Pyrrolidine Compounds

Abstract

To provide additional treatment options for patients suffering from cardiovascular diseases and, in particular, patients suffering from lipid disorders or dyslipidemia.SOLUTION: Compounds of the structure in the figure are provided to reduce plasma Lp(a) levels.SELECTED DRAWING: None

Description

[Technical field]

[0001] The present invention relates to pyrrolidine compounds, pharma- ceutically acceptable salts thereof, pharmaceutical compositions, and The present invention relates to the therapeutic use of the compounds of the present invention.

[0002] There have been great advances in the treatment of cardiovascular disease (CVD). Despite these advances, Patients continue to experience cardiovascular events such as angina, myocardial infarction, and stroke, and are being treated. Lipid disorders or dyslipidemia remain the major risk factor for CVD. Lipid disorders are caused by four common risk factors: low-density lipoprotein cholesterol (LDL) Increase in high-density lipoprotein cholesterol (HDL-c), decrease in triglyceride These can be divided into increases in triglycerides (TG) and increases in lipoprotein (a) (Lp(a)). Various drugs targeting high LDL-c, low HDL-c, and high triglycerides are available. There are few approved treatment options for patients with high Lp(a) levels. In some cases, apheresis is used to filter the blood to remove LDL and Lp(a). may remove the sebum, but the effect is temporary and usually needs to be repeated every 2 weeks. There are no approved drug treatments to lower Lp(a) levels. Although the physiological functions of Lp(a) are complex, elevated plasma levels of Lp(a) are an independent risk factor for CVD. It has been reported that Lp(a)1 is a contributing factor. Drug treatment is necessary for patients with elevated Lp(a)1. It is.

[0003] For patients with cardiovascular disease, especially those with lipid disorders or dyslipidemia, Additional treatment options are desirable. Diet, exercise, and / or statins, fibrinogens, and other anti-inflammatory drugs are also currently available. Patients who have been diagnosed with cardiovascular disease using the current standard of care, such as the use of one or more medications such as niacin or niacin. Additional treatment options are needed for patients whose cancer is not adequately controlled. Offers another treatment option for patients with VD. Reduces plasma Lp(a) levels Therefore, there is a need for pharmacologic acceptable compounds and treatment options.

[0004] A compound of formula I', [ka] During the ceremony, L is -CH2NHCH2-, -CH2NH-, -NH-, -S-, -S(O)-, -S (O)2-, -O-, -OCH2-, -OCH2CH2O-, -NHSO2NH-, [ka] is selected from the group consisting of R 1 , R 2 , R 3 , R 4 , R 5 , and R 6 are each independently converted from H and CH3 or a compound of formula I' selected from the group consisting of Pharmaceutically acceptable salts thereof are provided.

[0005] Compounds of Formula I'' [ka] ,or Pharmaceutically acceptable salts thereof are provided.

[0006] A compound of formula 1, [ka] or a pharma- ceutically acceptable salt thereof is provided.

[0007] One embodiment is a compound of formula 2: [ka] or a pharma- ceutically acceptable salt thereof.

[0008] In one embodiment, the compound is of formula I', I'', 1 or formula 2, and the compound is In one embodiment, the compound is of Formula I', Formula 1, or Formula 2, In one embodiment, the compound of formula 1 or formula 2 is a hydrochloride salt. is the tetrahydrochloride salt.

[0009] In one embodiment, the compound of Formula I', Formula 1, or Formula 2 is a monohydrochloride salt. In one embodiment, the hydrochloride is a hydrochloride salt selected from the group consisting of a dihydrochloride, a trihydrochloride, and a tetrahydrochloride. In the form, the compound of Formula 1 or Formula 2 is a zwitterion.

[0010] A compound of formula I', wherein L is -CH2NHCH2-, -CH2NH-, -N H-, -S-, -S(O)-, -S(O)2-, -O-, -OCH2-, -OCH2CH 2O-, -NHSO2NH-, or a pharma- ceutically acceptable salt thereof. Acceptable salts are provided.

[0011] A compound of formula I', wherein R 1 and R 3 are H, and L is -CH 2 NHCH 2 -, -CH2NH-, -NH-, -S-, -S(O)-, -S(O)2-, - Selected from the group consisting of O-, -OCH2-, -OCH2CH2O-, -NHSO2NH- The present invention provides a compound, or a pharma- ceutically acceptable salt thereof, which is

[0012] A compound of formula I', wherein R 1 and R 3 are H and R 2 and R 4 are CH3, and L is -CH2NHCH2- 、 -CHNH-, -NH -、- S-、-S(O)-、-S(O)2 -, -O-, -OCH2-, -OCH2CH2O-, -NHSO2NH-, or a pharma- ceutically acceptable salt thereof. Salt is provided.

[0013] A compound of formula I', wherein R 1 and R 3 are H and R 2 and R 4 are H, and L is -CH 2 NHCH 2 -, -CH 2 NH-, -NH-, -S- , -S(O)-, -S(O) 2 -, -O-, -OCH2-, -OCH2CH2O-, -N HSO2NH-, or a pharma- ceutically acceptable salt thereof Provided.

[0014] A compound of formula I', wherein L is [ka] or a pharma- ceutically acceptable salt thereof is provided.

[0015] A compound of formula I', wherein R 1 , R 3 , and R 5 are H, and L is [ka] or a pharma- ceutically acceptable salt thereof is provided.

[0016] A compound of formula I', wherein R 1 , R 3 , and R 5 are H and R 2 , R 4 , and R 6 are each H, or a pharma- ceutically acceptable salt thereof. will be done.

[0017] A compound of formula I', wherein R 1 , R 3 , and R 5 are H and R 2 , R 4 , and R 6 are each CH3, or a pharma- ceutically acceptable salt thereof; Provided.

[0018] A compound of formula I', wherein L is [ka] and R 1 , R 3 , and R 5 are H and R 2 , R 4 , and R 6 Each or a pharma- ceutically acceptable salt thereof is provided.

[0019] In one embodiment, a compound of Formula I', Formula I'', Formula 1, or Formula 2, or a pharmaceutical agent thereof. and at least one pharma- ceutically acceptable carrier, diluent, or excipient. and

[0020] In one embodiment, the method is a method of treating a patient in need of treatment for cardiovascular disease, an effective amount of a compound selected from the group consisting of formula I', formula 1, and formula 2, or a combination thereof In one embodiment, the method comprises administering a physiologically acceptable salt thereof to a subject in need of treatment for cardiovascular disease. The present invention relates to a method of treating a patient suffering from a rheumatoid arthritis, the method comprising administering to said patient an effective amount of a compound of formula I″ or a pharmaceutical agent thereof. In one embodiment, the method comprises administering a physiologically acceptable salt thereof to the subject. The present invention relates to a method for treating a patient in need of treatment for hypercalcaemia, the method comprising administering to said patient an effective amount of a compound of formula I', formula I', or a compound selected from the group consisting of formula 2, or a pharma- ceutically acceptable salt thereof. In one embodiment, a patient in need of treatment for elevated plasma levels of Lp(a) is The method comprises administering an effective amount of a compound of formula I″ or a pharma- ceutical acceptable salt thereof to a patient having a medical condition, comprising administering to said patient an effective amount of a compound of formula I″ or a pharma- ceutical acceptable salt thereof. This includes administering a salt thereof.

[0021] In one embodiment, a compound selected from the group consisting of Formula I', Formula 1, and Formula 2 is provided for use in therapy. In one embodiment, the compound is used in the treatment The compound of formula I″, or a pharma- ceutically acceptable salt thereof, for use in

[0022] In one embodiment, a compound of formula I', formula 1, and formula 2 is provided for use in the treatment of cardiovascular disease. or a pharma- ceutically acceptable salt thereof. The present invention relates to a compound of formula I″, or a pharma- ceutical acceptable salt thereof, for use in the treatment of cardiovascular disease. It is an acceptable salt.

[0023] In one embodiment, a compound of formula I', formula 1 is provided for use in the treatment of elevated Lp(a) plasma levels. and 2, or a pharma- ceutically acceptable salt thereof. do.

[0024] In one embodiment, a compound of formula I', formula 1, and formula 2 is used in the manufacture of a medicament for cardiovascular disease. The present invention relates to the use of a compound selected from the group, or a pharma- ceutically acceptable salt thereof.

[0025] Lp(a) has both prothrombotic and antithrombotic properties, as well as atherothrombotic characteristics. Lp(a) inhibits fibrinolysis and accumulates in the vascular wall, causing thrombus formation and atherosclerosis. It can induce atherosclerotic lesions. Plasma levels of Lp(a) vary substantially between individuals. Unlike other risk factors, Lp(a) plasma levels are significantly altered by diet and exercise. Lp(a) plasma levels may be related to genetic predisposition.

[0026] Lp(a) contains the LDL lipid core with associated apolipoprotein B (apoB). Lp(a) is similar to LDL-c in that it binds to disulfides. Specific apolipoprotein(a) (apo(a)) bound to apoB via a peptide bond Apo(a) is synthesized in the liver. Apo(a) and LDL particles are synthesized in the liver. Aggregations of p(a) can occur within liver cells, on the cell walls, or in the plasma. Inhibition of particle assembly with apo(a) can reduce Lp(a) levels.

[0027] As used herein, the term "elevated Lp(a) plasma levels" refers to an elevation of approximately 50 mg / mL. The compounds provided herein are intended to provide Lp(a) plasma levels of 0.1 g / dL or greater. It can be used therapeutically to reduce levels.

[0028] As used herein, the term "pharmaceutical acceptable salt" refers to a clinically and / or Refers to salts of compounds that are acceptable for veterinary use. Pharmaceutically acceptable salts and their preparation An example of a general methodology for this is given in the “Handbook of Pharmacology” cal Salts:Properties,Selection and Use”P .Stahl,et al.,2nd Revised Edition,Wiley- VCH, 2011 and S. M. Berge, et al., “Pharmaceutical Cal Salts”, Journal of Pharmaceutical Sci. Ences, 1977, 66(1), 1-19. Formula I', I'', 1 or The compounds of formula 2 may be zwitterions, monoacid, diacid, or triacid addition salts. The compounds of formula 1 or 2 may be mono-, di-, or tri-base addition salts.

[0029] The pharmaceutical compositions of the present invention may be prepared using pharma- ceutically acceptable excipients. The term "physically acceptable" means that the compound is compatible with the other ingredients of the composition and is not pharmacologically harmful to the patient. The term "compound" refers to one or more carriers, diluents, and / or excipients that are not part of the pharmaceutical composition. Examples of processes for their preparation are well known to those skilled in the art and are described, for example, in "Reming n:The Science and Practice of Pharmacy”, Loyd, V.,et al.Eds.,22nd Ed.,Mack Publish can be found in http: / / www.ingco.jp / , 2012.

[0030] As used herein, the term "effective amount" refers to an amount effective to treat a disorder. The effective amount for a particular patient can be determined by a skilled medical practitioner. do.

[0031] As used herein, the terms "treating," "treat," or "treatment" The term refers to a treatment that slows or reduces the progression or severity of an existing symptom, disorder, condition, or disease. As used herein, "cardiac atherosclerosis" includes causing, preventing, or reversing a cardiac atherosclerosis. "Treating vascular disease" means slowing, mitigating, or preventing the progression of heart or vascular disease. The compound of formula I', formula 1, or formula 2 is used to prevent or reverse the The present invention provides a method of treating myocardial infarction comprising administering to a patient in need thereof a

[0032] As used herein, the term "patient" refers to a mammal. The subject is a human being.

[0033] The pharmaceutical composition may be in the form of a tablet or capsule for oral administration, a solution for oral administration, or In one embodiment, the composition is suitable for oral administration. be.

[0034] Specific abbreviations are defined as follows: "Apo" refers to apolipoprotein; "B "OC" stands for tert-butoxycarbonyl, and "BSA" stands for bovine serum albumin. "DAD" stands for Diode Array Detector, and "DCM" stands for Dichloromethane or or methylene chloride, "de" refers to diastereomeric excess, and "DMEA" refers to "Dimethylethylamine" refers to dimethyl ethylamine, and "DMEM" refers to Dulbecco's Modified Eagle's Medium. "DMF" refers to N,N-dimethylformamide, and "DMSO" refers to dimethylsulfate. "ee" stands for enantiomeric excess, and "EACA" stands for epsilon-amino Caproic acid or 6-aminocaproic acid, and "ELISA" stands for enzyme-linked immunosorbent assay. "Et2O" refers to diethyl ether. "EtOAc" refers to ethyl acetate, and "EtOH" refers to ethanol or ethyl acetate. "Ex" refers to an embodiment; "FBS" refers to fetal bovine serum; "HEC" refers to hydroxyethyl cellulose and "HEK" refers to human embryonic kidney. "HepG2" refers to a human hepatoma cell line, and "HEPES" refers to 4-(2-hydroxyethyl ether) 1-(ethyl)-1-piperazineethanesulfonic acid, and "HPLC" stands for high performance liquid chromatography. "HRP" stands for horseradish peroxidase, and "IC 50 "teeth, "min" refers to the concentration of a drug that produces 50% of the maximum inhibitory response possible for that drug, and "min" refers to the concentration of a drug that produces 50% of the maximum inhibitory response possible for that drug. (singular or plural), and "MeOH" refers to methanol or methyl alcohol. "MTBE" stands for methyl tert-butyl ether, and "RP-HPLC / MS" refers to reversed-phase high-performance liquid chromatography with mass spectrometry, and "RT" refers to room temperature. "SFC" stands for supercritical fluid chromatography, and "SPA" stands for scintillation chromatography. The term "t-proximity assay" refers to (R) " refers to retention time, and "THF" refers to tetrahydrofuran. "TMB" stands for 3,3',5,5'-teramethylbenzidine, and "T "ris" refers to tris(hydroxymethyl)aminomethane.

[0035] The individual isomers, enantiomers, and diastereomers can be readily understood by one of ordinary skill in the art to be At any convenient point in the synthesis of the compound to be synthesized, selective crystallization techniques or chiral crystalline structures may be used. They may be separated or resolved by methods such as chromatography.

