CN114213331B - URAT1 inhibitor and application thereof - Google Patents

URAT1 inhibitor and application thereof Download PDF

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CN114213331B
CN114213331B CN202210131754.5A CN202210131754A CN114213331B CN 114213331 B CN114213331 B CN 114213331B CN 202210131754 A CN202210131754 A CN 202210131754A CN 114213331 B CN114213331 B CN 114213331B
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compound
ethyl
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acid
pharmaceutically acceptable
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CN114213331A (en
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史东方
周禾
朱江华
李鹏飞
承曦
杨艳
顾杰
吴帆
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Jiangsu Atom Bioscience and Pharmaceutical Co Ltd
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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D231/00—Heterocyclic compounds containing 1,2-diazole or hydrogenated 1,2-diazole rings
    • C07D231/54—Heterocyclic compounds containing 1,2-diazole or hydrogenated 1,2-diazole rings condensed with carbocyclic rings or ring systems
    • C07D231/56—Benzopyrazoles; Hydrogenated benzopyrazoles
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D471/00—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, at least one ring being a six-membered ring with one nitrogen atom, not provided for by groups C07D451/00 - C07D463/00
    • C07D471/02—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, at least one ring being a six-membered ring with one nitrogen atom, not provided for by groups C07D451/00 - C07D463/00 in which the condensed system contains two hetero rings
    • C07D471/04—Ortho-condensed systems

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Abstract

The invention belongs to the field of pharmaceutical chemistry, and particularly relates to a URAT1 inhibitor and application thereof, wherein the URAT1 inhibitor is a compound with a structure shown in a formula (I) or a formula (II) or pharmaceutically acceptable salt thereof. Experiments show that the compound provided by the invention has very good inhibition effect on the transfer of uric acid by hURAT1 in HEK293 transfected cells, and shows that the compound has good application prospect in the aspect of treating hyperuricemia or gout.

Description

URAT1 inhibitor and application thereof
Technical Field
The invention belongs to the field of pharmaceutical chemistry, and particularly relates to a URAT1 inhibitor compound and application of the compound.
Background
Gout is a metabolic disease caused by hyperuricemia. When purine metabolism is disturbed or excessive high purine foods are ingested in human body, uric acid production is increased, and uric acid excretion by kidneys is not smooth, so that blood uric acid concentration exceeds normal level, saturation is achieved, uric acid salt is accumulated in a large amount and is crystallized, and the uric acid salt is deposited on joints, tendons, kidneys and other parts, and finally, repeated arthritic pain is caused (RICHETTE P, bardin T.gout.Lancet.2010,375 (9711): 318-328). Hyperuricemia is clinically defined when uric acid concentration in the blood of men is >7mg/dL or uric acid concentration in the blood of women is greater than 6 mg/dL. About 80-85% of patients with hyperuricemia have a etiology of poor uric acid excretion by the kidneys (Cheeseman C.Solute carrier family 2,member 9 and uric acid homeostasis.Current Opinion in Nephrology and Hypertension,2009,18(5):428-432).
Gout has serious harm. The patient will be acutely painful during the onset. If a large amount of sodium urate crystals are deposited subcutaneously in humans, a tophus can be formed, which can deform joints, lose functions and break epidermal cells. Tophus formed in the kidneys can lead to uric acid lithiasis and nephritis, and even to kidney damage. Gout may also have a certain interaction with various diseases such as hypertension, metabolic syndrome, hyperlipidemia, diabetes, insulin resistance and the like, and exacerbate the risk coefficient of the diseases .(Rho YH,Woo JH,Choi SJ,et al.Association between serum uric acid and the adult treatment panel III-defined metabolic syndrome:results from a single hospital database.Metabolism.2008,57:(1)71-76).
With the increasing level of human life, the trend of increasing diet and lifestyle, gout incidence has been the most common chronic disease, and has been listed by the united nations as one of the 20 biggest obsessions of the 21 st century (Grobner W, walter si treatent of hyperuricemia and gout. Med Monatsschr Pharm,2005, 28:159-164). Currently, there are nearly 1 hundred million gout patients worldwide, and the market is huge. Gout incidence in europe is about 1-2%, wherein the main incidence group is middle-aged and elderly men (Michael Doherty,Tim L Jansen,George Nuki,et al.Gout:why is this curable disease so seldom cured?.Annals of the Rheumatic Diseases.2012,71(11):1765-1770);, and the number of gout patients in the united states is 830 ten thousand; the number of patients suffering from hyperuricemia in China is about 1.2 hundred million, wherein patients suffering from gout are over 5000 ten thousand, and the incidence rate of gout and hyperuricemia in developed coastal areas (such as Qingdao, hong Kong in China, taiwan in China and the like) is high. According to statistics of certain hospitals in Taiwan of China in 2000-2007, the incidence rate of hyperuricemia is up to 22.8%.
Standardized treatment regimens for gout include uric acid lowering therapy (urate-lower therapy, ULT). It can make the concentration of body blood uric acid lower than saturation concentration without forming urate crystal, and can make urate crystal at focus position dissolve. After the crystallization of the uric acid salt disappears in vivo, gout is no longer formed. The purpose of the ULT is to reduce the patient's blood uric acid concentration below 6 mg/dL. For tophus patients, the target blood uric acid concentration needs to be reduced to 5mg/dL, so that the medicine for effectively reducing the urate sediment (Puja P Khanna,George Nuki,Thomas Bardin,et al.Tophi and frequent gout flares are associated with impairments to quality of life,productivity,and increased healthcare resource use:Results from a cross-sectional survey.Health and Quality of Life Outcomes.2012,10(117):117;Richette P,Frazier A,Bardin T.Pharmacokinetics considerations for gout treatments.Expert Opin Drug Metab Toxicol.2014,10(7):949-957).ULT of the patient can be divided into uric acid generation inhibiting medicines and uric acid excretion promoting medicines and uricase medicines within a certain time range. Allopurinol and febuxostat are xanthine oxidase inhibitors, are main uric acid generation inhibitors, but allopurinol is clinically used in large dosage and has various adverse reactions, such as skin rash which can cause fatal; febuxostat has very serious cardiovascular and gastrointestinal discomfort side effects and also has hepatotoxicity. Moreover, about 40-80% of patients in clinic cannot achieve the aim of controlling blood uric acid level through uric acid generation inhibiting drugs, such as polyethylene glycol recombinant uricase (pegloticase), and although the therapeutic effect is remarkable, the medicament can only be used for adult gout patients who are not effective in conventional treatment or cannot tolerate conventional treatment because the medicament has serious adverse reactions such as cardiovascular events, transfusion reactions, immunogenic reactions and the like, so the medicament is rarely used in clinic .(Lipsky PE,Calabrese LH,Kavanaugh A,et al.Pegloticase Immuneogenicity:the relationship between efficacy and antibody development in patients treated for refractory chronic gout.Arthritis Research Therapy.2014,16(2):60.)
