CN106810488B - One class is used as GPR17 and CysLT1Compound of dual antagonist, preparation method and application thereof - Google Patents

Abstract

The invention is used as GPR17 and CysLT₁A compound of a dual antagonist, a preparation method and application thereof relate to a compound shown in a general formula I, a preparation method and application thereof. The compounds have weak antagonistic activity on GPR17 and CysLT₁Has strong antagonistic effect, and can be used for preventing and treating central nervous system injury and neurodegenerative diseases (such as multiple sclerosis and leukodystrophy); can also be used for the treatment of CysLT₁Mediated asthma and rhinitis, allergies, other inflammatory diseases.

Description

One class is used as GPR17 and CysLT1Compound of dual antagonist, preparation method and application thereof

Technical Field

The present invention relates to a novel class of compounds useful as GPR17 and CysLT₁Compounds of dual antagonists, methods of preparation and uses thereof. The compounds have weak antagonistic activity on GPR17 and CysLT₁Has strong antagonistic effect, and can be used for preventing and treating central nervous system injury and neurodegenerative diseases (such as multiple sclerosis and leukodystrophy); can also be used for the treatment of CysLT₁Mediated asthma and rhinitis, allergies, other inflammatory diseases.

Background

The orphan receptor GPR17(the G protein-coupled receptor 17) belongs to the large family of rhodopsin-like A class G-protein coupled receptors (GPCRs), and GPR17 is located at P from the phylogenetic tree₂Y₁₂、P₂Y₁₃、P₂Y₁₄Subgroup and intermediate of the cysteinyl leukotrien receptor (CysLT receptor), GPR17 has a typical seven-helix transmembrane structure, human GPR17 maps to chromosome 2q21, and CysLT₁Has a homology of 31%. GPR17 is expressed predominantly in the central nervous system, especially in differentiated oligodendrocyte precursor cells, with GPR17 having two splice variants, short (339 amino acids) and long (28 amino acids more N-terminal), with GPR17 being expressed at different levels in different tissues, short GPR17 being abundant in the brain, approximately 8-23 times longer than long, and vice versa in heart and kidney.

In recent years, GPR17 was found to be involved in myelin formation, Chen et al reported that GPR17 is involved in the whole process from early differentiation of Oligodendrocyte Precursor Cells (OPCs) to oligodendrocyte maturation and myelination, as expressed by the gradual up-regulation of GPR17 expression during differentiation of NG2 positive cells into OPCs, and the highest expression during the pre-differentiation of OPCs, and as the expression of OPCs differentiated mature GPR17 is gradually reduced, GPR17 is not expressed by fully mature oligodendrocytes. The helix-loop-helix transcription factor Olig1 promotes oligodendrocyte maturation, thereby repairing myelin sheaths. Olig1 negatively regulated the expression of GPR 17. Over-expression of GPR17 was found to inhibit oligodendrocyte differentiation maturation, causing demyelination, both in vivo and in vitro, and early myelination in GPR17 knockout mice. In conclusion, GPR17 has positive regulation effect on the early differentiation of OPCs and negative regulation effect on the late differentiation and maturation of oligodendrocytes. The inhibition of GPR17 in the later period of cerebral ischemia and spinal cord injury can promote the repair of myelin sheath.

After cerebral ischemia and cerebral trauma in human and experimental animals, synthesis and release of CysLTs are increased, and the CysLTs can activate GPR17 and participate in cerebral injury lesion. Ischemic central region CysLT in rat focal cerebral ischemia model₁The expression of receptor mRNA is increased, and has two peaks 3-6h and 7-14d after ischemia, while the expression is only increased at 7-14d in the ischemic periphery area; ischemic central CysLT₁The receptor was expressed in neurons 24h after ischemia, microglia after 14d, and proliferating astrocytes after 14d in the ischemic periphery. The mouse model of persistent focal cerebral ischemia also demonstrates CysLT₁The receptors have similar expression characteristics in the acute phase (Fang SH et al. neuroscience,2006,140(3): 969-. This indicates that CysLT₁The receptors are associated with neuronal damage during the acute phase of cerebral ischemia and with late glial cell proliferation. In the overall experiment, CysLT has been demonstrated₁The receptor antagonist has protective effect on cerebral ischemia and cerebral trauma.

Taken together, GPR17 and CysLT₁Has close relation with central nervous system injury, and the development of antagonists thereof has important significance for treating central nervous system diseases. At present, no selective GPR17 antagonist has been reported. It was found that the GPR17 receptor may have two distant ligand binding sites (nucleotide binding site and cysteinyl leukotriene binding site) and that the nucleotide binding region of GPR17 binds to P₂The nucleotide binding regions of the Y receptor are in the same position, and Ciana et al do it with 1321N1 cell expressing hGPR17³⁵S]GTP γ S binding assay, P₂Y₁₂/P₂Y₁₃Antagonists of the receptor cangrelor and P₂Y₁The receptor antagonist MRS2179 can block UDP-glucose induced activation, IC, of hGPR17₅₀0.7nM and 508nM, respectively; CysLT₁Concentration-dependent inhibition of LTD by receptor antagonists montelukast and pranlukast₄Induced activation of hGPR17, IC₅₀60nM and 10.5nM, respectively; in 2013, Hennen et al found that MDL29951 selectively activated GPR17 and failed to activate P₂Y receptor and CysLT₁The receptor, pranlukast, inhibits MDL 29951-induced activation of GPR17, whereas montelukast does not. GV150526A has a weak activating effect on GPR 17. In 2014, Kirsten Ritter et al used radioligand [ alpha ], [ beta ] -N-hydroxyquinoline³H]PSB-12150 was subjected to binding experiments, and MDL29951 and GV150526A, pranlukast and montelukast were found to have similar affinities for GPR17, with Ki of 2.32. mu.M, 1.63. mu.M, 4.06. mu.M and 6.54. mu.M, respectively. Compared with MDL29951, GV150526A has similar affinity for GPR17, the agonism is greatly reduced, probably caused by phenyl of amido bond side chain, and GV150526A can be regarded as a partial agonist of GPR 17. The inventors thus conceived of the use of (E) -3- [2- [ 7-chloro-2-quinoline of montelukast]Vinyl radical]The phenyl replaces the phenyl of the amido bond side chain of GV150526A to design and synthesize CHY113, so that the high affinity of the CHY113 to GPR17 is kept, and meanwhile, the antagonistic action to GPR17 is realized, and the activity result shows that the CHY113 has weak antagonistic action to GPR17 and CysLT₁Has stronger antagonism, and the result proves that the design idea is correct. On the basis, the inventor conducts preliminary structure optimization and evaluation on CHY113, thereby obtaining a series of compounds.

Disclosure of Invention

The object of the present invention is to provide a novel class of compounds useful as GPR17 and CysLT₁A compound that is a dual receptor antagonist;

another object of the present invention is to provide a process for producing the above compound;

it is still another object of the present invention to provide the use of the above compounds in the preparation of a medicament for preventing or treating central nervous system injury and neurodegenerative diseases (e.g., multiple sclerosis, leukodystrophy) and in the preparation of a medicament for treating CysLT₁Mediated byThe application of the medicine in treating asthma, rhinitis, allergy and other inflammatory diseases.

According to one aspect of the invention, there is provided a method for use as GPR17 and CysLT₁A compound which is a dual receptor antagonist having the structure of formula i:

wherein:

y is absent, or is-O-or-NH-;

x is absent, or is C₁-C₆Alkylene radical, C₂-C₆Alkenylene or C₂-C₆Alkynylene; preferably X is absent, or is-CH₂CH₂-or-CH ═ CH-;

absent, or a benzene ring;

R₁is any one selected from the following groups: h; a halogen atom; a hydroxyl group; a carboxyl group; an amino group; c₁-C₁₀A linear or branched alkyl group; c₁-C₁₀A linear or branched alkoxy group; cyano group C₀-C₆An alkyl group; c₂-C₈An alkenyl group; c₂-C₆An alkynyl group; c₁-C₆An alkylamino group; 3-8 membered cycloalkyl C₀-C₁₀An alkyl group; 5-to 8-membered aryl C₀-C₆Alkyl, wherein the 5-to 8-membered aryl is unsubstituted or substituted by a group selected from:

R₁preferably C₁-C₆Linear or branched alkyl of (a); c₂-C₅An alkynyl group; 5-8 membered cycloalkyl C₀-C₆An alkyl group; phenyl radical C₀-C₆Alkyl, wherein the phenyl is unsubstituted or selected fromAnd (3) group substitution:

R₁further preferred is

Methyl, ethyl, propyl, n-butyl, isobutyl, pentyl, hexyl, phenyl C₁-C₆Alkyl, phenyl, 4-6 membered cycloalkyl C₀-C₆Alkyl, cyano C₁-C₆Alkyl, phenyl C₁-C₆An alkoxyphenyl group;

m is an integer from 0 to 10, preferably 0, 1,2, 3, 4, 5, 6, 7, 8, 9 or 10;

R₂and R₄Each independently is hydrogen, C₁-C₄Alkyl radical, C₁-C₄Alkoxy, halogen atom, hydroxyl or amino; preferably each independently hydrogen, C₁-C₃Alkyl radical, C₁-C₃Alkoxy, F, Cl, hydroxy, or amino;

