Pyrrolidine compounds having Lp(a)-reducing activity

Abstract

This disclosure provides compounds of formula: and pharmaceutically acceptable salts thereof, as well as pharmaceutical compositions comprising these compounds, and their use in the treatment of cardiovascular disease and elevated Lp(a) plasma levels. [Formula 1] TIFF2026525345000055.tif58128

Description

[Technical Field]

[0001] The present invention relates to pyrrolidine compounds, pharmaceutically acceptable salts thereof, pharmaceutical compositions, and the therapeutic use of these compounds, particularly their use in reducing lipoprotein(a) (Lp(a)) plasma levels. [Background technology]

[0002] There have been remarkable advances in the treatment of cardiovascular disease (CVD). Despite these advances, patients continue to experience cardiovascular events such as angina, myocardial infarction, and stroke, which can be fatal if left untreated. Dyslipidemia remains a major risk factor for CVD. Dyslipidemia can be divided into four common risk factors: elevated low-density lipoprotein cholesterol (LDL-c), decreased high-density lipoprotein cholesterol (HDL-c), elevated triglycerides (TG), and elevated Lp(a). There are various treatment regimens targeting high LDL-c, low HDL-c, and high triglycerides. There are few approved treatment options for patients with high Lp(a) levels. In some cases, apheresis can be used to filter the blood and remove LDL and Lp(a), however, its effect is temporary and typically needs to be repeated every two weeks. Currently, there are no approved drug treatments specifically for lowering Lp(a) levels.

[0003] Lp(a) is a genetically determined, independent cardiovascular risk factor. Elevated serum Lp(a) levels above 50 mg / dL or 125 nmol / L, found in approximately 20% of the population, are associated with at least a 1.6-fold risk of the first cardiovascular event (Kronenberg, F. Clin. Res. Cardiol. Suppl. 14, 5-12 (2019)) and have been shown to increase the risk of a second event by more than 1.42 times (Madsen, CM et al. Arterioscler. Thromb. Vasc. Biol. 40, 255-266 (2020)). Lp(a) can exhibit both prothrombinogenic and antithrombotic properties, as well as both atherogenic and atherothrombotic properties. Lp(a) inhibits fibrinolysis and can accumulate in the vascular wall, inducing thrombus formation and atherosclerotic lesions. Plasma levels of Lp(a) vary substantially among individuals. Unlike other risk factors, Lp(a) plasma levels do not change significantly with diet and exercise.

[0004] Lp(a) is similar to LDL-c in that it contains an LDL lipid core with an associated apolipoprotein B (apoB), but unlike LDL-c, Lp(a) contains its own apolipoprotein (a) (apo(a)) bound to apoB via a disulfide bond. apo(a) is synthesized in the liver. Aggregations of apo(a) and Lp(a) from LDL particles can occur in hepatocytes, on the cell wall, or in plasma. Inhibition of LDL particle aggregation with apo(a) lowers Lp(a) levels.

[0005] International Publication No. 2020 / 247429 discloses compounds that inhibit Lp(a) formation by blocking the interaction between Apo(a) and ApoB. [Overview of the project] [Problems that the invention aims to solve]

[0006] Patients with cardiovascular disease, particularly those with dyslipidemia or dyslipidemia, require additional treatment options. Additional treatment options are needed for patients whose cardiovascular risk is not adequately managed with current standard treatments, including diet, exercise, and / or the use of one or more medications such as statins, fibrates, and niacin. More specifically, there is a need for further alternative, pharmaceutically acceptable compounds that inhibit Lp(a) formation and thus lower plasma Lp(a) levels. [Means for solving the problem]

[0007] Therefore, in the first embodiment, Equation I:

[0008] [ka] (In the formula, L is a C2-C6 alkylene, a C2-C6 alkenylene, or -(CH2) n NHCONH(CH2) n -and, n is independently between 2 and 4 in each occurrence. In each occurrence, Y is independently CH2, O, or S. Z is independently H, C1-C4 alkyl, OH, or cyclopropyl in each instance. Compounds thereof, or pharmaceutically acceptable salts thereof, are provided.

[0009] In the second aspect, formula II:

[0010] [ka] (In the formula, L is a C2-C3 alkylene or a C2-C3 alkenylene. R 1 is -NHC(O)R 2 ,-NHC(O)NHR 2 COOH, -C(O)NH-cyclopropyl,

[0011]

Chem.

[0012] In a third aspect, a pharmaceutical composition is provided comprising a compound of formula I or II, or a pharmaceutically acceptable salt thereof, and at least one pharmaceutically acceptable carrier, diluent, or excipient.

[0013] In a fourth aspect, a method of treating a cardiovascular disease in a patient is provided comprising administering to the patient an effective amount of a compound of formula I or II, or a pharmaceutically acceptable salt thereof.

[0014] In a fifth aspect, a method of treating an elevated Lp(a) plasma level in a patient is provided comprising administering to the patient an effective amount of a compound of formula I or II, or a pharmaceutically acceptable salt thereof.

[0015] In a sixth aspect, a compound of formula I or II, or a pharmaceutically acceptable salt thereof, is provided for use in therapy.

[0016] In a seventh aspect, a compound of formula I or II, or a pharmaceutically acceptable salt thereof, is provided for use in the treatment of cardiovascular disease.

[0017] In the eighth aspect, compounds of formula I or II, or pharmaceutically acceptable salts thereof, are provided for use in the treatment of elevated Lp(a) plasma levels. [Modes for carrying out the invention]

[0018] In one embodiment of the compound of formula I, Z is H in each occurrence.

[0019] In one embodiment of the compound of formula I, Y is CH2 in each occurrence.

[0020] In one embodiment, formula Ia:

[0021] [ka] Compounds of or pharmaceutically acceptable salts thereof are provided. Formula I encompasses Formula Ia, and any following references to Formula I in therapeutic methods and therapeutic uses should also be interpreted as references to Formula Ia.

[0022] In one embodiment of the compound of formula I, L is a C2 alkylene. In another embodiment of the compound of formula I, L is a C2 alkenylene. In yet another embodiment of the compound of formula I, L is -(CH2)3NHCONH(CH2)3-.

[0023] In some embodiments, the compound of formula I is

[0024] [ka] Select from, or a pharmaceutically acceptable salt thereof.

[0025] In one embodiment of the compound of formula II, Z is H.

[0026] In one embodiment of the compound of formula II, Y is CH2.

[0027] In one embodiment, a compound of formula IIa:

[0028]

Chemical formula

[0029] In one embodiment of the compound of formula II, R[[ID=1十七]] 1 is -NHC(O)CH3, -NHC(O)NHCH2CH3, COOH, -C(O)NH-cyclopropyl,

[0030]

Chemical formula

[0031] In one embodiment, the compound of formula II is

[0032]

Chemical formula

[0033] In one embodiment, a method of treating a patient who requires treatment for cardiovascular disease is provided, comprising administering an effective amount of a compound of formula I or II, or a pharmaceutically acceptable salt thereof. In one embodiment, a method of treating a patient who requires treatment for an elevation in Lp(a) plasma levels is provided, comprising administering an effective amount of a compound of formula I or II, or a pharmaceutically acceptable salt thereof.

[0034] In one embodiment, a compound of formula I or II, or a pharmaceutically acceptable salt thereof, is provided for use in therapy.