[0036] The compounds of formula I', I'', 1, or 2 can be readily converted to pharma- ceutically acceptable forms. The formation of salts can be achieved by the addition of a pharma- ceutically acceptable acid. When forming a salt or by addition of a pharma- ceutically acceptable base to form a base addition salt Salts can also be prepared by deprotection of the nitrogen or oxygen, i.e., removal of a protecting group. Examples of salt formation, reactions and conditions are known to those skilled in the art. do.

[0037] A compound selected from the group consisting of formula I', formula I'', formula 1, and formula 2, or The salts of can be prepared by a variety of procedures, some of which are described below in detail. The examples show: Certain synthetic steps may be combined in different ways or in combination with steps from different routes. The products of each step in the following preparation may be extracted, evaporated, precipitated, chromatographed, or It can be recovered by conventional methods, including roughening, filtration, grinding, and crystallization.

[0038] In the following schemes, all substituents are as previously defined unless otherwise indicated. The reagents and starting materials are readily available to one of ordinary skill in the art. Without limiting the scope, the following schemes, preparations and examples further illustrate the invention. Compounds of Formula I', I'', Formula 1, and Formula 2, or salts thereof, are provided herein. by using starting materials or intermediates that have the corresponding desired stereochemical configuration. It can be prepared by the following steps.

[0039] Scheme 1 illustrates the preparation of intermediates that can provide access to the compounds of the present invention. The pyrrolidin-3-yl acetic acid derivative A is first converted to an acyloxazolidinone B. This can be achieved by first converting A to its acid chloride, followed by the reaction in the presence of lithium chloride at 10 °C with the (4S)- This is achieved by reacting with 4-benzyloxazolidin-2-one. Derivatives and bases such as lithium bis(trimethylsilyl)amide at 0°C Alkylation of ruxazolidinone B affords intermediate C in a diastereoselective manner. Those skilled in the art will appreciate that the stereochemistry of the oxazolidinone substitution affects the diastereoselectivity of the alkylation. In this synthesis, the oxazolidinyl group of the opposite stereochemical configuration or racemic stereochemical configuration is The use of non-derivative B gives either the opposite or diastereoselectivity, respectively. The conversion of acyl oxazolidinone C to the acid intermediate D can be achieved by This is accomplished using aqueous LiOH and H2O2 at 5-15 °C. The acid intermediate D can be The acid intermediate D can be obtained by, for example, reaction with tert-butyl ether at elevated temperature. tert-Bromo-1,3-diisopropylisourea to give intermediate E. It is protected as the ethyl ester.

[0040] Scheme 1 also shows the conversion of acid A to methyl ester F, which can be obtained by the addition of a carbonate base. This is accomplished by reacting A with iodomethane in the presence of - Benzyl bromide derivatives were synthesized using a base such as lithium bis(trimethylsilyl)amide at 78°C. The intermediate G is alkylated with lithium bis(trimethylsilyl) ether at -78°C to give intermediate G. Re-alkylation with iodomethane using a base such as ethylsilylamide, followed by esterification with ethylsilylamide. The ter is hydrolyzed with sodium hydroxide at high temperature to give the acid intermediate H. For example, reaction with tert-butyl-1,3-diisopropylisourea at high temperature gives te Protection as the rt-butyl ester gives intermediate I.

[0041] In particular, R a Intermediates D, E, H, and I, in which is bromine or -NO2, are compounds of formula I'. It is particularly useful for further transformations in the preparation of R a Intermediate D, where -H, is Alkylation of B with benzyl followed by hydrolysis of the acyloxazolidinone Or R a The intermediate D, in which is a bromide, is stirred with palladium on carbon under a hydrogen atmosphere to obtain It can be prepared either by a When -H, the pyrrolidinyl group on intermediate D Deprotection of the nitrogen affords compounds of formula I. [ka]

[0042] Scheme 2 shows the synthesis of a key intermediate (J, prepared as described in Scheme 1). The conversion of the bromide to the penultimate compound of the invention is shown. CO / H2), palladium(II) acetate, butyldi-1-adamantylphosphine, and and N,N,N',N'-tetramethylethylenediamine at high temperatures to give aldehyde K. The aldehyde K is then converted to aldehyde K by the addition of ammonia and sodium triacetoxyborohydride. Reductive amination of K with reducing agents such as thorium or sodium cyanoborohydride ( (2 or 3 equivalents, respectively) to a mixture of N and O, and then N and O are Alternatively, the dimeric compound O can be separated by chromatographic separation of the aldehyde K. Aldoxime L was converted to aldoxime L by flow hydrogenation using sponge nickel catalyst at high temperature. The amines L are reduced to amines M, followed by reductive amination of amines M with aldehydes K. Intermediate P is produced by the reaction of 3-fluoro-5-methoxybenzaldehyde with the dimer intermediate. It is prepared by reductive amination of form O. [ka]

[0043] Scheme 3 illustrates the further synthesis of bromide intermediate J for the preparation of the penultimate compounds of the present invention. Bromide J is tetrahydroxydiboron, chloro(2-dicyclohexyl) 2',4',6'-tri-i-propyl-1,1'-biphenyl )(2'-amino-1,1'-biphenyl-2-yl)palladium(II), X-PHO S, and potassium acetate to convert it to boronic acid Q. Boronic acid Q is then reacted with 5 Phenol T is converted to phenol T using H2O2 at 10 °C. Phenol T is converted to phenol T using copper iodide at high temperature. (I), N,N-dimethylglycine hydrochloride, and cesium carbonate were used to form bromide J. The biphenyl ether U is obtained by combining phenols T and T. Phenol T can also be converted to 1,2-dibromoethylene at high temperatures. The aldehyde intermediate K (as shown in Scheme 2) is reacted with benzene and a carbonate base to give V. The alcohol R was then reduced to the alcohol (prepared from the compound J) with sodium borohydride at 0°C, followed by photochemical reaction. Upon reaction, intermediate S is obtained. [ka]

[0044] Scheme 4 illustrates the further synthesis of bromide intermediate J for the preparation of the penultimate compounds of the present invention. The use of 2 equivalents of bromide J in bis(dibenzylideneacetone)palladium nitrile is shown. phosphate, tripotassium phosphate, 1,1'-bis(diphenylphosphino)ferrocene This is then coupled with potassium thioacetate to give diphenyl sulfide W. Sulfide W is then In either the sulfone X or the sulfoxide Y, 1 or 2 equivalents of methionine are added, respectively. It is converted using chloroperoxybenzoic acid. [ka]

[0045] Scheme 5 illustrates the synthesis of nitro intermediate Z (S) for the preparation of the penultimate compounds of the present invention. The nitro intermediate Z was prepared as in scheme 1. In the presence of naphthalene triacetoxyborohydride, aniline AA is reduced to naphthalene triacetoxyborohydride. Reduction with aldehyde K (prepared as in Scheme 2) using a reducing agent such as ammonium phosphate. This is converted to CC by selective amination. Oxy-2',4',6'-triisopropyl-1,1'-biphenyl)-2-(2'-a 1,1'-amino-biphenyl)]palladium(II) methanesulfonate (BrettPh os Pd G3) and potassium carbonate at high temperature to prepare aniline AA and bromide BB ( The diphenylamine derivative was prepared as described in Kiem 1 by the Buchwald reaction between Two equivalents of aniline AA also give the 1,4-diazabicyclo[2.2.2] Reacts with octane bis(sulfur dioxide) adduct (DABSO) and iodine to give sulfamide Get E.E. [ka]

[0046] Intermediates N, O, P, S, U, V, W, X, Y, CC, DD, and E in Schemes 2-5 Global deprotection of E gives the compound of formula I'. Pg 1 ) is -BOC, and the ester (Pg in Schemes 1 to 5) 2 ) is tert-butyl If the ester is an ester, the overall deprotection can be carried out using diethyl ether, dioxane, or isopropyl alcohol. This is accomplished in one step using a solution of HCl in an organic solvent such as propanol. The pyrrolidine nitrogen in the compound of formula I' is then reacted with paraformaldehyde and triacetamide. The methylation can be carried out by reductive amination with sodium borohydride.

[0047] Preparation 1 tert-Butyl (3R)-3-[2-[(4S)-4-benzyl-2-oxo-oxa Zolidine-3-yl]-2-oxo-ethyl]pyrrolidine-1-carboxylate [ka] Triethylamine (56.5 g, 77.9 mL, 559 mmol, 2.5 equiv.) was added to 1 2-[(3R)-1-tert-butoxycarbonyl] A solution of 1.05 equiv. After 5 minutes, pivaloyl chloride (33.7 g, 34.2 mL, 279 mmol, After 15 min, lithium chloride (11. 8 g, 279 mmol, 1.25 eq.) and (4S)-4-benzyloxazolidine- 2-one (40.0 g, 223 mmol, 1 equiv.) is added. The mixture is allowed to warm to room temperature. Stir for 24 hours. After 24 hours, add 1N aqueous HCl (500 mL) and the organic phase The organic phase is separated from the aqueous phase. The organic phase is washed with 1N aqueous NaOH (500 mL) and saturated aqueous NaCl. solution (500 mL), dried over MgSO4, filtered, and the solution was concentrated in vacuo. The residue is suspended in a mixture of MeOH / water (1:2, 575 mL) and stirred at room temperature overnight. The solid was filtered off, washed with hexane (2×150 mL), and the solid was dried to give the title compound. The product (65.7 g, 76%) was obtained. ES / MS (m / z): 333 (M+H-tert -butyl), 1 H NMR (400.13MHz, CDCl3)δ7.38-7.28( m, 3H), 7.25-7.20(m, 2H), 4.73-4.70(m, 1H), 4. 27-4.19(m, 2H), 3.75-3.66(m, 1H), 3.55-3.48( m, 1H), 3.38-3.30(m, 2H), 3.11-2.96(m, 3H), 2. 84-2.76(m, 1H), 2.74-2.65(m, 1H), 2.14-2.11( m, 1H), 1.64-1.58(m, 1H), 1.49(s, 9H).

[0048] Preparation 2 tert-Butyl (3S)-3-[2-[(4R)-4-benzyl-2-oxo-oxa Zolidine-3-yl]-2-oxo-ethyl]pyrrolidine-1-carboxylate [ka] 2-[(3S)-1-tert-butoxycarbonylpyrrolidin-3-yl]acetic acid and and (4R)-4-benzyloxazolidin-2-one, essentially as described in Preparation 1. Prepare the title compound as described. Use a gradient of 10-50% EtOAc in hexanes. The product is purified by silica gel chromatography using ES / MS (m / z ):333(M+H-tert-butyl)

[0049] Preparation 3 tert-Butyl(3R)-3-(2-methoxy-2-oxo-ethyl)pyrrolidine-1 -Carboxylate [ka] Iodomethane (2 mol / L in MTBE, 240 mL, 480 mmol, 1.1 equiv.) of 2-[(3R)-1-tert-butoxycarbonylpyrrolidone] in DMF (800 mL) Lysine-3-yl]acetic acid (100 g, 436 mmol) at room temperature. sodium (90.4 g, 654 mmol, 1.5 equiv.) was added and the resulting mixture was stirred for 4 h. Stir. Add water (1.5 L) and extract with MTBE (3 L). The organic phase is washed with ice / water (3 × 500 mL), the organic phase was dried over MgSO4, filtered and concentrated under reduced pressure to give The title compound is obtained (103 g, 97%). 1 H NMR (300 MHz, CDCl3) δ 3.68(S, 3H), 3.65-3.53(m, 1H), 3.52-3.36(m, 1 H), 3.35-3.23(m, 1H), 3.03-2.84(m, 1H), 2.64- 2.49(m, 1H), 2.45-2.30(m, 2H), 2.13-1.97(m, 1 H), 1.65-1.48(m, 1H), 1.45(S, 9H).

[0050] Preparation 4 tert-Butyl (3R)-3-[1-[(3-bromophenyl)methyl]-2-methoxy 2-Oxo-ethyl]pyrrolidine-1-carboxylate [ka] tert-Butyl (3R)-3-(2-methoxy- Solution of 2-oxo-ethyl)pyrrolidine-1-carboxylate (7 g, 28 mmol) to 300 ml of lithium bis(trimethylsilyl)amide (1 M solution in THF, 33.5 mL, 3 The mixture was stirred at -78°C for 1 h and 3-bromide was added. Bromobenzyl (8.37 g, 33.5 mmol, 1.2 equiv.) in THF (5 mL) Add 100 ml of 100% NaCl. Allow the mixture to warm to room temperature and stir overnight. The organics were washed with saturated aqueous NaCl and concentrated over MgSO4. Dry, filter and evaporate to dryness. A gradient of 10-40% EtOAc in hexanes is added. The residue was purified by silica gel chromatography using A mixture of omers (8.4 g, 73%) is obtained as a yellow oil. ES / MS (m / z): 356, 358 (M+H-tert-butyl).

[0051] Preparation 5 3-(3-bromophenyl)-2-[(3R)-1-tert-butoxycarbonylpyrrolidone Lysine-3-yl]-2-methyl-propanoic acid [ka] tert-Butyl (3R)-3-[ 1-[(3-bromophenyl)methyl]-2-methoxy-2-oxo-ethyl)pyrrolidine A solution of lithium bis(trimethylsilyl) (ethylsilyl)amide (1 M solution in THF, 41 mL, 41 mmol, 2 equiv.) was added. The reaction is stirred at -78 °C for 2 h. Iodomethane (25 mL, 410 mmol, 2 0 equiv.) and allow the mixture to warm to room temperature. Stir the mixture overnight. Saturated NH4Cl aqueous solution Add solution and extract with EtOAc. Wash the organic layer with saturated aqueous NaCl and MgSO 4, filtered and the solvent removed in vacuo. The residue was dissolved in MeOH (80 mL) and Dissolve in THF (80 mL) and then add sodium hydroxide (5 M aqueous solution, 81 mL, 41 0 mmol, 20 equiv.) is added and the resulting mixture is heated at 60° C. for 3 days. Cool to room temperature and add HCl (1N aqueous solution) to adjust the pH of the mixture to 2-3. The organic layer is extracted with EtOAc. The organic layer is dried over MgSO4, filtered, and concentrated in vacuo. The residue was subjected to chiral SFC under the following parameters: Column - Chiralp ak® AD (25×3 cm, 5 μm), mobile phase—solvent A=CO2, solvent B=M eOH + 0.2% v / v DMEA, gradient - isocratic 80:20 A:B, flow rate -120 mL / min). Isomer 1 (1.7 g, 27%) and Isomer 2 (3. 4g, 41%) is obtained as a white solid. ES / MS (m / z): 356 / 358 (M+ H-tert-butyl).

