CN112480117A

CN112480117A - Pyrrolopyrazole derivative, preparation method and medical application thereof

Info

Publication number: CN112480117A
Application number: CN201910866374.4A
Authority: CN
Inventors: 任峰; 徐咏梅; 王显连; 陈春麟; 蔡金娜
Original assignee: Medicilon Puya Medical Technology (shanghai) Co ltd; Shanghai Medicilon Inc
Current assignee: Medicilon Puya Medical Technology (shanghai) Co ltd; Shanghai Medicilon Inc
Priority date: 2019-09-12
Filing date: 2019-09-12
Publication date: 2021-03-12
Anticipated expiration: 2039-09-12
Also published as: CN112480117B

Abstract

The invention relates to a pyrrolopyrazole derivative, a preparation method thereof and application thereof in medicines. In particular, the invention relates to a novel pyrrolopyrazole derivative shown in a general formula (I), a preparation method thereof, a pharmaceutical composition containing the derivative and the application of the derivative as a therapeutic agent, in particular as a gastric acid secretion inhibitor and a potassium ion competitive agentThe use of competitive acid blockers (P-CABs) in biomedicine. Wherein each substituent (R) in the general formula (I)¹、R²、R³、R⁴) The definitions are the same as those in the specification.

Description

Pyrrolopyrazole derivative, preparation method and medical application thereof

Technical Field

The present invention relates to a new class of pyrrolopyrazole derivatives, to a process for their preparation and to their use as therapeutic agents, in particular as inhibitors of gastric acid secretion and potassium ion competitive acid blockers (P-CABs), or pharmaceutical compositions containing them.

Background

Peptic ulcer refers to mainly chronic ulcer occurring in stomach and duodenum. Although there are regional differences, the incidence of peptic ulcer usually accounts for 10% to 20% of the total population, and is a frequently occurring and common disease. Ulcer formation is due to various factors, and the digestive action of acidic gastric juice on the mucosa is the basic factor for ulcer formation. Therefore, inhibition of gastric acid secretion is becoming the first choice for the treatment of peptic ulcer diseases.

Since omeprazole, the first Proton Pump Inhibitors (PPIs) in 1988, several PPIs have been marketed worldwide to date, including lansoprazole, pantoprazole, rabeprazole, esomeprazole, and the like. PPIs have become the first choice drugs for the treatment of gastric acid related diseases including peptic ulcer, reflux esophagitis and zollinger-ellison syndrome. The Proton Pump (Proton Pump) is substantially H⁺/K⁺Adenosine triphosphatase (H)⁺/K⁺ATPase), which specifically pumps protons (H +) into the gastric lumen to form a strong acidity in the stomach. Proton pump inhibitors may inhibit the activity of proton pumps and thereby modulate proton pump mediated gastric acid secretion.

Potassium-Competitive Acid Blockers (Potassium-Competitive Acid blocks,P-CABs) are a novel class of gastric acid blockers by reversible, interaction with potassium ion (K)⁺) Competitive binding H⁺/K⁺ATPase thus acting to inhibit H⁺/K⁺-the effect of ATPase enzymatic activity. Compared with PPIs, P-CABs have the characteristics of lipophilicity, alkalescence, stability under acidic (low pH) conditions and the like. Meanwhile, the P-CABs have the advantages of quick response, easy acid inhibition effect and the like.

The first new P-cab drug, voronorazan, was marketed in japan in 2014 for the treatment of gastric acid related diseases such as peptic ulcers. A series of potassium ion competitive acid blocker structures have also been disclosed. There is still a need to develop new compounds with a wide variety of structural types that are better drug-forming.

Disclosure of Invention

In view of the above problems, the present invention aims to provide a compound for treating gastric acid-related diseases such as peptic ulcer, which is of a novel structural type and has excellent effects and actions.

In one aspect, the invention provides a compound of formula (I) or a pharmaceutically acceptable salt thereof,

wherein:

R¹selected from hydrogen atoms, halogens or alkyl groups;

R²selected from hydrogen, halogen, hydroxyl or alkyl;

R³selected from hydrogen atoms, halogens, hydroxyl groups, alkyl groups or alkoxy groups;

R⁴selected from OR^aOr NR^bR^cWherein R is^aSelected from hydrogen atoms or alkyl radicals, R^b、R^cEach independently selected from a hydrogen atom, a hydroxyl group, an amine group, an alkoxy group, an optionally substituted alkyl group.

Preferably, R¹Selected from halogens;

R²selected from hydrogen atoms, halogens or hydroxyl groups;

R³selected from a hydrogen atom, a fluorine atom or a hydroxyl group;

R⁴selected from OR^aOr NR^bR^cWherein R is^aSelected from hydrogen atoms or C_1～3Alkyl radical, R^b、R^cEach independently selected from hydrogen atom, hydroxyl group, optionally substituted C_1～3An alkyl group.

Preferably, R¹Selected from halogens;

R²selected from hydrogen atoms, halogens or hydroxyl groups;

R³selected from a hydrogen atom, a fluorine atom or a hydroxyl group;

R⁴selected from OR^aOr NR^bR^cWherein R is^aSelected from hydrogen atoms or C_1～3Alkyl radical, R^b、R^cEach independently selected from hydrogen atom, hydroxy, optionally substituted by one hydroxy, amino or-CONH₂Substituted C_1～3An alkyl group.

Preferably, R¹Is a fluorine atom;

R²selected from a hydrogen atom, a fluorine atom or a hydroxyl group;

R³is a hydrogen atom;

R⁴selected from OR^aOr NR^bR^cWherein R is^aSelected from hydrogen atoms or methyl radicals, R^b、R^cEach independently selected from hydrogen atom, hydroxy group, -CH₂CONH₂。

Preferably, the compound is selected from:

3 '- ((3- (2-fluorophenyl) -5-methyl-5, 6-dihydropyrrolo [3,4-c ] pyrazol-2 (4H) -yl) methyl) - [1,1' -biphenyl ] -3-carboxamide;

3 '- ((3- (2-fluorophenyl) -5-methyl-5, 6-dihydropyrrolo [3,4-c ] pyrazol-2 (4H) -yl) methyl) - [1,1' -biphenyl ] -3-carboxylic acid;

3 '- ((3- (2-fluorophenyl) -5-methyl-5, 6-dihydropyrrolo [3,4-c ] pyrazol-2 (4H) -yl) methyl) -N-hydroxy- [1,1' -biphenyl ] -3-carboxamide;

n- (2-amino-2-oxoethyl) -3 '- ((3- (2-fluorophenyl) -5-methyl-5, 6-dihydropyrrolo [3,4-c ] pyrazol-2 (4h) -yl) methyl) - [1,1' -biphenyl ] -3-carboxamide;

5 ' - ((3- (2-fluorophenyl) -5-methyl-5, 6-dihydropyrrolo [3,4-c ] pyrazol-2 (4h) -yl) methyl) -2' -hydroxy- [1,1' -biphenyl ] -3-carboxylic acid methyl ester;

5 ' - ((3- (2-fluorophenyl) -5-methyl-5, 6-dihydropyrrolo [3,4-c ] pyrazol-2 (4H) -yl) methyl) -2' -hydroxy- [1,1' -biphenyl ] -3-carboxylic acid;

5 ' - ((3- (2-fluorophenyl) -5-methyl-5, 6-dihydropyrrolo [3,4-c ] pyrazol-2 (4H) -yl) methyl) -N, 2' -dihydroxy- [1,1' -biphenyl ] -3-carboxamide;

2' -fluoro-5 ' - ((3- (2-fluorophenyl) -5-methyl-5, 6-dihydropyrrolo [3,4-c ] pyrazol-2 (4H) -yl) methyl) - [1,1' -biphenyl ] -3-carboxylic acid;

3 ' - ((3- (2-fluorophenyl) -5-methyl-5, 6-dihydropyrrolo [3,4-c ] pyrazol-2 (4H) -yl) methyl) -5 ' -hydroxy- [1,1' -biphenyl ] -3-carboxylic acid.

In a second aspect, the present invention provides a pharmaceutical composition comprising a compound represented by formula (I) or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier, excipient or diluent.

In a third aspect, the invention provides a compound shown in the general formula (I) or a pharmaceutically acceptable salt thereof or an application of the pharmaceutical composition in preparing a gastric acid secretion inhibitor.

In a fourth aspect, the present invention provides a compound of formula (I) or a pharmaceutically acceptable salt thereof or a pharmaceutical composition as described above in the preparation of H⁺/K⁺Adenosine triphosphatase (H)⁺/K⁺-ATPase) inhibitors.

In a fifth aspect, the invention provides the use of a compound represented by the general formula (I), or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition described above, in the preparation of potassium ion competitive acid blockers (P-CABs).

In a sixth aspect, the present invention provides the use of a compound represented by the general formula (I) or a pharmaceutically acceptable salt thereof or a pharmaceutical composition as described above for the manufacture of a medicament for the treatment and/or prevention of peptic ulcer, zollinger-ellison syndrome, gastritis, erosive esophagitis, reflux esophagitis, symptomatic gastroesophageal reflux disease, barrett's esophagitis, functional dyspepsia, helicobacter pylori infection, gastric cancer, gastric MALT lymphoma, ulcers caused by non-steroidal anti-inflammatory drugs or hyperacidity or ulcers caused by post-operative stress; or inhibiting upper gastrointestinal hemorrhage due to peptic ulcer, acute stress ulcer, hemorrhagic gastritis or invasive stress.