[0035] In one embodiment, a compound of formula I or II, or a pharmaceutically acceptable salt thereof, is provided for use in the treatment of cardiovascular disease. In another embodiment, a compound of formula I or II, or a pharmaceutically acceptable salt thereof, is provided for use in the treatment of elevated Lp(a) plasma levels.

[0036] In one embodiment, the use of a compound of formula I or II, or a pharmaceutically acceptable salt thereof, in the manufacture of a pharmaceutical for the treatment of cardiovascular disease is provided. In another embodiment, the use of a compound of formula I or II, or a pharmaceutically acceptable salt thereof, in the manufacture of a pharmaceutical for the treatment of elevated Lp(a) plasma levels is provided.

[0037] "C 1~ C n The term "alkyl" refers to a straight-chain or branched-chain saturated hydrocarbon containing 1 to n carbon atoms. 1~ Examples of C4 alkyl groups include, but are not limited to, methyl, ethyl, propyl, butyl, and tert-butyl. 1~ Examples of C3 alkyl groups include, but are not limited to, methyl, ethyl, and propyl.

[0038] "C 2~ The term "C6 alkylene" refers to a straight-chain or branched-chain saturated hydrocarbon containing 2 to 6 carbon atoms. 2~ The term "C3 alkylene" refers to a straight-chain or branched-chain saturated hydrocarbon containing two to three carbon atoms.

[0039] "C 2~ The term "C6 alkenylene" refers to a straight-chain or branched-chain hydrocarbon containing 2 to 6 carbon atoms and at least one double bond. 2~ The term "C3 alkenylene" refers to a straight-chain or branched-chain hydrocarbon containing two to three carbon atoms and at least one double bond.

[0040] As used herein, the term “elevated Lp(a) plasma levels” means Lp(a) plasma levels of approximately 50 mg / dL or higher. The compounds provided herein can be used for therapeutic purposes to lower Lp(a) plasma levels.

[0041] As used herein, the term “pharmaceutically acceptable salt” refers to a salt of a compound that is acceptable for clinical and / or veterinary use. Examples of pharmaceutically acceptable salts and general methodologies for preparing them can be found in “Handbook of Pharmaceutical Salts: Properties, Selection and Use” P. Stahl, et al., 2nd Revised Edition, Wiley-VCH, 2011 and SMBerge, et al., “Pharmaceutical Salts,” Journal of Pharmaceutical Sciences, 1977, 66(1), 1-19. In particular, the compound of formula I or II may be a zwitterionic, monoacid, diacid, or triacid addition salt.

[0042] The pharmaceutical compositions of the present invention may be prepared using pharmaceutically acceptable additives. The term "pharmaceutically acceptable" means one or more carriers, diluents, and / or excipients that are compatible with the other components of the composition and are not pharmaceutically harmful to the patient. Examples of pharmaceutical compositions and processes for their preparation are well known to those skilled in the art and can be found, for example, in "Remington: The Science and Practice of Pharmacy," Loyd, V., et al. Eds., 22nd Ed., Mack Publishing Co., 2012.

[0043] As used herein, the term “effective dose” refers to the dosage that is effective in treating a disorder. The effective dose for a particular patient can be determined by a skilled healthcare professional.

[0044] As used herein, the terms “to treat,” “to cure,” or “to treat” include slowing, reducing, preventing, or reversing the progression or severity of an existing symptom, disorder, condition, or disease. As used herein, “to treat a cardiovascular disease” means slowing, reducing, preventing, or reversing the progression of a disease of the heart or blood vessels.

[0045] As used herein, the term “patient” refers to a mammal; preferably, the patient is a human.

[0046] The pharmaceutical composition can be formulated as tablets or capsules for oral administration, as a solution for oral administration, or as an injectable solution. In one embodiment, the composition is suitable for oral administration.

[0047] The present invention aims to provide all individual enantiomers, diastereomers, mixtures thereof, and racemates of the compounds of formulas I and II, as well as pharmaceutically acceptable salts thereof.

[0048] Certain abbreviations may refer to the following: "ACN" refers to acetonitrile, "Apo" refers to apolipoprotein, "BOC" refers to tert-butoxycarbonyl, "DCM" refers to dichloromethane, "DMA" refers to dimethylacetamide, "DMEM" refers to Dulbecco's Modified Eagle's Medium, "DMF" refers to dimethylformamide, "DMSO" refers to dimethyl sulfoxide, "EACA" refers to epsilon-aminocaproic acid or 6-aminocaproic acid, "ELISA" refers to enzyme-linked immunosorbent assay, "Â" refers to ethyl acetate, and "FBS" refers to fetal bovine serum. "HEC" refers to hydroxyethyl cellulose, "HEK" refers to human embryonic kidney, "HepG2" refers to human hepatocellular carcinoma cell line, "HEPES" refers to 4-(2-hydroxyethyl)-1-piperazine ethanesulfonic acid, "HLB" refers to hydrophilic lipophilic balance, "hr / s" refers to hour / hours, "HRP" refers to horseradish peroxidase, and "IC 50" refers to the concentration of the drug that produces 50% of the maximum possible inhibitory response for that drug, "MeCN" refers to acetonitrile, "min" refers to minutes (minutes / s), "MTBE" refers to methyl tert-butyl ether, "RT" refers to room temperature, "TEA" refers to triethylamine, "TFA" refers to trifluoroacetic acid, "THF" refers to tetrahydrofuran, and "TMB" refers to 3,3',5,5'-teramethylbenzidine.

[0049] Individual isomers, enantiomers, and diastereomers may be separated or divided by methods known to those skilled in the art, such as selective crystallization techniques or chiral chromatography, at any convenient point in the synthesis of the compounds listed below.

[0050] Compounds of formula I or II are readily convertible to pharmaceutically acceptable salts and can be isolated as pharmaceutically acceptable salts. Salt formation can be carried out by adding a pharmaceutically acceptable acid to form an acid addition salt, or by adding a pharmaceutically acceptable base to form a base addition salt. Salts can also be formed simultaneously with the deprotection of nitrogen or oxygen, i.e., the removal of the protecting group. Examples, reactions, and conditions for salt formation are known to those skilled in the art.

[0051] Compounds of formula I or II, or any of the illustrated formulas, or salts thereof, may be prepared by various procedures, some of which are illustrated in the following preparations and examples. The compounds or salts of the present invention may be prepared by combining specific synthesis steps of each described pathway in different ways, or by combining steps of different pathways. The products of each step in the following preparations can be recovered by conventional methods, including extraction, evaporation, precipitation, chromatography, filtration, grinding, and crystallization.

[0052] In the following schemes, all substituents are as previously defined unless otherwise indicated. Reagents and starting materials are readily available to those skilled in the art. Without limiting the scope of the present invention, the following schemes, preparations, and examples are provided to further illustrate the invention. Compounds of the formulas shown above, or salts thereof, can be prepared by using starting materials or intermediates having the corresponding desired stereochemical configuration.

[0053] [ka]

[0054] In Scheme 1, step A represents the conversion of compound (1) to compound (2). Compound (2) is obtained by reacting compound (1) with (E)-1,4-dibromobuto-2-ene in the presence of lithium bis(trimethylsilyl)amide as a base and a solvent such as THF.

[0055] Step B shows the formation of compound (3) by reacting compound (2) with a solvent such as potassium acetate and MeCN.

