1, 2-substituted 3-oxopyrazolidine derivatives as prostaglandin E2 receptor 4 (EP4) agonists for treatment of gastrointestinal and lung diseases

Abstract

The present invention relates to compounds of formula I as prostaglandin E2 receptor 4 (EP4) agonists for use in methods of treating gastrointestinal and pulmonary diseases or conditions. Exemplary compounds such as 4-(2-(2-(3-hydroxy-3-(4 '-hydroxy-2'-methyl-[1, 1 '-biphenyl]-3-yl) propyl)-5-oxopyrazolidin-1-yl) ethyl) benzoic acid (embodiment 1), for example, pharmacological data are provided. # imgabs 0 #

Description

[0001] The present application relates to novel compounds and their use as prostaglandin E2 receptor 4 (EP4) agonists. The compounds described herein can be used to treat or prevent diseases in which the EP4 receptor is involved. The present application also relates to pharmaceutical compositions comprising these compounds, and the preparation and use of these compounds and compositions in preventing or treating such diseases in which the EP4 receptor is involved. Background Art

[0002] Prostaglandins (comprising prostaglandins and thromboxanes) are metabolic derivatives of arachidonic acid that play a key role in cellular physiological functions.Arachidonic acid is the indispensable component of the membrane phospholipids released by the activity of phospholipase A2 (phospholipase A2, PLA2).The biosynthesis of prostaglandins is mediated by cyclooxygenase (cyclooxygenase, COX), and the cyclooxygenase catalysis arachidonic acid is converted into unstable intermediate (PGH2) and causes the generation of prostaglandins including PGE2.PGE2 represents the most widely produced prostaglandins, and its activity is mediated by acting on 4 kinds of different receptor subtypes EP1-4 of functions.

[0003] EP receptors belong to the G protein coupled receptor (GPCR) family of integral membrane proteins with seven transmembrane domains. Such receptors can be broadly classified according to their signal transduction pathways: i) Gs-coupled receptors (adenylate cyclase activation and cyclic adenosine monophosphate (cAMP) production), EP2 and EP4; ii) Gq-coupled receptors (PLC activation), EP1; and iii) Gi-coupled receptors (adenylate cyclase inhibition), EP3.

[0004] The EP4 receptor signals through Gs and positively couples to adenylate cyclase to increase cAMP levels. The receptor was originally described in 1993, where an EP2-like receptor was identified that positively couples to adenylate cyclase but does not bind butaprost (A Honda et al. J. Biol. Chem. 1993, 268, 7759-7762).

[0005] EP4 receptors play a key role in a variety of physiological functions, including gastrointestinal homeostasis, regulation of vascular tone, renal function, inflammation, fever, and carcinogenesis. The potent biological effects of PGE2 have stimulated interest in developing subtype-selective EP4 agonists and antagonists for the treatment of a wide range of indications.

[0006] Functional gastrointestinal diseases (FGIDs), including chronic constipation, are common gastrointestinal disorders encountered by primary care physicians and gastroenterologists. The prevalence of chronic constipation is 1% to 8% and can negatively impact quality of life (QoL), leading to a serious social and economic burden. Chronic constipation can cause discomfort to patients and affect their daily lives. Symptoms include hard and lumpy stools, straining during defecation, and a feeling of incomplete defecation. The use of laxatives and stool softeners remains high, although many patients do not gain substantial benefit. There is still a need for treatments that can provide complete and sustainable symptom relief.

[0007] Intestinal mucosa plays an important role in the homeostasis of anion and body fluid maintenance. These functions are mediated by coordinated ion transport via membrane-bound transporters and channels on the top and basolateral membranes of intestinal epithelial cells. PGE2 is a recognized secretagogue that can directly promote intestinal epithelial cells to secrete chloride. The secretory effect of PGE2 is partly mediated by EP4 receptors, which can stimulate anion chloride secretion through intestinal mucosa. Lubiprostone (lubiprostone) (bicyclic fatty acid derivatives of PGE1) approved clinically for the treatment of chronic constipation and IBS-C has been shown to promote fluid secretion and gastrointestinal motility by activating prostaglandin (EP4) receptors. EP4 selective agonists can have therapeutic value in promoting the intestinal fluid homeostasis in chronic constipation disorders.

[0008] Inflammatory bowel disease (IBD) is a chronic debilitating gastrointestinal disorder, including ulcerative colitis and Crohn's disease. Patients with IBD usually show symptoms including diarrhea, abdominal pain, weight loss, rectal bleeding and fever. Clinical management includes strategies for controlling the abnormal dysregulated immune response of the intestinal mucosa. A wide range of agents are used to induce and maintain remission, depending on the severity of the disease. These include aminosalicylates, corticosteroids, immunosuppressants, antibiotics and biologics. This can be given in a stepwise approach, wherein treatment is strengthened according to the severity and progression of the disease. However, it is clear that some patients are refractory to drug therapy, or cannot tolerate drug therapy due to systemic side effects. Only a portion of patients achieve long-term remission due to current treatment strategies, which indicates the need for improved treatment.

[0009] Intestinal barrier dysfunction plays a key pathogenic role in IBD, and there is interest in the development of agents that restore barrier function (mucosal healing). EP4 is expressed in many cell types, including gastrointestinal (GI) epithelial cells, lamina propria mononuclear cells, and colon innervating sensory neurons, and may provide benefits in suppressing inappropriate mucosal immune responses and protecting the GI mucosal barrier. PGE2 signaling through EP4 promotes cell differentiation into a wound-associated epithelial cell phenotype, which is important for wound repair (Miyoshi, H. et al. EMBO J. 2017, 36, 5–24). Administration of EP4 agonists provides benefits in a chemically induced colitis model (Kabashima, K. et al. J. Clin. Invest. 2002, 109, 883–893; Watanabe, Y. et al. Eur. J. Pharmacol. 2015, 754, 179–189; Nitta, M. et al. Scand. J. Immunol. 2002, 56, 66–75), while mutant mice lacking EP4 develop severe dextran sulfate sodium-induced colitis, characterized by impaired mucosal barrier function, increased epithelial cell loss, crypt damage, and increased immune cell infiltration (Kabashima, K. et al. J. Clin. Invest. 2002, 109, 883–893). In a small PhII study, the EP4 agonist ONO-4819CD was evaluated in patients with mild to moderate ulcerative colitis refractory to 5-ASA (Nakase, H. et al. Inflamm. Bowel Dis. 2010, 16, 731-733). Although the study did not demonstrate efficacy, patients treated with ONO-4819CD showed signs of improved disease activity index (DAI) scores and histological scores. These data support the potential clinical benefits of EP4 agonist treatment in IBD.

[0010] Asthma and chronic obstructive pulmonary disease are inflammatory diseases of the airways characterized by airflow limitation. It is estimated that about 300 million people suffer from asthma and represent the most common chronic diseases in children. Although progress has been made in asthma management, the disease of a considerable number of patients is not controlled, which can lead to death and morbidity. Still need to achieve effective control of asthma symptoms and reduce the treatment of future exacerbation risks for long-term management. Known PGE2 has bronchodilator and anti-inflammatory effects in the airway smooth muscle isolated from rodents and people. It has been shown that the PGE2 inhaled is beneficial to the inflammation and airway diameter (calibre) of the patient suffering from chronic bronchitis and asthma. However, PGE2 also induces reflex cough, potentially through EP3 receptor-mediated upper airway stimulation. This makes efforts to find EP receptor selective agents for the treatment of airway diseases. Interestingly, it is reported that there are key species differences in the receptor subtypes responsible for regulating airway smooth muscle relaxation. In guinea pigs, monkeys and mice, EP2 agonists can induce relaxation of airway smooth muscle, while in humans this is mediated through the EP4 receptor (Buckley, J. et al. Thorax 2011, 66, 1029-1035). Selective EP4 receptor agonists may have potential therapeutic value in airway diseases.

[0011] It has been shown that EP4 receptors play a role in blood pressure regulation. EP4 is expressed on smooth muscle and endothelial cells, and may induce vasodilation through nitric oxide (nitric oxide, NO) mediated by endothelial nitric oxide synthase (endothelial nitric oxide synthase, eNOS). It has been shown that PGE2 relaxes smooth muscle in aortic rings in a dose-dependent manner, and this effect is eliminated in EP4 knockout mice. In dogs anesthetized by halothane, EP4 selective agonist ONO-AE1-329 induces vasodepressor response (Honda, A. et al. Eur. J. Pharmacol. 2016, 775, 130–137) and similarly, hypotension is reported as one of the adverse drug reactions of IBD patients receiving ONO-4819CD (Nakase, H. et al. Inflamm. Bowel Dis. 2010, 16, 731–733). EP4 agonists that can be used to treat a range of gastrointestinal and airway disorders without cardiovascular side effects may have potential therapeutic value. In particular, EP4 agonists that can be used to treat a range of gastrointestinal disorders without cardiovascular side effects may have potential therapeutic value. There is a need to discover safe and effective EP4 selective agents. Summary of the invention

[0012] The present invention provides compounds having activity as prostaglandin E2 receptor 4 (EP4) agonists.

[0013] In one aspect, provided herein is a compound of Formula I or a pharmaceutically acceptable salt, solvate, hydrate, tautomer or optical isomer thereof:

[0014]

[0015] in:

[0016] A is OR', C(O)R', CO2R', C(O)N(R')2, C(O)N(R')S(O)2R 3 ,S(O)2R',S(O)2OR',SO2N(R')2,C 1-8 alkyl, cycloalkyl, heterocycloalkyl, aryl or heteroaryl;

[0017] X is halogen, OR', COOR' or C 1-6 alkyl;

[0018] L and L' are each independently C 2-4 Alkylene;

[0019] R 1 H, halogen, CN, NO2, OR', SR', COOR', C 1-6 Alkoxy or C 1-6 alkyl;

[0020] R 2 is OR', OC(O)R 3 ,OC(O)OR 3 , CO2R', CON(R')2, SO2N(R')2, SO2R 3 ,OSO2R 3 or OSO2N(R')2;

[0021] R 3 It is C 1-6 Alkyl, C 3-6 Cycloalkyl or phenyl;

[0022] R' is H, C 1-6 Alkyl or C 3-6 cycloalkyl; and

[0023] n is 0, 1, 2, or 3;

[0024] wherein at each occurrence, alkyl, alkylene, alkoxy and cycloalkyl are each optionally and independently substituted by OH, SH, CN, NO2, COOH, halogen or COOC 1-4 Up to 3 instances of substitution in the alkyl group;

[0025] wherein heterocycloalkyl, aryl and heteroaryl are each optionally and independently substituted at each occurrence with up to 3 instances of OR', SR', CN, NO2, CO2R', halogen, C1-4 alkyl or oxo.

[0026] In another aspect, the present invention includes a pharmaceutical composition comprising a compound described herein and a pharmaceutically acceptable excipient.

[0027] In another aspect, the invention includes a kit comprising a compound described herein and at least one additional therapeutic agent selected from the group consisting of an aminosalicylate, a corticosteroid, an immunomodulator, and combinations thereof.

[0028] In another aspect, the present invention includes a compound described herein, a composition described herein, or a kit described herein for use as a medicament.

[0029] In another aspect, the invention includes a compound described herein, a composition described herein, or a kit described herein for use in treating an EP4 receptor mediated disease.

[0030] In another aspect, the invention includes a method of modulating EP4 receptor agonist activity in a biological sample, the method comprising contacting the EP4 receptor with a compound or composition described herein.

[0031] In another aspect, the invention includes a method of treating a disease mediated by the EP4 receptor, comprising administering to a patient in need thereof a compound or composition described herein.

[0032] The compounds herein can be used as EP4 receptor agonists. The compounds herein can selectively act on the EP4 receptor. The compounds can be used to prepare compositions or drugs. The compounds, compositions or drugs can be used to treat, prevent, improve, control or reduce the risk of diseases or conditions involving the EP4 receptor. The compounds, compositions or medicaments can be used to treat, prevent, ameliorate, control or reduce the risk of gastrointestinal disorders and conditions, including but not limited to constipation disorders, constipation-dominant irritable bowel syndrome, mixed irritable bowel syndrome, chronic idiopathic constipation, gastrointestinal symptoms associated with Parkinson's disease, gastrointestinal symptoms associated with cystic fibrosis, intestinal motility disorders, postoperative ileus, food allergy or food intolerance, celiac disease, gastrointestinal motility disorders, functional gastrointestinal disorders, drug-induced enteropathy, NSAID-induced gastric and intestinal damage, chemotherapy-induced mucositis, gastroesophageal reflux disease (GERD), duodenogastric reflux, diarrheal diseases, immune-mediated gastrointestinal diseases, Crohn's disease, ulcerative colitis, inflammatory bowel disease and ischemic colitis.

[0033] The compounds, compositions or drugs can also be used to treat, prevent, improve, control or reduce the risk of lung diseases and disorders, such as chronic obstructive pulmonary disease, asthma, chronic bronchitis, cystic fibrosis, emphysema, chronic idiopathic cough, overactive airway disorders and idiopathic pulmonary fibrosis. DETAILED DESCRIPTION

[0034] The present invention relates to novel compounds. The present invention also relates to the use of novel compounds as EP4 receptor agonists. The present invention also relates to the use of novel compounds in the preparation of medicaments used as EP4 receptor agonists, and treatment methods comprising administering the compounds of the present invention as EP4 receptor agonists.

[0035] The compounds of formula (I) can be used to treat, prevent, improve, control or reduce the risk of diseases or conditions involving EP4 receptors. The compounds of formula (I) can be used to treat, prevent, improve, control or reduce the risk of gastrointestinal disorders and conditions, including but not limited to constipation disorders, constipation-dominant irritable bowel syndrome, mixed irritable bowel syndrome, chronic idiopathic constipation, gastrointestinal symptoms associated with Parkinson's disease, gastrointestinal symptoms associated with cystic fibrosis, intestinal motility disorders, postoperative ileus, food allergy or food intolerance, celiac disease, gastrointestinal motility disorders, functional gastrointestinal disorders, drug-induced enteropathy, NSAID-induced gastric and intestinal damage, chemotherapy-induced mucositis, gastroesophageal reflux disease (GERD), duodenogastric reflux, diarrheal diseases, immune-mediated gastrointestinal diseases, Crohn's disease, ulcerative colitis, inflammatory bowel disease and ischemic colitis.

[0036] The compounds of formula (I) may also be used to treat, prevent, ameliorate, control or reduce the risk of lung diseases and conditions, such as chronic obstructive pulmonary disease, asthma, chronic bronchitis, cystic fibrosis, emphysema, chronic idiopathic cough, overactive airways disorders and idiopathic pulmonary fibrosis.

[0037] Certain novel compounds of the present invention show particularly high activity as EP4 receptor agonists.

[0038] The compounds of the present invention have been shown to have activity as EP4 receptor agonists. As shown by Caco-2 studies, the compounds of the present invention also have low gastrointestinal permeability. Therefore, it is believed that when administered orally, the compounds of the present invention exhibit low systemic bioavailability. Functional agonists of EP4 receptors expressed in the gastrointestinal tract have the potential to treat a range of gastrointestinal disorders. The combination of EP4 receptor agonist activity and low gastrointestinal permeability indicates that the compounds of the present invention can be used to treat a range of gastrointestinal disorders without cardiovascular side effects caused by systemic distribution.

[0039] In one aspect, the present invention comprises a compound of formula I or a pharmaceutically acceptable salt, solvate, hydrate, tautomer or optical isomer thereof:

[0040]

[0041] in:

[0042] A is OR', C(O)R', CO2R', C(O)N(R')2, C(O)N(R')S(O)2R 3 ,S(O)2R',S(O)2OR',SO2N(R')2,C 1-8alkyl, cycloalkyl, heterocycloalkyl, aryl or heteroaryl;

[0043] X is halogen, OR', COOR' or C 1-6 alkyl;

[0044] L and L' are each independently C 2-4 Alkylene;

[0045] R 1 H, halogen, CN, NO2, OR', SR', COOR', C 1-6 Alkoxy or C 1-6 alkyl;

[0046] R 2 is OR', OC(O)R 3 ,OC(O)OR 3 , CO2R', CON(R')2, SO2N(R')2, SO2R 3 ,OSO2R 3 or OSO2N(R')2;

[0047] R 3 It is C 1-6 Alkyl, C 3-6 Cycloalkyl or phenyl;

[0048] R' is H, C 1-6 Alkyl or C 3-6 cycloalkyl; and

[0049] n is 0, 1, 2, or 3;

[0050] wherein at each occurrence, alkyl, alkylene, alkoxy and cycloalkyl are each optionally and independently substituted by OH, SH, CN, NO2, COOH, halogen or COOC 1-4 Up to 3 instances of substitution in the alkyl group;

[0051] wherein at each occurrence, heterocycloalkyl, aryl and heteroaryl are each optionally and independently substituted by OR', SR', CN, NO2, CO2R', halogen, C 1-4 Up to 3 instances of alkyl or oxo are substituted.

[0052] In some embodiments, the compound is a compound of Formula (I), (Ia), (Ib), (Ic), (1), (1a), (1b), (1c), (2a), (2b), (2c), (2d), (3a), (3b), (3c), (3d), (3e), (3f), (5), (5a), (5b), (6), (6a), (6b) or a pharmaceutically acceptable salt or tautomer thereof.

[0053] In some embodiments, the compound is a compound of Formula (I), (Ia), (Ib), (Ic), (1), (1a), (1b), (1c), (2a), (2b), (2c), (2d), (3a), (3b), (3c), (3d), (3e), (3f), (5), (5a), (5b), (6), (6a), (6b) or a pharmaceutically acceptable salt thereof.

[0054] In some embodiments, L' is a group

[0055]

[0056] In some embodiments, L is a group

[0057]

[0058] In one embodiment of this aspect, the compound is a compound of formula (1):

[0059]

[0060] or a pharmaceutically acceptable salt, solvate, hydrate, tautomer or optical isomer thereof.

[0061] In some embodiments, A is selected from C(O)OR', C(O)N(R')S(O)2R 3 ,S(O)2OR',C 1- alkyl, heterocycloalkyl or heteroaryl, wherein heterocycloalkyl and heteroaryl are each optionally and independently substituted by OR', SR', halogen, C 1-4 Up to 3 instances of alkyl or oxo are substituted.

[0062] In some embodiments, A is selected from C(O)OR' and heteroaryl.

[0063] In some embodiments, A is selected from:

[0064]

[0065] In some embodiments, A is:

[0066]

[0067] In some embodiments, A is

[0068]

[0069] In some embodiments, the compound of Formula (I) is a compound of Formula (Ia), Formula (Ib), or Formula (Ic):

[0070]

[0071] or a pharmaceutically acceptable salt, solvate, hydrate, tautomer or optical isomer thereof.

[0072] In some embodiments, the compound of formula (I) is a compound of formula (1a), formula (1b), or formula (1c):

[0073]

[0074] or a pharmaceutically acceptable salt, solvate, hydrate, tautomer or optical isomer thereof.

[0075] In some embodiments, the compound of formula (I) is a compound of formula (3a), formula (3b), formula (3c), formula (3d), formula (3e), or formula (3f):

[0076]

[0077] or a pharmaceutically acceptable salt, solvate, hydrate, tautomer or optical isomer thereof.

[0078] In some embodiments, X is halogen or OR'.

[0079] In some embodiments, X is F, Cl, or OH.

[0080] In some embodiments, X is F or OH, and n is 0, 1, or 2.

[0081] In some embodiments, X is F, Cl, or OH, and n is 1 or 2.

[0082] In some embodiments, X is F or OH, and n is 1 or 2.

[0083] In some embodiments, n is 0.

[0084] In some embodiments, R 1 is H, OH, halogen, CN, C optionally substituted by 1 to 3 fluorine atoms 1-6 alkoxy, or C optionally substituted by 1 to 3 fluorine atoms 1-6 alkyl.

[0085] In some embodiments, R 1 is H, OH, halogen, CN, C optionally substituted by 1 to 3 fluorine atoms 1-6 alkyl, or C 1-6 alkyl.

[0086] In some embodiments, R1 It's methyl.

[0087] In some embodiments, R 2 It is OR', CON(R')2, SO2N(R')2 or OSO2N(R')2.

[0088] In some embodiments, R 2 It is OH, CONH2, SO2NH2 or OSO2NH2.

[0089] In some embodiments, R 2 It is CON(R')2, SO2N(R')2, OSO2R3 or OSO2N(R')2.

[0090] In some embodiments, R 2 It is CONH2, SO2NH2 or OSO2NH2.

[0091] In some embodiments, R 2 It is CONH2.

[0092] In some embodiments, provided herein are compounds of formula (2a), formula (2b), formula (2c), or formula (2d):

[0093]

[0094] or a pharmaceutically acceptable salt, solvate, hydrate or tautomer thereof.

[0095] In some embodiments, R 1 Is H or C 1-6 In another embodiment, R 1 In another embodiment, n is 0.

[0096] In some embodiments, provided herein are compounds of formula (5), formula (5a), formula (5b), formula (6), formula (6a), and formula (6b):

[0097]

[0098] or a pharmaceutically acceptable salt, solvate, hydrate or tautomer thereof.

[0099] In some embodiments, R 2 is CO2R', CON(R')2, SO2N(R')2 or OSO2N(R')2. In another embodiment, R 2 It is CONH2, SO2NH2 or OSO2NH2.