[0052] Preparation 6 tert-Butyl (3R)-3-[(1S)-2-[(4S)-4-benzyl-2-oxo So-oxazolidin-3-yl]-1-[(3-bromophenyl)methyl]-2-oxo -Ethyl]pyrrolidine-1-carboxylate [ka] Lithium bis(trimethylsilyl)amide (1M in THF, 818 mL, 818 mmol) l, 1.2 equiv.) in a 3 L three-neck round-bottom flask in an ice bath under nitrogen in THF. tert-Butyl (3R)-3-[2-[(4S)-4-benzyl] -2-oxo-oxazolidin-3-yl]-2-oxo-ethyl]pyrrolidine-1-carboxylate To a solution of carboxylate (265 g, 682 mmol) at 0 °C, 2 The mixture is stirred at 0.6° C. for 47 min. Then THF (450 ml) is added. 1-Bromo-3-(bromomethyl)benzene (190 g, 760 mmol, 1. A solution of 12 eq. of 1,2-dichlorophenyl ether (NaOH) was added over 28 min and the reaction temperature was allowed to rise to 5.1° C. Stir overnight. Allow the mixture to warm to room temperature while stirring. Use an ice / water bath to cool the reaction mixture and then Add a saturated aqueous solution of NH4Cl (1 L) in 4 portions at such a rate as to keep the temperature below 21°C. The mixture was added with water (1 L) and extracted with MTBE (3.5 L). (1 L) and saturated aqueous NaCl (500 mL), then washed with saturated aqueous NaCl The organics are dried over Na2SO4, filtered and then washed with 100 mL of ethyl acetate. Concentrate in vacuum. Add hexane (1 L) to the residue, concentrate in vacuum, and then reconstitute under high vacuum. The title compound was obtained as an orange oil (416 g, >100%) after drying at 4°C for 10 min at 4°C for 2 h. The purity is estimated to be 90% by weight based on theoretical yield. ES / MS (m / z): 501 / 503 (M+H-tert-butyl).

[0053] Preparation 7 tert-Butyl (3S)-3-[(1R)-2-[(4R)-4-benzyl-2-oxo So-oxazolidin-3-yl]-1-[(3-bromophenyl)methyl]-2-oxo -Ethyl]pyrrolidine-1-carboxylate [ka] The title compound was synthesized by the reaction of tert-butyl(3S)-3-[2-[(4R)-4-benzyl-2 -Oxo-oxazolidin-3-yl]-2-oxo-ethyl]pyrrolidine-1-carbo Prepare the carboxylate essentially as described in Preparation 6. ES / MS (m / z): 501, 503 (M+H-tert-butyl).

[0054] Preparation 8 tert-Butyl (3R)-3-[(1S)-2-[(4S)-4-benzyl-2-oxo So-oxazolidin-3-yl]-1-[(3-nitrophenyl)methyl]-2-oxo -Ethyl]pyrrolidine-1-carboxylate [ka] Lithium bis(thiomethylsilyl)amide (1.0 M in THF, 46 mL, 46 mmol) , 1.2 equiv.) in THF (75 mL) 4S)-4-Benzyl-2-oxo-oxazolidin-3-yl]-2-oxo-ethyl ]pyrrolidine-1-carboxylate (15 g, 39 mmol, 1 equiv.) was added to a solution of -2 The mixture was stirred at -20°C for 20 min. 1-( Bromomethyl)-3-nitrobenzene (9.17 g, 42.5 mmol, 1.1 equiv.) Add the solution. Stir the solution for 2 hours and allow it to warm to room temperature. Dilute the mixture with MTBE and add saturated Quench with aqueous NH4Cl. Separate the phases and extract the aqueous phase with MTBE. Organic Extraction Combine the organics and wash the organics successively with water and saturated aqueous NaCl. Dry over Na2SO4. The residue is dissolved in MeOH / H2O (2:1, Mix with 150 mL of hexane. Stir the slurry overnight. Filter to collect the solids and The solid is dried under vacuum at 40° C. to give the title compound (19 g, 88%). ES / MS(m / z): 468(M+H-tert-butyl).

[0055] Preparation 9 tert-Butyl (3R)-3-[(1S)-1-benzyl-2-[(4S)-4-benzyl Dimethyl-2-oxo-oxazolidin-3-yl]-2-oxo-ethyl]pyrrolidine-1 -Carboxylate [ka] Benzyl bromide was used to prepare 76% pure 1,2-dichloro-1,2-diphenylmethane (1,2-dichloro-1,2-diphenylmethane) essentially as described in Preparation 6. The title compound is prepared: ES / MS (m / z): 423 (M+H-tert-butyl).

[0056] Preparation 10 (2S)-3-(3-bromophenyl)-2-[(3R)-1-tert-butoxy Bornylpyrrolidin-3-yl]propanoic acid [ka] A solution of hydrogen peroxide (0.88 M in water, 105 mL, 926 mmol, 1.5 equiv.) was added to tert-Butyl (3R)-3-[(1S)-2-[(4S)-4- Benzyl-2-oxo-oxazolidin-3-yl]-1-[(3-bromophenyl)methyl [ethyl]-2-oxo-ethyl]pyrrolidine-1-carboxylate (purity 90% by weight, 3 81 g, 615 mmol) in one portion to a mechanically stirred mixture equipped with an ice / water bath. Cool a 12 L three-neck round-bottom flask to 8.6°C. A solution of tetrahydrofuran monohydrate (38.7 g, 923 mmol, 1.5 equiv) was added over 25 min. The reaction temperature is increased to 12.8° C. The reaction is stirred for 2.5 hours and then cooled to 5.7° C. Sodium bisulfite (129.4 g, 1244 mmol, 2.02 mmol) in water (2 L) A solution of NaOH (5 eq.) is added over 40 min and the reaction temperature is allowed to rise to 14.7°C. The pH of the reaction mixture was raised to >12 by adding an aqueous solution of 100% Methylcellulose (1 L) and 100% Methylcellulose (1 L). Add BE (4 L). Separate the layers and extract the aqueous layer with MTBE (2 L). Combine and extract with water (1 L) and then add this aqueous extract to the bulk aqueous solution. Stir with MTBE (3 L) and cool the mixture to 5° C. Add an aqueous solution of hydrochloric acid (5 N) The pH of the mixture is brought to 3. Separate the layers and add saturated aqueous NaCl (1 L) and water (50 Wash the organic layer with a mixture of 100 mL of ethyl acetate (NaSO4). Dry the organics over Na2SO4, filter, and Concentrate in vacuo at 40° C. The residue is dried under high vacuum to give the title compound (221.5 g , 90%) as a white solid. ES / MS (m / z): 342 / 344 (M+H -tert-butyl).

[0057] Preparation 11 Ammonium;(2S)-3-(3-bromophenyl)-2-[(3R)-1-tert -Butoxycarbonylpyrrolidin-3-yl]propanoate [ka] (2S)-3-(3-bromophenyl)-2-[(3R)-1-tert-butoxycarbonyl] [Carbonylpyrrolidin-3-yl]propanoic acid (237.6 g, 596.5 mmol) and M Mix with TBE (3801 mL) to obtain a cloudy mixture containing solids. The solution was filtered through a glass fiber filter and diluted with ammonia (7 M) in MeOH (128 mL, 896 mm ol, 1.5 equiv.) is added to the filtrate under nitrogen with mechanical stirring. A white solid precipitates. The mixture becomes very thick. MTBE (800 mL) is added to the mixture to make it free-flowing. The mixture is stirred at room temperature for 1.5 h and then cooled to -5 to 0°C in an ice / acetone bath. Cool to °C and stir for 1.5 h. Transfer the mixture to a propylene mat and a 3 L glass fritted funnel. The solids were filtered by vacuum filtration under reduced pressure, the solids were rinsed with MTBE (1 L), and then nitrogen-purged. Vacuum was continued under the blanket at room temperature overnight. The solid was then subjected to HPLC analysis to obtain (2S)-3-(3 -bromophenyl)-2-[(3R)-1-tert-butoxycarbonylpyrrolidine- Two more batches of material were prepared in a similar manner starting with 47 3-yl]propanoic acid. .3g, 113mmol, and 53.0g, 120mmol). was suspended in acetonitrile (4300 mL) and incubated at 23° C. with mechanical stirring overnight under nitrogen. Stir the mixture with 500 mL of acetonitrile. and MTBE (1 L), then vacuum was applied under a nitrogen blanket at room temperature for 4 h. Continue to use the crude product to obtain the title compound as a white powder (249.5 g, 77%). m / z): 342 / 344 (M+H-tert-butyl).

[0058] Preparation 12 Ammonium;(2R)-3-(3-bromophenyl)-2-[(3S)-1-tert -Butoxycarbonylpyrrolidin-3-yl]propanoate [ka] tert-Butyl(3S)-3-[(1R)-2-[(4R)-4-benzyl-2-o 1-[(3-bromophenyl)methyl]-2-oxo-oxazolidin-3-yl Preparation 10 was essentially as described in Preparation 10 using [so-ethyl]pyrrolidine-1-carboxylate. The title compound was prepared as described in Preparation 11, followed by ammonium ionization as described in Preparation 12. Forms a tert-butyl salt. ES / MS (m / z): 342, 344 (M+H-tert-butyl) .

[0059] Preparation 13 (2S)-2-[(3R)-1-tert-butoxycarbonylpyrrolidin-3-yl] -3-(3-nitrophenyl)propanoic acid [ka] tert-Butyl(3R)-3-[(1S)-2-[(4S)-4-benzyl-2-o 1-[(3-nitrophenyl)methyl]-2-oxo-oxazolidin-3-yl 5-ethylpyrrolidine-1-carboxylate was used to work up the reaction. 1. Prepare a 10-mercaptoethanol solution essentially as described in Preparation 10, omitting the step of adding an aqueous solution of thorium. Prepare the title compound: ES / MS (m / z): 309 (M+H-tert-butyl).

[0060] Preparation 14 (2S)-2-[(3R)-1-tert-butoxycarbonylpyrrolidin-3-yl] -3-phenyl-propanoic acid [ka] tert-Butyl (3R)-3-[(1S)-1-benzyl-2-[(4S)-4-benzyl 2-oxo-2-oxazolidin-3-yl]-2-oxo-ethyl]pyrrolidine- The title compound was prepared essentially as described in Preparation 10 using the 1-carboxylate. The product is purified by trituration with 1:1 MeOH:water. MS(m / z):264(M+H-tert-butyl).

[0061] Preparation 15 (2R)-2-[(3S)-1-tert-butoxycarbonylpyrrolidin-3-yl] -3-phenyl-propanoic acid [ka] Ammonium;(2R)-3-(3-bromophenyl)-2-[(3S)-1-tert-butyl t-Butoxycarbonylpyrrolidin-3-yl]propanoate (830 mg, 2.08 20 mmol) in EtOH (20 mL) and palladium on carbon (10% w / w, 22 2 mg, 0.208 mmol, 0.1 equiv.) is added. The mixture is heated under a balloon of hydrogen. Stir at room temperature overnight. Filter the reaction mixture through a pad of diatomaceous earth. Concentrate the filtrate and add 1% acetic acid. Silica gel chromatography using a gradient of 10-40% in hexane with Purify the residue further to give the title compound (560 mg, 84%) as a white solid. / MS(m / z):264(M+H-tert-butyl).

[0062] Preparation 16 tert-Butyl(3R)-3-[(1S)-1-[(3-bromophenyl)methyl]- 2-tert-Butoxy-2-oxo-ethyl]pyrrolidine-1-carboxylate [ka] Into the reactor, add ammonium; (2S)-3-(3-bromophenyl)-2-[(3R)-1 -tert-butoxycarbonylpyrrolidin-3-yl]propanoate (500 g, 1 210 mmol), 2-methyltetrahydrofuran (4000 mL), then KHSO4 A solution (1M in water, 3000 mL) is added. The mixture is stirred for 30 minutes, during which the pH The reaction mixture was phase-separated and the aqueous layer was diluted with 2-methyltetrahydrofuran ( Extract with 1000 mL of ethyl acetate. Combine the organic phases and wash them with saturated aqueous NaCl solution. Dry the organic phase over MgSO4 and filter it. Transfer the solution to a reactor and add 2-tert -Butyl-1,3-diisopropylisourea (618.2 g, 3024 mmol, 2.5 The mixture is stirred at 65° C. for 3 hours and then further 2-tert-butyl-1, Add 3-diisopropylisourea (247.3 g, 1210 mmol, 1 eq). The mixture is stirred at 65° C. overnight. The mixture is cooled to room temperature. The solid is filtered off and the organic layer is Wash with saturated aqueous NaHCO3 (1000 mL). Dry the organic layer over MgSO4. The residue was added with MTBE (2000 mL) and the solids were filtered off. Concentrate the filtrate to give the title compound (483 g, 88%) as a white solid. S / MS(m / z):342 / 344(M+H-2×tert-butyl)

[0063] Preparation 17 tert-Butyl (3R)-3-[1-[(3-bromophenyl)methyl]-2-tert-butyl t-Butoxy-1-methyl-2-oxo-ethyl]pyrrolidine-1-carboxylate [ka] 3-(3-bromophenyl)-2-[ (3R)-1-tert-butoxycarbonylpyrrolidin-3-yl]-2-methyl- A solution of isopropyl alcohol, isomer 2 (3.4 g, 8.2 mmol) was added to the solution of 2-tert-butyl- ,3-Diisopropylisourea (5.1 g, 5.7 mL, 25 mmol, 3 equiv.) was added. The mixture is heated at 55° C. for 3.5 hours. Additional 2-tert-butyl-1,3-diisopropyl Isopropylisourea (5.1 g, 5.7 mL, 25 mmol, 3 equiv.) was added and the mixture was heated at 55° C. Continue heating the reaction at RT for 1.5 h. Add additional 2-tert-butyl-1,3-diisopropyl Add pyrisourea (2.5 g, 2.9 mL, 12 mmol, 1.5 equiv.) and heat at 55 °C. The reaction was heated overnight. The white solid was filtered off and washed with MTBE, and the filtrate was evaporated. Evaporate to dryness. Silica gel chromatography using a gradient of 0-100% EtOAc in hexanes. The residue was purified by chromatography to give the title compound (3.6 g, 93%) as a yellow oil. ES / MS(m / z): 356, 358 [M+H-(2×tert-butyl )].

