M+1]: 459. 1 H NMR (300 MHz, DMSO-d6) δ 12.60 (s, 1H), 11.15 (s, 1H), 7.87 (d, J = 7.6 Hz, 2H), 7.76 - 7.49 (m, 3H), 7.47 - 7.34 (m, 2H), 7.34 - 7.16 (m, 4H), 7.03 (td, J = 7.9, 5.0 Hz, 1H), 6.73 (dd, J = 11.8, 7.7 Hz, 1H), 4.36 (t, J = 8.5 Hz, 1H), 4.31 - 4.02 (m, 3H), 3.51 (q, J = 7.4 Hz, 1H), 1.31 (d, J = 7.0 Hz, 4H), 0.78 (s, 1H).

[0163] Synthesis scheme 16 [ka] (2S,3S)-2-Amino-3-(4-chloro-1H-indol-3-yl)butanoic acid A 1-liter three-necked round-bottom flask purged and maintained under an inert atmosphere of nitrogen was charged with 4-chloro-1H-indole (10 g, 1.00 equiv.), L-threonine (14.09 g, 1.8 equiv.), DMSO (100 mL), potassium phosphate buffer (0.2 M, 300 mL, pH=7.4), the reaction mixture was heated to 65° C., and then PfTrpB-7E6 (7.5 g, 25 wt %) and 3-hydroxy-2-methyl-5-([phosphonooxy]methyl)-4-pyridinecarboxaldehyde (174 mg, 0.01 equiv.) were added. The resulting solution was stirred at 65° C. for 36 hours. The mixture was cooled to room temperature and used directly in the next step. To the above reaction mixture was added THF (100 mL), sodium carbonate (20.9 g, 3.0 equiv.) and 2,5-dioxopyrrolidin-1-yl 9H-fluoren-9-yl methyl carbonate (31.0 g, 1.40 equiv.) at 0° C. The resulting solution was stirred at room temperature overnight. The pH was adjusted to 4 with 3M HCl and the solid precipitate was filtered off. The resulting solution was extracted with EtOAc (3×500 mL). The organic fractions were combined, washed with brine (1 L), dried over anhydrous sodium sulfate and concentrated under reduced pressure. HPLC-MS: (ES, m / z): [M+1]: 475.

[0164] Synthesis scheme 17 [ka] (2S,3S)-2-Amino-3-(naphthalen-1-yl)butanoic acid Step 1: To a solution of (2S,3R)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-hydroxybutanoic acid (250 g, 1.00 equiv.) in DMF (1.5 L) was added benzyl bromide (250 g, 2.00 equiv.) dropwise at 20° C. Cesium carbonate (477 g, 2.00 equiv.) was then added and the solution was stirred at 20° C. for 3 h. The reaction was poured into ice-HO (3 L) and extracted with EtOAc (3×500 mL). The organic layer was washed with 3% LiCl solution (2×500 mL) and brine (500 mL), dried over sodium sulfate, and concentrated under reduced pressure at 40° C. The crude product was triturated with methyl tert-butyl ether:PE=6:1. 1H NMR (400 MHz, CDCl3): δ 7.77 (d, J = 7.6 Hz, 1H), 7.40 (d, J = 8.0 Hz, 1H), 7.31-7.36 (m, 10H), 5.65-5.71 m, 1H), 5.13-5.31 (m, 3H), 4.39-4.43 (m, 3H), 4.22-4.25 (m, 1H), 1.25 (d, J = 6.4 Hz, 3H)

[0165] Step 2: To a three-necked round bottom flask was added (2S,3R)-benzyl 2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-hydroxybutanoate (125 g, 1.00 equiv.) and DCE (750 mL) under an inert atmosphere of nitrogen. The reaction was cooled to 0° C., then NIS (195 g, 3.00 equiv.) and PPh3 (228 g, 3.00 equiv.) were added. The temperature was raised to 50° C., and the reaction mixture was stirred for 3 h. The reaction was poured into ice-HO (500 mL) and extracted with DCM (2×500 mL). The organic layer was dried over sodium sulfate and concentrated under reduced pressure at 40° C. The residue was purified by silica gel column chromatography (PE / EtOAc=1 / 0 to 0 / 1). 1 H NMR (400 MHz, CDCl3): δ 7.78 (d, J = 7.6 Hz, 2H), 7.68 (d, J = 7.2 Hz, 2H), 7.33-7.43 (m, 9H), 5.27-5.68 (m, 1H), 5.21-5.23 (m, 2H), 4.39-4.52 (m, 3H), 4.25-4.38 (m, 1H), 1.91-1.95 (m, 3H).

[0166] Step 3: A three-necked round-bottom flask was charged with 2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-iodobutanoate, 1-iodonaphthalene (42.2 g, 1.20 equiv), TBAI (76.7 g, 1.50 equiv), Zn (19.0 g, 2.10 equiv), and DMA (750 mL). A second three-necked round-bottom flask was charged with picoline imidamide·2HCl (42.2 g, 1.20 equiv), NiCl2·glyme (7.61 g, 0.25 equiv), and DMA (750 mL) at 25 °C. The contents of the second flask were added to the first flask under argon. The resulting mixture was stirred at 25 °C for 12 h. The reaction was poured into iced HO (3 L) and extracted with EtOAc (2 x 1 L). The organic layer was dried over sodium sulfate and concentrated under reduced pressure at 40° C. The crude product was purified by reverse phase HPLC (MeCN:H2O). HPLC-MS: [M+23]: 564. 1 H NMR (400 MHz, CDCl3) δ: 8.17-8.24 (m, 1H), 8.15-8.17 (m, 1H), 7.77-7.87 (m, 2H), 7.76-7.77 (m, 4H), 7.30-7.41 (m, 10H), 5.30-5.38 (m, 1H), 4.96-5.04 (m, 3H), 4.85-4.87 (m, 1H), 4.30-4.34 (m, 1H), 4.18-4.26 (m, 4H), 1.43-1.45 (m, 3H).

[0167] Step 4: 143 g of (2S)-benzyl 2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(naphthalen-1-yl)butanoate was separated by SFC. The organic layer was concentrated under reduced pressure at 35° C.

[0168] Peak 1: (2S,3R)-benzyl 2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(naphthalen-1-yl)butanoate. 1H NMR (400 MHz, DMSO-d6): δ 8.11-8.12 (m, 2H), 8.10-8.11 (m, 1H), 7.88-7.90 (m, 2H), 7.54-7.88 (m, 1H), 7.44-7.53 (m, 2H), 7.42-7.44 (m, 4H), 7.33-7.42 (m, 3H), 7.27-7.33 (m, 6H), 7.08-7.09 (m, 2H), 4.91-4.94 (m, 1H), 4.79- 4.82 (m, 1H), 4.58 (t, J = 8.0 Hz), 4.17-4.25 (m, 4H), 1.39 (d, J = 6.8 Hz, 3H).Peak 2: (2S,3S)-benzyl 2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(naphthalen-1-yl)butanoate. 1 H NMR (400 MHz, DMSO-d6): δ 7.92-8.15 (m, 1H), 7.86-7.92 (m, 1H), 7.84-7.86 (m, 1H), 7.57-7.84 (m, 2H), 7.56-7.57 (m, 1H), 7.41-7.54 (m, 4H), 7.30-7.38 (m, 4H), 7.27-7.30 (m, 7H), 5.08-5.14 (m, 2H), 4.65 (t, J = 8.0 Hz), 4.23-4.26 (m, 1H), 4.05-4.18 (m, 3H), 1.30 (d, J = 6.8 Hz, 3H).