[0100] In some embodiments, provided herein is a compound selected from the group consisting of:

[0101]

[0102] or a pharmaceutically acceptable salt, solvate, hydrate, tautomer or optical isomer thereof; or a pharmaceutically acceptable salt thereof.

[0103] In some embodiments, the present invention encompasses a pharmaceutical composition comprising a compound described herein and a pharmaceutically acceptable excipient.

[0104] In some embodiments, the pharmaceutical composition further comprises at least one additional therapeutic agent selected from the group consisting of aminosalicylate, corticosteroid, immunomodulator, and combinations thereof.

[0105] In some embodiments, the invention includes a method of modulating EP4 receptor agonist activity in a biological sample, the method comprising contacting the EP4 receptor with a compound or composition described herein.

[0106] In some embodiments, the present invention includes methods of treating a disease mediated by the EP4 receptor comprising administering to a patient in need thereof a compound or composition described herein.

[0107] In some embodiments, the EP4 receptor-mediated disease is a gastrointestinal disorder. In another embodiment, the gastrointestinal disorder is selected from constipation disorders, constipation-dominant irritable bowel syndrome, mixed irritable bowel syndrome, chronic idiopathic constipation, gastrointestinal symptoms associated with Parkinson's disease, gastrointestinal symptoms associated with cystic fibrosis, intestinal motility disorders, postoperative ileus, food allergies or food intolerance, celiac disease, gastrointestinal motility disorders, functional gastrointestinal disorders, drug-induced bowel disease, NSAID-induced gastric and intestinal damage, chemotherapy-induced mucositis, gastroesophageal reflux disease (GERD), duodenogastric reflux, diarrheal diseases, immune-mediated gastrointestinal diseases, Crohn's disease, ulcerative colitis, inflammatory bowel disease and ischemic colitis, or lung diseases and disorders, such as chronic obstructive pulmonary disease, asthma, chronic bronchitis, cystic fibrosis, emphysema, chronic idiopathic cough, overactive airway disorders and idiopathic pulmonary fibrosis.

[0108] In some embodiments, the compound of Formula I is a compound listed in Table 1 or a pharmaceutically acceptable salt, solvate, hydrate, or tautomer thereof.

[0109] In some embodiments, provided herein is a compound selected from the compounds listed in Table 1 or a pharmaceutically acceptable salt thereof:

[0110] Table 1: Exemplary compounds of formula (1)

[0111]

[0112]

[0113]

[0114] or a pharmaceutically acceptable salt thereof.

[0115] abbreviation

[0116] aq aqueous solution

[0117] Bn benzyl

[0118] DCM dichloromethane

[0119] DMA Dimethylacetamide

[0120] DMF dimethylformamide

[0121] dppf 1,1'-Bis(diphenylphosphino)ferrocene

[0122] EDCI 1-Ethyl-3-[3-dimethylaminopropyl]carbodiimide

[0123] EtOAcEthyl acetate

[0124] FA Formic acid

[0125] HATU 1-[Bis(dimethylamino)methylene]-1H-1,2,3-triazolo[4,5-b]pyridine 3-Oxide Hexafluorophosphate

[0126] HOBt Hydroxybenzotriazole

[0127] HPLC High Performance Liquid Chromatography

[0128] hr hours

[0129] hrs hours

[0130] LCMS liquid chromatography-mass spectrometry

[0131] M Moore

[0132] MeCNAcetonitrile

[0133] MeOHMethanol

[0134] N equivalent (normal)

[0135] NBS N-Bromosuccinimide

[0136] prep HPLC preparative high performance liquid chromatography

[0137] RT

[0138] sat saturated

[0139] THF Tetrahydrofuran

[0140] UPLC High Performance Liquid Chromatography

[0141] definition

[0142] In this application, unless otherwise indicated, the following definitions apply.

[0143] The term "treatment" in connection with the use of any of the compounds described herein (including those of Formula (A)-(C), (la)-(la), (lb)-(lb), (lc), (1), (1a)-(1b), (1c), (2a)-(2b), (2c)-(2d), (3a)-(3b), (3c)-(3d), (3e)-(3f), (5)-(5a), (5b), (6), (6a)-(6b) and (6b)) is used to describe any form of intervention in which the compound is administered to a subject suffering from, at risk of, or potentially at risk of suffering from the disease or condition in question. Thus, the term "treatment" includes both prophylactic (preventive) treatment and treatment that results in measurable or detectable symptoms of a disease or condition.

[0144] The term "therapeutically effective amount" (e.g., in relation to a method of treating a disease or condition) refers to an amount of a compound that is effective to produce the desired therapeutic effect. For example, if the condition is pain, the therapeutically effective amount is an amount sufficient to provide a desired level of pain relief. The desired level of pain relief can be, for example, complete elimination of pain or a reduction in the severity of pain.

[0145] As used herein, the term "hydroxyl" or "hydroxy" refers to an -OH moiety.

[0146] As used herein, "alkyl" refers to a saturated aliphatic hydrocarbon group containing 1 to 12 (e.g., 1 to 8, 1 to 6, or 1 to 4) carbon atoms. The alkyl group can be straight chain or branched. Some examples of alkyl groups include, but are not limited to, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, n-heptyl, or 2-ethylhexyl. The alkyl group may be substituted (i.e., optionally substituted) with one or more substituents, such as halogen, phosphate, alicyclic [e.g., cycloalkyl or cycloalkenyl], heteroalicyclic [e.g., heterocycloalkyl or heterocycloalkenyl], aryl, heteroaryl, alkoxy, aroyl, heteroaroyl, acyl [e.g., (aliphatic)carbonyl, (alicyclic)carbonyl, or (heteroalicyclic)carbonyl], nitro, cyano, amide [e.g., (cycloalkylalkyl)carbonylamino, arylcarbonylamino, aralkylcarbonylamino, (heterocycloalkyl)carbonylamino, (heterocycloalkylalkyl)carbonylamino, heterocycloalkylalkyl)carbonylamino, [e.g., aliphatic amino, alicyclic amino, or heteroalicyclic amino], sulfonyl [e.g., aliphatic -SO2-], sulfinyl, sulfanyl, sulfoxy, urea, thiourea, sulfamoyl, sulfonamide, oxo, carboxyl, carbamoyl, alicyclic oxy, heteroalicyclic oxy, aryloxy, heteroaryloxy, aralkyloxy, heteroarylalkoxy, alkoxycarbonyl, alkylcarbonyloxy, or hydroxyl. Without limitation, some examples of substituted alkyl groups include carboxyalkyl (e.g., HOOC-alkyl, alkoxycarbonylalkyl, and alkylcarbonyloxyalkyl), cyanoalkyl, hydroxyalkyl, alkoxyalkyl, acylalkyl, aralkyl, (alkoxyaryl)alkyl, (sulfonylamino)alkyl (e.g., (alkyl-SO2-amino)alkyl), aminoalkyl, amidoalkyl, (alicyclic)alkyl, or haloalkyl.

[0147] As used herein, an "alkylene" group refers to a divalent branched or straight chain alkyl group containing 2 to 12 (e.g., 2 to 8, 2 to 6, or 2 to 4) carbon atoms and is used to connect two chemical moieties. Some examples of alkylene include, but are not limited to, methylene, ethylene, propylene, butylene, isopropylidene (methylethylene) and isobutylene (2-methylpropylene). Alkylene can be substituted (i.e., optionally substituted) by one or more substituents as defined in the alkyl group.

[0148] As used herein, "amido" encompasses both "aminocarbonyl" and "carbonylamino". These terms, when used alone or in conjunction with another group, refer to an amide group, for example, when used terminally, the term refers to -N(R X )-C(O)-R Y or -C(O)-N(R X)2, and when used internally, the term refers to -C(O)-N(R X )-or-N(R X )-C(O)-, where R X and R Y It can be aliphatic, alicyclic, aryl, araliphatic, heteroalicyclic, heteroaryl or heteroaraliphatic. Some examples of amide groups include alkylamide groups (e.g., alkylcarbonylamino or alkylaminocarbonyl), (heteroalicyclic)amide groups, (heteroaralkyl)amide groups, (heteroaryl)amide groups, (heterocycloalkyl)alkylamide groups, arylamide groups, aralkylamide groups, (cycloalkyl)alkylamide groups or cycloalkylamide groups.

[0149] As used herein, "amino" refers to -NR X R Y , where R X and R Y Each of is independently hydrogen, aliphatic, alicyclic, (alicyclic) aliphatic, aryl, araliphatic, heteroalicyclic, (heteroalicyclic) aliphatic, heteroaryl, carboxyl, sulfanyl, sulfinyl, sulfonyl, (aliphatic) carbonyl, (alicyclic) carbonyl, ((alicyclic) aliphatic) carbonyl, arylcarbonyl, (araliphatic) carbonyl, (heteroalicyclic) carbonyl, ((heteroalicyclic) aliphatic) carbonyl, (heteroaryl) carbonyl or (heteroaraliphatic) carbonyl, each of which is defined herein and is optionally substituted. Some examples of amino include alkylamino, dialkylamino or arylamino. When the term "amino" is not a terminal group (e.g., alkylcarbonylamino), it is represented by -NR X - indicates that R X Has the same meaning as defined above.

[0150] As used herein, "aryl" used alone or as part of a larger moiety in "aralkyl", "aralkyloxy" or "aryloxyalkyl" refers to monocyclic (e.g., phenyl); bicyclic (e.g., indenyl, naphthyl, tetrahydronaphthyl, tetrahydroindenyl); and tricyclic (e.g., fluorenyl, tetrahydrofluorenyl, or tetrahydroanthracenyl, anthracenyl) ring systems, wherein the monocyclic system is aromatic or at least one of the rings in the bicyclic or tricyclic system is aromatic. Bicyclic and tricyclic groups include benzo-fused 2- to 3-membered carbocyclic rings. For example, benzo-fused groups include benzo-fused 2- to 3-membered carbocyclic rings with two or more C 4-8The aryl group is optionally substituted with one or more substituents including: aliphatic [e.g., alkyl, alkenyl, or alkynyl]; alicyclic; (alicyclic) aliphatic; heteroalicyclic; (heteroalicyclic) aliphatic; aryl; heteroaryl; alkoxy; (alicyclic) oxy; (heteroalicyclic) oxy; aryloxy; heteroaryloxy; (aroliphatic) oxy; (heteroaroaliphatic) oxy; aroyl; heteroaroyl; amino; oxo (on the non-aromatic carbocyclic ring of the benzo-fused bicyclic or tricyclic aryl group); nitro; carboxyl; amide; acyl [e.g., For example, (aliphatic) carbonyl; (alicyclic) carbonyl; ((alicyclic) aliphatic) carbonyl; (aromatic) carbonyl; (heteroalicyclic) carbonyl; ((heteroalicyclic) aliphatic) carbonyl; or (heteroaromatic) carbonyl]; sulfonyl [e.g., aliphatic -SO2- or amino-SO2-]; sulfinyl [e.g., aliphatic -S(O)- or alicyclic -S(O)-]; sulfanyl [e.g., aliphatic -S-]; cyano; halogen; hydroxyl; mercapto; sulfoxyl; urea; thiourea; sulfamoyl; sulfonamide; or carbamoyl. Alternatively, the aryl group can be unsubstituted.

[0151] Some non-limiting examples of substituted aryl groups include haloaryl [e.g., monohaloaryl, dihaloaryl (e.g., p-, m-dihaloaryl), and (trihalo)aryl]; (carboxy)aryl [e.g., (alkoxycarbonyl)aryl, ((aralkyl)carbonyloxy)aryl, and (alkoxycarbonyl)aryl]; (amido)aryl [e.g., (aminocarbonyl)aryl, (((alkylamino)alkyl)aminocarbonyl)aryl, (alkylcarbonyl)aminoaryl, (arylaminocarbonyl)aryl, and (((heteroaryl)amino)carbonyl)aryl]; aminoaryl [e.g., ((alkylsulfonyl)amino)aryl or ((dialkyl)amino)aryl]; (cyanoalkyl)aryl; (alkoxy)aryl; (sulfamoyl)aryl )aryl [e.g., (aminosulfonyl)aryl]; (alkylsulfonyl)aryl; (cyano)aryl; (hydroxyalkyl)aryl; ((alkoxy)alkyl)aryl; (hydroxy)aryl, ((carboxy)alkyl)aryl; (((dialkyl)amino)alkyl)aryl; (nitroalkyl)aryl; (((alkylsulfonyl)amino)alkyl)aryl; ((heteroalicyclic)carbonyl)aryl; ((alkylsulfonyl)alkyl)aryl; (cyanoalkyl)aryl; (hydroxyalkyl)aryl; (alkylcarbonyl)aryl; alkylaryl; (trihaloalkyl)aryl; p-amino-m-alkoxycarbonylaryl; p-amino-m-cyanoaryl; p-halo-m-aminoaryl; or (m-(heteroalicyclic)-o-(alkyl))aryl.

[0152] As used herein, a "cycloalkyl" group refers to a saturated carbocyclic monocyclic or bicyclic (fused or bridged) ring of 3 to 10 (e.g., 5 to 10) carbon atoms. Some examples of cycloalkyl include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, adamantyl, norbornyl, cubicyl, octahydro-indenyl, decahydro-naphthyl, bicyclo[3.2.1]octyl, bicyclo[2.2.2]octyl, bicyclo[3.3.1]nonyl, bicyclo[3.3.2.]decyl, bicyclo[2.2.2]octyl, adamantyl, or ((aminocarbonyl)cycloalkyl)cycloalkyl.

[0153] The cycloalkyl radical may be optionally substituted with one or more substituents such as phosphate, aliphatic [e.g., alkyl, alkenyl, or alkynyl], cycloaliphatic, (cycloaliphatic)aliphatic, heterocycloaliphatic, (heterocycloaliphatic)aliphatic, aryl, heteroaryl, alkoxy, (cycloaliphatic)oxy, (heterocycloaliphatic)oxy, aryloxy, heteroaryloxy, (araliphatic)oxy, (heteroaroaliphatic)oxy, aroyl, heteroaroyl, amino, amido [e.g., (aliphatic)carbonylamino, (cycloaliphatic)carbonylamino, ((cycloaliphatic)aliphatic)carbonylamino, (aryl)carbonylamino, (araliphatic)carbonylamino, (heterocycloaliphatic)carbonylamino, ((heterocycloaliphatic)aliphatic) [e.g., alkyl-S-], sulfoxyl, urea, thiourea, sulfamoyl, sulfonyl, or carbamoyl groups.

[0154] As used herein, "heterocycloalkyl" refers to a 3- to 10-membered monocyclic or bicyclic (fused or bridged) (e.g., a 5- to 10-membered monocyclic or bicyclic) saturated ring structure in which one or more of the ring atoms is a heteroatom (e.g., N, O, S, or a combination thereof). Some examples of heterocycloalkyl include piperidinyl, piperazinyl, tetrahydropyranyl, tetrahydrofuranyl, 1,4-dioxolanyl, 1,4-dithianyl, 1,3-dioxolanyl, Oxazoline, iso oxazolinyl, morpholinyl, thiomorpholyl, octahydrobenzofuranyl, octahydrochromenyl, octahydrothiochromenyl, octahydroindolyl, octahydropyridinyl, decahydroquinolinyl, octahydrobenzo[b]thienyl, 2-oxa-bicyclo[2.2.2]octyl, 1-aza-bicyclo[2.2.2]octyl, 3-aza-bicyclo[3.2.1]octyl and 2,6-dioxa-tricyclo[3.3.1.0 3，7 A monocyclic heterocycloalkyl group may be fused with a phenyl moiety to form a structure such as tetrahydroisoquinoline, which would be classified as a heteroaryl group.

[0155] The heterocycloalkyl radical may be optionally substituted with one or more substituents such as phosphate, aliphatic [e.g., alkyl, alkenyl, or alkynyl], alicyclic, (alicyclic)aliphatic, heteroalicyclic, (heteroalicyclic)aliphatic, aryl, heteroaryl, alkoxy, (alicyclic)oxy, (heteroalicyclic)oxy, aryloxy, heteroaryloxy, (araliphatic)oxy, (heteroaraliphatic)oxy, aroyl, heteroaroyl, amino, amido [e.g., (aliphatic)carbonylamino, (alicyclic)carbonylamino, ((alicyclic)aliphatic)carbonylamino, (aryl)carbonylamino, (araliphatic)carbonylamino, (heteroalicyclic)carbonylamino, ((heteroalicyclic) [e.g., (cycloaliphatic)carbonyl, ((cycloaliphatic)aliphatic)carbonyl, (araliphatic)carbonyl, (heterocycloaliphatic)carbonyl, ((heterocycloaliphatic)aliphatic)carbonyl, or (heteroaroaliphatic)carbonyl], nitro, cyano, halogen, hydroxy, mercapto, sulfonyl [e.g., alkylsulfonyl or arylsulfonyl], sulfinyl [e.g., alkylsulfinyl], sulfanyl [e.g., alkylsulfanyl], sulfoxy, urea, thiourea, sulfamoyl, sulfonamide, oxo, or carbamoyl.

[0156] As used herein, "heteroaryl" refers to a monocyclic, bicyclic or tricyclic ring system having 4 to 15 ring atoms, wherein one or more of the ring atoms are heteroatoms (e.g., N, O, S or a combination thereof), and wherein the monocyclic ring system is aromatic, or at least one ring in the bicyclic or tricyclic ring system is aromatic. Heteroaryl includes benzo-fused ring systems having 2 to 3 rings. For example, a benzo-fused group includes a benzo fused to one or two 4 to 8-membered heteroalicyclic moieties (e.g., indolizyl, indolyl, isoindolyl, 3H-indolyl, dihydroindole, benzo[b]furanyl, benzo[b]thiophene-yl, quinolyl or isoquinolyl). Some examples of heteroaryl are azetidinyl, pyridyl, 1H-indazolyl, furyl, pyrrolyl, thienyl, thiazolyl, oxazolyl, imidazolyl, tetrazolyl, benzofuranyl, isoquinolyl, benzothiazolyl, anthenyl, thioxanthenyl, phenothiazinyl, indoline, benzo[1,3]dioxole, benzo[b]furanyl, benzo[b]thienyl, indazolyl, benzimidazolyl, benzothiazolyl, puryl, cinnolyl, quinolyl, quinazolinyl, cinnolinyl, phthalazyl, quinazolinyl, quinoxalyl, isoquinolyl, 4H-quinolizyl, benzo-1,2,5-thiadiazolyl or 1,8-naphthyridyl.

[0157] Without limitation, monocyclic heteroaryl groups include furanyl, thiophene-yl, 2H-pyrrolyl, pyrrolyl, oxazolyl, thiazolyl, imidazolyl, pyrazolyl, isocyanate oxazolyl, isothiazolyl, 1,3,4-thiadiazolyl, 2H-pyranyl, 4-H-pyranyl, pyridinyl, pyridazyl, pyrimidinyl, pyrazolyl, pyrazyl or 1,3,5-triazyl. Monocyclic heteroaryl groups are numbered according to standard chemical nomenclature.

[0158] Without limitation, bicyclic heteroaryl groups include indolizyl, indolyl, isoindolyl, 3H-indolyl, dihydroindole, benzo[b]furanyl, benzo[b]thienyl, quinolyl, isoquinolyl, indolizyl, isoindolyl, indolyl, benzo[b]furanyl, bexo[b]thienyl, indazolyl, benzimidazolyl, benzothiazolyl, purinyl, 4H-quinolizinyl, quinolyl, isoquinolyl, cinnolinyl, phthalazinyl, quinazolinyl, quinoxalinyl, 1,8-naphthyridinyl or pteridinyl. Bicyclic heteroaryl groups are numbered according to standard chemical nomenclature.

[0159] The heteroaryl group is optionally substituted by one or more substituents such as: aliphatic [e.g., alkyl, alkenyl, or alkynyl]; alicyclic; (alicyclic)aliphatic; heteroalicyclic; (heteroalicyclic)aliphatic; aryl; heteroaryl; alkoxy; (alicyclic)oxy; (heteroalicyclic)oxy; aryloxy; heteroaryloxy; (aroliphatic)oxy; (heteroaroaliphatic)oxy; aroyl; heteroaroyl; amino; oxo (on the non-aromatic carbocyclic or heterocyclic ring of the bicyclic or tricyclic heteroaryl); carboxyl; amide The heteroaryl group may be substituted with acyl, acyl [e.g., aliphatic carbonyl; (alicyclic) carbonyl; ((alicyclic) aliphatic) carbonyl; (aromatic) carbonyl; (heteroalicyclic) carbonyl; ((heteroalicyclic) aliphatic) carbonyl; or (heteroaromatic) carbonyl]; sulfonyl [e.g., aliphatic sulfonyl or aminosulfonyl]; sulfinyl [e.g., aliphatic sulfinyl]; sulfanyl [e.g., aliphatic sulfanyl]; nitro; cyano; halogen; hydroxyl; mercapto; sulfoxyl; urea; thiourea; sulfamoyl; sulfonamide; or carbamoyl. Alternatively, the heteroaryl group may be unsubstituted.