R₇selected from H, tetrazolyl, - (CH)₂)_nCOOH, wherein n is an integer of 0 to 3; preferably, R₇Selected from H, - (CH)₂)_nCOOH, wherein n is an integer from 0 to 2, preferably 0, 1 or 2;

when in use

In the absence of, R_5aAnd R_6aIs absent, R_5bAnd R_6bEach independently selected from H, halogen atom (e.g., F, Cl, etc.), hydroxy, carboxy, amino, C₁-C₁₀Straight or branched alkyl, C₁-C₁₀Straight or branched alkoxy, C₂-C₈Alkenyl radical, C₂-C₆Alkynyl, C₁-C₆An alkylamino group; preferably R_5bAnd R_6bEach independently of the other being H, a halogen atom, C₁-C₆An alkoxy group;

when in use

When it is a benzene ring, R_5bAnd R_6bIs absent, R_5aAnd R_6aEach independently selected from H, halogen atom (such as F, Cl), hydroxyl, carboxyl, amino, and C₁-C₁₀Straight or branched alkyl, C₁-C₁₀Straight or branched alkoxy, C₂-C₈Alkenyl radical, C₂-C₆Alkynyl, C₁-C₆An alkylamino group; preferably R_5aAnd R_6aEach independently of the other being H, a halogen atom, C₁-C₆An alkoxy group;

preferably, the compounds of the invention are used as GPR17 and CysLT₁The compound of the receptor dual antagonist has the following structure of a general formula II or a general formula IV:

X、Y、R₁、R₇、R_5a、R_6a、R_5band R_6bThe definitions of (A) and (B) are the same as previously defined;

in a more preferred embodiment of the invention, R in said formula II₁The structure of (A) is as follows:

y is-NH-;

x is-CH ═ CH-, the double bond has the configuration Z, E, preferably the E configuration;

wherein R is₇Preferably a carboxyl group;

preferably, said compounds of the invention are used as GPR17 and CysLT₁The compound of the receptor dual antagonist has the following structure of a general formula III:

wherein:

R_5aand R_6aThe definition of (1) is as above; preferably R_5aAnd R_6aEach independently H, halogen (e.g. F, Cl), C₁-C₆An alkoxy group;

further preferably, the compounds of the general formula i according to the invention have the following structure:

according to another aspect of the present invention, there is provided a pharmaceutical composition comprising a compound according to formula I of the present invention.

The pharmaceutical composition further comprises pharmaceutically acceptable auxiliary materials, excipients and the like.

According to a further aspect of the invention there is provided a composition as described herein for use as GPR17 and CysLT₁A preparation method of a compound of a receptor dual antagonist.

The preparation method of the compound shown in the general formula I comprises the following steps:

a) dissolving the compound shown as the general formula V in DMF, and adding R under the protection of nitrogen₁OH, condensing agent DCC or EDCI and DMAP, and carrying out esterification reaction at room temperature overnight;

or dissolving the compound shown as the general formula V in DMF, adding condensing agents HATU and DIPEA at room temperature, and adding R after 15 minutes to half an hour₁NH_2，Standing at room temperature overnight;

or dissolving the compound shown as the general formula V in DMF, adding alkali potassium carbonate or cesium carbonate in ice water bath under the protection of nitrogen, adding R after 15 minutes₁Br at room temperature orReacting at 60 ℃ for 4 to 16 hours;

or the compound shown in the general formula V is dissolved in DMF, cesium carbonate and R are added at room temperature₁OTs, overnight at room temperature;

when R is₉And R₇When not identical, further comprises a step b)

b) Make R₉Deprotection to give compounds of formula I. For example, when the protecting group is methyl or ethyl, ethanol or isopropanol is used as a solvent, lithium hydroxide or sodium hydroxide is used as an alkali, and the reaction is carried out at 60 ℃ for 4h to 16h to obtain acid; and when the protecting group is trimethylsiloxyethyl group, taking tetrahydrofuran as a solvent, adding TBAF (tert-butyl ether), and reacting at room temperature for 4 hours to obtain the acid.

Wherein X is absent or-CH₂CH₂-or-CH ═ CH-; y is absent, or-O-, -NH-); r₉Is hydrogen, or- (CH)₂)_nCOOR₁₀，R₁₀Being carboxyl protecting groups, e.g. C₁-C₆Alkyl, trimethylsilylethyl; r₇Is H, - (CH)₂)_nCOOH；

R₁、R_5a、R_6a、R_5b、R_6b、

And n is as defined above.

Wherein the compound shown in the general formula V is prepared by the following steps:

c) oxidizing the aldehyde group in the general formula VI by an oxidant to obtain a compound in a general formula V;

for example, the reaction conditions may be such that aldehyde is added to acetonitrile water (5:1, v/v), sodium dihydrogen phosphate dihydrate, hydrogen peroxide and sodium hypochlorite are added, and the reaction is carried out overnight at room temperature;

or

d) Refluxing a compound shown as a general formula VI and tert-butyloxycarbonyl methylene triphenylphosphine in toluene overnight to perform Horner-Wadsworth-Emmons reaction; then using formic acid to react at room temperature overnight or using trifluoroacetic acid to react at room temperature for 2 to 6 hours to selectively hydrolyze tert-butyl ester to obtain alpha, beta unsaturated carboxylic acid in the general formula V, and optionally, using a reducing agent Pd/C to catalyze and hydrogenate the alpha, beta unsaturated carboxylic acid in methanol and tetrahydrofuran (2:1, V/V) at room temperature for 4 hours to obtain a compound shown in the general formula V;

or

e) Reacting the aldehyde group in the general formula VI with malonic acid at 50 ℃ overnight under the action of weak base such as pyridine and piperidine to obtain a compound shown in the general formula V;

wherein X is absent or-CH₂CH₂-or-CH ═ CH-; r₉Is hydrogen, - (CH)₂)_nCOOH or- (CH)₂)_nCOOR₁₀，R₁₀Is a carboxyl protecting group;

R_5a、R_6a、R_5b、R_6b、

and n is as defined above;

more specifically, the compounds of formula I of the present invention may be prepared by a variety of procedures and synthetic routes, representative procedures and synthetic methods are as follows, but not limited to the following:

route one:

wherein R is_5a、R_6aAnd R₁The definition of (A) is the same as that described above;

specifically, Japp-Klingemann condensation is carried out on a compound 1 and ethyl 2-methylacetoacetate to generate a compound 2, the compound 2 is subjected to ring closing to generate an indole compound 3 under the action of polyphosphoric acid, the compound 3 is subjected to Vilsmeier-Haack formylation to obtain a compound 4, the compound 4 is hydrolyzed under the alkaline condition to obtain an acid 5, the acid 5 and 2-trimethylsilylethanol are subjected to condensation agents EDCI and DMAP to generate an ester compound 6, and the compound 6 and tert-butyloxycarbonyl methylene triphenylphosphine are subjected to condensationRaw Horner-Wadsworth-Emmons reaction to obtain unsaturated ester compound 7, selective hydrolysis of tert-butyl ester by formic acid to obtain acid compound 8, and introduction of ester group R into compound 8 by different methods₁Compound 9 is obtained, and deprotection of the compound 9 by TBAF gives the compound shown in the general formula 10.

Wherein compounds 8 to 9 can be converted by the following method: compound 8 and alcohol R₁OH is condensed under the conditions of DCC and DMAP; ② Compounds 8 and R₁OTs are performed under basic conditions (e.g., NaHCO)₃) Nucleophilic substitution is carried out to obtain; compound 8 and R₁Br under alkaline conditions (e.g. Cs)₂CO₃) Nucleophilic substitution is carried out to obtain;

and a second route:

wherein R is_5aAnd R_6aEach independently of the other being H, a halogen atom, C₁-C₄An alkoxy group; r₈Is methyl or ethyl; preferably R₁Comprises the following steps:

specifically, the compound 11 is formylated by Vilsmeier-Haack to obtain 12, the compound 12 reacts with tert-butyloxycarbonylmethylenetriphenylphosphine to obtain 13, the compound 13 reacts with trifluoroacetic acid to obtain 14, the compound 14 and amine are condensed into an amide compound 15 under the action of a condensing agent, and the compound 15 is hydrolyzed into acid under the alkaline condition to obtain the compound shown in the general formula 16.