[0056] Step C describes the reaction in which oxazolidinone in compound (3) is hydrolyzed using LiOH / H2O2 in the presence of a solvent such as THF to obtain compound (4).

[0057] Step D involves the reaction of compound (4) with 2-tert-butyl-1,3-diisopropylisourea in the presence of a solvent such as 2-methyltetrahydrofuran to form compound (5), which is a tert-butyl ester.

[0058] Step E shows the conversion of compound (5) to compound (6) in the presence of an iridium catalyst and a solvent such as acetone.

[0059] Step F shows the formation of compound (7) by oxidation of the aldehyde in compound (6) using selenium(IV) oxide in the presence of H2O2. In step 2F, compound (7) is converted to compound (7a) (compound of formula II) by acid-deprotection of compound (7) using excess HCl in diethyl ether to obtain the HCl salt compound.

[0060] Step G shows the step of reacting compound (7) with cyclopropylamine in the presence of solvents such as ethyl chloroformate and DCM to obtain compound (8). In Step 2G, compound (8) is converted to compound (8a) (compound of formula II) by acid deprotection of compound (8) using excess HCl in diethyl ether to obtain the HCl salt compound.

[0061] [ka]

[0062] In Scheme 2, in step A, the aldehyde compound (6) is reduced to the alcohol compound (9) using a reducing agent such as sodium borohydride in an alcohol solvent. In step B, the alcohol compound (9) is converted to a mesylate (10) using methanesulfonyl chloride in a solvent such as DCM in the presence of a base such as TEA. In step C, compound (10) is reacted with aniline (optionally substituted with one or two substituents (indicated by "R") selected from halo and OCH3) using a base that may contain potassium carbonate and sodium iodide, and a solvent such as DCM, to obtain compound (11). In step D, compound (11a) (compound of formula II) is obtained by BOC deprotection of (11) using HCl.

[0063] Alternatively, in step E, the mesylate (10) can be reacted with potassium cyanide in a solvent such as DMF to obtain the cyano compound (13). In step F, the cyano group is converted to a carboxylic acid by reaction with HCl, and the BOC group is removed to obtain compound (13a) (compound of formula II).

[0064] The mesylate (10) may be reacted with pyrazole in step G in the presence of a base such as potassium carbonate and a solvent such as MeCN to obtain compound (12), and then BOC deprotection in step H to obtain compound (12a) (compound of formula II).

[0065] [ka]

[0066] In Scheme 3, step A represents the step of converting compound (14) to compound (15) by reacting compound (14) with compound (1) in a solvent such as lithium bis(trimethylsilyl)amide and THF to obtain compound (15).

[0067] Step B describes a reaction in which oxazolidinone in compound (15) is cleaved using LiOH / H2O2 in the presence of a solvent such as THF to obtain compound (16).

[0068] Step C shows that compound (17) (compound of formula II), which is an HCl salt, is obtained by acid deprotection using excess HCl and diethyl ether.

[0069] Step D describes an alternative route to obtain compound (31) by reducing the double bond of compound (16) by hydrogenation using a palladium catalyst and hydrogen gas in an alcohol solvent such as ethanol.

[0070] Step E shows that compound (32) (compound of formula II), which is the HCl salt, is obtained by acid-deprotection of compound (31) using excess HCl and diethyl ether.

[0071] [ka]

[0072] In Scheme 4, step A represents a reductive amination reaction in which compound (6) is reacted with dibenzylamine in the presence of sodium triacetoxyborohydride and a solvent such as DCM to convert it to compound (18), thereby obtaining compound (18).

[0073] Step B shows the conversion of compound (18) to compound (19) in a hydrogenation reaction in the presence of a palladium catalyst on carbon and a solvent such as ethanol.

[0074] Step C involves converting the amine in compound (19) to an acid chloride [R 2 This demonstrates that compound (21) is converted to an amide via a reaction with a base such as -C(O)Cl] or TEA, and a solvent such as DCM.

[0075] Step D shows the formation of compound (20) by reacting 2 equivalents of compound (19) with 1 equivalent of 1,1'-carbonyldiimidazole and a solvent such as THF.

[0076] Step E involves converting the amine in compound (19) to an isocyanate (R) in the presence of a base such as TEA and a solvent such as DCM. 2 This shows that compound (22) can be obtained by reacting it with (-N=C=O).

[0077] Steps 2C, 2D, and 2E show that by removing BOC from compounds (21), (20), and (22) using excess HCl, the HCl salt compounds (24) (compound of formula II), (23) (compound of formula I), and (25) (compound of formula II) are obtained, respectively. 2 This is defined in equation II.

[0078] [ka]

[0079] In Scheme 5, step A shows that compound (1) is alkylated to obtain compound (26). Compound (26) is obtained by reacting compound (1) with allyl bromide in the presence of lithium bis(trimethylsilyl)amide as a base and a solvent such as THF.

[0080] Step B describes a hydrolysis reaction in which oxazolidinone in compound (26) is cleaved using LiOH / H2O2 in the presence of a solvent such as THF to obtain compound (27).

[0081] Step C represents an esterification reaction in which compound (27) is reacted with N,N-dimethylformamide di-tert-butyl acetal in the presence of a solvent such as toluene to obtain compound (28).

[0082] Step D shows a metathesis reaction in the formation of compound (29) by contacting compound (28) with a solvent such as a second-generation Grubbs catalyst and DCM.

[0083] Step E shows that compound (30) (compound of formula II), which is the HCl salt, is obtained by acid-deprotection of compound (29) using excess HCl and diethyl ether.

[0084] Preparation 1 tert-butyl(3R)-3-[2-[(4S)-4-benzyl-2-oxo-oxazolidine-3-yl]-2-oxo-ethyl]pyrrolidine-1-carboxylate

[0085] [ka]

[0086] TEA (56.5 g, 77.9 mL, 559 mmol) was added to a solution of (R)-N-boc-3-pyrrolidineacetic acid (CAS registry number 204688-60-8; 53.8 g, 235 mmol) in THF (540 mL) maintained at 10°C. After 5 minutes, pivaloyl chloride (33.7 g, 34.2 mL, 279 mmol) was added. After 15 minutes, lithium chloride (11.8 g, 279 mmol) and (4S)-4-benzyloxazolidine-2-one (40.0 g, 223 mmol) in THF (540 mL) were added. The mixture was heated to room temperature and stirred for 24 hours. After 24 hours, aqueous HCl (500 mL) was added to separate the organic phase from the aqueous phase. The organic phase was washed with aqueous NaOH solution (500 mL) and saturated aqueous NaCl solution (500 mL), dried over MgSO4, filtered, and concentrated under vacuum. The residue was suspended in a mixture of MeOH and H2O (1:2, 575 mL) and stirred at room temperature for 16-18 hours. The solid was filtered off, washed with hexane (2 × 150 mL), and dried to obtain the title compound (65.7 g, 76%) as a white solid. ES / MS (m / z): 333 (M+H-tert-butyl).