[0160] Some non-limiting examples of substituted heteroaryl groups include (halo)heteroaryl [e.g., monohaloheteroaryl and dihaloheteroaryl]; (carboxy)heteroaryl [e.g., (alkoxycarbonyl)heteroaryl]; cyanoheteroaryl; aminoheteroaryl [e.g., ((alkylsulfonyl)amino)heteroaryl and ((dialkyl)amino)heteroaryl]; (amido)heteroaryl [e.g., aminocarbonylheteroaryl, ((alkylcarbonyl)amino)heteroaryl, ((((alkyl)amino)alkyl)aminocarbonyl)heteroaryl, (((heteroaryl)amino)carbonyl)heteroaryl, ((heteroalicyclic)carbonyl)heteroaryl, and ((alkylcarbonyl)amino)heteroaryl]; (cyanoalkyl)heteroaryl; (alkoxy)heteroaryl; ((alkylsulfonyl)amino)heteroaryl; ((alkylsulfonyl)amino)heteroaryl; ((dialkyl)amino)heteroaryl) heteroaryl]; (alkyl)heteroaryl ...

[0161] As used herein, an "alkoxy" group refers to an alkyl-O- group, wherein "alkyl" has been previously defined.

[0162] As used herein, a "carboxyl" group, when used as a terminal group, refers to -COOH, -COOR X 、-OC(O)H、-OC(O)R X; or when used as an internal group, a "carboxy" group refers to -OC(O)- or -C(O)O-.

[0163] As used herein, a "thiol" group refers to -SH.

[0164] As used herein, a "sulfo" group when used terminally refers to -SO3H or -SO3R X , or -S(O)3- when used internally.

[0165] As used herein, a "sulfonamide" group when used terminally refers to a group of the structure -NR X -S(O)2-NR Y R Z , and when used internally refers to the structure -NR X -S(O)2-NR Y -, where R X , R Y and R Z Defined above.

[0166] As used herein, a "sulfamoyl" group refers to a group of the structure -OS(O)2-NR Y R Z , where R Y and R Z Defined above.

[0167] As used herein, a "sulfonamide" group when used terminally refers to the structure -S(O)2-NR x R y or -NR x -S(O)2-R z ; or when used internally, -S(O)2-NR x -or-NR x -S(O)2-, where R x , R y and R Z Defined above.

[0168] As used herein, a "sulfanyl" group when used terminally refers to -SR X , and when used internally refers to -S-, where R X Some examples of sulfanyl groups include aliphatic-S-, alicyclic-S-, aryl-S-, and the like.

[0169] As used herein, a "halogen" or "halo" group refers to fluorine, chlorine, bromine, or iodine.

[0170] As used herein, "oxo" refers to =0.

[0171] As used herein, the term "vicinal" generally refers to the positioning of substituents on a group containing two or more carbon atoms wherein the substituents are attached to adjacent carbon atoms.

[0172] As used herein, the term "geminal" generally refers to the position of substituents on a group containing two or more carbon atoms, wherein the substituents are attached to the same carbon atom.

[0173] The terms "terminal" and "internal" refer to the position of a group within a substituent. A group is terminal when it is present at the end of a substituent and is no longer bonded to the rest of the chemical structure. X O(O)C-alkyl) is an example of a carboxyl group used terminally. When the group is present in the middle of a substituent in a chemical structure, the group is internal. Alkylcarboxyl (e.g., alkyl-C(O)O- or alkyl-OC(O)-) and alkylcarboxyaryl (e.g., alkyl-C(O)O-aryl or alkyl-O(CO)-aryl-) are some examples of carboxyl groups used internally.

[0174] As used herein, "aliphatic chain" refers to a branched or straight chain aliphatic group (e.g., an alkyl, alkenyl, or alkynyl group). A straight chain aliphatic chain has the structure -[CH2] v -, where v is 1 to 12. A branched aliphatic chain is a straight aliphatic chain substituted with one or more aliphatic groups. A branched aliphatic chain has the structure -[CQQ] v -, wherein Q is independently hydrogen or an aliphatic group; however, in at least one instance, Q shall be an aliphatic group. The term aliphatic chain includes alkyl chains, alkenyl chains, and alkynyl chains, wherein alkyl, alkenyl, and alkynyl are defined above.

[0175] The phrase "optionally substituted" is used interchangeably herein with the phrase "substituted or unsubstituted". As described herein, the compounds of the invention may be optionally substituted with one or more substituents, such as are generally exemplified above, or exemplified by specific classes, subclasses, and species of the invention. Unless otherwise indicated, each specific group of variables described herein may be optionally substituted with one or more substituents described herein. Each substituent of a specific group is also optionally substituted with one to three of halogen, cyano, oxo, alkoxy, hydroxy, amino, nitro, aryl, alicyclic, heteroalicyclic, heteroaryl, haloalkyl, and alkyl. For example, an alkyl group may be substituted with an alkylsulfanyl group, and an alkylsulfanyl group may be optionally substituted with one to three of halogen, cyano, oxo, alkoxy, hydroxy, amino, nitro, aryl, haloalkyl, and alkyl. As another example, the cycloalkyl moiety of (cycloalkyl)carbonylamino can be optionally substituted by one to three of halogen, cyano, alkoxy, hydroxy, nitro, haloalkyl and alkyl. When two alkoxy groups are bound to the same atom or adjacent atoms, the two alkoxy groups can form a ring together with the atoms to which they are bound.

[0176] As used herein, the term "substituted", whether or not there is the term "optionally" in front, generally refers to that the hydrogen atoms in a given structure are replaced by the group of a specific substituent. Specific substituents are described in the above definition and in the following description of compounds and embodiments thereof. Unless otherwise indicated, the optionally substituted group may have a substituent at each substitutable position of the group, and when more than one position in any given structure may be substituted by more than one substituent selected from a specific group, the substituent at each position may be the same or different. Ring substituents such as heterocycloalkyl can be combined with another ring such as cycloalkyl to form a spiro-bicyclic system, such as two rings share a common atom. As will be appreciated by those of ordinary skill in the art, the combination of substituents contemplated by the present invention is those combinations that result in the formation of stable or chemically feasible compounds.

[0177] As used herein, the phrase "stable or chemically feasible" refers to a compound that is not substantially altered when subjected to conditions that allow its production, detection, and preferably its recovery, purification, and use for one or more purposes disclosed herein. In some embodiments, a stable compound or a chemically feasible compound is one that is not substantially altered when kept at 40° C. or less for at least one week in the absence of moisture or other chemically reactive conditions.

[0178] Insofar as any of the compounds described have chiral centers, the present invention extends to all optical isomers of such compounds, whether in racemic or resolved enantiomeric form. The invention described herein relates to all crystalline forms, solvates and hydrates of any disclosed compound, no matter how prepared. Insofar as any of the compounds disclosed herein have acid or basic centers such as carboxylate / ester or amino groups, all salt forms of the compounds are included herein. The invention described herein also relates to all tautomers of the compounds described herein. In the case of pharmaceutical use, the salts should be considered as pharmaceutically acceptable salts.

[0179] Salts or pharmaceutically acceptable salts that may be mentioned include acid addition salts and base addition salts. Such salts can be formed, for example, by the following conventional methods: by reacting the compound in free acid or free base form with one or more equivalents of a suitable acid or base, optionally in a solvent or medium in which the salt is not soluble, followed by removal of the solvent or medium using standard techniques (e.g. in vacuo, by freeze drying or by filtration). Salts can also be prepared by exchanging the counterion of the compound in salt form with another counterion, for example using a suitable ion exchange resin.

[0180] Some examples of pharmaceutically acceptable salts include acid addition salts derived from inorganic and organic acids, and salts derived from metals such as sodium, magnesium, potassium, and calcium.

[0181] Some examples of acid addition salts include acid addition salts formed with acetic acid, 2,2-dichloroacetic acid, adipic acid, alginic acid, arylsulfonic acids (e.g., benzenesulfonic acid, naphthalene-2-sulfonic acid, naphthalene-1,5-disulfonic acid, and p-toluenesulfonic acid), ascorbic acid (e.g., L-ascorbic acid), L-aspartic acid, benzoic acid, 4-acetamidobenzoic acid, butyric acid, (+) camphoric acid, camphorsulfonic acid, (+)-(1S)-camphor-10-sulfonic acid, decanoic acid, hexanoic acid, caprylic acid, cinnamic acid, citric acid, cyclohexanesulfamic acid, dodecylsulfuric acid, ethane-1,2-disulfonic acid, ethanesulfonic acid, 2-hydroxyethanesulfonic acid, formic acid, fumaric acid, galactaric acid, gentisic acid, glucoheptonic acid, gluconic acid (e.g., D-gluconic acid), glucuronic acid (e.g.,

[0013] Examples of the present invention include, but are not limited to, D-glucuronic acid), glutamic acid (e.g., L-glutamic acid), α-oxoglutaric acid, glycolic acid, hippuric acid, hydrobromic acid, hydrochloric acid, hydroiodic acid, isethionic acid, lactic acid (e.g., (+)-L-lactic acid and (±)-DL-lactic acid), lactobionic acid, maleic acid, malic acid (e.g., (-)-L-malic acid), malonic acid, (±)-DL-mandelic acid, metaphosphoric acid, methanesulfonic acid, 1-hydroxy-2-naphthoic acid, nicotinic acid, nitric acid, oleic acid, orotic acid, oxalic acid, palmitic acid, pamoic acid, phosphoric acid, propionic acid, L-pyroglutamic acid, salicylic acid, 4-aminosalicylic acid, sebacic acid, stearic acid, succinic acid, sulfuric acid, tannic acid, tartaric acid (e.g., (+)-L-tartaric acid), thiocyanic acid, undecylenic acid, and valeric acid.

[0182] Any solvate of the compound and its salt is also contemplated.Preferred solvates are solvates formed by incorporating nontoxic pharmaceutically acceptable solvent molecules (hereinafter referred to as solvating solvents) into the solid state structure (e.g., crystal structure) of the compound of the present invention.Some examples of such solvents include water, alcohols (e.g., ethanol, isopropanol and butanol) and dimethyl sulfoxide.Solvates can be prepared by recrystallizing the compound of the present invention with a solvent or a mixture of solvents containing a solvating solvent. Whether a solvate has been formed in any given case can be determined by analyzing the crystals of the compound using known and standard techniques such as thermogravimetric analysis (TGA), differential scanning calorimetry (DSC) and X-ray crystallography.

[0183] Solvates can be stoichiometric or non-stoichiometric solvates. Specific solvates can be hydrates, and some examples of hydrates include hemihydrates, monohydrates, and dihydrates. For a more detailed discussion of solvates and methods for preparing and characterizing them, see Bryn et al, Solid-State Chemistry of Drugs, Second Edition, published by SSCI, Inc of West Lafayette, IN, USA, 1999, ISBN 0-967-06710-3.

[0184] The term "pharmaceutical composition" in the context of the present invention means a composition comprising an active agent and additionally comprising one or more pharmaceutically acceptable carriers or pharmaceutically acceptable excipients. The composition may also comprise ingredients selected from, for example, diluents, adjuvants, excipients, carriers, preservatives, fillers, disintegrants, wetting agents, emulsifiers, suspending agents, sweeteners, flavoring agents, aromatics, antibacterial agents, antifungal agents, lubricants and dispersants, depending on the nature of the administration mode and dosage form. The composition may take the form of, for example, tablets, dragees, powders, elixirs, syrups, liquid preparations (including suspensions, sprays, inhalants, tablets, lozenges, emulsions, solutions, cachets, granules, capsules and suppositories) and liquid preparations for injection (including liposomal preparations).

[0185] The compounds of the present invention may contain one or more isotopic substitutions, and reference to a particular element includes within its scope all isotopes of that element. For example, reference to hydrogen includes within its scope all isotopes of that element. 1 H. 2 H(D) and 3 H(T). Similarly, references to carbon and oxygen include within their scope 12 C. 13C and 14 C and 16 O and 18 O. In a similar manner, references to specific functional groups also include isotopic variations within their scope unless the context indicates otherwise. For example, references to alkyl groups such as ethyl or alkoxy groups (e.g., methoxy) also include variants in which one or more hydrogen atoms in the group are in deuterated or tritium isotopic form, such as ethyl in which all five hydrogen atoms are in deuterated isotopic form (perdeuterated ethyl) or methoxy in which all three hydrogen atoms are in deuterated isotopic form (trideuterated methoxy). Isotopes may be radioactive or non-radioactive.

[0186] The therapeutic dose may vary depending on the patient's needs, the severity of the condition being treated, and the compound being employed. It is within the skill of the art to determine the appropriate dose for a particular situation. Typically, treatment begins with a smaller dose that is less than the optimal dose of the compound. Thereafter, the dose is increased in small increments until the optimal effect is achieved in the situation. For convenience, the total daily dose may be divided and administered in batches throughout the day if desired.

[0187] Of course, the size of the compound effective dose will vary with the severity of the condition to be treated and the specific compound and its route of administration. The selection of appropriate doses is within the capabilities of those of ordinary skill in the art, without excessive burden. Typically, daily dose ranges can be from about 10 μg to about 30 mg / kg human and non-human animal body weight, preferably from about 50 μg to about 30 mg / kg human and non-human animal body weight, for example, from about 50 μg to about 10 mg / kg human and non-human animal body weight, for example, from about 100 μg to about 30 mg / kg human and non-human animal body weight, for example, from about 100 μg to about 10 mg / kg human and non-human animal body weight, and most preferably from about 100 μg to about 1 mg / kg human and non-human animal body weight.

[0188] Combination therapy

[0189] In the methods or pharmaceutical compositions of the present invention, an effective amount can be achieved by using a compound of the present invention (including a pharmaceutically acceptable salt or solvate (e.g., hydrate)) alone or in combination with another suitable therapeutic agent (e.g., an antiviral agent or a vaccine). When "combination therapy" is employed, an effective amount can be achieved by using a first amount of a compound of the present invention and a second amount of another suitable therapeutic agent.

[0190] In another embodiment of the present invention, the compounds of the present invention and the additional therapeutic agent are each administered in an effective amount (i.e., if administered alone, each is administered in an amount that would be therapeutically effective). In another embodiment, the compounds of the present invention and the additional therapeutic agent are each administered in an amount that alone does not provide a therapeutic effect (a subtherapeutic dose). In another embodiment, the compounds of the present invention may be administered in an effective amount, while the additional therapeutic agent is administered in a subtherapeutic dose. In another embodiment, the compounds of the present invention may be administered in a subtherapeutic dose, while the additional therapeutic agent, such as a suitable cancer therapeutic agent, is administered in an effective amount.

[0191] As used herein, the terms "combination" or "co-administration" are used interchangeably to refer to the use of more than one treatment (e.g., one or more prophylactic and / or therapeutic agents). The use of these terms does not limit the order in which the treatments (e.g., prophylactic and / or therapeutic agents) are administered to a subject.

[0192] Co-administration encompasses administration of the co-administered first and second amounts of the compound in a substantially simultaneous manner, such as in a single pharmaceutical composition, e.g., a capsule or tablet having a fixed ratio of the first and second amounts, or in a plurality of separate capsules or tablets of each. In addition, such co-administration also encompasses the use of each compound in a sequential manner in either order.

[0193] In one embodiment, the compound of the invention and the additional therapeutic agent are administered to a subject separately, sequentially or simultaneously.

[0194] When co-administration involves separate administration of a first amount of a compound of the invention and a second amount of an additional therapeutic agent, the compound is administered in close enough time to have the desired therapeutic effect. For example, the time period between each administration that can produce the desired therapeutic effect can be from several minutes to several hours, and can be determined based on the properties of each compound (e.g., efficacy, solubility, bioavailability, plasma half-life, and kinetic profile). For example, the compound of the invention and the second therapeutic agent can be administered in any order within 24 hours of each other, within 16 hours of each other, within 8 hours of each other, within 4 hours of each other, within 1 hour of each other, or within 30 minutes of each other.

[0195] More specifically, a first therapy (e.g., a prophylactic or therapeutic agent, such as a compound of the invention) can be administered to a subject prior to (e.g., 5 minutes, 15 minutes, 30 minutes, 45 minutes, 1 hour, 2 hours, 4 hours, 6 hours, 12 hours, 24 hours, 48 ​​hours, 72 hours, 96 hours, 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 8 weeks, or 12 weeks before), concurrently with, or after (e.g., 5 minutes, 15 minutes, 30 minutes, 45 minutes, 1 hour, 2 hours, 4 hours, 6 hours, 12 hours, 24 hours, 48 ​​hours, 72 hours, 96 hours, 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 8 weeks, or 12 weeks after) the administration of a second therapy (e.g., a prophylactic or therapeutic agent, such as an anti-cancer agent).

[0196] It is understood that the method of co-administering a first amount of a compound of the present invention and a second amount of an additional therapeutic agent can produce an enhanced or synergistic therapeutic effect, wherein the combined effect is greater than the additive effect produced by separately administering the first amount of the compound of the present invention and the second amount of the additional therapeutic agent.

[0197] The term "synergistic" as used herein refers to a combination of a compound of the invention and another treatment (e.g., a prophylactic or therapeutic agent) that is more effective than the additive effect of the treatment. The synergistic effect of a combination of treatments (e.g., a combination of prophylactics or a combination of therapeutics) can allow the use of lower doses of one or more treatments and / or less frequent administration of the treatment to a subject. The ability to use lower doses of a treatment (e.g., a prophylactic or therapeutic agent) and / or less frequent administration of the treatment can reduce the toxicity associated with administering the treatment to a subject without reducing the effectiveness of the treatment in preventing, controlling or treating a condition. In addition, synergy can result in an improved effectiveness of a medicament in preventing, managing or treating a condition. Finally, the synergistic effect of a combination therapy (e.g., a combination of prophylactics or a combination of therapeutics) can avoid or reduce adverse or undesirable side effects associated with the use of any one treatment alone.

[0198] The presence of synergy can be determined using a suitable method for evaluating drug interactions. Suitable methods include, for example, Sigmoid-Emax equation (Holford, NHG and Scheiner, LB, Clin. Pharmacokinet. 6: 429-453 (1981)), Loewe additivity equation (Loewe, S. and Muischnek, H., Arc h. Exp. Pathol Pharmacol. 114: 313-326 (1926)) and median effect equation (Chou, TC and Talalay, P., Adv. Enzyme Regul. 22: 27-55 (1984)). Each equation mentioned above can be applied together with experimental data to produce corresponding figures for helping to evaluate the effect of drug combinations. The corresponding graphics associated with the equation mentioned above are concentration-action curves, isobologram curves and combination index curves, respectively.

[0199] In one aspect, the present invention provides a pharmaceutical composition comprising a compound of the present invention, a pharmaceutically acceptable salt, solvate, hydrate, tautomer or optical isomer, a pharmaceutically acceptable excipient, and at least one additional therapeutic agent. In some embodiments, the compound of the present invention, a pharmaceutically acceptable salt, solvate, hydrate, tautomer or optical isomer, and at least one additional therapeutic agent are co-formulated. In some embodiments, the compound of the present invention, a pharmaceutically acceptable salt, solvate, hydrate, tautomer or optical isomer, and at least one additional therapeutic agent are separately formulated.

[0200] In another aspect, the present invention provides a kit comprising a compound, a pharmaceutically acceptable salt, a solvate, a hydrate, a tautomer or an optical isomer of the present invention and at least one additional therapeutic agent. The kit may include instructions for administering a compound, a pharmaceutically acceptable salt, a solvate, a hydrate, a tautomer or an optical isomer of the present invention and at least one additional therapeutic agent to a subject in need thereof.

[0201] In another aspect, the present invention provides a combination therapy for use as a medicament, wherein the combination therapy comprises administering to a subject in need thereof a compound, pharmaceutically acceptable salt, solvate, hydrate, tautomer or optical isomer of the present invention, and administering to a subject in need thereof an additional therapeutic agent.

[0202] The compound, pharmaceutically acceptable salt, solvate, hydrate, tautomer or optical isomer of the present invention and the additional therapeutic agent may be administered to a subject separately, sequentially or simultaneously.

[0203] Pharmaceutical compositions, kits and / or combination therapies can be used to treat gastrointestinal disorders or pulmonary diseases or disorders. Gastrointestinal disorders can be selected from constipation disorders, constipation-dominant irritable bowel syndrome, mixed irritable bowel syndrome, chronic idiopathic constipation, gastrointestinal symptoms associated with Parkinson's disease, gastrointestinal symptoms associated with cystic fibrosis, intestinal motility disorders, postoperative ileus, food allergies or food intolerance, celiac disease, gastrointestinal motility disorders, functional gastrointestinal disorders, drug-induced enteropathy, NSAID-induced gastric and intestinal damage, chemotherapy-induced mucositis, gastroesophageal reflux disease (GERD), duodenogastric reflux, diarrheal diseases, immune-mediated gastrointestinal diseases, Crohn's disease, ulcerative colitis, inflammatory bowel disease and ischemic colitis. Pulmonary diseases or disorders can be selected from chronic obstructive pulmonary disease, asthma, chronic bronchitis, cystic fibrosis, emphysema, chronic idiopathic cough, overactive airway disorders and idiopathic pulmonary fibrosis.

[0204] At least one additional therapeutic agent may be selected from aminosalicylate, corticosteroid, immunomodulator and combination thereof. Aminosalicylate is also known as 5-aminosalicylate (5-ASA). At least one additional therapeutic agent may be aminosalicylate. Aminosalicylate may be mesalamine. Aminosalicylate may be sulfasalazine. At least one additional therapeutic agent may be a corticosteroid. A corticosteroid may be budesonide. At least one additional therapeutic agent may be an immunomodulator. An immunomodulator may be a thiopurine. An immunomodulator may be methotrexate.