For the commercially unavailable indole compound 11, the synthetic route is as follows:

1. synthesis of indoles by the method of Synthesis of Compound 3 in route one, which isThe compound 2 can also be in ZnCl₂Compound 3 is obtained under the condition of TsOH/Tol;

2、

reacting a compound 19 with sodium azide to generate a compound 20, reacting the compounds 17 and 20 under the condition of sodium methoxide to obtain a compound 18, and heating and ring closing the compound 18 to obtain an indole compound 11;

3、

the hydrochloride of the substituted phenylhydrazine or the substituted phenylhydrazine and pyruvate are condensed into a compound 22, and the compound 22 is subjected to ring closure in PPA to obtain an indole compound 11; sometimes the compound 21 and pyruvate directly obtain the indole compound 11 under the catalysis of acetic acid;

and a third route:

wherein R is_5aAnd R_6a、R₁The definition of (1) is as above; r₁Preferably C₁-C₆Alkyl, phenyl substituted C₁-C₆Alkyl, phenyl;

in particular, compound 23 and PPh₃Reacting to obtain a compound 24, reacting the compound 24 with the compound 4 to obtain a compound 25, and hydrolyzing the compound 25 under an alkaline condition to obtain a compound shown as a general formula 26;

and a fourth route:

wherein R is_5aAnd R_6aPreferably H, Cl, Y is preferably-NH-, R₁Preferably:

specifically, Pd/C catalyzed hydrogenation of compound 14 affords compound 27, compound 27 and R₁YH condensation to obtain 28, hydrolyzing the compound 28 under an alkaline condition to obtain the compound shown as the general formula 26;

and a fifth route:

wherein R is_5aAnd R_6aPreferably H, Cl, Y is preferably-NH-, R₁Preferably:

specifically, the aldehyde group at the 3-position of the indole ring of the compound 12 is oxidized into a carboxyl group by hydrogen peroxide or the like to obtain a compound 30, and the compound 30 and R₁YH condensation to obtain 31, hydrolyzing the compound 31 under an alkaline condition to obtain the compound shown as a general formula 32;

route six:

wherein R is_5aAnd R_6aPreferably H, Cl, Y is preferably-O-, R₁Preferably:

specifically, compound 5 is decarboxylated under the conditions of cuprous chloride and quinoline to obtain compound 33, compound 33 is reacted with malonic acid under the conditions of weak base to obtain 34, and acid 34 and bromide R₁Br is subjected to substitution reaction under alkaline condition to generate the compound shown in the general formula 35; the bromide is synthesized as follows:

compound 36 is reduced by sodium borohydride to alcohol 37, which is shown in NBS, PPh₃Is converted to bromide 38 under conditions.

A seventh route:

wherein R is_5bAnd R_6bPreferably H, methyl, ethyl, phenyl, R₁Preferably:

specifically, compound 39 is reacted with compound 46 to form compound 40, compound 40 is oxidized by PCC to compound 41, and compound 41 is converted to the compound of formula 45 according to the present invention using the method for synthesizing compound 16, route two.

According to another aspect of the present invention, there is provided the use of a compound of formula I for the preparation of a medicament for the prevention or treatment of central nervous system injury and neurodegenerative diseases (e.g. multiple sclerosis, leukodystrophy) and for the preparation of a medicament for the treatment of CysLT₁Mediated asthma, rhinitis, allergy, other inflammatory diseasesUse in medicine.

According to a further aspect of the invention there is provided the use of a compound of formula I for the prevention or treatment of central nervous system injury and neurodegenerative diseases (e.g. multiple sclerosis, leukodystrophy) and for the treatment of CysLT₁Mediated asthma, rhinitis, allergy, and other inflammatory diseases.

The invention designs and synthesizes a novel GPR17 receptor and CysLT₁The receptor dual antagonist, in vitro calcium flow experiment shows that the compound has weak antagonistic activity on GPR17 and CysLT₁Has strong antagonistic effect, and can be used for preventing and treating brain and spinal cord injury and neurodegenerative diseases (such as multiple sclerosis and leukodystrophy); can also be used for the treatment of CysLT₁Mediated asthma and rhinitis, allergies, other inflammatory diseases.

Detailed Description

The present invention will be further illustrated with reference to the following specific examples, but the present invention is not limited to these examples.

Examples of preparation of Compounds

In the following preparation examples, NMR was measured with a Mercury-Vx 300M instrument manufactured by Varian, and NMR was calibrated: Δ H7.26 ppm (CDCl)₃),2.50ppm(DMSO-d₆),2.05ppm(Acetone-d₆) (ii) a Reagents are mainly provided by Shanghai chemical reagents company; TLC thin layer chromatography silica gel plate is produced by Shandong tobacco Taihuyou silica gel development Co., Ltd, model number HSGF 254; the normal phase column chromatography silica gel used for compound purification is produced by Shandong Qingdao ocean chemical plant, model zcx-11, 200-300 mesh.

Preparation example one (Compound No. CHY90)

3, 5-dichloroaniline (5g,30.8mmol) is added into a single-neck round-bottom flask, water (20mL) is added, concentrated hydrochloric acid (20mL) is added dropwise under the condition of ice-water bath, and after dropwise addition is finished, ice water cooled sodium nitrite (2.34g,33.91mmol) water solution 1 is added dropwise1mL, stirring for 20 minutes under the condition of ice-water bath after dripping; in another round bottom flask, ethyl 2-methylacetoacetate (6.6g,45.78mmol) was added, ethanol (25mL) was added, 30mL of an aqueous solution of potassium hydroxide (15.5g,0.28mol) was added dropwise in an ice-water bath, after completion of addition, the mixture was stirred for 15 minutes in an ice-water bath, added dropwise to the first flask, stirred for 15 minutes at 40 ℃ and cooled to room temperature, extracted with ether (20 mL. times.3 times), the organic phases were combined, washed with saturated brine (20mL), and anhydrous MgSO 2₄Drying, removing the solvent under reduced pressure and purifying by silica gel column chromatography (PE/EtOAc ═ 100:1 → 50:1 → 20:1) to give product M2(2.876g, 33.9%, yellow solid).

¹H NMR(300MHz,CDCl₃)：δ7.63(s,1H),7.11(d,J＝1.8Hz,2H),6.94(d, J＝1.8Hz,1H),4.33(q,J＝7.1Hz,2H),2.10(s,3H),1.39(t,J＝7.1Hz,3H).

Compound M2(780mg,2.84mmol) was charged in a single neck round bottom flask, polyphosphoric acid (9.6g), toluene (10mL) were added, stirring was carried out overnight at 45 ℃, toluene was removed under reduced pressure, ice water was added, stirring was carried out for 15 minutes, a precipitate was precipitated, the filtrate was filtered, and the filter cake was washed with CH₂Cl₂Washed twice and dried under vacuum at 40 ℃ to give compound M3 (707mg, 96.5%, white solid). 1.876g of Compound M3 were prepared in a similar manner.¹H NMR(300MHz,DMSO-d₆)：δ7.43(s,1H),7.25(s,1H),7.09(s,1H),4.34(q,J ＝6.6Hz,2H),1.33(t,J＝6.9Hz,3H).

Placing a two-neck round-bottom flask provided with a condenser tube in an ice-water bath, adding DMF (3.26mL, 42.32mmol) under the protection of argon, and slowly dropwise adding POCl₃(3.65mL,39.92mmol), stirred at room temperature for 15 minutes, added M3(2.06g,7.98mmol) in 1, 2-dichloroethane (40mL), refluxed at 100 ℃ for 7 hours, cooled to room temperature, poured slowly into an aqueous solution of sodium acetate (50g AcONa,100mL H)₂O), a large amount of yellow solid precipitated, which was stirred overnight, filtered, the filter cake was washed with water and dichloromethane, respectively, and dried under vacuum at 40 ℃ to give compound M4(1.1g, 59.2%, grey solid).¹H NMR(300 MHz,DMSO-d₆)：δ10.63(s,1H),7.56(d,J＝1.8Hz,1H),7.42(d,J＝1.5Hz, 1H),4.44(q,J＝7.2Hz,2H),1.37(t,J＝7.2Hz,3H)。

Compound M4(1.1 g)3.86mmol) was added to a single neck round bottom flask, ethanol (60mL) and lithium hydroxide (616mg,14.67mmol) were added, and gradually dissolved by heating to 50 ℃ after 2 hours, the reaction was completed, ethanol was concentrated, diluted with water, pH adjusted to 2 with 1N hydrochloric acid, yellow precipitate was precipitated, filtered, the filter cake was washed with water 2 times, and dried under vacuum at 40 ℃ to give compound M5(920mg, 92.4%, yellow solid).¹H NMR(300 MHz,DMSO-d₆)：δ10.76(s,1H),7.50(d,J＝1.5Hz,1H),7.34(d,J＝2.1Hz, 1H).

Compound M5(600mg,2.33mmol), redistilled 1, 2-dichloroethane (20mL), 2-trimethylsilylethanol (332mg,2.79mmol), EDCI (671mg,3.50 mmol), DMAP (340mg,2.79mmol) were added in this order to a round-bottomed flask, and stirred at room temperature overnight, the insoluble matter was filtered off, the solvent was removed by evaporation under reduced pressure, and the resulting crude product was dissolved in ethyl acetate (30mL), washed with 0.5N hydrochloric acid (40 mL. times.3), washed with saturated brine (20mL), dried MgSO (MgSO) anhydrous MgSO₄Drying, concentration and purification by silica gel column chromatography (PE/EtOAc ═ 20:1) gave compound M6(200mg, 24%, white solid).¹H NMR(300MHz,CDCl3): δ10.81(s,1H),9.47(bs,NH),7.38(d,J＝1.5Hz,1H),7.34(d,J＝1.8Hz,1H), 4.56-4.51(m,2H),1.24-1.18(m,2H),0.11(s,9H).