[0169] Step 5: A three-necked round bottom flask was charged with (2S,3S)-benzyl 2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(naphthalen-1-yl)butanoate (40.0 g, 1.00 equiv.) and THF (200 mL). 10% wet Pd / C (7.00 g) was added and the reaction was purged with H2 three times and stirred under H2 (15 psi) at 25°C for 12 h. The reaction was filtered through a pad of Celite and concentrated under reduced pressure at 35°C. The crude product was triturated with PE at 25°C for 1 h. After filtration, the filter cake was dissolved in MeCN (100 mL) and concentrated under reduced pressure at 35°C to remove residual solvent. HPLC-MS: [M+23]: 474. 1 H NMR (400 MHz, DMSO-d6) δ 12.78 (s, 1H), 8.23 ​​(d, J = 7.6 Hz, 1H), 7.86-7.88 (m, 1H), 7.80-7.86 (m, 2H), 7.61-7.80 (m, 2H), 7.55-7.59 (m, 4H), 7.481-7.55 (m, 1H), 7.40-7.48 (m, 3H), 7.27-7.29 (m, 2H), 4.28-4.60 (m, 1H), 4.24-4.28 (m, 1H), 4.17-4.24 (m, 2H), 4.04-4.15 (m, 1H), 1.36 (d, J = 6.8 Hz, 3H).

[0170] Synthesis scheme 18 [ka] L-Tyrosine O-ethylacetamide or (S)-3-(4-(2-acetamidoethoxy)phenyl)-2-aminopropanoic acid Step 1: To a stirred solution of methyl (tert-butoxycarbonyl)-L-tyrosinate (10.0 g, 33.9 mmol), benzyl (2-bromoethyl)carbamate (26.2 g, 102 mmol) and TBAB (5.46 g, 16.93 mmol) in DMF (150 mL) was added potassium carbonate (14.04 g, 102 mmol) at room temperature. The mixture was stirred at 50° C. for 24 hours. The mixture was cooled to room temperature, quenched with water (250 mL) and extracted with EtOAc (2×500 mL). The combined organic layers were washed with brine (3×150 mL), dried over anhydrous sodium sulfate and filtered. The filtrate was concentrated under reduced pressure and the residue was purified by silica gel chromatography eluting with 0% to 30% EtOAc / PE. MS ESI:C 25 H 32 N2O7[M+Na] + Calculated value: 495.22, Measured value: 495.10; 1 H NMR (300 MHz, CDCl3) δ 7.38 - 7.32 (m, 5H), 7.04 (d, J = 8.4 Hz, 2H), 6.81 (d, J = 8.4 Hz, 2H), 5.31 (br, 1H), 5.21 (s, 2H), 4.97 (br, 1H), 4.56 - 4.53 (m, 1H), 4.03 (t, J = 5.0 Hz, 2H), 3.72 (s, 3H), 3.64 - 3.58 (m, 2H), 3.06 - 3.01 (m, 2H), 1.43 (s, 9H).

[0171] Step 2: To a stirred solution of methyl (S)-3-(4-(2-(((benzyloxy)carbonyl)amino)ethoxy)phenyl)-2-((tert-butoxycarbonyl)amino)propanoate (16.0 g, 33.9 mmol) and acetic anhydride (6.39 mL, 67.7 mmol) in THF (200 mL) was added Pd / C (3.60 g, 33.9 mmol, dry, 10 wt%) at room temperature under nitrogen atmosphere. The mixture was degassed with hydrogen three times and stirred at 20° C. for 4 h. DIPEA (17.74 mL, 102 mmol) was added to the mixture and stirred at 20° C. for 1 h. The mixture was filtered and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel chromatography eluting with 0-3% MeOH / DCM. MS ESI:C 19 H 28 N2O6[M+Na] + Calculated value: 403.19, Measured value: 403.10; 1 H NMR (300 MHz, CDCl3) δ 7.05 (d, J = 8.4 Hz, 2H), 6.85 - 6.80 (m, 2H), 5.99 (br, 1H), 5.32 (br, 1H), 4.99 - 4.97 (m, 1H), 4.02 (t, J = 5.0 Hz, 2H), 3.72 (s, 3H), 3.69 - 3.63 (m, 2H), 3.10 - 2.89 (m, 2H), 2.02 (s, 3H), 1.42 (s, 9H).

[0172] Step 3: To a stirred solution of methyl (S)-3-(4-(2-acetamidoethoxy)phenyl)-2-((tert-butoxycarbonyl)amino)propanoate (12.5 g, 32.9 mmol) in THF (100 mL) at room temperature was added lithium hydroxide (65.7 mL, 65.7 mmol, 1N aqueous solution). The solution was stirred at 20° C. for 2 h. The pH of the solution was adjusted to 3 with 1N HCl. The aqueous layer was extracted with EtOAc (2×250 mL). The combined organic layers were washed with brine (150 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure.

[0173] MS ESI [M+H] +:367.10.

[0174] Step 4: To a stirred solution of (S)-3-(4-(2-acetamidoethoxy)phenyl)-2-((tert-butoxycarbonyl)amino)propanoic acid (12.5 g, 30.7 mmol) in THF (20 mL) was added 4N HCl / dioxane (200 mL) at room temperature. The solution was stirred at 20° C. for 1 h. The solvent was concentrated under reduced pressure. MS ESI [M+H] + :267.05.

[0175] Step 5: To a stirred mixture of (S)-3-(4-(2-acetamidoethoxy)phenyl)-2-aminopropanoic acid hydrochloride (9.50 g, 25.1 mmol) and NaHCO3 (10.54 g, 126 mmol) in THF (100 mL) and water (100 mL) was added Fmoc-OSu (7.62 g, 22.59 mmol) at room temperature. The mixture was stirred at 20° C. for 1 h. The pH value of the solution was adjusted to 3 with 1N HCl. The aqueous layer was extracted with EtOAc (2×500 mL). The combined organic layers were washed with brine (150 mL), dried over anhydrous sodium bicarbonate, and filtered. The filtrate was concentrated under reduced pressure, and the residue was recrystallized from EtOAc (200 mL). The solid was collected by filtration and dried in vacuum. MS ESI [M+H] + :489.05; 1 H NMR (300 MHz, methanol-d4) δ 7.79 (d, J = 7.6 Hz, 2H), 7.62-7.57 (m, 2H), 7.42-7.26 (m, 4H), 7.17-7.14 (m, 2H), 6.83 (d, J = 8.4 Hz, 2H), 4.41-4.31 (m, 2H), 4.29-4.10 (m, 2H), 3.96 (t, J = 4.8 Hz, 2H), 3.51 (t, J = 5.4 Hz, 2H), 3.19-3.13 (m, 1H), 2.92-2.84 (m, 1H), 1.94 (s, 3H).