[0087] Preparation 2 tert-butyl(3R)-3-[(E,1S)-5-acetoxy-1-[(4S)-4-benzyl-2-oxo-oxazolidine-3-carbonyl]penta-3-enyl]pyrrolidine-1-carboxylate

[0088] [ka]

[0089] 5.59 g, 10.7 mmol of tert-butyl(3R)-3-[(E,1S)-1-[(4S)-4-benzyl-2-oxo-oxazolidine-3-carbonyl]-5-bromo-penta-3-enyl]pyrrolidine-1-carboxylate (5.59 g, 10.7 mmol) was dissolved in MeCN (75 mL), and potassium acetate (1.6 g, 16 mmol) was added. The mixture was stirred at 70°C for 16-18 hours. The solvent was evaporated, the residue was dissolved in diethyl ether, and washed with H2O. The organic layer was dried over Na2SO4, filtered, and concentrated under vacuum to obtain the title compound as a pale yellow foamy substance (5.36 g, 99+%). ES / MS (m / z): 401 (M+H-Boc).

[0090] Preparation 3 tert-butyl(3R)-3-[(E,1S)-1-tert-butoxycarbonyl-5-hydroxypenta-3-enyl]pyrrolidine-1-carboxylate

[0091] [ka]

[0092] (E,2S)-2-[3R)-1-tert-butoxycarbonylpyrrolidine-3-yl]-6-hydroxyhexa-4-enoic acid (0.28 g, 0.935 mmol) was purged under N2, and then 2-methyltetrahydrofuran (5.5 mL) and 2-tert-butyl-1,3-diisopropylisourea (830 μL) were added. The reaction mixture was stirred at room temperature for 16-18 hours. An additional 2-tert-butyl-1,3-diisopropylisourea (150 mL) was added, and the reaction mixture was stirred at room temperature for approximately 4 hours. The reaction mixture was then diluted with ELISA and filtered through diatomaceous earth. The solid was further washed with diethyl ether (x3), and the filtrate was evaporated. The residue was subjected to three further cycles of solubilization in diethyl ether, filtration, and drying under vacuum. Next, the substance was purified by silica gel chromatography using elution with DCM (3:1) in ethyl acetate to obtain the title compound (0.26 g, 70%) as a concentrated yellow oil. ES / MS m / z: 298 (MH, negative ionization mode).

[0093] Preparation 4 tert-butyl(3R)-3-[(1S)-1-tert-butoxycarbonyl-5-oxopentyl]pyrrolidine-1-carboxylate

[0094] [ka]

[0095] tert-butyl(3R)-3-[(E,1S)-1-tert-butoxycarbonyl-5-hydroxypenta-3-enyl]pyrrolidine-1-carboxylate (0.195 g, 0.56 mmol) was dissolved in acetone and H2O. Dichloro(pentamethylcyclopentadienyl)iridium(III) dimer catalyst (0.006 g, 0.008 mmol) was added, and the reaction mixture was stirred at room temperature for 16-18 hours. The acetone was evaporated, and the residue was partitioned between diethyl ether and H2O. After separation, the organic matter was dried over Na2SO4, evaporated, and dried under vacuum for about 1 hour to obtain a pale yellow, concentrated oily substance (0.186 g). The crude product was purified by silica gel chromatography using elution with DCM (9:1) in toluene, evaporated, and dried under vacuum to obtain the title compound (0.14 g, 75%) as a very concentrated pale yellow oil. ES / MS m / z: 244 (M+H-2×tert-butyl).

[0096] Preparation 5 tert-butyl(3R)-3-[(1S)-1-tert-butoxycarbonyl-5-hydroxypentyl]pyrrolidine-1-carboxylate

[0097] [ka]

[0098] 0.58 g, 1.631 mmol of tert-butyl(3R)-3-[(1S)-1-tert-butoxycarbonyl-5-oxopentyl]pyrrolidine-1-carboxylate was dissolved in 7 mL of EtOH. 0.049 g, 1.29 mmol of sodium borohydride was added, and the reaction mixture was stirred at room temperature for 0.5 hours. The reaction mixture was diluted with ethyl acetate and washed with aqueous solutions of H2O and NH4Cl. The organic matter was dried over Na2SO4, evaporated, and dried under vacuum to obtain a pale yellow, concentrated oil (0.55 g). The crude product was purified by silica gel chromatography using elution with DCM (2.7:1) in ethyl acetate, evaporated, and dried under vacuum to obtain the title compound (0.48 g, 80%) as a concentrated pale yellow oil. ES / MS m / z: 246 (M + H - 2 × tert-butyl).

[0099] Preparation 6 tert-butyl(3R)-3-[(1S)-1-tert-butoxycarbonyl-5-methylsulfonyloxypentyl]pyrrolidine-1-carboxylate

[0100] [ka]

[0101] tert-butyl(3R)-3-[(1S)-1-tert-butoxycarbonyl-5-hydroxypentyl]pyrrolidine-1-carboxylate (0.24 g, 0.671 mmol) and TEA (160 μL) were dissolved in dry DCM under N2. The mixture was cooled to 0°C, and methanesulfonyl chloride (70 μL) was added dropwise. The reaction mixture was stirred at 0°C for 1 hour. The reaction mixture was diluted in DCM and washed with H2O. After separation, the organic matter was dried over Na2SO4, evaporated, and dried under vacuum to obtain the title compound as a pale yellow, very concentrated oil (0.29 g, 90%). ES / MS m / z: 324 (M + H - 2 × tert-butyl).

[0102] Preparation 7 tert-butyl(3R)-3-[(1S)-1-tert-butoxycarbonyl-5-(3-fluoro-5-methoxy-anilino)pentyl]pyrrolidine-1-carboxylate

[0103] [ka]

[0104] 3-Fluoro-5-methoxyaniline and tert-butyl(3R)-3-[(1S)-1-tert-butoxycarbonyl-5-methylsulfonyloxypentyl]pyrrolidine-1-carboxylate (0.162 g, 0.371 mmol) were purged under N2 with potassium carbonate (0.075 g, 0.542 mmol) and sodium iodide (0.008 g, 0.053 mmol). DMF was added, and the reaction mixture was heated at 85°C for 16-18 hours. The reaction mixture was poured into H2O and extracted with diethyl ether. The organic matter was washed with H2O (×2), dried over Na2SO4, evaporated, and dried under vacuum. The crude product was purified by silica gel chromatography with elution in DCM (95:5) in diethyl ether to obtain a very concentrated pale yellow oily substance (0.07 g). The product was purified by reversed-phase chromatography (12g column; A: water, B: MeCN; 70% 2CV, 70-80% 4CV, 80% 4CV) to obtain the title compound as a concentrated pale yellow oily substance (0.038g, 22.91%). ES / MS m / z: 481 (M+H).

[0105] Preparation 8 (5S)-6-tert-butoxy-5-[(3R)-1-tert-butoxycarbonylpyrrolidine-3-yl]-6-oxohexanoic acid

[0106] [ka]

[0107] To a solution of tert-butyl(3R)-3-[(1S)-1-tert-butoxycarbonyl-5-oxopentyl]pyrrolidine-1-carboxylate (0.15 g, 0.422 mmol) in THF (2 mL), selenium(IV) dioxide (0.003 g, 0.270 mmol), followed by H2O2 (34% aqueous solution, 100 μL), was added, and the reaction mixture was heated at 70°C for approximately 2.5 hours. The reaction mixture was diluted with ethyl and washed with H2O. After separation, the organic matter was washed with H2O, dried over Na2SO4, evaporated, and dried under vacuum to obtain a clear, concentrated oil (160 mg). The crude product was purified by silica gel chromatography using elution with DCM (9:1) in diethyl ether, evaporated, and dried under vacuum to obtain the title compound as a concentrated, clear oil (0.125 g, 80%). ES / MS m / z: 260 (M + H - 2 × tert-butyl).