Compound M6(200mg,0.56mmol) was added to a single neck round bottom flask, tert-butoxycarbonylmethylenetriphenylphosphine (315mg,0.84mmol) was added, redistilled toluene (20mL) was added under nitrogen, reflux overnight, the solvent was removed under reduced pressure, the resulting crude product was dissolved in dichloromethane (10mL), and purified by column chromatography on silica gel (PE/EtOAc ═ 30:1) to give compound M7(203mg, 79.6%, yellow solid).¹H NMR(300MHz,CDCl₃):δ9.15(bs,NH),8.32(d,J＝16.5Hz,1H),7.32(d,J＝ 1.5Hz,1H),7.20(d,J＝1.5Hz,1H),6.48(d,J＝16.2Hz,1H),4.49-4.43(m, 2H),1.26-1.15(m,2H),0.08(s,9H).

Compound M7(190mg,0.42mmol) was added to a single neck round bottom flask, 98% formic acid (50mL) was added, stirred at room temperature overnight, the solvent was spun off under reduced pressure, the crude product was dissolved in acetone (10mL), and purified by column chromatography on silica gel (PE/EtOAc ═ 20:1) to give 62mg of compound M7, (PE/EtOAc ═ 4:1) to give compound M8(78mg, 46.4%, white solid).¹H NMR(300MHz,Acetone-d₆)):δ11.63 (bs,1H),10.69(bs,1H),8.50(d,J＝16.5Hz,1H),7.59(s,1H),7.27(s,1H),6.64 (d,J＝15.9Hz,1H),4.48(t,J＝9.0Hz,2H),1.19(t,J＝9.0Hz,2H),0.09(s, 9H).

Dissolving compound M8(10mg,0.025mmol) in 0.5mL DMF, adding cesium carbonate (12mg,0.038mmol) under ice-water bath, adding benzyl bromide (4.7mg,0.028mmol) in DMF (0.2 mL) dropwise, reacting at room temperature for 4 hours, adding ethyl acetate (5mL), water (5mL), shaking, standing for layering, washing the organic phase with saturated common salt water, anhydrous MgSO₄Drying, concentration and purification by silica gel column chromatography (PE/EtOAc ═ 20:1) gave compound M9(10mg, 83.3%, yellow solid).¹H NMR(300MHz,CDCl3): δ9.20(bs,NH),8.49(d,J＝16.2Hz,1H),7.44-7.21(m,7H),6.64(d,J＝16.2 Hz,1H),5.28(s,2H),4.48-4.42(m,2H),1.17-1.11(m,2H),0.09(s,9H).

Compound M9(10mg,0.020mmol) was dissolved in 0.3mL of THF, TBAF (0.2mL,1M in THF) was slowly added dropwise, the reaction was carried out at room temperature for 3 hours, THF was removed by evaporation under reduced pressure, water (2mL) was added, 1N hydrochloric acid was added to adjust pH to 2, extraction was carried out with ethyl acetate (4 mL. times.2), the organic phases were combined, washed with saturated brine, anhydrous MgSO 2₄Drying, concentration and purification by silica gel column chromatography (PE/EtOAc ═ 1:1) gave product CHY90 (8mg, 100%, yellow solid). Wherein the double bond is mainly in E configuration and contains a trace of Z configuration.¹H NMR (300MHz,Acetone-d₆):δ11.66(bs,1H),8.62(d,J＝16.2Hz,1H),7.62(d,J＝ 2.1Hz,1H),7.49-7.33(m,5H),7.27(d,J＝1.5Hz,1H),6.79(d,J＝16.2Hz, 1H),5.27(s,2H).

The following compounds were synthesized using a procedure similar to that of preparation example one:

preparation example two (Compound No. CHY98)

Adding the compound 38(100mg,0.37mmol) into 5mL of methanol to form turbid, adding sodium borohydride (28mg,0.73mmol) in an ice water bath, reacting for 8 hours at room temperature, detecting by TLC that the reaction is not complete, supplementing 50mg of sodium borohydride, reacting overnight at room temperature, detecting by TLC that the reaction is complete, removing methanol by decompression, adding water (10mL), diluting, filtering, and drying to obtain the compound 37(100mg, 100%, white solid).¹H NMR (300MHz,DMSO-d₆):δ8.40(d,J＝9.0Hz,1H),8.03-7.31(m,10H),5.26(s, 1H),4.56(s,2H).

Compound M8(20mg,0.050mmol) was dissolved in anhydrous DMF (1mL), and compound 37(16mg,0.058mmol), DCC (16mg,0.078mmol), DMAP (4mg,0.033 mmol) were added, respectively, reacted at room temperature overnight, diluted with ethyl acetate (5mL), washed with water (5 mL. times.2), washed with saturated brine, and anhydrous MgSO₄Drying, concentration and purification by silica gel column chromatography (PE/EtOAc 10:1) afforded compound M10(25mg, 74%, yellow solid).¹H NMR(300MHz,CDCl3):δ9.20(bs, NH),8.54(d,J＝15.9Hz,1H),8.11(d,J＝8.7Hz,1H),8.07(s,1H),7.77-7.22 (m,11H),6.69(d,J＝15.9Hz,1H),5.33(s,2H),4.46(m,2H),1.15(m,2H), 0.06(s,9H).

Dissolving a compound M10(25mg,0.037mmol) in 1mL THF, slowly dropwise adding TBAF (0.3 mL,1M in THF), reacting for 2 hours at room temperature, detecting complete reaction by TLC, removing THF by decompression, adding water (2mL), adjusting pH to 2 by 1N hydrochloric acid, precipitating yellow solid, filtering, washing a filter cake with water and dichloromethane respectively, drying in vacuum at 40 ℃ to obtain 15mg of a crude product, and pulping with acetone to obtain a product CHY98(10mg, 47.6%, yellow solid) containing a small amount of impurities.¹H NMR(300MHz,DMSO-d₆):δ8.63(d, J＝7.8Hz,1H),8.54(d,J＝15.9Hz,1H),8.17-8.06(m,5H),7.81(s,1H), 7.75-7.50(m,5H),7.32(s,1H),6.74(d,J＝15.9Hz,1H),5.31(s,2H).

CHY109 was synthesized by a procedure similar to that of the preparation example.¹H NMR(300MHz, Acetone-d₆):δ11.73(bs,1H),8.77(d,J＝15.9Hz,1H),7.65(s,1H),7.45-7.22 (m,6H),6.99(d,J＝16.2Hz,1H).

Using a procedure similar to that of the preparation example, CHY111 was synthesized.¹H NMR(300MHz, CDCl3):δ9.53(s,1H),9.19(s,1H),8.83(d,J＝16.5Hz,1H),8.36(s,1H),8.18 (d,J＝8.7Hz,1H),8.10(d,J＝8.1Hz,1H),7.92(d,J＝8.7Hz,1H),7.82-7.70 (m,2H),7.59(s,1H),7.53-7.21(m,8H),6.78(d,J＝16.5Hz,1H),5.35(s,2H).

Preparation example three (Compound No. CHY96)

4-Pentyyn-1-ol (100mg,1.19mmol) and pyridine (141mg,1.57mmol) were sequentially added to anhydrous dichloromethane (2mL), and a solution of p-toluenesulfonyl chloride (248mg,1.30mmol) in dichloromethane (2mL) was added dropwise in an ice-water bath, reacted at room temperature for 2 hours, washed with 1N hydrochloric acid and a saturated aqueous sodium bicarbonate solution, respectively, and anhydrous MgSO₄Drying, concentration and purification by silica gel column chromatography (PE/EtOAc 10:1) gave compound M11(73mg, 23.8%, colorless oil).¹H NMR(300MHz,CDCl3):δ7.81-7.78(m, 2H),7.36-7.33(m,2H),4.15(t,J＝6Hz,2H),2.45(s,3H),2.25(td,J＝6.9,2.7 Hz,2H),1.90-1.81(m,2H),1.58(s,2H).

Dissolving compound M11(13mg,0.055mmol) in 0.5mL DMF, adding compound M8(20mg,0.050mmol) and sodium bicarbonate (12mg,0.15mmol), reacting overnight at room temperature, detecting by TLC to remove reaction, adding cesium carbonate (24mg,0.074mmol), reacting overnight at 60 deg.C, adding ethyl acetate (5mL), water (5mL), adjusting pH to 2 with 1N hydrochloric acid, shaking, standing for layering, washing the organic phase with saturated saline, anhydrous MgSO₄Drying, concentration and purification by silica gel column chromatography (PE/EtOAc ═ 20:1) gave compound M12(16mg, 69.6%, white solid). The nuclear magnetism contains a small amount of Z configuration products, and E configuration is the main.¹H NMR(300MHz,CDCl3):δ9.42(s,1H),8.44(d,J＝16.2Hz, 1H),7.33(d,J＝1.5Hz,1H),7.20(d,J＝1.5Hz,1H),6.57(d,J＝15.9Hz,1H), 4.50-4.42(m,2H),4.34(t,J＝6.3Hz,2H),2.35(td,J＝7.2,3.0Hz,2H), 2.04-1.91(m,2H),1.64(s,2H),1.22-1.12(m,2H),0.09(s,9H).