[0205] Pharmaceutical preparations

[0206] While it is possible for the active compound to be administered alone, it is preferable to present it as a pharmaceutical composition (eg, formulation).

[0207] Therefore, in another embodiment of the present invention, there is provided a pharmaceutical composition comprising at least one compound of formula (I) as defined above and at least one pharmaceutically acceptable excipient.

[0208] When the pharmaceutical composition comprises at least one additional therapeutic agent, the compound of the invention, pharmaceutically acceptable salt, solvate, hydrate, tautomer or optical isomer and the at least one additional therapeutic agent may be co-formulated, or the compound of the invention, pharmaceutically acceptable salt, solvate, hydrate, tautomer or optical isomer and the at least one additional therapeutic agent may be formulated separately.

[0209] The composition may be a tablet composition.

[0210] The composition may be a capsule composition.

[0211] Pharmaceutically acceptable excipients may be selected, for example, from carriers (e.g., solid, liquid or semisolid carriers), excipients, diluents (e.g., solid diluents, such as fillers or extenders; and liquid diluents, such as solvents and co-solvents), granulating agents, binders, glidants, coating agents, release controlling agents (e.g., release-delaying or -retarding polymers or waxes), adhesives, disintegrants, buffers, lubricants, preservatives, antifungal and antibacterial agents, antioxidants, buffers, tonicity modifiers, thickeners, flavoring agents, sweeteners, pigments, plasticizers, taste-masking agents, stabilizers or any other excipients conventionally used in pharmaceutical compositions.

[0212] The term "pharmaceutically acceptable" as used herein means compounds, materials, compositions and / or dosage forms that are, within the scope of sound medical judgment, suitable for contact with the tissues of subjects (e.g., human subjects) without excessive toxicity, irritation, allergic response, or other problems or complications, commensurate with a reasonable benefit / risk ratio. Each excipient must also be "acceptable" in the sense of being compatible with the other ingredients of the formulation.

[0213] Pharmaceutical compositions containing compounds of formula (I) may be formulated according to known techniques, see for example Remington's Pharmaceutical Sciences, Mack Publishing Company, Easton, PA, USA.

[0214] The pharmaceutical composition may be in any form suitable for oral, parenteral, topical, intranasal, intrabronchial, sublingual, ocular, otic, rectal, intravaginal or transdermal administration.

[0215] Suitable pharmaceutical dosage forms for oral administration include tablets (coated or uncoated), capsules (hard or soft shell), caplets, pills, lozenges, syrups, solutions, powders, granules, elixirs and suspensions, sublingual tablets, wafers or patches, such as a buccal patch.

[0216] Tablet compositions may contain a unit dose of active compound and an inert diluent or carrier (e.g., a sugar or sugar alcohol (e.g., lactose, sucrose, sorbitol, or mannitol); and / or a non-sugar derived diluent (e.g., sodium carbonate, calcium phosphate, calcium carbonate), or a cellulose or derivative thereof (e.g., microcrystalline cellulose (MCC), methylcellulose, ethylcellulose, hydroxypropylmethylcellulose), and starch (e.g., corn starch)). Tablets may also contain standard ingredients such as binders and granulating agents, such as polyvinylpyrrolidone, disintegrants (e.g., swellable cross-linked polymers such as cross-linked carboxymethylcellulose), lubricants (e.g., stearates), preservatives (e.g., parabens), antioxidants (e.g., BHT), buffers (e.g., phosphate or citrate buffers), and effervescent agents (e.g., citrate / bicarbonate mixtures). Such excipients are well known and need not be discussed in detail herein.

[0217] Tablets may be designed to release the drug upon contact with gastric fluid (immediate release tablets), or in a controlled manner over a longer period of time (controlled release tablets), or upon contact with a specific area of ​​the GI tract.

[0218] The pharmaceutical composition generally comprises about 1% (w / w) to about 95%, preferably % (w / w) of active ingredient and 99% (w / w) to 5% (w / w) of pharmaceutically acceptable excipients (e.g., as defined above) or a combination of such excipients. Preferably, the composition comprises about 20% (w / w) to about 90% (w / w) of active ingredient and 80% (w / w) to 10% of pharmaceutical excipients or a combination of excipients. The pharmaceutical composition comprises about 1% to about 95%, preferably about 20% to about 90% of active ingredient. The pharmaceutical composition according to the present invention can be, for example, in the form of a unit dose, such as an ampoule, a vial, a suppository, a prefilled syringe, a dragee, a powder, a tablet or a capsule.

[0219] Tablets and capsules may contain, for example, 0% to 20% disintegrants, 0% to 5% lubricants, 0% to 5% glidants and / or 0% to 99% (w / w) fillers and / or bulking agents (depending on the drug dose). They may also contain 0% to 10% (w / w) polymer binders, 0% to 5% (w / w) antioxidants, 0% to 5% (w / w) pigments. In addition, sustained-release tablets typically contain 0% to 99% (w / w) controlled-release (e.g., delayed-release) polymers (depending on the dose). The film coating of a tablet or capsule typically contains 0% to 10% (w / w) polymers, 0% to 3% (w / w) pigments and / or 0% to 2% (w / w) plasticizers.

[0220] Parenteral formulations typically contain 0% to 20% (w / w) buffer, 0% to 50% (w / w) cosolvent and / or 0% to 99% (w / w) water for injection (WFI) (depending on the dose and whether lyophilized). Formulations for intramuscular depots may also contain 0% to 99% (w / w) oil.

[0221] Pharmaceutical preparations may be provided to patients in "patient packs" which contain an entire course of treatment in a single package, usually a blister pack.

[0222] The compound of formula (I) is usually present in a unit dosage form, and therefore usually contains enough compound to provide the desired level of biological activity. For example, the formulation may contain 1 nanogram to 2 grams of active ingredient, such as 1 nanogram to 2 milligrams of active ingredient. Within these ranges, a specific subrange of the compound is 0.1 milligram to 2 grams of active ingredient (more generally 10 milligrams to 1 g, such as 50 milligrams to 500 milligrams), or 1 microgram to 20 milligrams (e.g., 1 microgram to 10 milligrams, such as 0.1 milligram to 2 milligrams of active ingredient).

[0223] For oral compositions, unit dosage forms may contain 1 mg to 2 g, more usually 10 mg to 1 g, for example 50 mg to 1 g, for example 100 mg to 1 g, of active compound.

[0224] The active compound is administered to a patient (eg, a human or animal patient) in need thereof in an amount sufficient to achieve the desired therapeutic effect (effective amount). The precise amount of compound administered can be determined by the attending physician according to standard procedures.

[0225] Synthesis of the compound of formula (1)

[0226] LC / MS Method 1

[0227] Instrument: Acquity UPLC with photodiode array detector and QDA mass detector; Column: Acquity C-18, 1.6 micron, 50×2.1 mm; Gradient [time (min) / solvent B in A (%)]: 0.00 / 10, 0.75 / 10, 2.80 / 90, 4.50 / 100, 4.60 / 100, 4.70 / 10; Solvents: Solvent A = 0.1% formic acid in water; Solvent B = 0.1% formic acid in water / acetonitrile (10:90); Column temperature 35° C.; Flow rate 0.8 mL / min.

[0228] LC / MS Method 2

[0229] Instrument: Acquity UPLC with photodiode array detector and QDA mass detector; Column: Acquity C-18, 1.6 micron, 50×1.6 mm; Gradient [time (min) / solvent B in A (%)]: 0.00 / 3, 0.20 / 3, 2.70 / 98, 3.00 / 100, 3.50 / 100, 3.51 / 3, 4.00 / 3; Solvents: Solvent A = 0.1% formic acid in water; Solvent B = 0.1% formic acid in water / acetonitrile (10:90); Column temperature 35° C.; Flow rate 0.9 mL / min.

[0230] LC / MS Method 3

[0231] Instrument: Water 2690 with photodiode array detector and QDA mass detector; Column: X-Bridge C-18, 5 micron, 100×1.6 mm; Gradient [time (min) / solvent B in A (%)]: 0.01 / 10, 1.00 / 10, 5.00 / 100, 7.00 / 100, 7.50 / 10, 8.00 / 10; Solvent: Solvent A = 0.1% formic acid and 10 mM ammonium carbonate in water; Solvent B = acetonitrile; Column temperature 35°C; Flow rate 0.9 mL / min.

[0232] General synthetic strategy

[0233] Compounds of formula G-4 can be synthesized according to Scheme 1 by first 1) alkylating tert-butyl carbazate with a bromide compound of formula G-1a, or 2) contacting tert-butyl carbazate with an aldehyde of formula G-1b under reductive amination conditions to provide hydrazine compound G-2. Contacting G-2 with the divalent compound 3-bromopropionyl chloride provides a cyclized and boc-protected compound of formula G-3. Deprotection of G-4 provides a compound of formula G-4.

[0234] Solution 1

[0235]

[0236] Compounds of formula G-4 can be alkylated according to Scheme 2 via a conjugate addition reaction with compounds of formula G-5 to provide compounds of formula G-6. G-6 can then be coupled with an aryl bromide of formula G-7, wherein R x and R x’ are each independently a lower alkyl group, or R x and R x’The two together with the boron atom and oxygen atom to which they are connected form a 5-yuan or 6-yuan heterocycle. The coupling of G-6 and G-7 is usually carried out in the presence of a palladium catalyst such as PdCl2 (dppf). DCM to provide a biaryl compound of formula G-8. G-8 can be further reduced to provide a G-10 compound. At this point in the synthesis, the mutual conversion of other functional groups is possible, for example, when A is an ester group, G-8 or G-10 compounds can be further hydrolyzed. Similarly, reduction (and / or other FGI, such as hydrolysis) can be carried out by a compound of formula G-9 before the Pd-catalyzed coupling according to Scheme 2.

[0237] Solution 2

[0238]

[0239] The phenolic compound of formula G-11 can be further derivatized to the compound of formula G-12 according to Scheme 3 by reaction with a sulfonamidating agent such as sulfamoyl chloride.

[0240] Solution 3

[0241]

[0242] Example

[0243] Additional embodiments are disclosed in further detail in the following examples, which are not intended to limit the scope of the claims in any way. In the exemplary chiral separations of racemic compounds in the following examples, "Isomer 1" refers to the isomer that is eluted first from the chiral column, and "Isomer 2" refers to the isomer that is eluted second from the chiral column.

[0244] Synthesis of intermediates

[0245] Intermediate 1 :Synthesis of methyl 4-(2-(5-oxopyrazolidin-1-yl)ethyl)benzoate (Intermediate 1)

[0246]

[0247] Step (i): At room temperature, methyl 4-(2-bromoethyl)benzoate (20.00 g, 82.65 mmol), tert-butyl carbazate (12.01 g, 90.92 mmol), NaHCO (27.78 g, 330.60 mmol) and NaI (1.24 g, 8.27 mmol) were suspended in MeCN (200 mL) and the reaction mixture was stirred at 80° C. for 24 hours. The reaction mixture was concentrated under vacuum and the residue was distributed between water (1000 mL) and EtOAc (800 mL), and the aqueous layer was further extracted with EtOAc (3×300 mL). The organic layers were combined and dried (Na SO). The solvent was removed in vacuo and the crude product was purified by gradient column chromatography (normal phase, silica), eluting the product at 0% to 18% EtOAc in hexanes to give an impure product, which was further purified by gradient reverse phase flash column chromatography (reverse phase, C18 silica), eluting the product at 0% to 55% MeCN in water to give tert-butyl 2-(4-(methoxycarbonyl)phenethyl)hydrazine-1-carboxylate (5.6 g, 23.%) as a yellow oil. The product was identified by LCMS (Method 3), m / z 239 (ES+, M+H-tBu) at 2.20 min.

[0248] Step (ii): tert-Butyl 2-(4-(methoxycarbonyl)phenethyl)hydrazine-1-carboxylate (5.60 g, 19.04 mmol) was dissolved in MeCN (60 mL), and potassium carbonate (13.16 g, 95.19 mmol) was added to the reaction mixture at room temperature, and the reaction mixture was stirred at room temperature for 10 minutes. After this, 3-bromopropionyl chloride (2.90 mL, 28.56 mmol) was added dropwise at room temperature, and the reaction mixture was stirred at room temperature for 24 hours. The reaction mixture was concentrated under vacuum, and the obtained residue was distributed between water (800 mL) and EtOAc (500 mL), and the aqueous layer was further extracted with EtOAc (2×200 mL). The organic layers were combined and dried (Na2SO4). The solvent was removed in vacuo and the product was purified by gradient column chromatography (normal phase, silica) eluting the product at 0% to 22% EtOAc in hexanes to give an impure product which was further purified by gradient reverse phase flash column chromatography (reverse phase, C18 silica) eluting the product at 0% to 45% MeCN in water to give tert-butyl 2-(4-(methoxycarbonyl)phenethyl)-3-oxopyrazolidine-1-carboxylate (6.5 g, 98%) as a yellow oil. The product was identified by LCMS (Method_3), m / z 293.(ES+, M+HtBu) at 2.43 min.

[0249] Step (iii): tert-Butyl 2-(4-(methoxycarbonyl)phenethyl)-3-oxopyrazolidine-1-carboxylate (6.5 g, 18.7 mmol) was dissolved in dioxane (70 mL) and 4N HCl in dioxane (70 mL) was added dropwise to the reaction mixture at 0°C and the reaction mixture was stirred at room temperature for 4 hours. The solvent was removed in vacuo to obtain a crude product which was purified by trituration with 20% MeOH in ether to give methyl 4-(2-(5-oxopyrazolidin-1-yl)ethyl)benzoate (4.8 g, 90.%) as an off-white powder. The product was determined by LCMS (Method_3), m / z 249 (ES+, M+H) at 1.57 minutes.

[0250] 1 H NMR: (400MHz, DMSO) δ: 2.62-2.58 (t, 2H, J=8.2Hz), 3.06-3.02 (t, 2H, J=7.4Hz), 3.58-3.56 (t, 2H , J=4.4Hz), 3.74-3.70 (t, 2H, J=7.2Hz), 7.46-7.44 (d, 2H, J=8.4Hz), 7.90-7.89 (d, 2H, J=8.0Hz).

[0251] Intermediate 2 :Synthesis of methyl 4-(2-(2-(3-(3-bromophenyl)-3-hydroxypropyl)-5-oxopyrazolidin-1-yl)ethyl)-2,6-difluorobenzoate (Intermediate 2)

[0252]

[0253] Step (i): 4-bromo-2,6-difluorobenzoic acid methyl ester (40.00 g, 159.36 mmol), potassium vinyl trifluoroborate (32.00 g, 239.04 mmol) and TEA (42 mL, 318.72 mmol) were dissolved in IPA (400 mL) and purged with nitrogen at room temperature for 30 minutes. After this, PdCl2(dppf).DCM (13.01 g, 15.93 mmol) was added and the reaction mixture was stirred at 80°C for 3 hours. The reaction mixture was then distributed between water (1000 mL) and EtOAc (1000 mL). The aqueous layer was further extracted with EtOAc (2×500 mL). The organic layers were combined and dried (Na2SO4). The solvent was removed in vacuo and the crude product was purified by gradient flash column chromatography (normal phase, silica) eluting the product at 0% to 5% EtOAc in hexanes to give methyl 2,6-difluoro-4-vinylbenzoate (25.0 g, 79%) as a yellow oil.

[0254] Step (ii) at room temperature, 2,6-difluoro-4-vinyl benzoic acid methyl ester (25.0g, 126.19mmol) is dissolved in the mixture of tert-butyl alcohol (130mL) and water (175mL). After this, NBS (26.94g, 151.43mmol) is added in batches at room temperature and stirred at 40 ℃ for 16 hours. After being cooled to 5 ℃, NaOH solution (10.09g, 252.23mmol) is added and stirred at room temperature for 30 minutes. Then the reaction mixture is distributed between water (800mL) and EtOAc (500mL), and the water layer is further extracted with EtOAc (2×300mL). The organic layers are merged and dried (Na2SO4). Remove solvent in a vacuum to obtain thick 2,6-difluoro-4-(oxirane-2-yl) benzoic acid methyl ester (22.8g, 84%) as a yellow oil.

[0255] Step (iii): 2,6-difluoro-4-(oxirane-2-yl) methyl benzoate (22.8g, 106.54mmol) is dissolved in methanol (500mL), and 10% palladium carbon (6.9g) containing 50% moisture is added. After this, ammonium formate (67.22g, 1065.42mmol) is added at room temperature, and the reaction mixture is stirred at room temperature for 16 hours. After completion, the reaction mixture is filtered through a diatomaceous earth bed, washed with MeOH (3000mL), and the filtrate is concentrated in vacuo. The reaction mixture is then distributed between water (1000mL) and EtOAc (500mL). The water layer is further extracted with EtOAc (2×300mL). The organic layers are combined and dried (Na2SO4). The solvent was removed in vacuo to give the crude product, which was purified by gradient flash column chromatography (normal phase, silica) eluting the product at 0% to 26% EtOAc in hexanes to give methyl 2,6-difluoro-4-(2-hydroxyethyl)benzoate (11 g, 47.80%) as a yellow oil.

[0256] Step (iv): at room temperature, 2,6-difluoro-4-(2-hydroxyethyl) methyl benzoate (11.00g, 50.87mmol) was dissolved in DCM (300mL). After that, Dess-Martin periodinane (32.36g, 76.31mmol) was added in batches at room temperature, and the reaction mixture was stirred at room temperature for 2 hours, and then the reaction mixture was distributed between saturated NaHCO3 aqueous solution (500mL) and DCM (800mL). The water layer was further extracted with EtOAc (2×500mL). The organic layers were combined and dried (Na2SO4). The solvent was removed in vacuo to obtain the crude 2,6-difluoro-4-(2-oxyethyl) methyl benzoate (7.96g, 73%) as a white solid. It can be used in the next step without further purification.

[0257] Step (v): Under nitrogen atmosphere, methyl 2,6-difluoro-4-(2-oxoethyl)benzoate (7.92 g, 37.00 mmol) and tert-butyl carbazate (4.89 g, 37.00 mmol) were dissolved in methanol (100 mL), and further added Molecular sieves are to maintain anhydrous conditions.After this, glacial acetic acid (0.3mL, 3.70mmol) is added at room temperature, and stirred at room temperature for 3 hours.After this, the reaction mixture is cooled at 0 ° C and sodium cyanoborohydride (2.78g, 44.40mmol) is added in batches, and stirred at room temperature for 12 hours.The reaction mixture is concentrated under vacuum to obtain a residue.The residue is distributed between saturated NaHCO3 aqueous solution (500mL) and EtOAc (300mL), and the water layer is further extracted with EtOAc (2 × 200mL).The organic layers are merged and dried (Na2SO4).The solvent is removed in a vacuum to obtain a crude product, which is purified by gradient column chromatography (normal phase, silica), and the product is eluted under 0% to 3% EtOAc in DCM to obtain a crude 2- (3,5-difluoro-4- (methoxycarbonyl) phenethyl) hydrazine-1-carboxylic acid tert-butyl ester (9.10g, 65%) as a white solid. The product was identified by LCMS (Method 2), m / z 275 (ES+, M+H-tBu) at 2.32 min.

[0258] Step (vi): tert-Butyl 2-(3,5-difluoro-4-(methoxycarbonyl)phenethyl)hydrazine-1-carboxylate (9.00 g, 27.26 mmol) was dissolved in DMF (20 mL), and potassium carbonate (18.83 g, 136.30 mmol) was added to the reaction mixture at room temperature and stirred at room temperature for 10 minutes. After this, 4-bromobutyryl chloride (3.4 mL, 40.89 mmol) was added dropwise at room temperature, and the reaction mixture was stirred at room temperature for 3 hours and then at 40° C. for 16 hours. The reaction mixture was partitioned between water (500 mL) and EtOAc (700 mL), and the aqueous layer was further extracted with EtOAc (2×250 mL). The organic layers were combined and dried (Na 2 SO 4 ). The solvent was removed in vacuo and the product was purified by gradient column chromatography (reverse phase, C18 silica) eluting the product at 0% to 50% MeCN in water to give (3.5 g, 33%) as a yellow oil. The product was identified by LCMS (Method 2), m / z 329 (ES+, M+H-tBu) at 2.48 minutes.

[0259] Step (vii): tert-Butyl 2-(3,5-difluoro-4-(methoxycarbonyl)phenethyl)-3-oxopyrazolidine-1-carboxylate (3.5 g, 9.11 mmol) was dissolved in dioxane (40 mL) and 4N HCl in dioxane (20 mL) was added dropwise to the reaction mixture at 0°C. The reaction mixture was stirred at room temperature for 18 hours. The solvent was removed in vacuo to obtain a crude product which was purified by trituration with 20% MeOH in ether to give methyl 2,6-difluoro-4-(2-(5-oxopyrazolidine-1-yl)ethyl)benzoate-HCl-salt (2.62 g, 79%) as an off-white amorphous powder. The product was determined by LCMS (Method 2), m / z 285 (ES+, M+H) at 1.67 minutes.

[0260] Step (vii): tert-Butyl 2-(3,5-difluoro-4-(methoxycarbonyl)phenethyl)-3-oxopyrazolidine-1-carboxylate (3.5 g, 9.11 mmol) was dissolved in dioxane (40 mL) and 4N HCl in dioxane (20 mL) was added dropwise to the reaction mixture at 0°C. The reaction mixture was stirred at room temperature for 18 hours. The solvent was removed in vacuo to obtain a crude product which was purified by trituration with 20% MeOH in ether to give methyl 2,6-difluoro-4-(2-(5-oxopyrazolidine-1-yl)ethyl)benzoate-HCl-salt) (2.62 g, 79%) as an off-white amorphous powder. The product was determined by LCMS (Method 2), m / z 285 (ES+, M+H) at 1.67 minutes.