Another ULT class of drugs is that which promote uric acid excretion. The mechanism of action is to inhibit the transfer effect of human uric acid anion transporter 1 (human urate anion transporter 1, hURAT1) of kidney proximal tubular epithelial cells on uric acid, reduce the reabsorption of uric acid in kidney proximal tubular, and thus promote the excretion of uric acid by kidney. hURAT1 is specifically expressed on brush-like border membrane of human kidney proximal tubular epithelial cells, is the most main uric acid reabsorption protein in human body, and controls the reabsorption of uric acid after glomerular filtration by more than about 90% (Michael FW,Jutabha P,Quada B.Developing potent human uric acid transporter 1(hURAT1)inhibitors.Journal of Medicinal Chemistry.2011,54:2701-2713).
The uric acid excretion-promoting drugs URAT1 inhibitors which are mainly used for treating gout in clinic at present comprise tribromouron (Benzbromarone), zurampic, probenecid and benzenesulfonyl. Zurampic of aslicon was approved by the united states and europe at 200 mg/day for use with allopurinol at 2015, 12 and 2016, 2, and was far less effective than benzbromarone. Clinical trials show that Lesinurad has a plurality of toxic and side effects: (1) The composition may cause serious cardiovascular adverse reaction such as fatal cardiovascular diseases, non-fatal myocardial infarction or brain pruritus. (2) Adverse reactions related to renal function can occur immediately after Lesinurad starts treatment, and when 400mg is taken alone, the occurrence rate of serious adverse events is highest, so that high-dose single use is forbidden clinically, and the renal function needs to be detected periodically before and after treatment. (3) the medicine can cause light and medium liver injury. Thus, the FDA requires Lesinurad to mark its severe renal toxicity in the specification with a black box. The traditional Chinese medicine probenecid and the benzenesulfonyl oxazolone have very poor curative effect, large dosage and large side effect.
Tribromone is a typical class of URAT1 selective inhibitors, the most potent uric acid excretion-promoting agent in the market at present, developed by the company Snaofi-synthetic in france and marketed in 1976. 92% of patients with gout who are not effective in allopurinol treatment are reported to have the chemical property that after the continuous use of the tribromoclone for 2 months, the serum uric acid value of the tribromoclone can be reduced to 5mg/dl(Halevy S,Ghislain PD,Mockenhaupt M,et al.Allopurinol is the most common cause of Stevens-Johnson syndrome and toxic epidermal necrolysis in Europe and Israel.Journal of the American Academy of Dermatology.2008,58(1):25-32)., the tribromoclone is easily oxidized and metabolized into 6-hydroxy tribromoclone by CYP2C9, and further metabolized into a6, 7-or 5, 6-o-benzobisquinone product (Matthew G.McDonald,Rettie AE.Sequential metabolism and bioactivation of the hepatotoxin benzbromarone:formation of glutathione adducts from a catechol intermediate.Chemical Research in Toxicology.2007,20(12):1833-1842);, and the tribromoclone is oxidized and generates 2, 6-dibromohydroquinone by CYP2C9 through ipso-substitution reaction due to the fact that the tribromo is metabolized into a p-benzobisquinone product (Yumina Kitagawara,Tomoyuki Ohe,Kumiko Tachibana.Novel Bioactivation Pathway of Benzbromarone Mediated by Cytochrome P450.ASPET Journal.2015)., and the chemical property of the tribromo is active by conjugation and addition with sulfhydryl on cysteine residues of proteins or polypeptides, so that hepatotoxicity can be caused. In addition, the benzbromarone has strong inhibition on CYP2C 9. The medicine cannot enter the U.S. market, is returned to (Jansen TL,Reinders MK,van Roon EN,et al.Benzbromarone withdrawn from the European market:another case of"absence of evidence is evidence of absence".Clinical Experimental Rheumatology,2004,22(5):651). from part of European countries in 2003, but is still widely used in more than 20 countries such as China, germany, japan, brazil, new Zealand and the like due to the lack of good anti-gout medicines in the market.
Disclosure of Invention
The invention aims to provide a series of novel compounds based on the prior art, and aims to obtain URAT1 inhibitors with low toxicity and better drug effect for treating hyperuricemia or gout diseases.
The aim of the invention can be achieved by the following measures:
A compound with a structure shown as a formula (I) or a formula (II) or pharmaceutically acceptable salts thereof,
Wherein,
A 1、A2、A3 or A 4 is CH or N;
g is carbonyl, sulfur, sulfonyl, sulfoxide, or optionally substituted methylene or imino;
R 1 is selected from one or more of hydrogen, deuterium, halogen, cyano, hydroxyl, nitro, amino, carboxyl, substituted amino or substituted or unsubstituted groups: c 1-5 alkyl, C 1-5 alkoxy or C 1-5 alkylthio;
R 2 is selected from one or more of hydrogen, deuterium, halogen, cyano, hydroxy, nitro, substituted amino, C 2-3 alkenyl, C 2-3 alkynyl or substituted or unsubstituted groups: c 1-4 alkyl, C 1-5 alkoxy or C 1-5 alkylthio;
R 3 is selected from the following groups, substituted or unsubstituted: c 1-4 alkyl or C 3-4 cycloalkyl, the substituents of which are selected from deuterium, halogen, C 1-2 alkyl or C 3-4 cycloalkyl;
m is an integer of 0 to 3;
n is an integer of 1 to 3;
The substituent in group G is selected from hydroxy, cyano, nitro, amino, carboxy or C 1-3 alkoxy, and the substituent in R 1 or R 2 is selected from deuterium, halogen, cyano, hydroxy, nitro, amino, C 1-3 alkyl, C 3-4 cycloalkyl or C 1-3 alkoxy.
When m is 2 or 3 in the present application, it means that the compound contains two R 1 groups, and the two R 1 groups may be the same or different groups defined by R 1 in the present application may be used respectively. When the R 1 group is attached to one or more of A 1、A2、A3 and A 4 ring atoms, and when the ring atom is CH, the group at the ring atom is C-R 1.
When n in the present application is 2 or 3, it means that the compound contains two R 2 groups, and these two R 2 groups may be the same or different groups defined as R 2 in the present application may be used respectively.
The six-membered ring containing A 1、A2、A3、A4 constitutes an aromatic ring, preferably, the aromatic ring is a benzene ring, a pyridazine ring, a pyrimidine ring, a pyrazine ring or a pyridine ring.
In a preferred embodiment, a 1 is CH, a 2 is CH or N, a 3 or a 4 is CH.
In another preferred embodiment, a 1 and a 4 are CH, and a 2 and a 3 are each independently CH or N.
In one embodiment, R 1 is selected from one or more of hydrogen, deuterium, fluorine, chlorine, bromine, cyano, hydroxy, C 1-3 alkyl, C 1-3 haloalkyl, C 1-3 deuteroalkyl, C 1-3 alkoxy, or C 1-3 deuteroalkoxy; m is 0, 1,2 or 3.
In a preferred embodiment, R 1 is selected from one or more of hydrogen, deuterium, fluorine, chlorine, bromine, cyano, methyl, ethyl, methoxy, deuterated methoxy, ethoxy; m is 0, 1,2 or 3.
In one embodiment, R 2 is selected from one or more of hydrogen, deuterium, halogen, cyano, nitro, vinyl, ethynyl, C 1-2 alkyl, substituted C 1-2 alkyl, C 1-2 alkoxy, substituted C 1-2 alkoxy, C 1-2 alkylthio, substituted C 1-2 alkylthio; the substituent is selected from deuterium, halogen, C 1-2 alkyl, C 3-4 cycloalkyl or C 1-3 alkoxy; n is 1 or 2.