Compound M12(16mg,0.034mmol) was dissolved in 0.3mL THF, TBAF (0.2mL,1M in THF) was slowly added dropwise, the reaction was allowed to proceed at room temperature for 4 hours, TLC was used to check completion of the reaction, THF was removed under reduced pressure, water was added(2mL), pH was adjusted to 2 with 1N hydrochloric acid, extraction was performed with ethyl acetate (4 mL. times.2), and the organic phases were combined, washed with saturated brine, and anhydrous MgSO₄Drying, concentration and purification by silica gel column chromatography (PE/EtOAc ═ 2:1) gave product CHY96(6mg, 50%, yellow solid). Wherein the double bond is mainly in E configuration and contains a trace of Z configuration.¹H NMR(300MHz,Acetone-d₆):δ11.63(bs,1H),8.55(d,J＝15.9 Hz,1H),7.61(s,1H),7.26(s,1H),6.72(d,J＝16.2Hz,1H),4.30(t,J＝6.3Hz, 2H),2.35(m,2H),2.05(s,1H),1.93(m,2H).

Preparation example four (Compound No. CHY113)

Compound M4 was prepared as described in example one.

Compound M13(850mg,5.63mmol) and compound M14(1g,5.63mmol) were added to a bottle, acetic anhydride (6mL) was added, reacted at 60 ℃ under nitrogen for 4.5 hours, cooled to room temperature to precipitate a yellow solid, filtered, and the filter cake was washed twice with water and ether to give compound M15(1.52g, 86.9%, yellow solid).¹H NMR(300MHz,CDCl3):δ8.50(t,J＝2.1Hz,1H),8.20-8.15 (m,2H),8.09(d,J＝2.1Hz,1H),7.94(d,J＝7.5Hz,1H),7.80(d,J＝16.2Hz, 1H),7.75(d,J＝8.7Hz,1H),7.64(d,J＝8.7Hz,1H),7.59(t,J＝8.1Hz,1H), 7.48(dd,J＝8.7,2.1Hz,1H),7.47(d,J＝16.5Hz,1H).

Dissolving stannous chloride dihydrate (1.81g,8.05mmol) in concentrated hydrochloric acid (4mL), dropwise adding acetic acid solution (8mL) of compound M15(500mg,1.61mmol) at room temperature, reacting at 60 ℃ for 2 hours to obtain yellow slurry, concentrating to remove acetic acid, diluting with 20mL of water, adjusting pH to 9 with 10N sodium hydroxide aqueous solution, extracting with ethyl acetate (50 mL. times.2), combining organic phases, washing with saturated salt water, and removing anhydrous MgSO (MgSO) MgSO₄After drying and concentration, compound M16(413mg, 91.8%, yellow solid) was obtained. ESIMS 281.0[ M + H ]]⁺；¹H NMR(300MHz,DMSO-d₆):δ8.38(d,J＝8.7Hz,1H),8.02(d,J＝1.8Hz,1H), 7.99(d,J＝8.7Hz,1H),7.90(d,J＝8.7Hz,1H),7.71(d,J＝16.2Hz,1H),7.58 (dd,J＝8.7,2.4Hz,1H),7.28(d,J＝16.2Hz,1H),7.09(t,J＝7.8Hz,1H),6.89 (s,1H),6.88(d,J＝8.4Hz,1H),6.58(d,J＝9.0Hz,1H),5.17(s,2H).

Compound M4(500mg,1.75mmol) was added to a single neck round bottom flask, tert-butoxycarbonylmethylenetriphenylphosphine (987mg,2.63mmol) was added, redistilled toluene (20mL) was added under nitrogen, reflux overnight, the solvent was removed under reduced pressure, the crude product was dissolved in dichloromethane (30mL), and purified by column chromatography on silica gel (PE/EtOAc ═ 8:1) to give compound M17(828mg, yield > 100%, yellow solid).¹H NMR(300MHz,CDCl₃):δ9.18(bs,NH),8.33(d,J＝16.2Hz,1H),7.34(s, 1H),7.21(s,1H),6.48(d,J＝16.2Hz,1H),4.44(q,J＝7.2Hz,2H),1.44(t,J＝ 7.2Hz,3H).

Dissolving compound M17(417mg,1.08mmol) in dichloromethane (15mL), adding trifluoroacetic acid (2mL,26.93mmol) dropwise in ice-water bath, reacting at room temperature for 1 hr, detecting by TLC, and purifying by silica gel column chromatography (CHCl)₃/CH₃OH 30:1) to give compound M18(340mg, 96.0%, yellow solid).¹H NMR(300MHz,DMSO-d₆):δ12.66(s, 1H),8.28(d,J＝15.9Hz,1H),7.51(s,1H),7.33(s,1H),6.45(d,J＝16.2Hz, 1H),4.37(q,J＝6.9Hz,2H),1.35(t,J＝6.9Hz,3H).

Compound M18(80mg,0.24mmol) was dissolved in DMF (1mL), HATU (139 mg,0.37mmol) and DIPEA (74mg,0.57mmol) were added, reacted at room temperature for half an hour, a solution of compound M16(68mg,0.24mmol) in dichloromethane (1mL) was added dropwise, the flask wall was washed with 1mL DMF, added to the reaction mixture, stirred at room temperature overnight, dichloromethane was concentrated to remove dichloromethane, the resulting crude product was diluted with ethyl acetate (10mL), washed with water (10mL × 2), the organic phase was washed with brine, part of ethyl acetate was concentrated, a precipitate was precipitated, filtered, the cake was washed twice with dichloromethane, and dried to give compound M19(55mg, 39.0%, grey solid).¹H NMR(300MHz,DMSO-d₆):δ10.41(s,1H),8.44(d,J＝9.0Hz, 1H),8.33(d,J＝15.6Hz,1H),8.22(s,1H),8.06(s,1H),8.03-7.98(m,2H),7.89 (d,J＝16.5Hz,1H),7.66(d,J＝7.8Hz,1H),7.61(d,J＝8.7Hz,1H),7.54(s, 1H),7.50-7.36(m,4H),6.86(d,J＝15.6Hz,1H),4.41(q,J＝6.9Hz,2H),1.38 (t,J＝6.9Hz,3H).

Compound M19(50mg,0.085mmol) was added to a flask, ethanol (5mL), sodium hydroxide (22mg,0.55mmol) was added, reaction was carried out at 60 ℃ for 18 hours, concentration was carried out to remove ethanol, dilution was carried out with 2mL of water, pH was adjusted to 2 with 1N hydrochloric acid, 5mL of ethyl acetate was added, stirring was carried out, filtration was carried out, and the filter cake was dried to obtain CHY113(24mg, 53.5%, red solid).¹H NMR(300MHz,DMSO-d₆):δ10.40(s, 1H),8.55(d,J＝8.7Hz,1H),8.36(d,J＝15.6Hz,1H),8.27(s,1H),8.11(s,1H), 8.09-8.06(m,2H),7.98(d,J＝16.5Hz,1H),7.69-7.64(m,2H),7.51-7.34(m, 5H),6.97(d,J＝15.6Hz,1H).

The following compounds were synthesized using a procedure similar to that of preparation example four:

wherein commercially unavailable indole derivatives can be prepared by:

method A see the synthesis of compound M3 in preparation example I, and 4, 6-difluoroindole-2-carboxylic acid ethyl ester can also be synthesized by this method.

The method B comprises the following steps:

the compound M21 is synthesized by taking o-anisidine as a raw material and adopting a method for synthesizing the compound M2.

¹H NMR(300MHz,CDCl₃):δ8.12(s,1H),7.59(d,J＝7.5Hz,1H), 6.70-6.85(m,3H),4.32(q,J＝7.2Hz,2H),3.90(s,3H),2.35(s,3H),1.38(t,J＝ 7.2Hz,3H).

Dissolving compound M21(760mg,3.22mmol) in acetic acid (5mL), adding toluene (5mL), zinc chloride (1.088g,8.09mmol), reacting at 110 deg.C for 12 hr, concentrating toluene, adding water and diethyl ether each 10mL, adjusting pH to neutral with 2N sodium hydroxide, separating layers, and extracting the aqueous phase with diethyl ether (10mL)Collecting, mixing organic phases, washing with water, saturated salt water, and anhydrous MgSO₄Drying, concentration and purification by silica gel column chromatography (PE/EtOAc 50:1to 20:1) gave compound M22(200mg, 28.4%, white solid).¹H NMR (300MHz,CDCl₃):δ9.04(bs,1H),7.27(d,J＝8.1Hz,1H),7.20(s,1H),7.06(t, J＝8.1Hz,1H),6.72(d,J＝7.5Hz,1H),4.41(q,J＝7.2Hz,2H),3.96(s,3H), 1.41(t,J＝7.2Hz,3H).