[0176] Synthesis scheme 19 [ka] (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(4-(2-morpholinoethoxy)phenyl)propanoic acid Step 1: To a solution of tert-butyl (tert-butoxycarbonyl)-L-tyrosinate (400 mg, 1.185 mmol) in DMF (4.742 mL) at 0° C. was added cesium carbonate (1 g, 3.08 mmol). The reaction was stirred for 15 min, 2-morpholinoethyl 4-methylbenzenesulfonate (474 ​​mg, 1.660 mmol) was added and stirred at 0° C. for 2.5 h. The reaction was diluted with EtOAc and washed with 10% aqueous LiCl. The organic layer was washed with 1M HCl, saturated aqueous NaHCO3, and aqueous NaCl, then dried over sodium sulfate, filtered, and concentrated. The crude material was purified by column chromatography on an Isco using a RediSep Rf Gold silica 24 g prepacked column eluting with a 0% to 100% EtOAc / hexane gradient over 30 min to give the product. MW: 450.576; MSESI [M+H] + :451.3.

[0177] Step 2: tert-Butyl (S)-2-((tert-butoxycarbonyl)amino)-3-(4-(2-morpholinoethoxy)phenyl)propanoate (525 mg, 1.165 mmol) was dissolved in DCM (3 mL) at ambient temperature. TFA (3 mL) was added and allowed to stir at ambient temperature for 1.5 hours. The reaction was concentrated under reduced pressure and carried forward as crude. MW: 294.351, Found: MSESI [M+H] + :295.1.

[0178] Step 3: (S)-2-Amino-3-(4-(2-morpholinoethoxy)phenyl)propanoic acid (354 mg, 1.203 mmol) was dissolved in tetrahydrofuran (3 mL) and water (3 mL) at ambient temperature. The reaction was cooled to 0° C. Sodium carbonate (382 mg, 3.61 mmol) was added and the solution was stirred at 0° C. for 5 min. (9H-fluoren-9-yl)methyl (2,5-dioxopyrrolidin-1-yl)carbonate (406 mg, 1.203 mmol) was added and stirred for 2 h. THF was removed under reduced pressure. The aqueous solution was acidified to pH 3 with 1N HCl. The reaction was extracted with EtOAc, washed with water, dried over sodium sulfate, filtered and concentrated. The crude material was purified by reverse phase column chromatography using a 50 g C18 pre-packed column on an Isco eluting with a 10% to 100% gradient of TFA-conditioned MeCN / TFA-conditioned water over a 30 min period to give the product in multiple fractions. The fractions were combined, frozen in a -78°C dry ice / acetone bath, and lyophilized to give the product. MW: 516.94, Found: MSESI [M+H] + :517.2.

[0179] Synthesis diagram 20 [ka] (R)-2,4-Diamino-2-methylbutanal Step 1: To a stirred solution of (R)-2-amino-2-methylpent-4-enoic acid (1.3 g, 10.07 mmol) and DIPEA (5.27 mL, 30.2 mmol) in dioxane (20 mL) and water (20 mL) was added Fmoc-OSu (3.73 g, 11.07 mmol) at room temperature. The resulting solution was stirred at 25° C. for 16 h. The pH was adjusted to 3 with 1N HCl and the solution was purified by reverse-phase flash column chromatography with the following conditions: C18 column, 330 g, 5%-5% 5 min, 5% to 50% in 30 min, 98%-98% 5 min, MeCN / water (0.05% TFA) to give (R)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-2-methylpent-4-enoic acid. MS ESI:C 21 H 21NO4Na[M+Na] + Calculated value: 374.15, actual value: 374.05. 1 H NMR (400 MHz, CDCl3) δ 7.79 (d, J = 7.5 Hz, 2H), 7.61 (d, J = 7.5 Hz, 2H), 7.48 - 7.29 (m, 4H), 5.71 (s, 1H), 5.48 (s, 1H), 5.20 - 5.16 (m, 2H), 4.44 - 4.40 (m, 2H), 4.25 (t, J = 6.7 Hz, 1H), 2.82-2.71 (m, 2H).

[0180] Step 2: To a stirred solution of (R)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-2-methylpent-4-enoic acid (3 g, 8.54 mmol) and N-methylmorpholine N-oxide (2.2 g, 9.39 mmol, 50% aqueous solution) in acetone (60 mL) at room temperature was added OsO4 (2.170 g, 0.854 mmol, 10% aqueous solution). The resulting solution was stirred at 25° C. for 4 h. To the solution was added an aqueous solution (20 mL water) of sodium periodate (2.00 g, 9.39 mmol) and the resulting mixture was stirred at 25° C. for 16 h. The reaction mixture was diluted with water (50 mL) and extracted with EtOAc (3×200 mL). The combined organic layers were washed with brine (2×100 mL), dried over anhydrous sodium sulfate, and filtered. The filtrate was then concentrated under reduced pressure to give (R)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-2-methyl-4-oxobutanoic acid. MS ESI:C 20 H 20 NO5[M+H] + Calculated value: 354.13, actual value: 354.00.

[0181] Step 3: To a stirred solution of (R)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-2-methyl-4-oxobutanoic acid (3 g, 6.79 mmol) in toluene (120 mL) at room temperature was added tert-butyl carbamate (4.77 g, 40.8 mmol) and TFA (2.32 g, 20.38 mmol). The resulting solution was stirred at 25° C. for 2 h. The solution was concentrated under reduced pressure to give (R,Z)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-4-((tert-butoxycarbonyl)imino)-2-methylbutanoic acid. MS ESI:C 25 H 27 N2O6[MH] + Calculated value: 451.19, actual value: 450.90.

[0182] Step 4: To a stirred solution of (R,Z)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-4-((tert-butoxycarbonyl)imino)-2-methylbutanoic acid (3.1 g, 5.48 mmol) and dimethyl(phenyl)silane (1.87 g, 13.7 mmol) in toluene (120 mL) at room temperature was added tris(pentafluorophenyl)borane (0.281 g, 0.548 mmol). The solution was stirred at 25° C. for 16 h. The solvent was concentrated under reduced pressure and the residue was purified by reverse-phase flash chromatography using the following conditions: 330 g C18 column, 5%-5% 5 min, 5% to 55% in 30 min, 98%-98% 5 min, MeCN / water (0.05% TFA), RT=35 min to give (R)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-4-((tert-butoxycarbonyl)amino)-2-methylbutanoic acid. MS ESI:C 25 H 31 N2O6[M+H] + Calculated value: 455.21, Measured value: 455.10; 1H NMR (400 MHz, DMSO-d6) δ 12.48 (s, 1H), 7.90 (d, J = 7.2 Hz, 2H), 7.73 (d, J = 7.2 Hz, 2H), 7.54 (s, 1H), 7.44 - 7.41 (m, 2H), 7.36 - 7.32 (m, 2H), 6.78 (s, 1H), 4.25 - 4.23 (m, 3H), 2.92 - 2.91 (m, 2H), 1.99 - 1.93 (m, 1H), 1.87 - 1.81 (m, 1H), 1.37 - 1.34 (m, 9H).