[0064] Preparation 18 tert-Butyl (3R)-3-[(1S)-2-tert-butoxy-1-[(3-di 2-Oxo-ethyl-1-phenylmethylpyrrolidine-1-carboxylate [ka] (2S)-2-[(3R)-1-tert-butoxycarbonylpyrrolidin-3-yl ]-3-(3-nitrophenyl)propanoic acid in toluene as the reaction solvent. Prepare the title compound essentially as described in Preparation 17, carrying out the reaction at 80° C. Silica gel chromatography using a gradient of 5-20% EtOAc in hexanes. The crude product was purified by HPLC. ES / MS (m / z): 309 [M+H-(2×tert-butyl ether] t-butyl).

[0065] Preparation 19 tert-Butyl (3R)-3-[(1S)-2-tert-butoxy-1-[(3-H 2-(2-oxo-ethyl)pyrrolidine-1-carboxylate [ka] In a Parr reactor, tert-butyl ether (3R) in toluene (3000 mL) was added. -3-[(1S)-1-[(3-bromophenyl)methyl]-2-tert-butoxy- 2-Oxo-ethyl]pyrrolidine-1-carboxylate (300g, 660mmol) followed by a solution of palladium(II) acetate (7.41 g, 33.0 mmol, 0.05 equiv. ), butyldi-1-adamantylphosphine (24.92 g, 66.02 mmol, 0. 1 equivalent) and N,N,N',N'-tetramethylethylenediamine (115 g, 149 1 mL, 990 mmol, 1.5 equiv.) and the mixture was pressurized with 70 psi of syngas (CO Pressurize with 1:1 H2O / H2 and stir at 100°C overnight. Cool the mixture to room temperature and evaporate the solvent. The residue was dissolved in EtOAc and filtered through silica gel to give the title compound (273 g, 97%) as an orange oil. ES / MS (m / z): 426 (M+N a).

[0066] Preparation 20 tert-Butyl (3R)-3-[2-tert-butoxy-1-[(3-formylphenyl) Nyl)methyl]-1-methyl-2-oxo-ethyl]pyrrolidine-1-carboxylate [ka] tert-Butyl (3R)-3-[1-[(3-bromophenyl)methyl]-2-te rt-Butoxy-1-methyl-2-oxo-ethyl]pyrrolidine-1-carboxylate Prepare the title compound essentially as described in Preparation 19 using hexa The crude product was purified by silica gel chromatography using a gradient of 10-30% acetone in ethanol. The product was purified. ES / MS (m / z): 262 (M+H-tert-butyl-BOC) .

[0067] Preparation 21 tert-Butyl (3R)-3-[(1S)-2-tert-butoxy-1-[[3-[ Hydroxyiminomethyl]phenyl]methyl]-2-oxo-ethyl]pyrrolidine-1- Carboxylate [ka] tert-Butyl(3R)-3-[(1S)-2-tert-butoxy-1-[(3- Formylphenyl)methyl]-2-oxo-ethyl]pyrrolidine-1-carboxylate (50 g, 105 mmol, 85% purity), EtOH (400 mL), pyridine (17. 0 mL, 211 mmol, 2 eq.) and hydroxylamine hydrochloride (10.98 g, 1 Mix 58.0 mmol, 1.5 eq.) and stir the mixture at room temperature for 2 h. Allow to evaporate, add KHSO4 (1M aqueous solution, 300 mL) and MTBE (500 mL). The organic layer is separated and washed with saturated aqueous NaHCO3, filtered, and concentrated over MgSO4. Dry at 40° C. Filter through Celite® and concentrate to dryness to give a yellow oil The title compound is obtained as 49 g (98% yield, 88% w / w purity). z): 319 (M+H-Boc)

[0068] Preparation 22 tert-Butyl(3R)-3-[(1S)-1-[[3-(aminomethyl)phenyl] Methyl]-2-tert-butoxy-2-oxo-ethyl]pyrrolidine-1-carboxy rate [ka] Sponge nickel catalyst (2 g) and glass beads (6 g, 212-300 microns) A stainless steel packed bed reactor column (1.2 cm inner diameter × 10 cm length) containing Prepare a lowCAT high pressure flow chemistry reactor (HEL Ltd). The reactor was equipped with gas and liquid flow controllers and a heating jacket around the column. The following parameters were used to flush the column with MeOH and hydrogen gas: Equilibrate the ram: Liquid flow rate-4mL / min, H2 flow rate-60mL / min, pressure-50 bar The reactor column jacket temperature was -120°C (the internal reaction temperature was maintained at 50°C).

[0069] tert-Butyl (3R)-3-[(1S)-2-tert-butoxy-1-[[3- [Hydroxyiminomethyl]phenyl]methyl]-2-oxo-ethyl]pyrrolidine-1 -Carboxylate (49 g, 103 mmol, 88% w / w purity) was dissolved in ammonia (Me OH, 7M solution, 606 mL) and equilibrated with hydrogen gas using column equilibration parameters. and recovering the effluent from the reactor column. The column was flushed again with ammonia (7 M solution in MeOH) for 20 min, and the reactor column was The reactor column eluate fractions are combined and the mixture is concentrated in vacuo to give The title compound is obtained as an oil (46.9 g, 88% yield, purity 78% w / w). MS(m / z): 405(M+H).

[0070] Preparation 23 tert-Butyl(3R)-3-[(1S)-1-[(3-aminophenyl)methyl]- 2-tert-Butoxy-2-oxo-ethyl]pyrrolidine-1-carboxylate [ka] Palladium on carbon (10% w / w, 380 mg, 0.36 mmol, 0.05 equiv.) of tert-butyl (3R)-3-[(1S)-2-tetramethylphenyl]phenylacetamide in EtOAc (71 mL) rt-Butoxy-1-[(3-nitrophenyl)methyl]-2-oxo-ethyl]pyrrol To a stirred solution of 1,2-diphenyl-1-carboxylate (3 g, 7.1 mmol) was added under a nitrogen atmosphere. The mixture is purged with hydrogen and stirred under a balloon of hydrogen at room temperature overnight. The reaction mixture is filtered through a pad of algal earth. The filtrate is concentrated in vacuo to give the title compound (2.62 g, 94%) as a white solid. ES / MS (m / z): 291 (M+H-BO C).

[0071] Preparation 24 tert-Butyl (3R)-3-[(1S)-2-tert-butoxy-1-[[3-( Hydroxymethyl)phenyl]methyl]-2-oxo-ethyl]pyrrolidine-1-carbo Xylate [ka] Sodium borohydride (0.208 g, 5.50 mmol, 1.2 equiv.) was dissolved in MeO tert-Butyl (3R)-3-[(1S)-2-tert-butyl]acetate in H (25 mL) 1-[(3-formylphenyl)methyl]-2-oxo-ethyl]pyrrolidine-1- Add to a solution of carboxylate (1.85 g, 4.58 mmol) at 0° C. After 10 min. The solvent is evaporated. Saturated aqueous NaHCO3 is added and the aqueous layer is extracted with EtOAc. Wash the organics with water and saturated aqueous NaCl. Dry the organics over MgSO4. Concentrate in vacuo to give the title compound (2.94 g, 97.5%) as a colorless oil. / MS(m / z):428(M+Na).

[0072] Preparation 25 [3-[(2S)-3-tert-butoxy-2-[(3R)-1-tert-butoxy Carbonylpyrrolidin-3-yl]-3-oxo-propyl]phenyl]boronic acid [ka] tert-Butyl(3R)-3-[(1S)-1-[(3-bromophenyl)methyl] -2-tert-butoxy-2-oxo-ethyl]pyrrolidine-1-carboxylate ( 16.4g, 36.1mmol), tetrahydroxydiboron (5.00g, 54.1m mol, 1.5 equivalents), chloro(2-dicyclohexylphosphino-2',4',6'- Triisopropyl-1,1'-biphenyl) [2-(2'-amino-1,1'-biphenyl aryl)]palladium(II) (XPhos Pd G2, 0.145g, 0.180mm ol, 0.005 equivalents), 2-dicyclohexylphosphino-2',4',6'-triyl isopropylbiphenyl (XPhos, 0.176 g, 0.361 mmol, 0.01 equiv. ), potassium acetate (10.6 g, 108 mmol, 3 equiv.), EtOH (246 mL) and and ethylene glycol (6.10 mL, 108 mmol, 3 equiv.). Purge for 5 min. Stir the mixture at 90° C. overnight. Cool the reaction to room temperature and add 2-methyl Add tetrahydrofuran (158 mL) and filter. Concentrate and further purify. Instead, use the residue from preparation 26. ES / MS (m / z): 308 (M+H-2x t ert-butyl).

[0073] Preparation 26 tert-Butyl (3R)-3-[(1S)-2-tert-butoxy-1-[(3-butyl hydroxyphenyl)methyl]-2-oxo-ethyl]pyrrolidine-1-carboxylate [ka] [3-[(2S)-3-tert-butoxy-2-[(3R) -1-tert-butoxycarbonylpyrrolidin-3-yl]-3-oxo-propyl] A solution of 1,2-diphenylboronic acid (3.8 g, 8.4 mmol) was added to a solution of hydrogen peroxide (3 mL in water). 0%, 9.1 mL, 84 mmol, 10 equiv.) and stir the reaction at room temperature overnight. A solution of sodium bisulfite (20% w / v in water, 120 mL) was added to the reaction. The mixture was slowly quenched. Extraction was performed twice with EtOAc and the combined organics were washed with saturated Na Wash with aqueous Cl. Dry the organics over Na2SO4, filter, and concentrate in vacuo. Silica gel chromatography using a gradient of 10-40% acetone in hexanes was used. The residue was purified by HPLC to give the title compound (3.05 g, 90%) as a beige solid. ES / MS(m / z):280(M+H-2x tert-butyl)

[0074] Preparation 27 tert-Butyl (3R)-3-[(1S)-2-tert-butoxy-1-[[3-[ [[[3-[(2S)-3-tert-butoxy-2-[(3R)-1-tert-butoxy 3-Oxo-propyl]phenyl]methyl]a Amino]methyl]phenyl]methyl]-2-oxo-ethyl]pyrrolidine-1-carboxy rate [ka] tert-Butyl(3R)-3-[(1S)-2-tert-butoxy-1-[(3- Formylphenyl)methyl]-2-oxo-ethyl]pyrrolidine-1-carboxylate (48 g, 100 mmol, 85% w / w purity, 1.1 equiv.), isopropanol (32 8 mL) and tert-butyl (3R)-3-[(1S)-1-[[3-(aminomethyl (phenyl)methyl)-2-tert-butoxy-2-oxo-ethyl)pyrrolidine 1-Carboxylate (46.9 g, 93 mmol, 78% purity, 1 equiv.) was placed in a round-bottom flask. The mixture is stirred at room temperature for 1 hour. The mixture is cooled to 0°C and triacetoxy water is added. Sodium borohydride (59 g, 280 mmol, 3 equiv.) was added and the mixture was stirred at room temperature for 2 Stir for 3 days. Remove the solvent under reduced pressure. Water (200 mL), saturated aqueous NaHCO3 ( Add 100 mL of hexane (300 mL) and EtOAc (500 mL). Separate the organic layer and dilute it in hexane. The mixture was purified by silica gel chromatography using a gradient of 50–100% EtOAc. The title compound is obtained as a colorless oil (36 g, 42% yield, purity 85% w / w ). ES / MS(m / z): 792(M+H).

[0075] Preparation 28 tert-Butyl(3R)-3-[(1S)-1-[[3-[[bis[[3-[(2S) -3-tert-butoxy-2-[(3R)-1-tert-butoxycarbonylpyrrolidone] phenyl]methyl]amino]methyl]phenyl 2-tert-butoxy-2-oxo-ethyl]pyrrolidine-1-carboxamide xylate and tert-butyl (3R)-3-[(1S)-2-tert-butoxy -1-[[3-[[[3-[(2S)-3-tert-butoxy-2-[(3R)-1 -tert-butoxycarbonylpyrrolidin-3-yl]-3-oxo-propyl]phen Nyl]methyl]amino]methyl]phenyl]methyl]-2-oxo-ethyl]pyrrolidine -1-carboxylate [ka] A method for preparing the title compounds as a mixture and then separating them by chromatography. 1 In a round-bottom flask, add tert-butyl (3R)-3-[(1S)-2-tert-butoxy] 1-[(3-formylphenyl)methyl]-2-oxo-ethyl]pyrrolidine-1- Carboxylate (222 g, 550 mmol), 2-propanol (888 mL) and and ammonia solution (2M in 2-propanol, 302.6 mL, 605.2 mmol, 1 Add 1 eq. of triacetoxyborohydride. Cool the mixture to 0-5°C in an ice-water bath. Sodium chloride (116.6 g, 550.2 mmol, 1 equiv.) was added in four portions, The portions are added at 40 min intervals. The mixture is stirred at room temperature overnight. The solvent is evaporated to dryness. To the residue, water (200 mL) and K2HPO4 aqueous solution (300 mL) were added, and MTB Extract the aqueous layer with E (2 x 500 mL). Dry the organic layer over MgSO4, filter and concentrate. Dry. Chromatograph on silica gel using a gradient of 20-80% EtOAc in hexanes. The residue was purified by HPLC to give tert-butyl (3R)-3-[(1S)-1- [[3-[[biS[[3-[(2S)-3-tert-butoxy-2-[(3R)-1 -tert-butoxycarbonylpyrrolidin-3-yl]-3-oxo-propyl]phen Nyl]methyl]amino]methyl]phenyl]methyl]-2-tert-butoxy-2-o xo-ethyl]pyrrolidine-1-carboxylate (57.8 g, 27%) as a white solid Obtain as. 1 H-NMR (400MHz, CDCl3)δ7.30-7.24(m, 6 H), 7.12(S, 3H), 7.04(S, 3H), 3.75-3.43(m, 12H) ), 3.30-3.21(m, 3H), 3.10-2.96(m, 3H), 2.89-2 .76(m, 6H), 2.49(d, J=4.7Hz, 3H), 2.37(dd, J=7 .2, 14.4Hz, 3H), 1.98-1.90(m, 3H), 1.74-1.61( m, 3H), 1.48(S, 27H), 1.22(S, 27H).