The method C comprises the following steps:

ethyl pyruvate (3.19g,27.59mmol) was dissolved in ethanol (60mL), acetic acid (517mg, 8.62mmol) and p-methoxyphenylhydrazine hydrochloride (3g,17.24mmol) were added, a condenser tube was added and the reaction was carried out at 80 ℃ for 2 hours, the solid gradually dissolved in a black solution, the reaction was continued for 3 hours, ethanol was concentrated to precipitate a white solid, filtration was carried out, and the filtrate was purified by silica gel column chromatography (PE/EtOAc 25:1to 10:1) to obtain an intermediate point, i.e., compound M24(456mg, 12.1%, white solid).¹H NMR(300MHz,CDCl₃):δ 11.72(bs,1H),7.32(d,J＝8.7Hz,1H),7.07(d,J＝2.1Hz,1H),7.02(s,1H), 6.89(dd,J＝9.0,2.4Hz,1H),4.30(q,J＝7.2Hz,2H),3.73(s,3H),1.30(t,J＝ 7.2Hz,3H).

The method D comprises the following steps:

dissolving ethyl pyruvate (3.1g,26.82mmol) in ethanol (60mL), adding acetic acid (498mg, 8.38mmol) and o-chlorophenylhydrazine hydrochloride (3g,17.76mmol), adding a condenser tube, reacting at 80 ℃ for 2 hours, cooling to room temperature, concentrating ethanol, precipitating a solid, filtering, washing a filter cake with ethanol for 2 times to obtain 1.36g of colorless needle-shaped crystals, concentrating a mother solution to precipitate a solid, filtering, washing the filter cake with ethanol to obtain 660mg of colorless needle-shaped crystals, wherein the yield is 47.3%.¹H NMR(300MHz,DMSO-d₆)：δ12.30(s,1H),7.55 (d,J＝8.4Hz,1H),7.42(d,J＝7.8Hz,1H),7.31(t,J＝7.5Hz,1H),6.93(d,J＝ 7.5Hz,1H),4.27(q,J＝7.2Hz,2H),2.13(s,3H),1.29(t,J＝7.2Hz,3H)。

Adding compound M26(1g, 4.15mmol) into a bottle, adding polyphosphoric acid (10g), reacting at 45 deg.C for 1 hr, adding ice water, stirring for 10 min, extracting with diethyl ether, and extracting with anhydrous MgSO₄Drying, concentration and purification by silica gel column chromatography (PE/EtOAc 50:1) afforded compound M27(300mg, 32%, white solid).¹H NMR(300MHz,DMSO-d₆):δ12.13(s,1H),7.65(d,J＝7.8Hz,1H),7.35(d, J＝7.5Hz,1H),7.26(s,1H),7.09(t,J＝8.1Hz,1H),4.35(q,J＝6.9Hz,2H), 1.34(t,J＝7.2Hz,3H)。

The method E comprises the following steps:

sodium azide (1.08g, 16.62mmol) was added to a bottle, DMF (4.5mL) was added, Compound M28(2.5g,14.97mmol) was added dropwise at room temperature, reaction was carried out at room temperature for 2 hours, water was added to dissolve the white precipitate, ether (10 mL. times.3) was extracted, the organic phases were combined, washed with water (10 mL. times.6), saturated brine, anhydrous MgSO 2₄After drying and concentration, compound M29(1.66g, 85.9%, colorless oil) was obtained.¹H NMR(300MHz, CDCl₃):δ4.26(q,J＝7.2Hz,2H),3.86(s,2H),1.30(t,J＝7.2Hz,3H).

Metallic sodium (280mg,12.17mmol) was added in portions to anhydrous methanol (9.5mL) at room temperature, after the sodium lumps disappeared, refluxing was carried out for 1 hour, cooling was carried out to-20 ℃, a methanol solution (2.5mL) of p-methoxybenzaldehyde (500mg,3.67 mmol) was added dropwise, then compound M29(1.184g,9.18mmol) was added dropwise, after reaction for half an hour at-20 ℃, reaction was carried out for 12 hours at-3 ℃, ice water was added, a white precipitate was precipitated by stirring, filtration was carried out, the filter cake was washed with ice water 2 times, and drying was carried out to obtain compound M31(412mg, 48.1%, white solid).¹H NMR(300MHz,CDCl₃):δ7.79(d,J＝7.5Hz,2H),6.91(d,J＝9.0Hz,2H), 6.89(s,1H),3.90(s,3H),3.84(s,3H).

Dissolving compound M31(412mg, 1.77mmol) in M-xylene (50mL), refluxing under argon at 160 deg.C for half an hour, concentrating under reduced pressure to remove M-xylene, dissolving crude product with dichloromethane, and purifying by silica gel column chromatography (PE/EtOAc 5:1) to obtain the final productCompound M32(359mg, 98.9%, white solid). ESIMS: 228.1[ M + Na ]]⁺。

¹H NMR(300MHz,Acetone-d₆):δ10.70(bs,1H),7.55(d,J＝9.0Hz,1H), 7.11(s,1H),7.00(s,1H),6.77(dd,J＝9.0,2.1Hz,1H),3.86(s,3H),3.83(s, 3H).

Preparation example five (Compound No. GP523)

Dissolving compound M33(639mg,3mmol), p-nitrophenol (500.8mg,3.6mmol) and potassium carbonate (829mg,6mmol) in DMF (15mL), reacting overnight at 60 deg.C, detecting by TLC that the raw materials disappear, diluting with water, extracting with ethyl acetate for 2 times, combining the organic phases, washing the organic phase with water for 3 times, washing with saturated common salt water, and anhydrous Na₂SO₄Drying, concentration and purification by silica gel column chromatography (PE/EtOAc 10:1) afforded compound M34(850mg, > 100% yield, colorless solid). ESIMS of 272.1[ M + H ]]⁺。

¹H NMR(300MHz,CDCl₃):δ8.19(d,J＝9.0Hz,2H),7.32-7.19(m,5H), 6.92(d,J＝9.3Hz,2H),4.06(t,J＝6.0Hz,2H),2.71(t,J＝6.9Hz,2H),1.83(m, 4H).

Compound M34(170mg,0.63mmol) was dissolved in methanol (10mL), and after 3 times replacement with nitrogen, Pa/C (20mg) was added, and after 3 times replacement with hydrogen, the reaction was carried out at room temperature for 50 minutes, and the reaction was completed by TLC. Filtration through celite and concentration to remove the solvent afforded compound M35(136mg, 89.5%, colorless liquid).

ESIMS:242.1[M+H]⁺。¹H NMR(300MHz,CDCl₃):δ7.31-7.18(m,5H), 6.74(d,J＝8.7Hz,2H),6.64(d,J＝8.7Hz,2H),3.90(t,2H),2.67(t,2H),1.79 (m,4H).

Dissolving compound M18(65mg, 0.20mmol) in dry DMF (4mL), adding compound M36(62mg, 0.28mmol) and triphenylphosphine (73mg, 0.28mmol) under nitrogen protection, reacting at room temperature for 3 hours, detecting many raw materials by TLC, adding 10 equivalents of compound M36 and triphenylphosphine, reacting for 2 hours, adding compound M35(67mg, 0.28mmol), reacting at 80 deg.C overnight, adding water, extracting with ethyl acetate, collecting the organic phase, filtering, concentrating the filtrate, purifying by silica gel column chromatography (PE/Acetone ═ 5:1) to obtain crude product, adding methanol to precipitate solid, filtering, and mixing the solid to obtain compound M37(5mg, 4.5%, white solid).

¹H NMR(300MHz,CD₃OD):δ8.40(d,J＝15.6Hz,1H),7.63(s,1H),7.55 (d,J＝8.7Hz,2H),7.41(d,J＝1.8Hz,1H),7.27-7.13(m,5H),6.85(d,J＝9.0 Hz,2H),6.78(d,J＝15.6Hz,1H),4.41(q,J＝6.9Hz,2H),3.96(t,2H),2.67(t, 2H),1.79(m,4H),1.41(t,J＝7.2Hz,3H).

Compound M37(5mg,0.0091mmol) was dissolved in isopropanol, hydrolyzed with sodium hydroxide (2.5mg,0.063 mmol), adjusted to pH 2 with 1N hydrochloric acid, concentrated to remove isopropanol, added with water to precipitate a solid, filtered and dried to give the product GP523(2mg, 41.7%, yellow solid).

¹H NMR(300MHz,DMSO-d₆):δ12.54(s,1H),10.07(s,1H),8.23(d,J＝ 15.6Hz,1H),7.62(d,J＝9.3Hz,1H),7.48(s,1H),7.39(s,1H),7.31-7.17(m, 5H),6.88(d,J＝9.0Hz,2H),6.75(d,J＝15.6Hz,1H),3.96(s,2H),2.64(s,2H), 1.71(s,4H).

Preparation example six (Compound No. CHY110)

Dissolving compound M38(500mg, 2.07mmol) in benzene (10mL), adding triphenylphosphine (580mg, 2.21mmol) solution in benzene (10mL) dropwise at room temperature, reacting at room temperature for 48 hr to precipitate white precipitate, filtering, washing filter cake with n-hexane, dissolving obtained solid in dichloromethane (30mL), adding 1% sodium hydroxide aqueous solution (30mL), stirring at room temperature for 30 min, separating layers, and collecting organic phase with anhydrous MgSO₄After drying and concentration, compound M39 (colorless oil) was obtained and directly charged into the next reaction.¹H NMR(300 MHz,CDCl₃):δ7.71-7.41(m,20H),7.27(s,1H),3.00(t,J＝7.2Hz,2H),2.65(t, J＝7.2Hz,2H).