[0183] Synthesis scheme 21 [ka] TIFF0007645434000045.tif34135 [(R)-3-Amino-3-carboxypropyl]-trimethylazanium Step 1: To a mixture of (R)-4-amino-2-((tert-butoxycarbonyl)amino)butanoic acid (7 g, 32.1 mmol) in MeOH (30 mL) at ambient temperature was added CHI (10.03 mL, 160 mmol) and KHCO (16.05 g, 160 mmol). The reaction was warmed to 35° C. for 12 h. The reaction mixture was filtered and the filtrate was concentrated under reduced pressure to give the crude product. The residue was dissolved in ethanol and filtered again. The filtrate was concentrated under reduced pressure to give (R)-3-((tert-butoxycarbonyl)amino)-3-carboxy-N,N,N-trimethylpropan-1-aminium iodide. MS ESI:C 12 H 25 IN2O4[MI] + Calculated value: 261.18, actual value: 261.30.

[0184] Step 2: To a mixture of (R)-3-((tert-butoxycarbonyl)amino)-3-carboxy-N,N,N-trimethylpropan-1-aminium iodide (15 g, 23.18 mmol) in DCM (60 mL) at ambient temperature was added TFA (30 mL, 389 mmol). The reaction was stirred at ambient temperature for 1 h and then concentrated under reduced pressure to give (R)-3-amino-3-carboxy-N,N,N-trimethylpropan-1-aminium 2,2,2-trifluoroacetate. MS ESI: CH 17 F3N2O4[M-CF3COO] + Calculated value: 161.13, actual value: 161.30.

[0185] Step 3: To a mixture of (R)-3-amino-3-carboxy-N,N,N-trimethylpropan-1-aminium 2,2,2-trifluoroacetate (11.5 g, 20.97 mmol) in THF (40 mL) and water (40 mL) was added NaHCO3 (8.81 g, 105 mmol) and Fmoc-OSu (6.37 g, 18.87 mmol). The reaction was stirred at ambient temperature for 2 h, acidified to pH 3-4 with aqueous HCl, and filtered to give the crude product. The crude product was purified by reverse phase column chromatography under the following conditions: 330 g C18 column; mobile phase A: water (0.05% TFA), mobile phase B: MeCN; (gradient: hold 0% B for 5 min, to 33% B in 19 min, hold 38% B for 7 min; to 95% B in 5 min, hold 95% B for 5 min); flow rate: 80 mL / min. Product-containing fractions were collected and lyophilized to give (R)-3-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-carboxy-N,N,N-trimethylpropan-1-aminium 2,2,2-trifluoroacetate. MS ESI: C 24 H 27 F3N2O6[M-CF3COO] + Calculated value: 383.20, actual value: 383.25. 1H NMR (300 MHz, DMSO-d6) δ 7.91 (d, J = 7.2 Hz, 2H), 7.79 - 7.71 (m, 3H), 7.46 - 7.41 (m, 2H), 7.37 - 7.32 (m, 2H), 4.42 - 4.22 (m, 3H), 4.07 - 4.01 (m, 1H), 3.99 - 3.41 (m, 1H), 3.33 - 3.28 (m, 1H), 3.07 (s, 9H), 2.21 - 2.16 (m, 1H), 2.08 - 2.04 (m, 1H). 19 F-NMR (282 MHz, DMSO-d6) δ 73.931.

[0186] Synthesis scheme 22 [ka] 3-[4-[(S)-2-amino-2-carboxyethyl]phenyl]bicyclo[1.1.1]pentane-1-carboxylic acid Step 1: To a stirred mixture of (S)-2-((tert-butoxycarbonyl)amino)-3-(4-iodophenyl)propanoic acid (3.91 g, 10 mmol) in DMF (40 mL) at 0° C. under argon atmosphere was added 3-bromoprop-1-ene (3.63 g, 30.0 mmol) and NaHCO3 (0.840 g, 10.00 mmol). The resulting mixture was stirred at 40° C. for 16 h. The reaction mixture was cooled to room temperature, quenched with water (200 mL), and extracted with EtOAc (2×200 mL). The combined organic layers were washed with brine (3×100 mL), dried over anhydrous sodium sulfate, and filtered. The filtrate was concentrated under reduced pressure, and the residue was purified by silica gel column chromatography (eluted with 0-40% EtOAc / PE), and the product-containing fractions were collected and evaporated under reduced pressure to give allyl (S)-2-((tert-butoxycarbonyl)amino)-3-(4-iodophenyl)propanoate. MS ESI:C 27 H 25 INO4[M+H] + Calculated value: 554.08, actual value: 554.20.

[0187] Step 2: To a stirred solution of 3-(methoxycarbonyl)bicyclo[1.1.1]pentane-1-carboxylic acid (8 g, 47.0 mmol) in DCM (100 mL) at room temperature was added tert-butyl (Z)-N,N'-diisopropylcarbamimidate (37.7 g, 188 mmol). The resulting solution was stirred at 40°C for 2 h. The mixture was then cooled to room temperature. The solids were filtered off and the filtrate was concentrated under reduced pressure to give 1-(tert-butyl)3-methylbicyclo[1.1.1]pentane-1,3-dicarboxylate. 1 H NMR (400 MHz, DMSO-d6) δ 3.61 (s, 3H), 2.17 (s, 6H), 1.39 (s, 9H).

[0188] Step 3: To a stirred solution of 1-(tert-butyl) 3-methylbicyclo[1.1.1]pentane-1,3-dicarboxylate (15 g, 39.8 mmol) in THF (150 mL) was added LiOH (119 mL, 119 mmol, 1N aqueous solution) at room temperature. The resulting solution was stirred at 25° C. for 5 h. The pH of the solution was adjusted to 3 with 1N HCl and extracted with EtOAc (3×100 mL). The combined organic layers were washed with brine (2×50 mL), dried over anhydrous sodium sulfate, and filtered. The filtrate was then concentrated under reduced pressure to give 3-(tert-butoxycarbonyl)bicyclo[1.1.1]pentane-1-carboxylic acid. MS ESI:C 11 H 15 O4[MH] - Calculated value: 211.10, actual value: 211.10.