[0076] From the above silica gel chromatography, tert-butyl(3R)-3-[(1S )-2-tert-butoxy-1-[[3-[[[[3-[(2S)-3-tert-butoxy Tox-2-[(3R)-1-tert-butoxycarbonylpyrrolidin-3-yl]- 3-Oxo-propyl]phenyl]methyl]amino]methyl]phenyl]methyl]-2- oxo-ethyl]pyrrolidine-1-carboxylate (82.50 g, 34%) was also Obtained as a colorless oil. ES / MS (m / z): 792 (M+H), HPLC: 90 wt. The purity in % is indicated.

[0077] tert-Butyl(3R)-3-[(1S)-1-[[3-[[bis[[3-[(2S) -3-tert-butoxy-2-[(3R)-1-tert-butoxycarbonylpyrrolidone] phenyl]methyl]amino]methyl]phenyl 2-tert-butoxy-2-oxo-ethyl]pyrrolidine-1-carboxamide Method 2 for the preparation of xylates In a round-bottom flask, add tert-butyl (3R)-3-[(1S)-2-tert-butoxy] -1-[[3-[[[3-[(2S)-3-tert-butoxy-2-[(3R)-1 -tert-butoxycarbonylpyrrolidin-3-yl]-3-oxo-propyl]phen Nyl]methyl]amino]methyl]phenyl]methyl]-2-oxo-ethyl]pyrrolidine -1-carboxylate (90% wt. purity, 81.5 g, 92.6 mmol), ter t-Butyl (3R)-3-[(1S)-2-tert-butoxy-1-[(3-formyl Phenyl)methyl]-2-oxo-ethyl]pyrrolidine-1-carboxylate (purity 8 5 wt%, 50.6 g, 106 mmol, 1.15 equiv), 2-propanol (652 m L) and acetic acid (5.31 mL, 92.6 mmol, 1 equiv.), and the mixture was added for 30 min. Stir. Sodium triacetoxyborohydride (2 equiv., 185 mmol, 39.3 g) is added to the mixture and stirred at room temperature for 2 hours, then the reaction mixture is concentrated in vacuo. Water (200 mL) and MTBE (300 mL) were added to the residue, followed by concentrated ammonium hydroxide. Add aqueous sodium hydroxide to adjust the pH to 9-10. Separate the organic phase and dry over MgSO4. The mixture is then filtered and concentrated to dryness. A gradient of 20-40% EtOAc in hexanes is used. The residue was purified by silica gel chromatography to give tert-butyl (3R)-3 -[(1S)-1-[[3-[[biS[[3-[(2S)-3-tert-butoxy- 2-[(3R)-1-tert-butoxycarbonylpyrrolidin-3-yl]-3-oxo so-propyl]phenyl]methyl]amino]methyl]phenyl]methyl]-2-tert -Butoxy-2-oxo-ethyl]pyrrolidine-1-carboxylate (89g, 82% ) as a white solid.

[0078] Preparation 29 tert-Butyl (3R)-3-[(1S)-2-tert-butoxy-1-[[3-[ [[3-[(2S)-3-tert-butoxy-2-[(3R)-1-tert-butoxy Dicarbonylpyrrolidin-3-yl]-3-oxo-propyl]phenyl]methyl-[( 3-Fluoro-5-methoxy-phenyl)methyl]amino]methyl]phenyl]methyl] -2-oxo-ethyl]pyrrolidine-1-carboxylate [ka] tert-Butyl (3R)-3-[(1S)-2-tert-butoxy-1-[[3- [[[[3-[(2S)-3-tert-butoxy-2-[(3R)-1-tert-butoxy 3-(3-oxo-propyl)phenyl)methyl Amino]methyl]phenyl]methyl]-2-oxo-ethyl]pyrrolidine-1-carboxamide Silane (36 g, 39 mmol, 85% purity), 3-fluoro-5-methoxybenzaldehyde aldehyde (6.684 g, 42.49 mmol, 1.1 equiv.), isopropanol (28 8 mL) and sodium triacetoxyborohydride (16.38 g, 77.26 mm ol, 2 eq.) and stir the mixture at room temperature for 2 h. Evaporate the solvent. Saturated Na Aqueous HCO3 (500 mL) is added and the aqueous layer is extracted with EtOAc (500 mL). The organic layer is dried over MgSO4, then filtered and concentrated to dryness. Purification by silica gel chromatography using a gradient of 40% EtOAc gave the compound shown in Table The title compound is obtained as a colorless oil (32 g, 76% yield, purity 85% w / w). MS(m / z):931(M+H).

[0079] Preparation 30 tert-Butyl (3R)-3-[1-[[3-[[bis[[3-[3-tert-butanoate 2-[(3R)-1-tert-butoxycarbonylpyrrolidin-3-yl]-2 -Methyl-3-oxo-propyl]phenyl]methyl]amino]methyl]phenyl]methyl [2-tert-butoxy-1-methyl-2-oxo-ethyl]pyrrolidine-1-carboxylate Ruboxylate [ka] tert-Butyl (3R)-3-[2-tert-butoxy-1-[(3-formyl) Phenyl)methyl]-1-methyl-2-oxo-ethyl]pyrrolidine-1-carboxylate Prepare the title compound essentially as described in Preparation 28 using hexanes. The elution was performed by silica gel chromatography using a gradient of 20-40% EtOAc in water. Purify the crude material. 1 H NMR (400.21MHz, d6-DMSO): δ7.2 7-7.19(m, 6H), 7.13(S, 3H), 7.08-7.00(m, 3H), 3.57-3.24 (m, 9H, in solvent), 3.22-2.98 (m, 9H), 2.63 -2.38(m, 9H, under solvent), 1.86-1.68(m, 6H), 1.40(S, 2 7H), 1.31(S, 27H), 0.93(S, 9H).

[0080] Preparation 31 tert-Butyl (3R)-3-[(1S)-2-tert-butoxy-1-[[3-[ 3-[(2S)-3-tert-butoxy-2-[(3R)-1-tert-butoxy] [Carbonylpyrrolidin-3-yl]-3-oxo-propyl]phenoxy]phenyl]methyl [2-oxo-ethyl]pyrrolidine-1-carboxylate [ka] tert-Butyl(3R)-3-[(1S)-1-[(3-bromophenyl)methyl] -2-tert-butoxy-2-oxo-ethyl]pyrrolidine-1-carboxylate ( 150mg, 0.330mmol), tert-butyl(3R)-3-[(1S)-2- tert-Butoxy-1-[(3-hydroxyphenyl)methyl]-2-oxo-ethyl ]pyrrolidine-1-carboxylate (0.155 g, 0.396 mmol, 1.2 equiv. ), cuprous iodide (6 mg, 0.03 mmol, 0.1 equiv.), N,N-dimethylglycine Hydrochloride (0.0138 g, 0.0990 mmol, 0.3 equiv.) and cesium carbonate ( 0.215 g, 0.660 mmol, 2 eq.) are mixed in DMF (2.3 mL). The mixture is stirred under nitrogen at 110° C. overnight. The reaction is cooled and then passed through a pad of diatomaceous earth. The mixture is filtered and washed with DCM and MeOH. The filtrate is concentrated in vacuo. The resulting residue is purified by RP-HPLC / MS using column X. Bridge(TM) C18 (19 x 100 mm, 5 μm), mobile phase - solvent A = 20 in water mM ammonium bicarbonate (pH 9), solvent B = acetonitrile, gradient - 0:100 to 1 00:0 B:A, flow rate - 25 mL / min. Obtain the title compound (43 mg, 17%). ES / MS(m / z):665(M+H-BOC).

[0081] Preparation 32 tert-Butyl (3R)-3-[(1S)-2-tert-butoxy-1-[[3-[ 2-[3-[(2S)-3-tert-butoxy-2-[(3R)-1-tert-butoxy [oxycarbonylpyrrolidin-3-yl]-3-oxo-propyl]phenoxy]ethoxy ]phenyl]methyl]-2-oxo-ethyl]pyrrolidine-1-carboxylate [ka] tert-Butyl(3R)-3-[(1S)-2-tert-butoxy-1-[(3- Hydroxyphenyl)methyl]-2-oxo-ethyl]pyrrolidine-1-carboxylate Dissolve cesium carbonate (275 mg, 0.70 mmol) in DMF (1.2 mL). (285 mg, 0.86 mmol) and 1,2-dibromoethane (0.030 mL, 0.34 mmol) and stir at room temperature for 3 days. Over the next 9 days, cesium carbonate or Potassium carbonate and 1,2-dibromoethane were added three more times, and the temperature was increased to 70°C and The temperature is gradually increased to 110°C. The mixture is cooled to room temperature and a saturated aqueous solution of NH4Cl is added. The aqueous layer is extracted with EtOAc, and the organic phase is washed with saturated aqueous NaCl and water. Dry the organic phase over MgSO4, filter, and concentrate the solution in vacuo. The residue was purified by silica gel chromatography using a gradient of 0% EtOAc. The title compound (33.4 mg, 11%) is obtained as a colorless oil. z):709(M+H-BOC).

[0082] Preparation 33 tert-Butyl (3R)-3-[(1S)-2-tert-butoxy-1-[[3-[ [3-[(2S)-3-tert-butoxy-2-[(3R)-1-tert-butoxy Carbonylpyrrolidin-3-yl]-3-oxo-propyl]phenoxy]methyl]phen Nyl]methyl]-2-oxo-ethyl]pyrrolidine-1-carboxylate [ka] Triphenylphosphine (0.5733 g, 2.164 mmol, 1.5 equiv.) was added to T tert-Butyl (3R)-3-[(1S)-2-tert-butanoate] in HF (14 mL) 1-[[3-(hydroxymethyl)phenyl]methyl]-2-oxo-ethyl]pi Rolidin-1-carboxylate (585 mg, 1.443 mmol) and tert- Butyl (3R)-3-[(1S)-2-tert-butoxy-1-[(3-hydroxyphenyl) 2-Oxo-ethyl]pyrrolidine-1-carboxamide in (phenyl)methyl)-THF (14 mL) Add to a solution of the carboxylate (0.8472 g, 2.164 mmol, 1.5 equiv). After purging with nitrogen, diethyl azodicarboxylate (0.34 mL, 2.2 mmol, 1 Add 0.5 eq. of 1000 ml dropwise. Stir the mixture overnight. Filter the reaction through a pad of diatomaceous earth. The mixture was washed with DCM and MeOH. The filtrate was concentrated in vacuo. The residue obtained is purified by SFC using a column - Chiracel® OD ( 5μm, 2x25cm), mobile phase-solvent A=CO2, solvent B=MeOH+DMEA(1. 0% v / v), Gradient-Isocratic 80:20 A:B, Flow-80mL / min, Pressure -120 bar, column temperature -40°C. Title compound (315 mg, 28%). ES / MS (m / z): 679 (M+H-BOC).

[0083] Preparation 34 tert-Butyl (3R)-3-[(1S)-2-tert-butoxy-1-[[3-[ 3-[(2S)-3-tert-butoxy-2-[(3R)-1-tert-butoxy] [Carbonylpyrrolidin-3-yl]-3-oxo-propyl]phenyl]sulfanylphenyl Nyl]methyl]-2-oxo-ethyl]pyrrolidine-1-carboxylate [ka] Dry toluene (0.9 mL) and acetone (1.8 mL) were dissolved in tert-butyl(3 R)-3-[(1S)-1-[(3-bromophenyl)methyl]-2-tert-butoxy 1-(2-oxo-ethyl)pyrrolidine-1-carboxylate (796 mg, 1.75 m mol), potassium phosphate tribasic (229 mg, 1.06 mmol), bis(dibenzylidene (diphenylacetone)palladium (50 mg, 0.09 mmol), 1,1'-bis(diphenyl phosphino)ferrocene (70 mg, 0.12 mmol) and potassium thioacetate (10 The resulting mixture was stirred for 5 min under nitrogen atmosphere. Sonicate for 20 min and then stir at 110 °C for 6 h. Add saturated aqueous NH4Cl and EtO Ac is added. The organic layer is washed with water. The organic phase is dried over MgSO4, filtered and Concentrate in vacuo. Silica gel chromatography using a gradient of 0-40% EtOAc in hexanes. The residue is purified by chromatography to give the title compound as a yellow oil (370 mg, 52%). ES / MS(m / z):781(M+H).

[0084] Preparation 35 tert-Butyl (3R)-3-[(1S)-2-tert-butoxy-1-[[3-[ 3-[(2S)-3-tert-butoxy-2-[(3R)-1-tert-butoxy] [Carbonylpyrrolidin-3-yl]-3-oxo-propyl]phenyl]sulfinylphenyl Nyl]methyl]-2-oxo-ethyl]pyrrolidine-1-carboxylate [ka] tert-Butyl (3R)-3-[(1S)-2-tert-butoxy-1-[[3- [3-[(2S)-3-tert-butoxy-2-[(3R)-1-in DCM (2 mL) tert-Butoxycarbonylpyrrolidin-3-yl]-3-oxo-propyl]phenanthroline Nyl]sulfanylphenyl]methyl]-2-oxo-ethyl]pyrrolidine-1-carbo The 3-chloroacetate (111 mg, 0.14 mmol) was dissolved in DCM (2 mL). Add chloroperoxybenzoic acid (34 mg, 0.14 mmol, 1 eq.). Stir at room temperature for 3 h. Add saturated aqueous NaHCO3, followed by DCM and separate the layers. The organic phase is washed with NaOH solution (3% w / v in water) and water. The organic phase is evaporated over MgSO4. Dry with 0-60% EtOAc in hexanes, then filter and concentrate the solution in vacuo. The residue was purified by silica gel chromatography using a gradient of 1,2-dichlorophenyl ether (C) to give the title compound. (97.2 mg, 87%) was obtained as a colorless oil. ES / MS (m / z): 697( M+H-BOC).