[0261] Step (viii): At room temperature, methyl 2,6-difluoro-4-(2-(5-oxopyrazolidin-1-yl)ethyl)benzoate-HCl-salt) (0.80 g, 2.81 mmol) and 1-(3-bromophenyl)prop-2-en-1-one (1.60 g, 14.08 mmol) were suspended in MeOH (10 mL), and the reaction mixture was stirred at room temperature for 15 minutes. After this, TEA (0.70 mL, 5.27 mmol) was added at room temperature and stirred at 60° C. for 4 hours. The reaction mixture was partitioned between water (200 mL) and EtOAc (200 mL), and the aqueous layer was further extracted with EtOAc (2×100 mL). The organic layers were combined and dried (Na2SO4). The solvent was removed in vacuo and the crude product was further purified by gradient reverse phase flash column chromatography (reverse phase, C18 silica) eluting the product at 0% to 58% ACN in water to give methyl 4-(2-(2-(3-(3-bromophenyl)-3-oxopropyl)-5-oxopyrazolidin-1-yl)ethyl)-2,6-difluorobenzoate (0.65 g, 47%) as a colorless viscous material. The product was identified by LCMS (Method 2), m / z 495 (ES+, M+H) at 2.58 minutes.

[0262] Step (ix): Methyl 4-(2-(2-(3-(3-bromophenyl)-3-oxopropyl)-5-oxopyrazolidin-1-yl)ethyl)-2,6-difluorobenzoate (0.650 g, 1.31 mmol) was dissolved in ethanol (4 mL) and water (2 mL), and CeCl (0.97 g, 3.94 mmol) was added and the reaction mixture was stirred at 0°C for 5 minutes. Sodium borohydride (0.20 g, 0.5.26 mmol) was then added at 0°C and the reaction mixture was stirred at room temperature for 1 hour. The reaction mixture was then partitioned between water (100 mL) and EtOAc (150 mL), and the aqueous layer was further extracted with EtOAc (2×50 mL). The organic layers were combined and dried (Na2SO4). The solvent was removed in vacuo to give Intermediate 2: methyl 4-(2-(2-(3-(3-bromophenyl)-3-hydroxypropyl)-5-oxopyrazolidin-1-yl)ethyl)-2,6-difluorobenzoate (0.620 g, 95.%) as a brown solid. The product was identified by LCMS (Method 2), m / z 497 (ES+, M+H) at 2.04 min.

[0263] 1H NMR: (400Mz, DMSO) δ: 1.74-1.73 (d, 2H, J=4.8Hz), 2.88-2.76 (m, 4H), 3.12-3.10 (d, 2H, J=7.2Hz), 3.86 (s, 3H), 4.66 (s, 1 H), 5.44-5.43 (d, 1H, J=4.4Hz), 7.12-7.10 (d, 2H, J=9.6Hz), 7.36-7.27 (m, 2H), 7.43-7.41 (d, 1H, J=8.0Hz), 7.55 (s.1H).

[0264] Intermediate B :Synthesis of 1-(3-bromophenyl)prop-2-en-1-one (Intermediate B)

[0265]

[0266] Step (i): 3-Bromobenzaldehyde (12.0 g, 56.6 mmol) was dissolved in ether (30 mL) at 0°C, and vinylmagnesium bromide (1 M in THF) (200 mL) was added dropwise at 0°C under a nitrogen atmosphere, and stirred at room temperature for 3 hours. The reaction mixture was partitioned between saturated aqueous NH4Cl solution (1000 mL) and EtOAc (800 mL); the aqueous layer was further extracted with EtOAc (2×300 mL). The organic layers were combined and dried (Na2SO4). The solvent was removed in vacuo, and the crude product was purified by gradient column chromatography (normal phase, silica), eluting the product at 0% to 14% EtOAc in hexane to give 1-(3-bromophenyl)prop-2-en-1-ol (A) (12.0 g, 94%) as a colorless oil.

[0267] Step (ii): [1-(3-bromophenyl)prop-2-en-1-ol A] (12.0 g, 57.1 mmol) was dissolved in acetone (30 mL) at room temperature. Jones reagent (39 mL) was added dropwise thereto at -25°C and stirred at -25°C for 30 minutes. The reaction mixture was then warmed to 0°C and stirred for 30 minutes. The reaction mixture was partitioned between saturated aqueous NaHCO3 solution (100 mL) and EtOAc (800 mL), and the aqueous layer was further extracted with EtOAc (2×250 mL). The organic layers were combined and dried (Na2SO4). The solvent was removed in vacuo, and the crude product was purified by gradient column chromatography (normal phase, silica), eluting the product at 0% to 8% EtOAc in hexane to give 1-(3-bromophenyl)prop-2-en-1-one (8.0 g, 97%) as a yellow gum. The product was confirmed by LCMS, massless ion, 2.39 min.

[0268] Intermediate 3 :Synthesis of methyl 4-(2-(2-(3-(3-bromophenyl)-3-hydroxypropyl)-5-oxopyrazolidin-1-yl)ethyl)-2-hydroxybenzoate (Intermediate 3)

[0269]

[0270] Step (i): 4-bromo-2-methoxybenzoic acid methyl ester (30.00g, 156.07mmol), potassium vinyl trifluoroborate (41.81g, 312.16mmol) and TEA (64.93mL, 468.21mmol) were dissolved in isopropanol (300mL) and purged with nitrogen at room temperature for 30 minutes. After this, PdCl2(dppf)DCM (20.09g, 24.60mmol) was added, and the reaction mixture was stirred at 80°C for 3 hours. The reaction mixture was then distributed between water (1200mL) and EtOAc (700mL). The aqueous layer was further extracted with EtOAc (2×500mL). The organic layers were combined and dried (Na2SO4). The solvent was removed in vacuo and the crude product was purified by gradient flash column chromatography (normal phase, silica) eluting the product at 0% to 14% EtOAc in hexanes to afford methyl 2-methoxy-4-vinylbenzoate (20.00 g, 84%) as a yellow oil. The product was determined by LCMS (Method 1), m / z 193 (ES+, M+H) at 2.18 minutes.

[0271] Step (ii): At room temperature, methyl 2-methoxy-4-vinylbenzoate (20.00 g, 104.05 mmol) was dissolved in a mixture of tert-butyl alcohol (15 mL) and water (30 mL). After this, NBS (27.78 g, 156.08 mmol) was added in batches at room temperature, and the reaction mixture was stirred at 40 ° C for 2 hours. The reaction mixture was then distributed between water (1000 mL) and EtOAc (800 mL). The aqueous layer was further extracted with EtOAc (2×300 mL). The organic layers were combined and dried (Na2SO4). The solvent was removed in vacuo to obtain crude methyl 2-methoxy-4-(oxirane-2-yl)benzoate (20.00 g, quantitative yield) as a yellow oil.

[0272] Note: Since this product is unstable, proceed to the next step without purification.

[0273] Step (iii): 2-methoxy-4-(oxirane-2-yl) methyl benzoate (20.00g, 96.06mmol) is dissolved in methanol (200mL), and 10% palladium carbon (10.00g) containing 50% moisture is added. After this, ammonium formate (60.57g, 960.60mmol) is added at room temperature, and the reaction mixture is stirred at 60 ℃ for 4 hours. After completion, the reaction mixture is filtered through a diatomaceous earth bed and washed with MeOH (2000mL). The filtrate is concentrated in a vacuum. The crude product is purified by gradient flash column chromatography (normal phase, silica), and the product is eluted under 0% to 45% EtOAc in hexane to obtain a thick 4-(2-hydroxyethyl)-2-methoxy methyl benzoate (9.80g, 49%) as a colorless oil. The product was identified by LCMS (Method 1), m / z 211 (ES+) at 1.60 min.

[0274] Step (iv): at room temperature, 4-(2-hydroxyethyl)-2-methoxybenzoic acid methyl ester (9.80g, 46.62mmol) was dissolved in DCM (100mL). After that, Dess-Martin periodinane (39.54g, 93.24mmol) was added in batches at room temperature, and the reaction mixture was stirred at room temperature for 4 hours. The reaction mixture was then distributed between saturated NaHCO3 aqueous solution (800mL) and EtOAc (500mL). The water layer was further extracted with EtOAc (2×200mL). The organic layers were combined and dried (Na2SO4). The solvent was removed in vacuo to obtain 2-methoxy-4-(2-oxyethyl)benzoic acid methyl ester (10.0g, quantitative yield) as a yellow oil, which was used for the next step without further purification.

[0275] Step (v): Under nitrogen atmosphere, methyl 2-methoxy-4-(2-oxoethyl)benzoate (10.0 g, 48.03 mmol) and tert-butyl carbazate (7.62 g, 57.63 mmol) were dissolved in methanol (100 mL). Molecular sieves (1.00g) are to maintain moisture-free conditions. After that, glacial acetic acid (0.3mL, 4.80mmol) is added at room temperature, and the reaction mixture is stirred at room temperature for 3 hours. After that, the reaction mixture is cooled at 0 ° C, and sodium cyanoborohydride (4.53g, 72.05mmol) is added in batches, and the reaction mixture is stirred at room temperature for 16 hours. The reaction mixture is filtered under reduced pressure, and the filtrate is concentrated in a vacuum. The residue obtained is distributed between saturated NaHCO3 aqueous solution (700mL) and EtOAc (500mL). The water layer is further extracted with EtOAc (3 × 200mL). The organic layers are merged and dried (Na2SO4). The solvent was removed in vacuo to give a crude product which was purified by gradient column chromatography (normal phase, silica) eluting the product at 0% to 29% EtOAc in hexanes to give crude tert-butyl 2-(3-methoxy-4-(methoxycarbonyl)phenethyl)hydrazine-1-carboxylate (7.0 g, 45%) as a yellow viscous material. The product was determined by LCMS (Method 2), m / z 347 (ES+, M+Na), at 2.02.

[0276] Step (vi): tert-Butyl 2-(4-(methoxycarbonyl)phenethyl)hydrazine-1-carboxylate (7.00 g, 21.59 mmol) was dissolved in MeCN (70 mL), and potassium carbonate (14.92 g, 107.93 mmol) was added to the reaction mixture at room temperature. The reaction mixture was stirred at room temperature for 10 minutes. After that, 3-bromopropionyl chloride (3.26 mL, 32.39 mmol) was added dropwise at room temperature, and the reaction mixture was stirred at room temperature for 24 hours. After that, the reaction mixture was stirred at 40 ° C for 4 hours. The reaction mixture was concentrated under vacuum, and the obtained residue was distributed between water (700 mL) and EtOAc (500 mL). The aqueous layer was further extracted with EtOAc (2×200 mL). The organic layers were combined and dried (Na2SO4). The solvent was removed in vacuo and the product was purified by gradient column chromatography (normal phase, silica) eluting the product at 0% to 32% EtOAc in hexanes to give tert-butyl 2-(3-methoxy-4-(methoxycarbonyl)phenethyl)-3-oxopyrazolidine-1-carboxylate (2.0 g, 24%) as a yellow oil. The product was identified by LCMS (Method 2), m / z 323 (ES+, M+H-tBu) at 2.27 minutes.

[0277] Step (vii): tert-Butyl 2-(3-methoxy-4-(methoxycarbonyl)phenethyl)-3-oxopyrazolidine-1-carboxylate (2.00 g, 5.29 mmol) was dissolved in 1,4-dioxane (20 mL) and 4N HCl in dioxane (20 mL) was added dropwise to the reaction mixture at 0°C and the reaction mixture was stirred at room temperature for 8 hours. The solvent was removed in vacuo to obtain the crude product, which was purified by trituration with 20% MeOH in ether to give crude methyl 2-methoxy-4-(2-(5-oxopyrazolidin-1-yl)ethyl)benzoate HCl salt (1.50 g, 90.36%) as an off-white amorphous powder. The product was identified by LCMS, m / z 279.10 (ES+) at 1.415 minutes.

[0278] Step (viii): Methyl 2-methoxy-4-(2-(5-oxopyrazolidin-1-yl)ethyl)benzoate hydrochloride (1.00 g, 3.18 mmol), 1-(3-bromophenyl)prop-2-en-1-one (3.33 g, 15.88 mmol) and TEA (2.2 mL, 15.88 mmol) were suspended in MeOH (10 mL) at room temperature and the reaction mixture was stirred at 60° C. for 4 hours. The reaction mixture was partitioned between saturated aqueous NH4Cl solution (300 mL) and EtOAc (200 mL). The aqueous layer was further extracted with EtOAc (2×100 mL). The organic layers were combined and dried (Na2SO4). The solvent was removed in vacuo and the crude product was further purified by reverse phase gradient flash column chromatography (reverse phase, C18 silica) eluting the product at 0% to 65% MeCN in water to give methyl 4-(2-(2-(3-(3-bromophenyl)-3-oxopropyl)-5-oxopyrazolidin-1-yl)ethyl)-2-methoxybenzoate (1.20 g, 69%) as a colorless viscous material. The product was identified by LCMS (Method 2), m / z 489 (ES+, M+H) at 2.38 minutes.

[0279] Step (ix): (R)-methyl 4-((1-(3-bromobenzyl)pyrrolidine-2-carboxamido)methyl)-2-methoxybenzoate (0.25 g, 0.51 mmol) was dissolved in DCM (3 mL) and the reaction mixture was cooled to -78 °C. After this, BBr3 (1 M solution in DCM) (2.5 mL, 2.56 mmol) was added and the reaction mixture was stirred at -78 °C for 1 hour. The reaction mixture was then partitioned between saturated NaHCO3 aqueous solution (100 mL) and DCM (70 mL). The aqueous layer was extracted again with EtOAc (2 x 40 mL). The organic layers were combined and dried (Na2SO4). The solvent was removed in vacuo to give methyl 4-(2-(2-(3-(3-bromophenyl)-3-oxopropyl)-5-oxopyrazolidin-1-yl)ethyl)-2-hydroxybenzoate (0.18 g, 74%) as a yellow sticky substance. The product was identified by LCMS (Method 2), m / z 475 (ES+, M+H) at 2.61 min.

[0280] Step (x): Methyl 4-(2-(2-(3-(3-bromophenyl)-3-oxopropyl)-5-oxopyrazolidin-1-yl)ethyl)-2-hydroxybenzoate (0.16 g, 0.34 mmol) was dissolved in ethanol (2 mL) and water (2 mL). CeCl (0.25 g, 1.01 mmol) was added thereto, and the reaction mixture was stirred at 0° C. for 5 minutes. Thereafter, NaBH (0.085 g, 1.36 mmol) was added at 0° C., and the reaction mixture was stirred at room temperature for 1 hour. The reaction mixture was partitioned between saturated NaHCO aqueous solution (30 mL) and EtOAc (30 mL). The aqueous layer was further extracted with EtOAc (2×20 mL). The organic layers were combined and dried (Na2SO4). The solvent was removed in vacuo to give Intermediate 3 methyl 4-(2-(2-(3-(3-bromophenyl)-3-hydroxypropyl)-5-oxopyrazolidin-1-yl)ethyl)-2-hydroxybenzoate (0.15 g, 93%) as an off-white solid. The product was identified by LCMS (Method 2), m / z 478 (ES+, M+H) at 2.46 min.

[0281] Synthesis of compounds of formula I

[0282] Embodiment 1: Synthesis of 4-(2-(2-(3-hydroxy-3-(4'-hydroxy-2'-methyl-[1,1'-biphenyl]-3-yl)propyl)-5-oxopyrazolidin-1-yl)ethyl)benzoic acid (Compound 1)

[0283]

[0284] Step (i): Intermediate 1 4-(2-(5-oxopyrazolidin-1-yl)ethyl)benzoic acid methyl ester hydrochloride (2.00 g, 7.26 mmol) and 1-(3-bromophenyl)prop-2-en-1-one (1.50 g, 14.52 mmol) were suspended in IPA (20 mL) at room temperature, and the reaction mixture was stirred at room temperature for 15 minutes. After this, TEA (2.5 mL, 36.30 mmol) was added at room temperature and stirred at 80° C. for 12 hours. The reaction mixture was partitioned between saturated NH4Cl aqueous solution (500 mL) and EtOAc (300 mL), and the aqueous layer was further extracted with EtOAc (2×100 mL). The organic layers were combined and dried (Na2SO4). The solvent was removed in vacuo and the crude product was further purified by gradient reverse phase flash column chromatography (reverse phase, C18 silica) eluting the product at 0% to 75% MeCN in water to give methyl 4-(2-(2-(3-(3-bromophenyl)-3-oxopropyl)-5-oxopyrazolidin-1-yl)benzoate (0.80 g, 24%) as a colorless sticky material. The product was identified by LCMS (Method 3), m / z 459 (ES+, M+H) at 2.48 min.

[0285] Step (ii): Methyl 4-(2-(2-(3-(3-bromophenyl)-3-oxopropyl)-5-oxopyrazolidin-1-yl)ethyl)benzoate (0.25 g, 0.55 mmol), 4-hydroxy-2-methylphenyl)boronic acid (0.99 g, 0.65 mmol) and potassium carbonate (0.23 g, 1.65 mmol) were suspended in dioxane (3 mL) and water (2 mL) at room temperature, and the reaction mixture was degassed with nitrogen for 15 minutes at room temperature. After this, PdCl2(dppf).DCM (0.04 g, 0.05 mmol) was added at room temperature, and the reaction mixture was stirred at 100°C for 4 hours. The reaction mixture was partitioned between water (200 mL) and EtOAc (100 mL), and the aqueous layer was further extracted with EtOAc (2×50 mL). The organic layers were combined and dried (Na2SO4). The solvent was removed in vacuo and the crude product was purified by gradient column chromatography (normal phase, silica) eluting the product at 0% to 80% EtOAc in hexanes to afford methyl 4-(2-(2-(3-(4'-hydroxy-2'-methyl-[1,1'-biphenyl]-3-yl)-3-oxopropyl)-5-oxopyrazolidin-1-yl)ethyl)benzoate intermediate 4 (0.21 g, 81%) as an off-white solid. The product was identified by LCMS (Method 3), m / z 509 (ES+, M+Na) at 2.33 min.

[0286] Step (iii): At room temperature, 4-(2-(2-(3-(3-bromophenyl)-3-oxopropyl)-5-oxopyrazolidine-1-yl)ethyl)benzoic acid methyl ester (0.21 g, 0.43 mmol) and cerium chloride (iii) (0.32 g, 1.30 mmol) were dissolved in ethanol (3 mL) and water (3 mL), and the reaction mixture was stirred at room temperature for 5 minutes. After this, sodium borohydride (0.65 g, 1.73 mmol) was added at 0°C, and the reaction mixture was stirred at room temperature for 2 hours. The reaction mixture was distributed between saturated NaHCO3 aqueous solution (100 mL) and EtOAc (50 mL), and the aqueous layer was further extracted with EtOAc (2×30 mL). The organic layers were combined and dried (Na2SO4). The solvent was removed in vacuo to give methyl 4-(2-(2-(3-hydroxy-3-(4'-hydroxy-2'-methyl-[1,1'-biphenyl]-3-yl)propyl)-5-oxopyrazolidin-1-yl)ethyl)benzoate (0.16 g, 76%) as an off-white solid. The product was identified by LCMS (Method 3), m / z 489 (ES+, M+H) at 2.19 min.

[0287] Step (iv): Methyl 4-(2-(2-(3-hydroxy-3-(4'-hydroxy-2'-methyl-[1,1'-biphenyl]-3-yl)propyl)-5-oxopyrazolidin-1-yl)ethyl)benzoate (0.16 g, 0.33 mmol) was dissolved in dioxane (2 mL) and water (2 mL). LiOH monohydrate (0.073 g, 1.75 mmol) was added thereto at room temperature and stirred for 3 hours at room temperature. The reaction mixture was then partitioned between water (80 mL) and EtOAc (2×50 mL). The aqueous layer was further acidified with 1N aqueous HCl (about 20 mL), the pH was adjusted to about 3 and extracted with EtOAc (2×70 mL). The organic layers were combined and dried (Na2SO4). The solvent was removed in vacuo to give 4-(2-(2-(3-hydroxy-3-(4'-hydroxy-2'-methyl-[1,1'-biphenyl]-3-yl)propyl)-5-oxopyrazolidin-1-yl)ethyl)benzoic acid) (0.125 g, 81%) as a brown solid. The product was identified by LCMS (Method 3), m / z 475 (ES+, M+H) at 1.97 min.

[0288] 1H NMR: (400MHz, DMSO) δ: 1.26-1.23 (m, 3H), 1.82-1.80 (d, 2H, J=6.4Hz), 2.16 (s, 3H), 2.88-2.85 (t, 3H, J=6.0Hz), 3.17 (s, 3H), 4.08 (s, 1H), 4.71-4.69 ( d, 1H, J=6.0Hz), 6.87-6.83 (m, 2H), 7.01-6.99 (d, 1H, J=8.4Hz), 7.16-7.14 (d, 1H, J=7.6Hz), 7.37-7.26 (m, 5H), 7.85-7.83 (d, 2H, 8.4Hz), 9.43 (s, 2H).