[0189] Step 4: To a stirred solution of 3-(tert-butoxycarbonyl)bicyclo[1.1.1]pentane-1-carboxylic acid (10 g, 28.3 mmol), 2-hydroxyisoindoline-1,3-dione (6.00 g, 36.7 mmol) and DMAP (0.345 g, 2.83 mmol) in DCM (100 mL) was added N,N′-dicyclohexylcarbodiimide (6.42 g, 31.1 mmol) at room temperature. The resulting mixture was stirred at 25° C. for 16 h. The solids were filtered off and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel chromatography eluting with 0-40% EtOAc / PE to give 1-(tert-butyl)3-(1,3-dioxoisoindolin-2-yl)bicyclo[1.1.1]pentane-1,3-dicarboxylate. 1 H NMR (400 MHz, CDCl3) δ 7.89 (dd, J = 5.5, 3.1 Hz, 2H), 7.79 (dd, J = 5.5, 3.1 Hz, 2H), 2.50 (s, 6H), 1.47 (s, 9H).

[0190] Step 5: To a stirred solution of NiBr2·3H2O (0.791 g, 2.90 mmol) in DMA (80 mL) was added 4,4′-di-tert-butyl-2,2′-dipyridine (0.973 g, 3.63 mmol) at room temperature under a nitrogen atmosphere. The resulting mixture was stirred at 50 °C for 30 min, after which it was cooled to room temperature. 1-(tert-butyl) 3-(1,3-dioxoisoindolin-2-yl)bicyclo[1.1.1]pentane-1,3-dicarboxylate (4.8 g, 12.09 mmol), allyl (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(4-iodophenyl)propanoate (6.69 g, 12.09 mmol), trimethylsilyl chloride (0.131 g, 1.209 mmol), and zinc (3.95 g, 60.4 mmol) were added to the above mixture at room temperature. The resulting mixture was stirred at 25° C. for 2 h. The reaction was quenched with brine (150 mL) and extracted with EtOAc (3×100 mL). The combined organic layers were washed with brine (3×100 mL), dried over anhydrous sodium sulfate, and filtered. The filtrate was then concentrated under reduced pressure and the residue was purified by silica gel chromatography eluting with 0-40% EtOAc / PE to give tert-butyl (S)-3-(4-(2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(allyloxy)-3-oxopropyl)phenyl)bicyclo[1.1.1]pentane-1-carboxylate. MS ESI:C 37 H 40 NO6[M+H] + Calculated value: 594.28, actual value: 594.30.

[0191] Step 6: To a stirred solution of tert-butyl (S)-3-(4-(2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(allyloxy)-3-oxopropyl)phenyl)bicyclo[1.1.1]pentane-1-carboxylate (2.3 g, 1.55 mmol) and phenylsilane (0.335 g, 3.10 mmol) in THF (30 mL) was added Pd(PhP) (0.090 g, 0.077 mmol) at room temperature under a nitrogen atmosphere. The resulting mixture was stirred at room temperature for 1 h, then concentrated under reduced pressure and the residue was purified by reverse-phase column chromatography with the following conditions: C18 column, 330 g, 5%-5% 5 min, 5% to 70% MeCN / water (0.05% TFA) in 40 min to give (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(4-(3-(tert-butoxycarbonyl)bicyclo[1.1.1]pentan-1-yl)phenyl)propanoic acid. MS ESI:C 34 H 34 NO6[MH] - Calculated value: 552.25, actual value: 552.30. 1 H NMR (400 MHz, methanol-d4) δ 7.80 - 7.77 (m, 2H), 7.67 - 7.65 (m, 2H), 7.48 - 7.23 (m, 4H), 7.19 - 7.17 (m, 2H), 7.10 - 7.07 (m, 2H), 4.45 - 4.44 (m, 1H), 4.42 - 4.41 (m, 1H), 4.34 - 4.29 (m, 2H), 3.24 - 3.18 (m, 1H), 2.94 - 2.86 (m, 1H), 2.12 (s, 6H), 1.45 (s, 9H).

[0192] Composite diagram 23 [ka] TIFF0007645434000048.tif35137 (1R,4s)-4-(4-((S)-2-amino-2-carboxyethyl)phenyl)cyclohexane-1-carboxylic acid Step 1: To a mixture of tert-butyl 4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)cyclohex-3-ene-1-carboxylate (29.5 g, 96 mmol) in THF (30 mL) at room temperature was added methyl (S)-2-((tert-butoxycarbonyl)amino)-3-(4-iodophenyl)propanoate (15.5 g, 38.2 mmol) and Pd(Ph3P)4 (2.21 g, 1.91 mmol). The reaction was warmed to 60° C. for 4 h. The resulting solution was quenched with water (100 mL) and extracted with EtOAc (3×300 mL). The organic layers were combined, washed twice with brine (200 mL), dried over anhydrous sodium sulfate, and filtered. The filtrate was concentrated under reduced pressure to give the crude product. The residue was purified by silica gel chromatography eluting with a gradient of ethyl acetate / petroleum ether (0% to 25%) to give tert-butyl 4'-((S)-2-((tert-butoxycarbonyl)amino)-3-methoxy-3-oxopropyl)-2,3,4,5-tetrahydro-[1,1'-biphenyl]-4-carboxylate. MS ESI:C 26 H 38 NO6Na[M+Na] + Calculated value: 482.26, actual value: 482.10.

[0193] Step 2: To a mixture of tert-butyl 4'-((S)-2-((tert-butoxycarbonyl)imino)-3-methoxy-3-oxopropyl)-2,3,4,5-tetrahydro-[1,1'-biphenyl]-4-carboxylate (16 g, 34.8 mmol) in methanol (160 mL) was added Pd-C (10% on carbon, approx. 55% water wet, 5.3 g, 4.98 mmol) at room temperature. The solution was degassed with H2 three times and stirred at room temperature under H2 atmosphere (1.5 atm) for 1 h. The resulting solution was filtered and the filtrate was concentrated under reduced pressure to give tert-butyl (S)-4-(4-(2-((tert-butoxycarbonyl)amino)-3-methoxy-3-oxopropyl)phenyl)cyclohexane-1-carboxylate. MS ESI: C 26 H 41 NO6[M+H] +Calculated value: 462.28, actual value: 462.30.

[0194] Step 3: To a stirred solution of tert-butyl (S)-4-(4-(2-((tert-butoxycarbonyl)amino)-3-methoxy-3-oxopropyl)phenyl)cyclohexane-1-carboxylate (15 g, 32.5 mmol) in THF (300 mL) was added LiOH (65.0 mL, 65.0 mmol) at room temperature. The solution was stirred at 20° C. for 1 h. The pH of the solution was adjusted to 3 with 1N HCl. The reaction was concentrated under reduced pressure to give (S)-2-((tert-butoxycarbonyl)amino)-3-(4-(4-(tert-butoxycarbonyl)cyclohexyl)phenyl)propanoic acid. MS ESI: C 25 H 38 NO6Na[M+Na] + Calculated value: 470.26, actual value: 470.30.