In a preferred embodiment, R 2 is selected from one or more of hydrogen, deuterium, halogen, cyano, C 1-2 alkyl, C 1-2 haloalkyl, C 1-2 alkoxy, or C 1-2 alkylthio; n is 1 or 2.
In a more preferred embodiment, R 2 is selected from one or more of bromine, chlorine, cyano; n is 1 or 2.
In one embodiment, R 3 is selected from methyl, ethyl, n-propyl, isopropyl, cyclopropyl, or cyclobutyl.
In a more preferred embodiment, the compound of the invention, or a pharmaceutically acceptable salt thereof, wherein the compound is selected from the group consisting of:
The preparation method of the compound with the structure shown in the formula (I) or the formula (II) comprises the following steps:
The substituted indazole (or pyridopyrazole) is reacted with a halide to give the corresponding 1-substituted compound (I-A) or 2-substituted compound (II-A). The product obtained by the reaction of the I-A and the aryl aldehyde is subjected to oxidation reaction and hydroxyl deprotection reaction to obtain a corresponding hydroxyl compound, wherein the compound can be a final product, and the corresponding target product (I) can also be obtained through halogenation reaction, reduction reaction or other reactions. The II-A obtained above is reacted with acyl chloride, and then hydroxy deprotection reaction is carried out to obtain corresponding hydroxy compound, wherein the compound can be a final product, and the corresponding target product (II) can be obtained through halogenation reaction, reduction reaction or other reactions. A1, A2, A3, A4, R 1、R2 and R 3 are defined as the general formula in the claims.
Unless otherwise indicated, the following terms used in the claims and specification have the following meanings:
"six-membered aromatic ring" refers to a fused ring group having a conjugated planar ring structure composed of six ring atoms, which is aromatic and the ring atoms may be other atoms than carbon atoms, i.e., heteroatoms. When a heteroatom is contained in a six-membered aromatic ring, the heteroatom may be N, S or O, and the number of heteroatoms is not limited to one, and may be two, three, or the like. Six-membered aromatic rings containing heteroatoms in the present invention include, but are not limited to, pyridine rings, pyrimidine rings, pyrazine rings, and the like.
"Hydrogen" refers to protium (1H), which is the primary stable isotope of hydrogen.
"Deuterium" refers to a stable form isotope of hydrogen, also known as deuterium, with the elemental symbol D.
"Halogen" means a fluorine atom, a chlorine atom, a bromine atom or an iodine atom.
"Alkyl" means a saturated aliphatic radical of 1 to 20 carbon atoms and includes straight and branched chain groups (the numerical ranges mentioned herein, e.g., "1 to 20", refer to such groups, which in this case are alkyl groups, which may contain 1 carbon atom, 2 carbon atoms, 3 carbon atoms, etc., up to and including 20 carbon atoms). Alkyl groups containing 1 to 4 carbon atoms are referred to as lower alkyl groups. When the lower alkyl group has no substituent, it is referred to as an unsubstituted lower alkyl group. More preferably, the alkyl group is a medium size alkyl group having 2 to 5 carbon atoms. Alkyl groups in the present application are, for example, methyl, ethyl, propyl, 2-propyl, n-butyl, isobutyl, t-butyl, pentyl and the like. Preferably, the alkyl group is a lower alkyl group having 1 to 4 carbon atoms, such as methyl, ethyl, propyl, 2-propyl, n-butyl, isobutyl, tert-butyl, or the like. Alkyl groups may be substituted or unsubstituted.
"Alkoxy" means both-O- (unsubstituted alkyl) and-O- (unsubstituted cycloalkyl) groups, it further represents-O- (unsubstituted alkyl). Representative examples include, but are not limited to, methoxy, ethoxy, propoxy, butoxy, cyclopropoxy, cyclobutoxy, cyclopentoxy, cyclohexyloxy, and the like.
"Carbonyl" means a c=o group.
"Sulfone" means the-S (O) 2 -group.
"Sulfoxide" means an-S (O) -group.
"Methylene" means the-CH 2 -group.
"Imino" means an-NH-group.
"Hydroxy" means an-OH group.
"Nitro" means a-NO 2 group.
"Amino" means the-NH 2 group.
"Carboxy" means a-COOH group.
"Cyano" means a-CN group.
"Pharmaceutically acceptable salts" are salts comprising the compounds of formula (I) with organic or inorganic acids, meaning those salts which retain the biological effectiveness and properties of the parent compound. Such salts include:
(1) Salified with acids, obtained by reaction of the free base of the parent compound with an inorganic acid such as, but not limited to, hydrochloric acid, hydrobromic acid, nitric acid, phosphoric acid, metaphosphoric acid, sulfuric acid, sulfurous acid, perchloric acid, and the like, or an organic acid such as, but not limited to, acetic acid, propionic acid, acrylic acid, oxalic acid, (D) or (L) malic acid, fumaric acid, maleic acid, hydroxybenzoic acid, gamma-hydroxybutyric acid, methoxybenzoic acid, phthalic acid, methanesulfonic acid, ethanesulfonic acid, naphthalene-1-sulfonic acid, naphthalene-2-sulfonic acid, p-toluenesulfonic acid, salicylic acid, tartaric acid, citric acid, lactic acid, mandelic acid, succinic acid, malonic acid, and the like.
(2) The acidic protons present in the parent compound are replaced by metal ions, such as alkali metal ions, alkaline earth metal ions or aluminum ions, or salts formed by complexation with organic bases, such as ethanolamine, diethanolamine, triethanolamine, tromethamine, N-methylglucamine, and the like.
"Pharmaceutical composition" refers to a mixture of one or more compounds described herein or pharmaceutically acceptable salts and prodrugs thereof with other chemical components, such as pharmaceutically acceptable carriers and excipients. The purpose of the pharmaceutical composition is to facilitate the administration of the compound to the organism.
The invention comprises a pharmaceutical composition which comprises any one of the compounds, pharmaceutically acceptable salts or easily hydrolyzed prodrug esters thereof as an active ingredient, or further comprises other compounds with pharmacodynamic activity as one of the active ingredients and is assisted by pharmaceutically acceptable auxiliary materials.
The compounds or the pharmaceutically acceptable salts thereof can be applied to the preparation of uric acid excretion promoting medicaments, in particular to the preparation of medicaments for treating or preventing hyperuricemia, nephropathy or gout. Experiments show that the compound provided by the invention has very good inhibition effect on the transfer of uric acid by hURAT1 in HEK293 transfected cells, and shows that the compound has good application prospect in the aspect of treating hyperuricemia or gout.
Detailed Description
The present invention will be further described with reference to examples, but the scope of the present invention is not limited to the examples.