Detailed Description

The present invention is further illustrated by the following examples, which are to be understood as merely illustrative and not restrictive.

Unless stated to the contrary, the following terms used in the specification and claims have the following meanings.

The term "alkyl" refers to a saturated aliphatic hydrocarbon group, including straight or branched chain groups of 1 to 10 carbon atoms. Alkyl groups having 1 to 5 carbon atoms are preferred. More preferred are alkyl groups having 1 to 3 carbon atoms, such as methyl, ethyl, n-propyl, isopropyl.

The carbon atom content of various hydrocarbon-containing moieties is represented by the prefix designating the minimum and maximum number of carbon atoms for that moiety, i.e., prefix C_i～jThe number of carbon atoms representing the moiety is from the integer "i" to the integer "j" (inclusive). Thus, for example, C_1～3Alkyl refers to alkyl groups of 1 to 3 carbon atoms (including 1 and 3).

The term "alkoxy" refers to an-O-alkyl group, wherein the alkyl group is as defined herein.

The term "hydroxy" refers to an-OH group.

The term "halogen" refers to fluorine, chlorine, bromine or iodine.

The term "optionally substituted" means that the substitution is optional and, thus, includes both unsubstituted atoms and groups and substituted atoms and groups. "substituted" an atom or group means that any hydrogen on the designated atom or group can be replaced by a choice from the designated group of substituents (up to the point where each hydrogen on the designated atom or group is replaced by a choice from the designated group of substituents), provided that the designated atom or group's normal valency is not exceeded, and that the substitution results in a stable compound. For example,if methyl (i.e., CH)₃) Is optionally substituted, up to 3 hydrogen atoms on a carbon atom may be replaced by a substituent. If an atom or group is described as optionally substituted with one or more non-hydrogen substituents, it may be substituted with up to the maximum number of non-hydrogen substituents on the atom or group that can be substituted.

Unless otherwise specified, all occurrences of a compound in the present invention are intended to include all possible isomers, such as tautomers, enantiomers, diastereomers, and mixtures thereof.

The term "compound of the present invention" means a compound represented by the general formula (I). The term also includes various crystalline forms, pharmaceutically acceptable salts, hydrates or solvates of the compounds of general formula (I).

The term "pharmaceutically acceptable salt" refers to a salt of a compound of the present invention with an acid or base that is suitable for use as a pharmaceutical. Pharmaceutically acceptable salts include inorganic and organic salts. One preferred class of salts is that formed by reacting a compound of the present invention with an acid. Suitable acids for forming salts include, but are not limited to: inorganic acids such as hydrochloric acid, hydrobromic acid, hydrofluoric acid, sulfuric acid, nitric acid, phosphoric acid, etc., organic acids such as formic acid, acetic acid, propionic acid, oxalic acid, malonic acid, succinic acid, fumaric acid, maleic acid, lactic acid, malic acid, tartaric acid, citric acid, picric acid, methanesulfonic acid, phenylmethanesulfonic acid, benzenesulfonic acid, etc.; and acidic amino acids such as aspartic acid and glutamic acid.

The term "pharmaceutically acceptable carrier" refers to carriers that can be used in the preparation of pharmaceutical compositions, which are generally safe, non-toxic, not biologically or otherwise undesirable, and includes carriers that are pharmaceutically acceptable to animals and humans. As used in the specification and claims, a "pharmaceutically acceptable carrier" includes one or more of such carriers.

The terms "comprising," "including," or "including" mean that the various ingredients may be used together in a mixture or composition of the invention. Thus, the terms "consisting essentially of and" consisting of are encompassed by the term "comprising.

The term "preventing" refers, for example, to the prevention of the development of clinical symptoms of a disease in a mammal that may be exposed to or predisposed to the disease but has not yet experienced or exhibited symptoms of the disease.

The term "treating" may refer to inhibiting a disease, e.g., arresting or reducing the development of a disease or clinical symptoms thereof, or ameliorating a disease, e.g., causing regression of a disease or clinical symptoms thereof.

A compound of the general formula (I)

In some embodiments of the invention, R¹Selected from hydrogen atoms, halogens or alkyl groups. In a preferred embodiment, R¹Selected from halogens. In a more preferred embodiment, R¹Is a fluorine atom. R¹The substitution site of (3) is preferably the 2-position.

In some embodiments of the invention, R²Selected from hydrogen atoms, halogens, hydroxyl groups or alkyl groups. In a preferred embodiment, R²Selected from hydrogen atoms, halogens or hydroxyl groups. In a more preferred embodiment, R²Is selected from a hydrogen atom, a fluorine atom or a hydroxyl group. In a preferred embodiment, R²Through CH which may be located on a benzene ring₂Meta or para to the carbon atom to which the pyrazole ring is attached.

In some embodiments of the invention, R³Selected from hydrogen atoms, halogens, hydroxyl groups, alkyl groups or alkoxy groups. In a preferred embodiment, R³Selected from a hydrogen atom, a fluorine atom or a hydroxyl group. In a more preferred embodiment, R³Is a hydrogen atom.

In some embodiments of the invention, R⁴Selected from OR^aOr NR^bR^c。

Wherein R is^aSelected from hydrogen atoms or alkyl groups. In a preferred embodiment, R^aSelected from hydrogen atoms or C_1～3An alkyl group, more preferably a hydrogen atom or a methyl group.

R^b、R^cEach independently selected from a hydrogen atom, a hydroxyl group, an amine group, an alkoxy group, an optionally substituted alkyl group.

Said optionally substituted alkyl being, for example, optionally substituted by one hydroxy, amino or-CONH₂A substituted alkyl group.

In a preferred embodiment, the optionally substituted alkyl is optionally substituted C_1～3An alkyl group.

In a preferred embodiment, R^b、R^cEach independently selected from hydrogen atom, hydroxy, optionally substituted by one hydroxy, amino or-CONH₂Substituted C_1～3An alkyl group. In a more preferred embodiment, R^b、R^cEach independently selected from hydrogen atom, hydroxy group, -CH₂CONH₂。

In a preferred embodiment, -C (═ O) R₄On a benzene ring and containing R₂Meta to the carbon atom to which the benzene ring of (a) is attached.

In some embodiments of the invention, the compound of formula (I) is selected from the compounds shown in Table 1.

TABLE 1

Process for the preparation of compounds of the general formula (I)

In some embodiments of the present invention, the compounds of formula (I) may be prepared using the following general synthetic route:

wherein R is¹、R²、R³、R⁴Is as defined above.

P₁The group may be an amino protecting group known in the art, and may be selected from, for example, C7-11 aralkyl group which may be substituted, such as benzyl group, p-methoxyphenylmethyl group, o-nitrophenylmethyl group, etc.; acetyl, trifluoroacetylC1-6 alkylcarbonyl which may be substituted; benzoyl and the like optionally substituted C6-10 arylcarbonyl; c1-6 alkoxycarbonyl which may be substituted such as methoxycarbonyl, ethoxycarbonyl, Boc (tert-butoxycarbonyl), Cbz (benzyloxycarbonyl), Fmoc (fluorenylmethoxycarbonyl), Teoc (trimethylsilylethoxycarbonyl) and the like; an alkenyloxycarbonyl group such as an Alloc group (allyloxycarbonyl group); alkylsulfonyl such as methylsulfonyl; p-toluenesulfonyl, and the like, C6-10 arylsulfonyl which may be substituted.

X₁The radical may be a leaving group known in the art, and may be selected from a halogen atom such as a fluorine atom, a chlorine atom, a bromine atom, an iodine atom, and the like.

X₂The group is selected from halogen atoms such as chlorine atom, bromine atom, iodine atom, etc.

X₃The group is selected from halogen atoms such as chlorine atom, bromine atom, iodine atom, etc.

In step (a), the compound of formula I-1 is reacted with the compound of formula I-2 to give the compound of formula I-3.

The molar ratio of the compound of formula I-1 to the compound of formula I-2 can be 1: (0.5 to 3.0). The reaction solvent can be acetonitrile, acetone, tetrahydrofuran, dioxane, N-dimethylformamide and the like. The reaction of step (a) may be carried out in the presence of a base. The base may be selected from: cesium carbonate, potassium carbonate, sodium carbonate, potassium hydroxide, sodium hydroxide, and the like. The molar ratio of the compound of formula I-1 to the base can be 1: (1.0-6.0). The reaction temperature in step (a) may be suitably set by those skilled in the art, and may be, for example, 0 to 100 ℃.

In step (b), the compound of formula I-2 is reacted with the compound of formula I-3 to provide the compound of formula I-4.

The molar ratio of the compound of formula I-2 to the compound of formula I-3 can be 1: (0.5 to 3.0). The reaction solvent may be N, N-dimethylacetamide, N-dimethylformamide, toluene, acetonitrile, or the like. Step (b) may be carried out in the presence of a palladium catalyst. The palladium catalyst may be selected from: allylpalladium (II) chloride dimer, tris (dibenzylideneacetone) dipalladium, [1,1' -bis (diphenylphosphino) ferrocene ] dichloropalladium, palladium chloride, and the like. Alternatively, the reaction of step (b) may be carried out in the presence of a base. The base may be selected from: potassium acetate, sodium acetate, potassium phosphate, potassium dihydrogen phosphate, potassium bistrimethylsilyl amide, sodium bistrimethylsilyl amide, and the like. The molar ratio of the compound of formula I-3 to the base can be 1: (0.5 to 3.0). The reaction temperature in step (b) may be suitably set by those skilled in the art, and may be, for example, 40 to 150 ℃.