[0195] Step 4: (S)-2-((tert-butoxycarbonyl)amino)-3-(4-(4-(tert-butoxycarbonyl)cyclohexyl)phenyl)propanoic acid (14 g, 31.3 mmol) is separated using preparative supercritical fluid chromatography under the following conditions: Column: CHIRAL ART Cellulose-SB, 3×25 cm, 5 μm; Mobile phase A: CO2, Mobile phase B: MeOH (0.1% 2M NH3-MeOH); Flow rate: 80 mL / min; Gradient: 10% B; 220 nm; Peak 1 retention time: 7.45; Peak 2 retention time: 8.38 to give the first eluting peak (S)-2-((tert-butoxycarbonyl)amino)-3-(4-((1s,4R)-4-(tert-butoxycarbonyl)cyclohexyl)phenyl)propanoic acid. MS ESI: C 25 H 38 NO6Na[M+Na] +Calculated for: 470.26, found 470.30. 1H NMR (300 MHz, CDCl3) δ 7.11 (d, J = 3.0 Hz, 4H), 4.27 (s, 1H), 3.18 - 3.12 (m, 1H), 2.91 (s, 1H), 2.60 (s, 1H), 2.49 (s, 1H), 2.20 (d, J = 10.9 Hz, 2H), 1.78 - 1.52 (m, 6H), 1.49 (d, J = 0.7 Hz, 9H), 1.36 (s, 9H); And the second eluting peak (S)-2-((tert-butoxycarbonyl)amino)-3-(4-((1r,4S)-4-(tert-butoxycarbonyl)cyclohexyl)phenyl)propanoic acid. MS ESI:C 25 H 38 NO6Na[M+Na] + Calculated value: 470.26, actual value: 470.30. 1H NMR (300 MHz, CDCl3) δ 7.14 - 7.06 (m, 4H), 4.25 (s, 1H), 3.14 (d, J = 12.3 Hz, 1H), 2.88 (s, 1H), 2.49 - 2.41 (m, 1H), 2.29 - 2.15 (m, 1H), 2.05 (d, J = 12.2 Hz, 2H), 1.90 (d, J = 12.1 Hz, 2H), 1.69 - 1.47 (m, 2H), 1.47 (s, 11H), 1.34 (s, 9H). obtained.

[0196] Step 5: To a solution of (S)-2-((tert-butoxycarbonyl)amino)-3-(4-((1s,4R)-4-(tert-butoxycarbonyl)cyclohexyl)phenyl)propanoic acid (8.3 g, 18.54 mmol) in THF (80 mL) at room temperature was added HCl (9.27 mL, 18.54 mmol) in small portions. The reaction was concentrated under reduced pressure to give (S)-2-amino-3-(4-((1s,4R)-4-(tert-butoxycarbonyl)cyclohexyl)phenyl)propanoic acid. MS ESI: C 20 H 30 NO4[M+H]+ Calculated value: 348.21, actual value: 348.25.

[0197] Step 6: To a stirred solution of (S)-2-amino-3-(4-((1s,4R)-4-(tert-butoxycarbonyl)cyclohexyl)phenyl)propanoic acid (6 g, 17.27 mmol) and NaHCO3 (7.25 g, 86 mmol) in THF (60 mL) and water (60.0 mL) was added Fmoc-OSu (5.24 g, 15.54 mmol). The mixture was stirred at room temperature for 1 h. The pH of the solution was adjusted to 3 with 1N HCl. The aqueous layer was extracted with EtOAc (2 x 200 mL). The combined organic layers were washed with brine (150 mL), dried over anhydrous sodium sulfate, and filtered. The filtrate was concentrated under reduced pressure and the residue was purified by reverse phase column chromatography to give (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(4-((1s,4R)-4-(tert-butoxycarbonyl)cyclohexyl)phenyl)propanoic acid. MS ESI:C 35 H 41 NO6[M+H] + Calculated value: 570.28, actual value: 570.15. 1 H NMR (300 MHz, methanol-d4) δ 7.81 (d, J = 7.5 Hz, 2H), 7.61 (d, J = 7.5 Hz, 2H), 7.43 - 7.27 (m, 4H), 7.16 (d, J = 7.8 Hz, 2H), 7.07 (d, J = 7.9 Hz, 2H), 4.45-4.33 (m, 2H), 4.22 - 4.06 (m, 2H), 3.26 - 3.14 (m, 1H), 2.96 - 2.88 (m, 1H), 2.59 (s, 1H), 2.48 (s, 1H), 2.15 (s, 2H), 1.61 (d, J = 5.5 Hz, 6H), 1.49 (s, 9H).

[0198] Synthesis scheme 24 [ka] (1S,4r)-4-(4-((S)-2-amino-2-carboxyethyl)phenyl)cyclohexane-1-carboxylic acid Step 1: To a mixture of (S)-2-((tert-butoxycarbonyl)amino)-3-(4-((1r,4S)-4-(tert-butoxycarbonyl)cyclohexyl)phenyl)propanoic acid (3 g, 6.70 mmol) in THF (30 mL) was added hydrogen chloride (30 mL, 60.0 mmol) in small portions at room temperature. The reaction was concentrated under reduced pressure to give (S)-2-amino-3-(4-((1r,4S)-4-(tert-butoxycarbonyl)cyclohexyl)phenyl)propanoic acid. MS ESI:C 20 H 31 NO4[M+H] + Calculated value: 348.21, actual value: 348.25.

[0199] Step 2: To a stirred solution of (S)-2-amino-3-(4-((1r,4S)-4-(tert-butoxycarbonyl)cyclohexyl)phenyl)propanoic acid (2 g, 5.76 mmol) (1.748 g, 5.18 mmol) and NaHCO3 (2.418 g, 28.8 mmol) in THF (20 mL) and water (20 mL) was added Fmoc-OSu (1.748 g, 5.18 mmol) at room temperature. The mixture was stirred at 20° C. for 1 h. The pH of the solution was adjusted to 3 with 1N HCl. The aqueous layer was extracted with EtOAc (2×200 mL). The combined organic layers were washed with brine (3×50 mL), dried over anhydrous sodium sulfate, and filtered. The filtrate was concentrated under reduced pressure and the residue was purified by reverse phase column chromatography to give (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(4-((1r,4S)-4-(tert-butoxycarbonyl)cyclohexyl)phenyl)propanoic acid. MS ESI:C 35 H 41 NO6[M+H] + Calculated value: 570.28, actual value: 570.35. 1H NMR (300 MHz, methanol-d4) δ 7.81 (d, J = 7.6 Hz, 2H), 7.61 (d, J = 7.5 Hz, 2H), 7.43 - 7.38 (m, 2H), 7.34 - 7.27 (m, 2H), 7.17 (d, J = 8.0 Hz, 2H), 7.08 (d, J = 7.8 Hz, 2H), 4.47 - 4.43 (m, 1H), 4.35 - 4.30 (m, 1H), 4.21 - 4.02 (m, 2H), 3.24 - 3.18 (m, 1H), 2.94 - 2.87 (m, 1H), 2.40 (s, 1H), 2.17 (d, J = 11.9 Hz, 1H), 1.96 (s, 3H), 1.82 - 1.55 (m, 2H), 1.47 (s, 13H).