Example 1: synthesis of (3, 5-dibromo-4-hydroxyphenyl) (2-ethyl-2H-indazol-3-yl) methanone (6)
Step A: a mixture containing indazole (5.00 g,42.3 mmol), iodoethane (13.2 g,90.3 mmol), potassium hydroxide (5.50 g,98.0 mmol) and ethanol (60 mL) was stirred at 65℃for 5 hours. Cooled to room temperature and insoluble materials were removed by filtration. The solvent was distilled off under reduced pressure, followed by addition of water (30 mL), extraction with methylene chloride (20 mL. Times.3) and drying over anhydrous sodium sulfate. The solvent was distilled off under reduced pressure, and the product was purified by column chromatography (200-300 mesh silica gel, ethyl acetate: petroleum ether=1:20-1:5 elution) to give 2-ethyl-2H-indazole (1) (1.56 g) and 1-ethyl-1H-indazole (2) (3.06 g). The yields were 25.2% and 49.5%, respectively. Compound 1:1H NMR(DMSO-d6,400MHz)δ8.37(s,1H),7.69(d,J＝8.4Hz,1H),7.60(d,J＝8.4Hz,1H),7.24-7.20(m,1H),7.05-7.01(m,1H),4.46(q,J＝7.2Hz,2H),1.51(t,J＝7.2Hz,3H). Compound 2:1H NMR(DMSO-d6,400MHz)δ8.05(s,1H),7.76(d,J＝8.0Hz,1H),7.67(dd,J＝0.8,8.4Hz,1H),7.41-7.36(m,1H),7.15-7.11(m,1H),4.45(q,J＝7.2Hz,2H),1.40(t,J＝7.2Hz,3H).
And (B) step (B): to a solution of compound 1 (480 mg,3.28 mmol) and diisopropylamine (432 mg,4.27 mmol) in anhydrous THF (10 mL) at-70-80℃was added dropwise a 2.5M n-hexane solution of n-butyllithium (1.7 mL,4.25 mmol). After the addition, stirring was continued for 10 minutes at this temperature, and then slowly warmed to-10 to-20 ℃ over about 20 minutes, and stirring was continued for 15 minutes at this temperature. Cooled again to-70-80℃and 4-methoxybenzaldehyde (514 mg,3.77 mmol) was added via syringe. After stirring for about 15 minutes, naturally warm to room temperature and continue stirring overnight. Water (25 mL) was added, extracted with ethyl acetate (20 mL. Times.3), and the combined organic phases were washed with saturated brine (15 mL) and dried over anhydrous sodium sulfate. The solvent was distilled off under reduced pressure, and the product was purified by column chromatography (200-300 mesh silica gel, ethyl acetate: petroleum ether=1:20-1:3 elution) to give (2-ethyl-2H-indazol-3-yl) (4-methoxyphenyl) methanol (3) (536 mg). Yield is as follows 57.9％.1H NMR(DMSO-d6,400MHz)δ7.52(d,J＝8.4Hz,1H),7.42(d,J＝8.4Hz,1H),7.30(d,J＝8.4Hz,2H),7.19-7.15(m,1H),6.93-6.89(m,3H),6.35(d,J＝4.4Hz,1H),6.30(d,J＝4.4Hz,1H),4.40(q,J＝7.2Hz,2H),3.73(s,3H),1.30(t,J＝7.2Hz,3H).
Step C: to a solution of compound 3 (320 mg,1.13 mmol) in DMSO (10 mL) was added 2-iodoxybenzoic acid (416 mg,1.49 mmol) and the resulting mixture was stirred at room temperature overnight. Water (40 mL) was added, extracted with ethyl acetate (30 mL. Times.3), and the combined organic phases were washed with water (20 mL. Times.2) and saturated brine (15 mL) and dried over anhydrous sodium sulfate. The solvent was distilled off under reduced pressure to give (2-ethyl-2H-indazol-3-yl) (4-methoxyphenyl) methanone (4) (275 mg). The yield thereof was found to be 86.8%.
Step D: a mixture containing compound 4 (267 mg,0.952 mmol), 1M boron tribromide in toluene (2.9 mL) and methylene chloride (15 mL) was stirred at room temperature overnight. The reaction was slowly poured into crushed ice (40 g), the pH was adjusted to 6 to 7 with 2M aqueous sodium hydroxide solution, extracted with methylene chloride (20 mL. Times.3), and the combined organic phases were washed successively with saturated aqueous sodium bicarbonate (10 mL) and saturated brine (10 mL), and dried over anhydrous sodium sulfate. The solvent was distilled off under reduced pressure to give (2-ethyl-2H-indazol-3-yl) (4-hydroxyphenyl) methanone (5) (200 mg). The yield thereof was found to be 78.9%.
Step E: a solution of bromine (257 mg,1.61 mmol) in acetic acid (5 mL) was added dropwise to a solution of compound 5 (195 mg, 0.730 mmol) and sodium acetate (180 mg,2.19 mmol) in acetic acid (10 mL), and after the addition was complete, the resulting mixture was stirred at room temperature for 1 hour. The reaction was quenched with aqueous sodium bisulfite and then most of the solvent was distilled off under reduced pressure. Water (20 mL) was added, insoluble matter was separated out, and the mixture was filtered, and the cake was recrystallized from petroleum ether/ethyl acetate to give (3, 5-dibromo-4-hydroxyphenyl) (2-ethyl-2H-indazol-3-yl) methanone (6) (236 mg). Yield is as follows 76.0％.1H NMR(DMSO-d6,400MHz)δ7.95(s,2H),7.82(d,J＝8.8Hz,1H),7.38-7.34(m,1H),7.22-7.18(m,1H),7.12(d,J＝8.4Hz,1H),4.69(q,J＝7.2Hz,2H),1.54(t,J＝7.2Hz,3H).MS(EI,m/z)：422.9[M-H]-.
Example 2: synthesis of 2, 6-dibromo-4- [ (2-ethyl-2H-indazol-3-yl) hydroxymethyl ] phenol (7)
To a solution of compound 6 (100 mg,0.236 mmol) in methanol (10 mL) was added sodium borohydride (90 mg,2.38 mmol), and after the addition, the resulting mixture was stirred under reflux for 30 minutes, sodium borohydride (90 mg,2.38 mmol) was added, and stirring was continued under reflux for 1 hour. Water (20 mL) was added, the pH was adjusted to 6-7 with 2M aqueous citric acid, extracted with ethyl acetate (20 mL. Times.3), and the combined organic phases were washed with water (20 mL) and dried over anhydrous sodium sulfate. The solvent is distilled off under reduced pressure, and the product is purified by column chromatography (200-300 meshes of silica gel, ethyl acetate: petroleum ether=1:20-1:3 elution) to obtain 2, 6-dibromo-4- [ (2-ethyl-2H-indazol-3-yl) hydroxymethyl ] phenol (7).1H NMR(DMSO-d6,400MHz)δ9.99(s,1H),7.55(d,J＝8.8Hz,1H),7.51(s,2H),7.38(d,J＝8.8Hz,1H),7.21-7.17(m,1H),6.97-6.93(m,1H),6.51(d,J＝4.4Hz,1H),6.35(d,J＝4.4Hz,1H),4.43(q,J＝7.2Hz,2H),1.35(t,J＝7.2Hz,3H).MS(EI,m/z)：427.0[M+H]+.