In step (c), P is removed₁Protecting group to obtain the compound of formula I-6. The reaction conditions may be those commonly used in the art for the removal of amino protecting groups. For example, P₁In the case of Boc, treatment with a protic acid (e.g., trifluoroacetic acid) or a Lewis acid may be used.

In step (d), the compound of formula I-6 is subjected to a aminomethylation reaction to obtain the compound of formula I-7. This step may employ aminomethylation reaction conditions well known in the art. In some embodiments, the compound of formula I-6 is stirred with formaldehyde for a period of time to form a Schiff base, and then a reducing agent, such as sodium borohydride acetate, is added for a period of time to react to provide the compound of formula I-7.

In step (e), the compound of formula I-7 is reacted with the compound of formula I-8 to give the compound of formula (I).

The reaction conditions of the compound of formula I-7 and the compound of formula I-8 can be those commonly used in Suzuki coupling reactions. The molar ratio of the compound of formula I-7 to the compound of formula I-8 can be 1: (0.5-5.0). The reaction solvent may be dioxane, tetrahydrofuran, toluene, N-dimethylformamide, etc. Step (e) may be carried out in the presence of a palladium catalyst. The palladium catalyst may be selected from: tris (dibenzylideneacetone) dipalladium (Pd)₂(dba)₃) Tetrakistriphenylphosphine palladium (Pd (PPh3)4), [1,1' -bis (diphenylphosphino) ferrocene]Palladium dichloride, palladium acetate, and the like. In addition, a phosphine ligand such as 2-dicyclohexylphosphine-2 ',4',6' -triisopropylbiphenyl (X-phos), 4, 5-bisdiphenylphosphine-9, 9-dimethylxanthene (Xant-phos), 1' -binaphthyl-2, 2' -Bisdiphenylphosphine (BINAP), triphenylphosphine, tricyclohexylphosphine, or the like may be added to the reaction in the step (e). The reaction of step (e) may be carried out in the presence of a base. The base may be selected from: potassium carbonate, sodium bicarbonate, cesium carbonate, sodium carbonate, and the like. The molar ratio of the compound of formula I-7 to the base can be 1: (0.5-5.0). The reaction temperature in step (e) can be suitably set by those skilled in the art, and may be, for example, 40 to 150 ℃.

Use of compounds of general formula (I)

The compounds of general formula (I) are useful as inhibitors of gastric acid secretion.

Compounds of formula (I) can be used as H⁺/K⁺Adenosine triphosphatase (H)⁺/K⁺-ATPase) inhibitors.

The compounds of the general formula (I) can be used as potassium ion competitive acid retarders (P-CABs).

The compounds of the general formula (I) can be used for treating and/or preventing peptic ulcer, Zollinger-Ellison syndrome, gastritis, erosive esophagitis, reflux esophagitis, symptomatic gastroesophageal reflux disease, barrett's esophagitis, functional dyspepsia, helicobacter pylori infection, gastric cancer, gastric MALT lymphoma, ulcer caused by non-steroidal anti-inflammatory drugs or hyperacidity or ulcer caused by postoperative stress; or inhibiting upper gastrointestinal hemorrhage due to peptic ulcer, acute stress ulcer, hemorrhagic gastritis or invasive stress. Such peptic ulcers include, but are not limited to, gastric ulcers, duodenal ulcers, or stomal ulcers. Symptomatic gastroesophageal reflux disease includes, but is not limited to, non-erosive reflux disease or gastroesophageal reflux disease without esophagitis.

Pharmaceutical composition

The pharmaceutical composition comprises an effective amount of the compound shown in the general formula (I) or tautomers, enantiomers, diastereomers and mixture forms thereof, and pharmaceutically acceptable salts thereof, and pharmaceutically acceptable carriers or excipients or diluents thereof.

By "effective amount" is meant a compound of the invention: (i) treating a particular disease, condition, or disorder, (ii) attenuating, ameliorating, or eliminating one or more symptoms of a particular disease, condition, or disorder, or (iii) preventing or delaying the onset of one or more symptoms of a particular disease, condition, or disorder described herein.

Examples of pharmaceutically acceptable carrier moieties are cellulose and its derivatives (e.g. sodium carboxymethylcellulose, sodium ethylcellulose, cellulose acetate, etc.), gelatin, talc, solid lubricants (e.g. stearic acid, magnesium stearate), calcium sulfate, vegetable oils (e.g. soybean oil, sesame oil, peanut oil)Oil, olive oil, etc.), polyol (such as propylene glycol, glycerin, mannitol, sorbitol, etc.), emulsifier (such as propylene glycol, glycerin, mannitol, sorbitol, etc.), and water

) Wetting agents (e.g., sodium lauryl sulfate), coloring agents, flavoring agents, stabilizers, antioxidants, preservatives, pyrogen-free water, and the like.

The mode of administration of the compounds or pharmaceutical compositions of the present invention is not particularly limited, and representative modes of administration include (but are not limited to): oral, intratumoral, rectal, parenteral (intravenous, intramuscular or subcutaneous), and topical administration.

The compounds of the present invention may be administered alone or in combination with other pharmaceutically acceptable compounds.

Another aspect of the present invention relates to a method for inhibiting gastric acid secretion, which comprises administering to a patient in need thereof an effective amount of a compound of formula (I) or a tautomer, enantiomer, diastereomer, or mixture thereof, or a pharmaceutically acceptable salt or pharmaceutical composition thereof.

Another aspect of the invention relates to H suppression⁺/K⁺Adenosine triphosphatase (H)⁺/K⁺-ATPase) comprising administering to a patient in need thereof an effective dose of a compound of formula (I) or its tautomers, enantiomers, diastereomers and mixtures thereof, and pharmaceutically acceptable salts thereof or pharmaceutical compositions thereof.

The invention will be further illustrated with reference to the following specific examples. It should be understood that these examples are for illustrative purposes only and are not intended to limit the scope of the present invention. The experimental procedures, in which specific conditions are not noted in the following examples, are generally carried out under conventional conditions or conditions recommended by the manufacturers. Unless otherwise indicated, percentages and parts are by weight.

The structure of the compound is determined by Nuclear Magnetic Resonance (NMR) or Mass Spectrometry (MS), and the purity of the compound is determined by liquid high pressure chromatography (HPLC). NMR was measured using a Bruker AVANCE-400 NMR spectrometer with deuterated dimethyl sulfoxide (DMSO-d6) or deuterated methanol (MeOH-d4) as solvent and Tetramethylsilane (TMS) as internal standard with chemical shifts in ppm. MS was determined using an Agilent 6120 mass spectrometer. HPLC was measured using agilent 1200DAD high pressure liquid chromatograph.

Example 1: 3 '- ((3- (2-fluorophenyl) -5-methyl-5, 6-dihydropyrrolo [3,4-c ] pyrazol-2 (4H) -yl) methyl) - [1,1' -biphenyl ] -3-carboxamide

The first step is as follows: tert-butyl (E) -3- ((dimethylamino) methylene) -4-oxopyrrolidine-1-carboxylate

1a (500g, 2.7mol) was dissolved in N, N-dimethylformamide dimethyl acetal (3.5L), stirred at reflux for 5 hours and LCMS monitored for reaction completion. The reaction was cooled to room temperature, the solvent was spun down, cyclohexane (500mL) was added to the residue, slurried, filtered, the solid washed with cyclohexane (1L. times.3) and dried under vacuum at 40 ℃ for 4 hours to give 1b (444g, yellow solid, 69% yield).¹H NMR(400MHz,CDCl₃)δ7.31(s,1H),4.57(s,2H),3.81(s,2H),3.09(s,6H),1.48(s,9H)。MS m/z(ESI)：241.3[M+H]。

The second step is that: tert-butyl 6 a-hydroxy-3 a,4,6,6 a-tetrahydropyrrole [3,4-c ] pyrazole-5 (1H) -carboxylate

To a solution of 1b (10g, 41.61mmol) in toluene (50mL) was carefully added hydrazine hydrate (2.4mL) dropwise. After dropping, the reaction flask was placed in an oil bath at 45 ℃ to react for 16 hours. LCMS showed the reaction was complete and after returning the reaction to room temperature, a large amount of solid precipitated. Suction filtration, solid washing with cyclohexane (40mL x 3), 40 degrees C vacuum drying for 1 hours, 1c (8.2g, light yellow solid, yield 87%).¹H NMR(400MHz,CDCl₃)δ6.77(s,1H),6.05(br,1H),3.96–3.67(m,2H),3.57(d,J＝11.7Hz,1H),3.45-3.40(m,1H),3.37–3.27(m,1H),1.44(s,9H)。MS m/z(ESI)：228.3[M+H]。

The third step: 2, 6-dihydropyrrolo [3,4-c ] pyrazole-5 (4H) -carboxylic acid tert-butyl ester