[0200] Preparation of final compound: A. Generalized Procedure for the Synthesis of Linear Peptide Precursors The peptides in Table 1 were synthesized using standard solid phase synthesis using Fmoc / tBu chemistry as exemplified in Chan, WC;White, PD “Fmoc Solid-Phase Synthesis: a Practical Approach”, Oxford University Press, Oxford, 2000;Steward, J.;Young, J. “Solid Phase Peptide Synthesis”, Pierce Chemical Company, Rockford, 1984.;Benoiton, NL “Chemistry of Peptide Synthesis”, CRC Press, New York, 2006; and Lloyd-Williams, P.;Albericio, F.;Giralt, E. “Chemical Approaches to the Synthesis of Peptides and Proteins”, CRC Press, New York, 1997.

[0201] During peptide chain elongation, the α-amino group of each amino acid was protected with a 9H-fluoren-9-ylmethoxycarbonyl group (Fmoc). To avoid side reactions during the chain elongation step, the reactive amino acid side chains also carried acid-labile protecting groups, which effectively mask the reactive groups until they were removed by treatment with strong acid. After completion of each coupling step, the Fmoc group of the N-terminal amino acid was removed with piperidine or 4-methylpiperidine, and the resin was washed extensively in preparation for the subsequent coupling of the Fmoc-protected amino acid derivative.

[0202] The side chain protecting groups used were tert-butyl (tBu) for S, hY, Bip4CO2H, Phe4COOH, F4pcCCA, F4ptCCA, F4bcpA; tert-butoxycarbonyl (Boc) for Dap, dDab, dK, dOrn, K, Prot4NH2, dDap, daMeDab; and β-methylpentyl ester (OMpe) for D.

[0203] Fmoc-protected amino acids were typically obtained from suppliers such as Sigma-Aldrich, Novabiochem, Chem-Impex, and Combi-Block.

[0204] B. Synthetic Procedures Used to Prepare Cyclic Peptides Composite diagram 25 [ka]

[0205] Solid Phase Peptide Synthesis Protocol A The peptides were synthesized on a Liberty Blue™ synthesizer from CEM using standard solid phase synthesis with Fmoc / tBu chemistry as outlined above in Scheme 25. N,N'-diisopropylcarbodiimide (DIC) and ethyl cyano(hydroxyimino)acetate (Oxyma) were used as coupling agents to form the amide bond between the free amino terminus of the resin-bound protected peptide and the carboxylic acid of the Fmoc-protected amino acid.

[0206] H-Gly loaded 2-chlorotrityl resin (200-400 mesh, 0.79 mmol / g loading, 1% cross-linked polystyrene, Novabiochem) was used for the synthesis. All amino acids were dissolved in DMF (N,N-dimethylformamide) at 0.2 M concentration. The amino acids were activated with an equimolar amount of Oxyma solution (0.5 M in DMF) and a 2-fold molar excess of DIC solution (1.0 M in DMF). Alternatively, the amino acids were dissolved in DMF (N,N-dimethylformamide) at 0.125 M concentration. The amino acids were activated with an equimolar amount of Oxyma Pure solution (0.125 M in DMF; containing 0.05 M DIEA) and a 2-fold molar excess of DIC solution (0.25 M in DMF). Reactions were typically performed on a 25 μmol scale.

[0207] Each synthesis cycle included deprotection of Fmoc amino acid with 20% piperidine in DMF (microwave heating at 90 °C, 2 min) and coupling with Fmoc-protected amino acid / DIC / Oxyma (5, 5, 10 equiv., microwave heating at 90 °C, 2 min or 4 min, respectively), which could be repeated twice for difficult couplings. Cycles of Fmoc-deprotection and coupling of Fmoc-protected amino acid were repeated for the desired monomers until the full-length linear peptide was formed.

[0208] Solid Phase Peptide Synthesis Protocol B Alternatively, peptides were synthesized on a Biotage® Syro II peptide synthesizer using standard solid phase synthesis using Fmoc / tBu chemistry as outlined above in Scheme 25. HATU and DIPEA were used as coupling agents to create an amide bond between the free amino terminus of the resin-bound protected peptide and the carboxylic acid of the Fmoc-protected amino acid. 2-Chlorotrityl resin loaded with H-Gly (200-400 mesh, 0.79 mmol / g loading, 1% cross-linked polystyrene, Novabiochem) was used for the synthesis. All amino acids were dissolved at 0.2 M concentration in 1:1 DMF:NMP. Reactions were typically performed on a 12 μmol scale.

[0209] Each synthesis cycle included: (1) coupling with Fmoc-protected amino acid / HATU / DIPEA (4, 4 and 8 equivalents, respectively; room temperature; 15 min) (repeated twice); the mixture was filtered and the peptidyl resin was washed with DMF (2 times with 1 mL); (2) Fmoc deprotection (repeated three times): 20% 4-methylpiperidine in DMF (1 mL; room temperature; 3 min); the mixture was filtered and the peptidyl resin was washed with DMF (4 times with 1 mL). Cycles of Fmoc deprotection and Fmoc-protected amino acid coupling were repeated with the desired monomers until the full-length linear peptide was formed.

[0210] Selective cleavage and macrolactamization of protected peptides. For cleavage of the protected linear peptide from the solid support, the peptidyl resin (approximately 16 mg) was treated with a solution of 25% hexafluoroisopropanol (HFIP) in DCM for 20 min at room temperature, filtered, and the solvent was removed under reduced pressure. The resulting residue was dissolved in DMF (5 mL). HATU (0.44 eq.) and DIPEA (2.5 eq.) were added. The mixture was stirred at room temperature for 5 min. Then, an additional 0.66 eq. of HATU was added. Upon completion of the macrolactamization, monitored by UPLC-MS, the solvent was removed under reduced pressure.

[0211] Final side chain deprotection A solution of TFA / HO / TIS (90 / 8 / 2, volume ratio, 1 mL) was added to the crude protected cyclic peptide. The mixture was stirred at room temperature for 10 min. Cold diethyl ether (15 mL) was added to the solution. The peptide was precipitated by centrifugation (3200 rpm, -10°C). The precipitate was washed with diethyl ether (2 x 10 mL) and dried in vacuum overnight to give the crude deprotected cyclic peptide as a solid.