[0108] Preparation 9 tert-butyl(3R)-3-[(1S)-1-tert-butoxycarbonyl-5-(cyclopropylamino)-5-oxopentyl]pyrrolidine-1-carboxylate

[0109] [ka]

[0110] A solution of (5S)-6-tert-butoxy-5-[(3R)-1-tert-butoxycarbonylpyrrolidine-3-yl]-6-oxohexanoic acid (0.215 g, 0.579 mmol) and TEA (170 μL) in dry DCM (3.5 mL) was purged under N2. Ethyl chloroformate (80 μL) was added dropwise, and the mixture was stirred for 0.75 hours. Next, cyclopropylamine (60 μL) was added dropwise, and the reaction mixture was stirred at room temperature for approximately 2.5 hours. The reaction product was diluted with DCM and washed with H2O + 10% Na2CO3. After separating the phases, the organic phase was washed with H2O, dried over Na2SO4, evaporated, and dried under vacuum to obtain a pale yellow, concentrated oily substance (250 mg). The crude product was purified by silica gel chromatography using elution with DCM (4:1) in diethyl ether, evaporated, and dried under vacuum to obtain the title compound (0.13 g, 55%) as a very concentrated pale yellow oil. ES / MS m / z: 411 (M+H).

[0111] Preparation 10 tert-butyl(3R)-3-[(1S)-1-tert-butoxycarbonyl-5-cyanopentyl]pyrrolidine-1-carboxylate

[0112] [ka]

[0113] tert-butyl(3R)-3-[(1S)-1-tert-butoxycarbonyl-5-methylsulfonyloxypentyl]pyrrolidine-1-carboxylate (0.146 g) and potassium cyanide (0.028 g, 0.335 mmol) were purged under N2. Dry DMF (1.2 mL) was added, and the reaction mixture was heated at 90°C for 16-18 hours. The reaction mixture was diluted with diethyl ether and washed with H2O. After separation, the organic matter was washed with H2O (×2), dried over Na2SO4, evaporated, and dried under vacuum to obtain a pale yellow, concentrated oil (80 mg). The crude product was purified by silica gel chromatography with elution using DCM (95:5) in toluene, and dried under vacuum to obtain the title compound (0.058 g, 47%) as a very concentrated, clear oil. ES / MS m / z: 255 (M + H - 2 × tert-butyl).

[0114] Preparation 11 tert-butyl(3R)-3-[(1S)-1-tert-butoxycarbonyl-5-pyrazole-1-ylpentyl]pyrrolidine-1-carboxylate

[0115] [ka]

[0116] A mixture of tert-butyl(3R)-3-[1S)-1-tert-butoxycarbonyl-5-methylsulfonyloxypentyl]pyrrolidine-1-carboxylate (0.155 g, 0.356 mmol) and pyrazole (0.04 g, 0.588 mmol) was purged with potassium carbonate (0.095 g, 0.687 mmol) under N2 in a coolant. MeCN (2.5 mL) was added, and the reaction mixture was heated at 75°C for 16-18 hours and then under an N2 atmosphere for approximately 16 hours. The reaction mixture was then heated at 80°C under an N2 atmosphere for 16-18 hours. The reaction mixture was then diluted with toluene and washed with H2O. After separation, the organic matter was dried over Na2SO4, evaporated, and dried under vacuum to obtain a very pale yellow, concentrated oily substance (150 mg). The crude product was purified by silica gel chromatography using elution with DCM (9:1) in toluene. The solvent was evaporated, and the residue was dried under vacuum to obtain the title compound (0.071 g, 50%) as a very concentrated, clear oil. ES / MS m / z: 408 (M+H).

[0117] Preparation 12 tert-butyl(3R)-3-[(1S)-1-tert-butoxycarbonyl-5-(ethylcarbamoylamino)pentyl]pyrrolidine-1-carboxylate

[0118] [ka]

[0119] A mixture of tert-butyl(3R)-3-[(1S)-5-amino-1-tert-butoxycarbonylpentyl]pyrrolidine-1-carboxylate (0.145 g, 0.407 mmol) and TEA (120 μL) was dissolved in dry DCM under an N2 atmosphere. Isocyanoethane (60 μL) was then added dropwise, and the reaction mixture was stirred at room temperature for approximately 1.5 hours. The reaction mixture was further diluted with DCM and washed with H2O. The organic matter was dried over Na2SO4, evaporated, and dried under vacuum to obtain a concentrated pale yellow oil (190 mg). The crude product was purified by silica gel chromatography with elution in DCM (65:35) in MeOH. The solvent was evaporated, and the residue was dried under vacuum to obtain the title compound (0.124 g, 80%) as a very concentrated clear oil. ES / MS m / z: 428 (M+H).

[0120] Preparation 13 1-[(E)-4-bromobuta-2-enoxy]-3-fluoro-5-methoxybenzene

[0121] [ka]

[0122] A mixture of (E)-1,4-dibromobuta-2-ene (CAS registry no. 821-06-7; 0.00192 g, 1.92 mmol) and 3-fluoro-5-methoxyphenol (CAS registry no. 850793-25-8; 0.833 g) was dissolved in MeCN (50 mL) under an N2 atmosphere with a coolant. Potassium carbonate (0.00135 g, 0.0097 mmol) was added all at once, and the final reaction mixture was stirred at 60°C for about 4.5 hours. The solvent was evaporated, and the residue was partitioned between diethyl ether and H2O. After separating the phases, the organic matter was washed with water, dried over Na2SO4, filtered, and evaporated to obtain a pale yellow, concentrated oil (2 g). The crude product was purified by silica gel chromatography with elution in DCM (1:9) in hexane to obtain the title compound (0.702 g, 45%) as a very pale yellow oil. 1H NMR(400.13MHz,CDCl3)d 6.29-6.25(m,3H),6.14-6.09(m,1H),6.02-5.96(m,1H),4.53(dd,J=1.1,5.1Hz,2H),4.01(dd,J=0.8,7.3Hz,2H),3.79(s,3H).

[0123] Preparation 14 2-[(E)-4-bromobuta-2-enyl]isoindorin-1-one

[0124] [ka]

[0125] (E)-1,4-dibromobuta-2-ene (CAS registry no. 821-06-7; 1.12 g) was dissolved in dry THF (20 mL) under an N2 atmosphere. Then, sodium hydride (60% by mass, mineral oil, 0.175 g) was added at room temperature, and the mixture was stirred for 0.25 hours. Solid isoindorin-1-one (CAS registry no. 480-91-1; 0.551 g, 4.13 mmol) was added all at once, and the reaction mixture was stirred at room temperature for approximately 4.5 hours. The reaction mixture was diluted with diethyl ether, washed with H2O (×2), dried over Na2SO4, evaporated, and stirred under vacuum to obtain a crude yellow solid (1 g) residue. The crude product was purified by silica gel chromatography with elution using siRNA (100 mL). The siRNA was evaporated, and the residue was dried under vacuum to obtain the title compound (0.22 g, 19%) as a very concentrated pale yellow oil. ES / MS m / z: 266, 268 (M+H).