To a solution of 1c (8.2g, 36.08mmol) in dichloromethane (100mL) under ice bath was carefully added dropwise a solution of p-toluenesulfonic acid (0.646g) in methanol (10 mL). After dropping, the temperature was allowed to rise to room temperature and stirred overnight, and LCMS indicated completion of the reaction. After 5% aqueous sodium bicarbonate (100mL) was carefully added to the reaction mixture, and the mixture was stirred for 30 minutes, the mixture was allowed to stand for separation, and the organic phase was washed with saturated brine (100mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was taken up in dichloromethane (5mL) and, with stirring, cyclohexane (600mL) was added slowly, whereupon a large amount of solid gradually precipitated, filtered off with suction, washed with cyclohexane (40mL x 3) and dried under vacuum at 40 ℃ for 1 hour to give 1d (5.3g, yellow solid, 71% yield).¹H NMR(400MHz,CDCl₃)δ10.41(s,1H),7.32(s,1H),4.49(s,4H),1.52(s,9H)。MS m/z(ESI)：210.2[M+H]。

The fourth step: 2- (3-chlorobenzyl) -4, 6-dihydropyrrolo [3,4-c ] pyrazole-5 (2H) -carboxylic acid tert-butyl ester

1d (20g, 95.6mmol) was dissolved in acetonitrile (200mL), 1- (bromomethyl) -3-chlorobenzene (20g, 97.3mmol) and cesium carbonate (12.5g, 38.3mmol) were added, the mixture was purged 3 times with water, and the mixture was placed in an oil bath at 85 ℃ for reaction overnight. After cooling to room temperature, the reaction mixture was filtered and the filtrate was concentrated. Yield 1e (20g, yellow powder, 62.5% yield). MS m/z (ESI): 334[ M +1 ].

The fifth step: 2- (3-chlorobenzyl) -3- (2-fluorophenyl) -4, 6-dihydropyrrolo [3,4-c ] pyrazole-5 (2H) -carboxylic acid tert-butyl ester

1e (20g, 60.1mmol), potassium acetate (35.3g, 360.6mmol), allylpalladium (II) chloride dimer (2.19g, 6.01mmol), 2-fluoroiodobenzene (27g, 120.1mmol), and N, N-dimethylacetamide (60mL) were sequentially added to a 1L eggplant-shaped flask, and the flask was placed in an oil bath previously warmed to 100 ℃ after oil pump ventilation three times and reacted overnight. After cooling to room temperature, the reaction mixture was poured directly into water (200mL), followed by extraction with ethyl acetate (100mL × 3). The organic phases were combined and washed with saturated brine (50mL × 2), then dried over anhydrous sodium sulfate, filtered and concentrated. The residue was purified by column chromatography (petroleum ether: ethyl acetate ═ 5: 1) to give 1f (8g, yellow oil, 23.3% yield). MS m/z (ESI): 428.3[ M +1 ].

And a sixth step: 2- (3-chlorobenzyl) -3- (2-fluorophenyl) -2,4,5, 6-tetrahydropyrrole [3,4-c ] pyrazole

Trifluoroacetic acid (16mL) was added to a solution of 1f (8g, 18.7mmol) in dichloromethane (48mL) and reacted at room temperature for 1 hour. After the reaction was complete, it was directly concentrated to give 1g (7g, yellow oil, 110% yield) of crude product for the next reaction. MS m/z (ESI): 328.1[ M +1 ].

The seventh step: aqueous formaldehyde (37%, 4.28g, 10.7mmol) was added to a solution of 1g (7g, 21.5mmol) in dichloromethane/methanol (30mL, 2/1) and stirred at room temperature for half an hour. Sodium borohydride acetate (27.2g, 12.8mmol) was slowly added to the reaction solution, followed by reaction at room temperature overnight. The reaction was concentrated and prepared by HPLC (acetonitrile/water (containing 0.05% trifluoroacetic acid) gradient) to give 1h (6.0g, red oil, 51.3% yield). MS m/z (ESI): 342.3[ M +1]]。¹H NMR(400MHz,CDCl₃)δ7.38(dt,J＝9.5,3.7Hz,1H),7.17(d,J＝16.1Hz,1H),7.09(dd,J＝22.2,12.7Hz,2H),6.99–6.77(m,2H),5.15(s,2H),4.82(d,J＝15.1Hz,2H),4.03(dd,J＝28.1,12.0Hz,2H),3.04(s,4H)。

Eighth step: 3 '- ((3- (2-fluorophenyl) -5-methyl-5, 6-dihydropyrrolo [3,4-c ] pyrazol-2 (4H) -yl) methyl) - [1,1' -biphenyl ] -3-carboxamide

The compound (1h) (100mg, 0.293mmol), 1i (63mg, 0.381mmol), potassium carbonate (81mg, 0.586mmol), tris (dibenzylideneacetone) dipalladium (27mg, 0.030mmol), 2-dicyclohexylphosphorus-2, 4, 6-triisopropylbiphenyl (28mg, 0.057mmol), 1, 4-dioxane/water (2.0mL/0.4mL) were sequentially added to a reaction flask, and reacted at 100 ℃ overnight under an argon atmosphere. The reaction mixture was cooled to room temperature, poured into water (20mL), and extracted with ethyl acetate (10 mL. times.3). The organic phases were combined and washed with saturated brine (10mL × 2), dried over anhydrous sodium sulfate, filtered and concentrated. After concentration, preparative HPLC (acetonitrile/water (containing 0.05% NH3) gradient) gave compound 1(30mg, yellow powder, c yield: 24%).¹H NMR(400MHz,CDCl₃)δ7.92(s,1H),7.76(d,J＝7.8Hz,1H),7.65(d,J＝7.7Hz,1H),7.49(dd,J＝15.2,7.6Hz,3H),7.44-7.37(m,1H),7.37-7.27(m,2H),7.24-7.14(m,4H),7.02(d,J＝7.8Hz,1H),5.32(s,2H),3.99(d,J＝37.1Hz,4H),2.76(s,3H)。MS m/z(ESI)：427.2[M+1]。

Example 2: 3 '- ((3- (2-fluorophenyl) -5-methyl-5, 6-dihydropyrrolo [3,4-c ] pyrazol-2 (4H) -yl) methyl) - [1,1' -biphenyl ] -3-carboxylic acid

Compound 1(0.1g, 0.23mmol), 25N sodium hydroxide solution (0.4ml, 10mmol, 44eq) and water (0.8ml) were added to a sealed tube in this order and heated at 40 ℃ for reaction overnight. The reaction mixture was cooled to room temperature, poured into water (10mL), and extracted with ether (10 mL. times.3). The aqueous phase was adjusted to pH 7 with citric acid and extracted with dichloromethane (10mL × 2), the organic phases were combined and dried over anhydrous sodium sulfate, filtered, concentrated and prepared by HPLC (acetonitrile/water (containing 0.05% trifluoroacetic acid) to give crude which was purified by TLC plate (dichloromethane/methanol 10/1) to give compound 2 (trifluoroacetate salt series 2, 140mg, pale yellow solid, yield: 92.9%).¹H NMR(400MHz,CDCl₃)δ13.98(s,1H),8.17(s,1H),8.06(d,J＝7.6Hz,1H),7.73(d,J＝7.7Hz,1H),7.58-7.43(m,3H),7.38(t,J＝7.7Hz,1H),7.27(dd,J＝19.3,10.3Hz,4H),7.06(d,J＝7.5Hz,1H),5.36(s,2H),5.02(s,2H),4.11(d,J＝31.2Hz,2H),3.15(s,3H)。MS m/z(ESI)：428.1[M+1]。

Example 3: 3 '- ((3- (2-fluorophenyl) -5-methyl-5, 6-dihydropyrrolo [3,4-c ] pyrazol-2 (4H) -yl) methyl) -N-hydroxy- [1,1' -biphenyl ] -3-carboxamide

Weighing compound 2(200mg, 0.47mmol), placing in a eggplant-shaped bottle, adding tetrahydrofuran (1mL), adding EDCI (136mg, 0.71mmol), HOBT (82mg, 0.61mmol), reacting for 1h under the protection of argon, adding hydroxylamine hydrochloride (36mg, 0.52mmol) and triethylamine 0.84mL, stirring at room temperature for 2h, washing the organic phase with brine (5mL x 2) after the reaction is finished, drying with anhydrous sodium sulfate, filtering, concentratingAfter concentration, the crude product was prepared by hplc (acetonitrile/water (containing 0.05% trifluoroacetic acid)) to give 3 (trifluoroacetate salt, salt coefficient 1.6, 39.9mg, light yellow oil, yield 10.5%).¹H NMR(400MHz,MeOD)δ7.86(s,1H),7.70(d,J＝7.6Hz,1H),7.64(d,J＝7.8Hz,1H),7.50(t,J＝7.5Hz,3H),7.34(dd,J＝12.3,7.6Hz,2H),7.31–7.22(m,2H),7.18(s,1H),6.98(d,J＝7.6Hz,1H),5.35(s,2H),3.91(d,J＝37.6Hz,4H),2.70(s,3H)。MS m/z(ESI)：443.1[M+1]。