[0085] Preparation 36 tert-Butyl (3R)-3-[(1S)-2-tert-butoxy-1-[[3-[ 3-[(2S)-3-tert-butoxy-2-[(3R)-1-tert-butoxy] [Carbonylpyrrolidin-3-yl]-3-oxo-propyl]phenyl]sulfonylphenyl methyl-2-oxo-ethyl pyrrolidine-1-carboxylate [ka] The compound is prepared essentially similarly to that of Preparation 35, except that 2.5 equivalents of 3- Use chloroperoxybenzoic acid. Use a gradient of 20-60% EtOAc in hexane. The crude product was purified by silica gel chromatography using 500 rpm for 10 min to give the title compound (116. 6 mg, 82%) as a white solid. ES / MS (m / z): 713 (M+H- BOC).

[0086] Preparation 37 tert-Butyl (3R)-3-[(1S)-2-tert-butoxy-1-[[3-[ [3-[(2S)-3-tert-butoxy-2-[(3R)-1-tert-butoxy Carbonylpyrrolidin-3-yl]-3-oxo-propyl]anilino]methyl]phenyl methyl-2-oxo-ethyl pyrrolidine-1-carboxylate [ka] Sodium triacetoxyborohydride (543 mg, 2.56 mmol, 2 equiv.) , tert-butyl(3R)-3-[(1S)-1-[(3-aminophenyl)methyl] -2-tert-butoxy-2-oxo-ethyl]pyrrolidine-1-carboxylate ( 500mg, 1.28mmol), tert-butyl(3R)-3-[(1S)-2-t ert-Butoxy-1-[(3-formylphenyl)methyl]-2-oxo-ethyl]pi Rolidin-1-carboxylate (620 mg, 1.54 mmol, 1.2 equiv.) and Add to a solution of acetic acid (220 μL, 3.84 mmol, 3 equiv.) and stir at room temperature for 4 days. Saturated aqueous NaHCO3 is added and the aqueous layer is extracted with DCM. The combined organic layers are washed with Mg Dry over SO4, filter, and concentrate the solution in vacuo. The residue was purified by silica gel chromatography using a gradient of OAc to give the title compound. The product was obtained (850 mg, 72%). ES / MS (m / z): 778 (M+H).

[0087] Preparation 38 (2S)-3-[3-[3-[(2S)-3-tert-butoxy-2-[(3R)-1 -tert-butoxycarbonylpyrrolidin-3-yl]-3-oxo-propyl]aniline 2-[(3R)-1-tert-butoxycarbonylpyrrolidine-3-yl]phenyl]- -yl]propanoic acid [ka] (2S)-3-(3-bromophenyl)-2- in 1,4-dioxane (160 mL) [(3R)-1-tert-butoxycarbonylpyrrolidin-3-yl]propanoic acid (8 g, 20 mmol), tert-butyl(3R)-3-[(1S)-1-[(3-amino Phenyl)methyl]-2-tert-butoxy-2-oxo-ethyl]pyrrolidine-1- Carboxylate (8.628 g, 22.09 mmol, 1.1 equiv.) and potassium carbonate The mixture was heated under nitrogen for 60 Heat at ℃ for 15 minutes. [(2-di-cyclohexylphosphino-3,6-dimethoxy-2 ',4',6'-triisopropyl-1,1'-biphenyl)-2-(2'-amino-1 ,1'-biphenyl)]methanesulfonate palladium(II) BrettPhos Pd G3, 0.371 g, 0.402 mmol, 0.02 equiv.) was added and stirred overnight. Heat the reaction from 0 to 100 °C. Cool the reaction to room temperature, dilute with EtOAc and add aqueous HCl. The pH of the mixture is adjusted to less than 3 by adding Me-THF solution (1N). The layers are extracted three times. The combined organics are washed with saturated aqueous NaCl. The organics are washed with MgSO Dry over 4 and concentrate the solution in vacuo. Gradient of 0-100% EtOAc in hexanes The residue was purified by silica gel chromatography using , 67%). ES / MS(m / z): 608(M+H-BOC)

[0088] Preparation 39 Di-tert-butyl 3,3'-((2S,2'S)-((sulfonylbis(azanediyl) Bis(3,1-phenylene))bis(3-(tert-butoxy)-3-oxopyrene) (3R,3'R)-bis(pyrrolidine-1-carboxylate) to) [ka] 1,4-Diazabicyclo[2.2.2]octane in acetonitrile (12.8 mL) Bis(sulfur dioxide) adduct (DABSO, 0.6279 g, 2.561 mmol, 2 equiv. ) was cooled to 0° C. Iodine (0.4875 g, 1.921 mmol, 1.5 equiv. The mixture is stirred at 0° C. for 15 min, and then tert-butyl (3R)-3- [(1S)-1-[(3-aminophenyl)methyl]-2-tert-butoxy-2-o xo-ethyl]pyrrolidine-1-carboxylate (500 mg, 1.280 mmol) Heat the reaction at 80° C. overnight. Cool the reaction and pass it through a pad of diatomaceous earth. Filter and wash the pad with DCM and MeOH. Concentrate the filtrate in vacuo. The residue was purified by RP-HPLC / MS using the following parameters: Column - XBridge ( Mobile phase-solvent A = 20 mM ammonia bicarbonate aqueous solution (pH 9), Solvent B = acetonitrile, gradient - isocratic 80:20 The title compound (195 mg, 18%) was obtained as a pale yellow solid. ES / MS(m / z): 743(M+H-BOC).

[0089] Example 1 (2S)-3-[3-[[bis[[3-[(2S)-2-carboxy-2-[(3R)- Pyrrolidin-3-yl]ethyl]phenyl]methyl]amino]methyl]phenyl]-2- [(3R)-Pyrrolidin-3-yl]propanoic acid, tetrahydrochloride [ka] A solution of hydrochloric acid (4 M in 1,4-dioxane, 67 mL, 270 mmol, 20 equiv.) tert-Butyl(3R)-3-[(1S)-1-[[3-[[biS[[3-[(2S )-3-tert-butoxy-2-[(3R)-1-tert-butoxycarbonylpyrrolidone Lysin-3-yl]-3-oxo-propyl]phenyl]methyl]amino]methyl]phenyl nyl]methyl]-2-tert-butoxy-2-oxo-ethyl]pyrrolidine-1-carboxamide carboxylate (15.7 g, 13.3 mmol) was added and the mixture was stirred at 40° C. overnight. The mixture is cooled to room temperature and then concentrated to dryness. The residue is dissolved in water (40 mL) and frozen. The resulting solid was dissolved again in water (40 mL) and freeze-dried to give the title compound. The product (8.6 g, 75%) was obtained as a white foam. ES / MS (m / z): 711 (M +H); 1 H-NMR (500 MHz, D2O) δ7.35-7.10 (m, 12H) , 4.25-4.17(m, 6H), 3.55(dd, J=8.1, 11.5 Hz, 3 H), 3.38-3.33(m, 3H), 3.22-3.15(m, 3H), 3.03- 2.87(m, 12H), 2.56-2.45(m, 3H), 2.12-2.08(m, 3H), 1.73-1.63(m, 3H).

[0090] Example 2 (2S)-3-[3-[[bis[[3-[(2S)-2-carboxy-2-[(3R)- Pyrrolidin-3-yl]ethyl]phenyl]methyl]amino]methyl]phenyl]-2- [(3R)-Pyrrolidin-3-yl]propanoic acid [ka] In a round-bottom flask, add tert-butyl (3R)-3-[(1S)-1-[[3-[[bi S[[3-[(2S)-3-tert-butOxy-2-[(3R)-1-tert- Butoxycarbonylpyrrolidin-3-yl]-3-oxo-propyl]phenyl]methyl ]amino]methyl]phenyl]methyl]-2-tert-butoxy-2-oxo-ethyl ]pyrrolidine-1-carboxylate (499.3 g, 423.3 mmol), 1,4- Dioxane (1997 mL), and hydrochloric acid solution (12 M in water, 529.1 mL, 15 equiv. The mixture is stirred at 40° C. for 1 h, and then the mixture is concentrated in vacuo to give 1,4- The dioxane is removed, resulting in an aqueous slurry. The mixture is passed through a propylene filter. Filter to remove insoluble particles. Adjust the pH of the filtrate using a solution of NaOH (2M in water). Adjust the pH to 9-10. Stir the mixture at room temperature overnight. Filter the resulting solids using filter paper. Filter the material slowly (slow filtration, low vacuum). Wash the solid with water and evacuate at 45°C. Dry to obtain the title compound (281 g, 88%) as a white crystalline solid. MS(m / z):711(M+H); 1 H-NMR(500 MHz, DO) δ7.3 3(t, J=7.6Hz, 3H), 7.27(d, J=7.8Hz, 3H), 7.13( d, J=7.8Hz, 3H), 7.09(s, 3H), 4.20(s, 6H), 3.54 (dd, J=7.9, 11.6Hz, 3H), 3.39-3.34(m, 3H), 3.2 3-3.17(m, 3H), 3.02-2.98(m, 3H), 2.84(dd, J=4 .6, 13.7Hz, 3H), 2.76(dd, J=10.6, 13.3Hz, 3H), 2.60(td, J=9.9, 4.8Hz, 3H), 2.48(td, J=17.3, 9 .6Hz, 3H), 2.12-2.07(m, 3H), 1.73-1.65(m, 3H) .

[0091] Example 3 (2S)-3-[3-[[3-[(2S)-2-carboxy-2-[(3R)-pyrrolyl 3-[(3R-phenyl-3-yl)ethyl]phenyl]methylamino]methyl]phenyl]-2 ... )-Pyrrolidin-3-yl]propanoic acid, trihydrochloride [ka] tert-Butyl (3R)-3-[(1S)-2-tert-butoxy-1-[[3- [[[[3-[(2S)-3-tert-butoxy-2-[(3R)-1-tert-butoxy 3-(3-oxo-propyl)phenyl)methyl Amino]methyl]phenyl]methyl]-2-oxo-ethyl]pyrrolidine-1-carboxamide The title compound was essentially prepared using silyl group and 2M HCl in EtO at room temperature. Prepared as described in Example 1. ES / MS (m / z): 480 (M+H).

[0092] Example 4 (2S)-3-[3-[[3-[(2S)-2-carboxy-2-[(3R)-pyrrolyl phenyl]methyl-[(3-fluoro-5-methoxy-phenyl) (3R)-pyrrolidin-3-yl]phenyl Propanoic acid [ka] tert-Butyl (3R)-3-[(1S)-2-tert-butoxy-1-[[3- [[[3-[(2S)-3-tert-butoxy-2-[(3R)-1-tert-butoxy 3-Oxo-propyl]phenyl]methyl-[ (3-Fluoro-5-methoxy-phenyl)methyl]amino]methyl]phenyl]methyl ]-2-oxo-ethyl]pyrrolidine-1-carboxylate to give the title compound. Prepared as described in Example 2. ES / MS (m / z): 618 (M+H).

[0093] Example 5 (2S)-3-[3-[[3-[(2S)-2-carboxy-2-[(3R)-pyrrolyl phenyl]methyl-[(3-fluoro-5-methoxy-phenyl) (3R)-pyrrolidin-3-yl]phenyl Propanoic acid trihydrochloride [ka] tert-Butyl (3R)-3-[(1S)-2-tert-butoxy-1-[[3- [[[3-[(2S)-3-tert-butoxy-2-[(3R)-1-tert-butoxy 3-Oxo-propyl]phenyl]methyl-[ (3-Fluoro-5-methoxy-phenyl)methyl]amino]methyl]phenyl]methyl ]-2-Oxo-ethyl]pyrrolidine-1-carboxylate (541 mg, 0.582 mmol), HCl (2 M solution in diethyl ether, 5.8 mL) and water (0.5 mL ) are mixed. The mixture is stirred at room temperature for 5 hours. The solvent is removed under reduced pressure to give the title compound ( Yield: 434 mg, 102%). ES / MS (m / z): 618 (M+H).

[0094] Example 6 3-[3-[[bis[[3-[2-carboxy-2-[(3R)-pyrrolidin-3-yl ]propyl]phenyl]methyl]amino]methyl]phenyl]-2-methyl-2-[(3 R)-Pyrrolidin-3-yl]propanoic acid, tetrahydrochloride [ka] tert-Butyl (3R)-3-[1-[[3-[[bis[ [3-[3-tert-butoxy-2-[(3R)-1-tert-butoxycarbonyl Pyrrolidin-3-yl]-2-methyl-3-oxo-propyl]phenyl]methyl]amine 2-tert-butoxy-1-methyl-2-oxo- Ethyl]pyrrolidine-1-carboxylate (275 mg, 0.225 mmol) and Add HCl (2M solution in EtO, 3.4 mL, 6.7 mmol, 30 equiv.). The mixture is stirred at room temperature overnight. The reaction solvent is decanted from the white solid and the solid is dissolved in water. The mixture was evaporated to dryness under a stream of nitrogen and then dried in vacuum at 40° C. to give the title compound. The compound (190 mg, 86%) was obtained as a white solid. ES / MS (m / z): 753( M+H).