[0126] Preparation 15 3-[(E)-4-bromobuta-2-enyl]-1,3-benzoxazole-2-one

[0127] [ka]

[0128] In a refrigerator, (E)-1,4-dibromobuta-2-ene (CAS registry number 821-06-7; 1.5 g, 7.02 mmol) and 3H-1,3-benzoxazole-2-one (CAS registry number 59-49-4; 0.69 g) were dissolved in MeCN (40 mL) under N2. Potassium carbonate (0.99 g, 7.16 mmol) was then added all at once, and the reaction mixture was stirred at 50°C for approximately 4.25 hours. The solvent was evaporated, and the residue was partitioned between diethyl ether and water. After phase separation, the organic matter was washed with H2O, dried over Na2SO4, filtered, and evaporated to obtain the crude product as a brown solid (1.5 g). The crude product was purified by silica gel chromatography with elution in DCM (3:1) in hexane to obtain the title compound (0.72 g, 50%) as a very pale yellow solid. ES / MS m / z: 268,270 (M+H).

[0129] Preparation 16 tert-butyl(3R)-3-[(1S)-1-tert-butoxycarbonyl-5-(dibenzylamino)pentyl]pyrrolidine-1-carboxylate

[0130] [ka]

[0131] A mixture of tert-butyl(3R)-3-[(1S)-1-tert-butoxycarbonyl-5-oxopentyl]pyrrolidine-1-carboxylate (0.15 g, 0.52 mmol) and dibenzylamine (0.12 mL, 0.61 mmol) was dissolved in DCM (3 mL) and stirred at room temperature for 30 minutes. Sodium triacetoxyborohydride (0.175 g, 0.80 mmol) was added all at once and stirred at room temperature for 16-18 hours. The mixture was diluted with DCM, an aqueous solution of NaHCO3 was added, and the phases were separated. The organic phase was dried over Na2SO4, filtered, and concentrated under vacuum. The residue was purified by silica gel chromatography with elution in 5% diethyl ether in DCM to obtain the title compound (0.17 g, yield 54%) as a concentrated pale yellow oil. ES / MS (m / z): 537 (M+H).

[0132] Preparation 17 tert-butyl(3R)-3-[(1S)-5-amino-1-tert-butoxycarbonylpentyl]pyrrolidine-1-carboxylate

[0133] [ka]

[0134] tert-butyl(3R)-3-[(1S)-1-tert-butoxycarbonyl-5-(dibenzylamino)pentyl]pyrrolidine-1-carboxylate (0.170 g, 0.30 mmol) was dissolved in EtOH (3 mL) and purged with N2. Palladium carbon (5% w / w, 140 mg, 0.06 mmol) was added and purged with hydrogen. The mixture was pressurized with hydrogen at 70 psi, stirred at room temperature for 4.5 hours, filtered through a diatomaceous earth pad, and washed with DCM. The filtrate was evaporated to obtain the title compound (0.12 g, yield 99+%) as a thick, dark brown oil. ES / MS (m / z): 357 (M+H).

[0135] Preparation 18 tert-butyl(3R)-3-[(1S)-5-[[(5S)-6-tert-butoxy-5-[(3R)-1-tert-butoxycarbonylpyrrolidine-3-yl]-6-oxo-hexyl]carbamoylamino]-1-tert-butoxycarbonylpentyl]pyrrolidine-1-carboxylate

[0136] [ka]

[0137] tert-butyl(3R)-3-[(1S)-5-amino-1-tert-butoxycarbonylpentyl]pyrrolidine-1-carboxylate (0.12 g, 0.29 mmol) was dissolved in dry THF (3 mL) under N2. 1,1'-carbonyldiimidazole (0.025 g, 0.15 mmol) was added, and the mixture was stirred at 70°C for 5 hours. The reaction mixture was poured into water and extracted with diethyl ether (×2). The combined organic matter was washed with H2O, dried over Na2SO4, filtered, and concentrated under vacuum. The residue was purified by silica gel chromatography with sequential elution using DCM and diethyl ether (3:1) and ethyl to obtain the title compound (0.050 g, 49%). 1H NMR(400.21MHz,CDCl3):3.67-3.37(m,4H),3.32-3.07(m,6H),2.99-2.81(m,2H ),2.35-2.21(m,2H),2.18-2.08(m,2H),1.99-1.86(m,2H),1.69-1.23(m,50H).

[0138] Preparation 19 Ditert-butyl(E,2S,7S)-2,7-bis[(3R)-1-tert-butoxycarbonylpyrrolidine-3-yl]octa-4-endoioate

[0139] [ka]

[0140] tert-butyl(3R)-3-[(1S)-1-tert-butoxycarbonylbuta-3-enyl]pyrrolidine-1-carboxylate (0.204 g, 0.627 mmol) was purged in a coolant under an N2 atmosphere. Dry DCM was then added, followed by Grubbs Catalyst® M2a (C848, CAS registry no. 246047-72-3, 0.055 g, 0.065 mmol). The reaction mixture was then heated under reflux for 16-18 hours. The solvent was diluted with DCM and washed with H2O. The organic matter was dried over Na2SO4 and evaporated to dryness to obtain a dark brown, concentrated oily substance (0.3 g). The residue was purified by silica gel chromatography using elution with DCM (95:5) in toluene. The solvent was evaporated, and the residue was dried under vacuum to obtain the title compound (0.11 g, 55%) as a brown, foamy substance. ES / MS m / z: 623 (M+H).

[0141] Preparation 20 Ditert-butyl(2S,7S)-2,7-bis[(3R)-1-tert-butoxycarbonylpyrrolidine-3-yl]octanedioate

[0142] [ka]

[0143] A mixture of ditert-butyl(E,2S,7S)-2,7-bis[(3R)-1-tert-butoxycarbonylpyrrolidine-3-yl]octa-4-endioate (400 mg, 0.64 mmol) and palladium catalyst (5% by mass on carbon, 700 mg, 0.33 mmol) was purged under a nitrogen atmosphere. EtOH (7 mL) was added, and the mixture was purged with hydrogen while vigorously stirring. The mixture was stirred under a hydrogen gas balloon at room temperature for 2 hours, and then filtered through a diatomaceous earth pad. The pad was washed with DCM, and the filtrate was evaporated. The residue was dissolved in DCM, dried over Na2SO4, filtered, and concentrated. The residue was purified by silica gel chromatography using 10% siRNA in DCM to obtain the title compound (365 mg, 91%) as a white foamy solid. ES-MS m / z 525 (M+H-Boc).

[0144] Preparation 21 tert-butyl(3R)-3-[(E,1S)-1-[(4S)-4-benzyl-2-oxo-oxazolidine-3-carbonyl]-5-(3-fluoro-5-methoxy-phenoxy)penta-3-enyl]pyrrolidine-1-carboxylate

[0145] [ka]

[0146] tert-butyl(3R)-3-[2-[(4S)-4-benzyl-2-oxo-oxazolidine-3-yl]-2-oxo-ethyl]pyrrolidine-1-carboxylate (0.815 g; 2.1 mmol) was dissolved in dry THF (9 mL) under N2 and cooled to approximately 0°C. Lithium bis(trimethylsilyl)amide (2.3 mL, 1 M) was added dropwise, and the mixture was stirred for approximately 1 hour. Solid 1-[(E)-4-bromobuta-2-enoxy]-3-fluoro-5-methoxybenzene (0.607 g, 2.21 mmol) was added all at once, and the reaction mixture was stirred at 0°C for approximately 1 hour, then left at room temperature for approximately 16-18 hours. The reaction mixture was poured into saturated NaHCO3 and extracted with diethyl ether. The organic matter was further washed with H2O, dried with Na2SO4, and the solvent was evaporated to obtain a concentrated yellow oily substance (1.25 g). The crude substance was purified by silica gel chromatography using elution with DCM(96:4) in toluene. The solvent was removed by distillation, and the residue was dried under vacuum to obtain the title compound (0.53 g, 40%) as a white foamy substance. ES / MS m / z: 483 (M+H-Boc).