Example 4: n- (2-amino-2-oxoethyl) -3 '- ((3- (2-fluorophenyl) -5-methyl-5, 6-dihydropyrrolo [3,4-c ] pyrazol-2 (4h) -yl) methyl) - [1,1' -biphenyl ] -3-carboxamide

After stirring 3(140mg, 0.33mmol), THF (2ml), EDCI (95mg, 0.50mmol), HOBT (58mg, 0.43mmol) in a 25ml eggplant-shaped flask in this order for 1 hour at normal temperature, triethylamine (0.051ml), 2-aminoacetamide (26mg, 0.36mmol) were added and stirred for 3 hours at normal temperature, after the reaction was completed, the reaction mixture was filtered and dried, and the crude product was prepared by high pressure liquid chromatography (acetonitrile/water (containing 0.05% trifluoroacetic acid) to give 4 (trifluoroacetate salt, salt ratio 4, 39.9mg, pale yellow solid, yield 13.1%).¹H NMR(400MHz,MeOD)δ7.99(s,1H),7.84(d,J＝7.7Hz,1H),7.67(d,J＝7.7Hz,1H),7.53(dd,J＝13.8,6.9Hz,3H),7.42(t,J＝6.9Hz,1H),7.37–7.24(m,3H),7.20(s,1H),7.02(d,J＝7.6Hz,1H),5.44(s,2H),4.81–4.37(m,4H),4.07(s,2H),3.18(s,3H)。MS m/z(ESI)：484.2[M+1]。

Example 5: 1-amino-2-methyl-1-oxoprop-2-yl-3- ((3- (2-fluorophenyl) -5-methyl-5, 6-dihydropyrrolo [3,4-c ] pyrazol-2 (4H) -yl) methyl) benzoate

The first step is as follows: 2-bromo-4- (bromomethyl) -1-methoxybenzene

2-bromo-1-methoxy-4-toluene (5g, 24 g) was added to the eggplant-shaped bottle8mmol) was dissolved in 30mL of dichloromethane, N-bromosuccinimide (5.3g, 29.8mmol) and azobisisobutyronitrile (6.8g, 42mmol) were added, and the mixture was heated at 40 ℃ under reflux overnight. After the reaction was complete, the reaction was brought to room temperature and water (20mL) was added, washed with ethyl acetate (40mL x 3) and brine (40mL x 2), then dried over anhydrous sodium sulfate, filtered and concentrated to give compound 5b (5.6g, yellow solid, 81.5% yield).¹H NMR(400MHz,CDCl₃)δ7.59(d,J＝2.0Hz,1H),7.32-7.26(m,1H),6.86(d,J＝8.4Hz,1H),4.44(s,2H),3.89(s,3H).MS m/z(ESI)：278.8[M+H]。

The second step is that: 2- (3-bromo-4-methoxybenzyl) -4, 6-dihydropyrrolo [3,4-c ] pyrazole-5 (2H) -carboxylic acid tert-butyl ester

1d (2.8g, 13.5mmol) was dissolved in acetonitrile (50mL), 5b (4.5g, 16.2mmol) and cesium carbonate (32.5g, 47mmol) were added, the mixture was purged 3 times with nitrogen, and the mixture was put in an oil bath at 80 ℃ for 3 hours. The reaction solution was filtered, and the filtrate was concentrated. The residue was purified by column chromatography (petroleum ether: ethyl acetate ═ 2: 1) to give 5c (6g, yellow oil, 91% yield). MS m/z (ESI): 408.0[ M + H ].

The third step: 2- (3-bromo-4-methoxybenzyl) -3- (2-fluorophenyl) -4, 6-dihydropyrrolo [3,4-c ] pyrazole-5 (2H) -carboxylic acid tert-butyl ester

To an eggplant-shaped bottle were added 5c (5.6g, 13.7mmol), potassium acetate (8g, 82.2mmol), allylpalladium (II) chloride dimer (499mg, 1.37mmol), N-dimethylacetamide (50mL) and o-fluoroiodobenzene (6.1g, 27.4mmol) in this order. After the oil pump takes gas (argon) for four times, the mixture is put into an oil bath which is heated to 100 ℃ in advance to react for 3 hours. After the reaction was returned to room temperature, the reaction mixture was poured directly into water (80mL), and extracted with ethyl acetate (40mL × 3). The organic phase was washed with brine (40mL x 2), then dried over anhydrous sodium sulfate, filtered and concentrated. The crude product was isolated by column chromatography (3/l petroleum ether/ethyl acetate) to give 5d (729mg, yellow oil, 10.6% yield). MS m/z (ESI): 502.1[ M + H ].

The fourth step: 2- (3-bromo-4-methoxybenzyl) -3- (2-fluorophenyl) -2,4,5, 6-tetrahydropyrrole [3,4-c ] pyrazole

Trifluoroacetic acid (0.5mL) was added to a solution of 5d (729mg, 1.45mmol) in dichloromethane (1.5mL) and reacted at room temperature for 1 hour. After the reaction was complete, concentration directly gave crude 5e (581mg, brown oil, 100% yield). MS m/z (ESI): 403.2[ M + H ].

The fifth step: 2- (3-bromo-4-methoxybenzyl) -3- (2-fluorophenyl) -5-methyl-2, 4,5, 6-tetrahydropyrrole [3,4-c ] pyrazole

Aqueous formaldehyde (37%, 1.2g, 14.5mmol) was added to a solution of 5e (581mg, 1.45mmol) in dichloromethane/methanol (6mL, 2/1) and stirred at room temperature for half an hour. Sodium borohydride acetate (1.8g, 8.7mmol) was slowly added to the reaction solution, followed by reaction at room temperature overnight. The reaction mixture was concentrated, diluted with dichloromethane (10mL), washed with aqueous ammonia/water (10mL × 2, 1/5) and saturated brine (10mL), and the organic phase was dried over anhydrous sodium sulfate and concentrated. Prepared by hplc 50mg of crude product (acetonitrile/water with 0.05% trifluoroacetic acid) to give 5f (trifluoroacetate salt, salt series 1.3, 20.1mg, yellow oil, 24.5% yield).¹H NMR(400MHz,CDCl₃)δ7.50-7.46(m,1H),7.25–7.07(m,4H),6.97(d,J＝8.4Hz,1H),6.80(d,J＝8.4Hz,1H),5.17(s,2H),4.96(s,1H),4.09–4.01(m,2H),3.86(s,3H),3.18(s,3H)。MS m/z(ESI)：416.0[M+H]。

And a sixth step: 2-bromo-4- ((3- (2-fluorophenyl) -5-methyl-5, 6-dihydropyrrolo [3,4-c ] pyrazol-2 (4H) -yl) methyl) phenol

5f (50mg, 0.11mmol) was weighed out and put in an eggplant-shaped bottle, dichloromethane (1mL) was added, and boron tribromide (0.1mL) was added under ice-cooling to react for 2 hours. After the reaction was complete, the reaction was quenched with water (5mL) and extracted with dichloromethane (5mL x 3). The organic phase was washed with brine (5mL × 2), then dried over anhydrous sodium sulfate, filtered and concentrated, and the crude product was prepared by high pressure liquid chromatography (acetonitrile/water (containing 0.05% trifluoroacetic acid) to give 5g (trifluoroacetate salt, salt coefficient ═ 1.6, 29.9mg, light yellow oil, yield 46.5%).¹H NMR(400MHz,CDCl₃)δ7.52-7.47(m,1H),7.27–7.10(m,4H),6.87(q,J＝8.4Hz,2H),6.70(s,1H),5.15(s,2H),4.94(d,J＝12.8Hz,2H),4.12-4.02(m,2H),3.13(s,3H)。MS m/z(ESI)：402.2[M+H]。

The seventh step: 5 ' - ((3- (2-fluorophenyl) -5-methyl-5, 6-dihydropyrrolo [3,4-c ] pyrazol-2 (4h) -yl) methyl) -2' -hydroxy- [1,1' -biphenyl ] -3-carboxylic acid methyl ester

5g (200mg, 0.5mmol), potassium carbonate (138mg, 1.0mmol), tris (dibenzylideneacetone) dipalladium (46mg, 0.05mmol), 2-dicyclohexylphosphine-2 ',4',6' -triisopropylbiphenyl (48mg, 0.1mmol), 1, 4-dioxane (5mL) and 3-methoxycarbonylphenylboronic acid (270mg, 1.5mmol) were sequentially charged in an eggplant-shaped bottle. After the oil pump was purged four times with argon, it was put into an oil bath which was warmed up to 110 ℃ in advance to react for 16 hours. After the reaction was returned to room temperature, the reaction mixture was poured directly into water (10mL), and extracted with ethyl acetate (10mL × 3). The organic phase was washed with brine (10mL x 2), then dried over anhydrous sodium sulfate, filtered and concentrated to give 110mg of a black oil. Prepared by hplc 30mg of crude product (acetonitrile/water with 0.05% trifluoroacetic acid) to give compound 5 (trifluoroacetate salt series 2, 2.2mg, yellow oil, 4.2% yield).¹H NMR(400MHz,CDCl₃)δ8.05(d,J＝10.0Hz,2H),7.61(d,J＝7.2Hz,1H),7.57–7.46(m,2H),7.29(s,1H),7.25–7.17(m,2H),6.95(s,2H),6.88(d,J＝8.4Hz,1H),5.24(d,J＝6.0Hz,2H),4.97(t,J＝14.4Hz,2H),4.17-4.03(m,2H),3.94(s,3H),3.12(s,3H)。MS m/z(ESI)：458.1[M+H]。