Compound M4(20mg, 0.078mmol) was dissolved in anhydrous toluene (5mL), compound M39(41mg, 0.10mmol) was added, reacted overnight at 120 ℃, diluted with dichloromethane and purified by silica gel column chromatography (PE/EtOAc ═ 6:1) to give compound M40(22mg, 68.8%, white solid).

¹H NMR(300MHz,CDCl₃):δ9.47(bs,1H),8.31(d,J＝16.2Hz,1H), 7.34-7.20(m,7H),6.82(d,J＝16.5Hz,1H),4.42(q,J＝7.2Hz,2H),3.05(s, 4H),1.39(t,J＝7.2Hz,3H).

Adding a compound M40(22mg, 0.053mmol) into a bottle, adding ethanol (2mL) and lithium hydroxide monohydrate (8mg, 0.20mmol), reacting at 50 ℃ for 3.5 hours, detecting by TLC that the reaction is not finished, supplementing 1N LiOH (1mL), reacting at 50 ℃ for 1 hour, detecting by TLC that the reaction is complete, concentrating to remove ethanol, adding water for dilution, adjusting the pH to 2 by using 1N hydrochloric acid, filtering, and drying a filter cake to obtain a product CHY110(9mg, 43.7%, yellow solid).

¹H NMR(300MHz,Acetone-d₆):δ11.63(bs,1H),8.42(d,J＝16.5Hz,1H), 7.61(s,1H),7.30-7.15(m,6H),6.96(d,J＝16.5Hz,1H),3.08-2.96(m,4H).

Preparation example seven (Compound No. CHY112)

Adding compound M5(300mg, 1.16mmol) and cuprous chloride (53mg, 0.54mmol) into microwave tube, adding quinoline (2mL), purging with argon, reacting at 200 deg.C for 10 min, adding ethyl acetate (15mL), washing with 1N hydrochloric acid twice, the organic phase is milky, filtering, washing the filtrate with saturated saline, MgSO 2₄Drying, concentration and purification by silica gel column chromatography (PE/EtOAc 1:1) afforded compound M41(124mg, 50.0%, grey solid).

¹H NMR(300MHz,DMSO-d₆):δ10.42(s,1H),8.34(s,1H),7.59(s,1H), 7.41(s,1H).

Compound M41(63mg, 0.29mmol), malonic acid (59mg, 0.57mmol), pyridine (1.7mL), piperidine (246mg, 2.89mmol) were added sequentially to a bottle, reacted at 50 ℃ for 12 hours, concentrated to remove the solvent, and purified by silica gel column chromatography (PE/EtOAc ═ 1:1+ 1% formic acid) to give compound M42(50mg, 67.6%, grey solid).

¹H NMR(300MHz,Acetone-d₆):δ11.21(bs,1H),10.46(bs,1H),8.57(d,J ＝15.9Hz,1H),8.16(s,1H),7.53(s,1H),7.21(s,1H),6.36(d,J＝15.9Hz,1H).

Compound M37(100mg, 0.34mmol) was dissolved in anhydrous DMF (2.5mL), triphenylphosphine (108mg, 0.41mmol), NBS (72mg, 0.41mmol) were added under ice-water bath, reaction was carried out at room temperature for 3 hours, the solution became cloudy by clarification, ethyl acetate (10mL) was added, and the mixture was washed with 0.5N lithium hydroxide (10mL), water (10mL), saturated brine (10mL), MgSO 2₄Drying, concentration and purification by silica gel column chromatography (PE/EtOAc ═ 20:1) gave compound M43(66mg, 54.1%, light yellow solid). ESIMS: 357.1, 359.1.

¹H NMR(300MHz,CDCl₃):δ8.13-8.08(m,2H),7.74-7.56(m,6H), 7.47-7.35(m,3H),4.54(s,2H).

Compound M42(35mg, 0.14mmol) was dissolved in anhydrous DMF (3mL), potassium carbonate (39mg, 0.28mmol) was added under ice water bath, after stirring for 15 minutes, compound M43(50mg, 0.14mmol) was added, the solution turned from clear to yellow cloudy after overnight at room temperature, water and ethyl acetate (10mL each) were added to give a number of white flocs, which were filtered and dried to give product CHY112(20mg, 26.8%, white solid).

¹H NMR(300MHz,DMSO-d₆):δ8.51(d,J＝15.9Hz,1H),8.41(d,J＝8.4 Hz,1H),8.33(s,1H),8.02(s,1H),8.01(d,J＝8.4Hz,1H),7.94-7.87(m,3H), 7.80(s,1H),7.73(d,J＝6.9Hz,1H),7.60(d,J＝8.1Hz,1H),7.54(s,1H), 7.50-7.48(m,2H),7.43(t,J＝6.3Hz,1H),7.24(s,1H),6.54(d,J＝15.9Hz,1H), 5.26(s,2H).

Preparation example eight (Compound No. CHY117)

Starting from compound M44, compound M47 was synthesized by the method for synthesizing M18 in preparation example four.

¹H NMR(300MHz,DMSO-d₆):δ12.34(s,1H),8.54(d,J＝16.2Hz,1H), 7.99(d,J＝8.1Hz,1H),7.55(d,J＝8.1Hz,1H),7.37(d,J＝8.1Hz,1H),7.24 (d,J＝8.1Hz,1H),6.57(d,J＝16.5Hz,1H),4.41(q,J＝7.2Hz,2H),1.39(t,J ＝7.2Hz,1H).

Compound M47(100mg, 0.39mmol) was dissolved in a mixed solution of ethanol (5mL) and tetrahydrofuran (8mL), and after 3 times replacement with nitrogen, Pa/C (29mg) was added, and 2 times replacement with hydrogen was performed, reaction was performed at room temperature for 7 hours, and filtration was performed with celite, and the solvent was concentrated off to give compound M48(74mg, 73.3%, white solid).

ESIMS：260.1[M-H]⁺；¹H NMR(300MHz,Acetone-d₆):δ10.69(bs,1H), 7.77(d,J＝8.7Hz,1H),7.49(d,J＝8.4Hz,1H),7.28(ddd,J＝8.1,7.2,1.2Hz, 1H),7.10(ddd,J＝7.2,6.9,0.9Hz,1H),4.38(q,J＝7.2Hz,2H),3.41(t,J＝8.1 Hz,2H),2.66(t,J＝7.5Hz,2H),1.38(t,J＝7.2Hz,3H).

Compound M48 can be converted to CHY117 as a product by the method of preparation example IV for the synthesis of CHY 113.

ESIMS：494.2[M-H]⁺；¹H NMR(300MHz,DMSO-d₆):δ11.39(s,1H), 10.01(s,1H)，8.40(d,J＝8.7Hz,1H),8.17(s,1H),8.04(s,1H)，8.02-7.92(m, 3H),7.82(d,J＝16.5Hz,1H),7.74(d,J＝7.8Hz,1H),7.59(dd,J＝8.7,2.4Hz, 1H),7.50(d,J＝7.5Hz,1H),7.40-7.32(m,5H),7.21(t,J＝7.2Hz,1H),7.04(t, J＝7.2Hz,1H),3.40(t,J＝7.2Hz,2H),2.69(t,J＝7.5Hz,2H).

Preparation example nine (Compound No. CHY118)

Adding a compound M45(200mg, 0.92mmol) into a bottle, adding water (2mL) and acetonitrile (10mL) to form a white turbid solution, adding sodium dihydrogen phosphate dihydrate (186mg, 1.20mmol) and 30% hydrogen peroxide (125mg, 1.10mmol), dropwise adding an aqueous solution (0.5mL) of sodium chlorite (116mg, 1.29mmol) under an ice-water bath condition, reacting for 7 hours at room temperature, detecting that the reaction is not finished by TLC, adding water (2mL) and tetrahydrofuran (10mL), reacting overnight at room temperature, adjusting the pH to 2 by using 1N hydrochloric acid under an ice-water bath, precipitating a white precipitate, filtering, washing a filter cake for 2 times, and drying to obtain a compound M50(177mg, 82.7% and a white solid).

¹H NMR(300MHz,DMSO-d₆):δ12.71(s,1H),12.48(s,1H)，7.99(d,J＝ 8.4Hz,1H),7.49(d,J＝8.1Hz,1H),7.31(t,J＝7.8Hz,1H),7.20(d,J＝7.2Hz, 1H),4.37(q,J＝6.9Hz,1H),1.33(t,J＝7.2Hz,1H).

Compound M50 was converted to product CHY118 using the procedure for the synthesis of CHY113 in preparation example four.

¹H NMR(300MHz,DMSO-d₆):δ12.36(s,1H),11.63(s,1H)，8.43(d,J＝ 8.4Hz,1H),8.26(d,J＝8.1Hz,1H),8.20(s,1H)，8.06-7.99(m,3H),7.91(d,J＝ 16.2Hz,1H),7.68(d,J＝7.2Hz,1H),7.61(d,J＝8.7Hz,1H),7.57-7.42(m, 5H),7.36(t,J＝8.1Hz,1H),7.23(t,J＝7.2Hz,1H).