Example 3: synthesis of (3, 5-dibromo-4-hydroxyphenyl) (1-ethyl-1H-indazol-3-yl) methanone (10)
Step A: a mixture containing compound 2 (2.59 g,17.7 mmol), p-methoxybenzoyl chloride (3.02 g,17.7 mmol) and anhydrous aluminum trichloride (3.54 g,26.6 mmol) was stirred overnight at 105 ℃. Cooled to room temperature, water (30 mL) and ethyl acetate (30 mL) were added and stirred for about 5 minutes. The layers were separated, the aqueous layer was extracted with ethyl acetate (10 mL. Times.3), and the combined organic phases were washed with water (15 mL) and dried over anhydrous sodium sulfate. The solvent was distilled off under reduced pressure, and the product was purified by column chromatography (200-300 mesh silica gel, ethyl acetate: petroleum ether=1:25-1:15 elution) to give (1-ethyl-1H-indazol-3-yl) (4-methoxyphenyl) methanone (8) (990 mg). Yield is as follows 21.1％.1H NMR(DMSO-d6,400MHz)δ8.34-8.29(m,3H),7.88(d,J＝8.4Hz,1H),7.56-7.52(m,1H),7.41-7.37(m,1H),7.15-7.11(m,2H),4.63(q,J＝7.2Hz,2H),3.89(s,3H),1.51(t,J＝7.2Hz,3H).
And (B) step (B): a mixture containing compound 8 (200 mg, 0.514 mmol), a 1M solution of boron tribromide in toluene (2.3 mL) and methylene chloride (5 mL) was stirred at room temperature overnight. The reaction was slowly poured into crushed ice (40 g), the pH was adjusted to 6 to 7 with 2M aqueous sodium hydroxide solution, extracted with methylene chloride (20 mL. Times.3), and the combined organic phases were washed successively with saturated aqueous sodium bicarbonate (10 mL) and saturated brine (10 mL), and dried over anhydrous sodium sulfate. The solvent was distilled off under reduced pressure to give (1-ethyl-1H-indazol-3-yl) (4-hydroxyphenyl) methanone (9) (131 mg). The yield thereof was found to be 68.9%.
Step C: a solution of bromine (236 mg,1.48 mmol) in acetic acid (2 mL) was added dropwise to a solution of compound 9 (106 mg, 0.390 mmol) and sodium acetate (109 mg,1.33 mmol) in acetic acid (3 mL), and the resulting mixture was stirred overnight at room temperature. The reaction was quenched with aqueous sodium bisulfite and then most of the solvent was distilled off under reduced pressure. Water (20 mL) was added, the pH was adjusted to 7-8 with saturated aqueous sodium bicarbonate, extracted with ethyl acetate (20 mL. Times.2), and the combined organic phases were washed with water (10 mL) and dried over anhydrous sodium sulfate. The solvent was distilled off under reduced pressure to give (3, 5-dibromo-4-hydroxyphenyl) (1-ethyl-1H-indazol-3-yl) methanone (10) (70 mg). Yield is as follows 41.4％.1H NMR(DMSO-d6,400MHz)δ8.53(s,2H),8.28(d,J＝8.4Hz,1H),7.89(d,J＝8.4Hz,1H),7.54(d,J＝7.6Hz,1H),7.39(d,J＝7.6Hz,1H),4.63(q,J＝7.2Hz,2H),1.52(t,J＝7.2Hz,3H).MS(EI,m/z)：422.9[M-H]-.
Example 4: synthesis of 3- (3, 5-dibromo-4-hydroxybenzoyl) -1-ethyl-1H-indazole-5-carbonitrile (14)
Step A: select fluorine (695 mg,1.96 mmol) was added to a mixture containing compound 8 (550 mg,1.96 mmol), iodine (59 mg,1.02 mmol) and acetonitrile (10 mL) under an ice-water bath, and after the addition was completed, the resulting mixture was stirred at room temperature overnight. Water (30 mL) was added and 2M aqueous sodium thiosulfate was added dropwise until the color disappeared. The mixture was extracted with ethyl acetate (20 mL. Times.3), and the combined organic phases were washed with water (20 mL) and saturated brine (20 mL) and dried over anhydrous sodium sulfate. The solvent was distilled off under reduced pressure, and the product was purified by column chromatography (200-300 mesh silica gel, dichloromethane: petroleum ether=1:3 elution) to give (1-ethyl-5-iodo-1H-indazol-3-yl) (4-methoxyphenyl) methanone (11) (290 mg). Yield is as follows 36.4％.1H NMR(DMSO-d6,400MHz)δ8.67(s,1H),8.35(d,J＝8.8Hz,2H),7.82-7.76(m,2H),7.13(d,J＝8.8Hz,2H),4.61(q,J＝7.2Hz,2H),3.89(s,3H),1.49(t,J＝7.2Hz,3H).
And (B) step (B): a mixture containing compound 11 (190 mg, 0.463 mmol), cuprous cyanide (187 mg,2.09 mmol) and DMF (4 mL) was stirred overnight at 130 ℃. Cooled to room temperature, ethyl acetate (20 mL) was added, and insoluble matter was removed by filtration through celite. Water (20 mL) was added, the layers separated, the aqueous layer extracted with ethyl acetate (20 mL. Times.3), and the combined organic phases washed successively with water (20 mL. Times.2) and saturated brine (20 mL), dried over anhydrous sodium sulfate. The solvent was distilled off under reduced pressure to give crude 1-ethyl-3- (4-methoxybenzoyl) -1H-indazole-5-carbonitrile (12) (150 mg). The compound was used in the next reaction without purification .1H NMR(DMSO-d6,400MHz)δ8.72(s,1H),8.37(d,J＝8.8Hz,2H),8.13(d,J＝8.8Hz,1H),7.90(d,J＝8.8Hz,1H),7.15(d,J＝8.8Hz,2H),4.67(q,J＝7.2Hz,2H),3.90(s,3H),1.51(t,J＝7.2Hz,3H).
Step C: 1M boron tribromide in toluene (2 mL) was added dropwise to a solution of compound 12 (150 mg) in crude methylene chloride (5 mL) under an ice-water bath, and after the addition was completed, the resulting mixture was stirred at room temperature overnight. The reaction was slowly poured into ice water (20 mL), the pH was adjusted to 6-7 with 2M aqueous sodium hydroxide solution, extracted with methylene chloride (20 mL. Times.3), and the combined organic phases were washed successively with saturated aqueous sodium bicarbonate (10 mL) and saturated brine (10 mL), and dried over anhydrous sodium sulfate. The solvent was distilled off under reduced pressure, and the product was purified by column chromatography (200-300 mesh silica gel, ethyl acetate: petroleum ether=1:20-1:3 elution) to give 1-ethyl-3- (4-hydroxybenzoyl) -1H-indazole-5-carbonitrile (13) (72 mg). The total yield of the two reactions in steps B and C was 52.5%.
Step D: NBS (97 mg,0.545 mmol) was added to a solution of compound 13 (72 mg,0.247 mmol) in DMF (3 mL), and the resulting mixture was stirred at room temperature for 1.5 hours. Adding water (15 mL), filtering, rinsing the filter cake with a small amount of water, and recrystallizing the obtained solid with ethyl acetate to obtain 3- (3, 5-dibromo-4-hydroxybenzoyl) -1-ethyl-1H-indazole-5-carbonitrile (14).1H NMR(DMSO-d6,400MHz)δ8.71(s,1H),8.54(s,2H),8.15(d,J＝8.8Hz,1H),7.92(d,J＝8.8Hz,1H),4.69(q,J＝7.2Hz,2H),1.53(t,J＝7.2Hz,3H).MS(EI,m/z)：447.9[M-H]-.