[0095] Example 7 (2S)-3-[3-[3-[(2S)-2-Carboxy-2-[(3R)-pyrrolidine -3-yl]ethyl]phenoxy]phenyl]-2-[(3R)-pyrrolidin-3-yl ]Propionic acid, dihydrochloride [ka] tert-Butyl (3R)-3-[(1S)-2-tert-butoxy-1-[[3- [3-[(2S)-3-tert-butoxy-2-[(3R)-1-tert-butoxy Carbonylpyrrolidin-3-yl]-3-oxo-propyl]phenoxy]phenyl]methyl The title compound was prepared using ethyl-2-oxo-ethyl-pyrrolidine-1-carboxylate. The product is prepared essentially as described in Example 6. ES / MS (m / z): 453 (M +H).

[0096] Example 8 (2S)-3-[3-[2-[3-[(2S)-2-carboxy-2-[(3R)-pyrrolidone Lysine-3-yl]ethyl]phenoxy]ethoxy]phenyl]-2-[(3R)-pyrrolidinyl]ethyl]phenoxy]ethoxy]phenyl Lysine-3-yl]propanoic acid, dihydrochloride [ka] tert-Butyl (3R)-3-[(1S)-2-tert-butoxy-1-[[3- [2-[3-[(2S)-3-tert-butoxy-2-[(3R)-1-tert-butoxy [3-oxo-3-propyl]phenoxy]ethoxy [phenyl]methyl]-2-oxo-ethyl]pyrrolidine-1-carboxylate was used. The title compound is prepared essentially as described in Example 6 using MTBE. The product was isolated by elution with hexane. ES / MS (m / z): 497 (M+H).

[0097] Example 9 (2S)-3-[3-[[3-[(2S)-2-Carboxy-2-[(3R)-pyrrolidine 2-[(3R)-pyrrolidine -3-yl]propanoic acid, dihydrochloride [ka] tert-Butyl (3R)-3-[(1S)-2-tert-butoxy-1-[[3- [[3-[(2S)-3-tert-butoxy-2-[(3R)-1-tert-butoxy [Dicarbonylpyrrolidin-3-yl]-3-oxo-propyl]phenoxy]methyl] Using phenylmethyl-2-oxo-ethyl pyrrolidine-1-carboxylate The title compound is prepared essentially as described in Example 6. ES / MS (m / z): 467(M+H).

[0098] Example 10 (2S)-3-[3-[3-[(2S)-2-Carboxy-2-[(3R)-pyrrolidine -3-yl]ethyl]phenyl]sulfanylphenyl]-2-[(3R)-pyrrolidine -3-yl]propanoic acid, dihydrochloride [ka] tert-Butyl (3R)-3-[(1S)-2-tert-butoxy-1-[[3- [3-[(2S)-3-tert-butoxy-2-[(3R)-1-tert-butoxy Carbonylpyrrolidin-3-yl]-3-oxo-propyl]phenyl]sulfanyl Using phenylmethyl-2-oxo-ethyl pyrrolidine-1-carboxylate The title compound is prepared essentially as described in Example 6. The product is isolated by ES / MS (m / z): 469 (M+H).

[0099] Example 11 (2S)-3-[3-[3-[(2S)-2-Carboxy-2-[(3R)-pyrrolidine -3-yl]ethyl]phenyl]sulfinylphenyl]-2-[(3R)-pyrrolidine -3-yl]propanoic acid, dihydrochloride [ka] tert-Butyl (3R)-3-[(1S)-2-tert-butoxy-1-[[3- [3-[(2S)-3-tert-butoxy-2-[(3R)-1-tert-butoxy Carbonylpyrrolidin-3-yl]-3-oxo-propyl]phenyl]sulfinyl Using phenylmethyl-2-oxo-ethyl pyrrolidine-1-carboxylate The title compound is prepared essentially as described in Example 6. The product is triturated with EtO. and subsequently the product is isolated by RP-HPLC / MS using the following parameters: Column-Agilent ZORBAX Bonus RP, Mobile phase-Solvent A=0 in water. Solvent B = acetonitrile + 0.05% trifluoroacetic acid (pH 2.5), solvent B = acetonitrile + 0.05% trifluoroacetic acid Fluoroacetic acid, gradient - 5-30% solvent B in solvent A, flow rate: 25 mL / min. HCl (1 The product was taken up in a mixture of 100% w / v aqueous solution (100% w / v solution) and water, and the solvent was evaporated under a stream of nitrogen at 40°C. ES / MS(m / z): 485(M+H).

[0100] Example 12 (2S)-3-[3-[3-[(2S)-2-Carboxy-2-[(3R)-pyrrolidine -3-yl]ethyl]phenyl]sulfonylphenyl]-2-[(3R)-pyrrolidine- 3-yl]propanoic acid, dihydrochloride [ka] tert-Butyl (3R)-3-[(1S)-2-tert-butoxy-1-[[3- [3-[(2S)-3-tert-butoxy-2-[(3R)-1-tert-butoxy Carbonylpyrrolidin-3-yl]-3-oxo-propyl]phenyl]sulfonylphenyl nyl]methyl]-2-oxo-ethyl]pyrrolidine-1-carboxylate, The title compound is prepared essentially as described in Example 6. The product was isolated from the column. ES / MS (m / z): 501 (M+H).

[0101] Example 13 (2S)-3-[3-[[3-[(2S)-2-Carboxy-2-[(3R)-pyrrolidine 2-[(3R)-pyrrolidine- 3-yl]propanoic acid, trihydrochloride [ka] tert-Butyl (3R)-3-[(1S)-2-tert-butoxy-1-[[3- [[3-[(2S)-3-tert-butoxy-2-[(3R)-1-tert-butoxy Dicarbonylpyrrolidin-3-yl]-3-oxo-propyl]anilino]methyl]phenyl nyl]methyl]-2-oxo-ethyl]pyrrolidine-1-carboxylate, The title compound is prepared essentially as described in Example 6. After decanting the product, Purify by RP-HPLC / MS using the following parameters: Column - Waters XBridge™ C18 (19×100 mm, 5 μm), mobile phase-solvent A = 20 mM NH4HCO3 in water, solvent B = acetonitrile, flow rate 25 mL / min, gradient - The purified material was dissolved in aqueous HCl (1N) at room temperature. Stir for 1 h, evaporate the solvent under a stream of nitrogen, and dry the solid in vacuum at 40 °C. S / MS(m / z):466(M+H).

[0102] Example 14 (2S)-3-[3-[[3-[(2S)-2-Carboxy-2-[(3R)-pyrrolidine 2-[(3R)-pyrrolidin-3-yl]ethyl]anilino]phenyl]- ]Propionic acid, dihydrochloride [ka] (2S)-3-[3-[3-[(2S)-3-tert-butoxy-2-[(3R)- 1-tert-Butoxycarbonylpyrrolidin-3-yl]-3-oxo-propyl]a 2-[(3R)-1-tert-butoxycarbonylpyrrolidine- 3-yl]propanoic acid (9 g, 12.7 mmol), isopropanol (27 mL), and and HCl (5.5 M solution in isopropanol) and stirred at room temperature for 2.5 hours. The reaction is heated to 60° C. for 2.5 hours, then cooled to room temperature and stirred for 3 days. Heat again to 60 °C for 2 h, cool to room temperature, and evaporate the reaction mixture to dryness in vacuo. The solid residue was mixed with MTBE using a solvent-based ... Dry in vacuum. Mix the solid with concentrated aqueous hydrochloric acid and heat to 80° C. overnight, then at room temperature. Cool to rt and concentrate to dryness in vacuo. Dissolve the residue in a minimum amount of water and add aqueous NaOH. The pH is adjusted to 7.5 by adding 100 ml of 100 ml of 100% NaOH. The mixture is stirred at room temperature for 3 hours and then precipitated. The solid was filtered off. The solid was dissolved in aqueous HCl (1N) and stirred at room temperature for 15 min. The water is then removed in vacuo. The residue is dried in vacuum at 45° C. overnight to give the title compound (5 0.4 g, 81%). ES / MS (m / z): 452 (M+H).

[0103] Example 15 (2S)-3-[3-[[3-[(2S)-2-carboxy-2-[(3R)-1-methyl 2-[(3R)-1-methylpyrrolidin-3-yl]ethyl]anilino]phenyl] Pyrrolidin-3-yl]propanoic acid, dihydrochloride [ka] Paraformaldehyde (89 mg, 0.95 mmol) in MeOH (1.9 mL) (2S)-3-[3-[3-[(2S)-2-carboxy-2-[(3R)-pyrrolidine 2-[(3R)-pyrrolidin-3-yl]ethyl]anilino]phenyl]- ]propanoic acid, dihydrochloride (100 mg, 0.19 mmol) in MeOH (1.9 mL) Add sodium triacetoxyborohydride (2 Add 0.02 mg, 0.95 mmol) and stir at room temperature for 16 h. Concentrate the solution in vacuo. Reverse phase chromatography (column: Claricep C series silica bound C 18) using a gradient of 5–25% acetonitrile in aqueous NH4CO3 (pH 9). The residue was purified using 100% ethanol. The purified material was dissolved in aqueous hydrochloric acid (1N, 1 mL) and stirred at room temperature. Stir at rt for 6 h. Concentrate the solution in vacuo to give the title compound (30 mg, 27% yield). . ES / MS(m / z): 480(M+H).

[0104] Example 16 (2S,2'S)-3,3'-[sulfonylbis(azanediyl-3,1-phenylene) ]Bis{2-[(3R)-pyrrolidin-3-yl]propanoic acid}, dihydrochloride [ka] Di-tert-butyl 3,3'-((2S,2'S)-((sulfonylbis(azanediyl) 3-(tert-butoxy)-3-oxo-2,4-diphenyl-2,5-diphenyl-1,2-diphenyl-2,6-diphenyl-1,2 ... (3R,3'R)-bis(pyrrolidine-1-carboxylate) The title compound is prepared as described in Example 6 using m / z):531(M+H).

[0105] Example 17 (2S)-3-Phenyl-2-[(3R)-pyrrolidin-3-yl]propanoic acid, hydrochloride [ka] HCl (5.5 M in isopropanol, 511 mL, 2.81 mol, 13 equiv.) was added to (2S)-2-[(3R)-1-tert-butoxy]-2,4-dimethyl-1,5-diisopropyl-2,5-diisopropyl ...1,5-diisopropyl-2,5 Carbonylpyrrolidin-3-yl]-3-phenyl-propanoic acid (69 g, 216 mmol) The mixture was then added to a solution of 2:1 MTBE:Hexane (90 0 mL) and stir for 10 min. Filter the suspension and wash the solid with 1:1 MT Wash with BE:hexane (100 mL). Dry the solid under vacuum at 50° C. to give the title compound. The compound was obtained (51.3 g, 93%). ES / MS (m / z): 220 (M+H).

[0106] Example 18 (2R)-3-Phenyl-2-[(3S)-pyrrolidin-3-yl]propanoic acid, hydrochloride [ka] (2R)-2-[(3S)-1-tert-butoxycarbonylpyrrolidin-3-yl ]-3-phenyl-propanoic acid to prepare the title compound essentially as described in Example 6. The isolated product is stirred with HCl (2M in ether) and water for 3 h to prepare the pure product. Concentrate in air, then triturate with MTBE and dry in vacuum at 40°C. ES / MS (m / z):220(M+H).

[0107] Radiolabeling and low temperature (i.e. non-radioactive) for in vitro Apo(a) binding assays Synthesis of labeled) standards Preparation 40 1-(3-benzyloxyphenyl)imidazolidin-2-one [ka] Argon-sparged DMF (20 mL) was dissolved in ethylene urea (1.4 g, 16 mmol) l), 1-benzyloxy-3-iodobenzene (4.9 g, 16 mmol, 1.0 equiv. ), cuprous iodide (0.61 g, 3.1 mmol, 0.20 equiv.), potassium dihydrogen phosphate The resulting suspension was added to a mixture of N ,N'-Dimethylethylenediamine (0.33 mL, 0.28 g, 3.1 mmol, 0. Heat the blue suspension in a microwave reactor at 120° C. for 3 h. The reaction is cooled and filtered through a pad of silica gel and the pad is flushed with EtOAc. Concentrate the filtrate. Remove the precipitate with 5:20:75 MeOH:Acetone:EtOAc in hexanes. The residue was purified by silica gel chromatography using a 0-100% gradient of The title compound (1.10 g, 26%) is obtained as a yellow solid. ES / MS (m / z): 2 69(M+H)

[0108] Preparation 41 (2S)-3-[3-[3-(3-benzyloxyphenyl)-2-oxo-imidazoline 2-[(3R)-1-tert-butoxycarbonylpyridine-1-yl]phenyl]- Lysine-3-yl]propanoic acid [ka] 1-(3-benzyloxyphenyl)imidazolidin-2-one (3.33 g, 12. 4mmol, 1.50 equivalents) and ammonium; (2S)-3-(3-bromophenyl)- 2-[(3R)-1-tert-butoxycarbonylpyrrolidin-3-yl]propanoic acid A mixture of acetonitrile (41 mL) and ethyl acetate (3.30 g) was added under a nitrogen atmosphere. DMF (to aid solubility) is added. The suspension is subjected to a steady stream of argon for 15 minutes. Add potassium carbonate (3.75 g, 26.8 mmol, 3.24 equiv.), diisopropyl iodide (1.0 g, 1.0 eq.), and 100% sodium iodide (0.01 g, 0.01 eq.). Copper monooxide (0.316 g, 1.66 mmol, 0.2 equiv.) and N,N'-dimethylethyl Add diamine (0.360 mL, 3.31 mmol, 0.4 equiv.). Heat at 100° C. over the weekend. Add water and extract the aqueous layer twice with EtOAc. The phase was acidified with 0.5N HCl in water and the aqueous layer was washed with EtOAc, DCM and then EtOAc. The combined organic phases are dried over MgSO4, filtered and evaporated to dryness. Silica gel chromatography using 10-60% (1% acetic acid / acetone) in acetone. The residue is purified by HPLC to give the title compound (4 g, 82%). ES / MS (m / z): 4 86(M+H-BOC).