Preparation example ten (Compound No. CHY127)

Propargyl alcohol (560mg, 10mmol) was dissolved in dioxane (27mL), silver carbonate (184 mg, 0.67mmol) was added, heated to 80 ℃, compound 46(753mg, 6.66mmol) was slowly added dropwise, reacted at 80 ℃ for 2 hours, the solvent was concentrated off, dissolved with dichloromethane (20mL), the black insoluble material was filtered off with celite, and the filtrate was concentrated and purified by silica gel column chromatography (PE/EtOAc 10: 1to 5:1to 3:1) to give compound M51(53mg, 13.6%, white solid).¹H NMR(300MHz,CDCl₃):δ9.07(bs,1H), 6.85(s,1H),6.25(s,1H),4.75(d,J＝6.6Hz,1H),4.36(q,J＝7.2Hz,2H),3.52 (bs,1H),1.28(t,J＝8.1Hz,3H).

Compound M51(153mg, 0.91mmol) was dissolved in dichloromethane (4mL), PCC (291 mg, 1.36mmol) was added, reaction was carried out at room temperature for 2 hours, filtration was carried out, and the filtrate was concentrated and purified by silica gel column chromatography (PE/EtOAc ═ 5:1) to give compound M52(100mg, 65.8%, white solid).¹H NMR(300 MHz,CDCl₃):δ10.49(s,1H),9.48(bs,1H),6.94(t,J＝2.4Hz,1H),6.81(t,J＝ 3.0Hz,1H),4.44(q,J＝7.2Hz,2H),1.42(t,J＝7.2Hz,3H).

The procedure for converting compound M52 into product CHY127 is described in preparation example four.

ESIMS：442.0[M-H]⁺；¹H NMR(300MHz,DMSO-d₆):δ11.66(bs,1H), 10.13(s,1H),8.43-8.33(m,2H),8.14(s,1H),8.06-7.94(m,3H),7.85(d,J＝ 16.2Hz,1H),7.64-7.58(m,2H),7.44-7.39(m,3H),6.90(s,1H),6.54(d,J＝ 15.6Hz,1H),6.46(s,1H).

Examples of biological experiments

Firstly, the method comprises the following steps: GPR17, CysLT₁,CysLT₂Antagonistic Activity test experiment

1. Purpose of the experiment: GPR17, CysLT, for carrying out the Compounds of the invention₁,CysLT₂And (4) testing antagonistic activity.

2. Cells used for the experiments: stable expression of GPR17/G alpha 16, CysLT₁/Gα16,CysLT₂HEK293 cells of/G alpha 16 as a cell model for high-throughput screening and detection of compound antagonistic activity.

3. The experimental principle is as follows: by establishing a cell line for cotransforming a target receptor and the G alpha 16 protein, the activation of the G alpha 16 protein can be caused after the target receptor is activated, and then phospholipase C (PLC) is activated to generate IP₃And DAG, IP₃Can interact with IP on endoplasmic reticulum and mitochondria in cells₃Receptor binding, resulting in the release of intracellular calcium ions. Therefore, measurement of changes in intracellular calcium ion concentration can be used as a method for detecting the activation state of a target receptor. Fluo-4/AM is a calcium fluorescent probe indicator used for measuring calcium ions, is used as a nonpolar fat-soluble compound, and after entering cells, under the action of cell lipolytic enzyme, an AM group is dissociated to release Fluo-4; since Fluo-4 is a polar molecule and does not readily pass through a lipid bilayer membrane, it can retain Fluo-4 in the cell for a long period of time. The level of activation of the G.alpha.protein can ultimately be reflected by measuring the intensity of the fluorescence that is excited. If the compound to be screened is capable of agonizing the target receptor, the calcium flux response can be greatly increased. On the contrary, if the compound to be screened is capable of antagonizing the target receptorAnd the calcium flow reaction can be greatly reduced.

Experimental reagent: BSA (Roche), D-glucose (national pharmaceutical group chemical Co., Ltd.), sulfipyrazone (Sigma), Cremophor EL (Biochemika), Fluo-4, AM (Invitrogen), montelukast (TCI), pranlukast (TCI), zafirlukast (TCI), MDL29951(TCI).

4. The experimental steps are as follows:

sample treatment: the samples were dissolved in DMSO and stored at low temperature, and the concentration of DMSO in the final system was controlled within a range that did not affect the detection activity.

HEK293 cells stably expressing the target receptor/G α 16 were plated in 96-well plates and cultured overnight. The culture medium in the wells seeded with cells was aspirated, 40 mL/well of freshly prepared dye was added, and incubation was carried out in an incubator at 37 ℃ for 40 minutes. The drug to be tested was diluted with calcium buffer and mixed well. The dye was removed by blotting, washed once with freshly prepared calcium buffer, and replaced with 50mL of calcium buffer containing the drug to be tested dissolved. The fluorescence at 525nm was read by the FlexStation II instrument by automatically adding 25mL of calcium buffer with known agonist dissolved at 15 th second.

Thirdly, experimental results:

the results of the positive controls Montelukast, Pranlukast, Zafirlukast indicate that the test method and experimental results are reliable. Based on the results of the above cell level tests, the present invention provides a novel class of GPR17 and CysLT₁Dual receptor antagonists with micromolar antagonist activity for GPR17, on CysLT₁And CysLT₂Has selective antagonistic effect, and the antagonistic activity is at nanomolar level.

Claims (10)

1. a compound shown in general formula I,

Wherein, Y is -O- or -NH-; X is C ₂ -C ₆ alkenylene;

is a benzene ring; _R1 is the

Substituted 5- to 8-membered aryl; R ₄ is hydrogen, C ₁ -C ₄ alkyl, C ₁ -C ₄ alkoxy, halogen atom, hydroxyl or amino; R ₇ is -(CH ₂ ) _n COOH, wherein n is an integer from 0 to 3; R _5b and R _6b are absent, R _5a and R _6a are each independently selected from H, halogen atom, hydroxyl, carboxyl, amino, C ₁ -C ₁₀ straight or branched chain alkyl, C ₁ -C ₁₀ straight chain or Branched alkoxy, C ₂ -C ₈ alkenyl, C ₂ -C ₆ alkynyl, C ₁ -C ₆ alkylamino. 2. The compound of claim 1, wherein R _5a and R _6a are each independently H, a halogen atom, a C ₁ -C ₆ alkoxy group, 3. The compound of claim 1, wherein X is -CH=CH-. 4. The compound of claim 1, wherein, _R1 is

5. A compound having the structure shown in the following structural formula:

6. a preparation method of the compound shown in general formula I, it comprises the steps:

a) The compound represented by general formula V undergoes esterification reaction with R ₁ OH under the conditions of DCC and DMAP; Or the compound represented by general formula V and R ₁ NH ₂ undergo condensation reaction in the presence of a condensing agent; Or the compound represented by general formula V and R ₁ Br undergo a nucleophilic substitution reaction under basic conditions; Or the compound represented by general formula V reacts with R ₁ OTs in the presence of cesium carbonate; When R ₉ is not the same as R ₇ , it further comprises step b) b) _deprotecting R to give compounds of general formula I; Wherein, X is -CH=CH-; Y is -O- or -NH-; R ₉ is hydrogen, -(CH ₂ ) _n COOH or -(CH ₂ ) _n COR ₁₀ , R ₁₀ is a carboxyl protecting group; R ₇ is H, -(CH ₂ ) _n COOH; R ₁ , R _5a , R _6a , R _5b , R _6b ,

The definitions of and n are the same as those in claim 1 . 7. preparation method according to claim 6, wherein, the compound shown in described general formula V is prepared by following steps:

c) the aldehyde group in general formula VI is oxidized by oxidizing agent to obtain the compound of general formula V; or d) Horner-Wadsworth-Emmons reaction occurs between the compound shown in general formula VI and tert-butoxycarbonyl methylene triphenylphosphine; then the compound shown in general formula V is obtained through selective hydrolysis of tert-butyl ester, or e) the aldehyde group in general formula VI and malonic acid react under the action of weak base to obtain the compound shown in general formula V; Wherein, X is -CH=CH-; R ₉ is hydrogen, -(CH ₂ ) _n COOH or -(CH ₂ ) _n COR ₁₀ , and R ₁₀ is a carboxyl protecting group; R _5a , R _6a , R _5b , R _6b ,

The definitions of and n are the same as those in claim 1 . 8. A pharmaceutical composition comprising a therapeutically effective amount of one or more compounds represented by the general formula I of claim 1 and a pharmaceutically acceptable carrier. 9. Use of the compound according to any one of claims 1 to 5 or the pharmaceutical composition according to claim 8 in the preparation of a medicament for preventing or treating damage to the central nervous system and neurodegenerative diseases or in the preparation of a treatment Use in the medicament of CysLT ₁ mediated diseases. 10. The use according to claim 9, wherein the central nervous system injury and neurodegenerative disease is multiple sclerosis or leukodystrophy; and the CysLT ₁ mediated disease is asthma, rhinitis, Allergies or other inflammatory conditions.