Example 5: synthesis of 2, 6-dibromo-4- [ (2-ethyl-2H-indazol-3-yl) hydroxymethyl ] phenol (15)
A mixture containing compound 10 (150 mg,0.354 mmol), sodium borohydride (120 mg,3.17 mmol) and THF (5 mL) was stirred overnight at room temperature. Water (20 mL) was added, extracted with ethyl acetate (20 mL. Times.2), and the combined organic phases were washed with water (10 mL) and dried over anhydrous sodium sulfate. The solvent was distilled off under reduced pressure to give 2, 6-dibromo-4- [ (2-ethyl-2H-indazol-3-yl) hydroxymethyl ] phenol (15) (119 mg). Yield is as follows 78.9％.1H NMR(DMSO-d6,400MHz)δ9.84(s,1H),7.71(d,J＝8.4Hz,1H),7.61(d,J＝8.4Hz,1H),7.56(s,2H),7.37-7.33(m,1H),7.10-7.06(m,1H),6.23(d,J＝4.4Hz,1H),6.02(d,J＝4.4Hz,1H),4.40(q,J＝7.2Hz,2H),1.37(t,J＝7.2Hz,3H).MS(EI,m/z)：426.9[M+H]+.
Example 6: synthesis of (3-bromo-4-hydroxy-5-methylphenyl) (2-isopropyl-2H-indazol-3-yl) methanone (21)
Experimental procedures for Steps A, B, C and D the preparation method of steps A, B, C and D in example 1 was followed, wherein the ethyl iodide in step A of example 1 was replaced with bromoisopropyl alcohol and the 4-methoxybenzaldehyde in step B of example 1 was replaced with 3-methyl-4-methoxybenzaldehyde. Compounds of formula (I) 20:1H NMR(DMSO-d6,400MHz)δ10.57(s,1H),7.81-7.79(m,1H),7.64(s,1H),7.56-7.53(m,1H),7.34-7.30(m,1H),7.14-7.08(m,2H),6.95-6.93(m,1H),5.30-5.26(m,1H),2.17(s,3H),1.58(d,J＝6.4Hz,6H).
Step E: NBS (69 mg, 0.3838 mmol) was added to a solution of compound 20 (95 mg,0.323 mmol) in DMF (10 mL), and after completion the resulting mixture was stirred at room temperature for 1 hour. Water (40 mL) was added, extracted with ethyl acetate (20 mL. Times.3), and the combined organic phases were washed with water (10 mL. Times.2) and saturated brine (10 mL) and dried over anhydrous sodium sulfate. The solvent was distilled off under reduced pressure, and the product was purified by column chromatography (200-300 mesh silica gel, ethyl acetate: petroleum ether=1:5 elution) to give (3-bromo-4-hydroxy-5-methylphenyl) (2-isopropyl-2H-indazol-3-yl) methanone (21).1H NMR(DMSO-d6,400MHz)δ10.26(s,1H),7.84-7.80(m,2H),7.63(s,1H),7.35-7.32(m,1H),7.17-7.14(m,1H),7.08-7.06(m,1H),5.34-5.28(m,1H),2.29(s,3H),1.59(d,J＝6.4Hz,6H).MS(EI,m/z)：373.1[M+H]+.
Example 7: synthesis of 2-bromo-4- [ hydroxy (2-isopropyl-2H-indazol-3-yl) methyl ] -6-methylphenol (22)
Starting from compound 21, the synthesis of compound 22 was prepared according to the procedure of example 2 .1H NMR(DMSO-d6,400MHz)δ9.05(s,1H),7.56(d,J＝8.4Hz,1H),7.48(d,J＝8.4Hz,1H),7.30(s,1H),7.20-7.16(m,1H),7.03(s,1H),6.96-6.92(m,1H),6.38-6.33(m,2H),5.01-4.96(m,1H),2.17(s,3H),1.43(d,J＝6.4Hz,3H),1.31(d,J＝6.4Hz,3H).MS(EI,m/z)：375.1[M+H]+.
Example 8: synthesis of 3-bromo-5- (2-ethyl-2H-indazole-3-carbonyl) -2-hydroxybenzonitrile (28)
Step A: a mixture containing 2-bromo-4-hydroxybenzaldehyde (3.0 g,14.9 mmol), cuprous cyanide (1.74 g,19.4 mmol) and N-methylpyrrolidone (15 mL) was stirred at 180℃for 5 hours. Cooled to room temperature, ethyl acetate (50 mL) and water (50 mL) were added. After filtration through celite, the layers were separated, the aqueous phase was extracted with ethyl acetate (40 mL. Times.4), and the combined organic phases were washed with saturated brine (30 mL) and dried over anhydrous sodium sulfate. The solvent was distilled off under reduced pressure, and the product was purified by column chromatography (200 to 300 mesh silica gel, ethyl acetate: dichloromethane=1:15 elution) to give 5-formyl-2-hydroxybenzonitrile (23) and a small amount of residual N-methylpyrrolidone, the total weight of the mixture (2.40 g), and then directly used for the next reaction. MS (EI, m/z): 146.0[ M-H ] -.
And (B) step (B): chloromethyl methyl ether (1.26 g,15.7 mmol) was added to a solution of crude compound 23 (2.30 g) and diisopropylethylamine (2.60 g,20.1 mmol) in dichloromethane (25 mL) under ice-water bath, and after the addition was completed, the resulting mixture was stirred at room temperature overnight. Water (40 mL) was added, extracted with dichloromethane (40 mL. Times.3), and the combined organic phases were washed with saturated brine (30 mL) and dried over anhydrous sodium sulfate. The solvent was distilled off under reduced pressure, and the product was purified by column chromatography (200-300 mesh silica gel, dichloromethane: petroleum ether=1:10-20:1 elution) to give 5-formyl-2- (methoxymethoxy) benzonitrile (24) (673 mg). The total yield of the two reactions in steps A and B was 24.7%.
Experimental procedures for steps C and D5- (2-ethyl-2H-indazol-3-yl) hydroxymethyl-2- (methoxymethoxy) benzonitrile (26) was obtained according to the preparation methods in steps B and C of example 1.
Step E: to a solution of compound 26 (284 mg,0.845 mmol) in dichloromethane (5 mL) was added trifluoroacetic acid (1 mL), and the resulting mixture was stirred at room temperature overnight. Water (30 mL) was added, the pH was adjusted to 7-8 with saturated sodium bicarbonate solution, followed by extraction with ethyl acetate (30 mL. Times.3), and the combined organic phases were washed with saturated brine (20 mL) and dried over anhydrous sodium sulfate. The solvent was evaporated under reduced pressure to give 5- (2-ethyl-2H-indazole-3-carbonyl) -2-hydroxybenzonitrile (27) (230 mg). The yield thereof was found to be 93.4%.
Experimental procedure of step F3-bromo-5- (2-ethyl-2H-indazole-3-carbonyl) -2-hydroxybenzonitrile was obtained according to the preparation method of step E, example 6 (28).1H NMR(DMSO-d6,400MHz)δ8.00(d,J＝2.4Hz,1H),7.75(d,J＝8.8Hz,1H),7.68(d,J＝2.4Hz,1H),7.34-7.31(m,2H),7.19-7.15(m,1H),4.59(q,J＝7.2Hz,2H),1.49(t,J＝7.2Hz,3H).MS(EI,m/z)：368.0[M-H]-.