[0212] HPLC purification Purification was performed by preparative reversed-phase high performance liquid chromatography (RP-HPLC) on a Waters X-Bridge Prep C18 OBD Prep column (130 Å, 5 pm, column size 19×100 mm) using a Waters MS-Directed AutoPurification HPLC / MS system. Mobile phase: (A) 0.16% TFA / HPLC water and (B) 0.16% TFA / HPLC acetonitrile; flow rate: 25 mL / min; UV wavelength λ=215 nm; gradient: 25% to 50% B over 5 min. Alternatively, purification was performed on a Waters CSH-C18 column (19×250 mm, 5 μM) using an Agilent 1290 infinity II preparative LC system and LC-MSD XT mass spectrometer. Mobile phase: (A) 0.1% formic acid / HPLC water and (B) 0.1% formic acid / HPLC acetonitrile; flow rate: 25 mL / min; UV wavelength λ=215 nm; gradient: 20% B 2.5 min, 55% B 2.5 to 20 min. UV absorbing fractions containing the m / z ion of interest were collected and fractions containing the product were confirmed by LC / MS.

[0213] The identity and purity of the final compound was determined by UPLC-MS measured on a reversed-phase Waters Acquity UPLC-MS system. Column: Waters XSelect CSH C18 column (130 Å, 2.5 μm, column size 2.1×50 mm). Mobile phase: (A) 0.05% TFA / HPLC water and (B) 0.05% TFA / HPLC acetonitrile; injection volume: 1 μL; flow rate: 1 mL / min; UV wavelength λ=215 nm; gradient: 5% to 100% B in 5 min. The combined fractions containing the pure peptide were lyophilized to give the final cyclized product as a powder.

[0214] Biological Assays: Procedure for IL-6 assay in MRC5 cells Inhibition of IL-1β-induced IL-6 secretion was assessed in MRC5 cells. 80Recombinant human IL-1β (BioLegend 579404) at 100% concentration was prepared in seeding medium, EMEM (ATCC 30-2003), containing 0.025% BSA (Sigma A9576), 1x penicillin / streptomycin (Gibco 15070-063), 1x NEAA (Gibco 11140-050), 1x GlutaMax (Gibco 35050-061), and 1x sodium pyruvate (Gibco 11360-070). 20 μL of IL-1β was added to a 384-well collagen-coated plate (Corning 354664) and preincubated for 1 h at ambient temperature with 200 nL of compound dispensed using an ECHO555 liquid handler. Human lung fibroblast MRC5 cells (ATCC CCL-171) were added at a density of 3000 cells / 20 μL / well. The cells were prepared by passage three times in growth medium EMEM (ATCC 30-2003) containing 10% fetal bovine serum (Gibco 16140-071), 1× penicillin / streptomycin (Gibco 15070-063), 1× NEAA (Gibco 11140-050), 1× GlutaMax (Gibco 35050-061), and 1× sodium pyruvate (Gibco 11360-070) in collagen-coated T175 flasks (Greiner 661950) and harvested in seeding medium after 5 min of digestion with 0.25% trypsin-EDTA (Gibco 25200-056). 384-well collagen-coated plates with a final volume of 40 μL were incubated overnight at 37° C., 5% CO2. 5 μL of conditioned medium was transferred to a 384-well AlphaLISA plate (PerkinElmer 6005350) for detection of IL-6 using the Human AlphaLISA IL-6 Kit (PerkinElmer AL223F) according to the manufacturer's protocol. 20 μL of the acceptor bead / biotin-labeled antibody mix was added to the 384-well AlphaLISA plate and incubated for 1 hour at ambient temperature. The donor bead mix was protected from light and 25 μL was added to the plate and incubated for 30 minutes at ambient temperature.AlphaLISA plates were read on an EnVision multimode plate reader (PerkinElmer model 2104) using AlphaScreen settings (laser excitation at 680 nm and emission at 570 nm). Dose-response curves and IC were calculated using a four-parameter logistic equation in Spotfire software (Tibco, Palo Alto, CA). 50 The values ​​were analyzed.

[0215] Amino acid sequences of Example Nos. 1 to 215 (SEQ ID NOs. 1 to 215), biological activity (MRC IC 50 ), the calculated monoisotopic mass, molecular formula, calculated molecular weight and mass spectral data ([M+H]+ or [M+2H] / 2+) are provided in Table 1 below.

[0216] Table 1 [Table 2] TIFF0007645434000052.tif255163TIFF0007645434000053.tif255163TIFF000 7645434000054.tif255163TIFF0007645434000055.tif255164TIFF0007645434 000056.tif255164TIFF0007645434000057.tif255163TIFF0007645434000058. tif255165TIFF0007645434000059.tif255166TIFF0007645434000060.tif36170

Claims

1. A compound selected from the group consisting of the following (SEQ ID NOs: 78, 71-72, 67, 65, 70, 69, 68, 66, 64, 77, 79, 209, 193, 215, 212, and 210, respectively, in order of appearance), or a pharma- ceutically acceptable salt thereof. 【Chemistry 1】 【change】 【change】 【change】

2. 2. The compound of claim 1, wherein the compound is SEQ ID NO: 78, or a pharma- ceutically acceptable salt thereof. 【Chemistry 2】

3. 2. The compound of claim 1, wherein the compound is SEQ ID NO: 71, or a pharma- ceutically acceptable salt thereof. 【Chemistry 3】

4. 2. The compound of claim 1, wherein the compound is SEQ ID NO: 72, or a pharma- ceutically acceptable salt thereof. 【Chemistry 4】

5. 2. The compound of claim 1, wherein the compound is SEQ ID NO: 67, or a pharma- ceutically acceptable salt thereof. 【Chemistry 5】

6. 2. The compound of claim 1, wherein the compound is SEQ ID NO: 79, or a pharma- ceutically acceptable salt thereof. 【Chemistry 6】

7. 2. The compound of claim 1, wherein the compound is SEQ ID NO: 209, or a pharma- ceutically acceptable salt thereof. 【Chemistry 7】

8. A pharmaceutical composition comprising a compound according to any one of claims 1 to 7, or a pharma- ceutically acceptable salt thereof, and a pharma- ceutically acceptable carrier.

9. A pharmaceutical composition for the treatment of atherosclerosis in a subject in need thereof, comprising an effective amount of a compound according to any one of claims 1 to 7 or a pharma- ceutically acceptable salt thereof.

10. A pharmaceutical composition for the treatment of vascular inflammation in a subject in need thereof, comprising a compound according to any one of claims 1 to 7 or a pharma- ceutically acceptable salt thereof.

11. A pharmaceutical composition for the treatment of an inflammatory disorder in a subject in need thereof, comprising a compound according to any one of claims 1 to 7 or a pharma- ceutically acceptable salt thereof.

12. 10. The pharmaceutical composition of claim 9, wherein the effective amount of the compound or a pharma- ceutically acceptable salt thereof is orally administered to a subject.

13. The method of claim 10, wherein the effective amount of the compound or a pharma- ceutically acceptable salt thereof is orally administered to a subject.

14. The method of claim 11 , wherein the effective amount of the compound or a pharma- ceutically acceptable salt thereof is orally administered to a subject.

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