[0109] Preparation 42 tert-Butyl(3R)-3-[(1S)-1-[[3-[3-(3-benzyloxy) Phenyl)-2-oxo-imidazolidin-1-yl]phenyl]methyl]-2-tert-butyl t-Butoxy-2-oxo-ethyl]pyrrolidine-1-carboxylate [ka] (2S)-3-[3-[3-(3-benzyloxyphenyl) )-2-oxo-imidazolidin-1-yl]phenyl]-2-[(3R)-1-tert-butyl t-Butoxycarbonylpyrrolidin-3-yl]propanoic acid (4.0 g, 6.8 mmol ) solution at 70°C, N,N-dimethylformamide di-tert-butyl acetal ( 15 mL, 55 mmol, 8.0 equiv) and heat the reaction at 70 °C overnight. The residue was purified by silica gel chromatography using a gradient of 0-50% EtOAc in water. The residue is purified to give the title compound (1.56 g, 33%). ES / MS (m / z): 54 2(M+H-BOC).

[0110] Preparation 43 tert-Butyl (3R)-3-[(1S)-2-tert-butoxy-1-[[3-[ 3-(3-hydroxyphenyl)-2-oxo-imidazolidin-1-yl]phenyl] Methyl-2-oxo-ethyl pyrrolidine-1-carboxylate [ka] tert-Butyl (3R)-3-[(1S)-1-[[3-[3-(3-benzyloxy) 2-phenyl-2-oxo-imidazolidin-1-ylphenylmethyl-2-tert-butyl rt-Butoxy-2-oxo-ethyl]pyrrolidine-1-carboxylate (1.56 g , 2.43 mmol) in THF (80 mL) and palladium hydroxide (0.7 g, The reaction was degassed under vacuum and then diluted with hydrogen gas. The reaction is stirred under hydrogen (1 atm) at room temperature for 4 h. The reaction is concentrated and Silica gel chromatography using a gradient of 0-50% EtOAc in hexanes The residue is purified by HPLC to give the title compound (1.19 g, 87%). ES / MS (m / z) :452(M+H-BOC).

[0111] Preparation 44 tert-Butyl (3R)-3-[(1S)-2-tert-butoxy-2-oxo-1 -[[3-[2-oxo-3-[3-(trithiomethoxy)phenyl]imidazolide 1-phenyl-1-ylphenylmethylethylpyrrolidine-1-carboxylate [ka] tert-Butyl (3R)-3-[(1S)-2-tert -Butoxy-1-[[3-[3-(3-hydroxyphenyl)-2-oxo-imidazoline 1-phenyl-2-oxo-ethyl-pyrrolidine-1-carboxylate A solution of silane (4 mg, 0.007 mmol) was added to a solution of cesium carbonate (10 mg, 0.03 Add 1 mmol of 1H 2 O and stir the solution at room temperature for 10 min. 3 H]Methyl nosylate (5 Add 0.0 mCi) and stir at room temperature for 1.5 hours. The reaction mixture is purified by HPLC / MS: Column - Phenomenex® ) Gemini® C18 (250×10 mm) was used with mobile phase-solvent A = water + trimethylsilane. Fluoroacetic acid (0.1%), Solvent B = acetonitrile + trifluoroacetic acid (0.1%), Gradient – ​​50–100% B in A over 50 min, flow rate – 3 mL / min. The purified product was eluted in Et Dissolve in OH and proceed to preparation 45 without further characterization.

[0112] Preparation 45 (2S)-3-[3-[2-oxo-3-[3-(trithiomethoxy)phenyl]i [(3R)-pyrrolidin-3-yl]propane Acid, hydrochloride [ka] In vacuum, tert-butyl(3R)-3-[(1S)-2-tert-butoxy-2 -Oxo-1-[[3-[2-oxo-3-[3-(trithiomethoxy)phenyl] Imidazolidin-1-yl]phenyl]methyl]ethyl]pyrrolidine-1-carboxylate The solvent is removed from the solution and the residue is dissolved in hydrogen chloride (4 M solution in 1,4-dioxane). The mixture is stirred at room temperature overnight. RP-HPLC / MS analysis is performed using the following parameters: The reaction mixture is purified by S: column - Phenomenex® Gemin i® C18 (250 x 10 mm) with mobile phase - solvent A = water + trifluoroacetic acid (0.1%), solvent B = acetonitrile + trifluoroacetic acid (0.1%), gradient - 1 in A 0-70% B over 60 min, flow rate - 3 mL / min. Dissolve the purified product in ethanol. Mass spectrometry revealed inactive material [ES / MS (m / z): 410 (M+H)] and 63 Provides spectra consistent with specific activity in Ci / mmol.

[0113] Preparation 46 1-(3-Methoxyphenyl)imidazolidin-2-one [ka] Using 1-iodo-3-methoxybenzene and potassium phosphate tribasic as a base, Prepare the title compound essentially as described in Preparation 40. No conventional heating is used. The reaction is heated to 120° C. for 16 h. Dilute with EtOAc and add Celite®. The organics were filtered through a pad of ethyl acetate (labeled ethyl acetate) and the solution was washed with water, aqueous NH4OH, and saturated aqueous NaCl. Work up the reaction by washing with solutions: 20-100% EtOAc in hexane. The crude material is purified by silica gel chromatography using a gradient of c. ES / MS (m / z): 193 (M+H).

[0114] Preparation 47 (2S)-2-[(3R)-1-tert-butoxycarbonylpyrrolidin-3-yl] -3-[3-[3-(3-methoxyphenyl)-2-oxo-imidazolidin-1-yl ]phenyl]propanoic acid [ka] (2S)-3-(3-bromophenyl)-2-[(3R)-1-tert-butoxycarbonyl] 1,2-Dimethylpyrrolidin-3-yl]propanoic acid (190 mg, 0.4770 mmol), -(3-Methoxyphenyl)imidazolidin-2-one (0.2751 g, 1.431 m mol, 3 equivalents), [(2-di-tert-butylphosphino-2',4',6'-tri Isopropyl-1,1'-biphenyl)-2-(2'-amino-1,1'-biphenyl) ]Palladium(II) methanesulfonate (tBuXPhos-Pd-G3, 0.0379 g, 0.0477 mmol, 0.1 equiv.) and sodium tert-butoxide (0. 1418 g, 1.431 mmol, 3 equiv.) in 1,4-dioxane (4.8 mL) Mix and stir the mixture overnight at 100° C. under nitrogen. Dilute the mixture with EtOAc. Acidify with aqueous HCl (1N). Mix through a pad of Celite®. Filter the organics, separate the layers, and dry the organics over MgSO4. Filter and concentrate the organics. Then, a gradient of 40–70% acetonitrile in aqueous NH4CO3 (pH 9) was used. The residue was purified by reversed-phase flash chromatography (silica-bonded C18 column) using To give the title compound (70 mg, 28%) as a white solid, use the following parameters: To obtain an additional amount of the title compound (90 mg, 36%) by RP-HPLC / MS: Ram-Waters(TM) XBridge(TM) C18(19×100mm, 5μm ), mobile phase-solvent A = 20 mM NH4HCO3 in water, solvent B = acetonitrile, flow rate 2 5 mL / min, gradient - 30:70 to 50:50 B:A. ES / MS (m / z): 510 ( M+H).

[0115] Preparation 48 (2S)-3-[3-[2-oxo-3-[3-(methoxy)phenyl]imidazolidine -1-yl]phenyl]-2-[(3R)-pyrrolidin-3-yl]propanoic acid, hydrochloride [ka] (2S)-2-[(3R)-1-tert-butoxycarbonylpyrrolidin-3-yl ]-3-[3-[3-(3-methoxyphenyl)-2-oxo-imidazolidine-1-yl Preparation 45 was prepared using 1,2-diphenylpropanoic acid, essentially as described in Preparation 45. Compound prepared: ES / MS(m / z): 410(M+H).

[0116] Biological assays In vitro Apo(a) binding assay The in vitro binding affinity of compounds to the intended target human Apo(a) protein was determined by competitive assay. The human Apo(a) tag, which contains 17 kringle repeats, is tested in a binding assay. Proteins were avidly isolated from conditioned medium of transiently transfected HEK-293F cells. All reagents were prepared in 50 mM Tris-HCl pH 7.4, 0.1% B Binding assays are performed in each well of a clear bottom plate. In the tube, (1) a dilution series of test compounds (final concentrations 0.32 to 10,000 nM), (2) Ap (a) Protein (6 ng / well), (3) Resuspended wheat germ agglutinin polyvinyl chloride Ruen SPA beads (20 mg / mL), and (4) radioactive ligand, tritium-labeled. (2S)-3-[3-[2-oxo-3-[3-(trithiomethoxy)phenyl]i [(3R)-pyrrolidin-3-yl]propane This was performed by adding 50 μl of each of phosphate; Plates are incubated at room temperature for 60 minutes and counted on a TRILUX LSC. Non-specific binding, binding in the presence of 10 μM cold (i.e., non-radiolabeled) ligand (2S)-3-[3-[2-oxo-3-[3-(methoxy)phenyl ]imidazolidin-1-yl]phenyl]-2-[(3R)-pyrrolidin-3-yl]p Subtract propanoic acid, hydrochloride to determine specific binding. Standard single-site binding model Analyze the data by fitting ICs of exemplary test compounds 50 This is to determine The results are summarized in Table 1, which shows that examples of test compounds bind to the human Apo(a) protein. This indicates that apo(a)-induced L-gamma agonism is mediated by binding to the Apo(a) protein. Inhibition of DL particle assembly supports the reduction of Lp(a) levels. [Table 1]

[0117] In vitro Lp(a) assembly assay The ability of compounds to inhibit Lp(a) particle formation in vitro was assessed using a cell-free assembly assay. Conditioned medium (10% FBS, 20 mM HEPES, and 1x penicillin) was used to evaluate the DMEM supplemented with cyclosporine / streptomycin) was incubated at 37°C and 5% CO2 for 24 h. After 2 h of incubation, confluent wild-type HepG2 cells (which express ApoB endogenously) were cultured. source) and expressing human Apo(a) containing 17 kringle repeats. The cells were harvested from a HEK293 stable cell line (selected with 1 mg / ml Geneticin). Equal amounts of HepG2 and HEK293 conditioned media were diluted in series (final concentrations: 0.01–10 In vitro assembly assays were performed by combining the test compounds added at 0 nM (0.01 nM). The reaction was incubated at 37° C. for 2 hours and then 6-aminocaproic acid (EAC) was added. A) to a final concentration of 150 mM was added to stop the reaction. Lp(a) was detected using anti-Lp(a) capture antibody. ApoB was detected using a sandwich ELISA with the antibody and an anti-ApoB detection antibody conjugated to HRP. The ELISA was developed using TMB and stopped using 1N sulfuric acid, resulting in a signal is read at 450 nm on a Molecular Devices plate reader. The % inhibition of Lp(a) formed under each test condition was calculated based on the percentage of inhibition in the absence of inhibitor (DMSO concentration of 1 %) assembly reaction was set to 0% inhibition and a minimal amount of HepG2 conditioned medium ( 50-fold dilution) is set to 100% inhibition. The IC summarized in Table 2 50 To determine the value To do this, the data were fitted to a four-parameter curve. As summarized in Table 2, ApoB and the addition of an exemplary test compound to conditioned medium containing Apo(a) in vitro The results show that these compounds inhibit Apo(a) formation in a concentration-dependent manner. and inhibiting the assembly of Lp(a) from LDL particles. [Table 2]

[0118] In vivo Lp(a) inhibition in mice The ability of the compounds to reduce steady-state Lp(a) levels in vivo was evaluated using humanized Lp(a The effect of Lp(a) on the immune response of the host is evaluated in a transgenic mouse model capable of producing Lp(a) particles. In vivo effects of disruptor compounds were assessed in human apoB-100 and 17 clonidine kinase inhibitors. 7- to 17-month-old female double transgenic mice expressing human apo(a) containing the 1 repeat Test in transgenic mice: B6.SJL-Tg(APOB)1102Sgy Tg(A lb-LPA) 32 Arte. Mice were kept under a standard light cycle (12 h light / 12 h light). The mice were housed under a dark period (dark period) at room temperature of 72 ± 8°F and a relative humidity of 30-70%, and were provided with water and regular chow diet by themselves. (Harlan Tecklad diet 2014). The mice were randomly assigned to the study using the Block Random Allocation Tool (BRAT) to determine weight and Treatment groups (n=5 / group) were randomly assigned 3-5 days before the study according to baseline plasma Lp(a) concentrations. Mice were randomized to receive vehicle (oral administration vehicle: 10 mL / kg, 1% H EC, 0.25% Tween 80, 0.01% antifoam, subcutaneous administration vehicle: 5 mL / kg saline) or test compound at various doses twice daily (6:30 am and 3:30 pm). Administer orally (or subcutaneously where indicated) at 1:30 pm for 5 days. Blood was collected from the tail suction tube. Blood samples were collected via bleeding into heparin-coated capillary tubes. Transfer 20 μL of the sample to a DBS card (Whatman catalog number The remaining blood sample is centrifuged to separate the plasma. Plasma Lp(a) concentrations were determined using an in vitro Lp(a) aggregation assay as described above. Measured using a sandwich ELISA. Percent inhibition for each dose group is calculated as the mean of the vehicle control group. The mean Lp(a) level is determined by setting it at 0% inhibition. Table 3 shows the mean Lp(a) level after oral administration on the morning of the third day. The results of a dose-response study in which tail bleed samples were taken after 8 hours are shown. 50(50 The calculated effective dose for inhibiting 100% of Lp(a) was determined by a threshold minimal effective dose analysis. Table 4 shows the four-day period for oral (or subcutaneous, if indicated) dosing on the morning of the third day. Single-dose studies with tail bleed samples taken after 2 h (or 8 h where noted) These results, shown in Tables 3 and 4, demonstrate that the compounds are effective in reducing plasma The compound was shown to be effective in lowering Lp(a) levels and showed a significant reduction in Lp(a) plasma concentrations. This paper supports the proposition that it can be used to reduce [Table 3] [Table 4]

Claims

1. A compound of the following formula: 【Chemistry 1】 Compound.

2. The compound of the following formula: 【Chemistry 2】 Compound.

3. The compound of the following formula: 【Chemistry 3】 Compound.

4. A compound of the following formula: 【Chemistry 4】 Compound.

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