Example 9: synthesis of (3, 5-dibromo-4-hydroxyphenyl) (2-ethyl-2H-pyrazolo [3,4-c ] pyridin-3-yl) methanone (29)
The synthesis of compound 29 was prepared according to the procedure of example 1, wherein the indazole in example 1 step a was replaced with 1H-pyrazolo [3,4-c ] pyridine .1H NMR(DMSO-d6,400MHz)δ9.39(s,1H),8.20(d,J＝5.6Hz,1H),7.96(s,2H),7.10(d,J＝5.6Hz,1H),4.77(q,J＝7.2Hz,2H),1.57(t,J＝7.2Hz,3H).MS(EI,m/z)：424.0[M-H]-.
Example 10: synthesis of 2, 6-dibromo-4- { deuterium (2-ethyl-2H-pyrazolo [3,4-c ] pyridin-3-yl) hydroxymethyl } phenol (30)
To a solution of compound 29 (65 mg,0.154 mmol) in THF (6 mL) was added sodium borodeuteride (32 mg,0.765 mmol) and the resulting mixture was stirred at room temperature overnight. Water (20 mL) was added, the pH was adjusted to 7-8 with a 2M citric acid solution, followed by extraction with ethyl acetate (20 mL. Times.3), and the combined organic phases were washed with saturated brine (15 mL) and dried over anhydrous sodium sulfate. The solvent is distilled off under reduced pressure, and the product is purified by column chromatography (200-300 meshes of silica gel, ethyl acetate: petroleum ether=1:3-2:1 elution) to obtain 2, 6-dibromo-4- { deuterium (2-ethyl-2H-pyrazolo [3,4-c ] pyridine-3-yl) hydroxymethyl } phenol (30).1H NMR(DMSO-d6,400MHz)δ10.07(s,1H),9.24(s,1H),7.88(d,J＝5.6Hz,1H),7.57-7.55(m,3H),6.74(s,1H),4.61-4.55(m,2H),1.40(t,J＝7.2Hz,3H).MS(EI,m/z)：426.0[M-H]-.
Example 11: synthesis of (7-bromo-2-ethyl-6-methoxy-2H-indazol-3-yl) (3, 5-dibromo-4-hydroxyphenyl) methanone (37)
Step A: 2-fluoro-4-methoxybenzaldehyde (6.5 g,42.2 mmol) and 85% hydrazine hydrate (50 mL) were stirred for 30 hours at 120℃in a sealed tube. Water (50 mL) was added and extracted with ethyl acetate (50 mL. Times.3) and the combined organic phases were washed with water (20 mL. Times.2). Then water (100 mL) was added to the organic phase, the pH was adjusted to 1-2 with 2M hydrochloric acid, and the layers were separated and the product was in the aqueous phase. The aqueous phase was adjusted to pH 8-9 with 2M sodium hydroxide solution, then extracted with ethyl acetate (50 mL. Times.3), and dried over anhydrous sodium sulfate. The solvent was distilled off under reduced pressure, and the product was recrystallized from methyl tert-butyl ether/petroleum ether to give 6-methoxy-1H-indazole (31) (3.47 g). The yield thereof was found to be 55.5%.
And (B) step (B): a mixture containing compound (3.43 g,23.2 mmol), ethyl iodide (9.06 g,58.1 mmol), potassium hydroxide (3.41 g,60.8 mmol) and ethanol (25 mL) was stirred at reflux overnight. Most of the solvent was distilled off under reduced pressure, water (40 mL) was added, and the mixture was extracted with methylene chloride (40 mL. Times.5) and dried over anhydrous sodium sulfate. The solvent was distilled off under reduced pressure, and the product was purified by column chromatography (200-300 mesh silica gel, ethyl acetate: petroleum ether=1:20-1:5 elution) to give 2-ethyl-6-methoxy-2H-indazole (32) (1.48 g) and 1-ethyl-6-methoxy-1H-indazole (33) (2.44 g). The yields were 36.2% and 59.7%, respectively.
Experimental procedures for Steps C, D and E the preparation of Steps C, D and E was followed in example 8, wherein Compound 24 in step C of example 8 was replaced with 4-methoxymethoxybenzaldehyde to give (2-ethyl-6-methoxy-2H-indazol-3-yl) (4-hydroxyphenyl) methanone (36).1H NMR(DMSO-d6,400MHz)δ10.58(s,1H),7.72-7.68(m,2H),7.11(d,J＝4.0Hz,1H),6.98-6.90(m,3H),6.82-6.78(m,1H),4.58(q,J＝6.8Hz,2H),3.81(s,3H),1.48(t,J＝6.8Hz,3H).
Using compound 36 as a starting material, the experimental procedure of step F was followed in accordance with the preparation method of example 1, step E, to give (7-bromo-2-ethyl-6-methoxy-2H-indazol-3-yl) (3, 5-dibromo-4-hydroxyphenyl) methanone (37).1H NMR(DMSO-d6,400MHz)δ7.96(s,2H),7.23(d,J＝9.2Hz,1H),7.11(d,J＝9.2Hz,1H),4.64(q,J＝6.8Hz,2H),3.92(s,3H),1.52(t,J＝6.8Hz,3H).MS(EI,m/z)：530.8[M-H]-.
Example 12: synthesis of (7-bromo-2-ethyl-6-tridentate methoxy-2H-indazol-3-yl) (3, 5-dibromo-4-hydroxyphenyl) methanone (43)
Step A: a mixture containing compound 32 (800 mg,4.54 mmol), 1M boron tribromide in toluene (11 mL) and methylene chloride (20 mL) was stirred at room temperature overnight. The reaction was slowly poured into crushed ice (60 g), the pH was adjusted to 6 to 7 with 2M aqueous sodium hydroxide solution, extracted with methylene chloride (30 mL. Times.4), and the combined organic phases were washed successively with saturated aqueous sodium bicarbonate (25 mL) and saturated brine (20 mL), and dried over anhydrous sodium sulfate. The solvent was distilled off under reduced pressure to give 2-ethyl-2H-indazol-6-ol (38) (710 mg). The yield thereof was found to be 96.4%.
And (B) step (B): a mixture containing compound 38 (500 mg,3.08 mmol), potassium carbonate (850 mg,6.17 mmol), deuterated iodomethane (581 mg,4.01 mmol) and DMF (10 mL) was stirred overnight at 30 ℃. Water (40 mL) was added, extracted with dichloromethane (40 mL. Times.3), and dried over anhydrous sodium sulfate. The solvent was distilled off under reduced pressure, and the product was purified by column chromatography (200-300 mesh silica gel, ethyl acetate: petroleum ether=1:15-1:1 elution) to give 2-ethyl-6-tridentate methoxy-2H-indazole (39) (410 mg). The yield thereof was found to be 81.6%.
Experimental procedures for Steps C, D, E and F following the preparation methods of Steps C, D, E and F of example 8, (7-bromo-2-ethyl-6-tridentate methoxy-2H-indazol-3-yl) (3, 5-dibromo-4-hydroxyphenyl) methanone was obtained (43).1H NMR(DMSO-d6,400MHz)δ7.97(s,2H),7.22(d,J＝9.2Hz,1H),7.11(d,J＝9.2Hz,1H),4.65(q,J＝7.2Hz,2H),1.52(t,J＝7.2Hz,3H).MS(EI,m/z)：533.9[M-H]-.