[0147] Preparations 22-26 in Table 1 were prepared in a manner similar to that of preparation 21, using the corresponding appropriate reagents and adjusting the reaction time to reach completion.

[0148] [Table 1]

[0149] Preparation 27 (E,2S)-2-[(3R)-1-tert-butoxycarbonylpyrrolidine-3-yl]-6-hydroxyhexa-4-enoic acid

[0150] [ka]

[0151] A solution of tert-butyl(3R)-3-[(E,1S)-5-acetoxy-1-[(4S)-4-benzyl-2-oxo-oxazolidine-3-carbonyl]penta-3-enyl]pyrrolidine-1-carboxylate (5.36 g, 10.7 mmol) in THF (80 mL) was cooled to 0°C. H2O2 (34 wt%, 9.5 mL, 110 mmol) was added, followed by a solution of lithium hydroxide (750 mg, 30.7 mmol) in H2O (14 mL). The reaction mixture was stirred for 16-18 hours, during which time it was allowed to reach room temperature. The mixture was then poured into H2O, siRNA and 1 M aqueous HCl were added, and the mixture was shaken vigorously. The phases were separated, and the organic phase was washed with H2O (×2). The organic phase was dried over Na2SO4, filtered, and concentrated under vacuum. The residue was purified by silica gel chromatography using elution with 3% MeOH in DCM containing 0.25% acetic acid to obtain a concentrated oily substance containing a small amount of acetic acid. The residue was then dissolved in DCM and washed with H2O (×2). The organic phase was dried over Na2SO4, filtered, and concentrated under vacuum to obtain the title compound (2.1 g) as a concentrated oily substance. ES / MS (m / z): 298 (MH).

[0152] Compounds 28-33 in Table 2 were prepared in a manner similar to that of Preparation 27, using the corresponding appropriate reagents and adjusting the reaction time to complete the process.

[0153] [Table 2]

[0154] Preparation 32 (2S)-2-[(3R)-1-tert-butoxycarbonylpyrrolidine-3-yl]-5-phenylpentanoic acid

[0155] [ka]

[0156] A solution of (E,2S)-2-[(3R)-1-tert-butoxycarbonylpyrrolidine-3-yl]-5-phenyl-penta-4-enoic acid (155 mg, 0.449 mmol) in ethanol (7 mL) was purged with nitrogen, and then palladium (5% by mass on carbon, 205 mg, 0.096 mmol) was added. The reaction mixture was purged with hydrogen, and then stirred for 3.5 hours under a hydrogen gas balloon. The mixture was filtered through a diatomaceous earth pad, rinsed with DCM, and the filtrate was concentrated under reduced pressure. The residue was dissolved in DCM, dried over Na2SO4, filtered, and concentrated under reduced pressure to obtain the title compound (149 mg, 96%) as a white foam. ES-MS m / z 292 (M+H-tert-butyl).

[0157] Preparation 34 tert-butyl(3R)-3-[(1S)-1-tert-butoxycarbonylbuta-3-enyl]pyrrolidine-1-carboxylate

[0158] [ka]

[0159] To a solution of (2S)-2-[(3R)-1-tert-butoxycarbonylpyrrolidine-3-yl]penta-4-enoic acid (D, 10.5 mmol, 10.5 mmol, 2.83 g) in toluene (105 mL) at 80°C, N,N-dimethylformamide di-tert-butyl acetal (42.0 mmol, 90% by mass, 11.2 mL, 9.49 g) was added, and the mixture was heated at 80°C for 1 hour. Further addition of N,N-dimethylformamide di-tert-butyl acetal (42.0 mmol, 90% by mass, 11.2 mL, 9.49 g) was added, and the mixture was heated at 80°C for 3 hours. The reaction mixture was concentrated, and the residue was purified by silica gel chromatography (5-20% acetone gradient in hexane) to obtain the title compound (1.8 g, 53%) as a colorless oil. ES-MS m / z 348(M + Na + ).

[0160] Preparation 35 tert-butyl(3R)-3-[(1S)-5-acetamido-1-tert-butoxycarbonylpentyl]pyrrolidine-1-carboxylate

[0161] [ka]

[0162] To a mixture of tert-butyl(3R)-3-[(1S)-5-amino-1-tert-butoxycarbonylpentyl]pyrrolidine-1-carboxylate (110 mg, 0.308 mmol) and TEA (100 μL, 72.5 mg, 0.72 mmol) in DCM (2 mL), acetyl chloride (25 μL, 27.5 mg, 0.35 mmol) was added dropwise, and the mixture was stirred at room temperature for 2 hours. The mixture was diluted with DCM and washed with water. The organic matter was dried over Na2SO4, filtered, and concentrated under reduced pressure. The residue was purified by silica gel chromatography using a gradient of 33-100% diethyl ether in DCM to obtain the title compound (75 mg, 68%) as a pale yellow oil. ES-MS m / z 299 (M-BOC+H).

[0163] Preparation 36 tert-butyl(3R)-3-[(1S)-1-tert-butoxycarbonyl-5-(3-fluoro-5-methoxyphenoxy)pentyl]pyrrolidine-1-carboxylate

[0164] [ka]

[0165] A mixture of tert-butyl(3R)-3-[(1S)-1-tert-butoxycarbonyl-5-hydroxypentyl]pyrrolidine-1-carboxylate (265 mg, 0.74 mmol), 3-fluoro-5-methoxyphenol (129 mg, 0.91 mmol), and triphenylphosphine (241 mg, 0.92 mmol) was purged under a nitrogen atmosphere, then dry THF (4 mL) was added, and the mixture was cooled in an ice / water bath. Diisopropyl azodicarboxylate (190 μL, 196 mg, 0.97 mmol) was added dropwise, and the reaction mixture was stirred for 0.5 hours while cooling. The reaction mixture was stirred overnight at room temperature, then poured into a 10% Na2CO3 aqueous solution, and extracted with diethyl ether. The organic matter was dried over Na2SO4, filtered, and concentrated under reduced pressure. The residue was purified by silica gel chromatography (DCM) using the eluent as the eluent, and then re-purified twice by reverse-phase HPLC to obtain the title compound (96 mg, 27%) as a brown oily substance. ES-MS m / z 382 (M-BOC+H).