[0289] Compounds 1R and 1S: Racemic compound 1 (0.12 g) was enantiomerically separated using a Shimadzu Prep-HPLC system (PHENOMNENEX AMYLOSE-250*21.2 mm, 5 μm column) using a mobile phase of 43.0% 10 mM DEA and 57.0% IPA in heptane with no gradient and a flow rate of 16.00 mL / min. Isomer 1 : (0.004 g, 2.6%; off-white solid) The product was identified by LCMS (Method 3), m / z 475 (ES+, M+H) at 2.30 min.

[0290] 1 H NMR: (400MHz, MeOD) δ: 2.03-1.93 (m, 2H), 2.93-2.90 (t, 4H, J=6.8Hz), 3.07-3.03 (t, 3H, J=7.2Hz), 3.25(s, 2H), 3.37-3.32(m, 1H), 3.59-3.56(m, 1H), 3.71-3.69(m, 2H), 4.87-4.84 (t, 1H, J=6.0Hz), 6.68-6.66 (m, 1H), 6.73-6.72 (d, 1H, J=2.0z), 7.02-7.0 0 (d, 1H, J=8.0Hz), 7.25-7.19 (m, 3H), 7.41-7.32 (m, 5H), 7.91-7.89 (d, 2H, J=8.0Hz).

[0291] Isomer 2 : The product was identified by LCMS (Method 3), m / z 475 (ES+, M+H) at 2.30 min.

[0292] 1H NMR: (400MHz, MeOD) δ: 1.97-1.94 (t, 2H, J=6.4Hz), 2.93-2.90 (t, 4H, J=6.8Hz), 3.07-3.03 (m, 3H), 3.26 (s, 2H), 3.59-3.57 (t, 2H) J=4.6Hz), 3.71-3.69 (t, 2H.J=4.6Hz), 4.87-4.84 (t, 1H, J=6.4Hz), 6.68-6.65 (dd, 1H.J=2.4Hz&J=4.0Hz), 6.72-6. 72 (d, 1H, J=2.0Hz), 7.02-7.00 (d, 1H, J=8.0Hz), 7.24-7.19 (m, 3H), 7.41-7.32 (m, 5H), 7.91-7.89 (d, 2H, J=8.0Hz).

[0293] Implementation Example 2: Synthesis of 4-(2-(2-(3-(4'-carbamoyl-2'-methyl-[1,1'-biphenyl]-3-yl)-3-hydroxypropyl)-5-oxopyrazolidin-1-yl)ethyl)benzoic acid (Compound 2)

[0294]

[0295] Step (i): At room temperature, the intermediate 1,4-(2-(5-oxopyrazolidine-1-yl)ethyl)benzoic acid methyl ester hydrochloride (2.00g, 7.26mmol) and 1-(3-bromophenyl)prop-2-en-1-one (1.50g, 14.52mmol) were suspended in IPA (20mL), and the reaction mixture was stirred at room temperature for 15 minutes. After this, TEA (2.5mL, 36.30mmol) was added at room temperature and stirred at 80°C for 12 hours. The reaction mixture was distributed between saturated NH4Cl aqueous solution (500mL) and EtOAc (300mL), and the aqueous layer was further extracted with EtOAc (2×100mL). The organic layers were combined and dried (Na2SO4). The solvent was removed in vacuo and the crude product was further purified by gradient reverse phase flash column chromatography (reverse phase, C18 silica) eluting the product at 0% to 75% MeCN in water to give methyl 4-(2-(2-(3-(3-bromophenyl)-3-oxopropyl)-5-oxopyrazolidin-1-yl)benzoate (0.80 g, 24%) as a colorless sticky material. The product was identified by LCMS (Method 3), m / z 459 (ES+, M+H) at 2.48 min.

[0296] Step (ii): Methyl 4-(2-(2-(3-(3-bromophenyl)-3-oxopropyl)-5-oxopyrazolidin-1-yl)ethyl)benzoate (0.60 g, 1.30 mmol), 3-methyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzamide (1.36 g, 5.23 mmol) and KCO (0.54 g, 3.92 mmol) were dissolved in a mixture of dioxane:water (3:2) (5 mL) and purged with nitrogen at room temperature for 20 min. After this, PdCl(dppf)DCM (0.21 g, 0.26 mmol) was added and the reaction mixture was stirred at 80°C for 8 h. The reaction mixture was then distributed between water (100 mL) and EtOAc (100 mL), and the aqueous layer was further extracted with EtOAc (2 × 70 mL). The organic layers were combined and dried (Na2SO4). The solvent was removed in vacuo, and the crude product was purified by gradient flash column chromatography (reverse phase, C18 silica), eluting the product under 0% to 38% MeCN in water to obtain 4-(2-(2-(3-(4'-carbamoyl-2'-methyl-[1,1'-biphenyl]-3-yl)-3-oxopropyl)-5-oxopyrazolidine-1-yl)ethyl)benzoic acid methyl ester (0.60 g, 90%) as a white solid. LCMS: (Method 3): The product was determined, m / z 536 (ES+, M+Na), at 2.16 minutes.

[0297] Step (iii): Methyl 4-(2-(2-(3-(4'-carbamoyl-2'-methyl-[1,1'-biphenyl]-3-yl)-3-oxopropyl)-5-oxopyrazolidin-1-yl)ethyl)benzoate (0.60 g, 1.16 mmol) was dissolved in ethanol (3 mL) and water (2 mL), and CeCl3 (0.86 g, 3.50 mmol) was added to the reaction mixture. The reaction mixture was stirred at 0°C for 5 minutes. After this, sodium borohydride (0.17 g, 4.67 mmol) was added at 0°C, and the reaction mixture was stirred at room temperature for 2 hours. The reaction mixture was distributed between saturated NaHCO3 aqueous solution (100 mL) and EtOAc (80 mL), and the aqueous layer was further extracted with EtOAc (2×50 mL). The organic layers were combined and dried (Na2SO4). The solvent was removed in vacuo and the crude product was purified by gradient flash column chromatography (reverse phase, C18 silica) eluting the product at 0% to 35% MeCN in water to give pure methyl 4-(2-(2-(3-(4'-carbamoyl-2'-methyl-[1,1'-biphenyl]-3-yl)-3-hydroxypropyl)-5-oxopyrazolidin-1-yl)ethyl)benzoate (0.50 g, 83%) as an off-white solid. LCMS: (LCMS Method 3): The product was identified, m / z 516 (ES+, M+H) at 2.78 min.

[0298] Step (iv): Methyl 4-(2-(2-(3-(4'-carbamoyl-2'-methyl-[1,1'-biphenyl]-3-yl)-3-hydroxypropyl)-5-oxopyrazolidin-1-yl)ethyl)benzoate (1.80 g, 3.49 mmol) was dissolved in dioxane (15 mL) and water (8 mL). LiOH monohydrate (0.73 g, 17.46 mmol) was added at room temperature and the reaction mixture was stirred at room temperature for 3 hours. The reaction mixture was then partitioned between water (250 mL) and EtOAc (2×350 mL). The aqueous layer was further acidified with 4N aqueous HCl (60 mL) to adjust the pH to about 1 and extracted with EtOAc (2×300 mL). The organic layers were combined and dried (Na2SO4). The solvent was removed in vacuo to give pure 4-(2-(2-(3-(4'-carbamoyl-2'-methyl-[1,1'-biphenyl]-3-yl)-3-hydroxypropyl)-5-oxopyrazolidin-1-yl)ethyl)benzoic acid (1.47 g, 83%) as an off-white solid.

[0299] LCMS (Method 2): the product was identified, m / z 502 (ES+, M+H) at 1.53 min.

[0300] 1H NMR:

[0301] (400MHz, DMSO) δ: 1.76-1.84 (m, 2H,), 2.26 (s, 3H), 2.70-2.90 (m, 4H), 3.18 (s, 3H), 4.72 (brs, 1H), 5.37 (d, 1H) , J=4.0Hz), 7.21-7.45 (m, 8H), 7.74 (dd, 1H, J=7.9and1.9Hz), 7.80-7.86 (m, 3H), 7.98 (s, 1H), 12.86 (brs, 1H).

[0302] Compounds 2R and 2S: Enantiomer separation of racemic compound 2 was performed on a Waters Prep-HPLC system using the following method: column ADH-9.5*250mm 5 microns, solvent A: 0.1% TFA in n-heptane, B: IPA: methanol (70:30), isocratic at 75% A / 25% B, with variable flow (time / flow, 0.01 / 4.0, 5.00 / 4.0, 10.00 / 8.0, 100.00 / 8.0, min / (ml / min)).

[0303] Isomer 1 (0.489 g, 28%) (as off-white solid). LCMS: (Method 2): The product was identified, m / z 502 (ES+, M+H) at 1.56 min.

[0304] 1H NMR: (400MHz, DMSO): δ1.77-1.86 (m, 2H), 2.27 (s, 3H), 2.74-2.90 (m, 4H), 3.14 (brs, 2H), 4.73 (t, 1H, J=6 .4Hz), 7.21-7.47(m, 8H), 7.75(dd, 1H, J=7.9Hz, 1.9Hz), 7.80-7.86(m, 3H), 7.98(s, 1H), 12.86(brs, 1H).

[0305] (Note: some aliphatic protons are masked by DMSO- and / or water)

[0306] Isomer 2 (0.477 g, 27%) (as off-white solid). LCMS: (Method 2): The product was identified, m / z 502 (ES+, M+H) at 1.56 min.

[0307] 1H NMR: (400MHz, DMSO) δ1.77-1.86(m, 2H), 2.27(s, 3H), 2.76-2.90(m, 4H), 3.14(brs, 2H), 4.69-4.76(m, 1H), 5.3 7 (brs, 1H), 7.22-7.45 (m, 8H), 7.75 (dd, 1H, J=7.9Hz, 1.9Hz), 7.81-7.86 (m, 3H), 7.97 (s, 1H), 12.84 (brs, 1H).

[0308] (Note: some aliphatic protons are masked by DMSO- and / or water)

[0309] Embodiment 3: Synthesis of 4-(2-(2-(3-hydroxy-3-(2'-methyl-4'-sulfamoyl-[1,1'-biphenyl]-3-yl)propyl)-5-oxopyrazolidin-1-yl)ethyl)benzoic acid (Compound 3)

[0310] 3-Methyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzenesulfonamide was synthesized by the method described in Example 1, 3-Methyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzenesulfonamide was synthesized from 3-methyl-4-bromobenzenesulfonamide, PdCl2(dppf).DCM and KOAc in dioxane (80°C; 2 hours), and reverse phase flash chromatography was used in the last step to yield 4-(2-(2-(3-hydroxy-3-(2'-methyl-4'-sulfamoyl-[1,1'-biphenyl]-3-yl)propyl)-5-oxopyrazolidin-1-yl)ethyl)benzoic acid as an off-white solid; (0.027 g, 28%). The product was identified by LCMS (Method 3), m / z 538 (ES+, M+H) at 1.90 min.

[0311] 1 H NMR: (400MHz, DMSO) δ: 1.10-1.07 (t, 1H, J=7Hz), 1.82-1.80 (d, 2H, 5.6Hz), 2.08 (s, 1H), 2.29 (s, 4H), 2.87-2..85 (d, 4H, 6.4Hz), 3.13 (s, 3H), 4.73 (s, 1H) 5.39-5.38 (d, 1H, 4.4Hz), 7.35-7.24 (m, 3H), 7.46-7.37 (m, 6H), 7 .71-7.68(m, 1H), 7.75(s, 1H), 7.84-7.82(d, 2H, J=8.4Hz), 12.85(s, 1H).

[0312] Compounds 3R and 3S: The racemic compound 3 was enantiomerically separated using a Waters Prep-HPLC system (CROMEGACHIRAL CCO250*20 mm, 5 μm column) using a mobile phase of 75% of a mixture of 0.1% FA in heptane and 25% of IPA and methanol (50:50) with a flow rate of 18.00 mL / min and no gradient. Isomer 1: (0.019 g, 6.5%; off-white solid). The product was identified by LCMS (Method 3), m / z 538 (ES+, M+H) at 1.90 min.

[0313] 1 H NMR: (400MHz, DMSO) δ: 1.23 (s, 2H), 1.82-1.81 (d, 2H, J=6.4Hz), 2.33-2.29 (m, 4H), 2.83 (s, 4H), 3.16 (s, 5H), 4.74 (s, 1H), 5 .40(s, 1H), 7.25-7.23(m, 3H), 7.44-7.34(m, 6H), 7.71-7.68(m, 1H), 7.75-7.75(d, 1H, J=1.6Hz), 7.82-7.8-(d, 2H, J=8Hz).

[0314] Isomer 2: (0.017 g, 5.8%; off-white solid). The product was identified by LCMS (Method 3), m / z 538 (ES+, M+H) at 1.93 min.

[0315] 1 H NMR: (400MHz, DMSO) δ: 1.23-1.16 (d, 3H, J = 28.0Hz), 1.82 (s, 2H), 2.29 (s, 4H), 2.82 (s, 4H), 3.14 ( s, 2H), 4.74 (s, 1H), 5.39 (s, 1H) 7.24-7.21 (d, 3H, J=1.2Hz), 7.42-7.35 (q, 5H), 7.80-7.69 (m, 4H).

[0316] Embodiment 4: Synthesis of 4-(2-(2-(3-(4'-hydroxy-2'-methyl-[1,1'-biphenyl]-3-yl)-3-oxopropyl)-5-oxopyrazolidin-1-yl)ethyl)benzoate (Compound 4)

[0317]

[0318] Step (i): Intermediate 4 from Example 1 (methyl (4-(2-(2-(3-(4'-hydroxy-2'-methyl-[1,1'-biphenyl]-3-yl)-3-oxopropyl)-5-oxopyrazolidin-1-yl)ethyl)benzoate, 0.17 g, 0.34 mmol) was dissolved in dioxane (3 mL) and water (2 mL). LiOH monohydrate (0.073 g, 1.74 mmol) was added at room temperature and stirred for 3 hours at room temperature. The reaction mixture was then stirred in water (80 mL) and EtOAc (2×50 The mixture was stirred for 2 hours at 4 ℃ for 10 minutes at 4 ℃ for 2 hours.Then the mixture was stirred for 2 hours at 4 ℃ for 3 hours at 4 ℃ for 10 minutes ...

[0319] Step (ii): 4-(2-(2-(3-(4'-hydroxy-2'-methyl-[1,1'-biphenyl]-3-yl)-3-oxopropyl)-5-oxopyrazolidin-1-yl)ethyl)benzoic acid (0.130 g, 0.27 mmol) was dissolved in DMA (3 mL). Sulfamide chloride (0.095 g, 0.82 mmol) was added at room temperature and stirred for 3 hours at room temperature. The reaction mixture was then partitioned between water (70 mL) and EtOAc (30 mL). The aqueous layer was further extracted with EtOAc (2×20 mL). The organic layers were combined and dried (Na2SO4). The solvent was removed in vacuo and the crude product was purified by gradient flash column chromatography eluting the product at 0% to 5% methanol in DCM to give pure 4-(2-(2-(3-(2'-methyl-4'-(sulfamoyloxy)-[1,1'-biphenyl]-3-yl)-3-oxopropyl)-5-oxopyrazolidin-1-yl)ethyl)benzoic acid (0.15 g, 99%) as a white solid. The product was identified by LCMS (Method 1), m / z 552 (ES+) at 2.14 min.

[0320] Step (iiia) (racemic method): 4-(2-(2-(3-(2'-methyl-4'-(sulfamoyloxy)-[1,1'-biphenyl]-3-yl)-3-oxopropyl)-5-oxopyrazolidin-1-yl)ethyl)benzoic acid (0.03 g, 0.054 mmol) was dissolved in methanol (3 mL) and 10% palladium / carbon (0.02 g) containing 50% water was added. H2 gas was purged through the reaction mixture at room temperature for 2 hours. After completion, the reaction mixture was filtered through a bed of celite, washed with MeOH (30 mL), and the filtrate was concentrated in vacuo. The crude product was purified by gradient flash column chromatography (reverse phase, C18 silica) eluting the product at 0% to 35% MeCN in water to give 4-(2-(2-(3-hydroxy-3-(2'-methyl-4'-(sulfamoyloxy)-[1,1'-biphenyl]-3-yl)propyl)-5-oxopyrazolidin-1-yl)ethyl)benzoic acid (0.010 g, 33%) as a brown solid. The product was identified by LCMS (Method 1), m / z 554 (ES+, M+H) at 2.01 min.

[0321] 1 H NMR: (400Mz, MeOD): 1.98-1.95 (t, 3H, J=6.4Hz), 2.26 (s, 3H), 2.95-2.92 (m, 3H, J=6.8Hz), 3.26 (s, 2H), 3.36 (s, 1H), 3.58-3.56 (m, 1H), 3.70-3.68(m, 1H), 4.87-4.85(d, 2H, J=6.4Hz), 7.25-7.12(m, 5H), 7.25(s, 1H), 7.46-7.39(m, 2H), 7.92(s, 2H), 12.72(s, 1H).

[0322] Step (iiib) (Chiral separation method): 4-(2-(2-(3-(2'-methyl-4'-(aminosulfonyloxy)-1,1'-biphenyl)-3-yl)-3-oxopropyl)-5-oxopyrazolidin-1-yl)ethyl)benzoic acid (0.10 g, 0.18 mmol) was dissolved in ethanol (3 mL) and water (2 mL), and CeCl3 (0.13 g, 0.54 mmol) was added to the reaction mixture. The reaction mixture was stirred at 0°C for 5 minutes. After this, sodium borohydride (0.027 g, 0.72 mmol) was added at 0°C, and the reaction mixture was stirred at room temperature for 1 hour. The reaction mixture was then partitioned between water (30 mL) and EtOAc (2×20 mL). The aqueous layer was further acidified with 4N aqueous HCl solution (3 mL), the pH was adjusted to about 1 and extracted with EtOAc (2×50 mL). The organic layers were combined and dried (Na2SO4). The solvent was removed in vacuo to provide the crude product.

[0323] Compounds 4R and 4S: The racemic compound 4 was enantiomerically separated using a Schimadzu Prep-HPLC system (CROMEGACHIRALCCJ 25CM'20mm, 5μm column) using a mobile phase of 60% of 0.1% FA in heptane and 40% of a mixture of IPA and methanol (60:40) with a flow rate of 19.00 mL / min and no gradient. Isomer 1 : (0.0057 g, 24%; white solid) The product was confirmed by LCMS (Method 1), m / z 554 (ES+, M+H) at 2.02 minutes. Chiral HPLC: The purity of the product was confirmed at 16.23 minutes.

[0324] 1 H NMR: (400MHz, MeOD) δ: 1.98-1.93 (m, 2H), 2.27 (s, 3H), 2.97-2.91 (m, 4H), 3.2 7-3.27(m, 2H), 4.88-4.85(t, 2H, J=6.4Hz), 7.23-7.18(m, 4H), 7.34-7.31(m, 3H), 7.46-7.41 (m, 2H), 7.93-7.92 (d, 2H, J=6.8Hz) 7.63-7.61 (d, 2H, J=8.8Hz ), 7.73-7.71 (d, 1H, J=7.6Hz), 7.83-7.78 (m, 3H), 8.58-8.56 (t, 1H, J=5.2Hz).

[0325] Isomer 2: (0.0047 g, 26%; white solid) The product was confirmed by LCMS (Method 1), m / z 554 (ES+, M+H) at 2.02 minutes. Chiral HPLC: The purity of the product was confirmed at 10.62 minutes.

[0326] 1 H NMR: (400MHz, MeOD) δ: 1.98-193 (m, 3H), 2.27 (s, 3H), 2.97-2.93 (m, 4H), 3.27 (s, 2H), 4.88-4.85 (t, 2H, J=6.4Hz ), 7.24-7.25 (m, 4H), 7.31-7.29 (m, 2H), 7.34 (s, 1H), 7.46-7.39 (m, 2H) 7.93-7.91 (d, 2H, J=7.6Hz), 8.51 (s, 1H).

[0327] Embodiment 5: Synthesis of 4-(2-(2-(3-(4'-carbamoyl-2'-methyl-[1,1'-biphenyl]-3-yl)-3-hydroxypropyl)-5-oxopyrazolidin-1-yl)ethyl)-2,6-difluorobenzoic acid (Compound 5)

[0328]

[0329] Step (i): Intermediate 2 methyl 4-(2-(2-(3-(3-bromophenyl)-3-hydroxypropyl)-5-oxopyrazolidin-1-yl)ethyl)-2,6-difluorobenzoate (0.20 g, 0.60 mmol), 3-methyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzamide (0.21 g, 0.79 mmol) and KCO (0.17 g, 0.1.30 mmol) were dissolved in a mixture of dioxane:water (1:1) (5 mL) and purged with nitrogen at room temperature for 30 min. After this, PdCl(dppf).DCM (0.099 g, 0.12 mmol) was added and the reaction mixture was stirred at 80°C for 1 h. The reaction mixture is then distributed between water (70mL) and EtOAc (70mL), and water layer is further extracted with EtOAc (2 × 30mL). The organic layers are merged and dried (Na2SO4). The solvent is removed in a vacuum, and the crude product is purified by reversed-phase gradient flash column chromatography (reverse phase, C18 silica), eluted under 0% to 28%MeCN in water, to obtain 4-(2-(2-(3-(4'-carbamoyl-2'-methyl-[1,1'-biphenyl]-3-yl)-3-hydroxypropyl)-5-oxopyrazolidine-1-yl)ethyl)-2,6-difluorobenzoic acid methyl esters (0.29g, 87%) as a light brown solid. The product is determined by LCMS (method 2), m / z 552 (ES+, M+H), at 1.92 minutes.