Example 13: synthesis of (3, 5-dibromo-4-hydroxyphenyl) (2-ethyl-5-methyl-2H-indazol-3-yl) methanone (44)
The synthesis of compound 44 was prepared according to the procedure of example 1, wherein the indazole in example 1, step a, was replaced with a 5-methylindazole .1H NMR(DMSO-d6,400MHz)δ7.95(s,2H),7.73(d,J＝8.8Hz,1H),7.21(d,J＝8.8Hz,1H),6.90(s,1H),4.66(q,J＝7.2Hz,2H),2.30(s,3H),1.51(t,J＝7.2Hz,3H).MS(EI,m/z)：436.9[M-H]-.
Example 13: synthesis of (3, 5-dibromo-4-hydroxyphenyl) (2-ethyl-6-fluoro-2H-indazol-3-yl) methanone (45)
The synthesis of compound 45 was prepared according to the procedure of example 1, wherein the indazole in example 1, step a, was replaced with 6-fluoroindazole .1H NMR(DMSO-d6,400MHz)δ7.92(s,2H),7.59(d,J＝9.6Hz,1H),7.22-7.19(m,1H),7.14-7.10(m,1H),4.65(q,J＝7.2Hz,2H),1.52(t,J＝7.2Hz,3H).MS(ESI,m/z)：440.9[M-H]-.
Example 14 test of Compounds for inhibition of uric acid transported by hURAT1 in HEK293 transfected cell lines
1. Reagent name and source:
zurampic available from Chengdu-super pharmaceutical technologies Co., ltd; plasmid pCMV6-hURAT1 is purchased from Origene Technologies, inc; g418 is purchased from Bio-engineering Co., ltd; HEK293 cell lines were purchased from Shanghai life sciences research institute cell resource center, national academy of sciences; polylysine was purchased from Sigma-Aldrich Co.LLC; 14 C-uric acid is purchased from U.S. American Radiolabeled Chemicals, inc; sodium gluconate, potassium gluconate, calcium gluconate, KH 2PO4、MgSO4, glucose and HEPES were purchased from national pharmaceutical chemicals limited; DMEM medium, fetal bovine serum was purchased from Thermo FISHER SCIENTIFIC INC;
2. test methods and results:
1. Constructing HEK293 stable transgenic cell strain with high expression of hURAT 1: plasmid pCMV6-hURAT1 is transfected into HEK293 cells, and then G418 (final concentration 500 mug/mL) resistance screening is carried out to obtain stable transgenic cell strains, which highly express hURAT1 transport membrane proteins and can be used for in vitro inhibition tests of hURAT1 transport uric acid (Weaver YM,Ehresman DJ,Butenhoff JL,et al.Roles of rat renal organic anion transporters in transporting perfluorinated carboxylates with different chain lengths.Toxicological Sciences,2009,113(2):305-314).
2. Coating 24 pore plates: 0.1mg/mL polylysine was added at 200. Mu.l/well and left overnight. Removing polylysine, cleaning with sterile water, and air drying thoroughly.
3. HEK293-hURAT1 stably transfected cells were inoculated into a coated 24-well plate at 2X 10 5 cells/well, and cultured at 37℃under 5% CO 2 for 3 days.
Preparation of HBSS: weighing each reagent according to the final concentration of 125mM sodium gluconate, 4.8mM potassium gluconate, 1.3mM calcium gluconate, 1.2mM KH 2PO4、1.2mM MgSO4, 5.6mM glucose and 25mM HEPES, adding deionized water to fix the volume to the corresponding volume, fully and uniformly mixing to obtain the HBSS solution with the pH of 7.4, and storing in a refrigerator at the temperature of minus 20 ℃.
5. On the day of the experiment, HBSS was removed from the refrigerator and heated to 37 ℃ in a water bath. Taking out the 24-hole cell culture plate, cleaning the cell with HBSS for 2 times and sucking the cell, adding HBSS according to 160 mu l/hole, and adding a test compound with a final concentration of 500nM according to 20 mu l/hole to obtain a test compound hole; HBSS was added at 180 μl/well but no test compound was added as a blank well. Standing at room temperature for 10min.
6. 14 C uric acid was added at a final concentration of 50. Mu.M at 20. Mu.l/well and left at room temperature for 20min.
7. The solution was blotted off each well, and the cells were washed with pre-chilled HBSS and blotted off. Finally, 0.2M NaOH is added to lyse cells, cell fragments are collected, a proper amount of scintillation liquid is added, and then the scintillation liquid is placed on a PERKINELMER MICROBETA TRILUX liquid scintillation analyzer to detect the radiation intensity (CPM value) of the isotope 14 C uric acid.
8. In HEK293 transfected cell lines, the formula for calculating the inhibition rate of the compound on the uric acid transported by hURAT1 is shown as follows, and CPM value of the test compound is expressed as CPM ( Test compounds ); CPM values for the blank are shown as CPM ( blank control ). Test compounds were each set up in triplicate, the test results averaged, and the standard deviation SD was calculated. The test results are shown in Table 1.
3. Test results
Test compounds the compounds of the invention (in particular compounds 6, 7, 10, 15, 28, 29, 30 and 44) have very good inhibition of the uric acid transport by hURAT1 in HEK293 transfected cells compared to Zurampic at a concentration of 500 nM.
TABLE 1 inhibition of hURAT1 transfer uric acid by test compounds and Zurampic in HEK293 transfected cell lines
/>

Claims (7)

1. A compound of the structure shown in formula (I) or a pharmaceutically acceptable salt thereof,
Wherein,
A 1 is CH, A 2 is CH, A 3 is CH or N, and A 4 is CH;
g is a carbonyl or hydroxy-substituted methylene;
R 1 is selected from hydrogen, deuterium, halogen, cyano or substituted or unsubstituted C 1-5 alkyl;
R 2 is selected from halogen or cyano;
R 3 is selected from C 1-4 alkyl;
m is 0 or 1
N is 2;
The substituents in R 1 are selected from deuterium or halogen.
2. The compound of claim 1, or a pharmaceutically acceptable salt thereof, wherein R 1 is selected from one or more of hydrogen, deuterium, fluorine, chlorine, bromine, cyano, C 1-3 alkyl, C 1-3 haloalkyl, or C 1-3 deuterated alkyl; m is 0 or 1.
3. The compound according to claim 1, or a pharmaceutically acceptable salt thereof, wherein R 3 is selected from methyl, ethyl, n-propyl, or isopropyl.
4. A compound or a pharmaceutically acceptable salt thereof, wherein the compound is selected from the group consisting of:
。
5. A pharmaceutical composition comprising a compound according to any one of claims 1 to 4 or a pharmaceutically acceptable salt thereof as an active ingredient, together with pharmaceutically acceptable excipients.
6. Use of a compound according to any one of claims 1 to 4, or a pharmaceutically acceptable salt thereof, for the manufacture of a uric acid excretion-promoting drug.
7. Use of a compound according to any one of claims 1 to 4, or a pharmaceutically acceptable salt thereof, for the manufacture of a medicament for the treatment or prophylaxis of hyperuricemia, renal disease or gout.
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