[0166] Preparation 37 (2S)-2-[(3R)-1-tert-butoxycarbonylpyrrolidine-3-yl]-6-(2-oxo-1,3-benzoxazole-3-yl)hexanoic acid

[0167] [ka]

[0168] A mixture of (E,2S)-2-[(3R)-1-tert-butoxycarbonylpyrrolidine-3-yl]-6-(2-oxo-1,3-benzoxazole-3-yl)hexa-4-enoic acid (52 mg, 0.12 mmol) in ethanol (2 mL) was purged with nitrogen. Palladium (5% by mass on carbon, 75 mg, 0.035 mmol) was then added, and the reaction mixture was purged with hydrogen and maintained under a hydrogen balloon for 2 hours. The mixture was filtered through a diatomaceous earth pad, and the pad was washed with DCM. The filtrate was concentrated under reduced pressure, and the residue was purified by silica gel chromatography using a gradient of 3% MeOH in DCM to obtain the title compound (42 mg, purity 82%) as a white foam. ES-MS m / z 319 (M-BOC+H).

[0169] Example 1 (2S)-6-(3-fluoro-5-methoxyanilino)-2-[(3R)-pyrrolidine-3-yl]hexanoic acid; dihydrochloride

[0170] [ka]

[0171] 0.3 mL of concentrated HCl (35 wt% aqueous solution) was added to a solution of tert-butyl(3R)-3-[(1S)-1-tert-butoxycarbonyl-5-(3-fluoro-5-methoxyanilino)pentyl]pyrrolidine-1-carboxylic acid (0.038 g, 0.079 mmol) in diethyl ether (3 mL). The reaction mixture was stirred at room temperature for 16-18 hours. The ether phase was removed by pipette, and the aqueous phase was washed with diethyl ether (×2). Then, H2O was evaporated with a stream of N2, and the mixture was dried under vacuum to obtain the title compound (0.029 g, 95.17%) as a light brown solid. ES / MS m / z: 325 (M+H)

[0172] Examples 2-17 in Table 3 were prepared in essentially the same manner as described in Example 1, using appropriate BOC and tert-butyl ester protected starting materials to simultaneously deprotect the amine and acid functional groups, or using appropriate BOC protected starting materials having an unprotected carboxylic acid.

[0173] [Table 3-1]

[0174] [Table 3-2]

[0175] [Table 3-3] a. Starting material used: tert-butyl(3R)-3-[(1S)-1-tert-butoxycarbonyl-5-cyanopentyl]pyrrolidine-1-carboxylate

[0176] In vitro Lp(a) aggregation assay The ability of the compounds to inhibit Lp(a) particle formation in vitro was evaluated by a cell-free aggregation assay. Conditioning media (DMEM supplemented with 10% FBS, 20 mM HEPES, and 1x penicillin / streptomycin) were cultured at 37°C and 5% CO2 for 24 hours, and then collected from confluent wild-type HepG2 cells (source of endogenously expressed ApoB) and from HEK293 stable cell lines expressing human Apo(a) containing 17 kringle repeats (selected with 1 mg / mL Geneticin). In vitro aggregation assays were performed by combining the test compounds with equal volumes of HepG2 and HEK293 conditioning media in a dilution series (final concentrations 0.01–100 nM). The reaction mixtures were incubated at 37°C for 2 hours, followed by the addition of 6-aminocaproic acid (EACA) to a final concentration of 150 mM to stop the reaction. Lp(a) was detected using a sandwich ELISA with an anti-Lp(a) capture antibody and an HRP-conjugated anti-ApoB detection antibody. The ELISA was colorimetrically developed using TMB and stopped with 1N sulfuric acid, and the signal was read at 450 nm with a Molecular Devices plate reader. The inhibition percentage of Lp(a) formed under each test condition was set to 0% inhibition for the aggregate reaction in the absence of the inhibitor (corresponding to a 1% DMSO concentration) and to 100% inhibition for the minimum amount of HepG2 conditioned medium present (50-fold dilution). To determine the IC50 values ​​summarized in Table 4, the data were fitted to a four-parameter curve. As summarized in Table 4, the addition of exemplary test compounds to conditioned media containing ApoB and Apo(a) resulted in concentration-dependent inhibition of Lp(a) formation in vitro. The results indicate that these compounds inhibit the aggregation of Lp(a) from Apo(a) and LDL particles.

[0177] [Table 4]

Claims

1. formula: 【Chemistry 1】 (In the formula, L is C ２～ C ６ Alkylene, C ２～ C ６ Alkenylene, or -(CH ２ ) ｎ NHCONH (CH ２ ) ｎ - and n is independently between 2 and 4 in each occurrence. Y is, in each occurrence, independently, CH ２ , O, or S, and In each occurrence, Z is independently H and C １～ C ４ (It is alkyl, OH, or cyclopropyl.) Compounds thereof, or pharmaceutically acceptable salts thereof.

2. The compound according to claim 1, or a pharmaceutically acceptable salt thereof, wherein Z is H in each occurrence.

3. Y is CH in each appearance ２ The compound according to claim 1 or 2, or a pharmaceutically acceptable salt thereof.

4. L is C ２ A compound according to any one of claims 1 to 3, which is an alkylene, or a pharmaceutically acceptable salt thereof.

5. L is C ２ A compound according to any one of claims 1 to 3, which is an alkenylene, or a pharmaceutically acceptable salt thereof.

6. L is - (CH ２ ) ３ NHCONH (CH ２ ) ３ - The compound according to any one of claims 1 to 3, or a pharmaceutically acceptable salt thereof.

7. The aforementioned compound, 【Chemistry 2】 Selected from, The compound according to claim 1, or a pharmaceutically acceptable salt thereof.

8. formula: 【Transformation 3】 (In the formula, L １ C ２～ C ３ Alkylene, or C ２～ C ３ It is alkenylene, R １ is -NHC(O)R ２ , -NHC(O)NHR ２ COOH, -C(O)NH-cyclopropyl, 【Chemistry 4】 -O-phenyl, -NH-phenyl, or phenyl, where phenyl is a halo and OCH ３ It is optionally substituted with one or two substituents selected from the following: R ２ C １～ C ３ It is alkyl, Y is CH ２ , O, or S, Z is H, C １～ C ４ (It is alkyl, OH, or cyclopropyl.) Compounds thereof, or pharmaceutically acceptable salts thereof.

9. The compound according to claim 8, or a pharmaceutically acceptable salt thereof, wherein Z is H.

10. Y is CH ２ The compound according to claim 8 or 9, or a pharmaceutically acceptable salt thereof.

11. R １ is -NHC(O)CH ３ , -NHC(O)NHCH ２ CH ３ COOH, -C(O)NH-cyclopropyl, 【Transformation 5】 【Transformation 6】 A compound according to any one of claims 8 to 10, or a phenyl, or a pharmaceutically acceptable salt thereof.

12. A pharmaceutical composition comprising a compound according to any one of claims 1 to 11, or a pharmaceutically acceptable salt thereof, and at least one pharmaceutically acceptable carrier, diluent, or excipient.

13. A method for treating a cardiovascular disease in a patient, comprising administering to the patient an effective amount of a compound according to any one of claims 1 to 11, or a pharmaceutically acceptable salt thereof.

14. A method for treating an elevation of Lp(a) plasma levels in a patient, comprising administering to the patient an effective amount of a compound according to any one of claims 1 to 11, or a pharmaceutically acceptable salt thereof.

15. A compound according to any one of claims 1 to 11, or a pharmaceutically acceptable salt thereof, for use in therapy.

16. A compound according to any one of claims 1 to 11, or a pharmaceutically acceptable salt thereof, for use in the treatment of cardiovascular disease.

17. A compound according to any one of claims 1 to 11, or a pharmaceutically acceptable salt thereof, for use in the treatment of elevated Lp(a) plasma levels.

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