Example 6: 5 ' - ((3- (2-fluorophenyl) -5-methyl-5, 6-dihydropyrrolo [3,4-c ] pyrazol-2 (4H) -yl) methyl) -2' -hydroxy- [1,1' -biphenyl ] -3-carboxylic acid

The first step is as follows: 5 ' - ((3- (2-fluorophenyl) -5-methyl-5, 6-dihydropyrrolo [3,4-c ] pyrazol-2 (4H) -yl) methyl) -2' -methoxy- [1,1' -biphenyl ] -3-carboxylic acid methyl ester

To an eggplant-shaped bottle were added 5f (500mg, 1.2mmol), potassium carbonate (331mg, 2.4mmol), tris (dibenzylideneacetone) dipalladium (109mg, 0.12mmol), 2-dicyclohexylphosphine-2 ',4',6' -triisopropylbiphenyl (114mg, 0.24mmol), 1, 4-dioxane (5mL) and 3-methoxycarbonylphenylboronic acid (648mg, 3.6mmol) in this order. After the oil pump was purged four times with argon, it was put into an oil bath which was warmed up to 110 ℃ in advance to react for 16 hours. After the reaction was returned to room temperature, the reaction mixture was poured directly into water (10mL), and extracted with ethyl acetate (10mL × 3). The organic phase was washed with brine (10mL × 2), then dried over anhydrous sodium sulfate, filtered and concentrated. The crude product was isolated on preparative plates to give compound 6a (475mg, black solid, 84% yield). MS m/z (ESI): 471.2[ M + H ].

The second step is that: 5 ' - ((3- (2-fluorophenyl) -5-methyl-5, 6-dihydropyrrolo [3,4-c ] pyrazol-2 (4H) -yl) methyl) -2' -hydroxy- [1,1' -biphenyl ] -3-carboxylic acid

Compound 6a (200mg, 0.42mmol) was weighed out and placed in a bottle shaped like a eggplant, methylene chloride (1mL) was added, and boron tribromide (0.4mL) was added under ice-cooling to react for 2 hours. After the reaction, sodium hydroxide (84mg, 2.1mmol) was added for 2h, and after the reaction, dichloromethane (5mL × 3) was used for extraction. The organic phase was washed with brine (5mL × 2), then dried over anhydrous sodium sulfate, filtered and concentrated, and the crude product was prepared by high pressure liquid chromatography (acetonitrile/water (containing 0.05% trifluoroacetic acid) to give compound 6 (trifluoroacetate salt series ═ 1.3, 92.8mg, yellow solid, 37.3% yield).¹H NMR(400MHz,MeOD)δ8.05(s,1H),7.93(d,J＝7.6Hz,1H),7.66(d,J＝7.6Hz,1H),7.55–7.48(m,1H),7.46-7.38(m,2H),7.34-7.20(m,2H),6.87–6.78(m,2H),6.77(d,J＝8.0Hz,1H),5.26(s,2H),4.68–4.50(m,4H),3.14(s,3H)。MS m/z(ESI)：444.1[M+H]。

Example 7: 5 ' - ((3- (2-fluorophenyl) -5-methyl-5, 6-dihydropyrrolo [3,4-c ] pyrazol-2 (4H) -yl) methyl) -N, 2' -dihydroxy- [1,1' -biphenyl ] -3-carboxamide

Adding 5(40mg, 0.088mmol) into a eggplant-shaped bottle, dissolving in tetrahydrofuran (6mL), adding sodium hydroxide (17.6mg, 0.44mmol), stirring at normal temperature for 1h, after the reaction is finished, extracting with dichloromethane (10mL, 3), washing with brine (10mL, 2), drying with anhydrous sodium sulfate, filtering, concentrating, adding tetrahydrofuran (6mL), 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride (5mg, 0.132mmol), 1-hydroxybenzotriazole (15mg, 0.115mmol), reacting for 0.5h, adding hydroxylamine hydrochloride (6.7mg, 0.097mmol), one drop of triethylamine, stirring at normal temperature for 3h, after the reaction is finished, spin-drying, ethyl acetate (10mL, 3)Extraction, washing with sodium bicarbonate (10mL × 2), water (10mL × 2) and brine (10mL × 2) in this order, drying over anhydrous sodium sulfate, filtration and concentration. The crude product was prepared by high pressure liquid chromatography (acetonitrile/water with 0.05% trifluoroacetic acid) to give 7 (trifluoroacetate salt, salt number ═ 1.4, 2.5mg, yellow oil, 4.5% yield).¹H NMR(400MHz,MeOD)δ7.76(s,1H),7.63(s,2H),7.54(d,J＝6.8Hz,1H),7.43(t,J＝7.2Hz,2H),7.35-7.28(m,2H),6.88-6.78(m,2H),6.75(d,J＝8.0Hz,1H),5.30(s,2H),4.45(s,4H),3.16(s,3H)。MS m/z(ESI)：459.1[M+H]。

Example 8: 2' -fluoro-5 ' - ((3- (2-fluorophenyl) -5-methyl-5, 6-dihydropyrrolo [3,4-c ] pyrazol-2 (4H) -yl) methyl) - [1,1' -biphenyl ] -3-carboxylic acid

The first step is as follows: 2- (3-bromo-4-fluorobenzyl) -4, 6-dihydropyrrolo [3,4-c ] pyrazole-5 (2H) -carboxylic acid tert-butyl ester

1d (3.3g, 15.8mmol) was dissolved in acetonitrile (40mL), 3-bromo-4-fluorobenzyl bromide (5g, 18.9mmol) and cesium carbonate (18g, 55.3mmol) were added, nitrogen was substituted 3 times, and the mixture was placed in an oil bath at 80 ℃ for reaction for 3 hours. The reaction solution was filtered, and the filtrate was concentrated. The residue was purified by column chromatography (petroleum ether: ethyl acetate ═ 2: 1) to give 8a (6.1g, yellow solid, yield 97.7%). MS m/z (ESI): 396.0[ M + H ].

The second step is that: 2- (3-bromo-4-fluorobenzyl) -3- (2-fluorophenyl) -4, 6-dihydropyrrolo [3,4-c ] pyrazole-5 (2H) -carboxylic acid tert-butyl ester

To a solanaceous bottle were added 8a (6.1g, 15.4mmol), potassium acetate (9g, 92.4mmol), allylpalladium (II) chloride dimer (560mg, 1.54mmol), N-dimethylacetamide (30mL), and o-fluoroiodobenzene (6.8g, 30.8mmol) in that order. After the oil pump takes gas (argon) for four times, the mixture is put into an oil bath which is heated to 100 ℃ in advance to react for 3 hours. After the reaction was returned to room temperature, the reaction mixture was poured directly into water (80mL), and extracted with ethyl acetate (40mL × 3). The organic phase was washed with water (15mL x 2) and brine (15mL x 2) in sequence, then dried over anhydrous sodium sulfate, filtered and concentrated. The crude product was isolated by column chromatography (5/l petroleum ether/ethyl acetate) to yield 8b (1.1g, yellow-brown oil, 14.6% yield). MS m/z (ESI): 492.2[ M + H ].

The third step: 2- (3-bromo-4-fluorobenzyl) - (2, 3-fluorophenyl) -2,4,5,6- [3,4-c ] tetrahydropyrrolopyrazole

Trifluoroacetic acid (4mL) was added to a solution of 8b (1.1g, 2.25mmol) in dichloromethane (12mL) and reacted at room temperature for 1 hour. After the reaction was complete, concentration directly gave crude 8c (875mg, black liquid, 100% yield). MS m/z (ESI): 392.2[ M + H ].

The fourth step: 2- (3-bromo-4-fluorobenzyl) -3- (2-fluorophenyl) -5-methyl-2, 4,5, 6-tetrahydropyrrolo [3,4-c ] pyrazole

Aqueous formaldehyde (37%, 1.6g, 22.5mmol) was added to a solution of 8c (875mg, 2.25mmol) in dichloromethane/methanol (15mL, 2/1) and stirred at room temperature for half an hour. Sodium borohydride acetate (1.4g, 6.75mmol) was slowly added to the reaction solution, followed by reaction at room temperature overnight. The reaction was concentrated, diluted with dichloromethane (10mL), washed successively with ammonia/water (10mL × 2, 1/5) and saturated brine (10mL), and the organic phase was dried over anhydrous sodium sulfate and concentrated to give a crude product. Prepared by hplc 50mg of crude product (acetonitrile/water containing 0.05% trifluoroacetic acid) to give 8d (trifluoroacetate salt series 2.3, 16.2mg, pale yellow solid, 19.0% yield).¹H NMR(400MHz,CDCl₃)δ7.42(d,J＝5.6Hz,1H),7.22–7.11(m,4H),6.96(t,J＝8.0Hz,1H),6.89(s,1H),5.13(s,2H),4.89(s,2H),4.05(s,2H),3.05(s,3H)。MS m/z(ESI)：406.3[M+H]。

The fifth step: 2' -fluoro-5 ' - ((3- (2-fluorophenyl) -5-methyl-5, 6-dihydropyrrolo [3,4-c ] pyrazol-2 (4h) -yl) methyl) - [1,1' -biphenyl ] -3-carboxylic acid methyl ester

To an eggplant-shaped bottle were added 8d (250mg, 0.62mmol), potassium carbonate (171mg, 1.24mmol), tris (dibenzylideneacetone) dipalladium (56mg, 0.062mmol), 2-dicyclohexylphosphine-2 ',4',6' -triisopropylbiphenyl (58mg, 0.124mmol), 1, 4-dioxane (3mL), and 3-methoxycarbonylphenylboronic acid (334mg, 1.86mmol) in this order. After the oil pump was purged four times with argon, it was put into an oil bath which was warmed up to 110 ℃ in advance to react for 16 hours. After the reaction was cooled to room temperature, the reaction mixture was poured directly into water (20mL), and extracted with ethyl acetate (20mL × 3). The organic phase was washed with (10mL x 2) and brine (10mL x 2) in that order, then dried over anhydrous sodium sulfate, filtered and concentrated. The crude product was isolated on a preparative plate (dichloromethane: methanol ═ 10: 1) to give compound 8e (204mg, yellow oil, yield 60.0%). MS m/z (ESI): 460.4[ M + H ].