[0330] Step (ii): Methyl 4-(2-(2-(3-(4'-carbamoyl-2'-methyl-[1,1'-biphenyl]-3-yl)-3-hydroxypropyl)-5-oxopyrazolidin-1-yl)ethyl)-2,6-difluorobenzoate (0.29 g, 0.29 mmol) was dissolved in dioxane (4 mL) and water (2 mL). LiOH (0.11 g, 0.88 mmol) was added at room temperature and stirred at room temperature for 16 hours. The reaction mixture was acidified with 4N aqueous HCl (4 mL) to adjust the pH to about 1 and extracted with EtOAc (40 mL). The aqueous layer was further extracted with EtOAc (3×30 mL). The organic layers were combined and dried (Na2SO4). The solvent was removed in vacuo and the crude product was purified by reverse phase gradient flash column chromatography (reverse phase, C18 silica) eluting the product at 0% to 23% ACN in water to give 4-(2-(2-(3-(4'-carbamoyl-2'-methyl-[1,1'-biphenyl]-3-yl)-3-hydroxypropyl)-5-oxopyrazolidin-1-yl)ethyl)-2,6-difluorobenzoic acid (0.15 g, 53%) as an off-white solid. The product was identified by LCMS (Method 2), m / z 538 (ES+, M+H) at 1.49 min.

[0331] 1 H NMR: (400MHz, DMSO) δ: 1.23 (s, 2H), 1.81-1.80 (d, 2H, J=4.8Hz), 2.26 (s, 3H), 2 .75(s, 3H), 3.17-3.16(d, 4H, 3.6Hz), 4.12-4.11(d, 1H, J=4.8Hz), 4.71(s, 1H), 5.47-5.46 (d, 1H, J=3.6Hz), 6.75-6.73 (d, 2H, J=7.2Hz), 7.25-7.22 (m, 2H), 7.3 7-7.33(m, 2H), 7.43-7.39(m, 2H), 7.76-7.74(m, 1H), 7.82(s, 1H), 8.02(s, 1H).

[0332] Compounds 5R and 5S: The racemic mixture (70 mg) was separated by enantiomers using a Shimadzu Prep-HPLC system (CHROMEGACHIRAL CCO 250*25 mm, 5 μm column) using a mobile phase of 80% of 0.1% FA in heptane and 20% of a mixture of IPA and methanol (70:30) with no gradient and a flow rate of 20.00 mL / min. different Construct 1:(0.038 g, 13%; white solid) The product was confirmed by LCMS (Method 2), m / z 538 (ES+, M+H) at 1.59 minutes. Chiral HPLC: the purity of the product was confirmed at 14.48 minutes.

[0333] 1 H NMR: (400MHz, DMSO) δ: 1.81-1.80 (d, 2H, J=6.4Hz), 2.27 (s, 4H), 2.85 (s, 4H), 3.13-3.12 (d, 2H, J=5.6Hz), 4.73 (s, 1H), 5.36 (s, 1H), 7.10-7.04 (m, 2H), 7.27-7.22 (m, 2H), 7.43-7.34 (m, 4H), 7.76-7.74 (m, 1H), 7.82 (s, 1H), 7.97 (s, 1H), 13.78 (s, 1H).

[0334] Isomer 2: (0.052 g, 18%; off-white solid) The product was confirmed by LCMS (Method 2), m / z 538 (ES+, M+H) at 1.60 min. Chiral HPLC: the purity of the product was confirmed at 10.50 min.

[0335] 1 H NMR: (400MHz, DMSO) δ: 1.81-1.80 (d, 2H, J=5..6Hz), 2.27 (s, 3H), 2.85 (s, 3H), 3.14-3.12 (d, 2H, 7.6Hz), 4.72 (s, 1H), 5.37 (s, 1H) ), 7.06-7.04(d, 2H, J=8.8Hz), 7.27-7.22(m, 2H), 7.43-7.34(m, 4H), 7.76-7.74(m, 1H), 7.82(s, 1H), 7.97(s, 1H), 13..79(s, 1H).

[0336] Embodiment 6: Synthesis of 2-hydroxy-4-(2-(2-(3-hydroxy-3-(2'-methyl-4'-sulfamoyl-[1,1'-biphenyl]-3-yl)propyl)-5-oxopyrazolidin-1-yl)ethyl)benzoic acid (Compound 6)

[0337]

[0338] Step (i): Intermediate 3 4-(2-(2-(3-(3-bromophenyl)-3-hydroxypropyl)-5-oxopyrazolidin-1-yl)ethyl)-2-hydroxybenzoic acid methyl ester (0.15 g, 0.32 mmol), 3-methyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzenesulfonamide (0.14 g, 0.47 mmol) and K2CO3 (0.89 g, 0.64 mmol) were dissolved in 4 mL of a 1:1 dioxane:water mixture. Nitrogen was purged at room temperature for 30 minutes. After this, PdCl2(dppf).DCM (0.026 g, 0.032 mmol) was added and the reaction mixture was stirred at 80°C for 1 hour. The reaction mixture was then partitioned between water (50 mL) and EtOAc (50 mL). The aqueous layer is further extracted with EtOAc (2 × 20mL). The organic layers are merged and dried (Na2SO4). Remove solvent in a vacuum, and the crude product is purified by gradient flash column chromatography (reverse phase, C18 silica), eluted under 0% to 55%MeCN in water, to obtain 2-hydroxy-4-(2-(2-(3-hydroxy-3-(2'-methyl-4'-sulfamoyl-[1,1'-biphenyl]-3-yl)propyl)-5-oxopyrazolidine-1-yl)ethyl)methyl benzoate (0.13g, 73%) as a white solid. The product is determined by LCMS (method 2), m / z 568 (ES+) at 1.86 minutes.

[0339] Step (ii): Methyl 2-hydroxy-4-(2-(2-(3-hydroxy-3-(2'-methyl-4'-sulfamoyl-[1,1'-biphenyl]-3-yl)propyl)-5-oxopyrazolidin-1-yl)ethyl)benzoate (0.13 g, 0.23 mmol) was dissolved in 4 mL of a 1:1 dioxane:water mixture at room temperature, and LiOH monohydrate (0.048 g, 1.15 mmol) was added, and the reaction mixture was stirred at room temperature for 3 hours. The reaction mixture was partitioned between water (30 mL) and EtOAc (2×50 mL). The aqueous layer was further acidified with 4N aqueous HCl (2 mL) to adjust the pH to about 1, and then extracted with EtOAc (2×50 mL). The organic layers were combined and dried (Na2SO4). The solvent was removed in vacuo to give pure 2-hydroxy-4-(2-(2-(3-hydroxy-3-(2'-methyl-4'-sulfamoyl-[1,1'-biphenyl]-3-yl)propyl)-5-oxopyrazolidin-1-yl)ethyl)benzoic acid (0.050 g, 39%) as an off-white solid. The product was identified by LCMS (Method 2), m / z 554 (ES+, M+H) at 1.90 min.

[0340] Compounds 6R and 6S: The racemic compound 6 (50 mg) was separated by enantiomers using a Waters Prep-HPLC system (CHIRALPAKAD-H 250X10mm 5μm column) using a mobile phase of 80% 0.1% FA in heptane and 20% IPA:acetonitrile (70:30) with no gradient and a flow rate of 7.00 mL / min. Isomer 1: (0.006 g, 4.7%; sticky solid) The product was confirmed by LCMS (Method 2), m / z 554 (ES+, M+H) at 1.86 min. Chiral HPLC: the purity of the product was confirmed at 18.45 min.

[0341] 1 H NMR: (400MHz, MeOD) δ: 1.971-1.955 (d, 3H, J=6.4Hz), 2.331 (s, 3H), 2.908-2.853 (m, 4H), 3.59-3.56 (t, 3H, J=4.8Hz), 3.70-3.68 (t, 3H, J=

[0342] 4.6Hz), 6.81 (s, 2H), 7.27-7.25 (d, 1H, J=7.2Hz), 7.38-7.36 (d, 2H, J=8.0Hz), 7.49-7.43 (m,

[0343] 2H), 7.78-7.76 (d, 2H, J=7.6Hz), 7.84 (s, 1H).

[0344] Isomer 2: (0.007 g, 4.0%; sticky solid) The product was confirmed by LCMS (Method 3), m / z 554 (ES+, M+H) at 1.87 minutes. Chiral HPLC: the purity of the product was confirmed at 29.47 minutes.

[0345] 1 H NMR: (400MHz, MeOD) δ: 1.99-1.96 (m, 3H), 2.33-2.30 (d, 3H, J=10.4Hz), 2.91-2.86 (m, 4H), 3.58-3.56 (t, 2H, J=4.6Hz), 3.70-3.68 (t, 2H, J= 4.6Hz), 6.82 (s, 2H), 7.27-7.25 (d, 1H, J=7.2Hz), 7.38-7.36 (d, 2H, J=8.0Hz), 7.49-7.43 (m, 2H), 7.78-7.76 (d, 2H, J=8.0Hz), 7.83 (s, 1H).

[0346] Embodiment 7:Synthesis of 4-(2-(2-(3-(4'-carbamoyl-2'-methyl-[1,1'-biphenyl]-3-yl)-3-hydroxypropyl)-5-oxopyrazolidin-1-yl)ethyl)-2-hydroxybenzoic acid (Compound 7)

[0347] Compound 7 was synthesized by the method described in Example 6 using 3-methyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzamide to give 4-(2-(2-(3-(4'-carbamoyl-2'-methyl-[1,1'-biphenyl]-3-yl)-3-hydroxypropyl)-5-oxopyrazolidin-1-yl)ethyl)-2-hydroxybenzoic acid (0.075 g, 86%) as an off-white solid. The product was identified by LCMS (Method 2), m / z 518 (ES+, M+H) at 1.62 minutes.

[0348] Compounds 7R and 7S: Racemic compound 7 (75 mg) was separated using a Waters Prep-HPLC system (CHIRALPAKAD-H 250X10mm 5μm column) using a mobile phase of 75% 0.1% FA in heptane and 25% IPA in acetonitrile (70:30) at a flow rate of 8.00 mL / min with no gradient. Isomer 1: (0.016 g, 19%; off-white solid) The product was confirmed by LCMS (Method 2), m / z 518 (ES+, M+H) at 1.74 minutes. Chiral HPLC: the purity of the product was confirmed at 11.21 minutes.

[0349] 1 H NMR: (400MHz, DMSO) δ: 1.81-1.80 (d, 2H, J=6.0Hz), 2.27 (s, 3H), 2.54 (s, 1H), 2.80-2.7 (d, 4H, J=6.4Hz), 3.153 (s, 3H), 4.74-4.71 (t, 1H, J=6.0Hz), 5.32 (s, 1H), 6.78-6.74 (m , 2H) 7.27-7.22 (m, 2H), 7.36-7.33 (m, 2H), 7.43-7.38 (m, 2H), 7.68-7.66 (d, 1H, J=8.0H z), 7.76-7.73 (dd, 1H, J=1.6Hz), 7.81 (s, 1H), 7.97 (s, 1H), 11.20 (s, 1H), 13.84 (s, 1H).

[0350] Isomer 2:(0.025 g, 29%; off-white solid) The product was confirmed by LCMS (Method 2), m / z 518 (ES+, M+H) at 1.73 minutes. Chiral HPLC: the purity of the product was confirmed at 9.72 minutes.

[0351] 1 HNMR: (400MHz, DMSO) δ: 1.81-1.80 (d, 2H, J=6.4Hz), 2.27 (s, 4H), 2.80 (s, 4H), 3.15-3.14 (d, 3H, J=6.4Hz), 4.74-4.72 (d, 1H, J=6.0Hz), 5.31 (s, 1H), 6.76-6.74 (d, 1H, J=8.4Hz), 6. 78(s, 1H), 7.27-7.22(m, 2H), 7.36-7.33(m, 2H), 7.43-7.38(m, 2H), 7.68-7.66(d, 1H, J=8. 0Hz), 7.76-7.73 (dd, 1H, J=1.6Hz), 7.81 (s, 1H), 7.97 (s, 1H), 11.20 (s, 1H), 13.84 (s, 1H).

[0352] Embodiment 8: Synthesis of 2,6-difluoro-4-(2-(2-(3-hydroxy-3-(2'-methyl-4'-sulfamoyl-[1,1'-biphenyl]-3-yl)propyl)-5-oxopyrazolidin-1-yl)ethyl)benzoic acid (Compound 8)

[0353] Compound (8) was synthesized by the method described in Example 5 using 3-methyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzenesulfonamide to give 2,6-difluoro-4-(2-(2-(3-hydroxy-3-(2'-methyl-4'-sulfamoyl-[1,1'-biphenyl]-3-yl)propyl)-5-oxopyrazolidin-1-yl)ethyl)benzoic acid (0.040 g, 23%) as a white solid. The product was identified by LCMS (Method 2), m / z 574 (ES+, M+H) at 1.58 minutes.

[0354] Compounds 8R and 8S: The racemic compound (8) (40 mg) was separated by enantiomers using a Shimadzu Prep-HPLC system (CHROMEGACHIRAL CCO 250*25 mm, 5 μm column) using a mobile phase of 80% of a mixture of 0.1% FA in heptane and 20% of IPA and acetonitrile (70:30) with no gradient and a flow rate of 22.00 mL / min. Isomer 1:(0.010 g, 5.7%; viscous gum) The product was confirmed by LCMS (Method 2), m / z 574 (ES+, M+H) at 1.58 min. Chiral HPLC: the purity of the product was confirmed at 28.03 min.

[0355] 1 H NMR: (400MHz, MeOD) δ: 1.30

[0356] (s, 3H), 1.98-1.96 (d, 3H, J=6.0Hz), 2.34 (s, 4H), 2.96-2.93 (t, 6H, J=6.6Hz), 6.98-6.95 (d, 2H,

[0357] J=9.2Hz), 7.28-7.25(m, 1H), 7.38-7.36(m, 2H), 7.50-7.43(m, 2H), 7.78-7.76(dd, 1H, J=

[0358] 1.5 Hz), 7.84-7.84 (m, 1H).

[0359] Isomer 2 : (0.012 g, 6.8%; viscous gum) The product was confirmed by LCMS (Method 2), m / z 574 (ES+, M+H) at 1.58 min. Chiral HPLC: The purity of the product was confirmed at 20.05 min.

[0360] 1 H NMR: (400MHz, MeOD) δ: 1.30 (s, 3H), 1.97-1.96 (d, 2H, J=5.6Hz), 2.34-2.32 (m, 3H), 2.96-2.93 (t, 4H, J=6.4Hz), 6.98-6. 95 (d, 2H, J=9.2Hz), 7.28-7.26 (m, 1H), 7.38-7.36 (m, 2H), 7.48-7.45 (m, 2H), 7.78-7.76 (dd, 1H, J=1.5Hz), 7.84 (s, 1H).

[0361] Embodiment 9: Synthesis of (3'-(3-(2-(4-(1H-tetrazolyl-5-yl)phenethyl)-3-oxopyrazolidin-1-yl)-1-hydroxypropyl)-2-methyl-[1,1'-biphenyl]-4-carboxamide (Compound 9)

[0362]

[0363] Step (i): 1-Bromo-4-(2-bromoethyl)benzene (10.00 g, 38.18 mmol), tert-butylhydrazinecarboxylate (7.57 g, 57.27 mmol), NaHCO (12.84 g, 15.27 mmol) and NaI (0.57 g, 3.82 mmol) were suspended in MeCN (100 mL) at room temperature and the reaction mixture was stirred at 70° C. for 72 hours. The reaction mixture was concentrated under vacuum and the residue was partitioned between water (1000 mL) and EtOAc (800 mL), and the aqueous layer was further extracted with EtOAc (3×500 mL). The organic layers were combined and dried (Na 2 SO 4 ). The solvent was removed in vacuo and the crude product was purified by gradient column chromatography (normal phase, silica) eluting the product at 0% to 15% EtOAc in hexanes to afford tert-butyl 2-(4-bromophenethyl)hydrazine-1-carboxylate (5.0 g, 42%) as an off-white solid. The product was identified by LCMS (Method 2), m / z 260 (ES+, M+H-tBu) at 2.34 minutes.

[0364] Step (ii): At room temperature, tert-butyl 2-(4-bromophenethyl)hydrazine-1-formate (5.00 g, 15.89 mmol) was dissolved in DMF (50 mL), potassium carbonate (10.98 g, 79.47 mmol) was added, and stirred at room temperature for 15 minutes. Then 3-bromopropionyl chloride (2.40 mL, 23.56 mmol) was added dropwise at room temperature, and the reaction mixture was stirred at room temperature for 24 hours. The reaction mixture was concentrated under vacuum. The obtained residue was distributed between water (500 mL) and EtOAc (400 mL), and the aqueous layer was further extracted with EtOAc (2×300 mL). The organic layers were combined, dried over Na2SO4, and the solvent was removed in vacuo. The crude product was purified by gradient column chromatography (normal phase, silica) with 0% to 30% EtOAc in hexanes to give tert-butyl 2-(4-bromophenethyl)-3-oxopyrazolidine-1-carboxylate (4.0 g, 68%) as a yellow sticky solid. The product was identified by LCMS (Method 2), m / z 313 (ES+, M+H-tBu), 2.65 min.

[0365] Step (iii): tert-Butyl 2-(4-bromophenethyl)-3-oxopyrazolidine-1-carboxylate (2.00 g, 5.43 mmol) was dissolved in DMF (20 mL). Purged with nitrogen at room temperature for 15 minutes. After this, zinc cyanide (1.28 g, 10.87 mmol) and tetrakistriphenylphosphine-palladium 0 (0.63 g, 0.54 mmol) were added, and the reaction mixture was stirred in a microwave at 150 ° C for 1 hour (the reaction was divided into 4 reaction vials). The reaction mixture was distributed between water (400 mL) and EtOAc (300 mL). The aqueous layer was further extracted with EtOAc (2×200 mL). The organic layers were combined and dried (Na2SO4). The solvent was removed in vacuo and the crude product was purified by reverse phase gradient flash column chromatography (reverse phase, C18 silica), eluting the product at 0% to 68% MeCN in water to give 4-(2-(5-oxopyrazolidin-1-yl)ethyl)benzonitrile (0.65 g, 41%) as a yellow solid. The product was identified by LCMS (Method 2), m / z 216 (ES+, M+H) at 1.63 min.

[0366] Step (iv): 4-(2-(5-oxopyrazolidin-1-yl)ethyl (0.60 g, 2.79 mmol) and 1-(3-bromophenyl)prop-2-en-1-one (2.90 g, 13.95 mmol) were suspended in MeOH (6 mL) at room temperature, and the reaction mixture was stirred at room temperature for 15 minutes. After that, TEA (2.00 mL, 13.95 mmol) was added at room temperature and stirred at 60° C. for 4 hours. The reaction mixture was washed with water (200 mL) and EtOAc (150 mL). The mixture was stirred for 2 hours at 4 ℃ for 10 minutes.Then the mixture was stirred for 2 hours at 4 ℃ for 3 ...

[0367] Step (v): 4-(2-(2-(3-(3-bromophenyl)-3-oxopropyl)-5-oxopyrazolidin-1-yl)ethyl)benzonitrile (0.50 g, 1.18 mmol), 3-methyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzamide (0.46 g, 1.76 mmol) and KCO (0.33 g, 2.35 mmol) were dissolved in 1,4-dioxane (5 mL) and water (5 mL). Nitrogen was purged at room temperature for 20 minutes. After this, PdCl(dppf).DCM (0.096 g, 0.12 mmol) was added and the reaction mixture was stirred at 80° C. for 2 hours. The reaction mixture was partitioned between water (200 mL) and EtOAc (150 mL). The aqueous layer was further extracted with EtOAc (2×100 mL). The organic layers were combined and dried (Na2SO4). The solvent was removed in vacuo and the crude product was purified by reverse phase gradient flash column chromatography (reverse phase, C18 silica), eluting the product at 0% to 55% MeCN in water to obtain 3'-(3-(2-(4-cyanophenylethyl)-3-oxopyrazolidine-1-yl)propanoyl)-2-methyl-[1,1'-biphenyl]-4-carboxamide (0.25 g, 44%) as an off-white solid. The product was determined by LCMS (Method 2), m / z 481 (ES+, M+H), at 2.18 minutes.

[0368] Step (vi) 3'-(3-(2-(4-cyanophenethyl)-3-oxopyrazolidin-1-yl)propanoyl)-2-methyl-[1,1'-biphenyl]-4-carboxamide (0.20 g, 0.42 mmol) was dissolved in ethanol (2 mL) and water (2 mL). After this, NaBH4 (0.03 g, 0.83 mmol) was added at 0°C and stirred at room temperature for 1 hour. The reaction mixture was partitioned between water (50 mL) and EtOAc (50 mL), and the aqueous layer was further extracted with EtOAc (2×30 mL). The organic layers were combined and dried (Na2SO4). The solvent was removed in vacuo to give pure 3'-(3-(2-(4-cyanophenethyl)-3-oxopyrazolidin-1-yl)-1-hydroxypropyl)-2-methyl-[1,1'-biphenyl]-4-carboxamide (0.17 g, 85%) as an off-white solid. The product was identified by LCMS (Method 2), m / z 483 (ES+, M+H) at 2.14 min.

[0369] Step (vii): 3'-(3-(2-(4-cyanophenethyl)-3-oxopyrazolidin-1-yl)-1-hydroxypropyl)-2-methyl-[1,1'-biphenyl]-4-carboxamide (0.17 g, 0.35 mmol) was suspended in DMF (2 mL) at room temperature, and sodium azide (0.24 g, 3.50 mmol) and ammonium chloride (0.20 g, 3.50 mmol) were added and stirred at 100°C for 48 hours. The reaction mixture was partitioned between water (70 mL) and EtOAc (50 mL) and the aqueous layer was further extracted with EtOAc (3 x 50 mL). The organic layers were combined and dried (Na2SO4). The solvent was removed in vacuo and the crude product was purified by reverse phase gradient flash column chromatography (reverse phase, C18 silica) eluting the product at 0% to 45% MeCN in water to give (3'-(3-(2-(4-(1H-tetrazol-5-yl)phenethyl)-3-oxopyrazolidin-1-yl)-1-hydroxypropyl)-2-methyl-[1,1'-biphenyl]-4-carboxamide (0.10 g, 54%) as an off-white solid. The product was confirmed by LCMS (Method 2), m / z 526 (ES+) at 1.91 min. Chiral HPLC: product purity was confirmed at 13.67 min and 15.68 min.

[0370] 1 H NMR (400MHz, DMSO): 1.85-1.83 (d, 2H, J=6.4Hz), 2.29 (s, 3H), 2.89-2.86 (m, 4H), 3.18 (S, 4H), 4.76 (S, 1H), 5.40 (m, 1H), 7.29-7.24 (m, 2H), 7.45-7.36 (m, 6H), 7.76-7.76 (d, 1H, J=1.2Hz), 7.83 (s, 1H), 7.95-7.93 (d, 2H, J=8.0Hz), 8.00 (s, 1H).

[0371] Compounds 9R and 9S: The racemic compound 9 (90 mg) was separated by enantiomers using a Waters 600 Controller-HPLC system (CHIRALPAK_IG_SFC_21 mm*250 mm, 5 μm column). The mobile phase used was 90% heptane and 10% IPA and acetonitrile (70:30), with no gradient and a flow rate of 23.00 mL / min. Isomer 1: (0.028 g, 31%; white solid) The product was confirmed by LCMS (Method 2), m / z 526 (ES+, M+H) at 1.75 minutes. Chiral HPLC: the purity of the product was confirmed at 13.19 minutes.

[0372] 1 H NMR: (400MHz, DMSO) 1.82-1.81 (d, 2H, J=5.6Hz), 2.10 (s, 3H), 2.87-2.83 (t, 4H, J=6.6Hz), 3.15 (s, 4H), 4.73 (s, 1H), 5.3 6 (s, 1H), 7.26-7.22 (m, 2H), 7.42-7.33 (m, 6H), 7.75-7.73 (m, 1H), 7.81 (s, 1H), 7.92-7.90 (d, 2H, J=8.0Hz), 7.97 (s, 1H).

[0373] Isomer 2: (0.025 g, 28%; white solid) The product was confirmed by LCMS (Method 2), m / z 526 (ES+, M+H) at 1.76 minutes. Chiral HPLC: Product purity was confirmed at 14.89 minutes. (98%)

[0374] 1 H NMR: (400MHz, DMSO): 1.82-1.81 (d, 2H, J=6.0Hz), 2.26 (s, 3H), 2.83-2.66 (m, 4H), 3.07 (s, 4H), 4.73 (s, 1H) , 5.37 (s, 1H), 7.43-7.22 (m, 8H), 7.75-7.73 (m, 1H), 7.80 (s, 1H), 7.90-7.89 (d, 2H, J=8.0Hz), 7.98 (s, 1H).

[0375] Embodiment 10: Biological assays

[0376] Cloning, baculovirus production, large-scale infection and membrane preparation of HEK293 cells: Human prostaglandin E2 receptor 4 (EP4) was cloned into the pBacMam expression vector (GeneScript, UK). Transposition of EP4 DNA was performed using the Bac-to-Bac baculovirus expression system of Invitrogen. P0 baculovirus was produced by transfecting SF9 cells with bacmid DNA using Cellfectin II transfection reagent (ThermoFisherScientific, UK). After the P0 generation, P1 virus was then produced for large-scale infection and membrane preparation. HEK293 cells were cultured in DMEM (ThermoFisher Scientific, UK) supplemented with 10% heat-inactivated fetal bovine serum (fetal bovine serum, FBS). Cells were incubated at 500 cm 3Infection was carried out in flasks at an inoculation density of 3.5 million cells / mL under 5% v / v EP4Bacman. Expression was carried out for 36 hours at 37°C and 5% CO2. Cells were removed using PBS and a cell scraper. Cell culture was centrifuged at 2500RPM for 10 minutes at 4°C. The supernatant was then poured out and the precipitate was stored at -80°C. The precipitate was thawed and resuspended in 15mL homogenization buffer (20mM HEPES, 10mM EDTA, pH 7.4). Homogenization was then carried out in a mechanical homogenizer (VMR) for 10 seconds. The membrane was centrifuged at 40,000g for 15 minutes at 4°C in a centrifuge tube. The supernatant was poured out and resuspended in 15mL homogenization buffer. Homogenization was carried out for 20 seconds. The membrane was centrifuged at 40,000g for 45 minutes at 4°C. The membrane was resuspended in 3 mL of well-mixed storage buffer (20 mM HEPES, 0.1 mM EDTA, pH 7.4). The resulting membrane was then stored at -80°C.

[0377] cAMP Gs functional assay; cAMP production upon EP4 receptor activation was determined using the Homogeneous Time-Resolved Fluorescence (HTRF) cAMP dynamic-2 assay (Cisbio, France). HEK293 cells were transfected with 0.5% EP4 Bacmam virus for 36 hours, then cells were dissociated and frozen at -150°C.

[0378] On the day of the assay, increasing concentrations of test compounds were added to ProxiPlate-384Plus, white 384-shallow well microplates (PerkinElmer, USA) using an ECHO dispenser along with a positive control (10 uM PGE2 (Tocris, Abingdon, UK)) and a negative control (DMSO (Sigma-Aldrich, UK)).

[0379] The cells were thawed in a water bath and resuspended in DMEM supplemented with 10% FBS, then centrifuged at 1200 RPM for 5 minutes to form a pellet. The pellet was resuspended in assay buffer (DMEM + 0.5 mM IBMX (Tocris, Abingdon, UK)) to a concentration of 0.5 × 10 6 Cells / mL. Cell suspensions were added to pre-assigned assay plates using multidrops, with a final assay concentration of 5000 cells / well. The plates were then incubated at 37°C and 5% CO2 for 30 minutes. cAMP production was determined according to the manufacturer's instructions, and the plates were then read on a PheraStar fluorescence plate reader (BMG LabTech, Germany).

[0380] pEC was calculated using Dotmatics from the midpoint of the curve. 50 Values ​​(-Log M) are shown in Table 2.

[0381] Table 2: Activity of selected compounds of formula (1) at prostaglandin E2 receptor 4 (cAMP human EP4) (pEC 50 and Emax value)

[0382]

[0383]

[0384] One-way CACO-2 cell permeability assay

[0385] Caco-2 cells (ECACC) were cultured at 2×10 5 Cells were seeded into 24-well Transwell plates and used for confluent monolayers after culturing for 21 days at 37°C, 5% CO2. Test compounds were incubated in assay buffer (Hanks balanced salt solution supplemented with 25mM HEPES, adjusted to pH 6.5) at 10 μM, 0.2% DMSO final concentration, n=2. Hanks balanced salt solution supplemented with 25mM HEPES (0.2% DMSO final concentration) adjusted to pH 7.4 was used for the basolateral chamber (as a receiver).

[0386] Incubations were performed at 37°C with samples removed from both the donor and acceptor compartments at T=0 and 1 hour and compounds analyzed by mass spectrometry (LC-MS / MS) including an analytical internal standard (0.5 μM carbamazepine).

[0387] Apparent permeability (P app ) values ​​are shown in Table 3 and are determined according to the following relationship:

[0388]

[0389] Where V is the volume of each Transwell compartment (top 125 μL, basolateral 600 μL), and Concentration is the relative MS response of compound in the donor compartment before incubation and in the receptor compartment at the end of incubation (normalized to internal standard). Area = area of ​​cells exposed for drug transfer (0.33 cm 2 ).

[0390] Lucifer Yellow (LY) was added to the top buffer in all wells to assess the viability of the cell layer. Since LY cannot freely permeate the lipophilic barrier, a high degree of LY transport indicates poor integrity of the cell layer and rejection of LY.app >10×10 -6 cm / sec hole.

[0391] Compounds recovered from the wells were determined based on the MS responses in the donor and acceptor compartments at the end of the incubation compared to the response in the donor compartment before the incubation (normalized to an internal standard).

[0392] Table 3 - Average apparent permeability (P app )value

[0393]

[0394] It should be understood that although the invention has been described in conjunction with its detailed description, the foregoing description is intended to illustrate and not limit the scope of the invention, which is defined by the scope of the appended claims. Other aspects, advantages and modifications are within the scope of the appended claims.

[0395] Numbered implementation plan

[0396] 1. Compounds of formula I:

[0397]

[0398] or a pharmaceutically acceptable salt or tautomer thereof, wherein;

[0399] A is OR', C(O)R', CO2R', C(O)N(R')2, C(O)N(R')S(O)2R', S(O)2R', S(O)2OR', SO2N(R')2, C 1-8 alkyl, cycloalkyl, heterocycloalkyl, aryl or heteroaryl;

[0400] X is halogen, OR', COOR' or C 1-6 alkyl;

[0401] L and L' are each independently C 2-4 Alkylene;

[0402] R 1 H, halogen, CN, NO2, OR', SR', COOR', C 1-6 Alkoxy or C 1-6 alkyl;

[0403] R 2 is OR', OC(O)R 3 ,OC(O)OR 3 , CO2R', CON(R')2, SO2N(R')2, SO2R 3 ,OSO2R 3 , or OSO2N(R')2;

[0404] R3 It is C 1-6 Alkyl, C 3-6 Cycloalkyl or phenyl;

[0405] R' is H, C 1-6 Alkyl or C 3-6 cycloalkyl; and

[0406] n is 0, 1, 2, or 3;

[0407] wherein at each occurrence, alkyl, alkylene, alkoxy and cycloalkyl are each optionally and independently substituted by OH, SH, CN, NO2, COOH, halogen or COOC 1-4 Up to 3 instances of substitution in the alkyl group;

[0408] wherein at each occurrence, heterocycloalkyl, aryl and heteroaryl are each optionally and independently substituted by OR', SR', CN, NO2, CO2R', halogen, C 1-4 Up to 3 instances of alkyl or oxo are substituted.

[0409] 2. The compound according to embodiment 1, wherein the compound is a compound of formula (1):

[0410]

[0411] or a pharmaceutically acceptable salt or tautomer thereof, wherein A, X, R 1 , R 2 and n are the same as defined in Embodiment 1.

[0412] 3. A compound according to Embodiment 1 or Embodiment 2, wherein A is selected from C(O)OR', C(O)N(R')S(O)2R 3 ,S(O)2OR',C 1-8 alkyl, heterocycloalkyl or heteroaryl, wherein the heterocycloalkyl and heteroaryl are each optionally and independently substituted by OR', SR', halogen, C 1-4 Up to 3 instances of alkyl or oxo are substituted.

[0413] 4. A compound according to Embodiment 1 or Embodiment 2, wherein A is selected from:

[0414]

[0415] 5. A compound according to any one of embodiments 1 to 4, wherein A is:

[0416]

[0417] 6. A compound according to any one of embodiments 1 to 5, wherein X is halogen or OR'.

[0418] 7. A compound according to embodiment 6, wherein X is F or OH, and n is 1 or 2.

[0419] 8. A compound according to any one of embodiments 1 to 7, wherein R 1 is H, OH, halogen, CN, C optionally substituted by 1 to 3 fluorine atoms 1-6 alkyl, or C 1-6 alkyl.

[0420] 9. The compound according to embodiment 8, wherein R 1 It's methyl.

[0421] 10. A compound according to any one of embodiments 1 to 9, wherein R 2 It is CON(R')2, SO2N(R')2, OSO2R 3 Or OSO2N(R')2.

[0422] 11. The compound according to embodiment 10, wherein R 2 It is CONH2, SO2NH2 or OSO2NH2.

[0423] 12. A compound according to any one of embodiments 1 to 8, which is a compound of formula (2a), formula (2b), formula (2c), or formula (2d):

[0424]

[0425] or a pharmaceutically acceptable salt or tautomer thereof, wherein X, R 1 , R 2 and n are the same as defined in Embodiment 1.

[0426] 13. The compound according to Embodiment 1, which is a compound of Formula (5), Formula (5a), Formula (5b), Formula (6), Formula (6a), Formula (6b):

[0427]

[0428] or a pharmaceutically acceptable salt thereof.

[0429] 14. The compound according to embodiment 1, wherein the compound is selected from:

[0430]

[0431] or a pharmaceutically acceptable salt thereof.

[0432] 15. A pharmaceutical composition comprising a compound according to any one of embodiments 1 to 14 and a pharmaceutically acceptable excipient.

[0433] 16. A compound according to any one of embodiments 1 to 14 or a composition according to embodiment 15 for use in the treatment of an EP4 receptor mediated disease.

[0434] 17. The compound or composition for use according to embodiment 16, wherein the EP4 receptor-mediated disease is a gastrointestinal disorder.

[0435] 18. The compound or composition for use according to embodiment 17, wherein the gastrointestinal disorder is selected from constipation disorder, constipation-predominant irritable bowel syndrome, mixed irritable bowel syndrome, chronic idiopathic constipation, gastrointestinal symptoms associated with Parkinson's disease, gastrointestinal symptoms associated with cystic fibrosis, intestinal motility disorders, postoperative ileus, food allergy or food intolerance, celiac disease, gastrointestinal motility disorders, functional gastrointestinal disorders, drug-induced enteropathy, NSAID-induced gastric and intestinal damage, chemotherapy-induced mucositis, gastroesophageal reflux disease (GERD), duodenogastric reflux, diarrheal diseases, immune-mediated gastrointestinal diseases, Crohn's disease, ulcerative colitis, inflammatory bowel disease and ischemic colitis.

[0436] 19. The compound or composition for use according to embodiment 16, wherein the EP4 receptor-mediated disease is a pulmonary disease or disorder.

[0437] 20. The compound or composition for use according to embodiment 19, wherein the pulmonary disease or disorder is selected from chronic obstructive pulmonary disease, asthma, chronic bronchitis, cystic fibrosis, emphysema, chronic idiopathic cough, overactive airways disorder and idiopathic pulmonary fibrosis.

Claims

1. A compound of formula I or a pharmaceutically acceptable salt, solvate, hydrate, tautomer or optical isomer thereof: in: A is OR', C(O)R', CO2R', C(O)N(R')2, C(O)N(R')S(O)2R 3 ,S(O)2R',S(O)2OR',SO2N(R')2,C 1-8 alkyl, cycloalkyl, heterocycloalkyl, aryl or heteroaryl; X is halogen, OR', COOR' or C 1-6 alkyl; L and L' are each independently C 2-4 Alkylene; R 1 H, halogen, CN, NO2, OR', SR', COOR', C 1-6 Alkoxy or C 1-6 alkyl; R 2 is OR’, OC(O)R 3 OC(O)OR 3 CO2R’, CON(R’)2, SO2N(R’)2, SO2R 3 OSO2R 3 or OSO2N(R’)2; R 3 It is C 1-6 Alkyl, C 3-6 Cycloalkyl or phenyl; R' is H, C 1-6 Alkyl or C 3-6 cycloalkyl; and n is 0, 1, 2, or 3; wherein at each occurrence, alkyl, alkylene, alkoxy and cycloalkyl are each optionally and independently substituted by OH, SH, CN, NO2, COOH, halogen or COOC 1-4 Up to 3 instances of substitution in the alkyl group; wherein at each occurrence, heterocycloalkyl, aryl and heteroaryl are each optionally and independently substituted by OR', SR', CN, NO2, CO2R', halogen, C 1-4 Up to 3 instances of alkyl or oxo are substituted.

2. The compound according to claim 1, wherein: (i) L' is a group and / or (ii) L' is a group 3. The compound according to claim 1 or 2, wherein the compound is a compound of formula (1) or a pharmaceutically acceptable salt, solvate, hydrate, tautomer or optical isomer thereof: Among them, A, X, R 1 , R 2 and n are the same as defined in claim 1.

4. The compound according to any one of claims 1 to 3, wherein A is selected from C(O)OR', C(O)N(R')S(O)2R 3 、S(O)2OR'、C 1-8 alkyl, heterocycloalkyl or heteroaryl, wherein the heterocycloalkyl and heteroaryl are each optionally and independently substituted by OR', SR', halogen, C 1-4 Up to 3 instances of alkyl or oxo are substituted.

5. The compound according to any one of claims 1 to 4, wherein A is selected from: Preferably, wherein A is: More preferably, wherein A is 6. A compound according to any one of claims 1 to 5, wherein X is halogen or OR', preferably wherein X is F or OH.

7. The compound according to claim 6, wherein X is F or OH, and n is 1 or 2.

8. The compound according to any one of claims 1 to 5, wherein n is 0.

9. A compound according to any one of claims 1 to 8, wherein R 1 is H, OH, halogen, CN, C optionally substituted by 1 to 3 fluorine atoms 1-6 alkoxy, or C optionally substituted by 1 to 3 fluorine atoms 1-6 Alkyl; preferably wherein R 1 is C optionally substituted by 1 to 3 fluorine atoms 1-6 alkyl.

10. The compound according to claim 9, wherein R 1 It's methyl.

11. A compound according to any one of claims 1 to 10, wherein R 2 is OR', CON(R')2, SO2N(R')2 or OSO2N(R')2, preferably wherein R 2 It is OH, CONH2, SO2NH2 or OSO2NH2.

12. A compound according to any one of claims 1 to 10, wherein R 2 It is CON(R')2, SO2N(R')2, OSO2R 3 or OSO2N(R')2; preferably wherein R 2 It is CONH2, SO2NH2 or OSO2NH2.

13. A compound according to any one of claims 1 to 12, which is a compound of formula (2a), formula (2b), formula (2c), or formula (2d), or a pharmaceutically acceptable salt, solvate, hydrate, or tautomer thereof: Among them, X, R 1 , R 2 and n are the same as defined in any of the preceding claims.

14. The compound according to claim 1, which is a compound of formula (5), formula (5a), formula (5b), formula (6), formula (6a), formula (6b) or a pharmaceutically acceptable salt, solvate, hydrate or tautomer thereof:

15. The compound according to claim 1, wherein the compound is selected from: or a pharmaceutically acceptable salt, solvate, hydrate, tautomer or optical isomer thereof.

16. A pharmaceutical composition comprising a compound, a pharmaceutically acceptable salt, a solvate, a hydrate or a tautomer according to any one of claims 1 to 15, and a pharmaceutically acceptable excipient.

17. The pharmaceutical composition of claim 16, wherein the composition further comprises at least one additional therapeutic agent selected from the group consisting of aminosalicylate, corticosteroid, immunomodulator, and combinations thereof.

18. A kit comprising a compound, pharmaceutically acceptable salt, solvate, hydrate, tautomer or optical isomer according to any one of claims 1 to 15, and at least one additional therapeutic agent selected from the group consisting of aminosalicylate, corticosteroid, immunomodulator and combinations thereof.

19. A compound according to any one of claims 1 to 15, a composition according to claim 16 or 17, or a kit according to claim 18 for use as a medicament.

20. A compound according to any one of claims 1 to 15, a composition according to claim 16 or 17, or a kit according to claim 18 for use in the treatment of an EP4 receptor mediated disease.

21. The compound, composition or kit for use according to claim 20, wherein the EP4 receptor-mediated disease is a gastrointestinal disorder.

22. A compound, composition or kit for use according to claim 21, wherein the gastrointestinal disorder is selected from constipation disorders, constipation-predominant irritable bowel syndrome, mixed irritable bowel syndrome, chronic idiopathic constipation, gastrointestinal symptoms associated with Parkinson's disease, gastrointestinal symptoms associated with cystic fibrosis, intestinal motility disorders, postoperative ileus, food allergy or food intolerance, celiac disease, gastrointestinal motility disorders, functional gastrointestinal disorders, drug-induced enteropathy, NSAID-induced gastric and intestinal damage, chemotherapy-induced mucositis, gastroesophageal reflux disease (GERD), duodenogastric reflux, diarrheal diseases, immune-mediated gastrointestinal diseases, Crohn's disease, ulcerative colitis, inflammatory bowel disease and ischemic colitis.

23. The compound, composition or kit for use according to claim 20, wherein the EP4 receptor mediated disease is a pulmonary disease or disorder.

24. A compound, composition or kit for use according to claim 23, wherein the pulmonary disease or condition is selected from chronic obstructive pulmonary disease, asthma, chronic bronchitis, cystic fibrosis, emphysema, chronic idiopathic cough, overactive airways disorder and idiopathic pulmonary fibrosis.

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