And a sixth step: 2' -fluoro-5 ' - ((3- (2-fluorophenyl) -5-methyl-5, 6-dihydropyrrolo [3,4-c ] pyrazol-2 (4H) -yl) methyl) - [1,1' -biphenyl ] -3-carboxylic acid

After 8e (204mg, 0.44mmol), tetrahydrofuran (6mL) and water (1mL) were added to an eggplant-shaped bottle in this order to dissolve it, lithium hydroxide (36mg, 0.88mmol) was added thereto, and the mixture was stirred at room temperature for 3 hours, the pH was adjusted to 4 to 5 with 1N diluted hydrochloric acid under ice bath, followed by extraction with dichloromethane (25mL _ 3), washing of the organic phase with water (10mL _ 2) and brine (10mL _ 2) successively, followed by drying over anhydrous sodium sulfate, filtration and concentration. The crude product was prepared by high pressure liquid chromatography (acetonitrile/water containing 0.05% trifluoroacetic acid) to afford compound 8 (trifluoroacetate salt series 1.4, 101.1mg, white solid, 38.0% yield).¹H NMR(400MHz,CDCl₃)δ8.08(d,J＝8.0Hz,2H),7.73(d,J＝7.6Hz,1H),7.54-7.45(m,2H),7.28(s,1H),7.26–7.16(m,2H),7.10-7.04(m,3H),5.30(s,2H),5.01(s,2H),4.22–4.11(m,2H),3.14(s,3H)。MS m/z(ESI)：446.4[M+H]。

Example 9: 3 ' - ((3- (2-fluorophenyl) -5-methyl-5, 6-dihydropyrrolo [3,4-c ] pyrazol-2 (4H) -yl) methyl) -5 ' -hydroxy- [1,1' -biphenyl ] -3-carboxylic acid

The first step is as follows: 1-bromo-3-methoxy-5- (bromomethyl)

1-bromo-3-methoxy-5-methylbenzene 9a (22.8g, 113mmol) was sequentially added to a flask in the shape of eggplant, dissolved in methylene chloride (100mL), and N-bromosuccinimide (242g, 136mmol) and azobisisobutyronitrile (31.5g, 192mmol) were added thereto, followed by heating to 45 ℃ and refluxing overnight. After completion of the reaction, the reaction was returned to room temperature and extracted with ethyl acetate (40mL × 3). The organic phase was washed successively with water (40mL x 2) and brine (40mL x 2) and then with anhydrousDried over sodium sulfate, filtered and concentrated, and the crude product was isolated on normal phase silica gel column (petroleum ether: ethyl acetate ═ 20:1) to give 9b (20g, yellow solid, 63% yield).¹H NMR(400MHz,CDCl₃)δ7.12(s,1H),6.98(s,1H),6.85(s,1H),4.38(s,3H),3.80(s,4H)。MS m/z(ESI)：278.8[M+H]。

The second step is that: 1d (12g, 56.7mmol) was dissolved in acetonitrile (100mL), 9b (19g, 68.1mmol) and cesium carbonate (64.8g, 199mmol) were added, the mixture was replaced with nitrogen 3 times, and the mixture was put in an oil bath at 80 ℃ and reacted for 3 hours. The reaction solution was filtered, and the filtrate was concentrated. The residue was purified by column chromatography (petroleum ether: ethyl acetate ═ 2: 1) to give 9c (23g, yellow oil, yield 82.9%). MS m/z (ESI): 408.0[ M + H ].

The third step: 2- (3-bromo-5-methoxybenzyl) -3- (2-fluorophenyl) -4, 6-dihydropyrrolo [3,4-c ] pyrazole-5 (2H) -carboxylic acid tert-butyl ester

9c (22g, 54.1mmol), potassium acetate (31.8g, 324.6mmol), allylpalladium (II) chloride dimer (1.9mg, 5.41mmol), N-dimethylacetamide (100mL) and o-fluoroiodobenzene (24g, 108.2mmol) were sequentially added to an eggplant-shaped bottle. After the oil pump takes gas (argon) for four times, the mixture is put into an oil bath which is heated to 100 ℃ in advance to react for 3 hours. After the reaction was returned to room temperature, the reaction mixture was poured directly into water (80mL), and extracted with ethyl acetate (40mL × 3). The organic phase was washed successively with water (40mL x 2) and brine (40mL x 2), then dried over anhydrous sodium sulfate, filtered and concentrated. The crude product was isolated by column chromatography (3/l petroleum ether/ethyl acetate) to give 9d (2.7g, yellow oil, 9.1% yield). MS m/z (ESI): 502.1[ M + H ].

The fourth step: 2- (3-bromo-5-methoxybenzyl) -3- (2-fluorophenyl) -2,4,5, 6-tetrahydropyrrole [3,4-c ] pyrazole

Trifluoroacetic acid (7mL) was added to a solution of 9d (2.7g, 5.5mmol) in dichloromethane (21mL) and reacted at room temperature for 1 hour. After the reaction was complete, concentration directly gave crude 9e (2.1g, black liquid, 100% yield). MS m/z (ESI): 403.2[ M + H ].

The fifth step: 2- (3-bromo-5-methoxybenzyl) -3- (2-fluorophenyl) -5-methyl-2, 4,5, 6-tetrahydropyrrole [3,4-c ] pyrazole

An aqueous formaldehyde solution (37%, 1.39g,55mmol) was added to a solution of 9e (2.2g, 5.5mmol) in dichloromethane/methanol (20mL, 2/1) and stirred at room temperature for half an hour. Sodium borohydride acetate (6.9g, 33mmol) was slowly added to the reaction solution, followed by reaction at room temperature overnight. The reaction mixture was concentrated, diluted with dichloromethane (10mL), washed successively with ammonia/water (10mL × 2, 1/5) and saturated brine (10mL), and the organic phase was dried over anhydrous sodium sulfate and concentrated to give a crude product. Prepared by hplc 50mg of crude product (acetonitrile/water with 0.05% trifluoroacetic acid) to give 9f (trifluoroacetate salt, salt series 2, 4.1mg, yellow oil, 5.1% yield).¹H NMR(400MHz,CDCl₃)δ7.49(s,2H),7.25(s,2H),6.95(s,1H),6.77(s,1H),6.50(s,1H),5.20(s,2H),5.01(s,2H),4.22–3.97(m,2H),3.73(s,3H),3.16(s,3H)。MS m/z(ESI)：416.3[M+H]。

And a sixth step: 3-bromo-5- ((3- (2-fluorophenyl) -5-methyl-5, 6-dihydropyrrolo [3,4-c ] pyrazol-2 (4H) -yl) methyl) phenol

9f (6g, 14.5mmol) was weighed into a bottle shaped like a eggplant, methylene chloride (10mL) was added, and boron tribromide (12mL) was added under ice-cooling to react for 2 hours. After the reaction was complete, the reaction was quenched with water (10mL) and extracted with dichloromethane (15mL x 3). The organic phase was washed with brine (10mL × 2), then dried over anhydrous sodium sulfate, filtered and concentrated, and 50mg of the crude product was prepared by high pressure liquid chromatography (acetonitrile/water (containing 0.05% trifluoroacetic acid) to give 9g (trifluoroacetate salt, salt coefficient ═ 1.7, 23mg, light yellow oil, yield 30.5%).¹H NMR(400MHz,MeOD)δ7.58-7.53(m,1H),7.40-7.26(m,2H),6.80(s,1H),6.56(s,1H),6.39(s,1H),5.24(s,2H),4.60(s,4H),3.18(s,3H)。MS m/z(ESI)：404.2[M+H]。

The seventh step: 3 ' - ((3- (2-fluorophenyl) -5-methyl-5, 6-dihydropyrrolo [3,4-c ] pyrazol-2 (4h) -yl) methyl) -5 ' -hydroxy- [1,1' -biphenyl ] -3-carboxylic acid methyl ester

9g (200mg, 0.5mmol), potassium carbonate (138mg, 1.0mmol), tris (dibenzylideneacetone) dipalladium (46mg, 0.05mmol), 2-dicyclohexylphosphine-2 ',4',6' -triisopropylbiphenyl (47mg, 0.1mmol), 1, 4-dioxane (3mL) and 3-methoxycarbonylphenylboronic acid (270mg, 1.5mmol) were sequentially charged in an eggplant-shaped bottle. After the oil pump was purged four times with argon, it was put into an oil bath which was warmed up to 110 ℃ in advance to react for 16 hours. After the reaction was returned to room temperature, the reaction mixture was poured directly into water (15mL), and extracted with ethyl acetate (20mL × 3). The organic phase was washed successively with water (10mL x 2) and brine (10mL x 2), then dried over anhydrous sodium sulfate, filtered and concentrated. The crude product was isolated on a preparative plate (dichloromethane: methanol ═ 10: 1) yielding 9h (80mg, yellow oil, 34.9% yield). MS m/z (ESI): 458.4[ M + H ].

Eighth step: 3 ' - ((3- (2-fluorophenyl) -5-methyl-5, 6-dihydropyrrolo [3,4-c ] pyrazol-2 (4H) -yl) methyl) -5 ' -hydroxy- [1,1' -biphenyl ] -3-carboxylic acid

Compound 9h (80mg, 0.18mmol), tetrahydrofuran (6mL) and water (1mL) were added to an eggplant-shaped bottle in this order to be dissolved, lithium hydroxide (10.6mg, 0.26mmol) was added thereto, and the mixture was stirred at room temperature for 3h, and after completion of the reaction, the mixture was washed with dichloromethane (10 mL. about.3) and brine (10 mL. about.2), and then dried over anhydrous sodium sulfate, filtered and concentrated. The crude product was prepared by hplc (acetonitrile/water with 0.05% trifluoroacetic acid) to provide compound 9 (trifluoroacetate salt 0.5, 58.3mg, white solid, 64.8% yield).¹H NMR(400MHz,MeOD)δ8.05(s,1H),7.94(d,J＝7.6Hz,1H),7.61(d,J＝7.6Hz,1H),7.54–7.37(m,3H),7.30–7.23(m,2H),6.91(s,1H),6.63(s,1H),6.49(s,1H),5.30(s,2H),4.59(s,2H),4.50(s,2H),3.14(s,3H)。MS m/z(ESI)：444.4[M+H]。

Test example: assay for inhibition of H +/K + ATPase enzymatic Activity by Compounds

The following experiment was conducted to determine the inhibitory effect of the compounds of the present invention on the activity of H +/K + ATPase.

1. Experimental Material

Plate reader:SpectraMax M5(MD)

Malachite green (Sigma Aldrich,213020-25G)

Ammonium molybdate (Sigma Aldrich,277908-20G)

ATP(Sigma Aldrich,A1852-1VL)。

2. Buffer solution preparation

Enzyme working solution: titrating enzyme, diluting enzyme with buffer solution 1, and taking 5. mu.l of diluted solution to 50. mu.l of reaction system

ATP solution: k-free for 100mM ATP⁺buffer was diluted to 5mM and 5. mu.l of the dilution was added to 50. mu.l of the reaction system, i.e., the final ATP concentration was 500. mu.M

MLG developing solution: 0.12% MLG, 7.5% ammonium molybdate, 11% Tween-20 as 100: 25: 2, and adding 15 mul of the mixture into each hole during detection

Buffer 1:50mM Tris-HCl pH 6.5,5mM magnesium chloride (magnesium chloride), 10. mu.M valinomycin

Buffer 2:50mM Tris-HCl pH 6.5,5mM magnesium chloride (magnesium chloride), 10. mu.M valinomycin (valinomycin),20mM KCl

Homogenization buffer: 10mmol/L Tris-HCl, pH 6.8, 0.25M sucrose (sucrose),1mmol/L EDTA 7.5% Ficoll layering: homogenization buffer + 7.5% (W/W)

400 (ficoll 400).

3. Experimental procedure

3.1.H⁺/K⁺ATP enzyme extraction

(1) Separating stomach tissue of rabbit, washing blood stain with tap water, and removing food residue;

(2) thoroughly cleaning the fundus part by using a precooled NaCl solution to remove surface mucus;

(3) loading the stripped mucosa into a sample bag or a 50ml centrifuge tube, and quickly freezing in a liquid nitrogen tank;

(4) taking out tissue, cutting with surgical scissors, adding pre-cooled homogenization buffer (4ml/g tissue), and homogenizing in tissue homogenizer for 2-10 min;

(5) after homogenization, if larger tissue particles exist, centrifuging (600g for 10min), removing the supernatant, then transferring the supernatant into a clean centrifuge tube, centrifuging 20000g for 30min, then transferring the supernatant into a clean centrifuge tube, further centrifuging, centrifuging 100000g for 90min, and collecting precipitates;

(6) resuspending the precipitate with homogenate buffer, blowing uniformly, adding 7.5% Ficoll layering solution in equal proportion, centrifuging at 100000g for 90min, and collecting the precipitate;

(7) the pellet was resuspended in homogenization buffer, blown out evenly and the protein concentration was measured by Bradford. Freezing at-80 deg.C.

3.2.H⁺/K⁺ATP enzyme Activity assay

(1) Mu.l reaction buffer was added to each experimental well, followed by 35. mu.l buffer 1

(2) To the whole enzyme and buffer wells, 5. mu.l of buffer 1 containing 10% DMSO was added

(3) Adding 5 μ l 10X compound working solution into the compound hole, and mixing

(4) Mu.l of buffer 1 was added to the buffer wells

(5) Adding 5 μ l 10 Xase working solution into the rest wells, mixing, incubating at 37 deg.C for 30min

(6) Mu.l of 10XATP working solution was added to all experimental wells and mixed well and incubated at 37 ℃ for 20min

(7) Adding 15 μ l MLG developing solution into all experimental wells, mixing well, and incubating at room temperature for 5-30min

(8) The M5 instrument detects readings at 620 nm.

4. Data analysis

The inhibition ratio was calculated by the following formula:

inhibition rate (IC)₅₀) [ OD (sample well) -OD (potassium chloride-containing whole enzyme well) ]/[ OD (potassium chloride-containing whole enzyme well) - (OD (potassium chloride-free whole enzyme well) ] X100%

5. Results of the experiment

Inhibition ratio (IC) of each example compound₅₀) Shown in Table 2

TABLE 2

Compound numbering	IC₅₀(μM)
		Example 1	0.0604
Example 2	0.2071
		Example 3	0.1199
Example 4	0.2635
		Example 5	0.2706
Example 6	0.6746
		Example 7	0.3644
Example 8	0.6108
		Example 9	1.214

As can be seen from Table 2, the compounds of the present invention have excellent H⁺/K⁺ATPase enzyme inhibitory activity, and can be used for preparing gastric acid secretion inhibitor.

Claims

1. A compound represented by the general formula (I) or a pharmaceutically acceptable salt thereof,

wherein:

R¹selected from hydrogen atoms, halogens or alkyl groups;

R²selected from hydrogen, halogen, hydroxyl or alkyl;

2. The compound or pharmaceutically acceptable salt thereof according to claim 1,

R¹selected from halogens;

R²selected from hydrogen atoms, halogens or hydroxyl groups;

R³selected from a hydrogen atom, a fluorine atom or a hydroxyl group;

3. The compound according to claim 1 or 2, or a pharmaceutically acceptable salt thereof,

R¹selected from halogens;

R²selected from hydrogen atoms, halogens or hydroxyl groups;

R³selected from a hydrogen atom, a fluorine atom or a hydroxyl group;

4. A compound or pharmaceutically acceptable salt thereof according to any one of claims 1 to 3, wherein R is¹Is a fluorine atom;

R²selected from a hydrogen atom, a fluorine atom or a hydroxyl group;

R³is a hydrogen atom;

5. The compound or pharmaceutically acceptable salt thereof according to any one of claims 1 to 4, wherein the compound is selected from:

6. A pharmaceutical composition comprising a compound of any one of claims 1 to 5, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier, excipient or diluent.

7. Use of a compound according to any one of claims 1 to 5 or a pharmaceutically acceptable salt thereof or a pharmaceutical composition according to claim 6 in the manufacture of a gastric acid secretion inhibitor.

8. Use of a compound according to any one of claims 1 to 5 or a pharmaceutically acceptable salt thereof or a pharmaceutical composition according to claim 6 in the preparation of H⁺/K⁺-inhibitors of atpase.

9. Use of a compound of any one of claims 1 to 5, or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition of claim 6, in the preparation of a potassium ion competitive acid blocker.

10. Use of a compound according to any one of claims 1 to 5 or a pharmaceutically acceptable salt thereof or a pharmaceutical composition according to claim 6 in the manufacture of a medicament for the treatment and/or prophylaxis of peptic ulcer, zollinger-ellison syndrome, gastritis, erosive esophagitis, reflux esophagitis, symptomatic gastroesophageal reflux disease, barrett's esophagitis, functional dyspepsia, helicobacter pylori infection, gastric cancer, gastric MALT lymphoma, ulcers caused by non-steroidal anti-inflammatory drugs or hyperacidity or ulcers caused by post-operative stress; or inhibiting upper gastrointestinal hemorrhage due to peptic ulcer, acute stress ulcer, hemorrhagic gastritis or invasive stress.